Study Of Peptide SIKVAV Modified Chitosan Hydrogels In Skin Wound Healing

1.The effect of peptide SIKVAV on dermal fibroblasts proliferation,migration,secretion growth factors and synthesis collagen in vitro.1.1 ObjectiveTo investigate whether SIKVAV promotes fibroblasts proliferation,migration,secretion growth factors and synthesis collagen in vitro.1.2 MethodsPrimary fibroblasts were obtained from Neonatal C57BL/6 mice via trypsin digestion and tissue adherence.CCK-8 was performed to detect the effect of peptide SIKVAV on proliferation of fibroblasts.Transwell assay was used to detect effect of peptide SIKVAV on the longitudinal migration of fibroblasts.Scratch test was performed to detect the effect of peptide SIKVAV on the transversal migration of fibroblasts.ELISAS was used to determine the effect of peptide SIKVAV on fibroblasts secretion growth factors and synthesis collagen ? and ?.1.3 ResultsCCK-8 assay showed that peptide SIKVAV significantly promoted the proliferation of fibroblasts(P<0.05)in concentration-dependent manner.The result of Transwell assay also demonstrated that peptide SIKVAV significantly promoted the longitudinal migration of fibroblasts(P<0.05)in a concentration-dependent manner.The result of scratch test demonstrated that peptide SIKVAV significantly promoted the transversal migration of fibroblasts(P<0.05)in a concentration-dependent manner.The results of ELISA assay showed that SIKVAV promoted fibroblasts secretion EGF,bFGF,TGF-?1 and VEGF(P<0.05),and synthesis collagen ? and ?(P<0.05)in a concentration-dependent manner.1.4 ConclusionThe above results demonstrate that peptide SIKVAV promotes the proliferation,transversal and longitudinal migration,secretion growth factors and synthesis collagen ? and ?,which suggests that peptide SIKVAV can involve in skin wound healing by promoting fibroblast biological activity.2.The effect of peptide SIKVAV modified chitosan hydrogels on reepithelialization,angiogenesis,collagen deposition and inflammation of skin wound.2.1 ObjectiveTo explore whether the the peptide SIKVAV modified chitosan hydrogels can accelerate reepithelialization,angiogenesis,collagen deposition and inhibit inflammation in the skin wound.2.2 MethodsThe chitosan hydrogels and peptide SIKVAV modified chitosan hydrogels were prepared and characterized by 1H-NMR.After skin trauma model of mice was established,the mice were then randomly divided into four groups,including a negative control group and Peptide group and Chitosan group and Peptide+Chitosan group.General skin wound of mice was observated and residual area of wound was statistically analyzed.HE staining was performed to detect the wound reepithelialization.The deposition of collagen was observed by Masson trichrome staining.The expression of F4/80,?-SMA,K1,K6,CD86,CD163 and CD31 were detected by immunohistochemistry in the skin wound.2.3 ResultsBoth hydrogels also have a certain stiffness.Scanning electron microscopy(SEM)showed that the pore size was homogeneous,and the peptide SIKVAV was confirmed to be combined to chitosan by 1H-NMR.The result of general skin wound showed that the wound healing effect in Peptide+ Chitosan hydrogel group was better than the other three groups.Peptide-modified chitosan hydrogels statistically enhanced the wound closure compared with the other three groups(P<0.05).HE staining showed that the peptide-modified chitosan hydrogels promoted the reepithelialization of the skin wound than the other three groups(P<0.05).Masson trichrome staining demonstrated that the deposition of collagen fibers in Peptide+Chitosan group was much more than the other three groups.The results of immunohistochemistry showed that the newborn vessel was more in the Peptide+Chitosan group than in the other groups(P<0.05).The ?-SMA expression in Peptide+Chitosan group was significantly higher than in the other three groups(P<0.05).Immunohistochemical results showed that the expression of K6 and K1 in Peptide+Chitosan group were significantly higher than in the other three groups(P<0.05),the number of F4/80 positive and CD86 cells in Peptide+ Chitosan group were less than the other three groups,(P<0.05),but CD 163 positive cells were increased compared with the other three groups(P<0.05).2.4 ConclusionIn vivo studies showed that the peptide SIKVAV modified chitosan hydrogels promoted the proliferation,migration and differentiation of keratinocytes,angiogenesis and collagen deposition,and inhibited wound inflammation in the skin wound.Therefore,the SIKVAV modified chitosan hydrogel can be used as a synthetic material in the treatment of skin wound.It provides a new theoretical basis for wound dressing clinical research and application.