LPS Facilitates Osteogenic Differentiation of Porcine Aortic Valve Interstitial Cells by Upregulating BMP4
ZHU Mengying, SHI Qiong, AN Liqin, LIU Yan, WENG Yaguang*
This study aimed to investigate the effect and mechanism of LPS on osteogenic differentiation of porcine aortic VICs (valve interstitial cells) by upregulating BMP4 (bone morphogenetic protein 4), which could provide a theoretical basis for the intervention and treatment of CAVD (calcific aortic valve disease). The expression of BMP4 and Runx2 in the non-CAVD group and CAVD group were detected by immunohistochemistry and Western blot. The porcine VICs were isolated by collagenase (type I) after the digestion of the valve, and the phenotype was identified by immunofluorescence staining. VICs were treated with LPS and recombinant human BMP4 adenovirus. ALP (alkaline phosphatase) staining, alizarin red S staining, qRT-PCR, and Western blot were used to detect the early and late osteogenic differentiation abilities of cells. The protein levels of p-Smad1/5/8 and p-ERK1/2 were measured by Western blot. The results showed that the expression of BMP4 and Runx2 in the CAVD group was significantly higher than that in the non-CAVD group. The porcine primary VICs were successfully isolated. The staining of α-SMA and Vimentin were positive, while the staining of CD31 was negative. LPS significantly increased the ALP activity and the deposition of calcium salts in VICs. The mRNA and protein levels of calcification markers and BMP4 were increased. Besides, BMP4 also increased the ALP activity, the deposition of calcium salts, and the mRNA and protein levels of calcification makers. Meanwhile, the protein levels of p-Smad1/5/8 and p-ERK1/2 were increased by BMP4. In conclusion, LPS promotes osteoblastic differentiation of aortic valve cells by upregulating BMP4. The Smad1/5/8 and ERK1/2 signaling pathways may play important roles in these processes.
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