Study On The Mechanism Of Taohong Siwu Decoction Mediating TLR2 To Inhibit Osteoclast Differentiation And Promote Fracture Healin

Objective: In this study,a fracture model was constructed by mice to confirm that THSWD can accelerate the healing of fractures.Then,through cell experiments,it was explored whether THSWD inhibited osteoclast differentiation,delayed bone resorption,and finally accelerated fracture healing by mediating the expression of TLR2.From the perspective of TLR signaling pathway,the intrinsic molecular mechanism of THSWD-mediated TLR2 expression inhibiting osteoclastic differentiation and promoting fracture healing during fracture healing was studied,and the intrinsic mechanism and new target of THSWD promoting fracture healing were further revealed.Methods: Part I:48 healthy C57BL/6 mice with a body weight of(23±2)g were taken and divided into two groups according to the principle of random allocation,namely the model group and the THSWD group,with 24 mice in each group,and a mouse fracture model was constructed;Micro-CT imaging and quantitative analysis of BV and BV/TV were performed in the model group fed normal saline and the group with THSWD intervention.Part 2: RAW264.7 cells were randomly divided into control group,THSWD group,TLR2 group and THSWD+TLR2 group,a total of 4 groups,and the TRAP staining,q PCR and WB of the four groups were detected.Result:1.It can be seen intuitively through CT that compared with the model group,the effect of fracture healing in the THSWD group is significant.2.BV,BV/TV: Compared with the model group,BV and BV/TV were significantly increased in the THSWD group on the 7th and 14 th days of fracture,and the difference was statistically significant(P<0.01).3.TRAP staining: Compared with the control group,the number of active osteoclastic cytoplasmic nuclei in the THSWD group was reduced,a large number of nuclei could be seen in the osteoclast cytoplasm of the TLR2 group,and the volume increased significantly,compared with the TLR2 group,the number of nuclei in the osteoclast cytoplasm of the THSWD+TLR2group was reduced,and the volume was also significantly reduced.4.q PCR: Compared with the control group,the expression of TLR2 m RNA in the THSWD group was significantly down-regulated with a statistically significant difference(P<0.05),and compared with the TLR2 group,the expression of TLR2 m RNA in the THSWD+TLR2 group was significantly downregulated with a statistically significant difference(P<0.01).5.WB: Compared with the control group,the expression of TLR2 protein in THSWD group was significantly downregulated with a statistically significant difference(P<0.05),and compared with the TLR2 group,the expression of TLR2 protein in THSWD+TLR2 group was significantly downregulated with a statistically significant difference(P<0.01).Conclusion:1.THSWD can speed up the healing time of fractures in mice and can promote fracture healing.2.THSWD inhibits the expression of TLR2,inhibits osteoclast differentiation,delays bone resorption,and ultimately accelerates the speed of fracture healing.3.By elucidating the molecular mechanism of THSWD to promote fracture healing,it provides a potential entry point for the development of new fracture healing treatment drugs and intervention programs,and also combines the theory of “blood circulation and stasis” and“osteoclastic differentiation” in traditional Chinese medicine,enriching the modern connotation of “blood circulation and stasis”.