Sodium-glucose cotransporter 2 (SGLT2) inhibitors have pleiotropic properties beyond blood glucose–lowering effects and modify important nonglycemic pathways, leading to end-organ protection. SGLT2 inhibitors display renoprotective effects in diabetic kidney disease, which creates a rationale for testing the therapeutic potential of this drug class in nondiabetic chronic kidney disease. Here, we have shown that dapagliflozin provided glomerular protection in mice with protein-overload proteinuria induced by bovine serum albumin (BSA), to a similar extent as an ACE inhibitor used as standard therapy for comparison. Dapagliflozin limited proteinuria, glomerular lesions, and podocyte dysfunction and loss. We provide the observation that SGLT2 was expressed in podocytes and upregulated after BSA injections. Through in vitro studies with cultured podocytes loaded with albumin we have identified what we believe to be a novel mechanism of action for SGLT2 inhibitor that directly targets podocytes and relies on the maintenance of actin cytoskeleton architecture. Whether SGLT2 inhibitors represent a possible future therapeutic option for some patients with proteinuric glomerular disease who do not have as yet an effective treatment will require ad hoc clinical studies.

Renal activation of the complement system has been described in patients with diabetic nephropathy (DN), although its pathological relevance is still ill-defined. Here, we studied whether glomerular C3a, generated by uncontrolled complement activation, promotes podocyte damage, leading to proteinuria and renal injury in mice with type 2 diabetes. BTBR ob/ob mice exhibited podocyte loss, albuminuria, and glomerular injury accompanied by C3 deposits and increased C3a and C3a receptor (C3aR) levels. Decreased glomerular nephrin and α-actinin4 expression, coupled with integrin-linked kinase induction, were also observed. Treatment of DN mice with a C3aR antagonist enhanced podocyte density and preserved their phenotype, limiting proteinuria and glomerular injury. Mechanistically, ultrastructural and functional mitochondrial alterations, accompanied by downregulation of antioxidant superoxide dismutase 2 (SOD2) and increased protein oxidation, occurred in podocytes and were normalized by C3aR blockade. In cultured podocytes, C3a induced cAMP-dependent mitochondrial fragmentation. Alterations of mitochondrial membrane potential, SOD2 expression, and energetic metabolism were also found in response to C3a. Notably, C3a-induced podocyte motility was inhibited by SS-31, a peptide with mitochondrial protective effects. These data indicate that C3a blockade represents a potentially novel therapeutic strategy in DN for preserving podocyte integrity through the maintenance of mitochondrial functions.

Sirtuin 3 (SIRT3) is the primary mitochondrial deacetylase that controls the antioxidant pathway and energy metabolism. We previously found that renal Sirt3 expression and activity were reduced in mice with type 2 diabetic nephropathy associated with oxidative stress and mitochondrial abnormalities and that a specific SIRT3 activator improved renal damage. SIRT3 is modulated by diet, and to assess whether Sirt3 deficiency aggravates mitochondrial damage and accelerates kidney disease in response to nutrient overloads, wild-type (WT) and Sirt3^−/− mice were fed a high-fat-diet (HFD) or standard diet for 8 months. Sirt3^−/− mice on HFD exhibited earlier and more severe albuminuria compared to WT mice, accompanied by podocyte dysfunction and glomerular capillary rarefaction. Mesangial matrix expansion, tubular vacuolization and inflammation, associated with enhanced lipid accumulation, were more evident in Sirt3^−/− mice. After HFD, kidneys from Sirt3^−/− mice showed more oxidative stress than WT mice, mitochondria ultrastructural damage in tubular cells, and a reduction in mitochondrial mass and energy production. Our data demonstrate that Sirt3 deficiency renders mice more prone to developing oxidative stress and mitochondrial abnormalities in response to HFD, resulting in more severe kidney diseases, and this suggests that mitochondria protection may be a method to prevent HFD-induced renal injury.

A reduced nephron number at birth, due to critical gestational conditions, including maternal malnutrition, is associated with the risk of developing hypertension and chronic kidney disease in adulthood. No interventions are currently available to augment nephron number. We have recently shown that sirtuin 3 (SIRT3) has an important role in dictating proper nephron endowment. The present study explored whether SIRT3 stimulation, by means of supplementation with nicotinamide riboside (NR), a precursor of the SIRT3 co-substrate nicotinamide adenine dinucleotide (NAD⁺), was able to improve nephron number in a murine model of a low protein (LP) diet. Our findings show that reduced nephron number in newborn mice (day 1) born to mothers fed a LP diet was associated with impaired renal SIRT3 expression, which was restored through supplementation with NR. Glomerular podocyte density, as well as the rarefaction of renal capillaries, also improved through NR administration. In mechanistic terms, the restoration of SIRT3 expression through NR was mediated by the induction of proliferator-activated receptor γ (PPARγ) coactivator-1α (PGC-1α). Moreover, NR restored SIRT3 activity, as shown by the reduction of the acetylation of optic atrophy 1 (OPA1) and superoxide dismutase 2 (SOD2), which resulted in improved mitochondrial morphology and protection against oxidative damage in mice born to mothers fed the LP diet. Our results provide evidence that it is feasible to prevent nephron mass shortage at birth through SIRT3 boosting during nephrogenesis, thus providing a therapeutic option to possibly limit the long-term sequelae of reduced nephron number in adulthood.

Peritubular capillary rarefaction is a recurrent aspect of progressive nephropathies. We previously found that peritubular capillary density was reduced in BTBR ob/ob mice with type 2 diabetic nephropathy. In this model, we searched for abnormalities in the ultrastructure of peritubular capillaries, with a specific focus on the endothelial glycocalyx, and evaluated the impact of treatment with an angiotensin-converting enzyme inhibitor (ACEi). Mice were intracardially perfused with lanthanum to visualise the glycocalyx. Transmission electron microscopy analysis revealed endothelial cell abnormalities and basement membrane thickening in the peritubular capillaries of BTBR ob/ob mice compared to wild-type mice. Remodelling and focal loss of glycocalyx was observed in lanthanum-stained diabetic kidneys, associated with a reduction in glycocalyx components, including sialic acids, as detected through specific lectins. ACEi treatment preserved the endothelial glycocalyx and attenuated the ultrastructural abnormalities of peritubular capillaries. In diabetic mice, peritubular capillary damage was associated with an enhanced tubular expression of heparanase, which degrades heparan sulfate residues of the glycocalyx. Heparanase was also detected in renal interstitial macrophages that expressed tumor necrosis factor-α. All these abnormalities were mitigated by ACEi. Our findings suggest that, in experimental diabetic nephropathy, preserving the endothelial glycocalyx is important in order to protect peritubular capillaries from damage and loss.

Shiga toxin (Stx)-producing Escherichia coli is the predominant offending agent of post-diarrheal hemolytic uremic syndrome (HUS), a rare disorder of microvascular thrombosis and acute kidney injury possibly leading to long-term renal sequelae. We previously showed that C3a has a critical role in the development of glomerular damage in experimental HUS. Based on the evidence that activation of C3a/C3a receptor (C3aR) signaling induces mitochondrial dysregulation and cell injury, here we investigated whether C3a caused podocyte and tubular injury through induction of mitochondrial dysfunction in a mouse model of HUS. Mice coinjected with Stx2/LPS exhibited glomerular podocyte and tubular C3 deposits and C3aR overexpression associated with cell damage, which were limited by C3aR antagonist treatment. C3a promoted renal injury by affecting mitochondrial wellness as demonstrated by data showing that C3aR blockade reduced mitochondrial ultrastructural abnormalities and preserved mitochondrial mass and energy production. In cultured podocytes and tubular cells, C3a caused altered mitochondrial fragmentation and distribution, and reduced anti-oxidant SOD2 activity. Stx2 potentiated the responsiveness of renal cells to the detrimental effects of C3a through increased C3aR protein expression. These results indicate that C3aR may represent a novel target in Stx-associated HUS for the preservation of renal cell integrity through the maintenance of mitochondrial function.