Trehalose lyophilized platelets for wound healing

MMP-13 Plays a Role in Keratinocyte Migration, Angiogenesis, and Contraction in Mouse Skin Wound Healing

Topical treatment for wound management in fish is challenging due to their aquatic life. The objective of the present study was to assess the effect of a topical gel containing misoprostol and phenytoin on the healing of skin wounds in Brook Trout Salvelinus fontinalis. Four 16-×8-mm, full-thickness wounds were created in each trout. Fish were randomly assigned either to the misoprostol/phenytoin group (14 fish) or to the untreated control group (5 fish). In fish from the misoprostol/phenytoin group, two randomly selected wounds were topically treated with a misoprostol/phenytoin gel, while the other two wounds were left without topical treatment. Follow-up and treatment were performed every 5d for 120d. Different macroscopic healing indexes were recorded over time, and histological characteristics of each wound were scored at the end of the study. Treatment with misoprostol/phenytoin was associated with delayed epithelialization and wound maturation in comparison with contralateral untreated wounds. The treatment was also associated with lower histological inflammation scores and increased dermal neovascularization. The untreated wounds of fish in the misoprostol/phenytoin group showed delayed maturation and decreased healing speed and had lower inflammation scores and increased neovascularization compared to untreated control wounds. Topical treatment of wounds in Brook Trout with misoprostol/phenytoin gel at the concentration used had a local and systemic deleterious effect on wound healing. The results of the present study do not support the use of this gel at this dosage for the treatment of wounds in Brook Trout.

Over 1 million burn injuries are treated annually in the United States, and current tissue engineered skin fails to meet the need for full-thickness replacement. Bioprinting technology has allowed fabrication of full-thickness skin and has demonstrated the ability to close full-thickness wounds. However, analysis of collagen remodeling in wounds treated with bioprinted skin has not been reported. The purpose of this study is to demonstrate the utility of bioprinted skin for epidermal barrier formation and normal collagen remodeling in full-thickness wounds. Human keratinocytes, melanocytes, fibroblasts, dermal microvascular endothelial cells, follicle dermal papilla cells, and adipocytes were suspended in fibrinogen bioink and bioprinted to form a tri-layer skin structure. Bioprinted skin was implanted onto 2.5 × 2.5 cm full-thickness excisional wounds on athymic mice, compared with wounds treated with hydrogel only or untreated wounds. Total wound closure, epithelialization, and contraction were quantified, and skin samples were harvested at 21 days for histology. Picrosirius red staining was used to quantify collagen fiber orientation, length, and width. Immunohistochemical (IHC) staining was performed to confirm epidermal barrier formation, dermal maturation, vascularity, and human cell integration. All bioprinted skin treated wounds closed by day 21, compared with open control wounds. Wound closure in bioprinted skin treated wounds was primarily due to epithelialization. In contrast, control hydrogel and untreated groups had sparse wound coverage and incomplete closure driven primarily by contraction. Picrosirius red staining confirmed a normal basket weave collagen organization in bioprinted skin-treated wounds compared with parallel collagen fibers in hydrogel only and untreated wounds. IHC staining at day 21 demonstrated the presence of human cells in the regenerated dermis, the formation of a stratified epidermis, dermal maturation, and blood vessel formation in bioprinted skin, none of which was present in control hydrogel treated wounds. Bioprinted skin accelerated full-thickness wound closure by promoting epidermal barrier formation, without increasing contraction. This healing process is associated with human cells from the bioprinted skin laying down a healthy, basket-weave collagen network. The remodeled skin is phenotypically similar to human skin and composed of a composite of graft and infiltrating host cells. Impact statement We have demonstrated the ability of bioprinted skin to enhance closure of full-thickness wounds through epithelialization and normal collagen remodeling. To our knowledge, this article is the first to quantify collagen remodeling by bioprinted skin in full-thickness wounds. Our methods and results can be used to guide further investigation of collagen remodeling by tissue engineered skin products to improve ongoing and future bioprinting skin studies. Ultimately, our skin bioprinting technology could translate into a new treatment for full-thickness wounds in human patients with the ability to recapitulate normal collagen remodeling in full-thickness wounds.

Objective: To explore the effect of P311 microspheres-loaded thermosensitive chitosan hydrogel on the wound healing of full-thickness skin defects in rats. Methods: The method of experimental study was adopted. The polyvinyl alcohol/sodium alginate microspheres (simple microspheres), P311 microspheres, and bovine serum albumin labeled with fluorescein isothiocyanate (FITC-BSA) microspheres were prepared by water-in-oil emulsification, and then their morphology was observed under a light microscope/inverted fluorescence microscope. Chitosan solution was prepared, chitosan solution and β-glycerol phosphate disodium hydrate were mixed to prepare simple thermosensitive hydrogels, and thermosensitive hydrogels loaded with simple microspheres or P311 microspheres were prepared by adding corresponding substances in simple thermosensitive hydrogels. The morphological changes of the prepared four liquids in the state of tilt was observed at 37 ℃. After being freeze-dried, the micromorphology of the prepared four liquids was observed under a scanning electron microscope. Eighteen 3-4-week-old male Sprague-Dawley rats were divided into normal group without any treatment, dressing group, chitosan group, hydrogel alone group, simple microspheres-loaded hydrogel group, and P311 microspheres-loaded hydrogel group, which were inflicted with one full-thickness skin defect wound on both sides of the back spine and were dealt correspondingly, with 3 rats in each group. Rats with full-thickness skin defects in the five groups were collected, the wound healing was observed on post injury day (PID) 0 (immediately), 5, 10, and 15, and the wound healing rates on PID 5, 10, and 15 were calculated. The wound and wound margin tissue of rats with full-thickness skin defects in the five groups on PID 15 and normal skin tissue in the same site of rats in normal group were collected, hematoxylin and eosin staining was conducted to observe the histological changes, immunohistochemical staining was performed to observe the expressions of CD31 and vascular endothelial growth factor (VEGF), and Western blotting was conducted to detect the protein expressions of CD31 and VEGF. The number of samples was all three. Data were statistically analyzed with one-way analysis of variance, analysis of variance for repeated measurement, and Bonferroni correction. Results: Simple microspheres were spherical, with loose and porous surface. The surfaces of P311 microspheres and FITC-BSA microspheres were smooth without pores, and the FITC-BSA microspheres emitted uniform green fluorescence. The diameters of the three microspheres were basically consistent, being 33.1 to 37.7 μm. Compared with chitosan solution and simple thermosensitive hydrogel, the structures of the two microspheres-loaded hydrogels were more stable in the state of tilt at 37 ℃. The two microspheres-loaded hydrogels had denser network structures than those of chitosan solution and simple thermosensitive hydrogel, and in the cross section of which microspheres with a diameter of about 30 μm could be seen. Within PID 15, the wounds of rats in the five groups were healed to different degrees, and the wound healing of rats in P311 microspheres-loaded hydrogel group was the best. On PID 5, 10, and 15, the wound healing rates of rats in dressing group and chitosan group were (26.6±2.4)%, (38.5±3.1)%, (50.9±1.5)%, (47.6±2.0)%, (58.5±3.6)%, and (66.7±4.1)%, respectively, which were significantly lower than (59.3±4.8)%, (87.6±3.2)%, (97.2±1.0)% in P311 microspheres-loaded hydrogel group (P<0.05 or P<0.01). The wound healing rates of rats in hydrogel alone group on PID 10 and 15, and in simple microspheres-loaded hydrogel group on PID 15 were (76.0±3.3)%, (84.5±3.6)%, and (88.0±2.6)%, respectively, which were significantly lower than those in P311 microspheres-loaded hydrogel group (P<0.05). The epidermis, hair follicles, and sebaceous glands could be seen in the normal skin of rats in normal group, without positive expressions of CD31 or VEGF. The wounds of rats in P311 microspheres-loaded hydrogel group on PID 15 were almost completely epithelialized, with more blood vessels, hair follicles, sebaceous glands, and positive expressions of CD31 and VEGF in the wounds than those of rats with full-thickness skin defects in the other four groups, and more protein expressions of CD31 and VEGF than those of rats in the other five groups. Conclusions: The P311 microspheres-loaded thermosensitive chitosan hydrogel can release the encapsulated drug slowly, prolong the drug action time, and promote wound healing in rats with full-thickness skin defects by promoting wound angiogenesis and re-epithelialization.

Botulinum toxin is a neurotoxin that has been utilized to induce chemo-denervation of muscles. Cutaneous wounds represent a special situation in which the tensile forces applied by these muscles on wound edges might have deleterious effects on the healing process. The aim of this review was to investigate such an effect and to review other mechanisms this toxin might have on the healing process. We also reviewed the role of botulinum toxin in the management of hypertrophic scars and cleft lip repair.

Background:The developmental abnormality spina bifida is hallmarked by missing tissues (e.g. skin) and exposure of the spinal cord to the amniotic fluid, which can negatively impact neurological development. Surgical closure of the skin in utero limits neurological damage, but in large defects this results in scarring and contractures. Stimulating skin regeneration in utero would greatly benefit treatment outcome. Previously, we demonstrated that a porous type I collagen (COL) scaffold, functionalized with heparin (HEP), fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor (VEGF) (COL-HEP/GF) improved pre- and postnatal skin regeneration in a fetal sheep full thickness wound model. In this study we uncover the early events associated with enhanced skin regeneration.Methods:We investigated the gene expression profiles of healing fetal skin wounds two weeks after implantation of the COL(-HEP/GF) scaffolds. Using laser dissection and microarrays, differentially expressed genes (DEG) were identified in the epidermis and dermis between untreated wounds, COL-treated wounds and wounds treated with COL-HEP/GF. Biological processes were identified using gene enrichment analysis and DEG were clustered using protein–protein-interaction networks.Results:COL-HEP/GF influences various interesting biological processes involved in wound healing. Although the changes were modest, using protein–protein-interaction networks we identified a variety of clustered genes that indicate COL-HEP/GF induces a tight but subtle control over cell signaling and extracellular matrix organization.Conclusion:These data offer a novel perspective on the key processes involved in (fetal) wound healing, where a targeted and early interference during wound healing can result in long-term enhanced effects on skin regeneration.

In facial plastic surgery, the potential for direct delivery of growth factors from platelet preparations has been of particular interest for use in facial rejuvenation, recovery after facial surgery, and wound healing. A literature search was conducted through PubMed for the terms PRP, PRFM, platelet-rich plasma, platelet-rich fibrin matrix, platelet preparations, platelet therapy, growth factors, platelet facial, platelet facial rejuvenation, platelet wound healing, platelet plastic surgery. Articles pertaining to the use of platelet preparations in facial surgery and wound healing in plastic surgery after 2001 were included. Thirteen in vitro studies showed use of platelet-rich plasma (PRP) and platelet-rich fibrin matrix (PRFM) had a significant effect on cellular activity. Twenty-four out of 28 animal studies exhibited favorable results with use of a platelet preparation, including five of six studies that showed enhanced fat graft survival with addition of a platelet preparation. Twenty-three case series and clinical trials were identified, only two of which showed no differences. Twenty-one reported favorable results with use of various platelet preparations. A total of 47 studies used PRP, four studies evaluated Leukocyte-rich PRP, and fourteen studies used PRFM. The vast majority of studies examined show a significant and measurable effect on cellular changes, wound healing, and facial esthetic outcomes with use of platelet preparations, both topical and injectable. One must also consider possible publication bias against null results that may have had an influence on the data that were available for review. However, the preponderance of studies suggests that platelet preparations might represent an as-of-yet untapped adjunct in facial plastic surgery.

Vascular endothelial growth factor (VEGF) is important in improving wound healing. We evaluated the effect of sensory (direct current, 600 microamperes) and motor (monophasic current, pulse duration 300 microseconds, 100 Hz, 2.5-3.0 mA) intensities of cathodal electrical stimulation (ES) current to release VEGF in muscle and skin in the wound site. We randomly assigned 48 male Sprague-Dawley rats into one control and two experimental groups (sensory and motor ESs). A full-thickness skin incision was made on each animal's dorsal region. The experimental groups received ES for 1 h/day every other day. In the control group, no current was applied. VEGF expression was measured in muscle and skin on the third and seventh days after surgical incision. Our outcomes demonstrated that no difference was found in the VEGF levels among groups on the third day. However, on the seventh day, the skin VEGF levels in the sensory group were significantly higher than those levels of the other groups (p < 0.05). No difference was found in the muscle VEGF levels on the third and seventh days. The results showed that sensory ES increases the release of more VEGF in skin. This mechanism may be one through which a sensory type of current is more effective in promoting wound healing.

Vascular Endothelial Growth Factor-C Accelerates Diabetic Wound Healing

The objective was to investigate the specific role and the regulatory mechanism of vascular endothelial growth factor (VEGF) during wound healing in diabetic foot ulcer (DFU). Streptozotocin-induced diabetic rats were used to establish a DFU animal model. VEGF and Axitinib (a specific inhibitor of VEGFR) were used for treatment invivo. The wounds at different time points were imaged and histological analysis of the wounds were performed by haematoxylin and eosin (H&E) staining and Masson's trichrome staining. Immunohistochemical staining was conducted to examine CD31 and eNOS expression in the wounds. Immunofluorescence assay and quantitative real-time PCR were performed to examine macrophage markers. In addition, THP-1 was differentiated to macrophages, and then treated with interleukin (IL)-4 to induce M2 macrophages, followed by VEGF treatment. The conditional medium (CM) from VEGF-mediated macrophages were collected to culture human dermal fibroblasts (HDFs). Cell viability and migration were measured by Cell Counting Kit (CCK)-8, wound-healing and Transwell assays, respectively. VEGF treatment remarkably accelerated wound healing of DFU rats. VEGF promoted collagen deposition and elevated CD31 and eNOS expression, confirming the pro-angiogenesis of VEGF around diabetic wound in rats. Meanwhile, VEGF restricted pro-inflammatory cytokines and increased F4/80 and CD206 expression, highlighting the activated macrophages and enhanced M2 macrophages following VEGF treatment in diabetic wounds of DFU rats. However, Axitinib exerted an opposite function to VEGF in DFU rats. Moreover, VEGF directly promoted macrophage polarization toward M2 phenotype invitro, and the CM from VEGF-mediated M2 macrophages markedly promoted HDFs proliferation, migration and collagen deposition. VEGF might accelerate the wound healing of DFU through promoting M2 macrophage polarization and fibroblast migration.

Regenerative medicine for the treatment of skin lesions is an innovative and rapidly developing field that aims to promote wound healing and restore the skin to its original condition before injury. Over the years, different topical treatments have been evaluated to improve skin wound healing and, among them, mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) have shown promising results for this purpose. This study sought to evaluate the quality of the healing process in experimentally induced full-thickness skin lesions treated with PRP associated or unassociated with MSCs in a sheep second intention wound healing model. After having surgically created full-thickness wounds on the back of three sheep, the wound healing process was assessed by performing clinical evaluations, histopathological examinations, and molecular analysis. Treated wounds showed a reduction of inflammation and contraction along with an increased re-epithelialization rate and better maturation of the granulation tissue compared to untreated lesions. In particular, the combined treatment regulated the expression of collagen types I and III resulting in a proper resolution of the granulation tissue contrary to what was observed in untreated wounds; moreover, it led to a better maturation and organization of skin adnexa and collagen fibers in the repaired skin compared to untreated and PRP-treated wounds. Overall, both treatments improved the wound healing process compared to untreated wounds. Wounds treated with PRP and MSCs showed a healing progression that qualitatively resembles a restitutio ad integrum of the repaired skin, showing features typical of a mature healthy dermis.

NF-κB signaling is key in the wound healing processes of silk fibroin.

Clinic-oriented injectable smart material for the treatment of diabetic wounds: Coordinating the release of GM-CSF and VEGF

Madhu ghrita is a herbal formulation containing honey and ghee (clarified butterfat) as its constituents. The aim of present study is to verify the wound healing and antiinflammatory claims of Madhu ghrita. Incision and excision wound models were used for evaluation of wound healing activity followed by histopathological study in which healing markers like keratinization, epithelization, fibrosis, neovascularisation and collagenation were evaluated in male Wistar rats. The results of Madhu ghrita were compared with the results of untreated control group and results of framycetine sulphate cream, considered as a positive control. The formulation was also tested for antiinflammatory activity, using carrageenan-induced paw oedema in male Wistar rats. The test formulation Madhu ghrita enhanced the tensile strength, which significantly improved over untreated wounds. The tensile strength of untreated control wound was 281±5.82, while with the Madhu ghrita and framycetine sulphate cream 1% w/w, it was 328±8.9 and 398±6.32, respectively. Treatment with Madhu ghrita alone promoted wound contraction and reduced the wound closure time, so increase in tensile strength and wound contraction shows the wound healing potential of Madhu ghrita . Histopathological study shows that proliferation of epithelial tissue promotes angiogenesis, multiplication of fibrous connective tissue due to treatment with Madhu ghrita . The test formulation Madhu ghrita also shows significant antiinflammatory activity when the results are compared with the activity of ibuprofen gel as reference standard. The present study demonstrates the wound healing and antiinflammatory potential of Madhu ghrita .

Vascular endothelial growth factor (VEGF) plays key roles in angiogenesis, vasculogenesis, and wound healing. In cancers, including triple negative breast cancer (TNBC), VEGF has been associated with increased invasion and metastasis, processes that require cancer cells to traverse through the extracellular matrix (ECM) and establish angiogenesis at distant sites. To further understand the role of VEGF in modifying the ECM, we characterized VEGF-mediated changes in the ECM of tumors derived from TNBC MDA-MB-231 cells engineered to overexpress VEGF. We established that increased VEGF expression by these cells resulted in tumors with reduced collagen 1 (Col1) fibers, fibronectin, and hyaluronan. Molecular characterization of tumors identified an increase of MMP1, uPAR, and LOX, and a decrease of MMP2, and ADAMTS1. α-SMA, a marker of cancer associated fibroblasts (CAFs), increased, and FAP-α, a marker of a subset of CAFs associated with immune suppression, decreased with VEGF overexpression. Analysis of human data from The Cancer Genome Atlas Program confirmed mRNA differences for several molecules when comparing TNBC with high and low VEGF expression. We additionally characterized enzymatic changes induced by VEGF overexpression in three different cancer cell lines that clearly identified autocrine-mediated changes, specifically uPAR, in these enzymes. Unlike the increase of Col1 fibers and fibronectin mediated by VEGF during wound healing, in the TNBC model, VEGF significantly reduced key protein components of the ECM. These results further expand our understanding of the role of VEGF in cancer progression and identify potential ECM-related targets to disrupt this progression.