Hypertension et régulation de l'expression moléculaire de l'angiotensinogène par la
ribonucléoprotéine hétérogène nucléaire K

Diabetes mellitus is a chronic metabolic disorder characterized by high plasma glucose caused by an impairment of insulin production, insulin action or both. Accumulating evidence has shown that chronic hyperglycemia can lead to dysfunction and failure of multiple organs including the heart, vascular system, eyes, and kidneys resulting in myocardial infarction, stroke, and retinal and renal complications, respectively. Diabetic nephropathy (DN) is the leading cause of end-stage renal disease affecting approximately 25–40% of diabetic patients. DN is invariably associated with an increased risk of stroke and cardiovascular dysfunction. Angiotensinogen (Agt) is the sole precursor for all types of angiotensins. In addition to systemic renin-angiotensin system (RAS), all the components of the intrarenal RAS are expressed in the kidney. Agt is highly expressed in the renal proximal tubular cells (RPTCs) and converted into biologically active angiotensin II (Ang II). In Diabetics, intrarenal Ang II level and RAS gene expression are upregulated, suggesting that intrarenal RAS activation plays an important role in the progression of DN. The mechanism (s) underlying the regulation of renal Agt by hyperglycemia and insulin are not completely understood. The overall aim of this thesis is to understand the molecular mechanism(s) that regulate renal Agt gene expression in an Akita mouse (a mouse model of type 1 diabetes). For this purpose, the first part of this thesis focuses on two transcription factors from the heterogenous nuclear ribonucleoprotein (hnRNPs) family. Previously, Chan’s group identified two nuclear proteins hnRNP F and hnRNP K of 48kD and 70kD, respectively. hnRNP F and hnRNP K form a heterodimer and bind to the insulin-responsive element (IRE) in the rat Agt gene promoter inhibiting Agt gene transcription in vitro. To determine whether hnRNP F / K mediate insulin inhibition of renal Agt expression in vivo, we used adult male Akita mice treated ± insulin implants for 4 weeks. Non-Akita mice served as controls. The Akita mice developed hypertension and exhibited renal hypertrophy. Insulin treatment normalized plasma glucose levels and systolic blood pressure (SBP), attenuated renal hypertrophy, decreased urinary albumin/creatinine ratio (ACR) and urinary Agt and Ang II levels in Akita mice. Furthermore, insulin treatment inhibited renal Agt expression but enhanced hnRNP F, hnRNP K and angiotensinconverting enzyme-2 (ACE2) expression. In vitro, insulin inhibited Agt but stimulated hnRNP F and hnRNP K expression in high-glucose media via p44/42 mitogen-activated protein kinase signaling in RPTCs. Transfection with hnRNP F and hnRNP K small interfering RNAs (siRNA) prevented the insulin inhibition of Agt expression in RPTCs. This study demonstrates that insulin prevents hypertension and attenuates kidney injury, at least in part, through suppressing renal Agt transcription via upregulation of hnRNP F and hnRNP K expression in diabetic Akita mice. In the second part of the thesis we focused on the nuclear factor erythroid 2-related factor 2 (Nrf2). Nrf2 is a transcription factor that regulates cellular antioxidant gene defense against oxidative stress or electrophiles. The purpose of this study is to investigate the impact of the overexpressing catalase (Cat) in RPTCs on Agt gene expression via Nrf2and the resulting effects on the development of hypertension and renal injury in diabetic Akita transgenic (Tg) mice. Our studies demonstrate that Cat overexpression normalizes SBP, attenuates renal injury, and inhibits RPTC Nrf2 and Agt gene expression in the Akita Cat- Tg compared to Akita mice. In vitro, high glucose (HG) and Oltipraz stimulated Nrf2 and Agt gene expression; these changes were blocked by Trigonelline (an inhibitor of Nrf2), siRNA against Nrf2, antioxidants, or pharmacological inhibitors of NF-kB and p38 mitogen-activated protein kinase. Moreover, deletion of Nrf2-responsive elements in the rat Agt gene promoter abolishes the stimulatory effect of Oltipraz. Finally,non transgenic adult male mice treated with the Nrf2 activator Oltipraz, upregulated Nrf2 and Agt expression in mouse RPTs, an effect that was normalized by Trigonelline. These data identify a novel mechanism via which Nrf2 mediates the stimulation of intrarenal Agt gene expression and activates the RAS through whichHG/reactive oxygen species (ROS) induce hypertension and renal injury in diabetic mice. Our findings demonstrate that the insulin induced hnRNP F and hnRNP K gene expression play a protective role in the preventing hypertension. Catalase overexpression, in RPT's, attenuates Nrf2 activation and lowers the SBP in Akita mice.

Keyphrases: HnRNP F, HnRNP K, Nrf2, Oltipraz, Trigonelline, Akita, Hypertension, Kidney, Angiotensinogen, HnRNP F, HnRNP K, Insulin, Nrf2, Oltipraz, Trigonelline, Akita, Hypertension