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An argon‐based low‐temperature plasma jet (LTPJ) was used to treat chronically infected wounds in Staphylococcus aureus‐laden mice. Based on physicochemical property analysis and in vitro antibacterial experiments, the effects of plasma parameters on the reactive nitrogen and oxygen species (RNOS) content and antibacterial capacity were determined, and the optimal treatment parameters were determined to be 4 standard litre per minute and 35 W. Additionally, the plasma‐treated activation solution had a bactericidal effect. Although RNOS are related to the antimicrobial effect of plasma, excess RNOS may be detrimental to wound remodelling. In vivo studies demonstrated that medium‐dose LTPJ promoted MMP‐9 expression and inhibited bacterial growth during the early stages of healing. Moreover, LTPJ increased collagen deposition, reduced inflammation, and restored blood vessel density and TGF‐β levels to normal in the later stages of wound healing. Therefore, when treating chronically infected wounds with LTPJ, selecting the medium dose of plasma is more advantageous for wound recovery. Overall, our study demonstrated that low‐temperature plasma jets may be a potential tool for the treatment of chronically infected wounds. [ABSTRACT FROM AUTHOR]

Skin injuries can have unexpected surfaces, leading to uneven wound surfaces and inadequate dressing contact with these irregular surfaces. This can decrease the dressing's haemostatic action and increase the healing period. This study recommends the use of sticky and flexible cryogel coverings to promote faster haemostasis and efficiently handle uneven skin wounds. Alginate cryogels have a fast haemostatic effect and shape flexibility due to their macroporous structure. The material demonstrates potent antibacterial characteristics and enhances skin adherence by adding grafted chitosan with gallic acid. In irregular defect wound models, cryogels can cling closely to uneven damage surfaces due to their amorphous nature. Furthermore, their macroporous structure allows for quick haemostasis by quickly absorbing blood and wound exudate. After giving the dressing a thorough rinse, its adhesive strength reduces and it is simple to remove without causing any damage to the wound. Cryogel demonstrated faster haemostasis than gauze in a wound model on a rat tail, indicating that it has considerable potential for use as a wound dressing in the biomedical area. [ABSTRACT FROM AUTHOR]

A wide variety of biomaterials has been developed to assist in wound healing, including acellular animal and human‐derived protein matrices. However, millions of patients worldwide still suffer from non‐healing chronic wounds, demonstrating a need for further innovation in wound care. To address this need, a novel biomaterial, the human keratin matrix (HKM), was developed, characterised, and tested in vitro and in vivo. HKM was found to be degradation‐resistant, and a proteomics analysis showed it to be greater than 99% human keratin proteins. PCR revealed adult human epidermal keratinocytes (HEKa) grown in contact with HKM showed increased gene expression of keratinocyte activations markers such as Epidermal Growth Factor (EGF). Additionally, a cytokine microarray demonstrated culture on HKM increased the release of cytokines involved in wound inflammatory modulation by both HEKa cells and adult human dermal fibroblasts (HDFa). Finally, in a murine chronic wound model, full‐thickness wounds treated weekly with HKM were smaller through the healing process than those treated with human amniotic membrane (AM), bovine dermis (BD), or porcine decellularized small intestinal submucosa (SIS). HKM‐treated wounds also closed significantly faster than AM‐ and SIS‐treated wounds. These data suggest that HKM is an effective novel treatment for chronic wounds. [ABSTRACT FROM AUTHOR]

Various preclinical and clinical studies have demonstrated the robust wound healing capacity of the natural anticoagulant activated protein C (APC). A bioengineered APC variant designated 3K3A‐APC retains APC's cytoprotective cell signalling actions with <10% anticoagulant activity. This study was aimed to provide preclinical evidence that 3K3A‐APC is efficacious and safe as a wound healing agent. 3K3A‐APC, like wild‐type APC, demonstrated positive effects on proliferation of human skin cells (keratinocytes, endothelial cells and fibroblasts). Similarly it also increased matrix metollaproteinase‐2 activation in keratinocytes and fibroblasts. Topical 3K3A‐APC treatment at 10 or 30 μg both accelerated mouse wound healing when culled on Day 11. And at 10 μg, it was superior to APC and had half the dermal wound gape compared to control. Further testing was conducted in excisional porcine wounds due to their congruence to human skin. Here, 3K3A‐APC advanced macroscopic healing in a dose‐dependent manner (100, 250 and 500 μg) when culled on Day 21. This was histologically corroborated by greater collagen maturity, suggesting more advanced remodelling. A non‐interference arm of this study found no evidence that topical 3K3A‐APC caused either any significant systemic side‐effects or any significant leakage into the circulation. However the female pigs exhibited transient and mild local reactions after treatments in week three, which did not impact healing. Overall these preclinical studies support the hypothesis that 3K3A‐APC merits future human wound studies. [ABSTRACT FROM AUTHOR]

Dendrobium officinale Kinura et Migo (DOKM) has a variety of medicinal applications; however, its ability to promote wound healing has not been previously reported. The purpose of this study is to investigate the proliferative phase of the wound−healing effect of DOKM glycoprotein (DOKMG) in rats and to elucidate its mechanism of action in vitro. In the present study, the ointment mixture containing DOKMG was applied to the dorsal skin wounds of the full‐thickness skin excision rat model, and the results showed that the wound healing speed was faster in the proliferative phase than vaseline. Histological analysis demonstrates that DOKMG promoted the re‐epithelialization of wound skin. Immunofluorescence staining and quantitative polymerase chain reaction assays revealed that DOKMG promotes the secretion of Fibronectin and inhibits the secretion of Collagen IV during the granulation tissue formation period, indicating that DOKMG could accelerate the formation of granulation tissue by precisely regulating extracellular matrix (ECM) secretion. In addition, we demonstrated that DOKMG enhanced the migration and proliferation of fibroblast (3T6 cell) in two‐dimensional trauma by regulating the secretion of ECM, via a mechanism that may implicate the AKT and JAK/STAT pathways under the control of epidermal growth factor receptor (EGFR) signalling. In summary, we have demonstrated that DOKMG promotes wound healing during the proliferative phase. Therefore, we suggest that DOKMG may have a potential therapeutic application for the treatment and management of cutaneous wounds. [ABSTRACT FROM AUTHOR]

Delayed tissue repair in the aged presents a major socio‐economic and clinical problem. Age‐associated delay in wound healing can be attributed to multiple factors, including an increased presence of senescent cells persisting in the wound. Although the transient presence of senescent cells is physiologic during the resolution phase of normal healing, increased senescent cell accumulation with age can negatively impact tissue repair. The objective of the study was to test interventional strategies that could mitigate the negative effect of senescent cell accumulation and possibly improve the age‐associated delay in wound healing. We utilised a 3D in vitro senescent fibroblast populated collagen matrix (FPCM) to study cellular events associated with senescence and delayed healing. Senescent fibroblasts showed an increase in anti‐apoptotic B‐cell lymphoma 2 (BCL‐2) family proteins. We hypothesized that reducing the senescent cell population and promoting non‐senescent cell functionality would mitigate the negative effect of senescence and improve healing kinetics. BCL‐2 inhibition and mitogen stimulation (FGF2) improved healing in the in vitro senescent models. These results were confirmed with an ex vivo human skin biopsy model. These data suggested that modulation of the senescent cell population with soluble factors improved the healing outcome in our in vitro and ex vivo healing models. [ABSTRACT FROM AUTHOR]

Recent evidence suggests that protecting human amnion and chorion matrices (HACM) during processing enhances the performance of HACM for wound repair and tissue regeneration. We utilised a diabetic (db/db) delayed wound healing mouse model. Treatment of db/db full‐thickness excisional wounds with HACM, processed with a polyampholyte preservative accentuated the proliferative phase of wound healing that decreased the time necessary to heal wounds. Polyampholyte protection improved the preservation of growth factors and cytokines during room temperature storage following E‐beam sterilisation and improved its function in wound healing applications. Our findings indicate protected HACM tissue up‐regulated MIP2, NF‐kB, TNF‐α, KI‐67, and Arg1 (0.6‐fold to 1.5‐fold) but those changes were not statistically significant. Immunofluorescent assessment identifying cell activity illustrated an induction of the proliferative phase of wound healing and a switch from an inflammatory macrophage phenotype (M1) to a pro‐regenerative macrophage phenotype (M2a). Genomic profiling of 282 genes was performed using Nanostring from co‐cultures of human macrophages and fibroblasts. The polyampholyte + HACM‐treated group, compared with the HACM or polyampholyte alone groups, had a statistically significant up‐regulation (32–368 fold) of 12 genes primarily involved in macrophage plasticity including CLC7, CD209, CD36, HSD11B1, ICAM1, IL1RN, IL3RA, ITGAX, LSP1, and PLXDC2 (adj. p‐value < 0.05). The polyampholyte alone group demonstrated statistically significant down‐regulation of four genes ADRA2, COL7A1, CSF3, and PTGS2 (adj. p < 0.05). The HACM alone group up‐regulated four genes ATG14, CXCL11, DNMT3A, and THBD, but the results were not statistically significant. Biomechanical measurements indicated that wounds treated with polyampholyte‐protected HACM had more tensile integrity compared with wounds treated with HACM alone. These findings indicate that better protection of HACM during processing stabilises the HACM matrix, which may lead to improved wound healing outcomes. [ABSTRACT FROM AUTHOR]

A new in vitro chronic wound biofilm model was recently published, which provided a layered scaffold simulating mammalian tissue composition on which topical wound care products could be tested. In this paper, we updated the model even further to mimic the dynamic influx of nutrients from below as is the case in a chronic wound. The modified in vitro model was created using collagen instead of agar as the main matrix component and contained both Staphylococcus aureus and Pseudomonas aeruginosa. The model was cast in transwell inserts and then placed in wound simulating media, which allowed for an exchange of nutrients and waste products across a filter. Three potential wound care products and chlorhexidine digluconate 2% solution as a positive control were used to evaluate the model. The tested products were composed of hydrogels made from completely biodegradable starch microspheres carrying different active compounds. The compounds were applied topically and left for 2–4 days. Profiles of oxygen concentration and pH were measured to assess the effect of treatments on bacterial activity. Confocal microscope images were obtained of the models to visualise the existence of microcolonies. Results showed that the modified in vitro model maintained a stable number of the two bacterial species over 6 days. In untreated models, steep oxygen gradients developed and pH increased to >8.0. Hydrogels containing active compounds alleviated the high oxygen consumption and decreased pH drastically. Moreover, all three hydrogels reduced the colony forming units significantly and to a larger extent than the chlorhexidine control treatment. Overall, the modified model expressed several characteristics similar to in vivo chronic wounds. [ABSTRACT FROM AUTHOR]

Immunosuppressive medications, which interfere with the activation and proliferation of T and B cells, increase the risk of wound healing complications. To address it, this study aimed to validate the feasibility of drug suspending during wound healing, whilst exploring the mechanisms exerted by T cells, which are important in the wound healing process. For this, a mouse skin wound model was set up. Tacrolimus (FK506) and fingolimod (FTY720) were both administered intraperitoneally prior to wounding to inhibit the T cell activation and migration, respectively. Flow‐cytometric analysis subsequently revealed the functional T cell subtypes detected during the healing process. A CD8a antibody was also administered to deplete CD8+ T cells in vivo to verify their specific function. It was found that FK506 or FTY720 administration delayed the early phase of wound healing by reducing collagen production, which was also supported by the downregulation of col1a1, col3a1 and tgfb1. However, there was no significant difference in the total healing period. Both spleen‐ and skin‐derived CD8+ T cells were proliferated and activated after injury without intervention, whereas CD4+ T cells showed no significant changes. Furthermore, selectively depleting CD8+ T cells retarded the healing process by downregulating collagen production‐associated genes (col1a1, col3a1, tgfβ1 and en1) and proteins (collagen type 1 and 3). In addition, the CD8a antibody decreased the expression of genes lta, tnfa, il13 and il13ra, and protein interleukin‐13Rα. In conclusion, suspending immunosuppressive drugs during wound healing was shown to be feasible through restraining the migration of activated T cells. CD8+ T cells represented the primary functional subtype positively associated with wound healing. [ABSTRACT FROM AUTHOR]

The venous lymph node flap (vLNF) is regarded as an arterialized or 'pure' venous flap, subject to which type of vessel provides blood supply to the flap. However, whether the vLNF drainage function is maintainable or sufficient for the treatment of lymphoedema is yet to be elucidated. In this study, the venous flow‐through lymph node flaps or 'pure' vLNF was investigated in a rat's tail (lymphoedema model), and the therapeutic effect was analysed to clarify its potential value for the treatment of lymphoedema. The subjects (rats) were divided into three groups, the experimental group ('pure' vLNF group), the control group (lymphoedema group), and the baseline group. The tail circumference and diameter were recorded for 5 weeks after the operation. Five weeks after surgery, the lymphatic drainage function was evaluated using indocyanine green (ICG) lymphography, while the skin thickness and collagen layer were assessed by histological analysis. The average diameter and circumference of the experimental group were significantly shorter than the control group (p < 0.01). When comparing ICG lymphography within the three groups, the baseline group showed that the ICG entered the inguinal lymph node and appeared in the liver while the ICG still accumulated in the oedematous tail with no fluorescence emerging in other parts of the rat's body, in the control group. In the experimental group, the ICG entered the vLNF via the flap junction with the rat's tail and fluorescence emerging in the rat's liver. When comparing the three groups above with the control group, the experimental group's skin and collagen layer thickness were significantly improved (p < 0.01). Lymphatic recanalization was shown to appear between 'pure' vLNF and the recipient area. Moreover, vLNF was shown to drain lymph fluid via the venous system, thus relieved swelling and fibrosis of the lymphoedematous tissue. Consequently, this could represent a new potential surgical approach for the treatment of lymphoedema. [ABSTRACT FROM AUTHOR]

Accurate burn depth assessment is crucial to determine treatment plans for burn patients. We have previously proposed a method for performing burn depth assessments based on photoacoustic (PA) imaging, and we have demonstrated the validity of this method, which allows the successful detection of PA signals originating from the blood under the bloodless burned tissue, using rat burn models. Based on these findings, we started a clinical study in which we faced two technical issues: (1) When the burn depth was shallow, PA signals due to skin contamination and/or melanin in the epidermis (surface signals) could not be distinguished from PA signals originating from the blood in the dermis; (2) the size of the system was too large. To solve these issues, we propose a burn depth diagnosis based on dual‐wavelength light emitting diodes (LEDs)‐excited PA imaging. The use of LEDs rendered the system compact compared to the previous one that used a conventional solid‐state laser. We replicated human burned skin by applying a titrated synthetic melanin solution onto the wound surface in albino rat burn models and measured their burn depths by PA excitation at 690 and 850 nm, where melanin and haemoglobin show greatly different absorption coefficients. As a result, the surface signals were eliminated by subtracting the PA signals at 690 nm from those at 850 nm. The resultant estimated burn depths were strongly correlated with the histological assessment results. The validity of the proposed method was also examined using a burn model of rats with real melanin. [ABSTRACT FROM AUTHOR]

Wound dehiscence, oftentimes a result of the poor tensile strength of early healing wounds, is a significant threat to the post‐operative patient, potentially causing life‐threatening complications. Vanadate, a protein tyrosine phosphatase inhibitor, has been shown to alter the organisation of deposited collagen in healing wounds and significantly improve the tensile strength of incisional wounds in rats. In this study, we sought to explore the effects of locally administered vanadate on tensile strength and collagen organisation in both the early and remodelling phases of excisional wound healing in a murine model. Wild‐type mice underwent stented excisional wounding on their dorsal skin and were divided equally into three treatment conditions: vanadate injection, saline injection control and an untreated control. Tensile strength testing, in vivo suction Cutometer analysis, gross wound measurements and histologic analysis were performed during healing, immediately upon wound closure, and after 4 weeks of remodelling. We found that vanadate treatment significantly increased the tensile strength of wounds and their stiffness relative to control wounds, both immediately upon healing and into the remodelling phase. Histologic analysis revealed that these biomechanical changes were likely the result of increased collagen deposition and an altered collagen organisation composed of thicker and distinctly organised collagen bundles. Given the risk that dehiscence poses to all operative patients, vanadate presents an interesting therapeutic avenue to improve the strength of post‐operative wounds and unstable chronic wounds to reduce the risk of dehiscence. [ABSTRACT FROM AUTHOR]

The process of wound healing is critical to maintaining homeostasis after injury. Although a considerable amount has been learned about this complex process, much remains unknown. Whereas, studies with human volunteers are ideal given the unique nature of the human skin anatomy and immune system, the lack of such clinical access has made animal models prime candidates for use in preclinical studies. This review aims to discuss the strengths and limitations of the commonly used mammalian species in wound healing studies: murine, rabbit and porcine. Thereafter, a survey of models of various acute wounds such as cutaneous, ear, and implant are presented and representative studies that use them are described. This review is intended to acquaint readers with the vast spectrum of models available, each of which has a distinct utility. At the same time, it highlights the importance of utilising clinical samples to complement investigations conducted in animal models. Through this strategy, it is hoped that forthcoming research may be more reflective of the acute wound healing process as it occurs in humans. [ABSTRACT FROM AUTHOR]

Why only certain species can regenerate their appendages (e.g. tails and limbs) remains one of the biggest mysteries of nature. Unlike anuran tadpoles and salamanders, humans and other mammals cannot regenerate their limbs, but can only regrow lost digit tips under specific circumstances. Numerous hypotheses have been postulated to explain regeneration‐incompetency in mammals. By studying model organisms that show varying regenerative abilities, we now have more opportunities to uncover what contributes to regeneration‐incompetency and functionally test which perturbations restore appendage regrowth. Particularly, Xenopus laevis tail and limb, and mouse digit tip model systems exhibit naturally occurring variations in regenerative capacities. Here, we discuss major hypotheses that are suggested to contribute to regeneration‐incompetency, and how species with varying regenerative abilities reflect on these hypotheses. [ABSTRACT FROM AUTHOR]

Infection is a major source of complications in delayed diabetic wound healing. Increased understanding of differential bacterial responses to diabetic wounds will enable us to better understand chronic wound pathogenesis. Here we create delayed‐healing wounds infected with Staphylococcus aureus in non‐diabetic and diabetic mice and used RNA‐seq to compare bacterial gene expression profiles 3 or 7 days after infection. Analysis at day 3 demonstrated substantial transcriptomic differences between bacteria colonising non‐diabetic and diabetic wound beds. Most of these transcriptional differences resolved by day 7, suggesting normalisation of many bacterial phenotypes later in the diabetic wound healing process. Lingering differentially expressed genes at day 7 were enriched for genes related to carbohydrate metabolism, which includes genes of the lac operon, and capsular polysaccharide synthesis, which includes the cap8 locus. These data encourage further research into host–pathogen interactions in wound healing and how they influence differential outcomes in the diabetic wound environment. [ABSTRACT FROM AUTHOR]

Cutaneous wound healing is an intricate orchestration of three overlapping phases of repair that encompass numerous cell types, signalling cascades, and microenvironment modifications to reach a successful resolution. Disruption of any of these steps will create an abnormal healing response resulting in either ulceration or excessive scarring. It has become evident that the extracellular matrix and its associated components are key orchestrators during this process. One of these essential matrix proteins is decorin, a small leucine‐rich proteoglycan (SLRP) that acts as a regulator of collagen fibrillogenesis and a non‐competitive inhibitor of multiple growth factors signalling cascades. Decorin is a necessary shut‐off switch for the pro‐reparative mechanism of the tissue replacement phase and limits the occurrence of hypertrophic scarring by preventing excessive repair. We investigated the use of decorin as a therapeutic by administering the matrix protein anchored in a slow‐release coacervate in a hypertrophic scarring mouse model. The results show that early wound healing phase measurements exhibit little difference in performance compared to our coacervate‐only baseline or HB‐EGF‐treated control mice. However, during the resolution phase of wound healing, the decorin‐treatment significantly reduces cutaneous thickness, enhances collagen alignment, and improves overall wound scoring in the mice. Thus, mice treated with decorin display better healing outcomes and could limit the hypertrophic scarring phenotype in the coacervate only, and HB‐EGF controls. These results suggest that decorin may be a promising tool and alternative therapy for patients who suffer from over‐exuberant matrix deposition during wound healing. [ABSTRACT FROM AUTHOR]

The US Food and Drug Administration (FDA) understands that innovative product development is essential to addressing the unmet medical need of non-healing chronic wounds. Barriers to product development for non-healing chronic wounds may involve but are not limited to a dearth of biological models, challenges in drug delivery, challenges in clinical trial execution, and limited commercial viability. This perspective article discusses FDA's renewed focus on non-healing chronic wounds and outlines efforts to address identified barriers to product development for nonhealing chronic wounds. In collaboration with key wound healing stakeholders including academia, professional associations, patient groups, reimbursement organizations and industry, FDA intends to help advance product development for non-healing chronic wounds for the ultimate betterment of patients. [ABSTRACT FROM AUTHOR]

Maximum survival area after perforator flap elevation is mainly achieved through vasodilation and angiogenesis, and endothelial Ca2+ signals play a pivotal role in both of them. Transient receptor potential (TRP) channels modulate many endothelial cell functions via mediating the extracellular Ca2+ entry. This study aims to investigate the correlation of TRPV4, TRPV1, and TRPA1 with vascular change after the inferior gluteal artery perforator flap elevation. A total of 50 adult male SD rats were used in this study. Ten rats were used in the part one to assess the flap viability on postoperative day 7. Twenty rats were used in the part two to evaluate blood flow change after flap elevation. The correlation of vascular change with TRPV1, TRPV4, and TRPA1 protein changes was investigated in 20 rats in the part three. The mean flap survival area percentage was 55 ± 5.7%. Blood flow in the overall flap and Zone II after the flap elevation markedly increased from the postoperative day 3. The most marked change of the vasodilation occurred on Days 3 and 5 after flap elevation. The angiogenesis occurred on Day 5 after flap elevation and the microvessel density peaked also on Day 5. Moreover, TRPA1 expression showed a trend towards continuous reduction over time. The expression of TRPV1 and TRPV4 reached the peak value on Day 3. The endothelial NO synthase expression showed an increasing trend at first, followed by a reduction over time, while VEGF expression reached the peak value on Day 3. The vascular changes after flap elevation might be associated with the changes in TRPV4, TRPV1, and TRPA1. [ABSTRACT FROM AUTHOR]

Biological scaffolds such as hydrogels provide an ideal, physio-mimetic of native extracellular matrix (ECM) that can improve wound healing outcomes after cutaneous injury. While most studies have focused on the benefits of hydrogels in accelerating wound healing, there are minimal data directly comparing different hydrogel material compositions. In this study, we utilized a splinted excisional wound model that recapitulates human-like wound healing in mice and treated wounds with three different collagen hydrogel dressings. We assessed the feasibility of applying each dressing and performed histologic and histopathologic analysis on the explanted scar tissues to assess variations in collagen architecture and alignment, as well as the tissue response. Our data indicate that the material properties of hydrogel dressings can significantly influence healing time, cellular response, and resulting architecture of healed scars. Specifically, our pullulan-collagen hydrogel dressing accelerated wound closure and promoted healed tissue with less dense, more randomly aligned, and shorter collagen fibres. Further understanding of how hydrogel properties affect the healing and resulting scar architecture of wounds may lead to novel insights and further optimization of the material properties of wound dressings. [ABSTRACT FROM AUTHOR]

Critical limb ischemia is a serious form of peripheral arterial disease (PAD). The consequences of lower limb ischemia are pain, claudication and chronic non‐healing wounds. Patients with diabetes are especially at a high risk for developing non‐healing ulcers. The most serious complication is major amputation. For this reason, there is a significant medical requirement to develop new therapies in order to prevent the progression of PAD. For research purposes, it is crucial to find an appropriate model of chronic ischemia to explore the processes of wound healing. According to recently acquired information, rodents are currently the most commonly used animals in these types of studies. The main advantage of using small animals is the low financial cost due to the relatively small demand for food, water and living space. The disadvantage is their anatomy, which is different from that of humans. Larger animals have a more human‐like anatomy and physiology, but they require more expense and space for housing. A bipedicle skin flap and its modifications are popular models for ischemic wounds. In order to secure healing through re‐epithelisation, as opposed to contraction in rodents, there is a need to remove the panniculus carnosus muscle. Wounds in other experimental animals heal primarily through re‐epithelisation. The application of a silicone mesh underneath the flap prevents vascular regrowth in ischemic tissue. There is an ongoing effort to create in vivo diabetic models for chronic ulcer research. This work presents an overview of existing animal models of ischemic wounds. [ABSTRACT FROM AUTHOR]

The application of photobiomodulation (PBM) in regenerative medicine has expanded to the treatment of alopecia caused by various reasons. However, the mechanisms responsible for its effects are poorly understood. Here, we aimed to investigate the effects of PBM on hair regeneration in injured skin and to explore the underlying mechanisms. The scratched epidermis or dermis models were established in C57 mice aged 7–8 weeks. We found that the scratched epidermis had no influence on hair regeneration, but the scratched dermis led to obvious hair follicle atrophy and significantly influenced hair regeneration. The wounds in scratched dermis models were treated with PBM (655 nm, 3 J/cm2 [10 min]) and the hair regeneration and cell proliferation in hair follicle were evaluated. Compared with control, the hair coverage level was significantly enhanced after PBM treatment. Sox9+ and PCNA+ cells in hair follicle were obviously observed in PBM‐treated group, but not in control. In vitro, the effects of PBM on the function of dermal papilla cells (DPCs) were investigated. The results showed that the migration of DPCs was increased significantly by PBM (655 nm, 3 J/cm2 [10 min]), whereas no effect was found on proliferation. Furthermore, we found that PBM promoted exosome secretion of DPCs, accompanied by the activation of Akt/GSK‐3β/β‐catenin pathway. AKT inhibitor MK‐2206 effectively blocked PBM‐induced migration and exosome secretion of DPCs. These findings suggest that the enhanced migration and exosome secretion of DPCs mediated by the Akt/GSK‐3β/β‐catenin pathway were responsible for the promotion of hair regeneration in injured skin by PBM. [ABSTRACT FROM AUTHOR]

In the skin‐healing field, porcine models are regarded as a useful analogue for human skin due to their numerous anatomical and physiological similarities. Despite the widespread use of porcine models in skin healing studies, the initial origin, recruitment and transition of fibroblasts to matrix‐secreting contractile myofibroblasts are not well defined for this model. In this review, we discuss the merit of the pig as an animal for studying myofibroblast origin, as well as the challenges associated with assessing their contributions to skin healing. Although a variety of wound types (incisional, partial thickness, full thickness, burns) have been investigated in pigs in attempts to mimic diverse injuries in humans, direct comparison of human healing profiles with regards to myofibroblasts shows evident differences. Following injury in porcine models, which often employ juvenile animals, myofibroblasts are described in the developing granulation tissue at 4 days, peaking at Days 7–14, and persisting at 60 days post‐wounding, although variations are evident depending on the specific pig breed. In human wounds, the presence of myofibroblasts is variable and does not correlate with the age of the wound or clinical contraction. Our comparison of porcine myofibroblast‐mediated healing processes with those in humans suggests that further validation of the pig model is essential. Moreover, we identify several limitations evident in experimental design that need to be better controlled, and standardisation of methodologies would be beneficial for the comparison and interpretation of results. In particular, we discuss anatomical location of the wounds, their size and depth, as well as the healing microenvironment (wet vs. moist vs. dry) in pigs and how this could influence myofibroblast recruitment. In summary, although a widespread model used in the skin healing field, further research is required to validate pigs as a useful analogue for human healing with regards to myofibroblasts. [ABSTRACT FROM AUTHOR]

This study aimed to establish a novel gouty ulcer rat model induced by monosodium urate (MSU) deposition and preliminarily explored how MSU crystals affected wound healing. MSU crystals were subcutaneously injected into the back of rats to simulate tophi formation and ulceration. Ultrasound was used to detect the formation of gouty tophi. MSU crystal deposition and histopathological changes were analysed by haematoxylin–eosin staining. After the skin over the tophi became broken in the model group, a full‐thickness tissue defect of the same area was made on the backs of the phosphate buffered saline (PBS) controls. On Days 3, 7, and 14 after wounding, the infiltration of neutrophils and macrophages and the expression of inflammatory markers, including interleukin‐1β (IL‐1β), tumour necrosis factor‐α (TNF‐α), and Nod‐like receptor protein 3 (NLRP3), were examined by immunohistochemical staining and Western blotting, respectively. After the first subcutaneous injection in rats, local tissues showed redness and swelling, indicating inflammation on approximately Day 14. Tophi‐like manifestations appeared on approximately Day 18. Tophi appeared heterogeneously hyperechoic by ultrasound. Swelling and redness in injured tissue areas increased on approximately Day 22, skin tissue necrosis was seen in a small area on approximately Day 26, and skin necrosis was enlarged and the tophi were ulcerated on approximately Day 32, accompanied by yellowish‐white, chalky secretions. Haematoxylin and eosin staining showed dermal deposition of needle‐like crystals with surrounding granulomatous inflammation. On Days 3, 7, and 14 after wounding, immunohistochemical staining showed the infiltration of neutrophils and macrophages, and the expression of inflammation‐related proteins (IL‐1β, TNF‐α, and NLRP3) were upregulated in gouty ulcers compared with those of PBS controls. The gouty ulcers were not completely healed by Day 14 compared with those in the PBS controls. In this study, a novel gouty ulcer rat model was constructed, which also revealed the existence of persistent chronic inflammation. [ABSTRACT FROM AUTHOR]

Negative pressure wound therapy (NPWT) is used clinically to promote tissue formation and wound closure. In this study, a porcine wound model was used to further investigate the mechanisms as to how NPWT modulates wound healing via utilization of a form of NPWT called the vacuum‐assisted closure. To observe the effect of NPWT more accurately, non‐NPWT control wounds containing GranuFoam™ dressings, without vacuum exposure, were utilized. In situ histological analysis revealed that NPWT enhanced plasma protein adsorption throughout the GranuFoam™, resulting in increased cellular colonization and tissue ingrowth. Gram staining revealed that NPWT decreased bacterial dissemination to adjacent tissue with greater bacterial localization within the GranuFoam™. Genomic analysis demonstrated the significant changes in gene expression across a number of genes between wounds treated with non‐NPWT and NPWT when compared against baseline tissue. However, minimal differences were noted between non‐NPWT and NPWT wounds, including no significant differences in expression of collagen, angiogenic, or key inflammatory genes. Similarly, significant increases in immune cell populations were observed from day 0 to day 9 for both non‐NPWT and NPWT wounds, though no differences were noted between non‐NPWT and NPWT wounds. Furthermore, histological analysis demonstrated the presence of a foreign body response (FBR), with giant cell formation and encapsulation of GranuFoam™ particles. The unique in situ histological evaluation and genomic comparison of non‐NPWT and NPWT wounds in this pilot study provided a never‐before‐shown perspective, offering novel insights into the physiological processes of NPWT and the potential role of a FBR in NPWT clinical outcomes. [ABSTRACT FROM AUTHOR]

Chronic wounds are a common and debilitating condition associated with aging populations that impact more than 6.5 million patients in the United States. We have previously demonstrated the efficacy of daily topical 1% valsartan in treating wounds in diabetic mouse and pig models. Despite these promising results, there remains a need to develop an extended‐release formulation that would reduce patient burden by decreasing the frequency of daily applications. Here, we used nanotechnology to self‐assemble valsartan amphiphiles into a filamentous structure (val‐filaments) that would serve as a scaffold in wound beds and allow for steady, localised and tunable release of valsartan amphiphiles over 24 days. Two topical treatments of this peptide‐based hydrogel on full‐thickness wounds in Zucker Diabetic Fatty rats resulted in faster rates of wound closure. By day 23, all val‐filament treated wounds were completely closed, as compared to one wound closed in the placebo group. Mechanistically, we observed enrichment of proteins involved in cell adhesion and energetics pathways, downregulation of Tgf‐β signalling pathway mediators (pSmad2, pSmad3 and Smad4) and increased mitochondrial metabolic pathway intermediates. This study demonstrates the successful synthesis of a sustained‐release valsartan filament hydrogel, its impact on mitochondrial energetics and efficacy in treating diabetic wounds. [ABSTRACT FROM AUTHOR]

Chronic wounds are a significant health problem worldwide. However, nothing is known about how chronic wounds initiate and develop. Here we use a chronic wound model in diabetic mice and a Systems Biology Approach using nanoString nCounter technology and weighted gene correlation network analysis (WGCNA), with tissues collected at 6, 12, 24 and 48 h post‐wounding, to identify metabolic signalling pathways involved in initiation of chronicity. Normalized counts obtained from the nanoString nCounter Mouse Metabolic Panel were used for the WGCNA, which groups genes into co‐expression modules to visualize the correlation network. Genes with significant module membership and gene trait significance (p < 0.05) were used to identify signalling pathways that are important for the development of chronicity. The pathway analysis using the Reactome database showed stabilization of PTEN, which down‐regulates PI3K/AKT1, which in turn down‐regulates Nrf2, as shown by ELISA, thus disabling antioxidant production, resulting in high oxidative stress levels. We find that pathways involved in inflammation, including those that generate pro‐inflammatory lipids derived from arachidonic acid metabolism, IFNγ and catecholamines, occur. Moreover, HIF3α is over‐expressed, potentially blocking Hif1α and preventing activation of growth factors and cytokines that promote granulation tissue formation. We also find that FGF1 is under‐expressed, while thrombospondin‐1 is over‐expressed, resulting in decreased angiogenesis, a process that is critical for healing. Finally, enzymes involved in glycolysis are down‐regulated, resulting in decreased production of pyruvate, a molecule critical for ATP production, leading to extensive cell death and wound paralysis. These findings offer new avenues of study that may lead to the development of novel treatments of CW to be administered right after debridement. [ABSTRACT FROM AUTHOR]

Microbial contamination of wounds is a significant problem that delays healing, particularly when bacterial biofilms are present. A novel combination of pectinic acid (PG) + caprylic acid (CAP) was previously found in vitro to be highly effective in eradicating various pathogens in biofilms with minimal cytotoxicity. In this study, a novel wound ointment was formulated with PG + CAP and first assessed in vitro using a well‐established biofilm eradication model. In vitro, the PG + CAP ointment was shown to be efficacious in reducing the microbial biofilms. This ointment was then tested in vivo in two pilot porcine wound healing models, with and without Staphylococcus aureus microbial challenge. Ointments were applied to each wound daily, and healing by wound closure area measurement was assessed weekly over 4 weeks. After 4 weeks, pigs were sacrificed and wounds were scored for reepithelialization, inflammation, granulation tissue, and collagen deposition. We compared PG + CAP to hydroxyethylcellulose + glycerol ointment base (control) and MediHoney (comparator). In the porcine microbial challenge model, the novel antimicrobial PG + CAP wound ointment rapidly eradicated bacterial organisms embedded in wounds, was safe and well‐tolerated, and was associated with enhanced healing compared to ointment base and MediHoney. Specifically, the cumulative histopathology, reepithelialization of epidermis, and mature granulation tissue in the wound bed was significantly better with PG + CAP than with control and MediHoney treatments. This ointment warrants further study as a non‐antibiotic ointment for use in treating a wide array of infected wounds. [ABSTRACT FROM AUTHOR]

Diabetes is a systemic disease in which the body cannot regulate the amount of sugar, namely glucose, in the blood. High glucose toxicity has been implicated in the dysfunction of diabetic wound healing, following insufficient production (Type 1) or inadequate usage (Type 2) of insulin. Chronic non‐healing diabetic wounds are one of the major complications of both types of diabetes, which are serious concerns for public health and can impact the life quality of patients significantly. In general, diabetic wounds are characterized by deficient chemokine production, an unusual inflammatory response, lack of angiogenesis and epithelialization, and dysfunction of fibroblasts. Increasing scientific evidence from available experimental studies on animal and cell models strongly associates impaired wound healing in diabetes with dysregulated fibroblast differentiation to myofibroblasts, interrupted myofibroblast activity, and inadequate extracellular matrix production. Myofibroblasts play an important role in tissue repair by producing and organizing extracellular matrix and subsequently promoting wound contraction. Based on these studies, hyperglycaemic conditions can interfere with cytokine signalling pathways (such as growth factor‐β pathway) affecting fibroblast differentiation, alter fibroblast apoptosis, dysregulate dermal lipolysis, and enhance hypoxia damage, thus leading to damaged microenvironment for myofibroblast formation, inappropriate extracellular matrix modulation, and weakened wound contraction. In this review, we will focus on the current available studies on the impact of diabetes on fibroblast differentiation and myofibroblast function, as well as potential treatments related to the affected pathways. [ABSTRACT FROM AUTHOR]

Myofibroblasts are mechanosensitive cells and a variety of their behaviours including differentiation, migration, force production and biosynthesis are regulated by the surrounding microenvironment. Engineered cell culture models have been developed to examine the effect of microenvironmental factors such as the substrate stiffness, the topography and strain of the extracellular matrix (ECM) and the shear stress on myofibroblast biology. These engineered models provide well‐mimicked, pathophysiologically relevant experimental conditions that are superior to those enabled by the conventional two‐dimensional (2D) culture models. In this perspective, we will review the recent advances in the development of engineered cell culture models for myofibroblasts and outline the findings on the myofibroblast mechanobiology under various microenvironmental conditions. These studies have demonstrated the power and utility of the engineered models for the study of microenvironment‐regulated cellular behaviours. The findings derived using these models contribute to a greater understanding of how myofibroblast behaviour is regulated in tissue repair and pathological scar formation. [ABSTRACT FROM AUTHOR]

Fibroblasts and myofibroblasts play a myriad of important roles in human tissue function, especially in wound repair and healing. Among all cells, fibroblasts are group of cells that decide the status of wound as they maintain tissue homeostasis. Currently, the increase in the deleterious effects of chronic wound and their morbidity rate has necessitated the need to understand the influence of fibroblasts and myofibroblasts, which chiefly originate locally from tissue‐resident fibroblasts to address the same. Wound pathophysiology is complex, herein, we have discussed fibroblast and myofibroblast heterogeneity in skin and different organs by understanding the phenotypical and functional properties of each of its sub‐populations in the process of wound healing. Recent advancements in fibroblast activation, differentiation to myofibroblasts, proliferation and migration are discussed in detail. Fibroblasts and myofibroblasts are key players in wound healing and wound remodelling, respectively, and their significance in wound repair is discussed. An increased understanding of their biology during wound healing also gives an opportunity to explore more of fibroblast and myofibroblast focused therapies to treat chronic wounds which are clinical challenges. In this regard, in the current review, we have described the different methods for isolation of primary fibroblasts and myofibroblasts from both animal models and humans, and their characterization. Additionally, we have also provided details on possible molecular targets for better understanding of prognosis, diagnosis and treatment of chronic wounds. Information will help both researchers and clinicians in providing molecular insight that enable them for effective chronic wound management. The knowledge of intimate dialogue between the fibroblast, sub‐populations like, myofibroblast and their microenvironment, will serve useful in determining novel, efficient and specific therapeutic targets to treat pathological wound conditions. [ABSTRACT FROM AUTHOR]

Negative pressure wound therapy is a widely used treatment for chronic, nonhealing wounds. Surprisingly, few studies have systematically evaluated the cellular and molecular effects of negative pressure treatment on human skin. In addition, no study to date has directly compared recently available single‐use negative pressure modalities to traditional negative pressure devices in a controlled setting. Here we developed a novel large‐scale ex vivo human skin culture system to effectively evaluate the efficacy of two different negative pressure wound therapy modalities. Single‐use and traditional negative pressure devices were applied to human ex vivo wounded skin sheets cultured over a period of 48 hours. Cellular tissue response to therapy was evaluated via a combination of histological analysis and transcriptional profiling, in samples collected from the wound edge, skin adjacent to the wound, and an extended skin region. Single‐use negative pressure wound therapy caused less damage to wound edge tissue than traditional application, demonstrated by improved skin barrier, reduced dermal‐epidermal junction disruption and a dampened damage response. Transcriptional profiling confirmed significantly less activation of multiple pro‐inflammatory markers in wound edge skin treated with single‐use vs traditional negative pressure therapy. These findings may help to explain the greater efficacy of sNPWT in the clinic, while offering a noninvasive system to develop improved NPWT‐based therapies. [ABSTRACT FROM AUTHOR]

Great progresses have been made in comprehension of tissue regeneration process. However, one of the central questions in regeneration research remains to be deciphered is what factors initiate regenerative process. In present study, we focused on systematic profiling of early regulators in tissue regeneration via high‐throughput screening on zebrafish caudal fin model. Firstly, 53 GO‐annotated regeneration‐related genes, which were specifically activated upon fin amputation, were identified according to the transcriptomic analysis. Moreover, qRT‐PCR analysis of a couple of randomly selected genes from the aforementioned gene list validated our sequencing results. These studies confirmed the reliability of transcriptome sequencing analysis. Fibroblast growth factor 20a (fgf20a) is a key initial factor in the regeneration of zebrafish. Through a gene expression correlation analysis, we discovered a collection of 70 genes correlating with fgf20a, whose expression increased promptly at 2 days post amputation (dpa) and went down to the basal level until the completion of fin regeneration. In addition, two genes, socs3b and nppc, were chosen to investigate their functions during the fin regeneration. Inhibition of either of those genes significantly delayed the regenerative process. Taken together, we provided a simple and effective time‐saving strategy that may serve as a tool for identifying early regulators in regeneration and identified 71 genes as early regulators of fin regeneration. [ABSTRACT FROM AUTHOR]

Complete extremity regeneration in mammals is restricted to distal amputations of the digit tip, the terminal phalanx (P3). In mice, P3 regeneration is mediated via the formation of a blastema, a transient population of progenitor cells that form from the blending of periosteal and endosteal/marrow compartmentalized cells that undergo differentiation to restore the amputated structures. Compartmentalized blastema cells are formed independently, and periosteal compartment‐derived cells are required for restoration of amputated skeletal length. P3 regenerative capacity is progressively attenuated at increasingly more proximal amputation levels, eventually resulting in regenerative failure. The continuum of regenerative capacity within the P3 wound milieu is a unique model to investigate mammalian blastema formation in response to distal amputation, as well as the healing response associated with regenerative failure at proximal amputation levels. We report that P3 proximal amputation healing, previously reported to result in regenerative failure, is not an example of complete regenerative failure, but instead is characterized by a limited bone regeneration response restricted to the endosteal/marrow compartment. The regeneration response is mediated by blastema formation within the endosteal/marrow compartment, and blastemal osteogenesis progresses through intramembranous ossification in a polarized proximal to distal sequence. Unlike bone regeneration following distal P3 amputation, osteogenesis within the periosteal compartment is not observed in response to proximal P3 amputation. We provide evidence that proximal P3 amputation initiates the formation of fibrotic tissue that isolates the endosteal/marrow compartment from the periosteal compartment and wound epidermis. While the fibrotic response is transient and later resolved, these studies demonstrate that blastema formation and fibrosis can occur in close proximity, with the regenerative response dominating the final outcome. Moreover, the results suggest that the attenuated proximal P3 regeneration response is associated with the absence of periosteal‐compartment participation in blastema formation and bone regeneration. [ABSTRACT FROM AUTHOR]

The process of reinnervation, an important component of functional restoration after flap transfer, is understudied, making it necessary to carry out a further investigation for delineation of the exact timeline. Seventy‐six Sprague‐Dawley rats were used as the experimental animals. An anatomic study was first conducted to clarify the pattern of nerve distribution in the dorsal skin of 16 rats. Afterward, a myocutaneous flap was harvested on the right flanks of 40 rats, which were then assigned into seven time points. At each time point, skin samples were harvested and immunofluorescent staining was performed using α‐Bungarotoxin, and antibodies against NF‐200, p75, α‐SMA, and TH. One‐way analysis of variance was adopted for comparison of nerve density after surgery. For evaluation of functional return, cutaneous trunci muscle reflex (CTMr) test was performed on 10 additional rats, and the Chi‐square test was used for comparison of reflex intensity among six time points after surgery. The outcomes revealed that the cutaneous branches from the intercostal nerves and the dorsothoracic nerve from the brachial plexus could be found entering the dorsal skin, distributed in the skin proper and the panniculus carnosus, respectively. After flap surgery, full spontaneous reinnervation of the skin proper and vessels within the flaps could be achieved at day 180. However, if the stumps of cutaneous branches of the intercostal nerves were damaged, the nerve density in the skin proper underwent a 2/3 decline. The panniculus carnosus in the cranial part had a much better reinnervation than that in the caudal part. The CTMr test showed that the flap could regain most of its sensate and motor activity. Our study shows that strong spontaneous reinnervation could be expected after flap surgery. The pattern of the original nerve distribution in both the recipient and donor sites may have a big impact on the reinnervation of the flap. [ABSTRACT FROM AUTHOR]

Impaired wound healing is a common complication of diabetes. In diabetic wounds, macrophages present dysfunctional efferocytosis and abnormal phenotypes, which could result in excessive neutrophil accumulation and prolonged inflammation, thereby eventually hindering wound repair. ANXA1 N‐terminal peptide Ac2‐26 exhibits a high potential in mitigating inflammation and improving repair; however, its efficacy in diabetic wound repair remains unclear. In this study, a cutaneous excisional wound model was built in genetically diabetic mice. Ac2‐26 or a vehicle solution was employed locally in wound sites. Subsequently, wound zones were measured and sampled at different time intervals post‐wounding. Using hematoxylin‐eosin and Masson's trichrome staining, we observed the histopathological variations and collagen deposition in wound samples. Based on immunohistochemistry and immunofluorescence, the numbers of neutrophils, macrophages, and CD206‐positive macrophages in the wound samples were determined. Cytokine expression in wound samples was studied by immunoblot assay. Results showed that Ac2‐26 treatment could facilitate diabetic wound closure, down‐regulate the number of neutrophils, and improve angiogenesis and collagen deposition. In addition, Ac2‐26 application expedited macrophage recruitment and up‐regulated the percentage of macrophages expressing CD206, which is a marker for M2 macrophages. Moreover, Ac2‐26 inhibited the expressions of TNF‐α and IL‐6 and up‐regulated the expressions of IL‐10, TGF‐β, and VEGFA during diabetic wound healing. Hence, based on the aforementioned findings, Ac2‐26 application in diabetic wounds could exert anti‐inflammatory and pro‐repair effects by reducing neutrophil accumulation and facilitating M2 macrophage development. [ABSTRACT FROM AUTHOR]

This study aimed to improve the conventional rat burn wound model and to validate its utility. In total, 60 Sprague‐Dawley rats were divided equally into the control and experimental groups. Altogether, 60 burn wound models with zones of stasis were created in each group. Gross visual assessments of the burn wounds were performed at 0, 24, and 48 hours after burn creation. The rates of necrosis in the zones of stasis were calculated, and the blood flow from the wounds was examined. Wound tissues were collected 48 hours after the burn and subjected to hematoxylin and eosin staining to determine whether the models were successfully established. The model success rates were calculated. The success rate of the burn wound models was significantly different between the control group and the experimental group (93.33% [56/60] vs 100%; P =.042). The Cronbach's alpha values and the respective correlation coefficients indicated that the stability of the zones of stasis in the models on the two sides of the spine was higher in the experimental group than in the control group. The standard deviations of the rate of necrosis, blood flow, and density of necrotic cells and apoptosis cell density, and inflammatory factor content in the zones of stasis were smaller in the experimental group than in the control group at 48 hours after model construction. This suggested that the stability of repeated procedures was higher in the experimental group than in the control group. The novel device for creating burns in animal models facilitated the effective creation of zones of stasis for rat burn wound models. Both the model success rate and stability were higher compared with the conventional model construction method. In addition, the use of the novel device can better align with the requirements of self‐controlled studies. [ABSTRACT FROM AUTHOR]

This systematic literature review was aimed to investigate the use of cell culture and animal models to evaluate the efficacy of alginate‐and chitosan‐based scaffolds on diabetic wound healing. We electronically searched the articles published until July 2019. The databases included five English databases such as PubMed, Web of Science, Embase, the Cochrane Library, CINAHL, and three Chinese databases like CNKI, WanFang Data, and VIP. The related articles were manually searched to identify studies that were not searched by electronic database searches. Twenty‐nine studies met the inclusion criteria. We divided the results into three groups: chitosan, alginate, and a combination of chitosan and alginate. Chitosan‐, alginate‐, and a combination of chitosan and alginate‐based scaffolds showed good intervention effects on wound healing. Chitosan‐based scaffolds were effective in diabetic skin wound healing. The effects of alginate and the combination of chitosan‐and alginate‐based scaffolds on diabetic skin wounds still need more research. However, due to the heterogeneity of animal and cell preclinical trials and the validity of the statistical analysis used in these studies, it is necessary to conduct a thorough study using well‐designed experiments to confirm these results. In addition, properly designed chitosan‐and/or alginate‐based scaffolds with thorough preclinical evaluations are required prior to clinical applications. [ABSTRACT FROM AUTHOR]

Translation of wound healing research is limited by the lack of an appropriate animal model, due to the anatomic and wound healing differences in animals and humans. Here, we characterize healing of grafted, full‐thickness human skin in an in vivo model of wound healing. Full‐thickness human skin, obtained from reconstructive operations, was grafted onto the dorsal flank of NOD.Cg‐KitW41JTyr + PrkdcscidIl2rgtm1Wjl/ThomJ mice. The xenografts were harvested 1 to 12 weeks after grafting, and histologic analyses were completed for viability, neovascularization, and hypoxia. Visual inspection of the xenograft shows drying and sloughing of the epidermis starting at week four. By week 12, the xenograft appears healed but has lost 63.05 ± 0.24% of the initial graft size. There is histologic evidence of epidermolysis as early as 2 weeks, which progresses until week 4, when new epidermis appears from the wound edges. Epidermal regeneration is complete by week 12, although the epidermis appears hypertrophied. An initial increase of infiltrating immune mouse cells into the xenograft normalizes to baseline 6 months after grafting. Neovascularization, as evidenced by positive staining for the proteins human CD31 and alpha smooth muscle actin, is present as early as 2 weeks after grafting at the interface between the xenograft and the mouse tissue. CD31 and alpha smooth muscle actin staining increased throughout the xenograft over the 12 weeks, leading to greater viability of the tissue. Likewise, there is increased Hypoxia Inducible Factor 1‐alpha expression at the interface of viable and nonviable tissue, which suggest a hypoxia‐driven process causing early graft loss. These findings illustrate human skin wound healing in an ischemic environment, providing a timeline for use of full thickness human skin after grafting in a murine model to study mechanisms underlying human skin wound healing. [ABSTRACT FROM AUTHOR]

Scar forming wounds are often characterized by higher levels of vascularity than non‐scarring wounds and normal skin, and inhibition of angiogenesis has been shown to inhibit scar formation in some model systems. The rabbit ear hypertrophic scar (HS) model has been widely used to study the mechanisms that underlie the development of HS as well as the effectiveness of various treatments. Although the rabbit ear HS model is well characterized in terms of scar formation, the rate and level of angiogenesis has not been investigated in this model, and the cause‐effect relationship between angiogenesis and rabbit HSs has not been examined. In the current study, full‐thickness excisional wounds were created on the ventral side of New Zealand White rabbit ears to induce HS formation, and the dynamic pattern of angiogenesis and the expression of angiogenic regulatory factors were examined over time. Blood vessel density was found to peak at 2.7% on day 14 post‐wounding, decreasing to 1.7% by day 28. mRNA levels of the proangiogenic factor VEGF‐A peaked at day 14, while the expression of the antiangiogenic factors pigment epithelium‐derived factor (PEDF) and thrombospondin 1 (TSP1) peaked at day 28 post‐wounding. To examine whether inhibition of angiogenesis influences HS formation in this model, wounds were treated with exogenous soluble antiangiogenic agents including recombinant PEDF (rPEDF) and a functional PEDF peptide (PEDF‐335). rPEDF and PEDF‐335 were administered intradermally from day 4 post‐wounding every 3 days until day 19. Intradermal injection of rPEDF or PEDF‐335 both led to decreased angiogenesis and decreased collagen deposition at the wound site. The results support the utility of antiangiogenic therapies, including rPEDF/PEDF‐335, as a potential new strategy for the prevention or treatment of HSs. [ABSTRACT FROM AUTHOR]

Ischemia‐reperfusion (I/R) is associated with various pathogenic conditions, and there has been increasing evidence that cutaneous I/R injury is associated with the pathogenesis of pressure ulcers (PUs), especially at the early stage presenting as non‐blanchable erythema. Several studies demonstrated that oxidative stress is a key player in I/R injury, and the inhibition of oxidative stress may be capable of protecting tissue damage after I/R injury in various organs including skin. Dimethyl fumarate (DMF) approved by the Food and Drug Administration is Nrf2 activator, and recent studies revealed the antioxidative and anti‐inflammatory effects of DMF on I/R injury in animal models. Our objective was to assess the effects of oral administration of DMF on the development of PUs after cutaneous I/R injury in mice. We found that DMF administration significantly decreased the size of PUs after cutaneous I/R. Cutaneous I/R‐induced oxidative stress was also significantly inhibited by DMF in OKD48 mice, in which oxidative stress can be visually assessed. In addition, DMF treatment decreased hypoxic area, the numbers of apoptotic cells, and vascular loss in I/R area. DMF treatment suppressed the infiltration of MPO+ neutrophils and the production of proinflammatory cytokines in I/R site after cutaneous I/R injury. in vitro experiments, DMF treatment suppressed the production of reactive oxygen species in pericyte‐like cells. These results suggest that DMF treatment might prevent the formation of PUs induced by cutaneous I/R injury via suppressing oxidative stress and subsequent inflammation. DMF treatment during the early phase of decubitus ulcers might protect against further progression. [ABSTRACT FROM AUTHOR]

Hypertrophic scar is an important clinical problem with limited therapeutic options. Aside from their roles as 3‐hydroxy‐3‐methylglutaryl‐coenzyme A reductase inhibitors, statins have also been demonstrated to decrease scarring by reducing connective tissue growth factor (CTGF) expression. However, poor penetrative ability limits their utility as topical treatments for hypertrophic scar. Here, we aim to develop novel statin formulations using liposomes to enhance dermal penetrative ability and to evaluate their efficacy against formation of hypertrophic scar utilizing our validated rabbit ear hypertrophic scar model. Liposomal simvastatin or pravastatin were compounded using a novel, flexible liposomal formulation and applied topically to rabbit ear hypertrophic scars daily from postoperation day (POD) 14 until POD 25. Scar color, including erythema and melanin, was measured using reflectance spectrophotometry on POD 28, and scar tissue was harvested for evaluation of scar elevation index as well as gene and protein expression. Human foreskin fibroblasts were also treated with statin formulations and CCN2 expression was determined by quantitative PCR. Both simvastatin and pravastatin were efficiently encapsulated in liposomes, forming nanometer‐scale particles possessing highly negative charges. Topical treatment with liposomal simvastatin and pravastatin at 6.5% concentration significantly reduced scar elevation index and decreased type I/III collagen content and myofibroblast persistence in the wound. The erythema/vascularity of scars was reduced by liposomal statin treatment, with concomitant decrease of CD31 expression as measured histologically. Expression levels of transcripts encoding CTGF, collagen I, and collagen III collagen in scar tissue were also decreased by liposomal pravastatin treatment, as were myofibroblast persistence and the type I/III collagen ratio as assessed by immunofluorescence and picrosirus red staining, respectively. Treatment of human foreskin fibroblasts with simvastatin or with liposome‐encapsulated pravastatin resulted in decreased expression of transcript encoding CTGF. Overall, our novel statin formulations encapsulated in liposomes were successfully delivered through topical application, significantly reducing hypertrophic scarring in a rabbit ear model. [ABSTRACT FROM AUTHOR]

Diabetic wound infection is a frequent complication that may result in limb amputation. To develop new treatment strategies in response to increasing bacterial resistance, animal models are needed. We created a diabetic mouse model with chronically infected wounds. Diabetes was induced using streptozotocin, and wounds were performed using a biopsy punch, and then infected with a clinical strain of Staphylococcus aureus. Chronification was reached by delaying healing thanks to chemical products (aminotriazole and mercaptosuccinic acid). Overall survival, as well as clinical, bacteriological and immunological data in skin, blood and spleens were collected at days 1, 7, and 14 after wounding. After a transient bacteremia proved by bacteria presence in spleen and kidneys in the first days after wounding, infected mice showed a chronic infection, with a bioburden impairing the healing process, and bacteria persistence compared to control mice. Infected mice showed gradual increasing skin levels of IL‐17A compared to control mice that resulted in an IL‐17/IFN‐γ inbalance, pointing out a localized Th17 polarization of the immune response. Whether infected or not, the skin level of IL‐10 decreased dramatically at days 1 and 7 after wounding, with an increase observed only in the control mice at day 14. After a decrease at day 1 in both groups, spleen IL‐10 showed a rather steady level at days 7 and 14 in the control group compared with the decrease observed in the infected group. The spleen IL‐10/IFN‐γ ratio showed a systemic inflammatory response with Th1 polarization. Therefore, this model provides useful data to study wound healing. It is easy to reproduce, affordable and offers clinical and biological tools to evaluate new therapeutics. [ABSTRACT FROM AUTHOR]

The field of regenerative medicine encounters different challenges. The success of tissue‐engineered implants is dependent on proper wound healing. Today, the process of normal urinary bladder wound healing is poorly characterized. We aspired to explore and elucidate the natural response to injury in an in vivo model in order to further optimize tissue regeneration in future studies. In this study, we aimed to characterize histological and molecular changes during normal healing in a rat model by performing a standardized incisional wound followed by surgical closure. We used a rodent model (n = 40) to follow the healing process in the urinary bladder for 28 days. Surgical exposure of the bladder without incision (n = 40) was performed in controls. Histological characterization and western blot analyses of proteins was carried out using specific staining and markers for inflammation, proliferation, angiogenesis, and tissue maturation. For the molecular characterization of gene expression total RNA was collected for RT2‐PCR in wound healing pathway arrays. Analysis of histology revealed distinct, but overlapping, phases of healing with a local inflammatory response (days 1‐8) simultaneous with a rapid formation of granulation tissue and proliferation (days 2‐8). We also identified significant changes in gene expression related to inflammation, proliferation, and extracellular matrix formation. Healing of an incisional wound in a rodent urinary bladder demonstrated that all the classical phases of wound healing: hemostasis, inflammation, proliferation followed by tissue maturation were present. Our data suggest that the bladder and the skin share similar molecular signaling during wound healing, although we noted differences in the duration of each phase compared to previous studies in rat skin. Further studies will address whether our findings can be extrapolated to the human bladder. [ABSTRACT FROM AUTHOR]

Human skin wound repair may result in various outcomes with most of them leading to scar formation. Commonly seen in many cutaneous wound healing cases, hypertrophic scars are considered as phenotypes of abnormal wound repair. To prevent the formation of hypertrophic scars, efforts have been made to understand the mechanism of scarring following wound closure. Numerous in vivo and in vitro models have been created to facilitate investigations into cutaneous scarring and the development of antiscarring treatments. To select the best model for a specific study, background knowledge of the current models of hypertrophic scars is necessary. In this review, we describe in vivo and in vitro models for studying hypertrophic scars, as well as the distinct characteristics of these models. The choice of models for a specific study should be based on the characteristics of the model and the goal of the study. In general, in vivo animal models are often used in phenotypical scar formation analysis, development of antiscarring treatment, and functional analyses of individual genes. In contrast, in vitro models are chosen to pathway identification during scar formation as well as in high‐throughput analysis in drug development. Besides helping investigators choose the best scarring model for their research, the goal of this review is to provide knowledge for improving the existing models and development of new models. These will contribute to the progress of scarring studies. [ABSTRACT FROM AUTHOR]

Ulcers including pressure ulcers and diabetic foot ulcers damage the skin and underlying tissue in people with compromised blood circulation. They are classified into four stages of severity and span from mild reddening of the skin to tissue damage and muscle/bone infections. Here, we used photoacoustic imaging as a noninvasive method for detecting early tissue damage that cannot be visually observed while also staging the disease using quantitative image analysis. We used a mouse model of pressure ulcers by implanting subdermal magnets in the dorsal flank and periodically applying an external magnet to the healed implant site. The magnet‐induced pressure was applied in cycles, and the extent of ulceration was dictated by the number of cycles. We used both laser‐ and light‐emitting diode (LED)‐based photoacoustic imaging tools with 690 nm excitation to evaluate the change in photoacoustic signal and depth of injury. Using laser‐based photoacoustic imaging system, we found a 4.4‐fold increase in the photoacoustic intensity in stage I vs. baseline (no pressure). We also evaluated the depth of injury using photoacoustics. We measured a photoacoustic ulcer depth of 0.38 ± 0.09 mm, 0.74 ± 0.11 mm, 1.63 ± 0.4 mm, and 2.7 ± 0.31 mm (n = 4) for stages I–IV, respectively. The photoacoustic depth differences between each stage were significant (p < 0.05). We also used an LED‐based photoacoustic imaging system to detect early stage (stage I) pressure ulcers and observed a 2.5‐fold increase in photoacoustic signal. Importantly, we confirmed the capacity of this technique to detect dysregulated skin even before stage I ulcers have erupted. We also observed significant changes in photoacoustic intensity during healing suggesting that this approach can monitor therapy. These findings were confirmed with histology. These results suggest that this photoacoustic‐based approach might have clinical value for monitoring skin diseases including pressure ulcers. [ABSTRACT FROM AUTHOR]

Proliferative and migratory abilities of fibroblasts are essential for wound healing at the skin surface. Cytoplasmic linker‐associated protein‐2 (CLASP2) was originally found to interact with cytoplasmic linker protein (CLIP)‐170. CLASP2 plays an important role in microtubule stabilization and the microtubule‐stabilizing activity of CLASP2 depends on its interactions with end binding (EB)‐1 and CLIP‐170. Although the microtubule‐stabilizing role of CLASP2 is well established, the effects of CLASP2 on the migration and proliferation of fibroblasts remain unclear in the context of wound healing. Therefore, we tested the utilization of CLASP2 as a directly applied protein drug to improve wound healing by promoting the migration of effector cells, including skin fibroblasts, to the site of repair or injury using an in vivo excisional wound mouse model and in vitro Hs27 skin fibroblast model. Epidermal growth factor, which is a recognized contributor to cell proliferation and migration, was used as positive control. In vitro and in vivo, CLASP2 treatment significantly enhanced cell migration and accelerated wound closure. Furthermore, in vivo, the CLASP2‐treated animal group displayed enhanced epidermal repair and collagen deposition. Next, we studied the mechanism of CLASP2 for wound healing. Increasing the abundance of intracellular free CLASP2 in skin fibroblasts by supplying exogenous CLASP2 seemed to stabilize microtubules through an interaction between CLASP2 and CLIP‐170, as well as EB1. Exogenous CLASP2 also showed direct binding with IQGAP1, increasing both cyclic adenosine monophosphate activity and phosphorylation of glycogen synthase kinase 3β, which in turn reinstated the binding between free CLASP2 and IQGAP1. In summary, exogenous CLASP2 increased Hs27 skin fibroblast migration by interacting with IQGAP1 and other cytoskeletal linker proteins, such as CLIP‐170 and EB1. Our results strongly suggest that CLASP2 can be developed in wound healing drugs for skin repair and/or regenerating cosmetic products. [ABSTRACT FROM AUTHOR]

The prevalence of infection in chronic wounds is well documented in the literature but not optimally studied due to the drawbacks of current methodologies. Here, we describe a tractable and simplified ex vivo human skin model of infection that addresses the critical drawbacks of high costs and limited translatability. Wounds were generated from excised abdominal skin from cosmetic procedures and cultured, inoculated with Staphylococcus aureus strain UAMS‐1, or under aseptic conditions. After three days, the infected wounds exhibited biofilm formation and significantly impaired reepithelialization compared to the control. Additionally, promigratory and proreparative genes were significantly downregulated, while proinflammatory genes were significantly upregulated, demonstrating molecular characterizations of impaired healing as in chronic wounds. This model allows for a simplified and versatile tool for the study of wound infection and subsequent development of novel therapies. [ABSTRACT FROM AUTHOR]

Keloid scars are described as benign fibro‐proliferative dermal outgrowths that commonly occur in pigmented skin post cutaneous injury, and continue to grow beyond the boundary of the original wound margin. There is a lack of thorough understanding of keloid pathogenesis and thus keloid therapeutic options remain ill‐defined. In view of the poor response to current therapy and high recurrence rates, there is an unmet need in improving our knowledge and therefore in identifying targeted and effective treatment strategies in management of keloids. Keloid research however, is hampered by a lack of relevant animal models as keloids do not spontaneously occur in animals and are unique to human skin. Therefore, developing novel animal models and nonanimal models for functional evaluation of keloid cells and tissue for better understanding their pathobiology and response to putative candidate therapies are essential. Here, we present the key concepts and relevant emerging research on two‐dimensional and three‐dimensional cell and tissue models for functional testing of keloid scars. We will describe in detail current models including in vitro mono‐ and co‐cultures, multi‐cellular spheroids (organoids) and organotyopic cultures, ex vivo whole skin keloid tissue organ culture models as well as in vivo human patient models. Finally, we discuss the role played by time as the fourth dimension in a novel model that involves sequential temporal biopsies of human patients with keloids (a so called 4D in vivo human model). The use of these unique models will no doubt prove pivotal in identification of new drug targets as well as biomarkers, in functional testing of emerging novel therapeutics, and in enhancing our understanding of keloid disease biology. [ABSTRACT FROM AUTHOR]

Current therapies available to treat and heal ocular surface injuries and periocular burns are frequently inadequate, costly, and labor intensive. To address these limitations, we have employed a flexible, semitransparent ocular wound chamber (OWC) to provide protection as well as a watertight seal to allow for the constant delivery of therapeutics to the ocular surface and surrounding periocular tissue. This study demonstrates the safety and utilization of the OWC on uninjured eyes and in our exposure keratopathy model. For initial safety studies (N = 3 per group), the eyelids remained intact and the eye uninjured. A blepharotomy (N = 6 per group) was performed to remove the upper and lower eyelids surrounding the left (OS) eye to create our exposure keratopathy model. Right (OD) eyes served as uninjured controls in all studies. Following OWC placement, 0.5 mL HPMC gel or balanced saline solution (BSS) was injected into the chamber. Animals were monitored daily and fully assessed via white light, fluorescein, and OCT imaging at least through 72 hours post OWC placement. In studies that included a blepharotomy, skin samples were analyzed by multiplex cytokine analysis. Results of safety experiments revealed no significant differences between treatment groups in corneal thickness, fluorescein staining, OCT imaging, or histological eye or skin sections when compared to control eyes. In our exposure keratopathy model, OWC treated eyes showed significantly less fluorescein uptake and also were found to have significantly lower levels of cytokines IL‐13 and IL‐5 in skin samples. These results demonstrate for the first time that treatment using the OWC device is not only safe, but significantly protects against blepharotomy‐induced exposure keratopathy. As a whole, this study advances our overall efforts to develop a feasible solution to treat ocular surface injuries, infections, and periocular burns. [ABSTRACT FROM AUTHOR]

The zebrafish is a popular animal model with well‐known regenerative capabilities. To study regeneration in this fish, the nitroreductase/metronidazole‐mediated system is widely used for targeted ablation of various cell types. Nevertheless, we highlight here some variability in ablation efficiencies with the metronidazole prodrug that led us to search for a more efficient and reliable compound. Herein, we present nifurpirinol, another nitroaromatic antibiotic, as a more potent prodrug compared to metronidazole to trigger cell‐ablation in nitroreductase expressing transgenic models. We show that nifurpirinol induces robust and reliable ablations at concentrations 2,000 fold lower than metronidazole and three times below its own toxic concentration. We confirmed the efficiency of nifurpirinol in triggering massive ablation of three different cell types: the pancreatic beta cells, osteoblasts, and dopaminergic neurons. Our results identify nifurpirinol as a very potent prodrug for the nitroreductase‐mediated ablation system and suggest that its use could be extended to many other cell types, especially if difficult to ablate, or when combined pharmacological treatments are desired. [ABSTRACT FROM AUTHOR]