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High glucose-induced Smad3 linker phosphorylation and CCN2 expression are inhibited by dapagliflozin in a diabetic tubule epithelial cell model.

Pan, X; Phanish, MK; Baines, DL; Dockrell, MEC (2021) High glucose-induced Smad3 linker phosphorylation and CCN2 expression are inhibited by dapagliflozin in a diabetic tubule epithelial cell model. Biosci Rep, 41 (6). ISSN 1573-4935 https://doi.org/10.1042/BSR20203947
SGUL Authors: Baines, Deborah

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Abstract

BACKGROUND: In the kidney glucose is freely filtered by the glomerulus and, mainly, reabsorbed by sodium glucose cotransporter 2 (SGLT2) expressed in the early proximal tubule. Human proximal tubule epithelial cells (PTECs) undergo pathological and fibrotic changes seen in diabetic kidney disease (DKD) in response to elevated glucose. We developed a specific in vitro model of DKD using primary human PTECs with exposure to high D-glucose and TGF-β1 and propose a role for SGLT2 inhibition in regulating fibrosis. METHODS: Western blotting was performed to detect cellular and secreted proteins as well as phosphorylated intracellular signalling proteins. qPCR was used to detect CCN2 RNA. Gamma glutamyl transferase (GT) activity staining was performed to confirm PTEC phenotype. SGLT2 and ERK inhibition on high D-glucose, 25 mM, and TGF-β1, 0.75 ng/ml, treated cells was explored using dapagliflozin and U0126, respectively. RESULTS: Only the combination of high D-glucose and TGF-β1 treatment significantly up-regulated CCN2 RNA and protein expression. This increase was significantly ameliorated by dapagliflozin. High D-glucose treatment raised phospho ERK which was also inhibited by dapagliflozin. TGF-β1 increased cellular phospho SSXS Smad3 serine 423 and 425, with and without high D-glucose. Glucose alone had no effect. Smad3 serine 204 phosphorylation was significantly raised by a combination of high D-glucose+TGF-β1; this rise was significantly reduced by both SGLT2 and MEK inhibition. CONCLUSIONS: We show that high D-glucose and TGF-β1 are both required for CCN2 expression. This treatment also caused Smad3 linker region phosphorylation. Both outcomes were inhibited by dapagliflozin. We have identified a novel SGLT2 -ERK mediated promotion of TGF-β1/Smad3 signalling inducing a pro-fibrotic growth factor secretion. Our data evince support for substantial renoprotective benefits of SGLT2 inhibition in the diabetic kidney.

Item Type: Article
Additional Information: © 2021 The Author(s). This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY).
Keywords: Dapagliflozin, Diabetic Kidney Disease, Fibrosis, Glucose, Glucose Transporters, Smad3, Biochemistry & Molecular Biology, 0601 Biochemistry and Cell Biology
SGUL Research Institute / Research Centre: Academic Structure > Infection and Immunity Research Institute (INII)
Journal or Publication Title: Biosci Rep
ISSN: 1573-4935
Language: eng
Dates:
DateEvent
25 June 2021Published
18 May 2021Accepted
Publisher License: Creative Commons: Attribution 4.0
Projects:
Project IDFunderFunder ID
ST_008_20151127Kidney Research UKhttp://dx.doi.org/10.13039/501100000291
TKF PhD2017South West Thames Kidney FundUNSPECIFIED
PubMed ID: 34003249
Go to PubMed abstract
URI: http://openaccess.sgul.ac.uk/id/eprint/113380
Publisher's version: https://doi.org/10.1042/BSR20203947

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