Mécanisme(s) d'action de l'insuline dans la prévention de l'hypertension et la progression de la tubulopathie dans le diabète : rôle de hnRNP F, Nrf2 et Bmf

Diabetes mellitus is a complex metabolic disorder characterized by abnormal glucose homeostasis, resulting in higher plasma glucose due to an absolute or relative deficit in insulin production or action. People with diabetes have an increased risk of developing complications including diabetic nephropathy (DN), which is the major cause of developing end stage renal disease (ESRD) and is associated with increased cardiovascular morbidity and mortality. Although, type I and type II diabetes (T1D and T2D) are developed by different mechanisms, there is no major pathophysiological difference between nephropathy progression and ESRD in both diabetes.  Tubulopathy including tubular apoptosis/atrophy and tubule-interstitial fibrosis is known to be final marker for DN progression. Hyperglycemia and oxidative stress are associated with hypertension and tubular injury; their precise molecular mechanisms remain unclear. Intensive insulin treatment for T1D patients, including daily insulin injections is the most effective therapy but is associated with drawbacks such as hypoglycemia. The aim of this thesis is to identify downstream target genes or molecules of insulin action as potential therapeutic targets to counter DN progression. Firstly, we investigated whether insulin affects renal Nrf2 expression in T1D and studied its underlying mechanism and reported that insulin treatment normalized hyperglycemia, hypertension, oxidative stress, and renal injury; inhibited renal Nrf2 and Agt gene expression; and upregulated heterogeneous nuclear ribonucleoprotein F and K (hnRNP F/K) expression in Akita mice. In vitro, insulin suppressed Nrf2 and Agt but stimulated hnRNP F/K gene transcription in HG via p44/42 mitogen-activated protein kinase (p44/42 MAPK) signalling in RPTCs. Inhibition with siRNAs of p44/42 MAPK, hnRNP F, or hnRNP K, reversed insulin inhibition of Nrf2 gene transcription. We further identified an insulin-responsive element (IRE) in rat Nrf2 promoter that binds to hnRNP F/K. In hyperinsulinemic-euglycemic clamp studies, renal Nrf2 and Agt expression were downregulated, whereas hnRNP F/K expression was upregulated, indicating insulin-mediated effects on Nrf2 and Agt expression largely occur independently of its glucose-lowering effect.  Secondly, the underlying mechanism of insulin action on preventing RPTC apoptosis was studied. In the present study, a Tg mouse overexpressing human Bcl2-modifying factor (Bmf) in RPTs was created and characterized. Furthermore, the molecular mechanism(s) of insulin action on preventing Bmf-induced RPTC apoptosis and loss in diabetic mice were investigated. To study the effect of insulin on Bmf expression, Akita mice implanted with insulin, specifically those overexpressing hnRNP F in their RPTCs, were used. Bmf-Tg mice exhibited higher systolic blood pressure (SBP), urinary albumin-creatinine ratio (ACR), RPTC apoptosis and more urinary RPTCs than non-Tg mice. Insulin treatment and hnRNP F-overexpression in Akita mice suppressed RPTC Bmf expression and apoptosis. In vitro, insulin inhibited HG-induced Bmf gene expression in RPTCs via p44/42 MAPK signaling. Transfection of hnRNP F siRNA prevented insulin inhibition of Bmf transcription. HnRNP F inhibited Bmf transcription via hnRNP F-responsive element in the Bmf promoter.  In summary, this thesis demonstrated that chronic Nrf2 activation by hyperglycemia aggravates renal dysfunction via enhanced intrarenal Agt expression and RAS activation in diabetes. It was demonstrated that insulin inhibits Agt, Nrf2, and Bmf expression in RPTCs via hnRNP F and hnRNP K expression and, subsequently, attenuates hypertension and kidney injury in Akita mice. This study identifies renal hnRNP F/K, Nrf2 and Bmf as potential targets for the treatment of hypertension and kidney injury in diabetes.

Keyphrases: Rein, Catalase, Hypertension, Bmf, Kidney, Renin angiotensin system, Angiotensinogen, Diabetic nephropathy, Diabetic Kidney Disease, Apoptosis, Tubulointerstitial fibrosis, Reactive oxygen species, Tubulopathy, Nrf2, hnRNP F, hnRNP K, ROS, Insulin