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Identification of dermal parameters for HTscar formation A) Thickness of dermis of SE (μm); B) relative matrix contraction of SE (surface area after 5 weeks of culture divided by surface at day 0); C) collagen 1 secretion into culture supernatants (ng/ml per equivalent per 24 h). Experiments were performed with SE constructed from three different donors each in duplicate. Keratinocytes, P-DSC, R-DSC, and ASC were all from the same donor within a single experiment. Data are presented as the mean (n=3 ±SEM) thickness of dermis, contraction, or secretion of collagen 1. Statistically significant differences were calculated using a repeated measures ANOVA test followed by Bonferroni's multiple comparison test. *, P<0.05; **, P<0.01.
Source publication
Adverse hypertrophic scars can form after healing of full-thickness skin wounds. Currently, reliable animal and in vitro models to identify and test novel scar reducing therapeutics are scarce. Here we describe the development and validation of a tissue-engineered human hypertrophic scar (HTscar) model based on reconstructed epidermis on a dermal m...
Contexts in source publication
Context 1
... healing, the development of HTscar is charac- terized by an overproduction of extracellular matrix, increased contraction, and augmented α-SMA expression compared to NTscar (Ehrlich et al., 1994). For this reason we first compared Se ASC with Se R-DSC and Se P-DSC with regards to thickness of the dermis, contraction, and collagen 1 secretion (Fig. ...
Context 2
... dermal thickness was not significantly different be- tween the three Se ( Fig. 2A). An increase in contraction is represented by a decrease in surface area of the SE and was ...
Context 3
... SE constructed from ASC populated matrixes not only visually represent HTscars, but also enabled quantifiable parameters to be identified, which are representative for HT- observed for SE ASC compared to Se R-DSC and Se P-DSC (Fig. 2B). Se ASC secreted significantly more collagen 1 compared to Se P-DSC (Fig. ...
Context 4
... SE constructed from ASC populated matrixes not only visually represent HTscars, but also enabled quantifiable parameters to be identified, which are representative for HT- observed for SE ASC compared to Se R-DSC and Se P-DSC (Fig. 2B). Se ASC secreted significantly more collagen 1 compared to Se P-DSC (Fig. ...
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The majority of studies on scar formation have mainly focused on the dermis and little is known of the involvement of the epidermis. Previous research has demonstrated that the scar tissue-derived keratinocytes are different from normal cells at both the genetic and cell biological levels; however, the mechanisms responsible for the fundamental abn...
Citations
... The topical application of 10% simvastatin cream alone, but not 2% simvastatin and 10% pravastatin creams, significantly attenuated the hypertrophy of resultant scars compared with vehicle cream alone. Data indicate that topical 10% simvastatin cream antagonizes dermal fibrosis and reduces hypertrophy [332]. ...
Keloids (KD) and hypertrophic scars (HTS), which are quite raised and pigmented and have increased vascularization and cellularity, are formed due to the impaired healing process of cutaneous injuries in some individuals having family history and genetic factors. These scars decrease the quality of life (QOL) of patients greatly, due to the pain, itching, contracture, cosmetic problems, and so on, depending on the location of the scars. Treatment/prevention that will satisfy patients’ QOL is still under development. In this article, we review pharmacotherapy for treating KD and HTS, including the prevention of postsurgical recurrence (especially KD). Pharmacotherapy involves monotherapy using a single drug and combination pharmacotherapy using multiple drugs, where drugs are administered orally, topically and/or through intralesional injection. In addition, pharmacotherapy for KD/HTS is sometimes combined with surgical excision and/or with physical therapy such as cryotherapy, laser therapy, radiotherapy including brachytherapy, and silicone gel/sheeting. The results regarding the clinical effectiveness of each mono-pharmacotherapy for KD/HTS are not always consistent but rather scattered among researchers. Multimodal combination pharmacotherapy that targets multiple sites simultaneously is more effective than mono-pharmacotherapy. The literature was searched using PubMed, Google Scholar, and Online search engines.
... Proinflammatory and anti-inflammatory cytokine secretion decreased considerably in adipose-RhS (and differentiated adipocyte monocultures). This is in line with previous studies conducted with an ASC-derived hypertrophic scar model and indicates that the addition of the adipose layer containing a mixture of stromal cells and adipocytes leads to a lower inflammatory state and contributes to tissue homeostasis [39]. It is also generally known that ASCs have immune-regulatory effects [40,41]. ...
Background
Dysregulation of skin metabolism is associated with a plethora of diseases such as psoriasis and dermatitis. Until now, reconstructed human skin (RhS) models lack the metabolic potential of native human skin, thereby limiting their relevance to study human healthy and diseased skin. We aimed to determine whether incorporation of an adipocyte-containing hypodermis into RhS improves its metabolic potential and to identify major metabolic pathways up-regulated in adipose-RhS.
Methods
Primary human keratinocytes, fibroblasts and differentiated adipose-derived stromal cells were co-cultured in a collagen/fibrin scaffold to create an adipose-RhS. The model was extensively characterized structurally in two- and three-dimensions, by cytokine secretion and RNA-sequencing for metabolic enzyme expression.
Results
Adipose-RhS showed increased secretion of adipokines. Both RhS and adipose-RhS expressed 29 of 35 metabolic genes expressed in ex vivo native human skin. Addition of the adipose layer resulted in up-regulation of 286 genes in the dermal-adipose fraction of which 7 were involved in phase I (CYP19A1, CYP4F22, CYP3A5, ALDH3B2, EPHX3) and phase II (SULT2B1, GPX3) metabolism. Vitamin A, D and carotenoid metabolic pathways were enriched. Additionally, pro-inflammatory (IL-1β, IL-18, IL-23, IL-33, IFN-α2, TNF-α) and anti-inflammatory cytokine (IL-10, IL-12p70) secretion was reduced in adipose-RhS.
Conclusions
Adipose-RhS mimics healthy native human skin more closely than traditional RhS since it has a less inflamed phenotype and a higher metabolic activity, indicating the contribution of adipocytes to tissue homeostasis. Therefore it is better suited to study onset of skin diseases and the effect of xenobiotics.
... Each sample was measured five times, and the average of those readings was used in the statistical analysis. The 10x objective's scale bar was calibrated against a known standard before measurements were taken, as detailed in previous studies [4,5]. ...
... Fibroblasts derived from deep dermal layers of healthy skin were reported to display characteristics similar to what was observed for hypertrophic scars, such as increased production of connective tissue growth factor (CTGF), collagen, and a-SMA, or decreased decorin amounts [74,136]. Moreover, hypertrophic scar characteristic traits, i.e., enhanced contraction, increased collagen, and a-SMA production, were observed in 3D models of dermal equivalents or HSEs using deep dermal fibroblasts [137], mechanical stress-activated fibroblasts [138], or cutaneous adipose tissue mesenchymal stem cells [139]. ...
Skin wound healing is essential to health and survival. Consequently, high amounts of research effort have been put into investigating the cellular and molecular components involved in the wound healing process. The use of animal experiments has contributed greatly to the knowledge of wound healing, skin diseases, and the exploration of treatment options. However, in addition to ethical concerns, anatomical and physiological inter-species differences often influence the translatability of animal-based studies. Human in vitro skin models, which include essential cellular and structural components for wound healing analyses, would improve the translatability of results and reduce animal experiments during the preclinical evaluation of novel therapy approaches. In this review, we summarize in vitro approaches, which are used to study wound healing as well as wound healing-pathologies such as chronic wounds, keloids, and hypertrophic scars in a human setting.
... Cells from different (disease-related) origins, such as skin cells derived from fetal, burn, or scar tissue, could be used to study their effect on skin regeneration. For example, van den Broek et al. developed a hypertrophic scar model using adipose-derived mesenchymal stem cells [55]. In these scar models, differences in contraction, epidermal thickness, and cytokine response were shown compared to models produced from dermal mesenchymal cells. ...
Healing of burn injury is a complex process that often leads to the development of functional and aesthetic complications. To study skin regeneration in more detail, organotypic skin models, such as full skin equivalents (FSEs) generated from dermal matrices, can be used. Here, FSEs were generated using de-epidermalized dermis (DED) and collagen matrices MatriDerm® and Mucomaix®. Our aim was to validate the MatriDerm- and Mucomaix-based FSEs for the use as in vitro models of wound healing. Therefore, we first characterized the FSEs in terms of skin development and cell proliferation. Proper dermal and epidermal morphogenesis was established in all FSEs and was comparable to ex vivo human skin models. Extension of culture time improved the organization of the epidermal layers and the basement membrane in MatriDerm-based FSE but resulted in rapid degradation of the Mucomaix-based FSE. After applying a standardized burn injury to the models, re-epithelization occurred in the DED- and MatriDerm-based FSEs at 2 weeks after injury, similar to ex vivo human skin. High levels of pro-inflammatory cytokines were present in the culture media of all models, but no significant differences were observed between models. We anticipate that these animal-free in vitro models can facilitate research on skin regeneration and can be used to test therapeutic interventions in a preclinical setting to improve wound healing.
... Collagen synthesis is promoted by the migrated fibroblasts from which myofibroblasts are differentiated, and, at the open wound bed, the myofibroblasts control bleeding by contraction of the ruptured vessels. However, overproduction of myofibroblasts causes scar and contracture in the wounds [6]: myofibroblasts express α-smooth muscle actins (α-SMA), which are mostly staining and covering the vessel wall but are also found in the lower dermis in the case of hypertrophic scars [7]. It is postulated that excess α-SMA levels upregulate fibroblast contractile activity [8,9]. ...
... In the past years, human physiologically relevant tissue-engineered skin models have been developed for wound healing advanced therapy medicinal products (ATMPs) and in vitro assays. [55][56][57][58] These models can be used for validation and improvement of computational models and to collect new preclinical data. However, skin substitutes do not contain all skin cell types and therefore do not fully represent original skin physiology. ...
Healthcare is undergoing a profound technological and digital transformation and has become increasingly complex. It is important for burns professionals and researchers to adapt to these developments which may require new ways of thinking and subsequent new strategies. As Einstein has put it: ‘We must learn to see the world anew'. The relatively new scientific discipline "Complexity science" can give more direction to this and is the metaphorical open door that should not go unnoticed in view of the burn care of the future.
Complexity sciences studies ‘why the whole is more than the sum of the parts’. It studies
how multiple separate components interact with each other and their environment and how these interactions lead to ‘behavior of the system’. Biological systems are always part of smaller and larger systems and exhibit the behavior of adaptivity, hence the name complex adaptive systems. From the perspective of complexity science, a severe burn injury is an extreme disruption of the 'human body system'. But this disruption also applies to the systems at the organ and cellular level. All these systems follow principles of complex systems. Awareness of the scaling process at multilevel helps to understand and manage the complex situation when dealing with severe burn cases.
The aim of this paper is to create awareness of the concept of complexity and to demonstrate the value and possibilities of complexity science methods and tools for the future of burn care through examples from preclinical, clinical, and organizational perspective in burn care.
... Muscle Actin (α-SMA). Hypertrophic scars contain abundant nodules containing myofibroblasts, which contain α-SMA [46] Myofibroblasts are differentiated fibroblasts found in granulation tissue and fibrotic lesions [28,29,47] and are responsible for skin contraction after wounding [46]. A large proportion of myofibroblasts express α-SMA, and persistence of myofibroblasts may lead to excess scarring, which impairs function and aesthetics [35,48]. ...
... Muscle Actin (α-SMA). Hypertrophic scars contain abundant nodules containing myofibroblasts, which contain α-SMA [46] Myofibroblasts are differentiated fibroblasts found in granulation tissue and fibrotic lesions [28,29,47] and are responsible for skin contraction after wounding [46]. A large proportion of myofibroblasts express α-SMA, and persistence of myofibroblasts may lead to excess scarring, which impairs function and aesthetics [35,48]. ...
... There was also a significant increase in α- 10 Analytical Cellular Pathology SMA stroma levels in the AlloDerm® compared to the Matri-derm® group (p < 0:05). van den Broek et al. stated that with hypertrophic scars, α-SMA positive staining was present not just around blood vessels but in single cells in the lower dermis as well [46]. In our study, there was no significant difference in α-SMA in the stroma between the Matriderm® group and in the normal skin in which both had low cell numbers compared to the other intervention groups (p > 0:05). ...
Background:
Post-burn hypertrophic scars commonly occur after burns. Studies that compare dermal substitutes with other treatment methods are insufficient. The purpose was to analyze the histopathological differences in hypertrophic burn scars after Matriderm®+split-thickness skin graft (STSG) and compare with AlloDerm®+STSG, STSG, full-thickness skin graft (FTSG), and normal skin.
Methods:
Samples of unburned, normal skin and deep 2nd or 3rd degree burns were obtained from patients who experienced a burn injury in the past to at least 6 months before biopsy, which was performed between 2011 and 2012. All subjects received >6 months of treatment before the biopsy. Intervention groups were normal (63), STSG (28), FTSG (6), Matriderm® (11), and AlloDerm® (18). Immunohistochemical analyses of elastin, collagen I, collagen III, cluster of differentiation 31 (CD31), smooth muscle actin (α-SMA), and laminin from scar and control tissues were performed and compared.
Results:
α-SMA vascular quantity and vessel width, stromal CD31, and basement membrane laminin expression were not significantly different between normal and intervention groups. Matriderm® group showed no significant difference in elastin, collagen III, stromal CD31 and α-SMA, CD31 vessel width, stromal α-SMA, vessel quantity and width, and laminin length compared to the normal group, meaning they were not significantly different from the normal skin traits.
Conclusion:
Dermal substitutes may be an optimal alternative to address the cosmetic and functional limitations posed by other treatment methods.
... These models are currently used to collect (pre-)clinical data, which can, in turn, be used for the systems biology approach in order to develop representative models (for example, dynamic mathematical models). [51][52][53] However, existing skin models still do not contain all kinds of skin cell types and do not provide original skin physiology. Three dimensional (3D) bioprinting, a computer-aided manufacturing process, provides sequential layer-by-layer delivery of vital cells within hydrogel-based scaffolds (i.e., 'bioink') to form a solid 3D structure. ...
A burn wound is a complex systemic disease at multiple levels. Current knowledge of scar formation after burn injury has come from traditional biological and clinical studies. These are normally focused on just a small part of the entire process, which has limited our ability to sufficiently understand the underlying mechanisms and to predict systems behaviour. Scar formation after burn injury is a result of a complex biological system—wound healing. It is a part of a larger whole. In this self-organising system, many components form networks of interactions with each other. These networks of interactions are typically non-linear and change their states dynamically, responding to the environment and showing emergent long-term behaviour. How molecular and cellular data relate to clinical phenomena, especially regarding effective therapies of burn wounds to achieve minimal scarring, is difficult to unravel and comprehend. Complexity science can help bridge this gap by integrating small parts into a larger whole, such that relevant biological mechanisms and data are combined in a computational model to better understand the complexity of the entire biological system. A better understanding of the complex biological system of post-burn scar formation could bring research and treatment regimens to the next level. The aim of this review/position paper is to create more awareness of complexity in scar formation after burn injury by describing the basic principles of complexity science and its potential for burn care professionals.
... The authors showed that they could mimic the formation of HTS and functionally evaluated the model with known therapeutic compounds. 60 Nonetheless the lack of inflammatory and vascular components in organotypic culture models continues to be a barrier in the construction of an ideal in vitro model. ...
Significance: The cutaneous repair process naturally results in different types of scarring that are classified as normal or pathological. Affected individuals are often affected from an esthetic, physical (functional), and psychosocial perspective. The distinct nature of scarring in humans, particularly the formation of pathological scars, makes the study of skin scarring a challenge for researchers in this area. Several established experimental models exist for studying scar formation. However, the increasing development and validation of newly emerging models have made it possible to carry out studies focused on different variables that influence this unique process. Recent Advances: Experimental models such as in vitro, ex vivo, and in vivo models have obtained different degrees of success in the reproduction of the scar formation in its native milieu and true environment. These models also differ in their ability to elucidate the molecular, cellular, and structural mechanisms involved in scarring, as well as for testing new agents and approaches for therapies. The models reviewed here, including cells derived from human skin and in vivo animal models, have contributed to the advancement of skin scarring research. Critical Issues and Future Directions: The absence of experimental models that faithfully reproduce the typical characteristics of the different types of human skin scars makes the improvement of validated models and the establishment of new ones a critical unmet need. The fields of wound healing research combined with tissue engineering have offered newer alternatives for experimental studies with the potential to provide clinically useful knowledge about scar formation.
