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Objective ultrastructure observation of a new pig collagen-based dermal bracket (PCDM), organizations and its cytotoxic detect, provide the basis for the application of experimental and clinical safety of this new type of biological material. Novel porcine collagen based dermal animal Norplant, the growth of vascular endothelial cells in the dermal fibroblast cells was observed, as well as dermal vascular, of modified heterologous dermal substitute, improve its histocompatibility method. Method 1) by HE staining and immunohistochemical staining and scanning electron microscopy porcine collagen-based dermal (PCDM) organizational structure and allogeneic acellular dermal matrix (HADM) compared to the organizational structure of porcine acellular dermal matrix (PADM). 2) obtain different concentrations of PCDM extract, positive control, negative control and blank control and L-929 cells for the detection of cells. Draw different inoculum concentrations the laboratory standard of the L-929 cell growth curve, determine the appropriate concentration of cells were seeded different concentrations of extracts of dermal cell proliferation was detected by MTT, medical biomaterials cytotoxicity, level standard rating. 3) 24 wistar rats Norplant dermal substitutes into the PCDM group and HADM group 3,7,14,30 days after the implants were removed tissue line graft generally observed by HE dyeing, CD34, factor VIII immunohistochemistry staining, into fiber cells, vascular endothelial cells in the distribution of the dermal matrix proliferation, vascularization process and histological changes were observed. 1) histological observation: complete removal of the light microscope PCDM epidermis, skin appendages no cellular components, blood vessels, hair follicles and sebaceous glands in the dermis, the three-dimensional structure of the collagen matrix remained intact. Smooth and complete nipple level, immunohistochemistry see above there is a very thin layer linear brown stained areas, suggesting that basement membrane integrity under parallel with the basement membrane, the regular arrangement of collagen fibers, papillary layer of collagen arranged relatively dense; net The rough surface like layer, visible broken stump of the collagen fibers, collagen bundles compared with papillary layer thick, loose. Scanning electron microscopy revealed dense ordered the PADM and PCDM of collagen fibers, structure extremely similar HADM, porcine dermal collagen fibers than the human dermal arranged more closely, suggesting that porcine dermal collagen fiber content greater than human dermal collagen fibers content. The cross-sectional scanning electron microscopy showed: PCDM collagen bundles arranged than HADM the PADM loose similar. 2) the number of cells inoculated set at 5 × 10 3 / well, and 1 × 10 4 / hole, within a week L929 growth curve can be seen was a positive linear correlation between the amount of cells with metabolic proliferation rate, good cell viability, measured The data is more stable, most appropriate Determination of PCDM cytotoxic. Cytotoxicity testing methods using standard PCDM, cell proliferation degrees with the blank control, and negative control was no significant difference, was significantly better than the positive control group, and the cytotoxic level score of 0 to 1, can be considered PCDM no significant cytotoxicity. 3) implants experiment general observation: the postoperative rats wound the skin around no obvious swelling and inflammation, wound healing good graft close contact wound. Histological observation: Norplant 3 days, groups were seen in fibroblasts (fibroblast, FB) invasive, neutrophils, lymphocytes, and a small amount of macrophage infiltration. Graft and surrounding tissue connection seen at the start hyperplasia tiny new blood capillaries bud, differences between groups were not statistically significant (P> 0.05); 7 days, the inflammatory cells in less than 3 days, shows a large number of groups transplant intradermal FB invasive, hyperplasia, along the direction of the distribution of collagen fibrils; extension of new blood capillaries within the graft. 14 days after the graft and demarcated by the bed surrounding tissue, inflammatory cells within the tissue to further reduce the infiltration of fibroblasts and endothelial cells (vascular endothelial cell, VEC) hyperplasia, newly formed capillaries reached graft internal, the number further increased, and over the three days, 7 days, the difference was statistically significant (P <0.05); 30 days, the distribution of blood vessels throughout the graft, fibroblasts, vascular endothelial cells to mature further, but the number of did not increase significantly, the difference was not statistically significant (P> 0.05), and basically did not see inflammatory cells. Conclusion 1) process through a variety of physical and chemical methods, this pig collagen-based dermal is a non-toxic, safe new natural xenogeneic dermal scaffold. 2) FB, VEC proliferation in porcine collagen-based dermal good, and visible cell moved into the stent internal, neovascularization mature porcine collagen based dermal has good tissue compatibility, there is no the xenoantigenic hinder fibroblasts immigration of vascular endothelial cells, is a good natural dermal substitutes.
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