| Skin injury is a common disease in animal clinic,which not only destroys the barrier function of skin,but also leads to the death of the sick animal,causing the animal owner to suffer varying degrees of economic loss.At present,there are some limitations in the clinical treatment of skin trauma.Bovine Achilles Collagen is a natural biomaterial with advantages of low toxicity,low immunogenicity and good biocompatibility.In recent years,its application in biomedical materials has attracted more and more attention.Mesenchymal stem cells(MSCs)are a kind of cells with the potential of self-renewal and multidirectional differentiation.They are widely derived,low immunogenicity and easy to obtain.In the process of culture,MSCs secreted exosomes and a variety of cytokines into the culture medium,which had the ability to promote the body wound repair.Therefore,in recent years,the application of mesenchymal stem cell exosomes as a new cell-free therapy in skin wound repair has attracted wide attention.As the supernatant of MSCs containing exosomes is highly liquid,it cannot stay at the wound site for a long time,making it difficult for exosomes to play a full role.This study by people between umbilical cord mesenchymal stem cells in supernatant combined with bovine tendon collagen production mixture,by in vitro cell scratch test and mouse skin trauma test,study whether this mixture has the role of promoting wound healing,to develop a new type of liquid skin wound repair,new treatment method for treatment of skin wound.The main results are as follows:(1)Collagen was extracted from bovine Achilles tendon by acetic acid and pepsin binding method,and the type of bovine Achilles tendon collagen was identified by SDS-PAGE gel electrophoresis.The results showed that the extracted bovine Achilles tendon collagen was type I collagen.BCA protein concentration kit was used to detect the concentration of bovine Achilles tendon collagen,and the results showed that the concentration of bovine Achilles tendon collagen extracted by this method was in the range of 1.25-2.8 mg/m L.(2)Human umbilical cord mesenchymal stem cells(HUCMSCs)were inoculated into a 24-well plate coated with 0.2 mg/m L bovine Achilles tendon collagen for osteogenic and chondrogenic differentiation.The results showed that bovine Achilles tendon collagen had no effect on the osteogenic and chondrogenic differentiation potential of HUCMSCs.HUCMSCs were inoculated into a 24-well plate coated with bovine Achilles tendon collagen for growth curve test.The results showed that bovine Achilles tendon collagen had no significant effect on the growth and proliferation of HUCMSCs.(3)MTT cytotoxicity kit was used to detect the cytotoxic effects of bovine Achilles tendon collagen on HUCMSCs,human skin fibroblasts and human skin keratinocytes,and the results showed that the extracted bovine Achilles tendon collagen had good biocompatibility to the above three kinds of cells and had no cytotoxic effects.(4)The scratch test of human skin fibroblasts and keratinocytes was carried out in a6-well plate coated with bovine Achilles tendon collagen.The results showed that bovine Achilles tendon collagen could promote the scratch healing of the two kinds of cells.HUCMSCs supernatant inhibited the scratch healing of human skin fibroblasts and promoted the scratch healing of human skin keratinocytes.(5)The mixture of bovine Achilles tendon collagen and HUCMSCs supernatant was used to treat skin wound in mice.The results showed that the mixture could significantly promote skin wound healing in mice compared with the control group on day 15. |
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