Exogenous PRAS40 reduces KLF4 expression and alleviates hypertrophic scar fibrosis and collagen deposition through inhibiting mTORC1.

BACKGROUND: To identify the anti-fibrosis effect of PRAS40 in scar, and its potential mechanism. METHODS: We constructed a rat model of hypertrophic scarthat was locally injected the PRAS40 overexpression adenoviruses, mTORC1 inhibitor MHY1485 and activator rapamycin, and further observed the pathological changes of skin tissue and the severity of fibrosis by HE, Masson and sirius red staining, and analyzed the deposition of a-SMA and collagen I by western blot and immunofluorescence test. Meanwhile, the co-localization of KLF4 with a-SMA and type I collagen was analyzed, as well as the regulatory effect of PRAS40 on KLF4. In addition, we also verified whether the inhibition of scar fibrosis by PRAS40 is related to mTORC1, and whether the upregulation of KLF4 is related to mTORC1. RESULTS: The results showed that the expression of PRAS40 was low and p-PRAS40 was high in scar skin tissue. After local injection of PRAS40 overexpression adenovirus, the expression of PRAS40 in skin tissue was increased. The overexpression of PRAS40 can inhibit scar skin fibrosis and reduce the content of a-SMA and collagen I. Further mechanism analysis confirms that the inhibitory effect of PRAS40 on skin fibrosis is related to mTORC1, and PRAS40 inhibits the activation of mTORC1. The expression of KLF4 is relatively low in scar tissue. PRAS40 administration upregulated the expression of KLF4, which is related to mTORC1 CONCLUSIONS: PRAS40 significantly improves fibrosis of scar skin tissue and increases the expression of KLF4 in scars. The anti-fibrotic effect of PRAS40 depends on mTORC1.

Research advances in prevention and treatment of burn wound deepening in early stage.

The burn wound is a dynamic living environment that is affected by many factors. It may present a progressive expansion of necrosis into the initially viable zone of stasis within a short time postburn. Therefore, how to salvage of the zone of stasis is of crucial importance in prevention and treatment strategies of burn wound progressive deepening. This review focuses on the cellular basis of tissue injury and the current progress of prevention and treatment strategies of burn wound progressive deepening, in order to provide references for the treatment of burn wounds in the early phase.

Protective Effect of Conditioned Media of Human Fetal Dermal Mesenchymal Stem Cells Can Inhibit Burn-induced Microvascular Hyperpermeability.

Burns often cause loss of skin barrier protection, fluid exudation, and local tissue edema, which hinder functional recovery. Effectively improving the quality of deep burn wound healing, shortening the wound healing time, and reducing tissue fluid leakage are urgent problems in the medical field. Human mesenchymal stem cells (MSCs) can effectively stabilize vascular endothelial injury. Fetal dermal MSCs (FDMSCs) are a newly discovered source of MSCs derived from the skin of accidentally aborted fetuses. However, the effect of FDMSCs on vascular permeability remains poorly understood. In this study, conditioned media from FDMSCs (F-CM) extracted from fetal skin tissue was prepared. The effect of F-CM on vascular permeability was evaluated using the internal circulation method FITC-dextran in vivo, and several in vitro assays, including cell viability assay, transwell permeability test, immunofluorescence, and western blotting. Altogether, our results demonstrate that F-CM could inhibit burn-induced microvascular hyperpermeability by increasing the protein expression levels of occludin and VE-cadherin, while restoring the expression of endothelial F-actin, and providing the foundation of a novel therapy for the treatment of burns with F-CM.

Bacterial cellulose membrane combined with BMSCs promotes wound healing by activating the notch signaling pathway.

Objective: The bacterial cellulose membrane (BCM) has been widely studied and applied as a new biomaterial for wound healing, but causes pain with frequent dressing changes. Local application of bone marrow mesenchymal stem cells (BMSCs) requires a niche. Furthermore, the effect and mechanism of the BCM combined with BMSCs have not been reported. Methods: Morphological and chemical identifications of BCMs were investigated by porosity analyses, scanning electron microscopy, and Fourier-transform infrared spectroscopy. Biological wound dressings (BWDs) were prepared by the BCM in combination with BMSCs. The biological effects of BWDs on human dermal fibroblast (HDF) and VEGF-A in human vascular endothelial cells (HuVECs) were detected in vitro, and the effect of BWDs on acute wounds in mice was detected in vivo. Collagen and angiogenesis were evaluated through hematoxylin-eosin staining and Masson staining. The expressions of COL-1 and VEGF-A and the activation of the Notch signaling pathway in vivo and in vitro were detected by quantitative reverse-transcriptase polymerase chain reaction. Results: The BCM had a nanoscale structure and provided a partial niche for the survival and proliferation of BMSCs. BWDs were successfully prepared and regulated the biological behaviors of wound healing-related cells in vitro and upregulated the expressions of COL-1 in HDF and VEGF-A in HuVECs. BWDs promoted wound healing by increasing collagen type I synthesis and angiogenesis in acute wounds in mice. Conclusions: BWDs prepared by the combination of nanomaterial BCMs and BMSCs facilitated acute wound healing, which may be regulated by activating the Notch signaling pathway.

Analysis of the Curative Effect and Influencing Factors of Collagen Sponge Combined with Autologous Skin Graft in the Treatment of Deep Burn Patients.

Burn is one of the common traumatic diseases in clinics. After deep burn, the complicated changes of the condition are caused by the burn wound, which ends with the repair of the wound. For patients with deep burns, whether the wound can be repaired as soon as possible is the key to the success of clinical treatment. For patients with deep burns, due to the lack of an autologous skin source, scar hyperplasia at donor site, skin graft repair at donor site, postoperative flap necrosis, and other problems in traditional surgical procedures, the method of improving function only by an autologous skin source has been unable to perform the later function reconstruction in patients with deep burns. In this study, collagen sponge combined with autologous skin graft was used to treat patients with deep burn, and the clinical efficacy of the patients was observed, and the related factors affecting the efficacy of the patients were analyzed. The results showed that collagen sponge combined with autologous skin graft was effective in the treatment of deep burn patients, and it was worth popularizing. Deep III-IV degree burns, wound infection, and hospital stay >3 months are all risk factors affecting the postoperative curative effect of patients. Therefore, in the clinical work, we should focus on patients with deep III-IV degree burns, perform surgery as soon as possible, and actively deal with wounds to prevent infection, which is beneficial to improve the curative effect.

Anti-aging effects of fetal dermal mesenchymal stem cells in a D-galactose-induced aging model of adult dermal fibroblasts.

The main characteristic of skin aging is the change in the composition of the dermis, mainly resulting from fibroblast senescence. Mesenchymal stem cells derived from fetal dermis are defined as fetal dermal mesenchymal stem cells; they reportedly exert wound healing effects on the skin and regulate keloid fibroblast proliferation. D-Galactose is widely used in animal aging models. In this study, we confirmed that D-galactose inhibits adult dermal fibroblast proliferation, and the inhibitory effect gradually increased with increasing concentration. Finally, we chose a concentration of 40 g/L D-galactose to induce adult dermal fibroblast senescence. D-Galactose increased the intensity of senescence-associated ß-galactosidase staining and the levels of reactive oxygen species in adult dermal fibroblasts. Furthermore, D-galactose increased the mRNA expression of p16, p21, and p53. The fetal dermal mesenchymal stem cell-conditioned medium improved the above-mentioned effects. Overall, fetal dermal mesenchymal stem cells exerted anti-aging effects against adult dermal fibroblasts induced by D-galactose via paracrine functions.

Antitumor effects of conditioned media of human fetal dermal mesenchymal stem cells on melanoma cells.

Background: Malignant melanoma is the most lethal form of cutaneous tumor and has a high metastatic rate and motility capacity. Owing to the poor prognosis, it is urgent to seek an effective therapeutic regimen. Human mesenchymal stem cells (MSCs) can home to tumor cells and have been shown to play important roles in both promoting and inhibiting tumor development. Fetal dermal MSCs (FDMSCs), derived from fetal skin are a novel source of MSCs. Nevertheless, the antitumor capacity of FDMSCs on malignant melanoma is not clearly understood. Materials and methods: FDMSCs were extracted from the dorsal skin of fetal tissues. A375 melanoma cells lines were obtained from American Type Culture Collection. The effects of conditioned media from FDMSCs (CM-FDMSC) on A375 melanoma cells were tested in vivo using tumor formation assay and in vitro using cell viability, 5-ethynyl-2'-deoxyuridine incorporation, flow cytometry, TdT-mediated dUTP Nick-End Labeling (TUNEL), wound healing, transwell invasion, and Western blotting. Results: CM-FDMSC inhibited A375 tumor formation in vivo. In vitro, CM-FDMSC inhibited the tumor-related activities of A375 melanoma cells, as evidenced reductions in viability, migration, and invasion. CM-FDMSC-treated A375 cells showed decreased phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and extracellular signal-regulated kinase (ERK) phosphorylation, and up-regulation of Bcl-2-Associated X (BAX) and down-regulation of B-cell lymphoma-2 (BCL-2) expression. Conclusion: CM-FDMSC can inhibit the tumor-forming behaviors of A375 melanoma cells and inhibit PI3K/AKT and mitogen-activated protein kinase signaling to shift their BCL-2/BAX ratio toward a proapoptotic state. Identification of the bioactive components in CM-FDMSC will be important for translating these findings into novel therapies for malignant melanoma.

Denatured acellular dermal matrix seeded with bone marrow mesenchymal stem cells for wound healing in mice.

It is the basic task of burn therapy to cover the wound with self-healthy skin timely and effectively. However, for patients with extensive burns, autologous skin is usually insufficient, and allogenic or heterogeneous skin leads to strong immune response. It is vital to choose an appropriate treatment for deep extensive burns. Nowadays, the dermal substitute combined with bone marrow mesenchymal stem cells (BM-MSCs) is a prospective strategy for burn wound healing. Denatured acellular dermal matrix (DADM), as one of dermal substitutes, which prepared by burn skin discarded in escharotomy, not only maintains a certain degree of 3D structure of collagen, but also has good biocompatibility. In this study, the preparation method of DADM was improved and DADM was seeded with BM-MSCs. Then BM-MSCs-seeded DADM (DADM/MSCs) was implanted into mice cutaneous wound, and the effect of DADM/MSCs dermal substitute was assessed on skin regeneration. As a result, BM-MSCs survived well and DADM/MSCs scaffolds significantly promoted wound healing in terms of angiogenesis, re-epithelialization and skin appendage regeneration. DADM/MSCs scaffold may represent an alternative promising therapy for wound healing in deep extensive burns.

Risk factors for necrosis of skin flap-like wounds after ED debridement and suture.

BACKGROUND: Skin flap-like wounds are common. These wound flaps are prone to avascular necrosis with simple debrided and sutured, and postoperative hyperplastic scarring and contracture of wound surfaces can adversely affect the patient's appearance. Here, we evaluate the data of cases with flap-like wounds to identify the causes of flap necrosis. METHODS: Six hundred patients with skin flap-like wounds between January 1, 2013 and December 31, 2016 were retrospectively reviewed. Their age, sex, injury reason, size of flap, length-width ratio of wound, thickness of pedicle, operation time, injury site, direction of blood perfusion in the flap and operating methods were recorded. The risks for flap necrosis were analyzed with one-factor analysis. RESULTS: A total success rate of 92.5% (555/600) for flap-like wound reconstruction was obtained. Among 67 flaps with vascular crisis, 22 were salvaged by subcutaneous injection of anisodamine, selective suture removal, and pressure dressing with elastic bandages. For the 45 patients with flap necrosis, there was no significant difference from patients without necrosis in terms of sex, age, and size of flap (P > 0.05). The incidence of flap necrosis was significantly different in terms of injury reason, length-width ratio of wound, thickness of pedicle, operation time, injury site, direction of blood perfusion in the flap and operating methods (P < 0.05). CONCLUSION: Injury reason, length-width ratio of wound, thickness of pedicle, operation time, injury site, direction of blood perfusion in the flap and operating methods, rather than age, sex and size of flap, were significant risk factors for necrosis of flap-like wounds.

Interleukin 17 (IL-17)-Induced Mesenchymal Stem Cells Prolong the Survival of Allogeneic Skin Grafts.

BACKGROUND Mesenchymal stem cells (MSCs) have the potential of self-renewal and multi-differentiation and have a wide application prospect in organ transplantation for the effect of inducing immune tolerance. It has found that interleukin 17 (IL-17) could enhance the inhibition effect of MSCs on T cell proliferation and increase the immunosuppressive effect of MSCs. In this study, we aimed to investigate the effect of IL-17-induced MSCs on allograft survival time after transplantation. MATERIAL AND METHODS BMSCs were characterized by differential staining. The allogenic skin transplantations were performed and the BMSCs pre-treated by IL-17 were injected. To assess the immunosuppressive function of IL-17-induced BMSCs, the morphology of the grafts, the homing ability of the BMSCs, and the survival time of the grafts were analyzed. RESULTS BMSCs from BALB/c have multidirectional differentiation potential to differentiate into osteogenic, chondrogenic, and adipogenic lineage cells. IL-17-induced BMSCs prolonged the survival time of allogeneic skin grafts dramatically. We found that there were more labeled MSCs in the skin grafts, and the Treg subpopulations percentage, IL-10, and TGF-ß were significantly increased, while the IFN-γ level was decreased compared to the control group and MSCs group. In conclusion, IL-17 can enhance the homing ability of MSCs and regulate the immunosuppressive function of MSC. CONCLUSIONS Our data demonstrate that IL-17 plays the crucial role in MSC homing behaviors and promotes immunosuppression of MSCs during transplantation procedures, suggesting that IL-17-pre-treated MSCs have potential to prolong graft survival and reduce transplant rejection.

Anthropometric growth study of the ear in a Chinese population.

BACKGROUND: A large number of anthropometric studies of the auricle have been reported in different nations, but little data were available in the Chinese population. The aim of this study was to analyze growth changes in the ear by measuring the width and length of ears in a Chinese population. METHODS: A total of 480 participants were enrolled and classified into 1-, 3-, 5-, 7-, 9-, 12-, 14-, and 18-year groups (half were boys and half were girls in each group). Ear length, ear width, body weight, and body length were measured and recorded; ear index was calculated according to ear length and ear width. The growth of auricle and differences between genders were analyzed. Growth of ear in relation to body height and weight and the degree of emphasis on the length and width of the auricle were also analyzed. RESULTS: Ear length and width increased with age. Ear length achieved its mature size in both 14-year-old males and females. Ear width reached its mature size in males at 7 years and in females at 5 years. Different trends of ear index were shown between males and females. People in this population paid more attention to the length than the width of the auricle. CONCLUSIONS: The data indicated that ear development followed increase in age. There were gender and ethnic difference in the development of ear. These results may have potential implications for the diagnosis of congenital malformations, syndromes, and planning of ear reconstruction surgery.

Inhibiting function of human fetal dermal mesenchymal stem cells on bioactivities of keloid fibroblasts.

BACKGROUND: Keloid is one kind of benign skin disease caused by hyperplasia of fibroblasts and collagen fibrils. It is refractory due to the lack of an effective treatment at present, which puts pressure on seeking a new therapeutic regimen. Mesenchymal stem cells (MSCs) from fetal skin are considered to play a crucial role in scarless healing. Nevertheless, the efficacy of them in keloid disorders remains poorly understood. METHODS: Keloid fibroblasts (KFs), human adult dermal fibroblasts (ADFs), and human fetal dermal mesenchymal stem cells (FDMSCs) were isolated to single cells and cultured in Dulbecco's modified Eagle's medium (DMEM). ADFs and FDMSCs were used to generate ADF-conditioned medium (A-CM) and FDMSC-conditioned medium (F-CM). The effects of A-CM and F-CM on KFs were tested using MTT assay, BrdU assay, TUNEL assay, quantitative polymerase chain reaction, Western blot, and annexin V-FITC/PI binding assay,. RESULTS: FDMSCs inhibited the bioactivity of KFs, downregulated the expression of the antiapoptotic protein BCL-2, and upregulated the expression of the proapoptotic protein BAX of KFs by secreting some soluble substances, thus accelerating the apoptosis of KFs. CONCLUSION: F-CM induces apoptosis of KFs, providing a novel treatment strategy for keloid disorders.

Immune Tolerance of Mesenchymal Stem Cells and Induction of Skin Allograft Tolerance.

BACKGROUND: Mesenchymal stem cells not only possess reparative properties, but also have immunomodulatory effect. Owing to the properties, they have been proposed to be hopeful candidates for cell therapy in the process of organ transplantation. In the preclinical researches, it shows that MSCs is capable of prolonging graft survival and inducing tolerance in some cases. Various mechanisms of immune tolerance were reported before, such as tolerogenic dendritic cells, induction of apoptosis, regulatory T cells, mixed chimerism, soluble factors and anergy. Furthermore, the induction of immune tolerance may be influenced by the dose, route and optimal timing of MSCs administration. Allograft of skin can provisionally restore the function of skin barrier and offer a good environment to expand micro-skin auto-graft; however, at the same time, it can cause strong immune rejection, which leads to the failure of skin graft. OBJECTIVE: The review aims at analyzing the mechanisms of immunosuppression mediated by MSCs, and the induction of tolerance of skin graft by MSCs. CONCLUSION: Mesenchymal stem cells are hopeful candidates for cell therapy in the process of organ transplantation, and can induce immune tolerance of skin graft to some extent.

Rib Cartilage Assessment Relative to the Healthy Ear in Young Children with Microtia Guiding Operative Timing.

BACKGROUND: The optimal age at which to initiate for auricular reconstruction is controversial. Rib cartilage growth is closely related to age and determines the feasibility and outcomes of auricular reconstruction. We developed a method to guide the timing of auricular reconstruction in children with microtia ranging in age from 5 to 10 years. METHODS: Rib cartilage and the healthy ear were assessed using low-dose multi-slice computed tomography. The lengths of the eighth rib cartilage and the helix of the healthy ear (from the helical crus to the joint of the helix and the earlobe) were measured. Surgery was performed when the two lengths were approximately equal. RESULTS: The preoperative eighth rib measurements significantly correlated with the intraoperative measurements (P < 0.05). From 5 to 10 years of age, eighth rib growth was not linear. In 76 (62.8%) of 121 patients, the eighth rib length was approximately equal to the helix length in the healthy ear; satisfactory outcomes were achieved in these patients. In 18 (14.9%) patients, the eighth rib was slightly shorter than the helix, helix fabrication was accomplished by adjusting the length of the helical crus of stent, and satisfactory outcomes were also achieved. Acceptable outcomes were achieved in 17 (14.0%) patients in whom helix fabrication was accomplished by cartilage splicing. In 9 (7.4%) patients with insufficient rib cartilage length, the operation was delayed. In one (0.8%) patient with insufficient rib cartilage length, which left no cartilage for helix splicing, the result was unsatisfactory. CONCLUSIONS: Eighth rib cartilage growth is variable. Rib cartilage assessment relative to the healthy ear can guide auricular reconstruction and personalize treatment in young patients with microtia.

[Advances in the research of the biological activities of degradation products of extracellular matrix].

ECM is a supporting structure for stabilizing the location of cells and preserving the structure of tissues. Recently, it has been discovered that ECM and its degradation products may exert profound influences on tissues and cells, such as activities of inflammatory cells and immune cells. Angiogenesis may be stimulated or inhibited by degradation products of ECM. Matrikines, liberated by partial proteolysis of ECM macromolecules, are found to regulate cell functional activities and play a significant role in wound healing or tumor invasion. Post-burn denatured dermal matrix is being studied in burn healing now. The study of post-burn denatured or necrotic dermal matrix should be emphasized in future.

[Construction of keratinocyte growth factor phage active peptides for the promotion of epidermal cell proliferation].

OBJECTIVE: To construct and display the keratinocyte growth factor (KGF) phage active peptides so as to detect the promoting effects of epidermal cell. METHODS: KGF sequences were chosen and their primers were designed. The selected genes of P1, P2 and P4 were obtained by reverse transcription (RT)-PCR. P3 was obtained by direct synthesis. And the KGF genes were subcloned into pComb3 vector. The technique of phage display was employed to display the genes on phage surface. Methyl thiazolyl tetrazolium (MTT) assay was used to evaluate the promoting effects of KGF phage active peptides on the proliferation of epidermal cell. Optical density (A) was determined at 570 nm. Immunofluorescent assay was employed to evaluate the cell affinity of KGF phage active peptides. RESULTS: The four KGF genes were obtained and subcloned into pComb3 vector. The proteins of the KGF genes were expressed on the surface of the pComb3 vector. The MTT data of optical density (A) showed that significant differences existed between the negative control and KGF control (0.293 ± 0.017 vs 0.520 ± 0.043) and KGF phage active peptide groups (0.293 ± 0.017 vs 0.469 ± 0.057, 0.441 ± 0.048, 0.438 ± 0.035, 0.446 ± 0.037) (all P < 0.01). The results of immunofluorescent assay indicated that KGF and KGF phage active peptides had excellent cell affinity. CONCLUSION: KGF phage active peptides are successfully constructed and displayed and they may promote the proliferation of epidermal cell.

Keratinocyte growth factor phage model peptides can promote human oral mucosal epithelial cell proliferation.

OBJECTIVE: The objective of this study was to find keratinocyte growth factor (KGF) mimic peptides by a phage display library screening and to analyze their effects on proliferation of human oral mucosal epithelial cells (HOMECs). STUDY DESIGN: A phage display library was screened by anti-KGF antibody. ELISA was performed to select monoclonal phages with higher binding activity. The promotion of the phage model peptides on HOMEC proliferation were analyzed by MTT and their cell affinities were confirmed by immunofluorescence assay. Their effect on KGFR, human beta-defensin 3, c-Fos, and c-Jun in HOMEC were analyzed by quantitative real-time PCR. RESULTS: Two model peptides with higher affinity with HOMEC were found to have promotive activity on cell proliferation, similar to that of KGF. These 2 model peptides have no KGF-like promotion effect on the expression of c-Fos and c-Jun. CONCLUSIONS: The 2 phage model peptides can promote the proliferation of HOMEC in vitro without tumorigenic effects, which suggests their possible usages in oral mucosal wound healing.

[Experimental study on the recycling of denatured acellular dermal matrix after burn].

OBJECTIVE: To explore the feasibility of burn denatured acellular dermal matrix (DADM) as dermal substitute in repairing wounds. METHODS: (1) Nine Wistar rats received a deep partial-thickness scald on the back. Full-thickness wounded skin was collected on post scald day (PBD) 1, 2, and 3 (with 3 rats at each time point), and it was treated with 2.5 g/L trypsin/0.5% Triton X-100 to remove cells to prepare DADM, respectively called DADM-1 d, DADM-2 d, and DADM-3 d. Another 3 rats without scald injury were treated with the same method as above to prepare acellular dermal matrix (ADM) to serve as control. Gross and histological observations and microbiological and biomechanical tests, including ultimate tensile strength, maximum tension, stretched length at breaking, stress-strain relationship, were conducted for the resulting ADM and DADM. (2) Another 64 rats were divided into ADM group and DADM-1 d, DADM-2 d, and DADM-3 d groups according to the random number table, with 16 rats in each group. A skin flap in size of 2.0 cm×1.8 cm was raised on the back of each rat. The above-mentioned ADM, DADM-1 d, DADM-2 d, and DADM-3 d were cut into pieces in the size of 1.8 cm×1.5 cm, and they were respectively implanted under the skin flaps of rats in corresponding group. At post surgery week (PSW) 1, 3, 5, or 9, 4 rats in each group were used to observe wound healing condition and change in implants with naked eye, and histological observation of the implants was conducted. Data were processed with one-way analysis of variance and t test. RESULTS: (1) The freshly prepared DADM was milky white, soft in texture with flexibility, but poor in elasticity as compared with ADM. No epithelial structure or cellular component was observed in ADM or DADM under light microscope. Collagen fibers of DADM were seen to be thickened unevenly and arranged in disorder and eosinophilic. All microbiological results of DADM were negative. There was no statistically significant difference among DADM-1 d, DADM-2 d, and DADM-3 d in levels of ultimate tensile strength, maximum tension, stretched length at breaking, and stress-strain relationship (with F values from 0.088 to 3.591, P values all above 0.05). Values of the above-mentioned four indexes were the highest in DADM-3 d, they were respectively (13.0 ± 2.4) MPa, (61 ± 4) N, (173 ± 7)%, (45.7 ± 2.0)%. Values of the four indexes of ADM were respectively (19.0 ± 2.6) MPa, (95 ± 4) N, (201 ± 5)%, (62.5 ± 2.2)%, which were higher than those of DADM-1 d, DADM-2 d, and DADM-3 d (with t values from 6.424 to 17.125, P values all below 0.01). (2) No exudate or swelling in the wounds of rats, and no contraction or curling of implants were observed in every group at PSW 1, but inflammatory cells infiltration and Fbs inward migration were observed in the wound. At PSW 3, the growth of hair was normal in the wound in DADM-1 d, DADM-2 d, and ADM groups, but few and scattered hair grew in DADM-3 d group. The inflammatory cells decreased, while Fbs increased, and new capillaries were found to grow inwardly in each group. The decrease in inflammatory cells was slightly delayed in DADM-3 d group. At PSW 5, hair growth became normal, and implants shrank and thinned with fiber membrane wrapped densely and bundles of ingrowing large caliber blood vessels in all groups. The dermal matrix in each group merged with the surrounding normal tissue. At PSW 9, ADM and DADM became white, thin, and soft tissue sheet which was closely connected with the inner side of the flap. There was no infiltration of inflammatory cells in implants in either group. The collagen fibers arranged regularly and densely, and they were integrated with normal collagen tissue. CONCLUSIONS: The burned DADM does not have obvious immunogenicity, but with good biocompatibility. It is prospective to become as a dermal substitute in repairing wounds.

Transforming growth factor-ß1 phage model peptides isolated from a phage display 7-mer peptide library can inhibit.

BACKGROUND: Transforming growth factor-ß1 (TGF-ß1) is known to have a role in keloid formation through the activation of fibroblasts and the acceleration of collagen deposition. The objective of this current study was to isolate TGF-ß1 phage model peptides from a phage display 7-mer peptide library to evaluate their therapeutic effect on inhibiting the activity of keloid fibroblasts. METHODS: A phage display 7-mer peptide library was screened using monoclonal anti-human TGF-ß1 as the target to obtain specific phages containing ectogenous model peptides similar to TGF-ß1. Enzyme-linked immunosorbent assay (ELISA) was performed to select monoclonal phages with good binding activity, which underwent DNA sequencing. MTT assay and apoptosis assessment were used to evaluate the biological effects of the phage model peptides on keloid fibroblasts. Immunofluorescence assay was employed to show the binding affinity of the model peptides on phages causing keloid fibroblasts. Quantitative real-time PCR analysis was carried out to detect the expressions of nuclear factor κB (NF-κB) mRNA, connective tissue growth factor (CTGF) mRNA and TGF-ß receptor II (TßRII) mRNA in keloid fibroblasts. RESULTS: Specific phages with good results of ELISA were beneficiated. Four phage model peptides were obtained. The data of MTT showed that TGF-ß1 and one phage model peptide (No. 4) could promote keloid fibroblasts proliferation, however, three phage model peptides (No. 1 - 3) could inhibit keloid fibroblasts proliferation. The results of apoptosis assessment showed that the three phage model peptides could slightly induce the apoptosis in keloid fibroblasts. The data of immunofluorescence assay revealed that the model peptides on phages rather than phages could bind to keloid fibroblasts. The findings of quantitative real-time PCR analysis suggested that the expressions of NF-κB mRNA and CTGF mRNA in the three phage model peptide groups decreased, while the expression of TßRII mRNA slightly increased. CONCLUSIONS: Three phage model peptides isolated from a phage display 7-mer peptide library can inhibit keloid fibroblasts proliferation and induce the apoptosis in keloid fibroblasts. They can inhibit the activity of keloid fibroblasts by blocking TGF-ß1 binding to its receptor and then regulating the expressions of NF-κB, CTGF and TßRII.