PCSK9 promotes osteogenic differentiation via ERK pathway and has no effect on osteoclast differentiation

Abstract

Background: Previous studies have demonstrated that PCSK9 was constitutively expressed in osteoclast precursors and played an important role in bone metabolism in periodontitis disease. But the function of PCSK9 in the osteogenesis of hBMSCs and specific mechanism still remains unknown. In this study, we aim to explore the function and mechanism of PCSK9 on regulating osteogenic differentiation and osteoclastic differentiation.Methods: The osteogenic effects of endogenous and exogenous PCSK9 on hBMSCs were performed, the effect on osteoclastic differentiation of mBMM too. A CCK-8 was used to assess the effect of PCSK9 on the proliferation of hBMSCs. qRT-PCR and Western blotting analysis were conducted to determine the expression of target genes and proteins. ALP, ARS staining were used to evaluate osteoblasts, the trap staining was used to evaluate osteoclasts. In vivo experiment, we constructed a femoral bone defect model, and injected Adeno associated virus at defect site. Radiographic analysis and histological evaluation were conducted to evaluate the union effect of PCSK9 at bone defect site. Student’s t test was used between two groups, one-way ANOVA or Bonferroni’s post-hoc test was used more than two groups, depending on the distribution of the tested population.Result: In vitro experiment, low dose exogenous and overexpress endogenous PCSK9 could enhance hBMSCs osteogenic differentiation via ERK signaling pathway, and knockdown PCSK9 would reduce this influence. The promoting effect could be weaken by ERK inhibitor PD98059 with an appropriate concentration. All of which was confirmed by q-PCR, Western blotting and immunofluorescence outcome. Besides, the ALP staining, ARS staining results prove that it could raise ALP activity and mineral deposits formation. But low dose PCSK9 has no obvious effect on osteoclastic differentiation. In vivo experiment, the radiographic, histological result prove overexpress PCSK9 could accelerate bone defect union. Conclusion: In summarize, PCSK9 could promote osteogenic differentiation via ERK pathway and has no effect on osteoclast differentiation. The vivo and vitro experiments outcomes indicate that it may be a novel target for treating bone defect union.