Calcium signaling mechanism in proximal tubule underlying calcium nephrolithiasis

The prevalence of nephrolithiasis affects more than 10% of Americans in their lifetime. Veterans’ health records show that nephrolithiasis is one of the leading causes of hospitalizations and is associated with increased morbidity. Calcium nephrolithiasis (CaNL), which has increased in the past few decades, comprises ~80% of patient stones. In CaNL patients stones usually consist of calcium oxalate (CaOx) with variable amounts of calcium phosphate (CaP) as a core component, signifying CaP as the focal point for CaP+CaOx (mixed) stone formation. Moreover, a recent surge in CaP content from CaNL patients suggests the further need for research on CaP stone formation. While the exact pathogenesis of initial CaP crystallization is unknown, it is proposed that the nucleation of CaP crystals starts with the supersaturation of Ca2+ and a relatively higher pH at the thin descending limb of the loop of Henle (LOH). Importantly, pH >6.5 increases CaP crystallization and pH <6.2 decreases its solubility. Thus, handling of PT luminal [Ca2+] and pH both can be critical to the formation of CaP at the LOH. The proximal tubule (PT), immediately upstream of LOH, is the major site (~70%) for Ca2+ reabsorption, where Ca2+ transport is described as paracellular. We found a transcellular Ca2+ transport machinery operated by transient receptor potential canonical 3 (TRPC3) Ca2+ channel at the PT apical brush border area. The mice lacking TRPC3 displayed disrupted Ca2+ transport by PT cells and presented with intraluminal CaP crystals at the LOH. Thus, our central hypothesis is that the interruption in PT transcellular Ca2+ transport, via TRPC3, and the rise in pH in PT luminal fluid, are critical to CaP crystal formation at the LOH, contributing to mixed stone formation. In Aim 1 we will investigate the renal Ca2+ clearance using TRPC3 global KO and PT-specific TRPC3 KO mice models to define the significance of PT transcellular versus paracellular Ca2+ reabsorption as well as to sort out the contribution of the TRPC3-mediated Ca2+ transport in the distal nephron. Importantly, the two main factors that favor CaP crystallization are alkaline urine and hypercalciuria. In fact, alkalinization of PT luminal fluid due to acidosis in women can facilitate CaP crystallization at the LOH; conversely, men are predisposed to idiopathic hypercalciuria, which is common in CaNL with mixed stones. Thus, in Aim 2, we will target the sex-dependent phenotype of CaP and mixed stone formation in our animal models to determine the susceptibility of CaP and mixed stone formation in response to high calcium diet/urine alkalization. Finally, in Aim 3 we will determine the impact of microenvironmental stress exerted by vascular calcification/inflammation on the PT cell function and the detrimental pathway activation to induce CaP and mixed stone formation in vitro and in vivo. Proposed research will find i) novel molecular targets/mechanism of Ca2+ regulation in the PT and its role in CaNL; ii) the PT-specific mechanism on initial intraluminal CaP crystallization site; iii) identify a new molecular mechanism handling higher alkaline conditions pertaining to the higher incidence CaP stone formation; iv) the impact of microenvironmental stress exerted by vascular/immune cells. These Aims will determine the basis for preventing the formation of CaP and mixed stones towards developing the therapy for pathophysiology-based intervention. Project Number: 1I01BX006849-01 | Fiscal Year: 2025 | NIH Institute/Center: Veterans Affairs (VA) | Principal Investigator: Bidhan Bandyopadhyay | Institution: U.S. DEPT/VETS AFFAIRS MEDICAL CENTER, WASHINGTON, DC | Activity Code: I01 | Study Section: Special Emphasis Panel[ZRD1 NEPH-N (01)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11050052