The endoplasmic reticulum (ER) stress response is activated in the diabetic kidney and functions to reduce ER protein accumulation and improve cellular function. We previously showed that tribbles homolog 3 (TRB3), an ER stress-associated protein, is upregulated in the diabetic kidney. Here, we investigated whether absence of TRB3 alters outcomes in diabetic nephropathy. Type 1 diabetes was induced in TRB3 wild-type and knockout ( 2/2 ) mice by low-dose streptozotocin, and the mice were followed for 12 weeks.Diabetic TRB3 2/2 mice developed higher levels of albuminuria and increased expression of inflammatory cytokine and chemokine mRNA in renal cortices relative to wild-type littermates, despite similar hyperglycemia. Diabetic TRB3 2/2 mice also expressed higher levels of ER stress-associated molecules in both the renal cortices and glomeruli. This change was associated with higher renal cortical phosphorylation of AKT at serine 473 (Ser 473 ), which is the AKT site phosphorylated by mammalian target of rapamycin complex-2 (mTORC2). We show in renal tubular cells that TRB3 binds to mTOR and the rapamycin-insensitive companion of mTOR (Rictor), a protein specific to mTORC2. Finally, we demonstrate in murine tubular cells that TRB3 can inhibit secretion of IL-6. Thus, TRB3 reduces albuminuria and inflammatory gene expression in diabetic kidney disease by a mechanism that may involve inhibition of the mTORC2/AKT pathway and may prove to be a novel therapeutic target.

Background/Aims: Acute kidney injury (AKI) contributes to significant morbidity and mortality in the intensive care unit (ICU). Plasma levels of interleukin (IL)-6 predict the development of AKI and are associated with higher mortality in ICU patients with AKI. Most studies in AKI have focused on the tubulo-interstitium, despite evidence of glomerular involvement. In the following study, our goals were to investigate the expression of IL-6 and its downstream mediators in septic-induced AKI. Methods: Podocytes were treated in vitro with lipopolysaccharide (LPS) and mice were treated with LPS, and we evaluated IL-6 expression by real-time PCR, ELISA and in situ RNA hybridization. Results: Following LPS stimulation, IL-6 is rapidly and highly induced in cultured podocytes and in vivo in glomeruli and infiltrating leukocytes. Surprisingly, in direct response to exogenous IL-6, podocytes produce lipocalin-2/neutrophil gelatinase-associated lipocalin (Lcn2/Ngal). LPS also potently induces Lcn2/Ngal expression in podocytes in culture and in glomeruli in vivo. Intense Lcn2/Ngal expression is also observed in IL-6 knockout mice, suggesting that while IL-6 may be sufficient to induce glomerular Lcn2/Ngal expression, it is not essential. Conclusions: The glomerulus is involved in septic AKI, and we demonstrate that podocytes secrete key mediators of AKI including IL-6 and Lcn2/Ngal.

Thiazolidinediones (TZDs), known as peroxisome proliferator-activated receptor (PPAR) agonists, are used to treat type 2 diabetes. However, ∼5% of patients experience the treatment-limiting side effect of edema. Studies have implicated activation of the epithelial sodium channel (ENaC) as a cause of TZD-induced fluid retention, although there have been conflicting reports. The goal of this study was to resolve the role of PPARγ in control of ENaC isoforms in the kidney. Herein, we demonstrate in mice that rosiglitazone (RGZ), a PPARγ ligand, increases body weight and abdominal fat pad fluid content and reduces hematocrit. Seven days of RGZ decreases ENaCα and ENaCβ mRNA and ENaCγ protein expression in the kidney cortex, and acute treatment for 5 h with pioglitazone, another potent TZD, does not increase renal ENaC isoform mRNA or protein expression. Pioglitazone also decreases ENaCα and ENaCγ mRNA expression in a cortical collecting duct cell line. As no direct transcriptional studies had been conducted, we examined the PPARγ-dependent regulation of ENaC. Pioglitazone represses ENaCγ promoter activity, and this repression is partially relieved by inhibition of protein synthesis. Chromatin immunoprecipitation assays revealed that repression is associated with a decrease in histone H4K5 acetylation at the proximal ENaCγ promoter. In summary, TZDs do not increase ENaC mRNA expression in the kidney, and in fact repress the ENaCγ promoter via an indirect transcriptional mechanism.