title: Melamine induces Ca2+-sensing receptor activation and elicits apoptosis in proximal tubular cells.
abstract: Melamine causes renal tubular cell injury through inflammation, fibrosis, and apoptosis. Although melamine affects the rise in intracellular Ca2+concentration ([Ca2+]i),reactive oxygen species (ROS) production, and proapoptotic pathway activation, the mechanism of upstream Ca2+signaling is unknown. Because melamine has some structural similarities with l-amino acids, which endogenously activate Ca2+-sensing receptors (CSR), we examined the effect of melamine on CSR-induced Ca2+signaling and apoptotic cell death. We show here that melamine activates CSR, causing a sustained Ca2+entry in the renal epithelial cell line, LLC-PK1. Moreover, such CSR stimulation resulted in a rise in [Ca2+]i, leading to enhanced ROS production. Furthermore, melamine-induced elevated [Ca2+]iand ROS production caused a dose-dependent increase in apoptotic (by DAPI staining, DNA laddering, and annexin V assay) and necrotic (propidium iodide staining) cell death. Upon examining the downstream mechanism, we found that transforming growth factor β1 (TGF-β1), which increases extracellular matrix genes and proapoptotic signaling, was also upregulated at lower doses of melamine, which could be due to an early event inducing apoptosis. Additionally, cells exposed to melamine displayed a rise in pERK activation and lactate dehydrogenase release resulting in cytotoxicity. These results offer a novel insight into the molecular mechanisms by which melamine exerts its effect on CSR, causing a sustained elevation of [Ca2+]i, leading to ROS generation, fibronectin production, proapoptotic pathway activation, and renal cell damage. Together, these results thus suggest that melamine-induced apoptosis and/or necrosis may subsequently result in acute kidney injury and promote kidney stone formation.
authors.affiliation: Calcium Signaling Laboratory, Research Service, Veterans Affairs Medical Center, Washington, District of Columbia.|Department of Pharmacology and Physiology, School of Medicine, George Washington University, Washington, District of Columbia; and.
authors.forename: Allen J
authors.initials: AJ
authors.lastname: Yiu
authors.affiliation: Calcium Signaling Laboratory, Research Service, Veterans Affairs Medical Center, Washington, District of Columbia.
authors.forename: Cliff-Lawrence
authors.initials: CL
authors.lastname: Ibeh
authors.affiliation: Calcium Signaling Laboratory, Research Service, Veterans Affairs Medical Center, Washington, District of Columbia.
authors.forename: Sanjit K
authors.initials: SK
authors.lastname: Roy
authors.affiliation: Calcium Signaling Laboratory, Research Service, Veterans Affairs Medical Center, Washington, District of Columbia; bidhan.bandyopadhyay@va.gov.|Department of Pharmacology and Physiology, School of Medicine, George Washington University, Washington, District of Columbia; and.|Department of Pharmacology and Physiology, Georgetown University, Washington, District of Columbia.
authors.forename: Bidhan C
authors.initials: BC
authors.lastname: Bandyopadhyay
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D053938:DNA Fragmentation; D004785:Environmental Pollutants / Q000633:toxicity*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000819:agonists / Q000378:metabolism; D044169:Receptors, Calcium-Sensing / Q000819:agonists* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence; D013552:Swine; D059008:Time-Lapse Imaging; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism; D014227:Triazines / Q000633:toxicity*
chemical_list: D004785:Environmental Pollutants; D005353:Fibronectins; D017382:Reactive Oxygen Species; D044169:Receptors, Calcium-Sensing; D053773:Transforming Growth Factor beta1; D014227:Triazines; D007770:L-Lactate Dehydrogenase; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C011907:melamine; D002118:Calcium
keywords: Ca2+-sensing receptor; acute kidney injury; apoptosis; cell cytotoxicity; intracellular Ca2+; kidney stone; kidney tubular cells; melamine