Search

Aims/Introduction The aim of this study was to elucidate whether sodium–glucose cotransporter 2 inhibitors (SGLT2is) treatment has any renoprotective effect for type 2 diabetes mellitus patients with an estimated glomerular filtration rate (eGFR) of, Administration of sodium–glucose cotransporter 2 inhibitors slowed the annual estimated glomerular filtration rate decline in type 2 diabetes patients with an estimated glomerular filtration rate of

Although Rathke's cleft cysts (RCCs) are common sellar/parasellar lesions, studies examining pituitary function in patients with nonsurgical RCC are limited. This study aimed to clarify the importance of RCCs, including small nonsurgical ones, as a cause of hypopituitarism by determining the prevalence of pituitary hormone secretion impairment and its relationship to cyst/tumor size in patients with RCC and in those with nonfunctioning pituitary adenoma (NFA). We retrospectively investigated the basal levels of each anterior pituitary hormone, its responses in the stimulation test(s), and cyst/tumor size in patients with RCC (n = 67) and NFA (n = 111) who were consecutively admitted to our hospital for endocrinological evaluation. RCCs were much smaller than NFAs (median height, 12 vs. 26 mm). The prevalence of gonadotropin, PRL, and GH secretion impairment in RCC was lower in comparison to NFA (19% vs. 44%, 34% vs. 61%, and 24% vs. 46%, respectively), whereas the prevalence of TSH and ACTH secretion impairment was comparable (21-27% and 17-24%, respectively). A significant positive relationship between cyst/tumor size and number of impaired hormones was observed in both groups, but smaller cysts could cause hormone secretion impairment in RCC. Stimulation tests suggested that most hormone secretion impairment was attributable to the interrupted hypothalamic-pituitary axis in both groups. Therefore, RCC, even small ones, can cause pituitary dysfunction. Different mechanisms may underlie hypothalamic-pituitary interruption in RCC and NFA.

Endothelial-to-mesenchymal transition (EndMT) has been shown to contribute to organ fibrogenesis. We have reported that N-acetyl-seryl-aspartyl- lysyl-proline (AcSDKP) restored levels of diabetes mellitus-suppressed FGFR1 (fibroblast growth factor receptor 1), the endothelial receptor essential for combating EndMT. However, the molecular regulation and biological/pathological significance of the AcSDKP-FGFR1 relationship has not been elucidated yet. Here, we demonstrated that endothelial FGFR1 deficiency led to AcSDKP-resistant EndMT and severe fibrosis associated with EndMT-stimulated fibrogenic programming in neighboring cells. Diabetes mellitus induced severe kidney fibrosis in endothelial FGFR1-deficient mice ( FGFR1 fl/fl ; VE-cadherin-Cre: FGFR1 EKO ) but not in control mice (FGFR1 fl/fl ); AcSDKP completely or partially suppressed kidney fibrosis in control or FGFR1 EKO mice. Severe fibrosis was also induced in hearts of diabetic FGFR1 EKO mice; however, AcSDKP had no effect on heart fibrosis in FGFR1 EKO mice. AcSDKP also had no effect on EndMT in either kidney or heart but partially suppressed epithelial-to-mesenchymal transition in kidneys of diabetic FGFR1 EKO mice. The medium from FGFR1-deficient endothelial cells stimulated TGFβ (transforming growth factor β)/Smad-dependent epithelial-to-mesenchymal transition in cultured human proximal tubule epithelial cell line, AcSDKP inhibited such epithelial-to-mesenchymal transition. These data demonstrated that endothelial FGFR1 is essential as an antifibrotic core molecule as the target of AcSDKP.

Aim To evaluate the effect of canagliflozin, a sodium-glucose co-transporter-2 (SGLT2) inhibitor, on albuminuria and the decline of estimated glomerular filtration rate (eGFR) in participants with type 2 diabetes and microalbuminuria. Methods The CANPIONE study is a multicentre, randomized, parallel-group and open-labelled study consisting of a unique 24-week preintervention period, during which the rate of eGFR decline before intervention is estimated, followed by a 52-week intervention and a 4-week washout period. Participants with a geometric mean urinary albumin-to-creatinine ratio (UACR) of 50 and higher and less than 300 mg/g in two consecutive first-morning voids at two different time points, and an eGFR of 45 ml/min/1.73m2 or higher, are randomly assigned to receive canagliflozin 100 mg daily or to continue guideline-recommended treatment, except for SGLT2 inhibitors. The first primary outcome is the change in UACR, and the second primary outcome is the change in eGFR slope. Results A total of 258 participants were screened and 98 were randomized at 21 sites in Japan from August 2018 to May 2021. The mean baseline age was 61.4 years and 25.8% were female. The mean HbA1c was 7.9%, mean eGFR was 74.1 ml/min/1.73m2 and median UACR was 104.2 mg/g. Conclusions The CANPIONE study will determine whether the SGLT2 inhibitor canagliflozin can reduce albuminuria and slow eGFR decline in participants with type 2 diabetes and microalbuminuria.

Aims/introduction To establish novel therapies to combat diabetic kidney disease, a human disease-relevant animal model is essential. However, a type 2 diabetic mouse model presenting progressive kidney fibrosis has not yet been established. Kidneys of streptozotocin-induced diabetic CD-1 mice showed severe fibrosis compared with other backgrounds of mice associated with the suppression of antifibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline. The BKS background (BKSdb / db ) is often utilized for diabetic kidney disease research; the kidney fibrosis in the BKSdb / db phenotype is minimal. Materials and methods We generated CD-1db / db mice by backcrossing the db gene into the CD-1 background, and analyzed phenotypic differences compared with BKSdb / db and CD-1db / m mice. Results Male CD-1db / db mice appeared to have elevated blood glucose levels compared with those of BKSdb / db mice. Fasting insulin levels declined in CD-1db / db mice. Plasma cystatin C levels tended to be elevated in CD-1db / db mice from 16 to 24 weeks-of-age. Male CD-1db / db mice showed significantly progressive kidney and heart fibrosis from 16 to 24 weeks-of-age when compared with that of age-matched BKSdb / db mice. The gene expression profile showed fibrogenic program-associated genes in male CD-1db / db mice. Male CD-1db / db mice displayed significantly lower urine antifibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline when compared to that of BKSdb / db at 24 weeks-of-age. The gene expression of prolyl oligopeptidase, the enzyme essential for antifibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline production from thymosin β4, was significantly lower in the CD-1 mice. Thymosin β4 levels were also lower in CD-1 mice. Conclusions These results suggest that CD-1db / db mice are a novel type 2 diabetic mouse model with progressive kidney and heart fibrosis.

Aims/Introduction We evaluated the efficacy of multifactorial intensive treatment (IT) on renal outcomes in patients with type 2 diabetes and advanced‐stage diabetic kidney disease (DKD). Materials and Methods The Diabetic Nephropathy Remission and Regression Team Trial in Japan (DNETT‐Japan) is a multicenter, open‐label, randomized controlled trial with a 5‐year follow‐up period. We randomly assigned 164 patients with advanced‐stage diabetic kidney disease (urinary albumin‐to‐creatinine ratio ≥300 mg/g creatinine, serum creatinine level 1.2–2.5 mg/dL in men and 1.0–2.5 mg/dL in women) to receive either IT or conventional treatment. The primary composite outcome was end‐stage kidney failure, doubling of serum creatinine or death from any cause, which was assessed in the intention‐to‐treat population. Results The IT tended to reduce the risk of primary end‐points as compared with conventional treatment, but the difference between treatment groups did not reach the statistically significant level (hazard ratio 0.69, 95% confidence interval 0.43–1.11; P = 0.13). Meanwhile, the decrease in serum low‐density lipoprotein cholesterol level and the use of statin were significantly associated with the decrease in primary outcome (hazard ratio 1.14; 95% confidence interval 1.05–1.23, P, The Diabetic Nephropathy Remission and Regression Team Trial in Japan was designed to clarify the beneficial effects of multifactorial intensified intervention by the team approach with medical staffs at each institution. There was an overall trend toward a lower risk on the development of kidney events in the intensive treatment group than in the conventional treatment group in this trial, but the benefit of intensive treatment could not be confirmed statistically. Lipid control by statin was associated with lower risk of kidney events in addition to strict control of blood glucose and blood pressure.

Diabetic kidney disease (DKD), a microvascular complication of diabetes, has been the leading cause of end-stage kidney disease (ESKD). Accordingly, patients with type 2 diabetes mellitus (T2DM) develop renal damage due to multiple metabolic and cardiorenal disease-related risk factors, including hyperglycemia, hypertension, dyslipidemia, hyperuricemia, and overnutrition/obesity. Despite multifactorial management including the administration of renin–angiotensin system inhibitors, patients often do not experience sufficient suppression of DKD progression and, thus, remain at risk for ESKD. Recent studies on cardiovascular outcomes among patients with T2DM have clearly shown that sodium–glucose cotransporter 2 (SGLT2) inhibitors, such as empagliflozin, canagliflozin, and dapagliflozin, have cardiorenal protective effects apart from their glucose-lowering effects. In particular, SGLT2 inhibitors have been found to improve renal outcomes, including ESKD, by slowing renal function decline and reducing urinary albumin excretion through their class effect. The proposed mechanisms for the renoprotective effects of SGLT2 inhibitors include the action of tubulo-glomerular feedback system and attenuation of hypoxia and metabolic stress in proximal tubular cells mediated through the inhibition of excessive glucose and sodium reabsorption, increased erythropoiesis, or increased ketone body production.

Mitochondrial oxidative stress is a significant contributor to the pathogenesis of diabetic kidney disease (DKD). We previously showed that mitochondrial oxidative stress in the kidneys of Zucker diabetic fatty rats is associated with a decreased intracellular NAD+/NADH ratio and NAD+-dependent deacetylase Sirt3 activity, and increased expression of the NAD+-degrading enzyme CD38. In this study, we used a CD38 inhibitor, apigenin, to investigate the role of CD38 in DKD. Apigenin significantly reduced renal injuries, including tubulointerstitial fibrosis, tubular cell damage, and pro-inflammatory gene expression in diabetic rats. In addition, apigenin down-regulated CD38 expression, and increased the intracellular NAD+/NADH ratio and Sirt3-mediated mitochondrial antioxidative enzyme activity in the kidneys of diabetic rats. In vitro, inhibition of CD38 activity by apigenin or CD38 knockdown increased the NAD+/NADH ratio and Sirt3 activity in renal proximal tubular HK-2 cells cultured under high-glucose conditions. Together, these results demonstrate that by inhibiting the Sirt3 activity and increasing mitochondrial oxidative stress in renal tubular cells, CD38 plays a crucial role in the pathogenesis of DKD.

Sodium-glucose cotransporter 2 inhibitors (SGLT2i) have been shown to display excellent renoprotective effects in diabetic kidney disease with macroalbuminuria/proteinuria. Regarding the renoprotective mechanism of SGLT2i, a sophisticated hypothesis was made by explaining the suppression of glomerular hypertension/hyperfiltration through the adenosine/adenosine type 1 receptor (A1R) signaling-mediated restoration of the tubuloglomerular feedback mechanism; however, how such A1R signaling is relevant for renoprotection by SGLT2i in diabetic kidney disease with proteinuria has not been elucidated.Streptozotocin-induced diabetic CD-1 mice were injected with bovine serum albumin (BSA) and treated with SGLT2i in the presence/absence of A1R inhibitor administration.We found that the influences of SGLT2i are essentially independent of the activation of A1R signaling in the kidney of BSA-overloaded streptozotocin-induced diabetic mice. BSA-overloaded diabetic mice showed the trend of kidney damage with higher glomerular filtration rate (GFR) and the significant induction of fibrogenic genes, such as transforming growth factor-β2 and collagen type III. SGLT2i TA-1887 suppressed diabetes-induced GFR in BSA-overloaded diabetic mice was associated with the significant suppression of transforming growth factor-β2 and collagen type III; A1R-specific inhibitor 8-cyclopentyl-1,3-dipropylxanthine did not cancel the effects of TA-1887 on either GFR or associated gene levels. Both TA-1887 and 8-cyclopentyl-1,3-dipropylxanthine-treated BSA-overloaded diabetic mice showed suppressed glycated hemoglobin levels associated with the increased food intake. When analyzing the association among histological evaluation, GFR and potential fibrogenic gene levels, each group of mice showed distinct correlation patterns.A1R signaling activation was not the dominant mechanism on the influence of SGLT2i in the kidney of BSA-overloaded diabetic mice.

Lifestyle improvement, including through exercise, has been recognized as an important mode of therapy for the suppression of diabetic kidney disease (DKD). However, the detailed molecular mechanisms by which exercise exerts beneficial effects in the suppression of DKD have not yet been fully elucidated. In this study, we investigate the effects of treadmill exercise training (TET) for 8 weeks (13 m/min, 30 min/day, 5 days/week) on kidney injuries of type 2 diabetic male rats with obesity (Wistar fatty (fa/fa) rats: WFRs) at 36 weeks of age. TET significantly suppressed the levels of albuminuria and urinary liver-type fatty-acid-binding protein (L-FABP), tubulointerstitial fibrosis, inflammation, and oxidative stress in the kidneys of WFRs. In addition, TET mitigated excessive apoptosis and restored autophagy in the renal cortex, as well as suppressed the development of morphological abnormalities in the mitochondria of proximal tubular cells, which were also accompanied by the restoration of AMP-activated kinase (AMPK) activity and suppression of the mechanistic target of rapamycin complex 1 (mTORC1). In conclusion, TET ameliorates diabetes-induced kidney injury in type 2 diabetic fatty rats.

Sodium-glucose cotransporter2 (SGLT2) inhibitors have a reno-protective effect in diabetic kidney disease. However, the detailed mechanism remains unclear. In this study, human proximal tubular cells (HK-2) were cultured in 5 mM glucose and 25 mM mannitol (control), 30 mM glucose (high glucose: HG), or HG and SGLT2 inhibitor, dapagliflozin-containing medium for 48 h. The autophagic flux was decreased, accompanied by the increased phosphorylation of S6 kinase ribosomal protein (p-S6RP) and the reduced phosphorylation of AMP-activated kinase (p-AMPK) expression in a HG condition. Compared to those of the control, dapagliflozin and SGLT2 knockdown ameliorated the HG-induced alterations of p-S6RP, p-AMPK, and autophagic flux. In addition, HG increased the nuclear translocation of nuclear factor-κB p65 (NF-κB) p65 and the cytoplasmic nucleotide-binding oligomerization domain-like receptor 3 (NLRP3), mature interleukin-1β (IL-1β), IL-6, and tumor necrosis factorα (TNFα) expression. Dapagliflozin, SGLT2 knockdown, and NF-κB p65 knockdown reduced the extent of these HG-induced inflammatory alterations. The inhibitory effect of dapagliflozin on the increase in the HG-induced nuclear translocation of NF-κB p65 was abrogated by knocking down AMPK. These data indicated that in diabetic renal proximal tubular cells, dapagliflozin ameliorates: (1) HG-induced autophagic flux reduction, via increased AMPK activity and mTOR suppression, and (2) inflammatory alterations due to NF-κB pathway suppression.

Aims To investigate the effects of sodium‐glucose co‐transporter‐2 (SGLT2) inhibitors vs. dipeptidyl peptidase‐4 (DPP‐4) inhibitors on renal function preservation (RFP) using real‐world data of patients with type 2 diabetes in Japan, and to identify which subgroups of patients obtained greater RFP benefits with SGLT2 inhibitors vs. DPP‐4 inhibitors. Methods We retrospectively analysed claims data recorded in the Medical Data Vision database in Japan of patients with type 2 diabetes (aged ≥18 years) prescribed any SGLT2 inhibitor or any DPP‐4 inhibitor between May 2014 and September 2016 (identification period), in whom estimated glomerular filtration rate (eGFR) was measured at least twice (baseline, up to 6 months before the index date; follow‐up, 9 to 15 months after the index date) with continuous treatment until the follow‐up eGFR. The endpoint was the percentage of patients with RFP, defined as no change or an increase in eGFR from baseline to follow‐up. A proprietary supervised learning algorithm (Q‐Finder; Quinten, Paris, France) was used to identify the profiles of patients with an additional RFP benefit of SGLT2 inhibitors vs. DPP‐4 inhibitors. Results Data were available for 990 patients prescribed SGLT2 inhibitors and 4257 prescribed DPP‐4 inhibitors. The proportion of patients with RFP was significantly greater in the SGLT2 inhibitor group (odds ratio 1.27; P = 0.01). The Q‐Finder algorithm identified four clinically relevant subgroups showing superior RFP with SGLT2 inhibitors (P

Aims/Introduction Diabetic kidney disease is characterized by increased albuminuria and/or a reduced glomerular filtration rate (GFR). We analyzed secular changes in the prevalence of albuminuria and reduced estimated GFR (eGFR) in Japanese patients with type 2 diabetes, and identified factors associated with these changes. Materials and Methods Using 1996, 2001, 2006 and 2014 cohort data from the Japanese serial cross‐sectional studies conducted at Shiga University of Medical Science, secular changes in the prevalence of diabetic kidney disease (albuminuria and/or reduced eGFR), patient characteristics and their associations were analyzed. Results The prevalence of microalbuminuria and macroalbuminuria decreased over time, whereas the prevalence of moderately reduced eGFR (30–60 mL/min/1.73 m2) and severely reduced eGFR (

Diabetic kidney disease (DKD) is appeared to be higher risk of declining kidney function compared to non-diabetic kidney disease with same magnitude of albuminuria. Epithelial-mesenchymal transition (EMT) program of tubular epithelial cells (TECs) could be important for the production of the extracellular matrix in the kidney. Caveolin-1 (CAV1), dipeptidyl peptidase-4 (DPP-4) and integrin β1 have shown to be involved in EMT program. Here, we found diabetic kidney is prone for albuminuria-induced TECs damage and DPP-4 plays a vital role in such parenchymal damages in diabetic mice. The bovine serum albumin (BSA) injection induced severe TECs damage and altered expression levels of DPP-4, integrin β1, CAV1, and EMT programs including relevant microRNAs in type 1 diabetic CD-1 mice when compared to non-diabetic mice; teneligliptin (TENE) ameliorated these alterations. TENE suppressed the close proximity among DPP-4, integrin β1 and CAV1 in a culture of HK-2 cells. These findings suggest that DPP-4 inhibition can be relevant for combating proteinuric DKD by targeting the EMT program induced by the crosstalk among DPP-4, integrin β1 and CAV1.

A biomarker, by which we can predict alterations of renal function in normoalbuminuric diabetic patients, is not available. Here, we report that endogenous anti-fibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) represents a potential biomarker to predict alterations in eGFR in normoalbuminuric diabetic patients. We analyzed 21 normoalbuminuric diabetic patients with eGFR ≥ 30 ml/min/1.73 m2 and measured AcSDKP levels in first morning void urine. We divided patients into two groups based on the median values: low or high urinary AcSDKP groups (uAcSDKP/Crlow or uAcSDKP/Crhigh). At baseline, no significant differences in sex, age, HbA1c, BMI, serum creatinine levels, etc., were observed between the two groups. During ~ 4 years, the alteration in eGFR [ΔeGFRop (ΔeGFR observational periods)] was significantly stable in uAcSDKP/Crhigh group compared with uAcSDKP/Crlow group over time (P = 0.003, χ2 = 8.58). We also evaluated urine kidney injury molecule-1 (uKim-1) levels and found that ΔeGFRop was also stable in low uKim-1 group compared with high uKim-1 group over time (P = 0.004, χ2 = 8.38). Patients who fulfilled the criteria for both uAcSDKP/Crhigh and uKim-1low exhibited stable ΔeGFRop (P

COMT (Catechol-O-methyl transferase), an enzyme that metabolizes catechol, requires magnesium (Mg 2+ ) to maintain its activity. Low COMT activity causes insufficient 2-methoxyestradiol (2-ME), a biologically active metabolite from hydroxyestradiol, which leads to hypertensive disorders, including preeclampsia. Hypoestrogenism increases the risk of salt-sensitive hypertension (SSH). SSH and preeclampsia are risk factors for each other; however, the molecular mechanism of this interaction is unclear. We focused on the interactive effect of Mg 2+ insufficiency and genetic COMT deficiency on SSH using 2 strains of mice with genetically distinct COMT activity. In male mice, BL6 (C57BL/6J), a high-activity COMT strain, displayed unaltered blood pressure regardless of the Mg 2+ and salt levels in food; DBA (DBA/2J), a low-activity COMT strain, developed SSH under low Mg 2+ and high-salt conditions. COMT inhibition in C57BL/6J strain also induced SSH. Treatment with 2-ME ameliorated SSH in both models. The ATR1 (angiotensin II type 1 receptor)–STE20-SPAK (serine-proline alanine-rich kinase)–NCC (sodium chloride cotransporter) axis, molecules associated with sodium reabsorption in distal convoluted tubules, was activated in mice that developed SSH. In female DBA mice, ovariectomized mice displayed SSH under low Mg 2+ associated with activation of ATR1-SPAK-NCC axis; 2-ME inhibited all, whereas the blood pressure of sham mice was unaltered regardless of any intervention. Our findings revealed that Mg 2+ insufficiency exaggerated the low COMT activity and induced SSH via the ATR1-SPAK-NCC axis due to 2-ME insufficiency, suggesting a new pathophysiological role that links COMT/2-ME deficiency with hypertensive syndrome.

Aims/Introduction Tubulointerstitial fibrosis is a hallmark of diabetic nephropathy and is associated with an epithelial‐to‐mesenchymal transition (EMT) program and aberrant glycolysis. Dimeric pyruvate kinase (PK) M2 (PKM2) acts as a key protein kinase in aberrant glycolysis by promoting the accumulation of hypoxia‐inducible factor (HIF)‐1α, while tetrameric PKM2 functions as a pyruvate kinase in oxidative phosphorylation. The aim of the research is to study the effect of PKM2 tetramer activation on preventing kidney fibrosis via suppression of aberrant glycolysis and the EMT program. Materials and methods In vivo: Streptozotocin (STZ) was utilized to induce diabetes in 8‐week‐old CD‐1 mice; 4 weeks after diabetes induction, proteinuria‐induced kidney fibrosis was developed by intraperitoneal injection of bovine serum albumin (BSA: 0.3 g/30 g BW) for 14 days; The PKM2 activator TEPP‐46 was also administered orally simultaneously. In vitro: HK2 cells were co‐treated with high‐glucose media or/and TGF‐β1 and TEPP46 for 48 h, cellular protein was extracted for evaluation. Results Diabetic mice developed kidney fibrosis associated with aberrant glycolysis and EMT; BSA injection accelerated kidney fibrosis in both the control and diabetic mice; TEPP‐46 rescued the kidney fibrosis. In HK2 cells, TEPP‐46 suppressed the EMT program induced by TGF‐β1 and/or high‐glucose incubation. TEPP‐46‐induced PKM2 tetramer formation and PK activity resulted in suppression of HIF‐1α and lactate accumulation. Specific siRNA‐mediated knockdown of HIF‐1α expression diminished high glucose‐induced mesenchymal protein levels. Conclusion PKM2 activation could restore the tubular phenotype via suppression of the EMT program and aberrant glycolysis, providing an alternative target to mitigate fibrosis in diabetic kidneys., Diabetic kidney disease displayed kidney fibrosis associated with aberrant glycolysis and EMT program, these alterations were related to the increased level of dimeric‐PKM2 formation, PKM2 activation by TEPP46 rescued the kidney fibrosis by promoting PKM2 tetramer formation, and inhibiting dimeric‐PKM2 which directly interacted with HIF‐1α and promoted its transcription.

The aim of this study was to investigate the effects of dietary supplementation with a nonalcoholic red wine extract (RWE), including resveratrol and polyphenols, on insulin sensitivity and Sirt1 expression in nondiabetic humans. The present study was a single-arm, open-label and prospective study. Twelve subjects received supplementation with RWE, including 19.2 mg resveratrol and 136 mg polyphenols, daily for 8 weeks. After 8 weeks, metabolic parameters, including glucose/lipid metabolism and inflammatory markers, were evaluated. mRNA expression of Sirt1 was evaluated in isolated peripheral blood mononuclear cells (PBMNCs). Additionally, Sirt1 and phosphorylated AMP-activated kinase (p-AMPK) expression were evaluated in cultured human monocytes (THP-1 cells). Supplementation with RWE for 8 weeks decreased the homeostasis model assessment for insulin resistance (HOMA-IR), which indicates an increase in insulin sensitivity. Serum low-density lipoprotein-cholesterol (LDL-C), triglyceride (TG) and interleukin-6 (IL-6) were significantly decreased by RWE supplementation for 8 weeks. Additionally, Sirt1 mRNA expression in isolated PBMNCs was significantly increased after 8 weeks of RWE supplementation. Moreover, the rate of increase in Sirt1 expression was positively correlated with the rate of change in HOMA-IR. The administration of RWE increased Sirt1 and p-AMPK expression in cultured THP-1 cells. Supplementation with RWE improved metabolism, such as insulin sensitivity, lipid profile and inflammation, in humans. Additionally, RWE supplementation induced an increase in Sirt1 expression in PBMNCs, which may be associated with an improvement in insulin sensitivity.

IntroductionGiven an increasing use of dipeptidyl peptidase-4 (DPP-4) inhibitors to treat patients with type 2 diabetes mellitus in the real-world setting, we conducted a prospective observational study (Japan-based Clinical Research Network for Diabetes Registry: J-BRAND Registry) to elucidate the safety and efficacy profile of long-term usage of alogliptin.Research design and methodsWe registered 5969 patients from April 2012 through September 2014, who started receiving alogliptin (group A) or other classes of oral hypoglycemic agents (OHAs; group B), and were followed for 3 years at 239 sites nationwide. Safety was the primary outcome. Symptomatic hypoglycemia, pancreatitis, skin disorders of non-extrinsic origin, severe infections, and cancer were collected as major adverse events (AEs). Efficacy assessment was the secondary outcome and included changes in hemoglobin A1c (HbA1c), fasting blood glucose, fasting insulin and urinary albumin.ResultsOf the registered, 5150 (group A: 3395 and group B: 1755) and 5096 (3358 and 1738) were included for safety and efficacy analysis, respectively. Group A patients mostly (>90%) continued to use alogliptin. In group B, biguanides were the primary agents, while DPP-4 inhibitors were added in up to ~36% of patients. The overall incidence of AEs was similar between the two groups (42.7% vs 42.2%). Kaplan-Meier analysis revealed the incidence of cancer was significantly higher in group A than in group B (7.4% vs 4.8%, p=0.040), while no significant incidence difference was observed in the individual cancer. Multivariate Cox regression analysis revealed that the imbalanced patient distribution (more elderly patients in group A than in group B), but not alogliptin usage per se, contributed to cancer development. The incidence of other major AE categories was with no between-group difference. Between-group difference was not detected, either, in the incidence of microvascular and macrovascular complications. HbA1c and fasting glucose decreased significantly at the 0.5-year visit and nearly plateaued thereafter in both groups.ConclusionsAlogliptin as a representative of DPP-4 inhibitors was safe and durably efficacious when used alone or with other OHAs for patients with type 2 diabetes in the real world setting.

Platelet-derived growth factor-B (PDGF-B) is a main factor to promote adipose tissue angiogenesis, which is responsible for the tissue expansion in obesity. In this process, PDGF-B induces the dissociation of pericytes from blood vessels; however, its regulatory mechanism remains unclear. In the present study, we found that stromal cell-derived factor 1 (SDF1) plays an essential role in this regulatory mechanism. SDF1 mRNA was increased in epididymal white adipose tissue (eWAT) of obese mice. Ex vivo pharmacological analyses using cultured adipose tissue demonstrated that physiological concentrations (1–100 pg/mL) of SDF1 inhibited the PDGF-B-induced pericyte dissociation from vessels via two cognate SDF1 receptors, CXCR4 and CXCR7. In contrast, higher concentrations (> 1 ng/mL) of SDF1 alone caused the dissociation of pericytes via CXCR4, and this effect disappeared in the cultured tissues from PDGF receptor β (PDGFRβ) knockout mice. To investigate the role of SDF1 in angiogenesis in vivo, the effects of anagliptin, an inhibitor of dipeptidyl peptidase 4 (DPP4) that degrades SDF1, were examined in mice fed a high-fat diet. Anagliptin increased the SDF1 levels in the serum and eWAT. These changes were associated with a reduction of pericyte dissociation and fat accumulation in eWAT. AMD3100, a CXCR4 antagonist, cancelled these anagliptin effects. In flow-cytometry analysis, anagliptin increased and decreased the PDGF-B expression in endothelial cells and macrophages, respectively, whereas anagliptin reduced the PDGFRβ expression in pericytes of eWAT. These results suggest that SDF1 negatively regulates the adipose tissue angiogenesis in obesity by altering the reactivity of pericytes to PDGF-B.

Sodium glucose cotransporter 2 (SGLT2) inhibitors are beneficial in halting diabetic kidney disease; however, the complete mechanisms have not yet been elucidated. The epithelial-mesenchymal transition (EMT) is associated with the suppression of sirtuin 3 (Sirt3) and aberrant glycolysis. Here, we hypothesized that the SGLT2 inhibitor empagliflozin restores normal kidney histology and function in association with the inhibition of aberrant glycolysis in diabetic kidneys. CD-1 mice with streptozotocin-induced diabetes displayed kidney fibrosis that was associated with the EMT at 4 months after diabetes induction. Empagliflozin intervention for 1 month restored all pathological changes; however, adjustment of blood glucose by insulin did not. Empagliflozin normalized the suppressed Sirt3 levels and aberrant glycolysis that was characterized by HIF-1α accumulation, hexokinase 2 induction, and pyruvate kinase isozyme M2 dimer formation in diabetic kidneys. Empagliflozin also suppressed the accumulation of glycolysis byproducts in diabetic kidneys. Another SGLT2 inhibitor, canagliflozin, demonstrated similar in vivo effects. High-glucose media induced the EMT, which was associated with Sirt3 suppression and aberrant glycolysis induction, in the HK2 proximal tubule cell line; SGLT2 knockdown suppressed the EMT, with restoration of all aberrant functions. SGLT2 suppression in tubular cells also inhibited the mesenchymal transition of neighboring endothelial cells. Taken together, SGLT2 inhibitors exhibit renoprotective potential that is partially dependent on the inhibition of glucose reabsorption and subsequent aberrant glycolysis in kidney tubules.

Aims/Introduction The objective of the present study was to elucidate the effect of switching to teneligliptin from other dipeptidyl peptidase‐4 (DPP‐4) inhibitors on glucose control and renoprotection in type 2 diabetes mellitus patients with diabetic kidney disease. Materials and Methods The present study was a single‐arm, open‐label, observational study. A total of 23 patients, who had urinary albumin/creatinine ratios (UACR) ≥30 mg/gCr in their first urine in the early morning, and received other DPP‐4 inhibitors and renin‐angiotensin system inhibitors, switched to teneligliptin 20 mg/day. After switching to teneligliptin for 24 weeks, we evaluated changes in glycated hemoglobin (HbA1c), fasting plasma glucose levels, plasma DPP‐4 activity and UACR. Results HbA1c, fasting plasma glucose and UACR values showed no significant change after 24 weeks compared with baseline. However, plasma DPP‐4 activity was significantly reduced after 24 weeks (0.57 ± 0.26 nmol/min/mL, P = 0.012, vs baseline), compared with baseline (1.49 ± 1.73 nmol/min/mL), and there was a positive relationship between the change rate of plasma DPP‐4 activity (Δ%DPP‐4) for 24 weeks and the levels of plasma DPP‐4 activity (r = −0.5997, P = 0.0025) and fasting plasma glucose (r = −0.4235, P = 0.0440) at baseline. Additionally, the Δ%DPP‐4 for 24 weeks was significantly correlated to the change rate of UACR (r = 0.556, P = 0.0059). However, there was no relationship between Δ%DPP‐4 and ΔHbA1c (amount of HbA1c change). Conclusions Switching to teneligliptin from other DPP‐4 inhibitors for 24 weeks reduces plasma DPP‐4 activity, which is associated with a reduction in albuminuria, independent of the change in glucose levels, in type 2 diabetes mellitus patients with diabetic kidney disease.

Muscle biology is important topic in diabetes research. We have reported that a diet with ketogenic amino acids rich replacement (KAAR) ameliorated high-fat diet (HFD)-induced hepatosteatosis via activation of the autophagy system. Here, we found that a KAAR ameliorated the mitochondrial morphological alterations and associated mitochondrial dysfunction induced by an HFD through induction of the AKT/4EBP1 and autophagy signaling pathways in both fast and slow muscles. The mice were fed with a standard HFD (30% fat in food) or an HFD with KAAR (HFDKAAR). In both the gastrocnemius and the soleus, HFDKAAR ameliorated HFD-impaired mitochondrial morphology and mitochondrial function, characterized by decreased mitofusin 2, optic atrophy 1, peroxisome proliferator-activated receptor (PPAR) γ coactivator-1α and PPARα levels and increased dynamin-related protein 1 levels. The decreased levels of phosphorylated AKT and 4EBP1 in the gastrocnemius and soleus of HFD-fed mice were remediated by HFDKAAR. Furthermore, the HFDKAAR ameliorated the HFD-induced autophagy defects in the gastrocnemius and soleus. These findings suggest that KAAR may be a novel strategy to combat obesity-induced mitochondrial dysfunction, likely through induction of the AKT/4EBP1 and autophagy pathways in skeletal muscle.

Key Clinical Message Vitamin D plays vital role for the health, and its deficiency has been implicated in the diverse pathological conditions such as hypomagnesemia and abnormal immune system. Here, we present a case of severe electrolytes disorders (hypokalemia and hypomagnesemia etc.) and kidney damages associated with vitamin D deficiency.

Background The objective of this study is to investigate the effects of a low-protein diet (LPD) starting from a young age on diabetic status and renal injury in a rat model of type 2 diabetes and obesity. Methods Diabetic male Wistar fatty (fa/fa) rats (WFRs) were fed a standard diet (23.84% protein) or an LPD (5.77% protein) for 24 weeks beginning at 6 weeks of age. We investigated the effects of the LPD on total body weight (BW); fat weight (FW); lower-limb muscle weight (MW); several measures of diabetic status, including fasting/random glucose levels, HOMA-IR and the IPITT; and renal injuries, including renal hypertrophy, albuminuria and histological changes. Additionally, autophagy and activation of mTORC1 were evaluated in the diabetic renal cortex. Furthermore, plasma FGF21 and high-molecular-weight (HMW) adiponectin levels, as well as UCP1 expression levels in brown adipose tissue (BAT), were evaluated. Results Increases in BW and FW in WFRs were significantly reduced by the LPD, and the LPD resulted in a significant reduction of lower-limb MW in WFRs. The LPD suppressed the elevation of glucose levels in WFRs through improvement of insulin resistance. The LPD also elevated the plasma FGF21 and HMW adiponectin of WFRs, as well as UCP1 expression in the BAT of the animals. Renal hypertrophy, albuminuria, renal histological changes, and increased expression of p62 and phospho-S6 ribosomal protein (p-S6RP) were observed in WFRs compared with the values from WLRs. The LPD clearly prevented the diabetic kidneys from sustaining any damage. Conclusions The LPD prevented the progression of diabetic status; this effect may have been associated with the reduction of FW and the elevation of plasma FGF21 and HMW adiponectin, as well as UCP1 expression in BAT, resulting in suppression of diabetic nephropathy. However, MW was decreased in rats by the consumption of an LPD from a young age; therefore, further research is needed to resolve the nutritional issue of LPD on decreasing in MW.

Objectives: Mitochondrial oxidative stress is involved in the pathogenesis of diabetic kidney disease. The objective of our study is to identify the mechanisms of renal mitochondrial oxidative stress, focusing on Sirt3, which is nicotinamide adenine dinucleotide (NAD+; oxidized NAD)-dependent deacetylase in mitochondria. Methods: Renal mitochondrial oxidative stress and Sirt3 activity, using Zucker diabetic fatty rats (ZDFRs) and cultured proximal tubular cells under high-glucose condition were evaluated. Results: At 28 weeks of age, ZDFRs exhibited the increased urinary albumin/liver-type fatty acid-binding protein (L-FABP)/8-hydroxy-2'-deoxyguanosine (8-OHdG) excretion, histological tubular cell damage, compared to non-diabetic Zucker Lean rats. In renal mitochondria, acetylated isocitrate dehydrogenase2 (IDH2) and superoxide dismutase2 (SOD2), accompanied with mitochondrial oxidative stress and mitochondrial morphologic alterations, were increased in ZDFRs, indicating inactivation of Sirt3. Additionally, expression of the NAD-degrading enzyme, CD38, was increased, and the NAD+/NADH (reduced NAD) ratio was reduced in the renal cortex of ZDFRs. High-glucose stimulation in cultured proximal tubular cells also resulted in an increase in acetylated IDH2/SOD2, CD38 overexpression and a reduction in the NAD+/NADH ratio. Conclusions: Enhancement of mitochondrial oxidative stress in the diabetic kidney was mediated by the reduction of Sirt3 activity. CD38 overexpression may be related to a reduction in the NAD+/NADH ratio in the diabetic kidney.

The natural history of typical and classical "diabetic nephropathy" has been described as high levels of albuminuria and subsequent renal function decline. However, recent decades, the cases, who show the reduced glomerular filtration rate (GFR) without the progression of albuminuria, has been increased. "Diabetic kidney disease (DKD)" is a concept that widely recognizes the pathophysiological change induced by diabetes as the onset and progressive factor of renal injury and renal function decline, regardless of the level of albuminuria. However, we may confuse that "chronic kidney disease (CKD) with diabetes" is "DKD". Therefore, to choose the appropriate treatment that should be prioritized in the clinical setting, we propose that "CKD with diabetes" is classified as "DKD", "non-DKD (NDKD) with diabetes" or "combined disease of DKD and NDKD".

Obesity-associated activation of the renin-angiotensin-aldosterone system is implicated in the pathogenesis of insulin resistance; however, influences of mineralocorticoid receptor (MR) inhibition remain unclear. Therefore, we aimed to clarify the anti-inflammatory mechanisms of MR inhibition using eplerenone, a selective MR antagonist, in C57BL/6 mice fed a high-fat diet (HFD) for 12 weeks. Eplerenone prevented excessive body weight gain and fat accumulation, ameliorated glucose intolerance and insulin resistance and enhanced energy metabolism. In the epididymal white adipose tissue (eWAT), eplerenone prevented obesity-induced accumulation of F4/80+CD11c+CD206−-M1-adipose tissue macrophage (ATM) and reduction of F4/80+CD11c−CD206+-M2-ATM. Interestingly, M1-macrophage exhibited lower expression levels of MR, compared with M2-macrophage, in the ATM of eWAT and in vitro-polarized bone marrow-derived macrophages (BMDM). Importantly, eplerenone and MR knockdown attenuated the increase in the expression levels of proIl1b, Il6 and Tnfa, in the eWAT and liver of HFD-fed mice and LPS-stimulated BMDM. Moreover, eplerenone suppressed IL1b secretion from eWAT of HFD-fed mice. To reveal the anti-inflammatory mechanism, we investigated the involvement of NLRP3-inflammasome activation, a key process of IL1b overproduction. Eplerenone suppressed the expression of the inflammasome components, Nlrp3 and Caspase1, in the eWAT and liver. Concerning the second triggering factors, ROS production and ATP- and nigericin-induced IL1b secretion were suppressed by eplerenone in the LPS-primed BMDM. These results indicate that eplerenone inhibited both the priming and triggering signals that promote NLRP3-inflammasome activation. Therefore, we consider MR to be a crucial target to prevent metabolic disorders by suppressing inflammasome-mediated chronic inflammation in the adipose tissue and liver under obese conditions.

A low-protein diet (LPD), particularly, very low-protein diet (VLPD) is expected for reno-protection in advanced chronic kidney disease, including diabetic nephropathy. We previously also demonstrated that a VLPD clearly improved advanced diabetic nephropathy in a type 2 diabetes and obesity rat. However, clinically, an everyday long-term VLPD contributes to poor adherence, which may be related to controvertial results of an LPD on the suppression for diabetic nephropathy, and has nutritional issues, such as sarcopenia or protein-energy wasting. The aim of this study is to elucidate the reno-protective effect of a cyclic and intermittent VLPD, not an everyday VLPD, against the advanced experimental diabetic nephropathy. Diabetic male Wistar fatty (fa/fa) rats (WFRs) were treated with a standard diet (STD; 23.84% protein) or a cyclic and intermittent VLPD (5.77% protein) consisting of an STD for 3 days and a VLPD for 4 days a week for 20 weeks beginning at 24 weeks of age. A cyclic and intermittent VLPD significantly improved renal hypertrophy, and significantly decreased urinary albumin and liver-type fatty acid binding protein (L-FABP) excretion without changes in body weight or exacerbation of HbA1c levels in diabetic rats. Additionally, diabetes-induced renal injuries including fibrosis, tubular cell damage and inflammation were significantly ameliorated by a cyclic and intermittent VLPD in diabetic rats. Thus, based on our experimental data, a cyclic and intermittent VLPD may be a dietary regimen that is easy to continue and has less risk of malnutrition, compared to an everyday long-term VLPD, against advanced diabetic nephropathy.

2-methoxyestradiol (2-ME), an estrogen metabolite generated via catechol-o-methyltransferase (COMT), is multifunctional methoxy-catechol. Here, we report that COMT deficiency leads to glucose intolerance and 2-ME rescues COMT-deficient-associated metabolic defects. Liver COMT protein was suppressed in high fat diet (HFD)-fed or in pregnant mice. COMT suppression, by Ro41-0960 or siRNA, in HFD fed mice or in pregnant mice exacerbated glucose intolerance; 2-ME intervention ameliorated these defects. 2-ME effects on glucose tolerance were associated with AMPK phosphorylation in the liver and in islet cells. Metformin restored liver COMT protein levels, and metformin-induced liver AMPK phosphorylation was abolished by COMT inhibition. The amelioration in glucose tolerance by 2-ME was associated with biphasic insulin secretion in an environment-dependent manner. 2-ME-induced insulin secretion was associated with the AMPK phosphorylation, PDX-1 phosphorylation, and MST-1 suppression in MIN-6 cells. Furthermore 2-ME displayed PPARγ agonist-like activity. These results suggest that COMT is an enzyme to maintain glucose homeostasis and 2-ME is a potential endogenous multi-target anti-diabetic candidate.

Platelet-derived growth factor (PDGF) is a key factor in angiogenesis; however, its role in adult obesity remains unclear. In order to clarify its pathophysiological role, we investigated the significance of PDGF receptor β (PDGFRβ) in adipose tissue expansion and glucose metabolism. Mature vessels in the epididymal white adipose tissue (eWAT) were tightly wrapped with pericytes in normal mice. Pericyte desorption from vessels and the subsequent proliferation of endothelial cells were markedly increased in the eWAT of diet-induced obese mice. Analyses with flow cytometry and adipose tissue cultures indicated that PDGF-B caused the detachment of pericytes from vessels in a concentration-dependent manner. M1-macrophages were a major type of cells expressing PDGF-B in obese adipose tissue. In contrast, pericyte detachment was attenuated and vascularity within eWAT was reduced in tamoxifen-inducible conditional Pdgfrb-knockout mice with decreases in adipocyte size and chronic inflammation. Furthermore, Pdgfrb-knockout mice showed enhanced energy expenditure. Consequently, diet-induced obesity and the associated deterioration of glucose metabolism in wild-type mice were absent in Pdgfrb-knockout mice. Therefore, PDGF-B–PDGFRβ signaling plays a significant role in the development of adipose tissue neovascularization and appears to be a fundamental target for the prevention of obesity and type 2 diabetes.

Hypoxia causes proximal tubular cell damage in diabetes, even though proximal tubular cells have an adaptive system to combat hypoxia involving induction of hypoxia factor-1 (HIF-1) and inhibition of mechanistic target of rapamycin complex 1 (mTORC1). Here, we examined the interference effect of altered glucose and lipid metabolism on the hypoxia responses in proximal tubular cells. In culture, hypoxia alone induced HIF-1 and inhibited mTORC1, preventing death in proximal tubular cells. However, hypoxia with high glucose and palmitate increased mTORC1 activity and promoted apoptosis in proximal tubular cells, which was inhibited by pharmacological and genetic inactivation of mTORC1. Since inhibition of all mTORC1's physiological functions regulated by growth factors including insulin causes various adverse effects, we screened for a microRNA that can inhibit only pro-apoptotic effects of mTORC1 to discover a safe therapeutic target. This screen found microRNA-148b-3p was able to specifically inhibit mTORC1-dependent apoptosis in hypoxic proximal tubular cells exposed to high glucose and palmitate, without affecting insulin-dependent mTORC1 activation. Furthermore, tumor necrosis factor receptor (TNFR) 2 was the target of microRNA-148b-3p and its suppression inhibited apoptosis. Finally, enhanced apoptosis with TNFR2 overexpression was found in hypoxic and mTORC1-activated proximal tubular cells in diabetic rats. Thus, diabetes activated mTORC1 even in hypoxic proximal tubular cells, leading to apoptosis by reducing microRNA-148b-3p expression. Modulating this pathogenic pathway may be a novel therapy for proximal tubular cell damage in diabetes.

Background: SGLT2 inhibitor (SGLT2i) displayed remarkable and early renal protection in overt nephropathy cases in clinical trials. However, detailed molecular mechanisms by which SGLT2i ameliorated renal damage in proteinuric diabetic patients were not elucidated yet. Here, we investigated that SGLT2i exerted renal protective effects via adenosine signal in diabetic mice with overt proteinuria. Methods: STZ (200mg/kgBW) was injected intraperitoneally to induce diabetes in 8-week-old CD-1 mice. Four weeks later, mice were treated with intraperitoneal injection of bovine serum albumin (BSA: 0.3g/30gBW) with or without oral administration of SGLT2i TA-1887 (30mg/kg). Result: In the STZ+BSA, pronounced renal tubular damages and interstitial fibrosis were observed; TA-1887 significantly ameliorated. The mRNA microarray analysis of kidney samples revealed that gene expressions related to TGF-β signaling pathway and epithelial-mesenchymal-transition program were increased in the STZ+BSA compared to the Control+BSA; TA-1887 suppressed them. Furthermore, the expression of ecto 5'-nucleotidase (5'-NT), which is essential for production of adenosine from AMP, was suppressed in STZ+BSA compared to Control+BSA; TA-1887 restored 5’-NT levels. In the immunohistological analysis, 5'-NT was expressed in glomeruli including mesangial cells and renal tubules of cortex/medulla in control kidney. However, 5’-NT was suppressed in all areas of STZ+BSA kidney; TA-1887 reversed the 5'-NT expression and distribution. Glomerular filtration rate (GFR) evaluated by inulin clearance was significantly greater in the STZ+BSA group and suppressed by TA-1887; the administration of the adenosine type 1 receptor inhibitor, DPCPX, cancelled the effects of TA-1887 on GFR and renal tubule damage. Conclusion: The adenosine signal plays an important role in the renal protective effects of SGLT2i in diabetic mice with proteinuria. Disclosure K. Shimada: None. Y. Takagaki: None. K. Kanasaki: Other Relationship; Self; Astellas Pharma Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Mitsubishi Tanabe Pharma Corporation, Ono Pharmaceutical Co., Ltd., Sanofi, Taisho Toyama Pharmaceutical. D. Koya: Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Mitsubishi Tanabe Pharma Corporation. Speaker's Bureau; Self; Astellas Pharma Inc., Astellas Pharma Inc., AstraZeneca, Merck & Co., Inc., Mitsubishi Tanabe Pharma Corporation, Taisho Pharmaceutical Co., Ltd. Funding Japan Society for the Promotion of Science

Background: In diabetic patients, the incident of heart failure is two to five times greater when compared to that of general population. The influence of DPP-4 inhibitor on the heart failure and cardiac damage in diabetes is not completely elucidated yet. Therefore, to understand the pathobiology of heart damage in diabetes and the influence of DPP-4 inhibitor is emerged as significant. Necroptosis, recently identified as programmed cell death, had shown to be involved in heart damage of ischemic heart disease model. Here we found that intervention with DPP-4 inhibitor linagliptin (LINA) ameliorated heart damage in diabetic mice associated with the suppression of necroptosis. Method: We utilized streptozotocin-induced diabetic CD-1 mice (STZ:200mg/kg). At 20 weeks after the onset of diabetes, diabetic mice were treated with LINA (STZ+LINA) for 4 weeks. Result: Blood pressure, blood glucose, body and heart weights were not different between STZ and STZ+LINA. Ultrasonic Echocardiography analysis revealed that LVDd and LVDs were significantly decreased in STZ and STZ+LINA compared with control. STZ-induced diabetic CD-1 mice exhibited perivascular fibrosis evaluated by picrosirius red staining and strong immunoreactivity for DPP-4 expression in cardiac interstitium; STZ+LINA exhibited suppression of DPP-4 and amelioration of heart fibrosis. Diabetic CD-1 mice exhibited deficiency in myocardial stripe structure and endothelial damage in electron microscopy; LINA normalized all. When necroptosis was evaluated by associated kinases, RIP3 and MLKL, diabetic CD-1 mice were exhibited a strong RIP3 and MLKL mRNA and protein expression levels in myocyte; LINA suppressed RIP3 and MLKL levels in myocyte. Conclusion: These results indicated that LINA restored cardiac structure associated with suppression of necroptosis in type 1 diabetic mice. Disclosure T. Hirai: None. Y. Takagaki: None. K. Kanasaki: Other Relationship; Self; Astellas Pharma Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Mitsubishi Tanabe Pharma Corporation, Ono Pharmaceutical Co., Ltd., Sanofi, Taisho Toyama Pharmaceutical. D. Koya: Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Mitsubishi Tanabe Pharma Corporation. Speaker's Bureau; Self; Astellas Pharma Inc., Astellas Pharma Inc., AstraZeneca, Merck & Co., Inc., Mitsubishi Tanabe Pharma Corporation, Taisho Pharmaceutical Co., Ltd. Funding Japan Society for the Promotion of Science

Background: Dietary interventions, including a low-protein diet (LPD) and methionine (Met) restriction, has shown longevity and metabolic health. LPD has a reno-protective effect against diabetic kidney disease (DKD) in rats with type 2 diabetes and obesity; however, it is unclear whether the beneficial effect of LPD is mediated by a low-Met intake. Methods: Rats were divided into four groups at 24 weeks of age: 1) non-diabetic Wistar lean (fa/-) rats fed a standard diet including 0.64% Met (STD) (Control); 2) Wistar fatty (fa/fa) rats with type 2 diabetes/obesity (WFRs) fed STD (0.64% Met) (Diabetes); 3) WFRs fed LPD (0.15% Met) (LPD); and 4) WFRs fed LP+Met (0.64% Met) (LP+Met). The dietary intervention was performed for 20 weeks. Findings: Accumulation of S-adenosylmethionine (SAM) in renal tubular cells of diabetic rats, associated with reduced glycine Nmethyltransferase (Gnmt) expression, led to mechanistic target of rapamycin complex1 (mTORC1) activation and impaired autophagy, resulting in the progression of DKD through mitochondrial abnormalities, oxidative stress, inflammation, tubular cell damage and tubulointerstitial fibrosis. Additionally, cystathionine γ-lyase (CGL) expression was decreased in the diabetic renal cortex. LPD reversed all of these alterations of the diabetic kidney; however, the LP+Met diet abrogated the effects of LPD in diabetic rats. Furthermore, LPD partially improved the diabetic state, while the LP+Met diet exacerbated it and was accompanied by changes in the LPD-induced expression of plasma fibroblast growth factor 21 (FGF21). Interpretation: LPD could exert a reno-protective effect through low-Met intake-mediated suppression of mTORC1 and restored autophagy, which is associated with a decrease in the accumulation of SAM due to reduced expression of Gnmt in diabetic kidneys. Funding Statement: This work was financially supported through a Grant-in-Aid for Scientific Research KAKENHI (C) (24591218), a grant for collaborative research with Kowa Hakko Kirin to MK. Declaration of Interests: Boehringer Ingelheim, Mitsubishi Tanabe Pharma, Kyowa Hakko Kirin, Taisho Toyama Pharmaceutical Co. and Ono Pharmaceutical Co. contributed to establishing the Division of Anticipatory Molecular Food Science and Technology. The authors declare that there is no duality of interest associated with this manuscript. Ethics Approval Statement: The Research Center for Animal Life Science of Kanazawa Medical University approved all experiments, and all experiments were performed in accordance with the relevant guidelines and regulations.

Diabetic nephropathy is the most common cause of end-stage renal disease. Therefore, novel therapies for the suppression of diabetic nephropathy must be developed. Rodent models are useful for elucidating the pathogenesis of diseases and testing novel therapies, and many type 1 and type 2 diabetic rodent models have been established for the study of diabetes and diabetic complications. Streptozotocin (STZ)-induced diabetic animals are widely used as a model of type 1 diabetes. Akita diabetic mice that have an Ins2+/C96Y mutation and OVE26 mice that overexpress calmodulin in pancreatic β-cells serve as a genetic model of type 1 diabetes. In addition, db/db mice, KK-Ay mice, Zucker diabetic fatty rats, Wistar fatty rats, Otsuka Long-Evans Tokushima Fatty rats and Goto-Kakizaki rats serve as rodent models of type 2 diabetes. An animal model of diabetic nephropathy should exhibit progressive albuminuria and a decrease in renal function, as well as the characteristic histological changes in the glomeruli and the tubulointerstitial lesions that are observed in cases of human diabetic nephropathy. A rodent model that strongly exhibits all these features of human diabetic nephropathy has not yet been developed. However, the currently available rodent models of diabetes can be useful in the study of diabetic nephropathy by increasing our understanding of the features of each diabetic rodent model. Furthermore, the genetic background and strain of each mouse model result in differences in susceptibility to diabetic nephropathy with albuminuria and the development of glomerular and tubulointerstitial lesions. Therefore, the validation of an animal model reproducing human diabetic nephropathy will significantly facilitate our understanding of the underlying genetic mechanisms that contribute to the development of diabetic nephropathy. In this review, we focus on rodent models of diabetes and discuss the utility and limitations of these models for the study of diabetic nephropathy.

Glucose-stimulated insulin secretion (GSIS) by pancreatic β cells is biphasic. However, the physiological significance of biphasic GSIS and its relationship to diabetes are not yet fully understood. This study demonstrated that impaired first-phase GSIS follows fasting, leading to increased blood glucose levels and brain glucose distribution in humans. Animal experiments to determine a possible network between the brain and β cells revealed that fasting-dependent hyperactivation of AMP-activated protein kinase in the hypothalamus inhibited first-phase GSIS by stimulating the α-adrenergic pancreatic nerve. Furthermore, abnormal excitability of this brain-β cell neural axis was involved in diabetes-related impairment of first-phase GSIS in diabetic animals. Finally, pancreatic denervation improved first-phase GSIS and glucose tolerance and ameliorated severe diabetes by preventing β cell loss in diabetic animals. These results indicate that impaired first-phase GSIS is critical for brain distribution of dietary glucose after fasting. Furthermore, β cells in individuals with diabetes mistakenly sense that they are under conditions that mimic prolonged fasting. The present study provides additional insight into both β cell physiology and the pathogenesis of β cell dysfunction in type 2 diabetes., Highlights • Fasting-induced hypothalamic AMPK activation inhibited first-phase GSIS by stimulating the α-adrenergic nerve. • The brain-pancreas neural axis was involved in β cell dysfunction and glucose intolerance in diabetes. • Pancreatic denervation improved first-phase GSIS, glucose tolerance and β cell survival in type 2 diabetic animals. Glucose-stimulated insulin secretion (GSIS) from pancreatic β cells is biphasic. Furthermore, first-phase GSIS is inhibited in type 2 diabetes. This study revealed that fasting reduced first-phase GSIS by signaling via the brain-pancreatic β cell neural axis, which is essential for maintaining glucose supply to the brain at re-feeding after fasting. Abnormal excitability of this neural axis was also associated with impaired first-phase GSIS in type 2 diabetes. Surgical pancreatic denervation improved diabetes in an animal study. The present data reveal that diabetic β cells exist under conditions that mimic starvation and provide a therapeutic potency of pancreatic denervation against diabetes., Graphical Abstract Image 2

The efficacy of a low-protein diet (LPD) on diabetic nephropathy is controversial. We aimed to investigate the renoprotective effects of an LPD and the underlying molecular mechanism in a rat model of type 2 diabetes and obesity.Diabetic male Wistar fatty (fa/fa) rats (WFRs) were treated with a standard diet (23.84% protein) or an LPD (5.77% protein) for 20 weeks from 24 weeks of age. We investigated the effect of the LPD on renal function, fibrosis, tubular cell damage, inflammation, mitochondrial morphology of proximal tubular cells (PTCs), apoptosis, autophagy and activation of mammalian target of rapamycin complex 1 (mTORC1).Kidney weight, albuminuria, excretion of urinary liver-type fatty acid binding protein, levels of plasma cystatin C and changes in renal histology, including fibrosis, tubular cell damage and inflammation, were aggravated in WFRs compared with non-diabetic Wistar lean rats (WLRs). Fragmented and swelling mitochondria accumulated in PTCs and apoptosis were enhanced in the kidney of WFRs. Immunohistochemical staining of p62 and p-S6 ribosomal protein (p-S6RP) in the tubular lesions of WFRs was increased compared with WLRs. The LPD intervention clearly ameliorated damage as shown by the assessment of renal function and histology, particularly tubulointerstitial damage in diabetic kidneys. Additionally, the 5.77% LPD, but not the 11.46% LPD, significantly suppressed p-S6RP levels and increased microtubule-associated protein light chain 3-II levels in the renal cortex. The LPD intervention partially decreased HbA1c levels in WFRs, and no differences in mean BP were observed among any of the groups.A very-low-protein diet improved advanced diabetic renal injuries, including tubulointerstitial damage, by restoring autophagy through the suppression of the mTORC1 pathway.

Kidney fibrosis is the final common pathway of progressive kidney diseases including diabetic nephropathy. Here, we report that the endogenous antifibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP), the substrate of angiotensin-converting enzyme (ACE), is an orally available peptide drug used to cure kidney fibrosis in diabetic mice. We utilized two mouse models of diabetic nephropathy, streptozotocin- (STZ-) induced type 1 diabetic CD-1 mice and type 2 diabetic nephropathy modeldb/dbmice. Intervention with the ACE inhibitor imidapril, oral AcSDKP, or imidapril + oral AcSDKP combination therapy increased urine AcSDKP levels. AcSDKP levels were significantly higher in the combination group compared to those of the other groups. AcSDKP oral administration, either AcSDKP alone or in addition to imidapril, ameliorated glomerulosclerosis and tubulointerstitial fibrosis. Plasma cystatin C levels were higher in both models, at euthanasia, and were restored by all the treatment groups. The levels of antifibrotic miRs, such as miR-29 or let-7, were suppressed in the kidneys of both models; all treatments, especially the combination of imidapril + oral AcSDKP, restored the antifibrotic miR levels to a normal value or even higher. AcSDKP may be an oral antifibrotic peptide drug that would be relevant to combating fibroproliferative kidney diseases such as diabetic nephropathy.

Diabetic nephropathy is a leading cause of end stage renal disease and its occurance is increasing worldwide. The most effective treatment strategy for the condition is intensive treatment to strictly control glycemia and blood pressure using renin-angiotensin system inhibitors. However, a fraction of patients still go on to reach end stage renal disease even under such intensive care. New therapeutic targets for diabetic nephropathy are, therefore, urgently needed. Autophagy is a major catabolic pathway by which mammalian cells degrade macromolecules and organelles to maintain intracellular homeostasis. The accumulation of damaged proteins and organelles is associated with the pathogenesis of diabetic nephropathy. Autophagy in the kidney is activated under some stress conditions, such as oxidative stress and hypoxia in proximal tubular cells, and occurs even under normal conditions in podocytes. These and other accumulating findings have led to a hypothesis that autophagy is involved in the pathogenesis of diabetic nephropathy. Here, we review recent findings underpinning this hypothesis and discuss the advantages of targeting autophagy for the treatment of diabetic nephropathy.

Free fatty acid-bound albumin (FFA-albumin)-related oxidative stress is involved in the pathogenesis of proximal tubular cell (PTC) damage and subsequent renal dysfunction in patients with refractory proteinuria. Nicotinamide adenine dinucleotide (NAD) metabolism has recently been focused on as a novel therapeutic target for several modern diseases, including diabetes. This study was designed to identify a novel molecule in NAD metabolism to protect PTCs from lipotoxicity-related oxidative stress. Among 19 candidate enzymes involved in mammalian NAD metabolism, the mRNA expression level of nicotinamide n-methyltransferase (NNMT) was significantly increased in both the kidneys of FFA-albumin-overloaded mice and cultured PTCs stimulated with palmitate-albumin. Knockdown of NNMT exacerbated palmitate-albumin-induced cell death in cultured PTCs, whereas overexpression of NNMT inhibited it. Intracellular concentration of 1-Methylnicotinamide (1-MNA), a metabolite of NNMT, increased and decreased in cultured NNMT-overexpressing and -knockdown PTCs, respectively. Treatment with 1-MNA inhibited palmitate-albumin-induced mitochondrial reactive oxygen species generation and cell death in cultured PTCs. Furthermore, oral administration of 1-MNA ameliorated oxidative stress, apoptosis, necrosis, inflammation, and fibrosis in the kidneys of FFA-albumin-overloaded mice. In conclusion, NNMT-derived 1-MNA can reduce lipotoxicity-mediated oxidative stress and cell damage in PTCs. Supplementation of 1-MNA may have potential as a new therapy in patients with refractory proteinuria.

Diabetic kidney disease is a leading cause of end stage kidney disease requiring renal transplantation or dialysis in not only Asian but also western countries. Recent data has also implicated that the prevalence of diabetic kidney disease was similar between 1988 and 2014 in cross-sectional studies of adults aged 20 years or older with diabetes mellitus participating in National Health and Nutrition Examination Surveys 1), although newly introduced treatments such as renin-angiotensin-aldosterone system inhibitors did affect the prevalence and clinical manifestations of diabetic kidney disease. Moreover, the prevalence of type 2 diabetes is still increasing strikingly all the world 2), thereby increasing the prevalence of diabetic kidney disease in future if we will not be able to discover new therapeutic strategies for high risk diabetic patients to develop and progress kidney disease. This article is protected by copyright. All rights reserved.

Acute kidney injury (AKI) is a syndrome which has a broad range of etiologic factors depending on different clinical settings. Because AKI has significant impacts on prognosis in any clinical settings, early detection and intervention are necessary to improve the outcomes of AKI patients. This clinical guideline for AKI was developed by a multidisciplinary approach with nephrology, intensive care medicine, blood purification, and pediatrics. Of note, clinical practice for AKI management which was widely performed in Japan was also evaluated with comprehensive literature search.

Kidney fibrosis is the final common pathway of progressive kidney disease including diabetic kidney disease (DKD). With the rapid increase of type 2 diabetes, DKD in type 2 diabetic patients has become a serious public health threat around the world. To establish novel therapy to combat DKD, human disease relevant animal model is essential. However, a type 2 diabetic mouse model with presenting progressive kidney fibrosis is not established yet. We have previously reported that kidneys of streptozotocin-induced diabetic CD-1 mice exhibited severe fibrosis compared to other backgrounds of mice associated with the suppression of anti-fibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline, fibroblast growth factor receptor 1, and their downstream target anti-fibrotic microRNAs (miRs) such as miR 29 and miR let-7. As the type 2 diabetic mice model, leptin receptor deficient mice in BKS background (BKSdb/db) are often utilized; the kidney fibrosis in BKSdb/db phenotype is minimum. Here we backcrossed db gene into potential fibrotic CD-1 background and found that male CD-1db/db exhibited severe kidney fibrosis when compared to male BKSdb/db. We have already performed the backcrosses for 11 generations. Body weight of CD-1db/db was comparable but drank up to 3 times more water compared to BKSdb/db. Blood glucose levels in CD-1db/db appeared to be higher when compared to BKSdb/db. When evaluated the per body weight ratio, kidney and heart weighed no significant differences between CD-1db/db and BKSdb/db. CD-1db/db exhibited severe tubulo-interstitial fibrosis, glomerulosclerosis, and tubular epithelial cells damage. Heart in CD-1db/db displayed fibrotic change as well. In conclusion, here we provide novel fibrotic type 2 DKD mouse model. CD-1db/db could be excellent model to perform experimental therapy for DKD. Disclosure Y. Mizunuma: None. K. Nitta: None. S. Takagi: None. K. Kanasaki: Advisory Panel; Self; Boehringer Ingelheim Pharmaceuticals, Inc.. Speaker's Bureau; Self; Eli Lilly and Company, Sanofi, Mitsubishi Tanabe Pharma Corporation. Research Support; Self; Boehringer Ingelheim GmbH. Speaker's Bureau; Self; Boehringer Ingelheim GmbH, Astellas, Astellas. Research Support; Self; Ono Pharmaceutical Co., Ltd., Mitsubishi Tanabe Pharma Corporation. D. Koya: None.

Diabetic kidney disease (DKD) is a major cause of end-stage renal disease worldwide. Although the detailed pathogenesis of DKD is not elucidated yet, aging is recognized as one of the risk factors for the development of end-stage renal failure due to chronic kidney disease including DKD. Therefore, investigation of aging-related mechanism would be a novel therapeutic target for DKD. NAD levels decrease during aging. Previous report demonstrated that expression and activity of the CD38 (NADase) increased with aging, and that CD38 is required for the age-related NAD decline and mitochondrial dysfunction via a pathway mediated at least in part by regulation of Sirt3 activity. However, the role of CD38 in the pathogenesis for DKD has never been investigated. In this study, we evaluated the role of CD38 on the mechanisms of renal mitochondrial oxidative stress which is related to changing Sirt3 activity, in Zucker Diabetic Fatty rats (ZDFRs) and in cultured HK2 cells under high-glucose condition. At 28 weeks of age, the ZDFRs exhibited elevated HbA1c levels, heavier kidney weight, increased urinary albumin, L-FABP and 8-OHdG excretion, histological tubulo-interstitial fibrosis and tubular cell damage, compared to nondiabetic Zucker Lean rats. Additionally, in renal mitochondria, the NAD+/NADH ratio was reduced, and acetylation levels of mitochondrial antioxidant enzymes, IDH2 and SOD2, which are regulated by Sirt3, were increased in ZDFRs. Similarly, in cultured HK2 cells under high-glucose condition, CD38 expression was increased, resulting in the reduction of NAD+/NADH ratio and Sirt3 activity, which increased in acetylation levels of IDH2 and SOD2. Administration of the CD38 inhibitor, apigenin, to ZDFRs and HK2 cells, restored the NAD+/NADH ratio, decreased the levels of IDH2 and SOD2 acetylation. Therefore, restoring Sirt3 activation by suppression of CD38 may be a novel therapeutic target for DKD. Disclosure Y. Ogura: None. M. Kitada: Research Support; Self; Astellas Pharma US, Inc., Kyowa Hakko Kirin Co., Ltd., Daiichi Sankyo Company, Limited, Ono Pharmaceutical Co., Ltd., Novartis Pharma K.K., Sumitomo Dainippon Pharma Co., Ltd., Boehringer Ingelheim GmbH, Taisho Pharmaceutical Co., Ltd., Mitsubishi Tanabe Pharma Corporation. I. Monno: None. A. Watanabe: None. D. Koya: None.