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Knockdown of HGF, followed by TGF-beta1 supplementation, is sufficient to differentiate OMFs into myofibroblasts.
Source publication
Oral mucosal wounds are characterized by rapid healing with minimal scarring, partly attributable to the “enhanced” wound healing properties of oral mucosal fibroblasts (OMFs).Hepatocyte growth factor (HGF) is a pleiotropic growth factor, with potential key roles in accelerating healing and preventing fibrosis. HGF can exist as full-length or trunc...
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Abstract Transforming growth factor-β1 (TGF-β1) is a potent inducer of fibroblast to myofibroblast differentiation and contributes to the pro-fibrotic microenvironment during cardiac remodeling. Fibroblast growth factor-2 (FGF-2) is a growth factor secreted by adipose tissue-derived stromal cells (ASC) which can antagonize TGF-β1 signaling. We hypo...
Citations
... When considering the normal amount of HGF present in the human body or in wound-healing applications, the physiological concentration of HGF can vary. In normal serum, HGF levels can be relatively low, typically under 0.5 ng/mL [27][28][29], but may increase significantly in tissue repair and wound-healing scenarios up to 140 ng/mL [30,31]. In therapeutic contexts, the concentrations of HGF could be elevated to promote healing. ...
Citation: Wein, S.; Jung, S.A.; Al Enezy-Ulbrich, M.A.; Reicher, L.; Rütten, S.; Kühnel, M.; Jonigk, D.; Jahnen-Dechent, W.; Pich, A.; Neuss, S. These authors contributed equally to this work. Abstract: A novel scaffold design has been created to enhance tissue engineering and regenerative medicine by optimizing the controlled, prolonged release of Hepatocyte Growth Factor (HGF), a powerful chemoattractant for endogenous mesenchymal stem cells. We present a new stacked scaffold that is made up of three different fibrin gel layers, each of which has HGF integrated into the matrix. The design attempts to preserve HGF's regenerative properties for long periods of time, which is necessary for complex tissue regeneration. These multi-layered fibrin gels have been mechanically evaluated using rheometry, and their degradation behavior has been studied using D-Dimer ELISA. Understanding the kinetics of HGF release from this novel scaffold configuration is essential for understanding HGF's long-term sustained bioactivity. A range of cell-based tests were carried out to verify the functionality of HGF following extended incorporation. These tests included 2-photon microscopy using phalloidin staining to examine cellular morphology, SEM analysis for scaffold-cell interactions, and scratch and scatter assays to assess migration and motility. The analyses show that the novel stacking scaffold promotes vital cellular processes for tissue regeneration in addition to supporting HGF's bioactivity. This scaffold design was developed for in situ tissue engineering. Using the body as a bioreactor, the scaffold should recruit mesenchymal stem cells from their niche, thus combining the regenerative abilities of HGF and MSCs to promote tissue remodeling and wound repair.
... TGF-β1 induces myofibroblast differentiation through the SMAD pathway [35] and HGF has previously been shown to inhibit myofibroblast differentiation in a number of different fibroblast types [30,32,36], including with HGF derived from ADSC [32]. HGF was found in our adipose tissue conditioned medium at a higher concentration than in ADSC conditioned medium and was therefore selected for further investigation. ...
Background
Hypertrophic scarring results from myofibroblast differentiation and persistence during wound healing. Currently no effective treatment for hypertrophic scarring exists however, autologous fat grafting has been shown to improve scar elasticity, appearance, and function. The aim of this study was to understand how paracrine factors from adipose tissues and adipose-derived stromal cells (ADSC) affect fibroblast to myofibroblast differentiation.
Methods
The transforming growth factor-β1 (TGF-β1) induced model of myofibroblast differentiation was used to test the effect of conditioned media from adipose tissue, ADSC or lipid on the proportion of fibroblasts and myofibroblasts.
Results
Adipose tissue conditioned media inhibited the differentiation of fibroblasts to myofibroblasts but this inhibition was not observed following treatment with ADSC or lipid conditioned media. Hepatocyte growth factor (HGF) was readily detected in the conditioned medium from adipose tissue but not ADSC. Cells treated with HGF, or fortinib to block HGF, demonstrated that HGF was not responsible for the inhibition of myofibroblast differentiation. Conditioned media from adipose tissue was shown to reduce the proportion of myofibroblasts when added to fibroblasts previously treated with TGF-β1, however, conditioned media treatment was unable to significantly reduce the proportion of myofibroblasts in cell populations isolated from scar tissue.
Conclusions
Cultured ADSC or adipocytes have been the focus of most studies, however, this work highlights the importance of considering whole adipose tissue to further our understanding of fat grafting. This study supports the use of autologous fat grafts for scar treatment and highlights the need for further investigation to determine the mechanism.
... When considering the normal amount of HGF present in the human body or in wound healing applications, the physiological concentration of HGF can vary. In normal serum, HGF levels can be relatively low, typically under 0.5 ng/ml [26][27][28], but may increase significantly in tissue repair and wound healing scenarios up to 140 ng/ml [29,30]. In therapeutic contexts, the concentrations of HGF could be elevated to promote healing. ...
A novel scaffold design has been created to enhance tissue engineering and regenerative medicine by optimizing the controlled, prolonged release of hepatocyte growth factor (HGF), a powerful chemoattractant for endogenous mesenchymal stem cells. We present a new stacked scaffold that is made up of three different fibrin gel layers, each of which has HGF integrated into the matrix. The design attempts to preserve HGF's regenerative properties for long periods of time, which is necessary for complex tissue regeneration. These multi-layered fibrin gels have been mechanically evaluated using rheometry, and their degradation behavior has been studied using D-Dimer ELISA. Understanding the kinetics of HGF release from this novel scaffold configuration is essential for understanding HGF's long-term sustained bioactivity. A range of cell-based tests were carried out to verify the functionality of HGF following extended incorporation. These tests included 2-photon microscopy using phalloidin staining to examine cellular morphology, SEM analysis for scaffold-cell interactions, and scratch and scatter assays to assess migration and motility. The analyses show that the novel stacking scaffold promotes vital cellular processes for tissue regeneration in addition to supporting HGF's bioactivity. This scaffold design was developed for in situ tissue engineering. Using the body as a bioreactor, the scaffold should recruit mesenchymal stem cells from their niche thus combining the regenerative abilities of HGF and MSCs to promote tissue remodeling and wound repair.
... Hepatocyte growth factor-cMET signaling is responsible for adhesion development following later follicular fluid exposure Hepatocyte growth factor (HGF) is a well-known growth factor responsible for tissue repair and regeneration. 17,25 Previously, we showed that HGF and its activator HGFA in FF are continuously activated once the coagulation protease cascade is activated after ovulation. 17 Through the activation of thrombin via the extrinsic coagulation pathway, HGFA cleaves pro-HGF to activate HGF. ...
... Hepatocyte growth factor splice variant NK1 (HGF/NK1) and bone morphogenetic protein 7 (BMP7) are two growth factors with demonstrated potent anti-fibrotic activities [22,23]. Huang and colleagues [15] employed H-RN peptide, derived from the kringle 1 domain present in HGF/NK1, to show in vitro inhibition of TGF-β2/Smad2/3-dependent induction of epithelial-mesenchymal transition (EMT) in lens epithelial cells, preventing their acquisition of myofibroblastic markers. ...
Myofibroblasts are the principal effector cells driving fibrosis, and their accumulation in tissues is a fundamental feature of fibrosis. Essential pathways have been identified as being central to promoting myofibroblast differentiation, revealing multiple targets for intervention. Compared with large proteins and antibodies, peptide-based therapies have transpired to serve as biocompatible and cost-effective solutions to exert biomimicry, agonistic, and antagonistic activities with a high degree of targeting specificity and selectivity. In this review, we summarize emergent antifibrotic peptides and their utilization for the targeted prevention of myofibroblasts. We then highlight recent studies on peptide inhibitors of upstream pathogenic processes that drive the formation of profibrotic cell phenotypes. We also briefly discuss peptides from non-mammalian origins that show promise as antifibrotic therapeutics. Finally, we discuss the future perspectives of peptide design and development in targeting myofibroblasts to mitigate fibrosis.
... Such contrasting genotypic profiles between OMFs and SFs contribute to the differential wound healing capabilities of oral mucosal and skin tissues, particularly in terms of the superior proliferative, migratory and matrix metalloproteinase (MMP)-mediated extracellular matrix (ECM) remodeling properties of OMFs, closely associated with their "younger" phenotype [4][5][6][7][8][9]. Furthermore, although SF-myofibroblast differentiation induced by profibrotic mediators, such as transforming growth factor-β1 (TGF-β1), is a pivotal response in facilitating normal wound closure, contraction, pro-fibrotic ECM deposition and scar formation in skin [10], OMFs exhibit lower TGF-β1 expression and resistance to TGF-β1driven myofibroblast differentiation compared to SFs, thereby retaining their nonscarring phenotype [5,[11][12][13]. ...
... Expression profiling comparisons between patient-matched OMFs and SFs have enhanced our understanding of the preferential wound healing responses of OMFs at a molecular level through identification of the key genes involved, such as hepatocyte growth factor (HGF) [5,9,[13][14][15][16][17]. However, another prominent regulator of normal and pathological wound healing and scarring mechanisms is oxidative stress, being particularly recognized in dermal tissues [18][19][20][21]. ...
... As previous studies have demonstrated that SFs are more vulnerable to cellular senescence in vitro compared to their patient-matched OMF counterparts due to their possession of "longer" telomeres [8], these findings support the concept that limited ROS generation and their accompanying resistance to oxidative biomolecular damage contribute to OMFs being genotypically and phenotypically "younger" than patient-matched SFs. Such properties, therefore, may be significant in relation to the maintenance of the preferential wound healing and minimal scarring properties of OMFs relative to patient-matched SFs [1,2,[4][5][6][7][8][9][10][11][12][13]. ...
Unlike skin, oral mucosal wounds are characterized by rapid healing and minimal scarring, attributable to the "enhanced" healing properties of oral mucosal fibroblasts (OMFs). As oxidative stress is increasingly implicated in regulating wound healing outcomes, this study compared oxidative stress biomarker and enzymic antioxidant profiles between patient-matched oral mucosal/skin tissues and OMFs/skin fibroblasts (SFs) to determine whether superior oral mucosal antioxidant capabilities and reduced oxidative stress contributed to these preferential healing properties. Oral mucosa and skin exhibited similar patterns of oxidative protein damage and lipid peroxidation, localized within the lamina propria/dermis and oral/skin epithelia, respectively. SOD1, SOD2, SOD3 and catalase were primarily localized within epithelial tissues overall. However, SOD3 was also widespread within the lamina propria localized to OMFs, vasculature and the extracellular matrix. OMFs were further identified as being more resistant to reactive oxygen species (ROS) generation and oxidative DNA/protein damage than SFs. Despite histological evaluation suggesting that oral mucosa possessed higher SOD3 expression, this was not fully substantiated for all OMFs examined due to inter-patient donor variability. Such findings suggest that enzymic antioxidants have limited roles in mediating privileged wound healing responses in OMFs, implying that other non-enzymic antioxidants could be involved in protecting OMFs from oxidative stress overall. Citation: Lohana, P.; Suryaprawira, A.; Woods, E.L.; Dally, J.; Gait-Carr, E.; Alaidaroos, N.Y.A.; Heard, C.M.; Lee, K.Y.; Ruge, F.; Farrier, J.N.; et al.
... These results support the important role of PG in the repair of the epithelial barrier and unveil its function in arresting epithelial regeneration through fibroblast-derived paracrine factors. Since IL-33 and Hepatocyte Growth Factor (HGF) have been demonstrated as factors released by fibroblasts involved in mucosal healing processes [22,23], the effect of PG on the release of both factors was tested by quantifying the levels of IL-33 and HGF in the supernatants of IBD fibroblasts stimulated for 48 h in presence or the absence of PG (5 µg/mL). IL-33 resulted undetectable in all experimental groups , whereas HGF levels significantly increased after PG stimulation ( Figure 7E). ...
... These data confirm that PG does not promote the accumulation and maturation in the gut of collagen fibers typical of fibrotic processes. Consistently with this, HGF, which exerts antifibrotic effects by modulating TGF-β signaling [23], was upregulated in primary fibroblasts stimulated with PG. HGF is a paracrine soluble factor that supports intestinal tissue repair controlling epithelial cell proliferation. ...
... HGF is a paracrine soluble factor that supports intestinal tissue repair controlling epithelial cell proliferation. Resident fibroblasts, indeed, besides producing extracellular matrix and participating in tissue ECM maintenance in homeostasis and remodeling after injury, play an important role in the regeneration of epithelial cells by secreting cytokines and several growth factors [41], including IL33 and HGF [22,23]. Different fibroblast subsets have been identified in the gut; those located around the crypts drive epithelial stem cell niche maintenance and crypt proliferation through Wnt signal-ing [42]. ...
Background: Pomegranate (Punica granatum) can be used to prepare a bioactive extract exerting anti-inflammatory activities. Clinical studies demonstrated an improvement in clinical response in inflammatory bowel disease (IBD) patients when pomegranate extract (PG) was taken as a complement to standard medications. However, the molecular mechanisms underlying its beneficial effects are still scarcely investigated. This study investigates the effect of PG on bacterial biofilm formation and the promotion of mucosal wound healing. Methods: The acute colitis model was induced in C57BL/6N mice by 3% dextran sodium sulfate administration in drinking water for 5 days. During the recovery phase of colitis, mice received saline or PG (200 mg/kg body weight) by oral gavage for 11 days. Colitis was scored daily by evaluating body weight loss, bleeding, and stool consistency. In vivo intestinal permeability was evaluated by fluorescein isothiocyanate-conjugated dextran assay, bacterial translocation was assessed by fluorescence in situ hybridization on tissues, whereas epithelial and mucus integrity were monitored by immunostaining for JAM-A and MUC-2 markers. Bacterial biofilm formation was assessed using microfluidic devices for 24 or 48 h. Primary fibroblasts were isolated from healthy and inflamed areas of 8 IBD patients, and Caco-2 cells were stimulated with or without PG (5 μg/mL). Inflammatory mediators were measured at the mRNA and protein level by RT-PCR, WB, or Bio-plex multiplex immunoassay, respectively. Results: In vivo, PG boosted the recovery phase of colitis, promoting a complete restoration of the intestinal barrier with the regeneration of the mucus layer, as also demonstrated by the absence of bacterial spread into the mucosa and the enrichment of crypt-associated fibroblasts. Microfluidic experiments did not highlight a specific effect of PG on Enterobacterales biofilm formation, even though Citrobacter freundii biofilm was slightly impaired in the presence of PG. In vitro, inflamed fibroblasts responded to PG by downregulating the release of metalloproteinases, IL-6, and IL-8 and upregulating the levels of HGF. Caco-2 cells cultured in a medium supplemented with PG increased the expression of SOX-9 and CD44, whereas in the presence of HGF or plated with a fibroblast-conditioned medium, they displayed a decrease in SOX-9 and CD44 expression and an increase in AXIN2, a negative regulator of Wnt signaling. Conclusions: These data provide new insight into the manifold effects of PG on promoting mucosal homeostasis in IBD by affecting pathogen biofilm formation and favoring the regeneration of the intestinal barrier through the regulation of the crosstalk between epithelial and stromal cells.
... For example, vascular endothelial (VEGF), fibroblast (FGF), and platelet-derived (PDGF) growth factors affect hemostasis and angiogenesis (Shen et al., 2016). Moreover, hepatocyte growth factor (HGF) contributes to the re epithelialization of injured tissues (Dally et al., 2017), while both FGF and epidermal growth factor (EGF) regulate the reepithelialization process (Seeger and Paller, 2015). HGF regulates re-epithelialization by binding to and activating the MET receptor tyrosine kinase (Chmielowiec et al., 2007). ...
Wounds can be divided into two categories, acute and chronic. Acute wounds heal through the normal wound healing process. However, chronic wounds take longer to heal, leading to inflammation, pain, serious complications, and an economic burden of treatment costs. In addition, diabetes and burns are common causes of chronic wounds that are difficult to treat. The rapid and thorough treatment of chronic wounds, including diabetes wounds and burns, represents a significant unmet medical need. Wound dressings play an essential role in chronic wound treatment. Various biomaterials for wound healing have been developed. Among these, hydrogels are widely used as wound care materials due to their good biocompatibility, moisturizing effect, adhesion, and ductility. Wound healing is a complex process influenced by multiple factors and regulatory mechanisms in which stem cells play an important role. With the deepening of stem cell and regenerative medicine research, chronic wound treatment using stem cells has become an important field in medical research. More importantly, the combination of stem cells and stem cell derivatives with hydrogel is an attractive research topic in hydrogel preparation that offers great potential in chronic wound treatment. This review will illustrate the development and application of advanced stem cell therapy-based hydrogels in chronic wound healing, especially in diabetic wounds and burns.
... IL-1ra blocks the inflammatory IL-1 receptors, resulting in less inflammation, PDGF-BB could be promoting proliferation and directing migration of mesenchymal cells and fibroblasts 76 and HGF may be contributing to cell growth and enhancing wound healing. 77 FGF-2 was also eluted reducing tissue death and encouraging cellular repair. 78 Not surprisingly, IL-10 was not detected in the membrane eluate. ...
Background
The favorable biological and mechanical properties of the most common components of the placenta, the amnion and chorion, have been explored for regenerative medical indications. The use of the combination of amnion and chorion has also become very popular. But, published data from placental tissues in their final, useable form is lacking. During treatment with membrane product, the tissue is usually sterile, intact and laid on a wound or treatment area. The factors available to the treatment area from the applied product need to be elucidated and presented in a relatable form. Current reporting for eluted growth factor results are typically expressed per milliliter, which is not informative with respect to the area of tissue covered by the actual membrane and may differ among techniques.
Methods
To address this inconsistency, amnion or amnion/chorion were isolated from human placentas and processed by a proprietary procedure. The final dry, sterilized product was evaluated for structural components and growth factor elution. Growth factors were quantified by multiplex panels and ELISAs and the values normalized to specific area and elution volume of finished product. This information allows extrapolation to all membrane sizes and affords cross‐study comparisons.
Results
Analysis of membrane supernatants show that dehydrated, sterilized amnion and amnion/chorion elute factors that are conducive to wound healing, which are available to recipient tissues. Importantly, these measurable factors eluted from dehydrated, sterilized membranes can be reported as a function of available factors per square centimeter of tissue.
Conclusions
The standardized characterization of dehydrated, sterilized amnion and amnion/chorion as delivered to recipient tissues permits understanding and comparison of the products across various graft sizes, types, and eluate volumes. Further, reporting this data as a function of cm² of dehydrated tissue allows extrapolation by independent scientists and clinicians.
... Other studies using OMLP-PC-derived sEVs have only focused on their effects on cell proliferation 50 and their impact on myofibroblast formation was previously unknown. It was known, however, that oral fibroblasts, that most probably contain some oral progenitors, are resistant to differentiation into myofibroblasts, 55,56 an effect that could, inherently, be due to OMLP-PC derived sEVs. Importantly, in terms of realworld functionality, in a murine wound model OMLP-PC L sEVs also demonstrated an ability to inhibit the formation of αSMA positive myofibroblasts and significantly reduced collagen deposition 7 days post wounding suggesting that such oral sEVs may potentiate longer-term scarring outcome. ...
Scar formation during wound repair can be devastating for affected individuals. Our group previously documented the therapeutic potential of novel progenitor cell populations from the non-scarring buccal mucosa. These Oral Mucosa Lamina Propria-Progenitor Cells (OMLP-PCs) are multipotent, immunosuppressive, and antibacterial. Small extracellular vesicles (sEVs) may play important roles in stem cell–mediated repair in varied settings; hence, we investigated sEVs from this source for wound repair. We created an hTERT immortalized OMLP-PC line (OMLP-PCL) and confirmed retention of morphology, lineage plasticity, surface markers, and functional properties. sEVs isolated from OMLP-PCL were analyzed by nanoparticle tracking analysis, Cryo-EM and flow cytometry. Compared to bone marrow–derived mesenchymal stromal cells (BM-MSC) sEVs, OMLP-PCL sEVs were more potent at driving wound healing functions, including cell proliferation and wound repopulation and downregulated myofibroblast formation. A reduced scarring potential was further demonstrated in a preclinical in vivo model. Manipulation of OMLP-PCL sEVs may provide novel options for non-scarring wound healing in clinical settings.
