Study On The Interactions Between Biological Tissues And Tilapia Collagen

In medical aid,it is very common situation that acute,superficial wounds may cause by bruises,cuts,and minor burns.Usually,these minor injuries do not require the help from medical practitioners,only need the uses of first-aid dressings to treat the wounds.Emergency dressings provide a protective barrier against dirt,bacterial contamination and pain caused by mechanical factors?stress and friction?.Despite the availability of modern treatments,chronic wounds such as venous congestion ulcers,arterial ulcers,diabetic ulcers,bedsores,traumatic ulcers and post-operative wounds are still one of the main causes of hospital visits and hospitalization.Wound healing is an important and complex,which involved many stack processes,including hemostasis/inflammation,proliferation,and remodeling.The utilization of fish processing waste is an ideal measure to maintain the environment.According to the current situation,waste utilization is mainly the recovery of valuable products from fish waste,because fish waste is a potential source of bioactive protein,mineral,fish oil and enzyme.Type I collagen,a major structural protein of vertebrates,has good biodegradability,biocompatibility and weak antigenicity,It is widely used in food,cosmetics,pharmaceuticals,tissue engineering and biomedicine.At home and abroad,the study of fish collagen is mainly about extraction,biochemical characteristics and mechanical properties.However,the role of Type I collagen in the biological role in the body has been reported by few researchers.Therefore,it is of great scientific significance and application value to study the implantation characteristics of marine collagen and the interaction between collagen and tissue-related cells.The histocompatibility of Tilapia type ? collagens such as pepsin soluble collagen?PSC?and acid soluble collagen?ASC?as biomaterials was studied in order to provide data support for their application in tissue engineering and biomedicine.In the first chapter,the structure and biochemical properties of type ? collagen were introduced.The review is about the interaction of type ? collagen with the surrounding environment.This part introduced several kinds of cells in skin and connective tissue,including fibroblasts,macrophages and mast cells,and describes the characteristics of these cells.The safety evaluation of biomaterials used in medical tissue engineering was introduced in order to ensure the safety and practicability in the process of application.The second chapter introduced the isolation and purification of Tilapia type ? collagen.Type I collagen in this whole project was extracted from Tilapia skin by pepsin,purified and freeze-dried.The purity and molecular weight of tilapia acid-soluble type ? collagen were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis?SDS-PAGE?.SDS-PAGE showed that tilapia collagen consisted of two?₁ chains and one?₂ strand,which has the characteristics of type ? collagen.The molecular weight of the?chains is about 116 to130 kDa,and the molecular weight of the polymerization?chain is about 200 kDa.There were no other obvious bands in the image,which indicated that the high purity of collagen extracted from Tilapia skin.Scanning electron microscopy?SEM?and atomic force microscopy?AFM?can be used for more in-depth research.From the results,it is confirmed that Tilapia collagen had the characteristics of type ? collagen.In the third chapter,the effects of Tilapia skin type ? collagen on mouse mast cell line P815 were studied.In the test of cell proliferation,when the concentration of collagen was 10?g/mL,the activity of P815 cells was decreased.In the process of cell degranulation,there was no significant difference between the PSC group and the control group,and the secretion of inflammatory cytokines by mast cells was not significantly increased in the PSC group,and the phenomenon of calcium influx was within an acceptable range.In chapter 4,the effects of Tilapia skin type ? collagen on mouse macrophage Ana-1 were studied.The results showed that Tilapia skin collagen and other types of collagen I had an effect on the growth of Ana-1 cells.Tilapia skin collagen could inhibit the growth of Tilapia cells at a specific concentration?1?g/mL?,and the type ? collagen from tilapia skin could inhibit the growth of mouse macrophage Ana-1 cells at a specific concentration?1?g/mL?.When the concentration of collagen was 10?g/mL,the collagen of tilapia skin promoted the growth of mouse macrophage Ana-1.Macrophage migration and phagocytosis after the action of collagen can be considered as the feedback of inflammatory response,on the other hand,it is also beneficial to the external response of the organism.Except for TNF-?,the secretion of IL-6 and IL-1?from macrophages stimulated by Tilapia collagen was lower than that of bovine calcaneus tendon collagen.In chapter 5,the effects of Tilapia skin collagen on human skin fibroblasts were studied.By scanning electron microscopy,we can see that HFF-1 can grow on collagen of tilapia skin.When the concentration of collagen was 10?g/mL,fibroblasts grow well,and the adhesion rate of collagen from tilapia and bovine calcaneus tendon collagen was high.In the cell migration experiment,fibroblasts were stimulated by collagen to extend out of the pseudopod and grow into the blank area.The amount of COL-I secreted by fibroblasts increased significantly to 9.89?g/L.ASC,on the other hand,significantly increasedthe amount of COL-III secretion.Tilapia skin collagen down-regulated the expression of MMP-1 and effectively controlled the process of wound healing.In chapter 6,the implantation characteristics of collagen from tilapia skin were studied.Intradermal stimulation was performed on the back of New Zealand white rabbits.Edema and erythema were measured by intradermal stimulation scoring system.Collagen and control samples were implanted on both sides of the spine of rabbits.The skin tissues were killed at 1^st,4^thh and 8^thh weeks after implantation.Biochemical indexes including IL-4 and IL-6 of intradermal stimulation test and implantation test were measured.The results showed that the Tilapia type ? collagen was low irritating and well implantable.A certain dose of Tilapia type ? collagen extract was injected into New Zealand white rabbits by ear vein.The results showed that the body temperature of New Zealand white rabbits was lower than 0.6?.The pyrogen reaction of Tilapia type ? collagen was determined to be in accordance with the regulations.The seventh chapter mainly includes the characteristics of antibacterial and biofilm formation of collagen from tilapia skin.The inhibitory effects of Tilapia type ? collagen on E.coli,S.aureus and P.aeruginosa were evaluated by inhibition circle test,growth curve test and conductivity of bacterial solution.The results showed that Tilapia type ? collagen had good bacteriostatic effect on S.aureus.Under the scanning electron microscope,it was found that collagen caused mechanical damage to the bacterial cell wall,and the bacteria shrank,resulting in morphological changes.The results of crystal violet test and colony count were consistent with the characteristic of anti-biofilm.The effect of Tilapia type ? collagen on E.coli was consistent.It did not inhibit the formation of biofilm,but inhibited S.aureus,P.aeruginosa and S.epidermidis to varying degrees.