The Therapeutic Effect And Mechanism Of Bone Marrow Mesenchymal Stem Cells On Deep Ⅱ Skin Burns In Hamsters

objectivesTo observe the growth and biological characteristics of rat bone marrow mesenchymal stem cells(BMSCS), establish the stable amplification system on rat BMSCS. Explore a simple and convenient,easy to control and high repeatability burns animal model, groping the method control of skin scaldburn area, the scalded time, scald the extent.To explore the mechanism of treatment of bonemesenchymal stem cells transplant for skin mast cell chymotrypsin skin deep Ⅱ degree burn.MethodBMSCS were isolated from the bone marrow of young rats were purified amplified by the method ofwhole bone marrow culture. Establish a stable amplification system, and identified P5cells byobserving the cultured cell morphology and biological characteristics9healthy adult hamster,intraperitoneal anesthesia with ketamine by Weight (45mg/kg), After the animals were anesthetized,shave and clean the abdominal area by sodium sulfide. Divided into three groups,65℃12s,75℃12s,85℃12s,3each group.Put50ml syringe to the back of the animal with a diameter of3cm to thebottom. Each group, by the time20ml hot water burns,30min observed skin changes after burnwound. And12hours after the burn wounds in each group derived histological examination, Andmodel comparison, identify the conditions for making deep Ⅱdegree burn model.40hamsters copyskin deep Ⅱ degree burn model and randomly divided into2groups: stem cell transplantation group（group A n=20）and model group（group B n=20）, cultured BMSCs in vitro to the5th generation（number of1×10~7/ml）, by local injection and surface smear supernatant transplanted to the burnarea of stem cell transplantation group, The control group given saline, observed wound change,granulation tissue growth and wound healing, histological observation were1,3,7,14days aftertransplantation, each group of five for hematoxylin the Seiichi eosin (HE) staining of tissuespecimens by conventional methods Different time points of wound tissue, expression level ofdetection by real-time PCR technology for skin mast cell chymotrypsin.resultSeparation of bone marrow cells by whole bone marrow, we can obtain a number of cells, more rapidproliferation of cells adherent, adherent cells covered the entire culture bottles at the bottom of thetime required for6-7d, Primary cells is not high purity, after passage, we can obtain higher purity ofMSCs, MSCs colony formation of fibroblast-like cells in about3-5days.Observation of the three modeling methods，all died after12hours in85°C for12seconds groups，65°C for12seconds and75°C for12seconds groups all survived.75℃12seconds group with poor mental state. Bypathological observation,65℃12seconds group failed to reach deep Ⅱdegree burn requirements.75℃12seconds group complies with deep Ⅱdegree burn model requires. Stem cell transplantationand control groups were observed animal skin wound healing after deep Ⅱdegree burn instantly,after Skin irritation6h, Skin irritation, Burns24h, Epidermis begins to harden,72h stem celltransplantation skin begins to form eschar, A, B groups had no significant difference in the scab time（P>0.05）. In terms of healing rate, To90%for the healing of the reference standard, Scald14d, A, Bbetween the two groups was significant.Stem cell transplantation group and control group tissuemorphology were observed after burn7d, fibroblasts were observed by the microscope, the number ofcapillaries increased,and formed abundance and granulate and collagen deposition.The chymotrypsin gene detection after burn wound1,3,7,14d organization by quantitative PCRmethod, chymotrypsin gene were detected in several time segments, the difference between the twogroups was not significant in terms of gene detection, compared the mRNA levels of chymotrypsindetection time point,3d is the highest expression,compared with control group, MRNA levels in thepresence of a significant difference in the transplantation group7,14d.conclusionThis study established the method to isolate、purify、amplify hamster bone marrow MSCs in vitroculture. A simple and convenient, easy to control and high repeatability scalded animal model weremade.It had been proved that BMSCs by stem cell transplantation can promote wound healing, skinmast cell chymotrypsin may be involved in the process of wound healing.