A Study Of Artificial Silk Ligament Composite Bone Marrow Mesenchymal Stem Cells To Repair Anterior Cruciate With Injury

ObjectiveArtificial silk ligament as biological material support, has good biologicalcompatibility, no cell toxicity, don’t cause inflammatory reaction, in tissue engineering hasbeen widely used. Silk degumming, through the many kinds of methods to increaseartificial silk ligament pore rate, improve its and peripheral things contact surface area, andthen increase the bone marrow mesenchymal stem cells in the silk ligament of adhesionrate. Through certain concentration bone marrow mesenchymal stem cells in the artificialsilk silk ligament on attached growth, make compound biological material, alternative NewZealand white rabbit of anterior cruciate ligament in knee joint micro environment andunder dynamic action, to observe the change of composite materials, and with normalligament do contrast. For artificial tendon and ligament tissue engineering materialproduction to provide certain feasibility.Methods1. Experimental process1.1Making artificial silk ligament, with pancreatic enzyme or soda ash fordegumming, then and Ⅰ type collagen composite, freeze drying, again with EDC (1-ethyl-3-(3-dimethyl ammonia propyl) carbon diimine)/NHS (N-hydroxy amber imide) ascrosslinking reagent for collagen cross-linking, again freeze drying, ethylene oxidesterilization standby.1.2Select about a week or so New Zealand small white rabbit, take the bonemarrow mesenchymal stem cells, subculture parallel flow cytometry instrument detectioncell types. Take the fourth generation of cells, and spare artificial silk ligament compound7days to form a composite material.1.3A closed group of New Zealand white rabbit only36, randomly divided into3groups (blank group, the experimental group and the control group), each group of12only. In the same conditions the anterior cruciate ligament were removed, retain residual end. Inthe experimental group, the use of composite reconstruction BMSC.2. Observation index2.1Different scheduled time the four animals were randomized to death, observe thecomposite material histology, pathology morphology change and bone and compositeconnection interface structure.2.2The scheduled time different random line experimental animal imaging andbiomechanics procuratorial.3. Statistical AnalysisSelected by using SPSS software, line matching t test, single sample t test,2x4factorial analysis, statistical analysis.Results1. Artificial silk ligament of the porositySilk sericin ligament surface because the existence of silk fiber tightly wrapped inbonded together to form a complete silk, silk surface that rough, but can see the single silkfiber (FIG.5); Degumming, silk surface of sericin removal and very smooth, here is asingle of silk fibers (figure6). After degumming artificial silk ligament and collagencomposite, forming on the surface with a layer of collagen layer, the structure is veryuneven, surface area is increased, making silk ligament surface become rough, porosity,and promoted the increase of BMSCs attached.2. Bone marrow mesenchymal stem cell cultureThe size of a small white rabbit put to death in New Zealand, immediately take itslimbs dry epiphysis end medullary cavity cell culture. After forty-eight hours first in liquid,with PBS buffer gently wash, remove all cells, purification, BMSCs were observed underinverted microscope, the cells attached growth, irregular spindle. Take some cells intosuspension after flow cytometry instrument testing, the results for CD34+cells CD44+10%and,42%and, for the performance of the stem cell characteristics (adherent culturemethod: CD34negative, CD44, CD50, STRO-1positive).3. Graft indexThe graft surrounded by a large number of soft tissue around the bag, there are a fewinflammatory cells infiltration. And the bone at the joints have Sharpey structuregeneration, bone cell growth is strong. ACL residual end encysted graft, pathologicalsection shows that structure close to normal ACL. And the control group in addition tohave a large number of inflammatory cells infiltration outside, no other corresponding performance, between the two groups have statistical significance (P <0.05).Conclusion1. Ⅰtype collagen protein and silk ligament composite can increase the surface areaof the ligament, improve the porosity, and promote the attached BMSCs.2. Artificial silk ligament and BMSCs after compound can promote ligament andbone healing, and in the knee joint micro environment there is normal ligament metaplasticsigns.