AIM To explore the role of 6-bisphosphatase 3 (PFKFB3) in phosphorus-induced calcification of primary vascular smooth muscle cells (VSMC) in rats and its underlying mechanism.
METHODS VSMC were randomly divided into 5 groups: control (Con) group, hyperphosphatemia induction (β-GP) group, β-GP+3-PO (a selective PFKFB3 inhibitor) group, Con+Lactate group, and Con+Lactate+3-PO group. Before and after the experiment, the contents of intracellular calcium, alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and lactate were detected. Alizarin red staining was used to detect the calcification degree of VSMC and CCK8 kit was used to detect cell viability. The expression levels of RUNX2, BMP-2, SM22a (as the key molecules of calcification-related proteins) and PFKFB3, HKⅡ, GLUT1, GLUT4 (as the key enzyme of glycolysis) were detected by Western bolt.
RESULTS Compared with the control group, the β-GP group showed a lower cell viability (P<0.01) and SM22a expression (P<0.01) and significantly higher RUNX2 and BMP-2 expressions (P<0.01). However, treatment with 3-PO, the above effects were abrogated (P<0.05). Further studies confirmed that glycolysis and PFKFB3 expressions were significantly increased in the β-GP group compared with those in the control group (P<0.01), while significantly lower expressions were found in the β-GP+3-PO group (P<0.01). The HKⅡ, GLUT1 and GLUT4 expressions were unaltered. Lactate treatment significantly increased VSMC calcification and the phenotypic transforming protein expression (P<0.01), while this phenomenon could be reversed by 3-PO administration. (P<0.05).
CONCLUSION Inhibition of PFKFB3 by 3-PO decreases VSMC calcification and increases cell viability by reducing glycolysis and PFKFB3 expression, which provides a potential drug target and strategy for clinical treatment of vascular calcification.
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