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Platelet-rich Plasmas: Growth Factor Content and Roles in Wound Healing
Abstract
Unlabelled:
Platelet-rich plasmas (PRPs) are used in a variety of clinical applications, based on the premise that higher growth factor content should promote better healing. In this study, we have determined the effects of calcium and thrombin on the release of EGF, TGF-alpha, IGF-1, Ang-2 and IL-1beta from PRPs, and assessed the mitogenic potential of PRP supernatants on osteoblast and endothelial cell division. ELISA assays indicate that (i) mean growth factor concentrations vary from traces (TGF-alpha) to 5.5 ng/mL (IGF-1), (ii) there are significant variations in growth factor concentrations between individuals, and (iii) calcium and thrombin regulate growth factor release, synthesis, and/or degradation in stereotyped patterns that are specific to each growth factor. PRP supernatants promote strong osteoblast and endothelial cell divisions, supporting the concept that PRPs may be beneficial in wound healing.
Abbreviations:
PRPs, platelet-rich plasmas; GFs, growth factors; EGF, epidermal growth factor; TGF-alpha, transforming growth factor-alpha; IGF-1, insulin-like growth factor-1; Ang-2, angiopoietin-2; IL-1beta, interleukin-1 beta; HUVECs, human umbilical vein endothelial cells; hFOB 1.19, human fetal osteoblasts; and FBS, fetal bovine serum.
... 6,7) Thus, the fact that PRP contains high concentrations of platelets with a large variety of bioactive proteins suggests that PRP can accelerate the tissue regeneration and healing processes. 8) Growth factors such as vascular endothelial growth factor, epidermal growth factor (EGF), platelet-derived growth factor (PDGF), insulinlike growth factor, and transforming growth factor-beta (TGF-β) are involved in tissue healing, which are included in α-granules of platelets. It has been revealed that PDGF and TGF-β levels increase as platelet levels become higher, which has been the basis for the clinical use of PRP. ...
... Despite recent studies on major growth factors in PRP that are related to tissue healing including EGF and PDGF, basic medical studies on their effects still remain to be conducted. 8,11) In addition, the compositions and expression patterns of active proteins in PRP need to be studied. Rapidly identifying and analyzing proteins, which used to be impossible, have become possible, and it became easy to access GO databases. ...
Background
Platelet-rich plasma (PRP) is a plasma component of autologous blood containing a high concentration of platelets. PRP is used to promote healing of damaged tissues. However, there are not many studies on the composition and expression patterns of active proteins in PRP. The purpose of this study was to identify unknown factors that contribute to tissue healing by proteomic analysis of proteins in PRP.
Methods
Three men in their 30s with no basal disease participated in this study. All identified proteins were classified for tissue healing-related functions on the basis of the gene ontology analysis of adhesion molecule with Ig-like domain 2 (AmiGO2). PRP was prepared by using the ACP kit and GPS III kit.
Results
We identified a total of 125 proteins related to wound healing, along with three proteins for angiogenesis involved in wound healing, two proteins for fibroblast migration, four proteins for …
... It has a high concentration of various growth factors, such as insulin-like, vascular, and fibroblast growth factors (Table 1). These factors can affect the development of specific cell types, which can be beneficial for the healing process of tissues [8,30]. The global market for platelet-rich plasma is expected to grow steadily over the next few years. ...
... Bone regeneration with platelet-rich plasma (PRP) is designed to stimulate a healing response at fracture and fusion sites around the foot and ankle using platelet-derived products. In several preclinical studies, PRP has been shown to improve osteogenesis [8,[21][22][23][24][25][26][27][28][29][30][31][141][142][143][144][145][146][147][148][149][150]. However, the translation of preclinical findings to in vivo bone repair has shown mixed results. ...
Advances in the Clinical Application of Platelet-Rich Plasma in the Foot and Ankle: A Review. Abstract: Autologous and recombinant biologic substances have been generated as a result of the research into the cellular features of the healing process. Orthobiologics are increasingly being used in sports medicine and musculoskeletal surgery. Nevertheless, clinical data are limited; consequently, further studies are required, particularly in foot and ankle pathologies. This review aims to provide evidence of the most recent literature results and ignite the interest of orthopedic specialists eager for an update about the most current discussion on platelet-rich plasma (PRP) clinical applications in the foot and ankle fields. Previous studies have shown that platelet-rich plasma can be beneficial in treating various conditions, such as chronic foot ulcers, osteoarthritis, Achilles tendinopathy, etc. Despite the positive effects of PRP on various musculoskeletal conditions, more prospective studies are needed to confirm its effectiveness at treating ankle and foot pathologies. In addition to clinical trials, other factors, such as the quality of the research and the procedures involved, must be considered before they can be used in patients. More long-term evaluations are needed to support or oppose its application in treating foot and ankle disorders. We present the most extensive review of PRP's clinical applications in the foot and ankle field.
... Considering EGF levels before and after treatment with both ASA and Clopidogrel, there was an increase in mean concentrations (compared to baseline) after treatment in both study groups, however, no Statistically significant differences were found in none of the platelet derivatives. The baseline values of this GF were similar to those reported by other authors [48][49][50], such as Eppley B et al. (2004), who found EGF in the PRP of 470 ± 320 pg / ml [48], these levels being approximate to baseline and posttreatment for the AAS group, but not for the Clopidogrel group. For the latter group, the concentrations were lower than those of ASA. ...
... The above has also been reported by other authors who indicate that each individual would need different concentrations of their cells, especially of the platelets, to achieve a secretion of GFs with comparable biological effect [44,48]; And that the interindividual variations found may not be correlated with the platelet count, thus, an individual with high levels of CF can exhibit the lowest concentrations of another GF [44,49]. ...
PRP is an useful bioproduct to tisular regeneration. The aim of study was evaluate the concentration of growth factors: PDGFBB (platelet-derived growth factor), EGF(epidermal growth factor) and VEGF (vascular endothelial growth factor) present in the Platelet-rich plasma (PRP) in subjects treated with drugs which inhibit platelet aggregation as acetylsalicylic acid (ASA) and clopidogrel before and after administration. We determined by ELISA PDGFBB, EGF and VEGF levels in PRP, Platelet Poor Plasma (PPP), lysate and exudate from 32 healthy subjects before and 24 hours after ingesting acid Acetyl salicylic acid (ASA) and clopidogrel as a single dose. The PRP and PPP were obtained by the method of Anitua by single centrifugation method. To analyze the results of student test and Pearson correlation was applied, with statistical significance level of p < 0.05. PPP and exudate (Clopidogrel: p < 0.001), PRP (Clopidogrel: p < 0.01) statistically significant differences for PDGFBB in PPP (p < 0.01 AAS) were found, and for VEGF in lysate (ASA and Clopidogrel: p < 0.05). No significant difference was found for EGF. Only was no correlation between baseline values of EGF in the ASA group and the respective PRP platelet count (r = 0.726). The results show that the average basal values of the three growth factors measured were considered particularly high in the PRP and lysate, showing the significant decrease for PDGFBB after antiplatelet therapy, especially of Clopidogrel and a significant increase for the VEGF only for the lysate. Although the behavior of three different soluble mediators was different to antiplatelet agents, the observed changes support the conclusion that a single dose of these drugs not markedly affect the secretion and availability of the three growth factors measured in various platelet derived obtained.
... Clinical outcomes have shown that the age is not a major limitation when using platelet and plasma-based products for the treatment of different dermatological pathologies, and major age-specificand sex-specific differences in individual growth-factor concentrations were not found [14]. ...
... As regards the different centrifugation methods, the double-spin low-centrifugation method showed a 140% increase in platelet count than the baseline value. These results are in agreement with Sabarish et al. [4], who demonstrated a 160% increase in basal platelet using the initial protocol by Marx, although Fréchette et al. [14] reported a higher increase up to 200-300%, this difference may be explained by the ethnic variation and the initial volume for PRP preparation. ...
... Currently, the classification of platelet-rich forms is presented as follows [1,2] In the preparation of PRP, the use of thrombin, calcium or other biologically safe anti-coagulants is often required [8][9][10]. These factors can have an adverse effect on the coagulation process and also lead to an immune response. ...
... Nguyen and Pham [13] also claimed that it can be concluded that in vitro human gingival fibroblast (hGF) traits depend on the concentration of PRP in the medium. So far, some recent studies have shown that increasing the levels of PRP leads to increased proliferation in cells [9,10]. ...
Blood derivates, such as platelet-rich plasma (PRP) and platelet-rich fibrin (PRF), are autogenous sources of many growth factors that are involved in the healing and regeneration of tissues, and for this reason, are used in dentistry treatments. This fact also contributes to the growing interest in these biomaterials in regenerative personalized medicine. The multitude of platelet-rich forms creates many possibilities for their use. This semi-systematic review describes and compares the methods of obtaining properties and potential uses of these materials in personalized treatments.
... 1,2) Recent studies suggest that growth factors promote tissue repair, bone formation, and regeneration. [2][3][4] As such, PRP has been increasingly used in recent years in the fields of orthopedics, orthopedic arthroscopic surgery, periodontics, maxillofacial surgery, urology, plastic surgery, and sports medicine, 5) where it facilitates healing in the tendons, ligaments, and cartilage. [6][7][8] However, although factors that originate from platelet α-granules influence tissue regeneration, catabolic signals from leukocytes may hinder healing in association with inflammatory cytokine expression. ...
Background
Leukocyte-poor platelet-rich plasma (LP-PRP) from peripheral blood is currently used as a concentrated source of growth factors to stimulate repair at sites of soft tissue injury. Fibroblasts are primary mediators of wound healing. Thus, we aimed to assess the positive effect of LP-PRP on human fibroblast proliferation in vitro.
Methods
LP-PRP was prepared from 49 donors. The fibroblasts were seeded, and at 24 hours after seeding, 1× 10 7/10 µL LP-PRP was added once to each well. The cells were harvested 10 times during study period at our planned points, and we examined cell proliferation using the water-soluble tetrazolium salt-1 assay. We collected the supernatants and measured the amount of growth factors such as platelet-derived growth factor (PDGF)-AB/BB, insulin-like growth factor-1 (IGF-1), transforming growth factor-β1 (TGF-β1), and vascular endothelial growth factor (VEGF), which are …
... It has a concentration of platelets five times greater than normal blood, at least 1,000,000 platelets/μl in 5 ml of plasma [8]. Platelets are best known for their function in hemostasis, but they also promote and control wound healing via a variety of growth factors, cytokines, and bioactive molecules [9,10]. During healing, growth factors have major impacts on cell regulation, differentiation, proliferation, migration, chemotaxis, angiogenesis, matrix formation and collagen synthesis [10,11]. ...
Purpose
Chronic rhinosinusitis (CRS) is a prevalent chronic disease observed on a global scale. The utilization of endoscopic sinus surgery (ESS) has gained significant recognition as an effective intervention for individuals with CRS and nasal polyps who have not responded to conventional treatments. The need (or not) for revision surgery frequently relies on the promotion of optimal wound healing. The impact of platelet-rich plasma (PRP) on tissue healing has been extensively examined in various surgical fields.
Methods
The present prospective study involved 30 patients suffering with nasal polyposis who underwent endoscopic sinus surgery. 15 patients were assigned to the PRP group, and 15 patients to the control group. The clinical follow-up of the patients took place at specific intervals, at weeks 1, 2, 3, 4, 8, and 12 after the surgical procedure. The evaluator identified the existence of adhesions, crusting, bleeding, granulation and infection using a visual analogue scale score. The patients also completed the SNOT 22 questionnaire prior to surgery and at each postoperative visit.
Results
The present study observed a lower incidence of adhesion, infection, hemorrhage and granulation in the PRP group. Furthermore, a statistically significant difference was detected between the groups.
Conclusion
Based on the findings of the present investigation, it seems that platelet-rich plasma (PRP) is beneficial on wound healing during the early stages following the surgical procedure. The technique is characterized by its limited invasiveness, which contributes to its low risk profile and the achievement of clinically good outcomes.
... This excludes any possibility of side effects that could be associated with the use of chemical substances, and excludes the possibility of cross infections. Compared to the other forementioned methods, platelet concentrates have the superiority for [1] being rich in growth factors that are important for bone healing [24][25][26], (2) contains variable blood proteins that are important for osteoblasts and fibroblasts chemotaxis [27], (3) considered a potent initiator for bone regeneration and apposition when injected at a local site [27][28][29][30][31]. ...
Background
Mid-Palatal suture expansion needs long retention period due to delayed bone formation in the expanded suture. Platelet-rich plasma (PRP) is a concentrated source of growth factors which increase bone formation. The aim of this study was to evaluate the effect of PRP injection on bone formation in expanded mid palatal suture in rabbits.
Methods
In this prospective randomized controlled animal study, Twenty male rabbits (8-weeks-old) were subjected to mid-palatal expansion for 5 days. Animals were afterwards randomly divided into control group A & study group B. PRP was prepared and injected in the mid-palatal suture in animals belonging to group B only. After 6 weeks of retention, all animals were euthanized, and premaxillae were prepared for histological, histomorphometric and immunohistochemical analysis. Student t-test and paired t-test were used to compare the means of the two groups and within the same group respectively. Significance level set at p ≤ 0.05.
Results
Histomorphometric analysis revealed a significant increase (p < 0.001) in the mean percentage of new bone in the study group (14.4%) compared to the control (1.4%). Suture width in study group was significantly wider than the control group (278.8 ± 9μms and 120.4 ± 3.4μms, p < 0.001). There was a significant increase in vascular density in study group than control group (309 ± 65.34 and 243.86 ± 48.1, p = 0.021). Osteopontin immuno-expression revealed a significant increase in optical density in study group than control group (0.21 ± 0.02 & 0.12 ± 0.01, p < 0.001).
Conclusions
In rabbit model, PRP injection can accelerate new bone formation in the expanded mid-palatal suture when compared to the control. This could hopefully result in a more stable midpalatal expansion and a reduced retention period.
... 27 The release profile of GFs was assessed as it corresponds to the proliferative and maturation phase of wound healing. 28 In addition to the activated platelets and leukocytes, the fibrin network in PRF acts as a reservoir for GFs, enabling a continuous release profile. 29 There is no significant difference in the mean levels of PDGF-BB, PDGF-AB, and PDGF-AA among the three groups in A-PRF preparation on intergroup comparison. ...
... Thrombin, calcium or other biologically acceptable anticoagulants are frequently needed for PRP preparation [13][14][15]. These components may interfere with the coagulation process and trigger an immunological reaction. ...
Recent research on the use of platelet concentration in medicinal dentistry has enhanced the potential for tissue regeneration. The ability of platelets to enhance tissue regeneration by using different forms of platelet concentration (blood component) such as platelet-rich plasma (PRP), platelet-rich fibrin (PRF), standard platelet-rich fibrin, advanced platelet-rich fibrin, etc. PRP became widely used once its potential for tissue regeneration was discovered; however, the use of anticoagulants restricted the easy use of PRP as compared to PRF; however, after the discovery of PRF because of the easy formulation, it became widely used in medicinal dentistry. The purpose of this review is to elaborate on the importance of platelet concentration in periodontal regeneration.
... However, it is noteworthy that platelets also play a significant part in the beginning and control of wound healing. This crucial activity involves the release and modulation of a wide array of cytokines, growth factors, and other bioactive molecules [7,8] . Platelets possess a recognized role in the process of haemostasis, but they also have the ability to produce several chemicals that facilitate tissue healing, angiogenesis, and inflammation [9] . ...
... Due to the unique biological properties described in the previous section, PRP therapy inherently lends itself to therapeutic utility in several clinical scenarios to promote wound healing [30]. Specifically, in the field of orthopedic surgery, evidence suggests that PRP therapy may be beneficial for patients that present with knee osteoarthritis, hip osteoarthritis, and lateral epicondylitis, as well as in the setting of TJA due to said conditions [25,31,32]. ...
Orthobiologics are playing an increasingly large role in the clinical setting across multiple fields of surgery. Particularly, in the field of orthopedic surgery, the employment of platelet-rich plasma (PRP) therapy in total joint arthroscopy (TJA) has become popular for its prorupted benefits of controlling pain, blood loss, and increased wound healing. PRP was originally used for thrombolytic conditions, however, the aforementioned potential benefits have led to its increased use across various fields of medicine including dermatology, neurosurgery, orthopedics, and sports medicine. Currently, there is a persisting gap in the literature surrounding the mechanism of action of PRP, as well as its true role in increasing positive patient outcomes in the context of TJA. Thus, this review aims to briefly highlight the physiological mechanisms underlining PRP therapy, evaluate recent preclinical and clinical data about its effects on TJA patient outcomes, and to describe its concomitant use in novel orthopedic applications.
... The fold concentration of thrombocytes after preparation by a single-or double-centrifugation process ranged from 2-to 33-fold. Platelets are rich in many growth factors involved in different healing processes: platelet-derived growth factors (PDGF-aa, PDGF-ab and PDFG-bb isomers), transforming growth factor-beta (TGFβ), VEGF, interleukin-1, platelet-derived angiogenesis factor, platelet-derived endothelial growth factor, insulin-like growth factor (IGF-1), osteocalcin and osteopontin [8,114]. Platelets are the first elements that release these growth factors at the fracture site, followed by macrophages, whose chemotaxis is stimulated by the factors released from the platelets. ...
The ideal biomaterials substitute for bone repair should possess the following characteristics: provide osteogenic, osteoinductive and osteoconductive properties; stimulate the neo-angiogenesis process; absence of antigenic, teratogenic or carcinogenic reactions; avoid the systemic toxicity complications; assure satisfactory support and stability from mechanical properties point of view; hydrophilic nature of the surface properties and good interface with human bone; good handling in clinical condition and ability to be easy sterilized; and able to be supplied in sufficient quantities with reduced costs. Despite years of effort, the perfect bone reconstruction material has not yet been developed; further effort is required to make this objective feasible. The aim of this article is to provide a contemporary and comprehensive overview of the grafting materials that can be applied for the treatment of bone defects by the clinicians from orthopedics surgery, neurosurgery and dentistry, discussing their properties, advantages and disadvantages, and illuminating present and future perspectives in the field of bone graft and biomaterials substitutes for bone repair, from clinical needs to advanced biomaterials research.
... However, despite the documented hyporeactivity of neonatal platelets, bleeding time and platelet closure time have actually been found to be shorter in full term infants compared to adults [62,67]. From a wound healing perspective, neonatal derived platelet-rich plasma (PRP) has been demonstrated to increase proliferation, differentiation, and migration of various cell types, including endothelial cells, mesenchymal stem cells, and dermal fibroblasts, all crucial for wound healing and may be a viable therapeutic for enhancing healing outcomes [68,69]. Developing pro-healing materials that modulate the concentrations of various clotting factors, use fetal fibrinogen, or employ neonatal PRP could leverage the advanced properties of the fetal and neonatal hemostatic system to create a fetal-like healing environment. ...
Chronic non-healing wounds are a significant healthcare challenge. Various biomaterials have been developed to treat chronic wounds but there are still opportunities for improvement of biomaterial therapeutics. This review discusses how fetal wound healing could be used as inspiration to develop pro-healing materials. Compared to adults, fetuses have enhanced wound healing outcomes and healing without scarring. Scarless fetal wound healing is associated with various key differences in several growth factors, cytokines, extracellular matrix components, and coagulation parameters. Mimicking the fetal wound healing environment through bioinspired materials could create improved therapeutics to treat chronic wounds. This review addresses the key differences between adult and fetal wound healing that allow for enhanced scarless fetal healing and discusses how these differences can be used to develop pro-healing materials.
... • As PRP is obtained from autologous whole blood, the therapeutic effects depend on the donor, and PRP processed from different donors vary substantially. [247] Additionally, pain practitioners can prepare PRP either by manual centrifugation or commercial kits which provide different PRPs with varying number of platelets and leukocytes. [248,249] • Var ious considerations are made du r ing PR P administration, including but not limited to: method of preparation, injection site, PRP volume, number and frequency of injection, concomitant treatment, and the use of anesthetics. ...
Introduction: Osteoarthritis knee (OAK) is the most common form of osteoarthritis and has a global prevalence of 22.9% among individuals
aged 40 years and above. Although primarily considered a disease of advancing age, OKA is known to affect the younger age group, leading
to substantial clinical, economic, and social burden. Although a plethora of therapies are available for OAK, patients ultimately become
refractory to them and require arthroplasty, thereby adding further to the burden. Over the decades, novel therapies have become available,
and accumulated researches suggest that platelet‑rich plasma (PRP) is both safe and effective in patients with OKA of varying severity. Thus,
we aimed to formulate the International Society for Musculoskeletal Ultrasound in Pain Medicine (ISMPM) guidelines for the management
of symptomatic OAK with intra‑articular PRP. Methods: An extensive literature review by pain experts was performed, resulting in 9
statements. Quality of evidence was assessed with the Third US Preventive Services Task Force guidance document, and the strength of the
recommendation was determined by the modified Delphi consensus process. Results: The level of evidence was I for four statements, II‑1 for
two, III for two, and II‑2 for one statement. Moreover, for most statements, the strength of consensus among the experts was strong (≥80%
of the experts). Conclusion: The ISMPM guidelines for the management of OAK are developed by expert pain practitioners. For nearly half
of the statements, the highest level of evidence was available, and consensus was strong. However, further high‑quality research is needed to
come up with more comprehensive guidelines in this promising pain medicine specialty.
Keywords: Guidelines, hyaluronic acid, knee, modified Delphi process, osteoarthritis, pain medicine, platelet‑rich plasma
... Khalmuratova et al. (15) guided further studies on the healing stages. The positive effects of PRP on wound healing by secreting various growth factors and cytokines have been shown in various studies (16)(17)(18). Therefore, the hypothesis of this study was that PRP would reduce crusting in the nasal mucosa and have a positive effect on postoperative pain. ...
Introduction:To evaluate the effect of submucosal platelet-rich plasma (PRP) therapy on intraoperative bleeding and early postoperative pain and crusting in patients undergoing inferior turbinate radiofrequency.Methods:A total of 70 adult patients with isolated inferior turbinate hypertrophy were included in this prospective study and randomized to the PRP or control groups. PRP was prepared for all patients. After applying submucosal radiofrequency to the inferior turbinates under local anesthesia, submucosal PRP was injected into the study group, and submucosal saline was injected into the control group. Patient controls were performed by another specialist. The patients and the specialist who performed the controls were blinded to which group they were in. All patients were evaluated endoscopically 1, 7, and 21 days after the operation.Results:The mean age of the patients was 33.37±11.92 years (range: 18 to 54). The two groups had no significant differences in intraoperative bleeding and mucociliary clearance values (p>0.05). The amount of crusting and the visual analog scale pain values were significantly lower in the submucosal PRP -injected group (p
... PRP gel is actually convenient biomaterial used for union of fracture site [6,22,23] in this single case study we noticed despite infection, union of nonunion fracture end were established by delivering PRP gel at gap of fracture site. Further randomized multicenter clinical trials need to be carried out. ...
... Platelet-rich plasma contains α-granules of platelets and several cytokines, including platelet-derived growth factor, TGF-β, VEGF, IGF, EGF, and bFGF. [22][23][24] Samberg et al 25 reported that human PRP increased the mRNA expression levels of VEGF-A and ANGPT-1 in cultured human ADSCs but decreased the mRNA levels of ANGPT-2. In FIGURE 5. Effects of the conditioned medium of PRP-treated ADSCs under hypoxia on NO production in HUVECs. ...
Platelet-rich plasma (PRP) and adipose-derived stem cells (ADSCs) are known to secrete angiogenic factors that contribute to the treatment of intractable ulcers. The combination of PRP and ADSCs may enhance their angiogenic effects. However, it remains unclear whether treatment of ADSCs with PRP influences angiogenesis. We studied whether the conditioned medium from PRP-treated ADSCs under hypoxic conditions exerts angiogenic effects. Although PRP stimulated the proliferation of ADSCs obtained from rats, it decreased the mRNA levels of vascular endothelial growth factor, hepatocyte growth factor, and TGF-β1, but not of basic fibroblast growth factor, under hypoxia. The conditioned medium of PRP-treated ADSCs inhibited endothelial nitric oxide synthase phosphorylation, decreased NO production, and suppressed tube formation in human umbilical vein endothelial cells. Transplantation of ADSCs alone increased both blood flow and capillary density of the ischemic limb; however, its combination with PRP did not further improve blood flow or capillary density. This suggests that both conditioned medium of ADSCs treated with PRP and combination of PRP with ADSCs transplantation may attenuate the phosphorylation of endothelial nitric oxide synthase and angiogenesis.
... Platelets contain high amounts of growth factors like transforming growth factor β-1 (TGFβ-1), platelet-derived growth factor (PDGF), epithelial growth factor (EGF), and vascular endothelial growth factors (VEGF). When used in combination with bone grafts, the platelet concentrates supposedly stimulate cellular proliferation and upregulate angiogenesis in order to accelerate bone graft healing over the bony defect [9]. The concentrated growth factor (CGF) is the third generation of the platelet concentrate product, which has been introduced in the recent past. ...
Background
The current study’s aim was to evaluate and compare demineralized freeze-dried bone allograft (DFDBA) with and without concentrated growth factor (CGF) membrane in the treatment of periodontal intrabony defects on both a clinical and radiographic level.Methods30 stage III grade B periodontitis patients with bilateral intrabony defects were involved in the split mouth randomized controlled trial, and they received either DFDBA alone (control group) or DFDBA plus CGF membrane treatment (test group). At baseline and 6 and 12 months, the gingival index (GI), plaque index (PI), probing pocket depth (PPD), clinical attachment level (CAL), and gingival recession (GR) were assessed, whereas cone beam computed tomography was used to assess the bone defect height (BDH), vertical bone loss (VBL), bone defect depth (BDD), mesio-distal bone defect width (MDBDW), bucco-lingual bone defect width (BLBDW), and bone defect volume (CBCT) at baseline and 12 months.ResultsThe radiographic measures BDH, BDD, MDBDW, BLBDW, and BDV in the test group likewise showed a higher reduction in PPD and gain in CAL than the control group. The results of the intergroup comparison showed that the test group had statistically significant differences in BDH, BDD, and MDBDW.Conclusion
The data show that the test group achieved better results, with greater reductions in PPD, gains in CAL, and decreases in BDH, BDD, MDBDW, BLBDW, and BDV.Clinical relevanceThe use of concentrated growth factor in conjunction with demineralized freeze-dried bone allograft may be suggested for the treatment of periodontal intrabony defects.
... Another important factor seems to be the proteomic analysis of the injected PRP and identification of proteins which contribute more to tissue healing. PRPs containing high concentration of platelets contain high quantity of bioactive proteins (such as growth factors and cytokines) which can promote tissue healing and regeneration and this has been and still is extensively researched [45,46]. ...
Objective
To assess the effect of PRP on knee articular cartilage content (thickness/volume) and examine the correlation between cartilage changes and clinical outcomes in patients with knee OA.
Method
A systematic literature search was performed using the Cochrane methodology in four online databases. Studies were included if they reported on cartilage content with cross-sectional imaging pre- and post-injection. A random-effects model meta-analysis was performed. Correlation with clinical outcomes was evaluated.
Results
14 studies (n = 1099 patients) from 1452 records met the inclusion criteria: seven RCTs (n = 688), one prospective (n = 50), one retrospective (n = 68), and four case-series (n = 224). The PRP preparation process and treatment protocol varied widely (follow-up 6–12 months). In meta-analysis, PRP treatment was not associated with a significant increase in cartilage thickness (4 studies, n = 187, standardized mean difference: Hedges g: 0.079; 95%CI: 0.358 - 0.516; p = 0.723). Meta-analysis of 3 RCTs (n = 112) showed no significant difference in the change of overall knee cartilage content with PRP injections compared with no PRP (Hedges’ g: 0.217; 95%CI: 0.177 – 0.611; P = 0.281).
Conclusion
The current literature does not support the PRP as chondrogenic in treatment of knee OA. However, there is substantial heterogeneity in the evaluated studies which limits the robustness of any conclusion. An adequately powered RCT, with a standardized PRP regime and standardized high-resolution MRI is needed to definitely define any effect of PRP on knee cartilage content and its relation to clinical outcomes. Until such high-quality evidence becomes available, we recommend that PRP is not administered with the intention of promoting chondrogenesis.
... The latter process also kicks start wound healing mediated in part by angiogenesis and fibroblastic activity. Platelets can be concentrated two and a half to eight folds into a platelet-rich plasma (PRP) preparation [1]. There is no prescribed standard concentration noted in the literature. ...
Background
Platelets are rich in cytokines and growth factors. Exposed tendons in wounds do not naturally heal by granulation and epithelization. The study aimed to explore the effects of PRP injection therapy on exposed tendons in open wounds and determine if the tendon could support wound healing.
Methods
A retrospective observational clinical study was undertaken from 2012 to 2018 to assess wound healing from exposed tendons in wounds in patients treated with PRP injections and occlusive dressings. Parameters studied included patient and management factors, wound and functional outcomes, wound healing progression, and the direct effects of PRP therapy on tissues.
Results
Twenty-three patients with several comorbidities received treatment. The average age of patients was 56 years, with an age range of 25 to 79 years. Twenty of the 23 patients (87%) reached complete healing. Eighteen of the 20 (90%) had good tendon excursion and associated joint movement for the limb’s function. The complication rate was low. PRP injection therapy produced a response of increased vascularity, the proliferation of granulation tissue from the tendon, and epithelialization from the surrounding skin.
Conclusions
Intra-tendinous PRP injections used with occlusive dressings can heal the exposed tendon and open wound by process of granulation and epithelization, restoring adequate limb function.
Level of evidence: Level IV, Therapeutic study.
... Albumin is one of the main constituents of such products; however, it is rarely considered a key element in the therapeutic efficacy of blood products. There have been several reports on the successful use of blood products in orthopedic and trauma surgery, plastic surgery, spinal surgery, heart bypass surgery, chronic cutaneous and soft tissue ulcerations, wound healing and burns, periodontal and oral surgery and maxillofacial surgery [124][125][126][127][128][129][130][131][132]. Blood products are used not only in vivo but also as a regenerative factor in numerous in vitro studies. ...
Albumin is a constitutional plasma protein, with well-known biological functions, e.g., a nutrient for stem cells in culture. However, albumin is underutilized as a biomaterial in regenerative medicine. This review summarizes the advanced therapeutic uses of albumin, focusing on novel compositions that take advantage of the excellent regenerative potential of this protein. Albumin coating can be used for enhancing the biocompatibility of various types of implants, such as bone grafts or sutures. Albumin is mainly known as an anti-attachment protein; however, using it on implantable surfaces is just the opposite: it enhances stem cell adhesion and proliferation. The anticoagulant, antimicrobial and anti-inflammatory properties of albumin allow fine-tuning of the biological reaction to implantable tissue-engineering constructs. Another potential use is combining albumin with natural or synthetic materials that results in novel composites suitable for cardiac, neural, hard and soft tissue engineering. Recent advances in materials have made it possible to electrospin the globular albumin protein, opening up new possibilities for albumin-based scaffolds for cell therapy. Several described technologies have already entered the clinical phase, making good use of the excellent biological, but also regulatory, manufacturing and clinical features of serum albumin.
... Generally, Platelet rich plasma (PRP) is an autologous blood derivative rich in active growth factors (GFs), some of which are hepatocyte growth factor (HGF); insulin-like growth factor-I (IGFI) and epidermal growth factor (Shoeib et al, 2018). PRP powerful therapeutic potentials is due to ability of del-ivering great variety of biologically active GFs to injury site as being simple, effective, safety and ability (Frechette et al, 2005). Also, it was related to powerful healing and an up-growing role in regenerative medicine (Alves and Grimalt, 2018). ...
... what constitutes PRP) [42] For instance: demographical variation in GF concentration (age, sex, platelet count and physical exertion prior to donation), centrifugation speed (e.g. due to apparatus variations), time from preparation to analysis, and in the case of Ca-PRP and PRF, the concentration and nature of the activating factor [27], [42], [43], [44], [45]. Moreover, it must be noted that platelets are precious clinical material. ...
Objectives: Platelet-derived products have been shown as promising novel therapeutic agents for chronic wounds. However, their use clinically requires a greater degree of method standardisation, part of which involved more extensive cataloguing of their biochemical composition. This study aimed to quantify and compare total protein and 6 angiogenically-active growth factors in three distinct platelet products.
Methods: Platelet Lysate (PL, n=5), Calcium-activated Platelet Rich Plasma (Ca-PRP, n=5) and Platelet-Rich Fibrin (PRF, n=5) were prepared from pooled platelet apheresis products (n=10). Ca-PRP and PRF were prepared from the same units (n=5) by activation with 20 mmolL-1 calcium chloride. PL was prepared from the remaining (n=5) units using an established lysate. Total protein was quantified with the Bradford Assay. Sandwich enzyme-linked immunosorbent assay was used to quantify six growth factors: epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), stromal cell de-rived growth factor-1α (SDF-1α), endostatin, and transforming growth factor-β1 (TGF-β1).
Results: Protein retrieval differed significantly (p<0.05) between the three prod-ucts: PL (11.35±0.80 mg/mL) < Ca-PRP (20.44±8.17 mg/mL) < PRF (40.67±3.13 mg/mL). Growth factor yield was considerable in all three products and differed significantly for: VEGF (PL<PRF); EGF (Ca-PRP<PRF); HFG (PL<Ca-PRP); Endostatin (PL<Ca-PRP, PRF<Ca-PRP, PL<PRF) and TGF-β1 (Ca-PRP<PL, Ca-PRP<PRF).
Conclusions: Platelet apheresis products contain a substantial quantity of the in-vestigated pro- and anti-angiogenic growth factors. Their release varies depending on the manufacturing protocol used. Clinically, alternate products could thus be combined to provide a therapeutically optimal mix of growth factors.
... In addition to exogenous administration of the above-mentioned GFs for promotion of wound angiogenesis, platelets have also received significant attention as a source of GFs and cytokines. Platelet-rich plasma (PRP) is obtained by gradient centrifugation of the patient's autologous whole blood [142], and is rich in PDGF-BB, TGF-β, FGF-2, VEGF-A, EGF, etc. [143]. Autologous PRP offers an affordable and safe source of multiple GFs and cytokines required for improved angiogenesis and accelerated wound healing [144]. ...
Management of skin wounds is of great clinical importance. Inducing the formation of new blood vessels (angiogenesis) can accelerate the healing process in patients suffering from acute and chronic wounds (e.g., burns and diabetic ulcers). It is currently accepted that angiogenesis plays a substantial role in all four overlapping phases of normal wound healing, that is, hemostasis, inflammation, proliferation, and remodeling. Many experimental studies have focused on the design and development of skin replacements capable of stimulating neovascularization within the skin defect regions. The use of stem/progenitor cells and bioactive molecules (e.g., growth factors and cytokines) in combination with three-dimensional (3D) scaffolds are among the most promising applied strategies for promoting angiogenesis, and subsequently accelerating wound healing. Each approach has its own pros and cons for managing skin wounds. In the present chapter, we first describe the physiology of skin tissue, and the diseases and disorders which affect wound healing. We then summarize current therapies with a focus on tissue-engineering (TE) approaches. The critical role of proangiogenic strategies are discussed to highlight their importance for future studies
... The finding of Sema4D released by platelets ( Figure 1A-C) corroborates the results of previous studies [26,27,31,32]. We also confirmed that platelets release IGF-1 ( Figure 1D), which is known as a factor that promotes osteoblastogenesis [33]. Indeed, IGF-1 is secreted by many tissues, such as liver, kidney and bone, acting as an endocrine hormone to promote bone formation [34,35]. ...
Effects of the antiosteoblastogenesis factor Semaphorin 4D (Sema4D), expressed by thrombin-activated platelets (TPs), on osteoblastogenesis, as well as osteoclastogenesis, were investigated in vitro. Intact platelets released both Sema4D and IGF-1. However, in response to stimulation with thrombin, platelets upregulated the release of Sema4D, but not IGF-1. Anti-Sema4D-neutralizing monoclonal antibody (mAb) upregulated TP-mediated osteoblastogenesis in MC3T3-E1 osteoblast precursors. MC3T3-E1 cells exposed to TPs induced phosphorylation of Akt and ERK further upregulated by the addition of anti-sema4D-mAb, suggesting the suppressive effects of TP-expressing Sema4D on osteoblastogenesis. On the other hand, TPs promoted RANKL-mediated osteoclastogenesis in the primary culture of bone-marrow-derived mononuclear cells (BMMCs). Among the known three receptors of Sema4D, including Plexin B1, Plexin B2 and CD72, little Plexin B2 was detected, and no Plexin B1 was detected, but a high level of CD72 mRNA was detected in RANKL-stimulated BMMCs by qPCR. Both anti-Sema4D-mAb and anti-CD72-mAb suppressed RANKL-induced osteoclast formation and bone resorptive activity, suggesting that Sema4D released by TPs promotes osteoclastogenesis via ligation to a CD72 receptor. This study demonstrated that Sema4D released by TPs suppresses osteogenic activity and promotes osteoclastogenesis, suggesting the novel property of platelets in bone-remodeling processes.
... PRP are small amount of autologus derived plasma rich in platelets (Roukis et al, 2006) that contain granules on its surface which release many factors and proteins that increase healing rate of the bone fractures complications and defect (Browner et al, 2014) by angiogensis stimulation, active proliferations and rapid differentiation of mesenchymal and osteoplasts cells (Frechette et al, 2005;Garcia et al, 2012). Cho et al (2013) and Roffi et al (2017) refer that improvement in bone regeneration in rabbits after grafting/implantation due to PRP application. ...
The experimental work planned for follow up the injection locally, autologus derived platelets rich plasma (PRP) on repair experimentally 3.5 mm hole wide in the femoral mid shift bone for this operation 24 adults mature rabbits were employed, after the operation the animals divided to two equal group each of (n-12), control group (CG) follow for normal bone repair and treatment group (TG) which injected the prepared autologus derived (PRP) locally in the hole, the results followed by weekly radiological image and histopathological finding. The radiological results showed clear and wide in sclerotic area at the hole defect ,with widening in the cortex diameter In the treated experimental animals more clear and rapid compare to the non treated animals, and the end results of the histological sections in the hole center showed profuse woven trabeculae bone formation in the end of the 1 st week, which converted to matures trabeculae bone with area of lamellar and compact bone formation after 14th and 21days from the surgical operation. clear and well marked in TG than CG. We conclude from the above results that using autologus derived (PRP) strongly and successfully repair the 3.5mm long bone defect.
... Хронічні дегенеративно-дистрофічні ураження хребта стають проблемою цивілізації, так званою «платою за прямоходіння» [1], тому пошук методів відтворення структурної цілісності міжхребцевих дисків та тканин, що їх оточують, є актуальним питанням сучасної медичної науки. З метою стимулювання регенерації міжхребцевих дисків останнє десятиріччя використовуюється збагачена тромбоцитами плазма, активно застосовується при лікуванні широкого кола патологічних процесів -при загоєнні ран [2], трансплантації щелепи [3], експериментальному фіброзі печінки [4] та ін. [5,6]. ...
Background. The world and domestic literature from time to time report about the problems of uncontrolled drug abuse. This problem is associated not only with the negative impact on the morphological structure, but also is a serious factor which, if it has been exposing for a long time, leads to disability and death. Despite a significant number of studies in this area, the problem of changes in the morphometric parameters of the intervertebral disc’s structural components under chronic exposure of opioid agents still remains unresolved. That is why the study of morphometric characteristics of the intervertebral disc’s structural components under experimental opioid exposure will be interesting for both - morphologists and practical traumatologists. Objective. To study the morphometric parameters of the structural components of the intervertebral disc in rats at different times of the experimental opioid exposure and after its withdrawal. Methods. The objects of the study were 61 mature outbred male rats, weight - 80-135g, age - 4.5-7.5 months. Animals were injected with nalbuphine intramuscularly once daily (at 10-11 am) for 42 days. Histological specimens were stained with alcyan blue (for the study of the collagen fibers thickness of the anterior and posterior longitudinal ligaments) and PAS-reaction (for the study of the gelatinous nucleus and its capsule thickness). For measurements of collagen fiber thickness, were taken images at x100 magnification, for measurements of the gelatinous nucleus and its capsule thickness - at x40 magnification. Measurements were performed with ImageJver software. 1.51 using the tool "straight" for linear measurements. Results. Thus, changes in the collagen fibers thickness of the anterior and posterior longitudinal ligaments during the experiment were uneven and had different tendencies. The thickness of the posterior longitudinal fibers changed mostly within the central trend of the control group with a significant decrease only at the 2nd, 3rd, and 4th weeks. However, more significant, as for pathogenetic changes, was a sharp decrease in the number of fibers thicker than 25 μm in animals of the experimental group after 2 weeks of the study, and the maximum thickness at 5th and 6th week was only 31.41 μm and 35.24 μm whereas the maximum thickness of the control group was 81.48 μm. In the withdrawal group, only a decrease in sites with nuclear edema can be considered positive, which is displayed by morphometrical decrease in the maximum value, but the central trend remains much lower than in the control group, which also indicates the predominance of decompensatory changes. Conclusion. During the whole experiment, we observed similar changes in the nucleus capsule thickness: non-systemic fluctuations of the central parameters up to the 5th week and a sharp decompensation at the 6th week, which was manifested by a sharp decrease in thickness. Simultaneously, we observed numerous sites of edema and thickening by more than 100 μm on the 2nd week. Despite a general decrease in median and quartile at this time, such sites were also noted at the 6th week. Thickness of the nucleus capsule in the withdrawal group did not demonstrate significant dynamics and remained similar to those at the 6th week. Although, this can be interpreted as the stabilization at a certain level without further negative dynamics.
... PDGF, hepatocyte growth factor, and fibroblast growth factor are stimuli of cell proliferation, whereas TGF-b seems to inhibit corneal epithelial cell proliferation and enhances apoptosis. 14,15 Generally, these biologic products of PRP are manufactured through a particular kit under good manufacturing practice conditions. When manufacturing a biologic PRP, it is essential to have established specifications to assure safety, purity, and potency. ...
PURPOSE: The objective of this study was to evaluate interleukin-6 (IL-6) tear concentration and clinical outcome in patients with moderate-to-severe bacterial corneal ulcers post autologous platelet-rich plasma (PRP) eye drop therapy.
MATERIALS AND METHODS: This was a pre–post designed study involving 21 moderate-severe corneal ulcer patients who got autologous PRP eye drop. Subjects were got autologous PRP eye drop as adjuvant therapy. Patients with moderate-to-severe infectious bacterial corneal ulcers were included in this study. Tear sampling was performed before therapy using sterile Schirmer paper from conjunctival inferior fornix. PRP therapy was performed for 7 days. Data recording and tear sampling were then performed at day 0 (pre-PRP), day 7 (D+7), and day 14 (D+14) after PRP therapy. Data recording included presence of pericorneal injection, blepharospasm, size of corneal defects, and hypopyon.
RESULTS: There was a decrease in IL-6 tear concentration by day 14 after PRP therapy (P < 0.001). IL-6 concentration at day 7 after therapy (7525.67 ± 7092 pg/mL) tended to be lower before therapy (10,599 ± 6158 pg/mL), but not statistically significant (P = 0.156). The size of corneal defects decreased significantly post PRP at day 7 (P = 0.035) and at day 14 (P = 0.001). There was a significant blepharospasm at day 7 (P = 0.012) and day 14 (P < 0.001). There was a significant pericorneal injection only at day 14 (P = 0.002). There was no significant decreased hypopyon.
CONCLUSION: There was a significant reduction in IL-6 tear concentration and clinical improvement in moderate-to-severe bacterial corneal ulcers which got autologous PRP eye drop as adjuvant therapy.
... Growth factors are proteins which bind to receptors on the cell surface and lead to better cell function and tissue regeneration [102][103][104] for example bone morphogenic proteins (BMPs) are used to induce bone formation [105]. Other growth factors which have shown to induce osteoblast differentiation are transforming growth factor beta (TGF-β), fibroblast growth factors (FGF), platelet-derived growth factors [106,107]. BMP is found to exhibit highest in vitro and in vivo osteogenic potential [108]. Jun et al. [109] proved Silica xerogel-chitosan hybrid coated BMP-2 with porous HA achieved in-vivo bone regeneration in rabbits. ...
Every year, millions of people around the world undergo bone graft and artificial prosthesis transplants which engenders the repair and/or replacement of native bone for treating bone defects and injury. As a result, bone has become the second most transplanted tissue globally and there is a growing need for advances in bone tissue engineering (BTE) to ensure improved quality of life. The advent of 3D bone scaffolds has led to a paradigm shift in this field. These scaffolds act as an extra cellular matrix (ECM) providing a 3D environment for cell adhesion, proliferation, and differentiation. The scaffolds so fabricated, must meet the mechanical and biological criteria for successful clinical translation. Initial iterations of these scaffolds were not constructed with precise geometries. This review looks at the various advances in scaffold designs, materials and 3D printing techniques employed, which enable us to fabricate scaffolds with tailored architecture and control its functionality. These advances allow us to optimize the parameters in the fabrication of these scaffolds, such that we can create structures with desired characteristics like controllable biodegradability using the least amount of material possible along with a precise pore size and high permeability for purposes of osteo-induction and osseointegration. This is followed by the computational analysis carried out on the samples to ensure they are suitable for a given application. Finally, this paper talks about regulatory hurdles (ethical, legal and social) faced by researchers, in addition to the various technical challenges, when it comes to the real-life implementation of this technology.
... | 7 factors, such as PDGF, TGFβ, or SDF1α; cytokines; and other proteins, inside α-granules platelets, which are key components for tissue repair (Anitua et al., 2012;Mussano et al., 2016). These molecules can only be released after breaking the plasma membrane of platelets (i.e., using calcium chloride), a process called activation or degranulation (Fréchette et al., 2005). Further, PRP can be easily obtained via centrifugation to create a gradient in which the lower part of the plasma fraction is enriched in platelets from a peripheral blood sample (Dohan Ehrenfest et al., 2018). ...
Endometrial function is essential for embryo implantation and pregnancy, but managing endometrial thickness that is too thin to support pregnancy or an endometrium of compromised functionality due to intrauterine adhesions is an ongoing challenge in reproductive medicine. Here, we review current and emerging therapeutic and experimental options for endometrial regeneration with a focus on animal models used to study solutions for Asherman's syndrome and endometrial atrophy, which both involve a damaged endometrium. A review of existing literature was performed that confirmed the lack of consensus on endometrial therapeutic options, though promising new alternatives have emerged in recent years (platelet‐rich plasma, exosomes derived from stem cells, bioengineering‐based techniques, endometrial organoids, among others). In the future, basic research using established experimental models of endometrial pathologies (combined with new high‐tech solutions) and human clinical trials with large population sizes are needed to evaluate these emerging and new endometrial therapies.
Numerous studies have been done on the use of biocompatible materials in regenerative medicine. Platelet concentrates, also known as concentrated growth factor, platelet-rich fibrin, and platelet-rich plasma, are the result of centrifuging blood to separate out the platelets. Platelet concentrations have generated a great deal of discussion in both soft and hard tissue engineering. In fact, growth factors, fibrin matrix, and platelets are among the components of autologous platelet concentrate that are essential for the healing of wounds. Modern techniques for tissue restoration by increasing the properties of autologous platelet concentrates are the subject of current research. The usage of platelet concentrates and their role in tissue regeneration are addressed in the current study, along with a number of new advances and its biological effects.
Human knee joint is a complex hinge joint of Saddle variety. Due to the degree of freedom imparted to it, the ligaments that stabilize the knee are subjected to constant stress. This results in increased incidence of ACL injuries particularly during sports. With the advent of Arthroscopy and better understating of ligament injuries with the use of MRI, the ligament reconstruction procedures have increased. The tunnels prepared during the surgery are subjected to constant biological and mechanical stress during post-operative period. The resultant widening leads to failure of graft integration and failure of ACLreconstruction surgery. In recent times biological augments have been used to prevent this tunnel widening. Various studies have been conducted to assess the outcomes of this augments to accelerate the graft healing.
Persistent pneumothorax is a life-threatening complication that can occur after congenital cardiac surgery. Traditional treatment such as chest tube drainage may not be effective in managing this condition. This study presents a new minimally invasive method for treating persistent pneumothorax using platelet-rich plasma–fibrin glue (PRP–FG). The method has been successful in treating postoperative chylothorax in previous studies, and its use has decreased morbidity, mortality, and hospital stay in chylothorax patients. Ten patients with persistent pneumothorax following cardiothoracic surgery (3 TAPVC, 2 d-TGA, 2 VSD + IAA,1 TRUNCUS + TAPVC, 1 VSD + COA, 1 GLENN), who did not respond to conservative management, underwent treatment with PRP–FG. Follow-up was done for a period of 1–4 years. The age and diagnoses of pneumothorax after surgery were 85.5 ± 36.0 days and 62.4 ± 34.3 h, respectively. Persistent pneumothorax of 8 patients (80%) was cured completely after PRP–FG injection. PRP–FG therapy was failed in two patients who died. All cured patients had a normal life without any complications during follow-up. After PRP–FG injection, 3 patients stopped bubbling at one-time injection, 3 patients stopped bubbling at two-time injection, and 2 patients stopped bubbling at three-time injection. Two patients died during treatment; in these cases, one-time injections were done which was not successful. Persistent pneumothorax after congenital–cardiac surgery can be treated successfully with PRP–FG. This bedside minimal-invasive procedure may significantly decrease the morbidity and mortality rate. Further research is needed to confirm the efficacy of this promising treatment through multicentre clinical trials.
Background:
Systemic administration of soluble factors from bone marrow-derived stem cells combined with activated platelet-rich plasma (SC-PRP) restored ovarian function, mediated through paracrine signaling, in murine models of chemotherapy-induced ovarian damage and human tissue from poor responder patients. However, the effects against age-related infertility and the efficacy of local administration have not yet been evaluated.
Objective:
To assess whether a single intraovarian dose of SC-PRP can recover ovarian function, oocyte quality and developmental competence in older mice.
Study design:
The effects of SC-PRP against age-related infertility were assessed following controlled ovarian stimulation, in an aging murine model reproducing three physiological stages of women´s reproductive life: young, advanced maternal age (AMA), and menopausal (n=12 animals per group). Female mice were randomized to receive a single intraovarian injection (10 μL/ovary) of saline, activated platelet-rich plasma (PRP), or SC-PRP. Seven days later, mice were stimulated, naturally mated, and sacrificed to harvest their ovaries for histological assessment and molecular analysis, and oviducts to evaluate oocyte maturation and assess early embryo development.
Results:
A single intraovarian injection of SC-PRP promoted follicle activation and development in young, advanced maternal age, and old mice. Further, SC-PRP rescued fertility in older mice, by enhancing the quantity and quality of ovulated mature oocytes and supporting early embryo development to the blastocyst stage in all the evaluated ages. These fertility outcomes were positively associated with mitochondrial quality, with treatment increasing mitochondrial DNA copy numbers and reducing oxidative damage and apoptosis. Finally, the effects observed by histological analysis were supported at the proteomic level. Functional proteomic analysis revealed molecular mechanisms involved in oocyte maturation and quality, mitochondrial function, and recovery of the ovarian stroma.
Conclusions:
SC-PRP is a promising treatment with the potential to improve the reproductive outcomes of women with age-related infertility, exceeding the restorative effects of PRP alone. Although further research in human ovarian samples is still required, the autologous nature of stem cell factors collected by non-invasive mobilization, their combination with PRP, and the local administration route, suggest that SC-PRP treatment could be a potentially effective and safe application for future clinical practice.
The bone microenvironment is a dynamic, specialized region composed of heterogeneous cells, extracellular matrix, soluble growth factors, and cytokines and where these components interact and produce physiological or pathological phenomena. It is the place where various components interact, function, and produce physiological or pathological phenomena. In this part, we introduce bone microenvironment, discuss its role in bone diseases, and review biomaterials used in bone microenvironment.
BACKGROUND CONTEXT
Platelet-rich plasma (PRP) can accelerate bone union in spinal fusion surgery with an autogenous bone graft. However, it is unclear whether bone union can be obtained by using artificial bone and PRP together in spinal interbody fusion surgery.
PURPOSE
This study aimed to determine whether interbody fusion can be achieved by transplanting porous hydroxyapatite/collagen(HAp/Col) which is an artificial bone material frequently used in spinal fusion surgery, together with PRP in the intervertebral disc space in rats.
STUDY DESIGN AND SETTING
A controlled laboratory study
METHODS
A total of 40 ten-week old Sprague-Dawley rats were used in this study and assigned to three groups as follow: disc curettage only (control group, n=10), disc curettage + HAp/Col transplant (H group, n=10), and disc curettage + HAp/Col + PRP transplant (H+P group, n=10). The other 10 rats were sacrificed as blood donors for acquisition of PRP. Microcomputed tomography (μCT) examinations were performed to evaluate bone union, bone volume (BV) and bone mineral density (BMD) at 4, 8 and 12 weeks following surgery. Twelve weeks postoperatively, each group of three of L4-L5 spines was harvested to perform histological examination (hematoxylin & eosin stain) and the others were subjected to biomechanical testing (compression properties).
RESULTS
The platelet count in PRP was approximately 4.1 times greater than that in whole blood (260.6±26.2 × 10⁴ mg/dL and 64.3±2.9 × 10⁴ mg/dL in PRP and whole blood, respectively). All the L4-L5 lumbar discs were fused in the H+P group, whereas only one case was fused in the H group and none in the control group at 12 weeks after surgery. BV was significantly higher in the H+P group than in the H group or control groups (both P < 0.01), although BMD was not significantly different among the three groups. Upon histological analysis, mature bone formation was observed at the transplanted space in all cases in the H+P group, whereas fibrous tissue was observed at the location in the H and control groups. Regarding biomechanical properties, the ultimate load to failure was significantly higher in the H+P group than in the H group or control group (P = 0.021 and 0.013, respectively), although stiffness was not significantly different between the three groups.
CONCLUSION
The combination of porous HAp/Col and PRP at an appropriate concentration can promote bone union in the intervertebral disc space without using an autologous bone graft in the rat model. Bone tissue formation was histologically confirmed, and it was mechanically strong.
CLINICAL SIGNIFICANCE
This preclinical study showed that porous HAp/Col, when combined with PRP at an appropriate concentration, can induce bone union without autologous bone grafts. The results may eliminate the need for autologous bone collection for spinal fusion surgery in the future.
Human periodontal ligament stem cells (hPDLSCs) are promising for tissue engineering applications but have received relatively little attention. Human platelet lysate (HPL) contains a cocktail of growth factors. To date, there has been no report on hPDLSC seeding on scaffolds loaded with HPL. The objectives of this study were to develop a calcium phosphate cement (CPC)-chitosan scaffold loaded with HPL and investigate their effects on hPDLSC viability, osteogenic differentiation and bone mineral synthesis for the first time. hPDLSCs were harvested from extracted human teeth. Scaffolds were formed by mixing CPC powder with a chitosan solution containing HPL. Four groups were tested: CPC-chitosan + 0% HPL (control); CPC-chitosan + 2.66% HPL; CPC-chitosan + 5.31% HPL; CPC-chitosan + 10.63% HPL. Scanning electron microscopy, live/dead staining, CCK-8, qRT-PCR, alkaline phosphatase and bone minerals assay were applied for hPDLSCs on scaffolds. hPDLSCs attached well on CPC-chitosan scaffold. Adding 10.63% HPL into CPC increased cell proliferation and viability (p < 0.05). ALP gene expression of CPC-chitosan + 10.63% HPL was 7-fold that of 0% HPL at 14 days. Runx2, OSX and Coll1 of CPC-chitosan + 10.63% HPL was 2-3 folds those at 0% HPL (p < 0.05). ALP activity of CPC-chitosan + 10.63% HPL was 2-fold that at 0% HPL (p < 0.05). Bone minerals synthesized by hPDLSCs for CPC-chitosan + 10.63% HPL was 3-fold that at 0% HPL (p < 0.05). This study showed that CPC-chitosan scaffold was a promising carrier for HPL delivery, and HPL in CPC exerted excellent promoting effects on hPDLSCs for bone tissue engineering for the first time. The novel hPDLSC-CPC-chitosan-HPL construct has great potential for orthopedic, dental and maxillofacial regenerative applications.
Objective
In the microenvironment of wound sites, naturally occurring growth factors are crucial for cell migration, opsonisation, chemotaxis, differentiation and angiogenesis. Exogenous growth factors, such as platelet-rich plasma (PRP) and adipose tissue, also improve healing.
Method
In the present within-subject study, we described the effects of PRP and adipose tissue extract (ATE) on skin graft donor site wound healing in patients requiring split-thickness skin grafts. Each patient, having at least two donor sites, received both control (no growth factor) and experimental (PRP or ATE) treatments. Wounds were evaluated on days 5, 7, 10, 15, 30 and 60. Digital photography and spectral images were used to analyse haemoglobin and melanin content, and re-epithelialisation area. Pain was assessed by visual analogue scale. Scar characteristics were scored on days 30 and 60. Biomaterial samples were analysed for growth factor and protein content.
Results
The study included 24 patients (18 male and six female; mean age: 59.1 years). PRP was topically applied to wounds in 11 patients (13 donor sites) and ATE in 13 patients (15 sites). ATE-treated donor sites exhibited significantly accelerated wound re-epithelialisation on days 5 and 7 compared with control sites (p=0.003 and 0.04, respectively). PRP accelerated healing on day 7 compared with control sites (p=0.001). Additionally, the application of ATE improved scar quality on days 30 and 60 (p=0.0005 and 0.02, respectively). Pain scores did not differ significantly between treatments.
Conclusion
In this study, both growth factor sources stimulated wound healing. ATE is an alternative source of growth factors that promote early wound healing and improve scar quality.
Now a days platelet rich plasma is the new approach to the tissue regeneration process along with it platelet rich plasma is found to be of valuable use in promoting healing in various periodontal, dental and oral surgical procedures. Platelet rich plasma is basically derived from the own blood of the patient, that is containing growth factors with in it, which promotes the healing process in periodontal or other dental procedures.
Background
Platelet-rich plasma (PRP) is reported as an effective treatment for lateral epicondylitis (LE). Theoretically, different types of PRP have different therapeutic effects. However, there is controversy on the effects of different types of PRP in the treatment of LE.
Purpose
To systematically compare the pain relief, functional improvement and successful rates on treatment of two different types of PRP, by reviewing and summarizing the data available in the current literature on LE after PRP injection.
Methods
The PubMed, Medline, Embase, Cochrane Library and Web of science were reviewed. A computerized literature search was performed for related studies published from inception to August 2021 by terms of lateral epicondylitis, tennis elbow, tendinopathy, lateral elbow pain, PRP. PRP involved in present study were divided into leukocyte-poor PRP and leukocyte-rich PRP groups according to different preparation methods. Outcomes of interest included characteristics of the subjects, types and preparations of PRP, clinical outcomes, successful rate and safety of treatment of short-term and long-term follow-up.
Results
A total of 33 studies included 2420 LE patients. There were 19 studies with LP-PRP, 13 studies with LR-PRP and 1 study involved both LP-PRP and LR-PRP. Patients had significant improved clinical outcomes post-treatment compared to pre-treatment in both groups of PRP. The mean of VAS was ranged from 6.1 to 8.0 before the treatment, 1.5 to 4.0 at short-term and 0.6 to 3.3 at the long-term follow-up in LR-PRP group. The mean of VAS was ranged from 4.2 to 8.4 before the treatment, 1.6 to 5.9 at short-term and 0.7 to 2.7 in the long-term follow-up in LP-PRP group. The DASH score of LR-PRP and LP-PRP were ranged from 47.0 to 54.3 and 30.0 to 67.7 separately before the treatment and 20.0 to 22.0 and 5.5 to 19.0 separately at long-term follow-up. LR-PRP and LP-PRP groups reflected successful rate ranged from 70%-100% and 36%-100% respectively. The complication rate lower in LP-PRP group (3.9%) than LR-PRP group (6.4%), with the major complication was temporary pain after PRP treatment (P = 0.029).
Conclusion
PRP treatment demonstrated a significant improvement with pain relief and functional improvement on lateral epicondylitis regardless types of PRP. There was no significant difference between LR-PRP and LP-PRP in pain relief and functional improvement. The major complication was temporary pain after PRP injection and the complication rate in LP-PRP was lower than LR-PRP.
Nowadays, many patients facing head and neck oncological surgery have a history of tissue irradiation. This represents an important risk factor for postsurgical complications, including dehiscences and fistulas. Platelet-rich plasma (PRP) obtained from the patient’s blood represents an easy, fast and inexpensive method for the prevention and treatment of such complications. We present three cases of previously irradiated patients in which PRP was successfully used to prevent and treat postsurgical complications.
Renal ischemia-reperfusion (I-R) injury is the main cause of acute renal failure. Acute pancreatitis is one of the fatal remote lesions that occurs in patients with renal I-R injury. Platelet-rich plasma (PRP) is a hopeful aiding therapy for tissue injuries. Forty adult rats were utilized in this study, ten for PRP preparation and thirty were divided into three groups; Control: subdivided into three equal subgroups, I-R group: exposed to bilateral renal pedicles clamping and I-R+ PRP group: were experienced identical procedures as I-R group then subcutaneously (S.C) injected with 0.5 ml PRP two times weekly for three weeks. Blood samples were taken for detection of serum urea and creatinine, blood glucose level and serum amylase. The pancreas was dissected and prepared for histopathological examination by light and electron microscope. Statistical analysis of all collected results was performed. Our biochemical results revealed deleterious effects of renal I-R on the pancreas as evidenced by a highly significant increase in serum amylase and blood glucose level. In I-R group, histopathological examination revealed wide septa and congested blood vessels, acinar cells showed disrupted rough endoplasmic reticulum and few secretory granules. Some islet cells showed pyknotic nuclei and vacuolated cytoplasm. PRP treated group showed nearly normal structure in the form of numerous acinar cells' granules, extensive rough endoplasmic reticulum and mitochondria. Most of Beta cells had euchromatic nuclei and numerous secretory granules. Accordingly, PRP treatment reduced the pancreatic biochemical and histopathological alterations caused by renal I-R injury and so considered a promising therapy for pancreatic damage.
Wound healing and tissue regeneration are a field of clinical medicine presenting high research interest, since various local and systematic factors can inhibit these processes and lead to an inferior result. New methods of healing enhancement constantly arise, which, however, require experimental validation before their establishment in everyday practice. Platelet-rich plasma (PRP) is a well-known autologous factor that promotes tissue healing in various surgical defects. PRP derives from the centrifugation of peripheral blood and has a high concentration of growth factors that promote healing. Recently, the use of adipose-derived mesenchymal stem cells (ADMSCs) has been thoroughly investigated as a form of wound healing enhancement. ADMSCs are autologous stem cells deriving from fat tissue, with a capability of differentiation in specific cells, depending on the micro-environment that they are exposed to. The aim of the present comprehensive review is to record the experimental studies that have been published and investigate the synergistic use of PRP and ADMSC in animal models. The technical aspects of experimentations, as well as the major results of each study, are discussed. In addition, the limited clinical studies including humans are also reported. Future perspectives are discussed, along with the limitations of current studies on the long-term follow up needed on efficacy and safety.
Wound healing is a complex phenomenon that involves different cell types with various functions, i.e., keratinocytes, fibroblasts, and endothelial cells, all influenced by the action of soluble mediators and rearrangement of the extracellular matrix (ECM). Physiological angiogenesis occurs in the granulation tissue during wound healing to allow oxygen and nutrient supply and waste product removal. Angiogenesis output comes from a balance between pro- and antiangiogenic factors, which is finely regulated in a spatial and time-dependent manner, in order to avoid insufficient or excessive nonreparative neovascularization. The understanding of the factors and mechanisms that control angiogenesis and their change following unloading conditions (in a real or simulated space environment) will allow to optimize the tissue response in case of traumatic injury or medical intervention. The potential countermeasures under development to optimize the reparative angiogenesis that contributes to tissue healing on Earth will be discussed in relation to their exploitability in space.
Peripheral arterial disease can involve tissue loss in up to 50% of patients with diabetic foot syndrome (DFS). Consequently, revascularization of narrowed or occluded arteries is one of the most common forms of comprehensive treatment. However, technically successful angioplasty does not always result in the healing of ulcers. The pathomechanism of this phenomenon is still not fully understood, but inadequate angiogenesis in tissue repair may play an essential role. Changes in pro- and anti-angiogenic factors among patients with DFS are not always clear and conclusive. In particular, some studies underline the role of decreased concentration of pro-angiogenic factors and higher levels of anti-angiogenic mediators. Nevertheless, there are still controversial issues, including the paradox of impaired wound healing despite high concentrations of some pro-angiogenic factors, dynamics of their expression during the healing process, and their mutual relationships. Exploring this process among diabetic patients may provide new insight into well-known methods of treatment and show their real benefits and chances for improving outcomes.
Remodeling of the human bony skeleton is constantly occurring with up to 10% annual bone volume turnover from osteoclastic and osteoblastic activity. A shift toward resorption can result in osteoporosis and pathologic fractures, while a shift toward deposition is required after traumatic, or surgical injury. Spinal fusion represents one such state, requiring a substantial regenerative response to immobilize adjacent vertebrae through bony union. Autologous bone grafts were used extensively prior to the advent of advanced therapeutics incorporating exogenous growth factors and biomaterials. Besides cost constraints, these applications have demonstrated patient safety concerns. This study evaluated the regenerative ability of a nanostructured, magnesium-doped, hydroxyapatite/type I collagen scaffold (MHA/Coll) augmented by autologous platelet-rich plasma (PRP) in an orthotopic model of posterolateral lumbar spinal fusion. After bilateral decortication, rabbits received either the scaffold alone (Group 1) or scaffold with PRP (Group 2) to the anatomic right side. Bone regeneration and fusion success compared to internal control were assessed by DynaCT with 3-D reconstruction at 2, 4, and 6 weeks postoperatively followed by comparative osteogenic gene expression and representative histopathology. Both groups formed significantly more new bone volume than control, and Group 2 subjects produced significantly more trabecular and cortical bone than Group 1 subjects. Successful fusion was seen in one Group 1 animal (12.5%) and 6/8 Group 2 animals (75%). This enhanced effect by autologous PRP treatment appears to occur via astounding upregulation of key osteogenic genes. Both groups demonstrated significant gene upregulation compared to vertebral bone controls for all genes. Group 1 averaged 2.21-fold upregulation of RUNX2 gene, 3.20-fold upregulation of SPARC gene, and 3.67-fold upregulation of SPP1 gene. Depending on anatomical subgroup (cranial, mid, caudal scaffold portions), Group 2 had significantly higher average expression of all genes than both control and Group 1–RUNX2 (8.23–19.74 fold), SPARC (18.67–55.44 fold), and SPP1 (46.09–90.65 fold). Our data collectively demonstrate the osteoinductive nature of a nanostructured MHA/Coll scaffold, a beneficial effect of augmentation with autologous PRP, and an ability to achieve clinical fusion when applied together in an orthotopic model. This has implications both for future study and biomedical innovation of bone-forming therapeutics.
Platelets contain mitogenic activities for MCF-7 human breast cancer cells when assayed under serum-free chemically defined conditions. Purification from outdated human platelets identified insulinlike growth factor I (IGF-I) as the most potent breast cancer cell mitogen in lysates (Karey KP, Sirbasku DA: see accompanying article, this issue). In this study the release and subcellular localization of IGF-I was investigated. Degranulation of platelets by thrombin treatment caused release of lysosomal enzymes (beta-glucuronidase and N-acetyl-D- glucosaminidase), alpha-granule proteins (beta-thromboglobulin and fibrinogen) as well as mitogenic activity for MCF-7 cells and IGF-I as measured by radioimmunoassay (RIA) and radioreceptor assay. Release of mitogenic activity and immunologically identified IGF-I was induced tenfold over controls by thrombin and was nearly complete as compared to platelets disrupted by repeated freezing and thawing. Disruption of platelets by nitrogen cavitation followed by separation of the organelles by sucrose density gradient sedimentation showed that IGF-I and mitogenic activity localized predominantly to fractions containing alpha-granules rather than soluble cellular components, lysosomes, or dense granules. The morphology of MCF-7 cells in serum-free medium supplemented with supernatants from thrombin-treated platelets also indicated the release of important cell-adhesion factors for human breast cancer cells.
Thrombin cleaves its G-protein-linked seven-transmembrane domain receptor, thereby releasing a 41-aa peptide and generating a new amino terminus that acts as a tethered ligand for the receptor. Peptides corresponding to the new amino terminal end of the proteolyzed seven-transmembrane domain thrombin receptor [TR42-55, SFLLRNPNDKYEPF, also known as TRAP (thrombin receptor-activating peptide)], previously have been demonstrated to activate the receptor. In this study, we demonstrate that the 41-aa cleaved peptide, TR1-41 (MGPRRLLLVAACFSLCGPLLSARTRARRPESKATNATLDPR) is a strong platelet agonist. TR1-41 induces platelet aggregation. In whole-blood flow cytometric studies, TR1-41 was shown to be more potent than TR42-55 and almost as potent as thrombin, as determined by the degree of increase in: (i) platelet surface expression of P-selectin (reflecting alpha granule secretion); (ii) exposure of the fibrinogen binding site on the glycoprotein (GP) IIb-IIIa complex; and (iii) fibrinogen binding to the activated GPIIb-IIIa complex. As determined by experiments with inhibitors [prostaglandin I2, staurosporine, wortmannin, the endothelium-derived relaxing factor congener S-nitroso-N-acetylcysteine (SNAC), EDTA, EGTA, and genestein], and with Bernard-Soulier or Glanzmann's platelets, we demonstrated that TR1-41-induced platelet activation is: (i) inhibited by cyclic AMP; (ii) mediated by protein kinase C, phosphatidyl inositol-3-kinase, myosin light chain kinase, and intracellular protein tyrosine kinases; (iii) dependent on extracellular calcium; and (iv) independent of the GPIb-IX and GPIIb-IIIa complexes. TR1-41-induced platelet activation was synergistic with TR42-55. In summary, the cleaved peptide of the seven-transmembrane domain TR (TR1-41) is a strong platelet agonist.
Platelets contain mitogenic activities for MCF-7 human breast cancer cells when assayed under serum-free chemically defined conditions. Purification from outdated human platelets identified insulinlike growth factor I (IGF-I) as the most potent breast cancer cell mitogen in lysates (Karey KP, Sirbasku DA: see accompanying article, this issue). In this study the release and subcellular localization of IGF-I was investigated. Degranulation of platelets by thrombin treatment caused release of lysosomal enzymes (beta-glucuronidase and N-acetyl-D-glucosaminidase), alpha-granule proteins (beta-thromboglobulin and fibrinogen) as well as mitogenic activity for MCF-7 cells and IGF-I as measured by radioimmunoassay (RIA) and radioreceptor assay. Release of mitogenic activity and immunologically identified IGF-I was induced tenfold over controls by thrombin and was nearly complete as compared to platelets disrupted by repeated freezing and thawing. Disruption of platelets by nitrogen cavitation followed by separation of the organelles by sucrose density gradient sedimentation showed that IGF-I and mitogenic activity localized predominantly to fractions containing alpha-granules rather than soluble cellular components, lysosomes, or dense granules. The morphology of MCF-7 cells in serum-free medium supplemented with supernatants from thrombin-treated platelets also indicated the release of important cell-adhesion factors for human breast cancer cells.
Platelet aggregation is a cardinal feature of both vascular repair and vascular disease. During aggregation platelets release a variety of vasoactive substances; some of these promote angiogenesis, endothelial permeability, and endothelial growth, actions shared by vascular endothelial growth factor (VEGF). This study was undertaken to investigate the hypothesis that VEGF is released by aggregating platelets. We found that VEGF was secreted during the in vitro aggregation of platelet-rich plasma induced by thrombin, collagen, epinephrine, and ADP (range 23-518 pg VEGF/ml). Furthermore, serum VEGF levels were elevated compared with plasma (230 +/- 63 vs. 38 +/- 8 pg VEGF/ml), indicative of VEGF release during whole blood coagulation. Lysates of apheresed, leukocyte-poor platelet units contained significant amounts of VEGF (2.4 +/- 0.8 pg VEGF/mg protein). VEGF message and protein were also present in a megakaryocytic cell line (Dami cell). These results suggest constitutive roles for platelet VEGF in the repair of intimal vessel injury and in the altered permeability and intimal proliferation seen at sites of platelet aggregation and thrombosis.
Platelet activation leads to secretion of granule contents and to the formation of microvesicles by shedding of membranes from the cell surface. Recently, we have described small internal vesicles in multivesicular bodies (MVBs) and alpha-granules, and suggested that these vesicles are secreted during platelet activation, analogous to the secretion of vesicles termed exosomes by other cell types. In the present study we report that two different types of membrane vesicles are released after stimulation of platelets with thrombin receptor agonist peptide SFLLRN (TRAP) or alpha-thrombin: microvesicles of 100 nm to 1 microm, and exosomes measuring 40 to 100 nm in diameter, similar in size as the internal vesicles in MVBs and alpha-granules. Microvesicles could be detected by flow cytometry but not the exosomes, probably because of the small size of the latter. Western blot analysis showed that isolated exosomes were selectively enriched in the tetraspan protein CD63. Whole-mount immuno-electron microscopy (IEM) confirmed this observation. Membrane proteins such as the integrin chains alpha(IIb)-beta(3) and beta(1), GPIbalpha, and P-selectin were predominantly present on the microvesicles. IEM of platelet aggregates showed CD63(+) internal vesicles in fusion profiles of MVBs, and in the extracellular space between platelet extensions. Annexin-V binding was mainly restricted to the microvesicles and to a low extent to exosomes. Binding of factor X and prothrombin was observed to the microvesicles but not to exosomes. These observations and the selective presence of CD63 suggest that released platelet exosomes may have an extracellular function other than the procoagulant activity, attributed to platelet microvesicles.
Platelet-rich plasma is an autologous product that is derived from whole blood through the process of gradient density centrifugation. The proposed value of this product in dental implantology and in bone augmentation procedures lies in the ability to incorporate high concentrations of the growth factors PDGF TGF-beta1, TGF-beta2, and IGF as well as fibrin, into the graft mixture. Research has shown an increased bone maturation rate and improved bone density when this product, or its recombinant growth factors, is added to small bony defects or to larger defects that use autogenous bone as the grafting material. This study tested the efficacy of platelet-rich plasma in three bilateral sinus graft cases with grafts of anorganic bovine bone that contained minimal or no autogenous bone. Histomorphometric analysis indicated that the addition of platelet-rich plasma to the grafts did not make a significant difference either in vital bone production or in interfacial bone contact on the test implants.
Acute wounds normally heal in a very orderly and efficient manner characterized by four distinct, but overlapping phases: hemostasis, inflammation, proliferation and remodeling. Specific biological markers characterize healing of acute wounds. Likewise, unique biologic markers also characterize pathologic responses resulting in fibrosis and chronic non-healing ulcers. This review describes the major biological processes associated with both normal and pathologic healing. The normal healing response begins the moment the tissue is injured. As the blood components spill into the site of injury, the platelets come into contact with exposed collagen and other elements of the extracellular matrix. This contact triggers the platelets to release clotting factors as well as essential growth factors and cytokines such as platelet-derived growth factor (PDGF) and transforming growth factor beta (TGF-beta). Following hemostasis, the neutrophils then enter the wound site and begin the critical task of phagocytosis to remove foreign materials, bacteria and damaged tissue. As part of this inflammatory phase, the macrophages appear and continue the process of phagocytosis as well as releasing more PDGF and TGF beta. Once the wound site is cleaned out, fibroblasts migrate in to begin the proliferative phase and deposit new extracellular matrix. The new collagen matrix then becomes cross-linked and organized during the final remodeling phase. In order for this efficient and highly controlled repair process to take place, there are numerous cell-signaling events that are required. In pathologic conditions such as non-healing pressure ulcers, this efficient and orderly process is lost and the ulcers are locked into a state of chronic inflammation characterized by abundant neutrophil infiltration with associated reactive oxygen species and destructive enzymes. Healing proceeds only after the inflammation is controlled. On the opposite end of the spectrum, fibrosis is characterized by excessive matrix deposition and reduced remodeling. Often fibrotic lesions are associated with increased densities of mast cells. By understanding the functional relationships of these biological processes of normal compared to abnormal wound healing, hopefully new strategies can be designed to treat the pathological conditions.
Platelet activation leads to secretion of granule contents and to the formation of microvesicles by shedding of membranes from the cell surface. Recently, we have described small internal vesicles in multivesicular bodies (MVBs) and -granules, and suggested that these vesicles are secreted during platelet activation, analogous to the secretion of vesicles termed exosomes by other cell types. In the present study we report that two different types of membrane vesicles are released after stimulation of platelets with thrombin receptor agonist peptide SFLLRN (TRAP) or -thrombin: microvesicles of 100 nm to 1 μm, and exosomes measuring 40 to 100 nm in diameter, similar in size as the internal vesicles in MVBs and -granules. Microvesicles could be detected by flow cytometry but not the exosomes, probably because of the small size of the latter. Western blot analysis showed that isolated exosomes were selectively enriched in the tetraspan protein CD63. Whole-mount immuno-electron microscopy (IEM) confirmed this observation. Membrane proteins such as the integrin chains IIb-β3 and β1, GPIb, and P-selectin were predominantly present on the microvesicles. IEM of platelet aggregates showed CD63+ internal vesicles in fusion profiles of MVBs, and in the extracellular space between platelet extensions. Annexin-V binding was mainly restricted to the microvesicles and to a low extent to exosomes. Binding of factor X and prothrombin was observed to the microvesicles but not to exosomes. These observations and the selective presence of CD63 suggest that released platelet exosomes may have an extracellular function other than the procoagulant activity, attributed to platelet microvesicles.
Werner, Sabine, and Richard Grose. Regulation of Wound Healing by Growth Factors and Cytokines. Physiol Rev 83: 835–870, 2003; 10.1152/physrev.00032.2002.—Cutaneous wound healing is a complex process involving blood clotting, inflammation, new tissue formation, and finally tissue remodeling. It is well described at the histological level, but the genes that regulate skin repair have only partially been identified. Many experimental and clinical studies have demonstrated varied, but in most cases beneficial, effects of exogenous growth factors on the healing process. However, the roles played by endogenous growth factors have remained largely unclear. Initial approaches at addressing this question focused on the expression analysis of various growth factors, cytokines, and their receptors in different wound models, with first functional data being obtained by applying neutralizing antibodies to wounds. During the past few years, the availability of genetically modified mice has allowed elucidation of the function of various genes in the healing process, and these studies have shed light onto the role of growth factors, cytokines, and their downstream effectors in wound repair. This review summarizes the results of expression studies that have been performed in rodents, pigs, and humans to localize growth factors and their receptors in skin wounds. Most importantly, we also report on genetic studies addressing the functions of endogenous growth factors in the wound repair process.
Platelet-rich plasma is an autologous source of platelet-derived growth factor and transforming growth factor beta that is obtained by sequestering and concentrating platelets by gradient density centrifugation. This technique produced a concentration of human platelets of 338% and identified platelet-derived growth factor and transforming growth factor beta within them. Monoclonal antibody assessment of cancellous cellular marrow grafts demonstrated cells that were capable of responding to the growth factors by bearing cell membrane receptors. The additional amounts of these growth factors obtained by adding platelet-rich plasma to grafts evidenced a radiographic maturation rate 1.62 to 2.16 times that of grafts without platelet-rich plasma. As assessed by histomorphometry, there was also a greater bone density in grafts in which platelet-rich plasma was added (74.0% ± 11%) than in grafts in which platelet-rich plasma was not added (55.1% ± 8%; p = 0.005).
Platelets contain a vast number of biologically active molecules within cytoplasmic granules which are classified according to their respective distinct ultrastructures, densities and content. The α-granule is a unique secretory organelle in that it exhibits further compartmentalization and acquires its protein content via two distinct mechanisms: (1) biosynthesis predominantly at the megakaryocyte (MK) level (with some vestigial platelet synthesis) (e.g. platelet factor 4) and (2) endocytosis and pinocytosis at both the MK and circulating platelet levels (e.g. fibrinogen (Fg) and IgG). The currently known list of α-granular proteins continues to enlarge and includes many adhesive proteins (e.g. Fg, von Willebrand factor (vWf) and thrombospondin (TSP)), plasma proteins (e.g. IgG and albumin), cellular mitogens (e.g. platelet derived growth factor and TGFβ), coagulation factors (e.g. factor V) and protease inhibitors (e.g. α2-macroglobulin and α2-antiplasmin). More recently the inner lining of the α-granule unit membrane has been demonstrated to contain a number of physiologically important receptors including glycoprotein IIb/IIIa (αIIbβ3) and P-selectin. The α-granules originate from small precursor granules which can be observed budding from the trans-Golgi network within the platelet precursor cell, the MK. During MK maturation the α-granules become very prominent and are ultimately packaged into platelets during thrombopoiesis. The α-granular contents are destined for release during platelet activation at sites of vessel wall injury and thus play an important role in haemostasis, inflammation, ultimate wound repair and in the pathogenesis of atherosclerosis.
Epidermal growth factor (EGF) is known to be present in the alpha granules of human platelets; however the source of this EGF, ie, whether it is taken up by the platelets from the circulation, or whether it is packaged into the platelets from the megakaryocyte during thrombopoiesis, is unknown. To determine whether EGF is taken up by platelets, platelets for EGF receptors were assayed and it was attempted to detect uptake of EGF by the platelets from culture medium. Platelets were found to lack EGF receptors, and no uptake of EGF from the culture medium was detected. To assess whether EGF is packaged into platelets from the megakaryocyte, megakaryocytes in frozen section bone marrow cores were stained for EGF protein by immunohistochemistry, and it was demonstrated that EGF is present in megakaryocytes. In addition, staining of megakaryocytes by in situ hybridization for EGF mRNA demonstrated its presence in these cells. Therefore it is concluded that the source of EGF in human platelets is the megakaryocyte and that this EGF is synthesized in the megakaryocyte rather than being taken up from its environment.
Human epidermal growth factor (hEGF) has previously been isolated from urine and probably is identical to human beta-urogastrone (hUG). Immunoreactive hEGF/UG has been found in the plasma of normal subjects. In this study, using immunoaffinity chromatography to extract hEGF/UG from plasma, we found that immunoreactive hEGF/UG in blood was associated with blood platelets. It was present in platelet-rich, but not platelet-poor plasma and serum, and was found predominantly in the platelet fraction of whole blood. Sephadex G-50 Fine gel-exclusion chromatography of an extract of outdated blood bank platelets revealed two hEGF/UG components, one of which eluted in the void volume, and the other of which coeluted with purified standard hEGF/UG. The former hEGF/UG component was a high-molecular weight form that was cleaved into hEGF/UG by incubation with either mouse EGF/UG-associated arginine esterase or trypsin. It appeared to be identical to the high-molecular weight hEGF/UG previously reported in human urine, except for its apparently equal activities in radioimmunoassay and radioreceptor assay. The latter hEGF/UG component was immunologically, biologically, and physiochemically indistinguishable from highly purified hEGF/UG from human urine and was immunologically different from purified human platelet-derived growth factor. Platelet-associated hEGF/UG may account for the mitogenic activity of serum in cell lines in which platelet-derived growth factor is not active. Since hEGF/UG appears to be liberated from platelets during coagulation, platelet-associated EGF/UG may be involved in normal vascular and tissue repair and in the pathogenesis of atherosclerotic lesions. The discovery that the EGF/UG in plasma is associated with blood platelets raises important new possibilities for its role in human health and disease.
Platelets contain a vast number of biologically active molecules within cytoplasmic granules which are classified according to their respective distinct ultrastructures, densities and content. The alpha-granule is a unique secretory organelle in that it exhibits further compartmentalization and acquires its protein content via two distinct mechanisms: (1) biosynthesis predominantly at the megakaryocyte (MK) level (with some vestigial platelet synthesis) (e.g. platelet factor 4) and (2) endocytosis and pinocytosis at both the MK and circulating platelet levels (e.g. fibrinogen (Fg) and IgG). The currently known list of alpha-granular proteins continues to enlarge and includes many adhesive proteins (e.g. Fg, von Willebrand factor (vWf) and thrombospodin (TSP)), plasma proteins (e.g. IgG and albumin), cellular mitogens (e.g. platelet derived growth factor and TGF beta), coagulation factors (e.g. factor V) and protease inhibitors (e.g. alpha 2-macroglobulin and alpha 2-antiplasmin). More recently the inner lining of the alpha-granule unit membrane has been demonstrated to contain a number of physiologically important receptors including glycoprotein IIb/IIIa (alpha IIb beta 3) and P-selectin. The alpha-granules originate from small precursor granules which can be observed budding from the trans-Golgi network within the platelet precursor cell the MK. During MK maturation the alpha-granules become very prominent and are ultimately packaged into platelets during thrombopoiesis. The alpha-granular contents are destined for release during platelet activation at sites of vessel wall injury and thus play an important role in haemostasis, inflammation, ultimate wound repair and in the pathogenesis of atherosclerosis.
Thrombin interacts with its receptor and thrombomodulin on endothelial cells. We evaluated the respective roles of these two proteins on human umbilical vein endothelial cell (HUVEC) growth by comparing thrombin, S195A (a mutant thrombin in which the serine of the charge stabilizing system had been replaced by alanine), and the receptor activating peptide (TRAP). Thrombin and TRAP induced DNA synthesis (half maximal cell proliferation with 5 nM and 25 microM, respectively), whereas S195A thrombin was inactive, inferring that growth is mediated through the thrombin receptor. Surprisingly, cells stimulated by TRAP exhibited a maximal proliferation twice greater than that obtained with thrombin. Combination of thrombin and TRAP resulted in a mitogenic response higher than by thrombin alone, but lower than by TRAP alone. The role of thrombomodulin was evaluated by adding an anti-thrombomodulin antibody, which prevents formation of the thrombin-thrombomodulin complex. Antibody did not interfere with cell proliferation induced by TRAP, but enhanced that induced by thrombin. We conclude that formation of the thrombin-thrombomodulin complex restrains HUVEC proliferation mediated through the thrombin receptor.
Platelet-rich plasma is an autologous source of platelet-derived growth factor and transforming growth factor beta that is obtained by sequestering and concentrating platelets by gradient density centrifugation. This technique produced a concentration of human platelets of 338% and identified platelet-derived growth factor and transforming growth factor beta within them. Monoclonal antibody assessment of cancellous cellular marrow grafts demonstrated cells that were capable of responding to the growth factors by bearing cell membrane receptors. The additional amounts of these growth factors obtained by adding platelet-rich plasma to grafts evidenced a radiographic maturation rate 1.62 to 2.16 times that of grafts without platelet-rich plasma. As assessed by histomorphometry, there was also a greater bone density in grafts in which platelet-rich plasma was added (74.0% +/- 11%) than in grafts in which platelet-rich plasma was not added (55.1% +/- 8%; p = 0.005).
Blood platelets contain angiopoietin-1, a growth factor essential for blood vessel development via stabilization of proliferating endothelial cells. It has recently been reported that angiopoietin-1 can act as a vascular stability factor (Nature Medicine 6:460, 2000). In investigating the normal tissue distribution of angiopoietin-1 from surgically-removed frozen specimens by RT-PCR, we found it consistently present in platelets and megakaryocytes, usually absent in relatively non-vascular tissue: breast, colon, lung, skin, kidney, thyroid, testicle, cervix and occasionally present in tissue enriched with vasculature: prostate, endometrium, ovary, under conditions in which mRNA stability was verified by the positive detection of internal control, actin mRNA. The consistent distribution in platelets and relatively absent distribution in non-vascular normal tissue suggested that the well-known role of platelets in maintaining vascular stability, may in part be due to platelet release of angiopoietin-1 following platelet activation. In this communication we report the incidence of Ang-1 in various normal tissues and demonstrate that thrombin-treated human platelets release angiopoietin-1 in vitro.
The role of thrombin in the stimulation of endothelial cell (EC) proliferation is controversial. The aim of this study was to investigate if thrombin regulates cell proliferation and production of platelet-derived growth factor (PDGF), bovine fibroblast growth factor (bFGF), and transforming growth factor beta(1) (TGF-beta(1)) by bovine aortic ECs.
ECs, obtained from thoracic aortas of calves, were stimulated with thrombin at various concentrations (from 0.05 to 1.0 IU/ml) in serum free culture. Mitogenic activity of thrombin on ECs was determined by tritiated thymidine uptake. The release of PDGF, bFGF, and TGF-beta(1) was assessed by ELISA. PDGF release was confirmed by Western blot and bFGF and TGF-beta(1) mRNA expression was determined by polymerase chain reaction (PCR).
Thrombin at high concentrations did not cause any increase in EC proliferation after 72 h of culture and induced inhibition of EC proliferation after 96 h and 8 days of culture. It induced a decrease in PDGF release and an increase in TGF-beta(1) release. Thrombin at low concentrations induced a significant increase in EC proliferation at 72 h, 96 h, and 8 days of culture. It induced an increase in PDGF release and a decrease in TGF-beta(1) release. bFGF release was higher than control at all thrombin concentrations. These data were confirmed by Western blot and PCR studies.
Thrombin regulates EC growth through the inhibition of EC proliferation at high concentrations and through the stimulation of EC proliferation at low physiological concentrations. EC proliferation is partially mediated by autocrine production of PDGF, bFGF, and TGF-beta(1).
To evaluate the effect of particulate dentin-plaster of Paris with and without platelet-rich plasma (PRP) on bone healing and new bone formation around titanium dental implants in a canine model. Histologic sections and histomorphometric analysis of the defects were obtained at 6 and 12 weeks after surgery.
Three circular bone defects were surgically prepared in iliac crest sites in each of 10 animals. A total of 30 Avana dental implants were placed in the animals. They were self-tapping, screw-type implants, 10 mm in length and 4 mm in diameter, all made of commercially pure titanium. A titanium implant was placed centrally in each defect. In each dog, the defects were treated with 1 of the following 3 treatment modalities: (1) no treatment (control); (2) grafting with particulate dentin-plaster of Paris; (3) grafting with particulate dentin-plaster of Paris and PRP.
Histologic analysis showed that all of the bone defects surrounding the implants that were treated with particulate dentin-plaster of Paris, with and without PRP, were filled with new bone. The defects that were not treated (control) demonstrated new bone formation only in the inferior threaded portion of the implants.
Histomorphometric results revealed a higher percentage of bone contact with particulate dentin-plaster of Paris and PRP compared to the control and particulate dentin-plaster of Paris.
These results suggested that bone defects around titanium implants can be treated successfully with particulate dentin-plaster of Paris, and that the outcome can be improved if PRP is also used.
Peripheral leukocytes and platelets (LAPs) contain many kinds of cells with the ability to secrete several growth factors and cytokines. We attempted to induce therapeutic angiogenesis by injecting self-LAPs into a rat ischemic hindlimb model.
Supernatants from cultured LAPs were used for the endothelial cell (EC) proliferation assay, and LAPs were used in a cornea model to evaluate angiogenic potency. LAPs were injected directly into the male Dark Agouti rat ischemic hindlimb model, after which a microangiogram was done and the capillary/muscle fiber ratio was examined histologically. ELISA revealed the levels of contributing growth factors and cytokines present in the ischemic muscles.
The EC proliferation assay showed that the supernatants of LAPs accelerated proliferation and that the LAPs induced angiogenesis in the cornea model. The microangiograms and histological evaluation revealed that angiogenesis was induced more effectively in the rats injected with LAPs (LAP group) than in the those injected with phosphate-buffered saline (PBS group). The levels of basic fibroblast growth factor (bFGF) in the ischemia, PBS, and LAP groups were significantly increased compared to those in the sham group. The level of interleukin-1beta (IL-1beta) in the LAP group was significantly more elevated than in the other groups.
The injection of self-LAPs induced angiogenesis in a rat ischemic hindlimb model. Ischemia caused an elevation in the level of bFGF and also in IL-1beta derived from LAPs, which contributed to angiogenesis. This is a novel, yet simple and safe method of inducing therapeutic angiogenesis.
Inflammatory cytokines in platelet concentrates (PC) may cause side-effects such as febrile non-haemolytic transfusion reactions. The maximum white blood cell (WBC) content tolerable to avoid the accumulation of cytokines, and whether these cytokines originate from degranulating leucocytes or de novo synthesis during storage, had not been investigated prior to this study.
We investigated the secretion of interleukin (IL)-1beta, IL-2, IL-6, IL-8, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) and quantified the appropriate expression of corresponding mRNA in PC with regard to different levels of WBC contamination and storage times. In addition we tested the viability of WBCs during PC storage (by staining with 7-aminoactinomycin D) and their ability to perform de novo cytokine synthesis (by using superantigen stimulation).
We detected a statistically significant increase of IL-1beta, IL-6, IL-8 and TNF-alpha in PC with > or = 108 WBCs. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) showed increasing mRNA expression of the respective cytokines depending on the number of WBC present. On day 5 of storage, WBC viability was > 80% and the leucocytes were still able to produce cytokines de novo.
These data show clear evidence for de novo synthesis of cytokines in PC. The cytokine pattern supports the hypothesis that activated monocytes are responsible for this cytokine synthesis. PC with a WBC contamination of > or = 108 contain inflammatory mediators in clinically relevant concentrations.
Platelet-rich plasma contains autologous thrombocyte growth factors and might be promising for acceleration of dentoalveolar bone regeneration. In this study, it was analysed for platelet counts and growth factor concentrations.
Platelet-rich plasma was isolated by discontinuous cell separation from 158 healthy men and 55 women aged 17-62 years. One hundred and fifteen specimens (stratified for age and gender of the donor) were analysed for growth factor concentrations and platelet count.
The platelet count in platelet-rich plasma (1,407,640+/-320,100/microl) was 5 times higher than in donor blood (266,040+/-60,530/microl). Platelet-derived growth factor AB (117+/-63 ng/ml), transforming growth factor (TGF) beta -1 (169+/-84 ng/ml), and insulin-like growth factor (IGF) I (84+/-23 ng/ml) were found in large amounts, while platelet-derived growth factor (PDGF) BB (10+/-8 ng/ml) and transforming growth factor beta -2 (0.4+/-0.3 ng/ml) were found in small amounts only. The growth factor content was not well correlated with the platelet count in whole blood nor with the platelet-rich plasma (r(p)=0.35). No influence of gender or age on platelet count or growth factor concentrations was discovered (except IGF-I).
While there was substantial variation in the growth factor content of platelet-rich plasma, the factors influencing this are still worthy of further investigation. Furthermore, a technique whereby the growth factor content could be rapidly assessed in platelet-rich plasma may be of therapeutic benefit.
The purpose was to evaluate the effect of platelet-rich plasma (PRP) on bone healing.
Fifteen rabbits were included in this randomized, blinded, prospective pilot study. Four equal 8 mm diameter cranial bone defects were created and immediately grafted with autogenous bone, PRP alone, autogenous bone and PRP, and no treatment as a control. The defects were evaluated by digital subtraction radiography with step-wedge calibration, histology, and histomorphometric analysis performed at 1, 2, and 4 months.
The results showed a significant increase in histomorphometric bone area and radiographic bone density in both bone and bone and PRP samples as compared with the control and PRP alone. No significant increase in bone formation was seen with the addition of PRP to autogenous bone. No significant difference in bone formation was seen between defects treated with PRP alone and control sites.
No significant improvement, radiographically or histomorphometrically, was seen with the addition of PRP in bone formation in noncritical sized defects in the rabbit cranial model. However, bone and bone and PRP showed a histomorphometric tendency toward increased bone formation at 1, 2, and 4 months.
The aim of the present study was to compare three different systems for preparing platelet concentrates: two commercially available bed-side techniques (Curasan system and PCCS) and a procedure used routinely in transfusion medicine. Platelet concentrates were prepared from venous blood of 12 healthy male volunteers using the three different systems. Platelet and leucocyte counts were performed and platelet derived growth factor and transforming growth factor beta were assayed by enzyme linked immunoassay. Handling was also considered. The three systems were able to collect 19.0 +/- 16.6% (laboratory system), 41.9 +/- 9.7% (Curasan system) and 49.6 +/- 21.0% (PCCS) of the absolute number of platelets which were originally in the venous blood volume within the platelet concentrate. Due to the amount of plasma which is left in the platelet concentrate portion, the platelet concentration could be increased between 1.4 +/- 1.3 times (laboratory system), 5.0 +/- 2.3 times (PCCS) and 11.7 +/- 2.4 times (Curasan system) compared to the venous blood. The amount of growth factors correlated with the number of platelets within the platelet concentrates. The two systems for intraoperative use are similar in their effects on the platelets. The absolute gain of platelets seems to be the highest with the PCCS; the highest concentration of platelets per micro L is gained with the Curasan system. The laboratory system may offer an alternative if an intraoperative system is not available.
In addition to its central role in blood coagulation and hemostasis, human alpha-thrombin is considered a pro-inflammatory molecule. We have previously demonstrated that differentiated monocytes express the proteolytically activated receptor for thrombin (PAR-1) and that thrombin enhances the release of interleukin (IL)-6 in human monocytes. In the present study we show that thrombin upregulates the production of both IL-1alpha and IL-1beta in phytohemagglutin (PHA)-activated human peripheral blood mononuclear cells (PBMC). Treating PHA-activated PBMC with the PAR-1 activation peptide, SFLLRN, mimics the effects of thrombin on IL-1alpha and IL-1beta production. Thus, it appears that these pro-inflammatory effects induced by thrombin may be mediated through activation of PAR-1. ELISA and RNase protection assays indicate that thrombin and SFLLRN peptide upregulates IL-1 expression at both protein and mRNA levels. Thrombin directly affects monocyte IL-1 expression, since treatment of differentiated U937 cells with thrombin and SFLLRN enhances IL-1 production. These results may help explain how thrombin can enhance IL-1 expression in normal tissue to initiate tissue repair and why thrombin and thrombin-like enzymes may contribute to inflammatory responses observed in several pathophysiological conditions.
Regulated exocytosis of secretory granules or dense-core granules has been examined in many well-characterized cell types including neurons, neuroendocrine, endocrine, exocrine, and hemopoietic cells and also in other less well-studied cell types. Secretory granule exocytosis occurs through mechanisms with many aspects in common with synaptic vesicle exocytosis and most likely uses the same basic protein components. Despite the widespread expression and conservation of a core exocytotic machinery, many variations occur in the control of secretory granule exocytosis that are related to the specialized physiological role of particular cell types. In this review we describe the wide range of cell types in which regulated secretory granule exocytosis occurs and assess the evidence for the expression of the conserved fusion machinery in these cells. The signals that trigger and regulate exocytosis are reviewed. Aspects of the control of exocytosis that are specific for secretory granules compared with synaptic vesicles or for particular cell types are described and compared to define the range of accessory control mechanisms that exert their effects on the core exocytotic machinery.
Platelets are generally believed to be inactive in terms of de novo protein synthesis. On the other hand, the presence of ribosomes and mRNA molecules is well established. Many studies have used reverse transcriptase (RT) -PCR for detection of gene transcripts in platelets. As RT-PCR is a very sensitive method, any leukocyte contamination of platelet preparations can lead to false results. We performed three filtration procedures to minimize leukocyte contamination of pooled buffy-coat platelet concentrates prior to RNA isolation. Furthermore, by applying a genomic PCR approach with 50 amplification cycles we demonstrated that nucleated cells were not detectable. Microarray hybridization was used to analyze 9,850 individual human genes in RNA from purified platelets. In total we identified 1,526 (15.5%) positive genes. The data were confirmed in six individual experiments each performed on a PC pooled from four individual blood donations. Genes specific for nucleated blood cells such as CD4, CD83 and others were negative and verified the purity of PC. Overrepresentation of positive genes was found in the functional categories of glycoproteins/integrins (22.6% vs. 15.5%, p=0.029) and receptors (20.7% vs. 15.5%, p<0.001). Gene transcripts encoding RANTES, GRO-alpha, MIP-1alpha, MIP-1beta, and others were found at high levels of signal intensity and confirmed literature data. This work provides a mRNA profile of human platelets and a complete list of results can be downloaded from the website of our institute www.ma.uni-heidelberg.de/inst/iti/plt_array.xls. The knowledge about gene transcripts may have an impact on the characterization of novel proteins and their functions in platelets.
The goal of periodontal therapy is to protect and maintain the patient's natural dentition for his or her lifetime. More specifically, after periodontal regenerative surgery, the aim is to achieve complete wound healing and regeneration of the periodontal unit. A recent innovation in dentistry is the preparation and use of platelet-rich plasma (PRP), a concentrated suspension of the growth factors found in platelets. These growth factors are involved in wound healing and are postulated as promoters of tissue regeneration. This clinical update outlines the specific effects of these growth factors, both in vitro and in vivo, on periodontal wound healing. The review focuses on current animal and human trials using PRP to promote tissue regeneration and alveolar bone repair. The article goes on to describe the clinical benefits of PRP and the step-by-step preparation of PRP in the dental office.
Autologous platelet concentrate (PC) is applied locally to improve wound healing and tissue repair. Previous measurements of the growth factor content of platelets have given conflicting results. To date, there is no information on the influence of different preanalytical sample-preparation methods on the detectable amount of growth factors.
We measured the level of growth factors in PCs obtained by plateletpheresis and by leukapheresis. We subjected aliquots of these components to six different preparation methods: freezing/thawing once or twice; dissolution in 0.5% Triton-X-100; and clot formation by the addition of calcium and thrombin with subsequent incubation for 1 h, for 24 h, or for 1 h followed by freezing and thawing.
In samples dissolved in Triton-X-100, higher levels of growth factors were detected than in the other specimens. In comparison to clot formation, freezing and thawing platelets twice was equivalent with respect to the release of platelet-derived growth factor (PDGF) but superior with respect to the release of transforming growth factor-beta1 (TGF-beta1). Overall, mean levels of 4.77 x 10(-16) g of PDGF-AB, 2.2 x 10(-17) g of PDGF-BB, and 2.41 x 10(-16) g of TGF-beta1 were found per single human platelet in white blood cell (WBC)-poor samples dissolved in Triton-X-100.
Dissolving PC in Triton-X-100 releases maximum quantities of growth factors from platelets. The release of each growth factor by any sample preparation method should be investigated and interpreted separately. The preanalytical sample-preparation method, as well as the platelet and WBC content, influence the measurable levels of growth factors in PCs. The results implicate the need to correct, considerably upwards, previous estimations of the PDGF content of platelets.
Clinical evidence suggests that platelet concentrate (PC) could have beneficial therapeutic effects on hard and soft tissue healing, due to the contents of growth factors (GFs) stored in the platelets. The objectives of this study were: 1) to determine the concentrations of platelet-derived growth factor-BB (PDGF-BB), transforming growth factor-beta1 (TGF-beta1), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) released from PCs and whole blood (WB), before and after the addition of various concentrations of calcium and thrombin, and 2) to assess the physiological importance of the released GFs on angiogenesis.
WB and PCs were harvested and prepared from three healthy volunteers. Enzyme-linked immunosorbent assay tests, specific for PDGF-BB, TGF-beta1, VEGF, and bFGF, were performed on WB and PC supernatants, collected before and 30 minutes after the addition of various concentrations of calcium and thrombin. The supernatants were also added to human umbilical vein endothelial cell (HUVEC) cultures in order to measure their effects on endothelial cell proliferation.
Growth factor concentrations detected in PC supernatants were significantly greater (280% to 800% increase) than concentrations present in WB supernatants. Calcium and thrombin induced immediate GF release from PCs in a dose-dependent fashion. Furthermore, PC supernatants led to greater HUVEC proliferation rates than WB supernatants. However, there was no correlation between the concentrations of specific GFs and HUVEC proliferation rates.
These results suggest that PCs could stimulate blood vessel formation. They also reinforce the relevance for using PCs in regenerative therapies.
Platelet concentrates (PCs) constitute new biological mediators used in osseous reconstructive surgery. In this study, we assessed (i) the effects of various concentrations of calcium and thrombin on the kinetics of platelet-derived growth factor (PDGF-BB), transforming growth factor-beta1(TGF-beta 1), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) release by PCs and (ii) the contribution of PC supernatants to endothelial cell proliferation. Our results indicate that high concentrations of calcium (Ca) and thrombin (Thr) trigger an immediate and significant increase in bFGF, TGF-beta 1 and PDGF-BB concentrations. Thereafter, PDGF-BB, VEGF and TGF-beta 1 levels remained generally constant over a 6-day period while a decrease in bFGF concentrations was noted after 24h. Lower Ca and Thr concentrations tended to reduce and delay growth factors release from PCs. Endothelial cell proliferation was greatly enhanced with PC supernatants (mean: 20-fold increase). This was especially evident when endothelial cells were treated with supernatants harvested early after PC treatment with high concentrations of Ca and Thr or later after PC treatment with low Ca and Thr concentrations. Additional research aiming to measure the effects of Ca and Thr on bone formation in vivo is needed.
Guided bone regeneration is an accepted surgical method employed in implant dentistry to increase the quantity and quality of the host bone in areas of localized alveolar defects. The lack of predictability in osseous regenerative procedures with various grafting materials suggests that improvement in the osteoinductive properties of these materials is highly desirable. Platelet-rich plasma (PRP), a modification of fibrin glue made from autologous blood, is being used to deliver growth factors in high concentration to sites requiring osseous grafting. Growth factors released from the platelets include platelet-derived growth factor, transforming growth factor beta, platelet-derived epidermal growth factor, platelet-derived angiogenesis factor, insulin-like growth factor 1, and platelet factor 4. These factors signal the local mesenchymal and epithelial cells to migrate, divide, and increase collagen and matrix synthesis. PRP has been suggested for use to increase the rate of bone deposition and quality of bone regeneration when augmenting sites prior to or in conjunction with dental implant placement Only 6 human studies using PRP have been found in the dental implant literature and 5 were case series or reports. Thus, there is clearly a lack of scientific evidence to support the use of PRP in combination with bone grafts during augmentation procedures. This novel and potentially promising technique requires well-designed, controlled studies to provide evidence of efficacy.
Cutaneous wound healing is a complex process involving blood clotting, inflammation, new tissue formation, and finally tissue remodeling. It is well described at the histological level, but the genes that regulate skin repair have only partially been identified. Many experimental and clinical studies have demonstrated varied, but in most cases beneficial, effects of exogenous growth factors on the healing process. However, the roles played by endogenous growth factors have remained largely unclear. Initial approaches at addressing this question focused on the expression analysis of various growth factors, cytokines, and their receptors in different wound models, with first functional data being obtained by applying neutralizing antibodies to wounds. During the past few years, the availability of genetically modified mice has allowed elucidation of the function of various genes in the healing process, and these studies have shed light onto the role of growth factors, cytokines, and their downstream effectors in wound repair. This review summarizes the results of expression studies that have been performed in rodents, pigs, and humans to localize growth factors and their receptors in skin wounds. Most importantly, we also report on genetic studies addressing the functions of endogenous growth factors in the wound repair process.