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Dysregulation of nicotinamide adenine dinucleotide (NAD +) metabolism contributes to the initiation and progression of age-associated diseases, including chronic kidney disease (CKD). Nicotinamide N-methyltransferase (NNMT), a nicotinamide (NAM) metabolizing enzyme, regulates both NAD + and methionine metabolism. Although NNMT is expressed abundantly in the kidney, its role in CKD and renal fibrosis remains unclear. We generated NNMT-deficient mice and a unilateral ureter obstruction (UUO) model and conducted two clinical studies on human CKD to investigate the role of NNMT in CKD and fibrosis. In UUO, renal NNMT expression and the degraded metabolites of NAM increased, while NAD + and NAD + precursors decreased. NNMT deficiency ameliorated renal fibrosis; mechanistically, it (1) increased the DNA methylation of connective tissue growth factor (CTGF), and (2) improved renal inflammation by increasing renal NAD + and Sirt1 and decreasing NF-κB acetylation. In humans, along with CKD progression, a trend toward a decrease in serum NAD + precursors was observed, while the final NAD + metabolites were accumulated, and the level of eGFR was an independent variable for serum NAM. In addition, NNMT was highly expressed in fibrotic areas of human kidney tissues. In conclusion, increased renal NNMT expression induces NAD + and methionine metabolism perturbation and contributes to renal fibrosis.

The progression of chronic kidney disease (CKD) cannot be completely inhibited. We first explored factors contributing to CKD progression in patients with CKD in a prospective observational study. In the next phase, we focused on the effects of aldosterone, conducting a single-blinded placebo-controlled study using the selective mineralocorticoid receptor antagonist (MRA), eplerenone (25 mg/day). We recruited patients with CKD stage 2 and 3 whose plasma aldosterone concentration was above 15 ng/dL based on the prior data of a prospective observational study. In the CKD cohort study (n = 141), baseline plasma aldosterone concentration was identified as an independent contributory factor for the future rate of change in estimated glomerular filtration rate (eGFR). When the cut-off value for aldosterone was set at 14.5 ng/dL, the decline rate was significantly higher in patients with higher plasma aldosterone concentration (− 1.22 ± 0.39 ml/min/1.73 m2/year vs. 0.39 ± 0.40 ml/min/1.73 m2/year, p = 0.0047). In the final intervention study, in the eplerenone group, eGFR dropped at 6 months after the initiation of the study, and thereafter eGFR was maintained until the end of the study. At 24 months and 36 months, eGFR was significantly higher in the eplerenone group than in the placebo group. In conclusion, MRA can be an effective strategy in preventing CKD progression, especially in patients with high plasma aldosterone.

Acid-resistance systems are essential for pathogenic Escherichia coli to survive in the strongly acidic environment of the human stomach (pH tolC mutant lacking the TolC outer membrane channel was defective in GAD induction. Here, we show that indole, a substrate of TolC-dependent efflux pumps and produced by the tryptophanase encoded by the tnaA gene, negatively regulates GAD expression. GAD expression was restored by deleting tnaA in the tolC mutant; in wild-type E. coli, it was suppressed by adding indole to the growth medium. RNA-sequencing revealed that tnaA mRNA levels drastically decreased upon exposure to moderately acidic conditions (pH 5.5). This decrease was suppressed by RNase E deficiency. Collectively, our results demonstrate that the RNase E-dependent degradation of tnaA mRNA is accelerated upon acid exposure, which decreases intracellular indole concentrations and triggers GAD induction.

Under diabetic conditions, sodium–glucose cotransporter 2 (SGLT2) for glucose uptake in proximal tubules (PTs) increases, whereas NAD+-dependent protein deacetylase silent mating type information regulation 2 homolog 1 (Sirtuin-1; SIRT1) for PT survival decreases. Therefore, we hypothesized that increased glucose influx by SGLT2 reduces SIRT1 expression. To test this hypothesis, db/db mice with diabetes and high-glucose (HG)-cultured porcine PT LLC-PK1 cells in a two-chamber system were treated with the SGLT2 inhibitor canagliflozin. We also examined SIRT1 and SGLT2 expression in human kidney biopsies. In db/db mice, SGLT2 expression increased with concomitant decreases in SIRT1, but was inhibited by canagliflozin. For determination of the polarity of SGLT2 and SIRT1 expression, LLC-PK1 cells were seeded into Transwell chambers (pore size, 0.4 µm; Becton Dickinson, Oxford, UK). HG medium was added to either or to both of the upper and lower chambers, which corresponded to the apical and basolateral sides of the cells, respectively. In this system, the lower chamber with HG showed increased SGLT2 and decreased SIRT1 expression. Canagliflozin reversed HG-induced SIRT1 downregulation. Gene silencing and inhibitors for glucose transporter 2 (GLUT2) blocked HG-induced SGLT2 expression upregulation. Gene silencing for the hepatic nuclear factor-1α (HNF-1α), whose nuclear translocation was enhanced by HG, blocked HG-induced SGLT2 expression upregulation. Similarly, gene silencing for importin-α1, a chaperone protein bound to GLUT2, blocked HG-induced HNF-1α nuclear translocation and SGLT2 expression upregulation. In human kidney, SIRT1 immunostaining was negatively correlated with SGLT2 immunostaining. Thus, under diabetic conditions, SIRT1 expression in PTs was downregulated by an increase in SGLT2 expression, which was stimulated by basolateral HG through activation of the GLUT2/importin-α1/HNF-1α pathway.

Nicotinamide N-methyltransferase (NNMT) catalyses the reaction between nicotinamide (NAM) and S-adenosylmethionine to produce 1-methylnicotinamide and S-adenosylhomocysteine. Recently, this enzyme has also been reported to modulate hepatic nutrient metabolism, but its role in the liver has not been fully elucidated. We developed transgenic mice overexpressing NNMT to elucidate its role in hepatic nutrient metabolism. When fed a high fat diet containing NAM, a precursor for nicotinamide adenine dinucleotide (NAD)

The involvement of tissue ischemia in obesity-induced kidney injury remains to be elucidated. Compared with low fat diet (LFD)-mice, high fat diet (HFD)-fed mice became obese with tubular enlargement, glomerulomegaly and peritubular capillary rarefaction, and exhibited both tubular and glomerular damages. In HFD-fed mice, despite the increase in renal pimonidazole-positive areas, the expressions of the hypoxia-responsive genes such as Prolyl-hydroxylase PHD2, a dominant oxygen sensor, and VEGFA were unchanged indicating impaired hypoxic response. Tamoxifen inducible proximal tubules (PT)-specific Phd2 knockout (Phd2-cKO) mice and their littermate control mice (Control) were created and fed HFD or LFD. Control mice on HFD (Control HFD) exhibited renal damages and renal ischemia with impaired hypoxic response compared with those on LFD. After tamoxifen treatment, HFD-fed knockout mice (Phd2-cKO HFD) had increased peritubular capillaries and the increased expressions of hypoxia responsive genes compared to Control HFD mice. Phd2-cKO HFD also exhibited the mitigation of tubular damages, albuminuria and glomerulomegaly. In human PT cells, the increased expressions of hypoxia-inducible genes in hypoxic condition were attenuated by free fatty acids. Thus, aberrant hypoxic responses due to dysfunction of PHD2 caused both glomerular and tubular damages in HFD-induced obese mice. Phd2-inactivation provides a novel strategy against obesity-induced kidney injury.