Visfatin reduces gap junction mediated cell-to-cell communication in proximal tubule-derived epithelial cells

Hills, Claire E. and Kerr, Michael I. and Wall, Mark J. and Squires, Paul E. (2013) Visfatin reduces gap junction mediated cell-to-cell communication in proximal tubule-derived epithelial cells. Cellular Physiology and Biochemistry, 32 (5). pp. 1200-1212. ISSN 1015-8987

Full content URL: http://dx.doi.org/10.1159/000354519

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Abstract

Background/Aims: In the current study we examined if the adipocytokine, visfatin, alters connexin-mediated intercellular communication in proximal tubule-derived epithelial cells. Methods: The effects of visfatin (10-200ng/mL) on cell viability and cytotoxicity in HK2-cells were assessed by MTT, crystal violet and lactate dehydrogenase assays. Western blot analysis was used to confirm expression of Cx26, Cx40 and Cx43. The effect of visfatin (10-200ng/mL) on TGF-β1 secretion was confirmed by ELISA, and the effects of both TGF-β1 (2-10ng/mL) and visfatin (10-200ng/mL) on connexin expression were assessed by western blot. Functional intercellular communication was determined using transfer of Lucifer Yellow and paired-whole cell patch clamp electrophysiology. Results: In low glucose (5mM), visfatin (10-200ng/mL) did not affect membrane integrity, cytotoxicity or cell viability at 48hrs, but did evoke a concentration-dependent reduction in Cx26 and Cx43 expression. The expression of Cx40 was unaffected. At 48hrs, visfatin (10-200ng/mL) increased the secretion of TGF-β1 and the visfatin-evoked changes in connexin expression were mimicked by exogenous application of the pro-fibrotic cytokine (2-10ng/ml). Visfatin reduced dye transfer between coupled cells and decreased functional conductance, with levels falling by 63% as compared to control. Although input resistance was increased following visfatin treatment by 166%, the change was not significant as compared to control. The effects of visfatin on Cx-expression and cell-coupling were blocked in the presence of a TGF-β1 specific neutralizing antibody. Conclusions: The adipocytokine visfatin selectively evoked a non-toxic reduction in connexin expression in HK2-cells. The loss in gap-junction associated proteins was mirrored by a loss in functional conductance between coupled cells. Visfatin increased TGF-β secretion and the pattern of change for connexins expression was mimicked by exogenous application of TGF-β1. The effect of visfatin on Cx-expression and dye transfer were negated in the presence of a TGF-β1 neutralising antibody. These data suggest that visfatin reduces connexin-mediated intercellular communication in proximal tubule-derived epithelial cells via a TGF-β dependent pathway.

© 2013 S. Karger AG, Basel

Keywords:Visfatin, Gap junctions, Proximal tubule, Transforming growth factor beta, Diabetic nephropathy
Subjects:C Biological Sciences > C720 Biological Chemistry
Divisions:College of Science > School of Life Sciences
ID Code:14131
Deposited On:29 May 2014 08:57

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