To Investigate The Effect Of Taohong Siwu Decoction On The Proliferation And Osteogenic Differentiation Of BMSCs Based On Hippo Signaling Pathway

Objective: To observe the effect of Taohong Siwu Decoction on proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs)by using Taohong Siwu Decoction containing serum and Hippo pathway inhibitor verteporfin.Methods: 1.Ten SD rats were selected and BMSCs were cultured in vitro by whole bone marrow adherent method;2.Forty healthy Wistar rats of one month old weighing about 0.3KG were randomly divided into four groups.They were given 10 times Taohong Siwu Decoction,5 times Taohong Siwu Decoction,2.5 times Taohong Siwu Decoction and normal saline respectively by gavage for 7 days.One hour after the last gavage,the rats were anesthetized and the blood was taken from abdominal aorta and centrifuged for 10 minutes In the first part of the experiment,BMSCs were randomly divided into four groups,which were intervened by high,medium and low concentrations of Taohong Siwu Decoction medicated serum and blank group medicated serum respectively to detect the proliferation activity,osteogenic differentiation ability and osteogenic key protein,After obtaining the best intervention concentration of Taohong Siwu Decoction medicated serum,the second part of the experiment was carried out.BMSCs were divided into Taohong Siwu Decoction medicated serum + verteporfin group,blank group serum group and Taohong Siwu Decoction medicated serum group,and the data were obtained by detection again.4.CCK-8 kit was used to determine the proliferation activity of BMSCs at 0,1,3 and 7 days of the two parts of the experiment,and BM was determined after 7 days of intervention Alkaline phosphatase(ALP),an early indicator of osteoblast differentiation,was used to observe the degree of osteogenic differentiation;5.After 21 days of treatment,alizarin red staining was used to observe the mineralization indexes of BMSCs,such as calcium accumulation,and WB was used to detect the osteogenic related factors;6.After obtaining the optimal serum concentration of Taohong Siwu Decoction,WB was used to detect the hippo Yap signal pathway between blank group and optimal concentration of Taohong Siwu Decoction The expression of Yap/TAZ was detected;Results:1.CCK-8 experiment confirmed that:(1)the proliferation activity of BMSCs in Taohongsiwu Decoction high concentration serum group was higher than that in blank serum group,medium and low concentration serum groups(P<0.05),(2)the proliferation ability of BMSCs in Taohongsiwu Decoction group was significantly higher than that in Taohongsiwu Decoction + verteporfin group(P<0.05).2.ALP test and alizarin red staining test confirmed that:(1)the osteogenic differentiation ability of BMSCs in Taohong Siwu Decoction high concentration serum group was higher than that in blank serum group,medium and low concentration serum groups(P<0.05)(2)the osteogenic differentiation ability of BMSCs in Taohong Siwu Decoction group was significantly higher than that in Taohong Siwu Decoction +verteporfin group(P<0.05).3.WB test confirmed that:(1)the protein expression of ALP,OCN,Runx2 and osterix in Taohong Siwu Decoction high,medium and low drug containing serum group was higher than that in blank serum group(P<0.05);(2)Taohong Siwu Decoction high,medium and low drug containing serum group The protein expression of ALP,OCN,Runx2 and osterix in high concentration serum group was higher than that in medium concentration and low concentration serum groups(P<0.05);(3)the protein expression of Yap in Taohong Siwu Decoction group was lower than that in control group,In blank serum group,the expression of TAZ protein in Taohong Siwu Decoction containing serum group was higher than that in blank serum group(P<0.05);(4)the expression of ALP,OCN,Runx2,osterix protein in Taohong Siwu Decoction containing serum group was significantly higher than that in Taohong Siwu Decoction + verteporfin group(P<0.05).Conclusion:1.The proliferation and osteogenesis differentiation of BMSCs were positively correlated with the intervention concentration of Taohong Siwu Decoction.2.Hippo signaling pathway participates in the process of Taohong Siwu Decoction containing serum promoting BMSCs proliferation and osteogenic differentiation,and plays a key role in it.The internal mechanism may be that Taohong Siwu Decoction stimulates Yap protein and TAZ protein,the key node of Hippo signaling pathway,promotes BMSCs proliferation and induces osteogenic differentiation gene transcription.