Nephrolithiasis is increasing in developed and developing countries at an alarming rate. With the global spike in kidney stone diseases, it is crucial to determine what risk factors are influencing the current global landscape for kidney stones. Our aims for this review are: to identity and analyze the four categories of risk factors in contributing to the global scale of stone formation: lifestyle, genetics, diet, and environment; and discuss preventative measures for kidney stone formation. We also performed data search through the published scientific literature, i.e., PubMed® and found that there is a significant link between lifestyle and obesity with cases of calcium stones. Food and Agriculture Organization of the United Nations and World Health Organization factor indicators for dietary intake and obesity, along with climate data were used to create the projected total risk world map model for nephrolithiasis risk. Complete global analyses of nephrolithiasis deplete of generalizations is nearly insurmountable due to limited sources of medical and demographic information, but we hope this review can provide further elucidation into confounding risk factors and preventative measures for global nephrolithiasis analysis.

Calcium crystal internalization into proximal tubular (PT) cells results in acute kidney injury, nephrocalcinosis, chronic kidney disease (CKD), and kidney-stone formation. Ca 2+ supersaturation in PT luminal fluid induces calcium crystal formation, leading to aberrant crystal internalization into PT cells. While such crystal internalization produces reactive oxygen species (ROS), cell membrane damage, and apoptosis; the upstream signaling events involving dysregulation of intracellular Ca 2+ homeostasis and ER stress, remain largely unknown. We have recently described a transepithelial Ca 2+ transport pathway regulated by receptor-operated Ca 2+ entry (ROCE) in PT cells. Therefore, we examined the pathophysiological consequence of internalization of stone-forming calcium crystals such as calcium phosphate (CaP), calcium oxalate (CaOx), and CaP + CaOx (mixed) crystals on the regulation of intracellular Ca 2+ signaling by measuring dynamic changes in Ca 2+ transients in HK2, human PT cells, using pharmacological and siRNA inhibitors. The subsequent effect on ER stress was measured by changes in ER morphology, ER stress-related gene expression, endogenous ROS production, apoptosis, and necrosis. Interestingly, our data show that crystal internalization induced G-protein-coupled receptor-mediated sustained rise in intracellular Ca 2+ concentration ([Ca 2+ ] i ) via store-operated Ca 2+ entry (SOCE); suggesting that the mode of Ca 2+ entry switches from ROCE to SOCE following crystal internalization. We found that SOCE components—stromal interacting molecules 1 and 2 (STIM1, STIM2) and ORAI3 (SOCE) channel were upregulated in these crystal-internalized cells, which induced ER stress, ROS production, and cell death. Finally, silencing those SOCE genes protected crystal-internalized cells from prolonged [Ca 2+ ] i rise and ER stress. Our data provide insight into the molecular mechanism of crystal-induced Ca 2+ dysregulation, ER stress, and PT cell death and thus could have a translational role in treating crystal nephropathies including kidney stones. Taken together, modulation of Ca 2+ signaling can be used as a tool to reverse the pathological consequence of crystal-induced conditions including cardiovascular calcification.

Melamine, which induces proximal tubular (PT) cell damage has a greater nephrotoxic effect when combined with cyanuric and uric acids; however, it is unknown whether such effect can stimulate calcium phosphate (CaP)/calcium oxalate (CaOx) stone formation. Here, we show that melamine acts as an inducer of CaP, CaOx and CaP + CaOx (mixed) crystal formations in a time and concentration-dependent manner by stabilizing those crystals and further co-aggregating with melamine. To explore the physiological relevance of such melamine-augmented calcium crystal formation, we used 2-dimensional (2D) and 3D microfluidic (MF) device, embedded with PT cells, which also resembled the effect of melamine-stimulated CaP, CaOx and mixed crystal formation. Significantly, addition of preformed CaP and/or CaOx crystal in the presence of melamine, further potentiated those crystal formations in 3D MFs, which helped the growth and aggregation of mixed crystals. Our data show that the mechanism of such predisposition of stone formation could be largely due to co-crystallization between melamine and CaP/CaOx and pronounced effect on induction of stone-forming pathway activation in 3D MF. Taken together, melamine-induced CaP and/or CaOx crystal formation ex-vivo will help us in understanding the larger role of melamine as an environmental toxicant in producing the pathology in similar cellular microenvironments.

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Proximal tubular (PT) acidosis, which alkalinizes the urinary filtrate, together with Ca2+ supersaturation in PT can induce luminal calcium phosphate (CaP) crystal formation. While such CaP crystals are known to act as a nidus for CaP/calcium oxalate (CaOx) mixed stone formation, the regulation of PT luminal Ca2+ concentration ([Ca2+]) under elevated pH and/or high [Ca2+] conditions are unknown. Since we found that transient receptor potential canonical 3 (TRPC3) knockout (KO; -/-) mice could produce mild hypercalciuria with CaP urine crystals, we alkalinized the tubular pH in TRPC3-/- mice by oral acetazolamide (0.08%) to develop mixed urinary crystals akin to clinical signs of calcium nephrolithiasis (CaNL). Our ratiometric (λ340/380) intracellular [Ca2+] measurements reveal that such alkalization not only upsurges Ca2+ influx into PT cells, but the mode of Ca2+ entry switches from receptor-operated to store-operated pathway. Electrophysiological experiments show enhanced bicarbonate related current activity in treated PT cells which may determine the stone-forming phenotypes (CaP or CaP/CaOx). Moreover, such alkalization promotes reactive oxygen species generation, and upregulation of calcification, inflammation, fibrosis, and apoptosis in PT cells, which were exacerbated in absence of TRPC3. Altogether, the pH-induced alteration of the Ca2+ signaling signature in PT cells from TRPC3 ablated mice exacerbated the pathophysiology of mixed urinary stone formation, which may aid in uncovering the downstream mechanism of CaNL.