The Research On Homing Of ADSCs And EPCs By Collagen-Adipose Tissue System

Objective:The collagen-fat joint system was constructed by the fusion of adipose tissue and collagen material.The labeled adipose stem cells(ADSCs)and endothelial progenitor cells(EPCs)were exogenously given to explore whether collagen could promote the homing of ADSCs and EPCs into adipose tissue and play the biological effects of ADSCs and EPCs.To improve the survival rate of fat transplantation.Methods:(1)SD rats were sacrificed after cervical dislocation,and bilateral femur was taken for isolation,culture and identification of endothelial progenitor cells(EPC),and bilateral inguinal adipose tissue was taken for isolation,culture and identification of adipose stem cells(ADSCs).(2)Stem cell homing observation:grouped according to animal experiment,fat in the groin area was taken and prepared to the size of granular fat.400ul of the prepared granular fat was fully mixed with 300ul of collagen(0.1mg/ml),and the mixture was transplanted into the back of experimental SD rats to prepare the collagen-fat combined system.Fluorescent and dye-labeled ADSC and EPC were injected from the tail 2 and 14 days after transplantation,respectively.In vivo imaging,electrophoresis test,RT-qPCR quantification,and confocal microscopy were used to observe the homing of stem cells.(3)Biological effect observation of stem cells:grouped according to animal experiments,and prepared collagen-fat combination system with homing experiment.The removal time after fat transplantation was divided into PBS group,collagen group,ADSC group and EPC group.The changes of fat transplantation in different groups were observed through the shape,volume,weight,HE,Masson and CD31 of fat transplantation.Results:(1)In vivo imaging detection:stem cells were injected 2 days and 14 days after transplantation,and fluorescence signals were concentrated in the abdomen and tail of the rats.(2)Electrophoresis showed that LUC gene was successfully amplified from transplanted fat in all groups.ADSC was injected 2 days after transplantation:the electrophoresis gray level of collagen group and PBS group was very shallow,and there was no difference.ADSC and EPC were injected 14 days after transplantation.The electrophoresis gray level of the collagen group was significantly deeper than that of the PBS group.(3)RT-qPCR quantification:Luc gene was successfully quantified from the transplanted fat in each group.ADSC was injected 2 days after transplantation:gene quantitation in the collagen group and PBS group was very low,and there was no difference.ADSC and EPC were injected 14 days after transplantation.Gene quantity in collagen group was significantly higher than that in PBS group.(4)Confocal microscopy:from the transplanted adipose tissue sections,the dye-labeled stem cells in the collagen group were significantly more than those in the PBS group.(5)Observation of stem cell biological effect:①He and Masson staining showed that the inflammation in the PBS group was severe,with many vacuoles and fewer adipocytes,while the inflammation in the other groups was significantly less,and there were many new adipocytes in the ADSC group.②CD31 immune tissue showed the number of blood vessels,ADSCgroup>EPCgroup>Collagen group>PBS group.③The volume and weight of transplanted fat in ADSC and EPC groups were higher than those in collagen and PBS groups.Conclusions:(1)For the luciferase labeling of ADSCS-Luc gene,the transfection efficiency of vector mCherry was better than that of ZSGREEN1;(2)Collagen-fat combination system can promote the homing of ADSC and EPC to the transplanted adipose tissue;(3)Fluorescence gene quantitative detection and confocal microscopy detection of stem cell homing is more sensitive than bioluminescence in vivo imaging.(4)Collagen combined with ADSCs and EPCs can stimulate vascular remodeling of the transplanted adipose tissue,improve the survival rate of the transplanted adipose tissue,reduce inflammation,and reduce the formation of vacuoles.(5)Collagen combined with ADSCs can significantly promote the formation of adipocytes.