Balancing calcium signals through TRPC5 and TRPC6 in podocytes

Harvard University, Cambridge, Massachusetts, United States
Journal of the American Society of Nephrology (Impact Factor: 9.34). 11/2011; 22(11):1969-80. DOI: 10.1681/ASN.2011040370
Source: PubMed


Calcium (Ca(2+)) ions are important mediators of cellular homeostasis owing to their ability to elicit a dynamic, transient, and tightly regulated range of biochemical responses. More than a decade ago, a nonselective, Ca(2+)-permeable, cationic conductance was identified in podocytes downstream of angiotensin II (Ang II) signaling, but its molecular structure remained elusive. Six years ago, transient receptor potential canonical 6 (TRPC6) mutations were found in families with hereditary FSGS, and TRPC5 and TRPC6 channels are now known as the Ca(2+) influx pathways for this previously described, nonselective, cationic current in podocytes. Ang II activation engages this Ca(2+) influx to modulate the actin cytoskeleton in podocytes. These discoveries dovetail with previously described regulation of actin dynamics by the Ca(2+)-activated phosphatase, calcineurin, and the emergence of Rho GTPases as critical regulators of podocyte function in health and disease. Understanding the interconnected signaling regulated by Ca(2+) currents offers potential new therapeutic targets and highlights the notion that synergistic therapies targeting multiple levels of biochemistry may be useful in treating proteinuric kidney disease.

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    • "Recent studies further revealed a role of TRPC5 in podocytes, demonstrating that the pathogenic remodelling in proteinuria mouse models can be attributed to the activation of TRPC5 channels. Accordingly, TRPC5-deficient mice were less sensitive to LPS-induced albuminuria (Greka and Mundel, 2011; Schaldecker et al., 2013), making a pharmacological inhibition of TRPC5 a promising target to protect from the destruction of the glomerular filter barrier in kidney injury. "
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    ABSTRACT: TRPC5 is a nonselective, Ca(2+) permeable cation channel which belongs to the large family of transient receptor potential channels. It is predominantly found in the central nervous system with a high expression density in the hippocampus, the amygdala and the frontal cortex. Several studies confirm that TRPC5 channels are implicated in the regulation of neurite length and growth cone morphology. We identified clemizole as a novel inhibitor of TRPC5 channels. Clemizole efficiently blocks TRPC5 currents and Ca(2+) entry in the low micromolar range (IC50 = 1.0 - 1.3 μM), as determined by fluorometric [Ca(2+)]i measurements and patch clamp recordings. Clemizole blocks TRPC5 currents irrespectively of the mode of activation, e.g. stimulation of GPCR, hypoosmotic buffer conditions or by the direct activator riluzole. Electrophysiological whole cell recordings revealed that the block was mostly reversible. Moreover, clemizole was still effective in blocking TRPC5 single channels in excised inside-out membrane patches, hinting to a direct block of TRPC5 by clemizole. Based on fluorometric [Ca(2+)]i measurements, clemizole exhibits a 6-fold selectivity for TRPC5 over TRPC4β (IC50 = 6.4 μM), the closest structural relative of TRPC5 and an almost 10-fold selectivity over TRPC3 (IC50 = 9.1 μM) and TRPC6 (IC50 = 11.3 μM). TRPM3 and M8 as well as TRPV1, V2, V3 and V4 channels were only weakly affected by markedly higher clemizole concentrations. Clemizole was not only effective in blocking heterologously expressed TRPC5 homomers but also TRPC1:TRPC5 heteromers as well as native TRPC5-like currents in the U-87 glioblastoma cell line.
    Molecular pharmacology 08/2014; 86(5). DOI:10.1124/mol.114.093229 · 4.13 Impact Factor
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    • "A vast body of evidence supports the important role of calcium in kidney disease. Mutations in transient receptor potential canonical 6 (TRPC6) channels and polycystin-2, a prototypical member of a subfamily of the TRPC channel superfamily, have been reported to cause familial focal segmental glomerulosclerosis and autosomal dominant polycystic kidney disease, respectively [36–41]. "
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    ABSTRACT: Taiwan has very high incidence and prevalence of chronic kidney disease (CKD), which easily progresses to end-stage renal disease (ESRD). The association between inflammation and CKD has been explored in several studies. ORAI1 functions as a pore-forming subunit of the store-operated calcium channels which are involved in the regulation of immune system. Hence, we conducted a case-control study to determine whether the genetic polymorphisms of ORAI1 gene is a susceptibility factor to CKD and its clinical features in a Taiwanese population. Five hundred seventy-nine CKD patients from a hospital-based CKD care program were included in the study. Five tagging single nucleotide polymorphisms (tSNPs) of ORAI1 were selected from the genotyping data of the Han Chinese population from the HapMap project. Among these polymorphisms, rs12313273 was found to be significantly associated with elevated serum calcium levels, which has been linked to increased risk of death in CKD patients. To have a better management of serum calcium, we suggest that ORAI1 polymorphisms might be used as a potential biomarker for initiating non-calcium-based phosphate binder in CKD patients in the future.
    03/2014; 2014:290863. DOI:10.1155/2014/290863
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    • "Since in podocytes, intracellular calcium is an important second messenger involved in crucial cell signaling pathways [4], [23], we examined if the two cell lines increased their intracellular calcium concentration in response to flufenamic acid (FFA, a TRPC6 agonist) and angiotensin II (Ang II), using fluorescence imaging (Fig. 5F–J). The increase in intracellular calcium in hAKPC-P in response to FFA was significantly greater than in hIPod; hAKPC-P also exhibited intense contraction after FFA stimulation as shown in the Supplementary Video. "
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    ABSTRACT: Amniotic fluid is in continuity with multiple developing organ systems, including the kidney. Committed, but still stem-like cells from these organs may thus appear in amniotic fluid. We report having established for the first time a stem-like cell population derived from human amniotic fluid and possessing characteristics of podocyte precursors. Using a method of triple positive selection we obtained a population of cells (hAKPC-P) that can be propagated in vitro for many passages without immortalization or genetic manipulation. Under specific culture conditions, these cells can be differentiated to mature podocytes. In this work we compared these cells with conditionally immortalized podocytes, the current gold standard for in vitro studies. After in vitro differentiation, both cell lines have similar expression of the major podocyte proteins, such as nephrin and type IV collagen, that are characteristic of mature functional podocytes. In addition, differentiated hAKPC-P respond to angiotensin II and the podocyte toxin, puromycin aminonucleoside, in a way typical of podocytes. In contrast to immortalized cells, hAKPC-P have a more nearly normal cell cycle regulation and a pronounced developmental pattern of specific protein expression, suggesting their suitability for studies of podocyte development for the first time in vitro. These novel progenitor cells appear to have several distinct advantages for studies of podocyte cell biology and potentially for translational therapies.
    PLoS ONE 12/2013; 8(12):e81812. DOI:10.1371/journal.pone.0081812 · 3.23 Impact Factor
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