| Objective1.By improving the physical and chemical method,the New collagen sponge scaffold(NCSS)and micronized acellular dermal matrix(MADM)were prepared,The feasibility of these two kinds of dermal collagen scaffolds as cell carriers was evaluated.2.NCSS and MADM combined with stromal vascular fraction(SVF)in the repair of full thickness skin wound in nude mice was investigated.Methods1.NCSS was obtained after repeated freeze-thaw combined with hot sodium hydroxide hydrolysate treatment,and MADM were obtained after filtration by freeze-drying and grinding method.The collagen spatial structure,pore cavity size,in vitro degradation time and cytotoxicity of the two kinds of dermal collagen scaffolds were detected by HE staining and scanning electron microscopy(SEM).2.Human SVF were extracted via enzyme digestion from human adipose tissue.3.The skin substitute was constructed using SVF combined with NCSS and MADM.15 mm~2full-thickness skin wounds were designed on the back of female nude mice,respectively.Mice were divided into 4 groups in the experiment:SVF-NCSS(group A),SVF-MADM(group B),simple SVF(group C)and blank control(group D).The wound healing rates were observed at 7?14 days after operation,and specimens were taken at 7 and 14 days for histological detection and immunohistochemical cluster of differentiation 31(CD31)vascular density detection,respectively.Results1.Improved methods can completely remove the components of the cell and cell debris,all both can maintain the integrity of the scaffold;NCSS was milky white or light yellow,showing soft,loose porous homogeneous shape,and the surface loose pores could be clearly observed;NCSS showed loose collagen fibers,slender fibers,complete structure of collagen fibers and uniform pore distribution,MADM showed that collagen was fine,discontinuous between collagen,loose structure,different forms of collagen,different diameters and sizes of collagen,mild red staining,loose and porous collagen bundles,The pore diameter(181.21±66.96)?m and porosity(93.1±1.02)%of the dermal reticular surface of NCSS,the pore diameter and porosity were significantly higher than that of ADM prepared by traditional methods,P<0.05 was statistically significant.In vitro degradation time of scaffold materials prepared by the two methods was not significantly different from that of traditional ADM.There was no obvious cytotoxicity in the infiltrating solution.2.At 3 days after SVF were inoculated onto NCSS and MADM,fluorescent dye staining showed that the infiltration of SVF could be seen within NCSS and MADM.the number of cells was also obviously increased.3.The wound healing rates at 7,and 14 days after operation in group A and group B were87%±2%?99%±1%;86%±3%?99%±1%,respectively,which were significantly higher than those in groups C-D;the differences were statistically significant(p<0.05).The detection of regenerated wound tissue thickness at 1 and 2 weeks after operation and CD31 vascular density at 1 week after operation showed that regenerated tissue thickness and vascular density in group A and group B were significantly higher than those in other groups;the differences were statistically significant(p<0.05).After the transplantation of tissue-engineered skin constructed by human SVF combined with NCSS and MADM,the quality of wound healing in nude mice can be significantly improved,and the wound repair can be promotedConclusion1.NCSS and MADM prepared by the improved method were completely removed from the cell components,and the 3d structure of the dermal scaffold was retained.Due to loose structure,high porosity,good cell compatibility and cell permeability,they can be used as a good transport carrier for SVF wound repair.2.NCSS and MADM with SVF can repair wounds with full-thickness defect,improve the wound healing rate,increase the thickness of new tissue and local microvascular density,promote skin healing,and improve the quality of wound healing. |
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