Proximal tubule (PT) transports most of the renal Ca²⁺, which was usually described as paracellular (passive). We found a regulated Ca²⁺ entry pathway in PT cells via the apical transient receptor potential canonical 3 (TRPC3) channel, which initiates transcellular Ca²⁺ transport. Although TRPC3 knockout (−/−) mice were mildly hypercalciuric and displayed luminal calcium phosphate (CaP) crystals at Loop of Henle (LOH), no CaP + calcium oxalate (CaOx) mixed urine crystals were spotted, which are mostly found in calcium nephrolithiasis (CaNL). Thus, we used oral calcium gluconate (CaG; 2%) to raise the PT luminal [Ca²⁺]_o further in TRPC3^−/− mice for developing such mixed stones to understand the mechanistic role of PT-Ca²⁺ signaling in CaNL. Expectedly, CaG-treated mice urine samples presented with numerous mixed crystals with remains of PT cells, which were pronounced in TRPC3^−/− mice, indicating PT cell damage. Notably, PT cells from CaG-treated groups switched their mode of Ca²⁺ entry from receptor-operated to store-operated pathway with a sustained rise in intracellular [Ca²⁺] ([Ca²⁺]_i), indicating the stagnation in PT Ca²⁺ transport. Moreover, those PT cells from CaG-treated groups demonstrated an upregulation of calcification, inflammation, fibrotic, oxidative stress, and apoptotic genes; effects of which were more robust in TRPC3 ablated condition. Furthermore, kidneys from CaG-treated groups exhibited fibrosis, tubular injury and calcifications with significant reactive oxygen species generation in the urine, thus, indicating in vivo CaNL. Taken together, excess PT luminal Ca²⁺ due to escalation of hypercalciuria in TRPC3 ablated mice induced surplus CaP crystal formation and caused stagnation of PT [Ca²⁺]_i, invoking PT cell injury, hence mixed stone formation.

近端小管（PT）运输大部分肾Ca ²⁺ ，通常被描述为细胞旁（被动）。我们发现 PT 细胞中通过顶端瞬时受体电位规范 3 (TRPC3) 通道调节 Ca ²⁺进入途径，该通道启动跨细胞 Ca ²⁺转运。尽管TRPC3敲除 (−/−) 小鼠存在轻度高尿钙，并且在亨利袢 (LOH) 处显示管腔磷酸钙 (CaP) 晶体，但未发现 CaP + 草酸钙 (CaOx) 混合尿液晶体，这主要见于钙肾结石（CaNL）。因此，我们使用口服葡萄糖酸钙（CaG；2%）来进一步提高TRPC3 ^−/−小鼠中的 PT 管腔 [Ca ²⁺ ] _o ，以形成此类混合结石，以了解 PT-Ca ²⁺信号传导在 CaNL 中的机制作用。不出所料，CaG 处理的小鼠尿液样本中出现大量含有 PT 细胞残留的混合晶体，这在TRPC3 ^−/−小鼠中很明显，表明 PT 细胞受损。值得注意的是，CaG 处理组的 PT 细胞将其 Ca ²⁺进入模式从受体操纵途径转变为钙池操纵途径，细胞内 [Ca ²⁺ ] ([Ca ²⁺ ]) 持续上升，表明 PT 停滞Ca ²⁺运输。此外，CaG 处理组的 PT 细胞表现出钙化、炎症、纤维化、氧化应激和凋亡基因的上调；在 TRPC3 消融条件下其效果更为强劲。 此外，CaG 治疗组的肾脏表现出纤维化、肾小管损伤和钙化，尿液中产生显着的活性氧，因此表明存在体内CaNL。总而言之，TRPC3 消融小鼠中高钙尿症升级导致过量 PT 管腔 Ca ²⁺诱导过量 CaP 晶体形成，并导致 PT [Ca ²⁺ ] 停滞，引起 PT 细胞损伤，从而形成混合结石。