Experimental Study On The Application Of Irisin In Tissue Engineering Bone

Objective: To investigate the effect of Irisin on osteogenic differentiation of rat BMSCs at the cell level;to study the tissues of Irisin-modified BMSCs in tissues of SD rats The effects of engineered bone healing were explored for the feasibility of Irisin as a tissue engineering bone growth factor and its effect on osteogenesis.Methods: Rat BMSCs were cultured in vitro,separated,purified,and identified.Different groups of Irisin(0,80ng/ml,100ng/ml,120ng/ml)were set,and the proliferation activity of BMSCs in each group was detected by CCK-8 method.The best Irisin was selected.Experimental concentration.Rat BMSCs were cultured with Irisin-containing culture medium as the experimental group,and the control group was cultured with simple medium.Alizarin red and von Kossa calcium staining were performed on days 3,7,and 14.Bolts detect mRNA and protein expression of osteogenesis-related genes.The rat models were divided into three groups: the experimental group: Irisin composite scaffold;the control group: no Irisin composite scaffold;Three groups of materials were implanted into the defect area of the rat model.MicroCT,HE staining and immunohistochemical analysis were performed to detect bone healing and cortical bone regeneration in the defect area.Result: 1.Flow cytometry showed that CD29(98.4%±2.1%)was expressed at a high level on the BMSC surface,and CD45(2.9%±0.4%)was expressed at a low level.2.The effect of Irisin at a concentration of 100ng/ml on promoting the proliferation of rat BMSCs cells was gradually apparent from 3 day(P<0.01).3.The staining depth,calcium nodule size and number of alizarin red and calcium salts in the experimental group were significantly higher than those in the control group.4.Compared with the control group,the expression levels of ALP,Lrp5,BMP2,and Smad1 were significantly increased,but Sost were significantly decreased,with significant differences(P<0.001).5.MicroCT results of animal experiments showed that the defect area of the blank group did not show newlyformed bones up to 3 months after surgery.In the experimental group,some scattered newly formed bones began to be seen 1 month after transplantation.Compared with the control group,the newly formed bones increased significantly and filled the entire defect area at 3 months after transplantation.The quantitative analysis of the defect area showed that compared with the blank group,and the bone volume fraction and bone minerals of the newly formed bones in the experimental group.The density and cortical bone volume were significantly higher than control group(P<0.05).The HE staining results showed that the stent material transplantation treatment of bone defects induced an increase in the amount of new bone,especially when the stent was compounded with Irisin,the use of Irisin composite scaffolds caused new bone tissue to surround and gradually replace the stent,which closed the surgical defect And more angiogenic areas.The immunoh-istochemical results showed that the positive expression of Sost protein could be seen in the transplanted area of each group(P <0.05).The mean optical density value of positive expression of Sost protein increased slightly with the extension of the transplantation time.Conclusions: 1.Irisin can promote proliferation and osteogenic differentiation of rat BMSCs in vitro,also Irisin can inhibit the expression of Sost gene.Wnt and BMP signaling pathways may play a role in Irisin promoting osteogenic differentiation of rat BMSCs.2.Irisin composite tissue engineering scaffold can promote the repair of critical bone defects.Irisin has good osteogenesis ability in vivo and in vitro,and is expected to become a growth factor for tissue engineering bone with promising prospects.