Klotho suppresses high phosphate-induced osteogenic responses in human aortic valve interstitial cells through inhibition of Sox9.

Elevated level of blood phosphate (Pi) associated with chronic kidney disease (CKD) is a risk factor of aortic valve calcification. Aortic valve interstitial cells (AVICs) display osteogenic responses to high Pi although the underlying mechanism is incompletely understood. Sox9 is a pro-chondrogenic factor and may play a role in ectopic tissue calcification. Circulating and kidney levels of Klotho are reduced in patients with CKD. We hypothesized that Sox9 mediates high Pi-induced osteogenic responses in human AVICs and that Klotho inhibits the responses. Treatment of human AVICs with high Pi increased protein levels of Runt-related transcription factor 2 (Runx2) and alkaline phosphatase (ALP), and a prolonged exposure to high Pi caused calcium deposition. High Pi induced Sox9 upregulation through PKD and Akt activation. Knockdown of Sox9 essentially abolished the effect of high Pi on the osteogenic responses. Lower Klotho levels were observed in calcified aortic valve tissues. Interestingly, high Pi decreased Klotho levels in AVICs from normal valves, and treatment with recombinant Klotho markedly reduced the effect of high Pi on the levels of Sox9, Runx2, and ALP and suppressed calcium deposition. We conclude that high Pi induces human AVIC osteogenic responses through Sox9. Human AVICs express Klotho, and its levels in AVICs are modulated by high Pi and valvular calcification. Importantly, Klotho suppresses the pro-osteogenic effect of high Pi on human AVICs. These novel findings indicate that modulation of Klotho may have therapeutic potential for mitigation of valvular calcification associated with CKD. KEY MESSAGES: CAVD associated with chronic kidney disease is a significant clinical problem. High phosphate upregulates Sox9 through AKT and PKD in human AVICs. Calcified human aortic valves have lower levels of Klotho. Klotho suppresses Sox9 upregulation and intranuclear translocation. Klotho inhibits high phosphate-induced osteogenic activity in human AVICs.