Serum Containing Compound Kidney-invigorating Granule Regulates the Osteogenic and Adipogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells Through the β-Catenin/Tribbles Pseudokinase 3 Pathway
Objective To investigate the mechanism of action of serum containing Compound Kidney-invigorating Granule (CKG)in regulating the osteogenic and adipogenic differentiation of human bone marrow mesenchymal stem cells (BMSCs ) through the β-catenin/tribbles pseudokinase 3(TRIB3)pathway.Methods Primary BMSCs were purified.BMSCs were trans-fected with β-catenin-siRNA,and after osteogenic induction,alkaline phosphatase (ALP)staining and alizarin red staining were used to observe the osteogenic differentiation of BMSCs;qPCR and Western blot were used to measure the mRNA and protein expression levels of β-catenin,TRIB3,osteopontin (OPN),Runt-related transcription factor 2 (RUNX2 ),and ALP;Western blot was used to observe the effect of CKG on the protein expression levels of β-catenin and TRIB3 in BMSCs.After BMSCs were transfected with β-catenin-siRNA and cultured with CKG-containing serum,ALP staining,alizarin red staining,and oil red O staining were used to observe the osteogenic and adipogenic differentiation of BMSCs,and qPCR and Western blot were used to measure the mRNA and protein expression levels of β-catenin,TRIB3,OPN,RUNX2,peroxisome proliferator-activated receptor gamma (PPARγ),and adiponectin.Results After BMSCs were transfected with β-catenin-siRNA,there were signifi-cant reductions in ALP activity (P<0.05),mineralized nodules,and the mRNA and protein expression levels of TRIB3,OPN,RUNX2,and ALP (P<0.05). CKG promoted the expression of β-catenin and TRIB3 in BMSCs (P<0.05). After BMSCs were transfected with β-catenin-siRNA and cultured with CKG-containing serum,the 5％ CKG-containing serum+siRNA group had significantly higher numbers of calcium-cobalt staining nodes and red calcified nodules than the 0％ CKG-containing serum group,as well as significantly lower numbers than the 5％ CKG-containing serum group;the 5％ CKG-containing serum+siRNA group had a significantly higher number of lipid droplets in cells than the 5％ CKG-containing serum group and a significantly lower number than the 0％ CKG-containing serum group;the 5％ CKG-containing serum+siRNA group had significantly higher mRNA and protein expression levels of TRIB3,OPN,and RUNX2 and a significantly higher King unit val-ue for ALP activity compared with the 0％ CKG-containing serum group,as well as significantly lower mRNA and protein ex-pression levels and King unit value than the 5％ MS group and the 5％ CKG-containing serum+negative control group (P<0.05);the 5％ CKG-containing serum+siRNA group had significantly lower mRNA and protein expression levels of PPARγand adiponectin than the 0％ CKG-containing serum group (P<0.05)and significantly higher levels than the 5％ CKG-contai-ning serum group and the 5％ CKG-containing serum+negative control group (P<0.05). Conclusion Endogenous β-catenin knockdown inhibits the osteogenic differentiation of BMSCs and reduces the expression of TRIB3,and CKG-containing serum promotes the expression of β-catenin and TRIB3 in BMSCs.CKG-containing serum can regulate the osteogenic and adipogenic differentiation of BMSCs,which may be associated with the β-catenin/TRIB3 pathway.
万方数据
维普
