Abstract
ObjectiveChronic hyperglycaemia, as seen in type II diabetes, results in both morphological and functional impairments of podocytes in the kidney. We investigated the effects of high glucose (HG) on the insulin signaling pathway, focusing on cell survival and apoptotic markers, in immortalized human glomerular cells (HGEC; podocytes) and isolated glomeruli from healthy rats.Methods and FindingsHGEC and isolated glomeruli were cultured for various time intervals under HG concentrations in the presence or absence of insulin. Our findings indicated that exposure of HGEC to HG led to downregulation of all insulin signaling markers tested (IR, p-IR, IRS-1, p-Akt, p-Fox01,03), as well as to increased sensitivity to apoptosis (as seen by increased PARP cleavage, Casp3 activation and DNA fragmentation). Short insulin pulse caused upregulation of insulin signaling markers (IR, p-IR, p-Akt, p-Fox01,03) in a greater extent in normoglycaemic cells compared to hyperglycaemic cells and for the case of p-Akt, in a PI3K-dependent manner. IRS-1 phosphorylation of HG-treated podocytes was negatively regulated, favoring serine versus tyrosine residues. Prolonged insulin treatment caused a significant decrease of IR levels, while alterations in glucose concentrations for various time intervals demonstrated changes of IR, p-IR and p-Akt levels, suggesting that the IR signaling pathway is regulated by glucose levels. Finally, HG exerted similar effects in isolated glomeruli.ConclusionsThese results suggest that HG compromises the insulin signaling pathway in the glomerulus, promoting a proapoptotic environment, with a possible critical step for this malfunction lying at the level of IRS-1 phosphorylation; thus we herein demonstrate glomerular insulin signaling as another target for investigation for the prevention and/ or treatment of diabetic nephropathy.
Highlights
Diabetes remains the most common cause of end-stage renal failure in the United States and worldwide with between 20 and 40% of those with diabetes mellitus developing some degree of nephropathy [1,2]
high glucose (HG) exerted similar effects in isolated glomeruli. These results suggest that HG compromises the insulin signaling pathway in the glomerulus, promoting a proapoptotic environment, with a possible critical step for this malfunction lying at the level of insulin receptor substrate (IRS)-1 phosphorylation; we demonstrate glomerular insulin signaling as another target for investigation for the prevention and/ or treatment of diabetic nephropathy
IRS-1 basal levels in HG-treated cells were reduced compared to cells which were grown in normal glucose (NG) (Fig 1C)
Summary
Diabetes remains the most common cause of end-stage renal failure in the United States and worldwide with between 20 and 40% of those with diabetes mellitus developing some degree of nephropathy [1,2]. At the onset of diabetes, the kidney becomes enlarged and the GFR increases [3] without overt clinical signs and symptoms. Current treatments aim only to slow the progression of the disease, and these include strict blood glucose and pressure control and inhibition of the renin-angiotensin system [4,5]. Podocyte loss is evident in early diabetic nephropathy in humans and largely determines how rapidly the disease will progress [6,7,8,9]. Various mechanisms are involved in high glucose (HG)-induced podocytic injury and apoptosis and include increased renin-angiotensin-aldosterone signaling [10,11], oxidative stress [12], advanced glycation end products (AGEs) formation [13,14], and PKC activation [15]. Nephrin expression, a protein which contributes to podocyte survival by inhibiting proapoptotic signaling [16], is reduced in diabetes [17] and highly secreted in urine [18], an indication of podocyte injury
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