Sodium-glucose cotransporter 2 (SGLT2) inhibitors are a new class of antidiabetic oral agents indicating promising effects on cardiovascular and renal end points. However, the renoprotective effects of SGLT2 inhibitors are not fully understood. Also, metabolic effects of SGLT2 inhibition on other organ systems, such as effects on hepatic steatosis, are not fully understood. This study sought to address these questions by treating 18-wk-old uninephrectomized db/db mice with the selective SGLT2 inhibitor dapagliflozin. Untreated db/db mice developed progressive albuminuria, glomerular mesangial matrix expansion, and fatty liver associated with increased renal expression of TGFβ1, PAI-1, type IV collagen and fibronectin, and liver deposition of fibronectin, type I and III collagen, and laminin. Treatment with dapagliflozin (1 mg·kg⁻¹·day⁻¹) via gel diet from 18 to 22 wk of age not only reduced blood glucose (371.14 ± 55.02 mg/dl in treated db/db vs. 573.53 ± 21.73 mg/dl in untreated db/db, P < 0.05) and Hb A_1c levels (9.47 ± 0.79% in treated db/db vs. 12.1 ± 0.73% in untreated db/db, P < 0.05) but also ameliorated the increases in albuminuria and markers of glomerulosclerosis and liver injury seen in untreated db/db mice. Furthermore, both renal expressions of NF-kB p65, MCP-1, Nox4, Nox2, and p47phox and urine TBARS levels and liver productions of myeloperoxidase and reactive oxygen species, the markers of tissue inflammation and oxidative stress, were increased in untreated db/db mice, which were reduced by dapagliflozin administration. These results demonstrate that dapagliflozin not only improves hyperglycemia but also slows the progression of diabetes-associated glomerulosclerosis and liver fibrosis by improving hyperglycemia-induced tissue inflammation and oxidative stress.

Recent identification of a counterregulatory axis of the renin-angiotensin system, called angiotensin-converting enzyme 2-angiotensin-(1–7) [ANG-(1–7)]-Mas receptor, may offer new targets for the treatment of renal fibrosis. We hypothesized that therapy with ANG-(1–7) would improve glomerulosclerosis through counteracting ANG II in experimental glomerulonephritis. Disease was induced in rats with the monoclonal anti-Thy-1 antibody, OX-7. Based on a three-dose pilot study, 576 μg·kg⁻¹·day⁻¹ ANG-(1–7) was continuously infused from day 1 using osmotic pumps. Measures of glomerulosclerosis include semiquantitative scoring of matrix proteins stained for periodic acid Schiff, collagen I, and fibronectin EDA+ (FN). ANG-(1–7) treatment reduced disease-induced increases in proteinuria by 75%, glomerular periodic acid Schiff staining by 48%, collagen I by 24%, and FN by 25%. The dramatic increases in transforming growth factor-β₁, plasminogen activator inhibitor-1, FN, and collagen I mRNAs seen in disease control animals compared with normal rats were all significantly reduced by ANG-(1–7) administration ( P < 0.05). These observations support our hypothesis that ANG-(1–7) has therapeutic potential for reversing glomerulosclerosis. Several results suggest ANG-(1–7) acts by counteracting ANG II effects: 1) renin expression in ANG-(1–7)-treated rats was dramatically increased as it is with ANG II blockade therapy; and 2) in vitro data indicate that ANG II-induced increases in mesangial cell proliferation and plasminogen activator inhibitor-1 overexpression are inhibited by ANG-(1–7) via its binding to a specific receptor known as Mas.

Higher doses of AngII (angiotensin II) blockers are intended to optimize albuminuria reduction rather than for blood pressure control in chronic kidney disease. However, the long-term renoprotection of high-dose AngII blockers has yet to be defined. The present study sought to determine whether doses of ARB (AngII receptor blocker) that maximally reduce proteinuria could slow the progression of glomerulosclerosis in the uninephrectomized db/db mouse, a model of Type 2 diabetes. Untreated uninephrectomized db/db mice had normal blood pressure, but developed progressive albuminuria and mesangial matrix expansion between 18 and 22 weeks of age, which was associated with increased renal expression of TGFβ₁ (transforming growth factor β₁), PAI-1 (plasminogen-activator inhibitor-1), type IV collagen and FN (fibronectin). Treatment with valsartan in the drinking water of db/db mice from 18 to 22 weeks of age, at a dose that was determined previously to maximally reduce proteinuria, prevented the increases in albuminuria and the markers of renal fibrosis seen in untreated db/db mice. In addition, WT-1 (Wilms tumour protein-1)-immunopositive podocyte numbers were found to be lower in the untreated glomeruli of mice with diabetes. The expression of podocin and nephrin were continually decreased in mice with diabetes between 18 and 22 weeks of age. These changes are indicative of podocyte injury and the administration of valsartan ameliorated them substantially. Renal expression of TNFα (tumour necrosis factor α), MCP-1 (monocyte chemoattractant protein-1), Nox2 (NADPH oxidase 2), p22^phox and p47^phox and urine TBARS (thiobarbituric acid-reacting substance) levels, the markers of renal inflammation and oxidative stress, were increased during disease progression in mice with diabetes. Valsartan treatment was shown to reduce these markers. Thus high doses of valsartan not only reduce albuminuria maximally, but also halt the progression of the glomerulosclerosis resulting from Type 2 diabetes via a reduction in podocyte injury and renal oxidative stress and inflammation.

Although elevated plasma prorenin levels are commonly found in diabetic patients and correlate with microvascular complications, the pathological role of these increases, if any, remains unclear. Prorenin/renin binding to the prorenin/renin receptor [(p)RR] enhances the efficiency of angiotensinogen cleavage by renin and unmasks prorenin catalytic activity. We asked whether plasma prorenin could be activated in local vascular tissue through receptor binding. Immunohistochemical staining showing localization of the (p)RR in the aorta to vascular smooth muscle cells (VSMCs). After cultured rat VSMCs were incubated with 10⁻⁷M inactive prorenin, cultured supernatant acquired the ability to generate ANG I from angiotensinogen, indicating that prorenin had been activated. Activated prorenin facilitated angiotensin generation in cultured VSMCs when exogenous angiotensinogen was added. Small interfering RNA (siRNA) against the (p)RR blocked this activation and subsequent angiotensin generation. Prorenin alone induced dose- and time-dependent increases in mRNA and protein for the profibrotic molecule plasminogen activator inhibitor (PAI)-1, effects that were blocked by siRNA, but not by the ANG II receptor antagonist saralasin. When inactive prorenin and angiotensinogen were incubated with cells, PAI-1 mRNA increased a striking 54-fold, 8-fold higher than the increase seen with prorenin alone. PAI-1 protein increased 2.75-fold. These effects were blocked by treatment with siRNA + saralasin. We conclude that prorenin at high concentration binds the (p)RR on VSMCs and is activated. This activation leads to increased expression of PAI-1 via ANG II-independent and -dependent mechanisms. These data provide a mechanism by which elevated prorenin levels in diabetes may contribute to the progression of fibrotic disease.

Adiponectin (ApN) is a multifunctional adipokine. However, high, rather than low, concentrations of ApN are unexpectedly found in patients with chronic kidney disease (CKD) via an as yet unknown mechanism, and the role of ApN in CKD is unclear. Herein, we investigated the effect of ApN overexpression on progressive renal injury resulting from deoxycorticosterone acetate-salt (DOCA) and angiotensin II (ANG II) infusion using a transgenic, inducible ApN-overexpressing mouse model. Three groups of mice [wild type receiving no infusion (WT) and WT and cytochrome P450 1a1 (cyp1a1)-ApN transgenic mice (ApN-Tg) receiving DOCA+ANG II infusion (WT/DOCA+ANG II and ApN-Tg/DOCA+ANG II)] were assigned to receive normal food containing 0.15% of the transgene inducer indole-3-carbinol (I3C) for 3 wk. In the I3C-induced ApN-Tg/DOCA+ANG II mice, not the WT or WT/DOCA+ANG II mice, overexpression of ApN in liver resulted in 3.15-fold increases in circulating ApN compared with nontransgenic controls. Of note, the transgenic mice receiving DOCA+ANG II infusion were still hypertensive but had much less albuminuria and glomerular and tubulointerstitial fibrosis, which were associated with ameliorated podocyte injury determined by ameliorated podocyte loss and foot process effacement, and alleviated tubular injury determined by ameliorated mRNA overexpression of kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin and mRNA decreases of cubilin and megalin in tubular cells, compared with WT/DOCA+ANG II mice. In addition, renal production of NF-κB-p65, NAPDH oxidase 2, and p47^phoxand MAPK-related cellular proliferation, which were induced in WT/DOCA+ANG II mice, were markedly reduced in ApN-Tg/DOCA+ANG II mice. These results indicate that elevated ApN in the CKD mouse model is renal protective. Enhancing ApN production or signaling may have therapeutic potential for CKD.

Eph-Ephrin receptor-ligand signaling has been implicated in the development of tissue fibrosis, though it has not been well defined in the kidney. We detected substantial upregulation of expression and phosphorylation of the EphB2 receptor tyrosine kinase in fibrotic kidney tissue obtained both from mice subjected to the unilateral renal ischemia-reperfusion (IR) model at 14 days and in patients suffering from chronic kidney disease (CKD). Knockout mice lacking EphB2 expression exhibited a normal renal structure and function, indicating no major role for this receptor in kidney development or action. Although IR injury is well known to cause tissue damage, fibrosis, and renal dysfunction, we found that kidneys from EphB2 knockout mice showed much less renal tubular injury and retained a more preserved renal function. IR-injured kidneys from EphB2 knockouts exhibited greatly reduced fibrosis and inflammation compared to injured wild-type (WT) littermates, and this correlated with a significant reduction in renal expression of pro-fibrotic molecules, inflammatory cytokines, NADPH oxidases, and markers for cell proliferation, tubular epithelial-to-mesenchymal transition, myofibroblast activation, and apoptosis. A panel of 760 fibrosis-associated genes were further assessed, revealing that 506 genes in WT mouse kidney following IR injury changed their expression. However, 70.9% of those genes were back to or close to normal in expression when EphB2 was deleted. These data indicate endogenous EphB2 expression and signaling are abnormally activated after kidney injury and subsequently contributes to the development of renal fibrosis via regulation of multiple pro-fibrotic pathways.