1. ¹Research Centre, Centre hospitalier de l'Université de Montréal (CHUM) – Hôtel-Dieu, Montreal, Quebec, Canada
2. ²Department of Medicine, Mount Sinai Hospital/University Health Network, 600 University Avenue, Lebovic Building, Toronto, Ontario, Canada
3. ³Harvard Medical School, Massachusetts General Hospital, Pediatric Nephrology Unit, Boston, Massachusetts, USA

Correspondence: JSD Chan, Research Centre, Centre hospitalier de l'Université de Montréal (CHUM) – Hôtel-Dieu, 3850 Saint Urbain Street, Montreal, Quebec, Canada H2W 1T8. E-mail: john.chan@umontreal.ca

⁴Current address: Graduate Institute of Medical Genetics, Kaohsiung Medical University, 100 Shih-Chuan 1st Rd, Kaohsiung 80708, Taiwan, ROC.

Received 6 January 2005; Revised 12 September 2005; Accepted 5 October 2005; Published online 1 February 2006.

Abstract

Progression of diabetic nephropathy appears directly related to renal tubulointerstitial injury, but the involved genes are incompletely delineated. To identify such genes, DNA microarray analysis was performed with RNA from renal proximal tubules (RPTs) of streptozotocin-induced diabetic Wistar rats, spontaneously diabetic BioBreeding rats, and rat immortalized renal proximal tubular cells (IRPTCs) exposed to high glucose (25 mM) medium for 2 weeks. Osteopontin (OPN) mRNA expression was quantified by real time-quantitative polymerase chain reaction (RT-qPCR) or conventional reverse transcriptase-polymerase chain reaction (RT-PCR). OPN mRNA expression was upregulated (5–70-fold increase) in diabetic rat RPTs and in IRPTCs chronically exposed to high glucose compared to control RPTs and IRPTCs. High glucose, angiotensin II, phorbol 12-myristate 13-acetate and transforming growth factor-beta 1 (TGF-₁) stimulated OPN mRNA expression in IRPTCs in a dose- and time-dependent manner. This effect was inhibited by tiron, taurine, diphenylene iodinium, losartan, perindopril, calphostin C, or LY 379196 but not PD123319. IRPTCs overexpressing dominant-negative protein kinase C-beta 1 (PKC-₁) cDNA or antisense TGF-₁ cDNA prevented the high glucose effect on OPN mRNA expression. We concluded that high glucose-mediated increases in OPN gene expression in diabetic rat RPTs and IRPTCs are mediated, at least in part, via reactive oxygen species generation, intrarenal rennin–angiotensin system activation, TGF-₁ expression, and PKC-₁ signaling.