Hills, Claire E., Kerr, Michael I., 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 & Biochemistry, Volume 32 (Number 5). pp. 1200-1212. doi:10.1159/000354519 ISSN 1015-8987.

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Abstract

Background/Aims: In the current study we examined if the adipocytokine, visfatin, alters connexinmediated
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.

Item Type:	Journal Article
Subjects:	Q Science > QH Natural history > QH301 Biology
Divisions:	Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- )
Library of Congress Subject Headings (LCSH):	Cell interaction, Diabetic nephropathies, Diabetes -- Complications , Epithelial cells
Journal or Publication Title:	Cellular Physiology & Biochemistry
Publisher:	S. Karger AG
ISSN:	1015-8987
Official Date:	2013
Dates:	Date Event 2013 Published
Volume:	Volume 32
Number:	Number 5
Page Range:	pp. 1200-1212
DOI:	10.1159/000354519
Status:	Peer Reviewed
Publication Status:	Published
Access rights to Published version:	Open Access (Creative Commons)
Date of first compliant deposit:	25 December 2015
Date of first compliant Open Access:	25 December 2015
Funder:	Diabetes UK, Warwick Private Hospitals Charitable Trust
Grant number:	BDA: 11/0004215 (DUK)
Related URLs:	Publisher

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