Exposure of rats to elevated dietary salt following recovery from acute kidney injury (AKI) accelerates the transition to chronic kidney disease (CKD), and is dependent on lymphocyte activity. Here we tested whether high salt diet triggers lymphocyte activation in post-ischemic kidneys to worsen renal inflammation and fibrosis. Male Sprague- Dawley rats on a 0.4% salt diet were subjected to left unilateral ischemia-reperfusion and allowed to recover for 5 weeks. This resulted in a mild elevation of CD4+ T-cells relative to sham animals. Contralateral unilateral nephrectomy and elevated dietary salt (4%) for 4 extra weeks hastened CKD and interstitial fibrosis. Activated T cells were increased in the kidney 3-fold after 4 weeks of elevated dietary salt exposure relative to post AKI rats prior to salt feeding. The T-cell subset was largely positive for IL-17, indicative of Th-17 cells. Because angiotensin II activity may influence lymphocyte activation, injured rats were given the AT1R antagonist, Losartan, along with high salt diet. This significantly reduced the number of renal Th-17 cells to levels of sham rats, and significantly reduced the salt-induced increase in fibrosis about half. In vitro studies in AKI-primed CD4+ T cells indicated angiotensin II and extracellular sodium enhanced, and Losartan inhibited IL-17 expression. Thus, dietary salt modulates immune cell activity in post ischemic recovering kidneys due to the activity of local RAS suggesting participation of these cells in CKD progression post AKI.

T cells have been implicated in the pathogenesis of acute kidney injury (AKI) and its progression to chronic kidney disease (CKD). Previous studies suggest that Th17 cells participate during the AKI-to-CKD transition, and inhibition of T cell activity by mycophenolate mofetil (MMF) or losartan attenuates the development of fibrosis following AKI. We hypothesized that T cell-deficient rats may have reduced levels of IL-17 cytokine leading to decreased fibrosis following AKI. Renal ischemis-reperfusion (I/R) was performed on T cell-deficient athymic rats (Foxn1) and control euthymic rats (Foxn1), and CKD progression was hastened by unilateral nephrectomy at and subsequent exposure to 4.0% sodium diet. Renal fibrosis developed in euthymic rats and was reduced by MMF treatment. Athymic rats exhibited a similar degree of fibrosis, but this was unaffected by MMF treatment. FACS analysis demonstrated that the number of IL-17 cells was similar between postischemic athymic vs. euthymic rats. The source of IL-17 production in euthymic rats was predominately from conventional T cells (CD3/CD161). In the absence of conventional T cells in athymic rats, a compensatory pathway involving natural killer cells (CD3/CD161) was the primary source of IL-17. Blockade of IL-17 activity using IL-17Rc receptor significantly decreased fibrosis and neutrophil recruitment in both euthymic and athymic rats compared with vehicle-treated controls. Taken together, these data suggest that IL-17 secretion participates in the pathogenesis of AKI-induced fibrosis possibly via the recruitment of neutrophils and that the source of IL-17 may be from either conventional T cells or NK cells.

Ischemic preconditioning confers organ-wide protection against subsequent ischemic stress. A substantial body of evidence underscores the importance of mitochondria adaptation as a critical component of cell protection from ischemia. To identify changes in mitochondria protein expression in response to ischemic preconditioning, we isolated mitochondria from ischemic preconditioned kidneys and sham-treated kidneys as a basis for comparison. The proteomic screen identified highly upregulated proteins, including NADP +-dependent isocitrate dehydrogenase 2 (IDH2), and we confirmed the ability of this protein to confer cellular protection from injury in murine S3 proximal tubule cells subjected to hypoxia. To further evaluate the role of IDH2 in cell protection, we performed detailed analysis of the effects of Idh2 gene delivery on kidney susceptibility to ischemia-reperfusion injury. Gene delivery of IDH2 before injury attenuated the injury-induced rise in serum creatinine (P,0.05) observed in controls and increased the mitochondria membrane potential (P,0.05), maximal respiratory capacity (P,0.05), and intracellular ATP levels (P,0.05) above those in controls. This communication shows that gene delivery of Idh2 can confer organ-wide protection against subsequent ischemiareperfusion injury and mimics ischemic preconditioning.

Damage to endothelial cells contributes to acute kidney injury (AKI) by leading to impaired perfusion. Endothelial colony-forming cells (ECFC) are endothelial precursor cells with high proliferative capacity, pro-angiogenic activity, and in vivo vessel forming potential. We hypothesized that ECFC may ameliorate the degree of AKI and/or promote repair of the renal vasculature following ischemia-reperfusion (I/R). Rat pulmonary microvascular endothelial cells (PMVEC) with high proliferative potential were compared with pulmonary artery endothelial cells (PAEC) with low proliferative potential in rats subjected to renal I/R. PMVEC administration reduced renal injury and hastened recovery as indicated by serum creatinine and tubular injury scores, while PAEC did not. Vehicle-treated control animals showed consistent reductions in renal medullary blood flow (MBF) within 2 h of reperfusion, while PMVEC protected against loss in MBF as measured by laser Doppler. Interestingly, PMVEC mediated protection occurred in the absence of homing to the kidney. Conditioned medium (CM) from human cultured cord blood ECFC also conveyed beneficial effects against I/R injury and loss of MBF. Moreover, ECFC-CM significantly reduced the expression of ICAM-1 and decreased the number of differentiated lymphocytes typically recruited into the kidney following renal ischemia. Taken together, these data suggest that ECFC secrete factors that preserve renal function post ischemia, in part, by preserving microvascular function.