Mechanism Of Kidney Senescence Induced By High-affinity Sodium-dependent Dicarboxylate Cotransporter

Background and objective: Normal aging alters renal function, the high frequency of end-stage renal disease (ESRD) in the elderly. The incidence of acute renal failure is higher in older subjects, and the clinical outcome of renal failure is significantly worse with regard to mortality, requirement of dialysis, and complete recovery. So it is important to study the mechanism of aging kidney. The pathological changes of tubular-interstitial are especially important because they determine the aggravate degree of renal function. High-affinity sodium-dependent dicarboxylate cotransporter (NaDC3) is expressed in basolateral membrane of proximal renal tubular epithelial cells primarily involved in the uptake and metabolism of Krebs cycle intermediates, which include a variety of di- and tricarboxylates such as succinate, citrate, and a-ketoglutarate. It induces the aging process. The pathway of NaDC3 mediating aging would provide the targets of delaying aging. But the mechanism is yet unclear. NaDC3 provides energy for renal tubular epithelial cells, and there are tight link between energy metabolism and lifespan. SIRT1, a NAD-dependent histone deacetylase, extends many organism lifespan and reduces aging-related diseases in caloric restriction. SIRT1 links energy metabolism and lifespan through NAD. We presumed NaDC3 probably regulated lifespan through inhibiting SIRT1 function. To determine the mechanism of aging induced by NaDC3, the study designed to investigate firstly the change of SIRT1 expression and activity in kidney with aging, and then detected the effects of NaDC3 on the level of SIRT1 expression and activity in HKC and WI38 cells, senescent-related phenotypes in WI38 cells, and NAD~+/NADH ratio.Methods: The kidneys from different month-old rats were used to investigate the change of SIRT1 and NaDC3 expression and SIRT1 activity. To determine the relation between NaDC3 and SIRT1, the expression and activity of SIRT1 protein were tested after transfecting NaDC3 gene in HKC and WI38 cells. To determine the effects of NaDC3 on SIRT1 in the aging process, aging-related phenotypes were tested after transfecting sense-NaDC3 in WI38 cells, including that the SA-β-gal staining positive ratio was detected by SA-β-gal staining, cells proliferation capacity was done by MTT, cell cycle was analysesed by flowcytometry system. Moreover, above parameters were also detected in WI38 transfecting sense-NaDC3 treated with resveratrol or CR serum. To determine the pathway of NaDC3 influencing SIRT1, we detected NAD~+/NADH ratio. Meanwhile, we tested aging-related phenotypes in WI38 transfecting sense-NaDC3 treated with NAD.Results: The results showed that the level of NaDC3 expression increased and SIRT1 expression and activity decreased in rat kidneys with aging (P<0.05). The level of SIRT1 expression and activity was significantly lower in HKC and WI38 cells transfecting sense-NaDC3 (P<0.05), while higher in HKC transfecting antisense-NaDC3 (ASNaDC3) than control cells (P<0.05). Cell prolification capacity and population doubling (PD) decreased, while the SA-β-gal staining positive ratio increased, and the cell number in G1 phase also increased in WI38 cells transfecting sense-NaDC3, compared with control group (P<0.05). But the aging-related phenotypes were reduced when WI38 cells transfecting sense-NaDC3 were treated with resveratrol or CR serum. NAD~+/NADH ratio was inhibited in WI38 cells transfecting sense-NaDC3, compared with control group (P<0.05). But when the cells transfecting sense-NaDC3 were treated with NAD~+, the aging-related phenotypes were reduced (P< 0.05).Conclusions: Our results have demonstrated that the level of SIRT1 expression and activity decreased in kidney with aging. NaDC3 promoted senescence by inhibiting SIRT1 expression and activity, which may provide a new target of delaying and preventing renal senescence.