Overexpression of acetyl CoA carboxylase ß exacerbates podocyte injury in the kidney of streptozotocin-induced diabetic mice.

A single nucleotide polymorphism (SNP) within the acetyl CoA carboxylase (ACC) ß gene (ACACB), rs2268388, has been shown to be associated with susceptibility to development of proteinuria in patients with type 2 diabetes. To investigate the biological roles of ACCß in the pathogenesis of diabetic nephropathy, we examined the effects of overexpression of ACACB using podocyte-specific ACACB-transgenic mice or ACACB-overexpressing murine podocytes. Podocyte-specific ACACB-transgenic mice or littermate mice were treated with streptozotocin (STZ) to induce diabetes, and 12 weeks after induction of diabetes, we examined the expression of podocyte markers to evaluate the degree of podocyte injury in these mice. We also examined the effects of ACCß on podocyte injury in ACACB- or LacZ-overexpressing murine podocytes. Podocyte-specific ACACB overexpression did not cause visible podocyte injury in non-diabetic mice. In STZ-induced diabetic mice, ACACB-transgenic mice showed a significant increase in urinary albumin excretion, accompanied by decreased synaptopodin expression and podocin mislocalization in podocytes, compared with wild-type mice. In cultured murine podocytes, overexpression of ACACB significantly decreased synaptopodin expression and reorganized stress fibers under high glucose conditions, but not in normal glucose conditions. The decrease of synaptopodin expression and reorganized stress fibers observed in ACACB overexpressing cells cultured under high glucose conditions was reversed by a treatment of 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR), activator of AMP-activated protein kinase (AMPK). The excess of ACCß might contribute to exacerbation of podocyte injury in the kidney of an animal model for diabetes mellitus, and the AMPK/ACCß pathway may be a novel therapeutic target for the prevention of diabetes-related podocyte injury.

Focal Segmental Glomerular Sclerosis Ameliorated by Long-term Hemodialysis Therapy with Low-density Lipoprotein Apheresis.

We report a case involving a 43-year-old Japanese woman with steroid-resistant focal segmental glomerular sclerosis (FSGS) and severe renal dysfunction, which was ameliorated by low-density lipoprotein apheresis (LDL-A). She had been treated with steroid therapy, but had experienced anuria for over 10 weeks and required hemodialysis. She was then treated with LDL-A, which resulted in improved urinary protein excretion and renal function. Her renal function recovered after 97 days of hemodialysis therapy. This case suggests that LDL-A may represent an effective rescue treatment in patients with FSGS and long-term anuria.

Oral glucose-stimulated serum C-peptide predicts successful switching from insulin therapy to liraglutide monotherapy in Japanese patients with type 2 diabetes and renal impairment.

AIMS/INTRODUCTION: In Japan, liraglutide was recently approved for patients with type 2 diabetes. To our knowledge, there are no markers predicting successful switching from insulin therapy to liraglutide monotherapy in Japanese patients with type 2 diabetes and renal impairment. We therefore assessed clinical characteristics predicting successful switching. MATERIALS AND METHODS: We analyzed 21 patients with type 2 diabetes and estimated glomerular filtration rates <60 mL/min/1.73 m(2) receiving long-term insulin in Shiga University of Medical Science Hospital, Otsu, Shiga, Japan. Their ß-cell function was assessed by measuring urinary C-peptide and C-peptide immunoreactivity (CPR) index, along with glucagon loading and oral glucose tolerance tests. Blood glucose concentration and blood pressure were measured daily before and after switching from insulin to liraglutide, and glycated hemoglobin (HbA1c; National Glycohemoglobin Standardization Program) was assessed 12 weeks after switching to liraglutide. RESULTS: Baseline HbA1c was significantly lower in successfully switched than in unsuccessfully switched patients. CPR index, urinary C-peptide concentration and 6-min post-glucagon increment in CPR (ΔCPR) did not differ significantly in the two groups. ΔCPR 120 min after 75 g oral glucose was significantly higher in successfully than unsuccessfully switched patients. Mean blood glucose concentrations before breakfast, after breakfast, before lunch and after dinner were significantly lower in successfully switched patients. HbA1c did not change significantly in either group. CONCLUSIONS: Measurement of oral glucose-stimulated ΔCPR120 min is recommended when considering switching Japanese type 2 diabetes patients with renal impairment from insulin to liraglutide monotherapy.

Predictive properties of plasma amino acid profile for cardiovascular disease in patients with type 2 diabetes.

Prevention of cardiovascular disease (CVD) is an important therapeutic object of diabetes care. This study assessed whether an index based on plasma free amino acid (PFAA) profiles could predict the onset of CVD in diabetic patients. The baseline concentrations of 31 PFAAs were measured with high-performance liquid chromatography-electrospray ionization-mass spectrometry in 385 Japanese patients with type 2 diabetes registered in 2001 for our prospective observational follow-up study. During 10 years of follow-up, 63 patients developed cardiovascular composite endpoints (myocardial infarction, angina pectoris, worsening of heart failure and stroke). Using the PFAA profiles and clinical information, an index (CVD-AI) consisting of six amino acids to predict the onset of any endpoints was retrospectively constructed. CVD-AI levels were significantly higher in patients who did than did not develop CVD. The area under the receiver-operator characteristic curve of CVD-AI (0.72 [95% confidence interval (CI): 0.64-0.79]) showed equal or slightly better discriminatory capacity than urinary albumin excretion rate (0.69 [95% CI: 0.62-0.77]) on predicting endpoints. A multivariate Cox proportional hazards regression analysis showed that the high level of CVD-AI was identified as an independent risk factor for CVD (adjusted hazard ratio: 2.86 [95% CI: 1.57-5.19]). This predictive effect of CVD-AI was observed even in patients with normoalbuminuria, as well as those with albuminuria. In conclusion, these results suggest that CVD-AI based on PFAA profiles is useful for identifying diabetic patients at risk for CVD regardless of the degree of albuminuria, or for improving the discriminative capability by combining it with albuminuria.

Fatty acids are novel nutrient factors to regulate mTORC1 lysosomal localization and apoptosis in podocytes.

Podocyte apoptosis is a potent mechanism of proteinuria in diabetic nephropathy. More detailed mechanistic insight into podocyte apoptosis is needed to better understand the pathogenesis of diabetic nephropathy. An elevated level of serum free fatty acid (FFA), as well as hyperglycemia, is a clinical characteristic in diabetes, although its causal role in podocyte apoptosis remains unclear. This study examined the effect of three types of FFAs, saturated, monounsaturated and polyunsaturated FFAs, on podocyte apoptosis. Palmitate, a saturated FFA, induced endoplasmic reticulum (ER) stress-dependent apoptosis in podocytes. Oleate, a monounsaturated FFA, and eicosapentaenoic acid (EPA), an ω-3 polyunsaturated FFA did not induce apoptosis; rather, they antagonized palmitate-induced apoptosis. Palmitate activated mammalian target of rapamycin (mTOR) complex 1 (mTORC1), a nutrient-sensing kinase regulating a wide range of cell biology. Furthermore, inhibition of mTORC1 activity by rapamycin or siRNA for Raptor, a component of mTORC1, ameliorated palmitate-induced ER stress and apoptosis in podocytes. Activity of mTORC1 is regulated by upstream kinases and Rag/Ragulator-dependent recruitment of mTOR onto lysosomal membranes. Palmitate activated mTORC1 by enhancing recruitment of mTOR onto lysosomal membranes, which was inhibited by co-incubation with oleate or EPA. Inhibition of mTOR translocation onto lysosomes by transfection with dominant-negative forms of Rag ameliorated palmitate-induced apoptosis. This study suggests that saturated and unsaturated FFAs have opposite effects on podocyte apoptosis by regulating mTORC1 activity via its translocation onto lysosomal membranes, and the results provide a better understanding of the pathogenesis in diabetic nephropathy and a novel role of mTORC1 in cell apoptosis.

Glycated albumin predicts the risk of mortality in type 2 diabetic patients on hemodialysis: evaluation of a target level for improving survival.

Glycated albumin (GA) is considered a more reliable marker than glycated hemoglobin (HbA1c) for monitoring glycemic control, particularly in diabetic hemodialysis patients. We investigated the associations of GA, HbA1c, and random serum glucose levels with survival, and evaluated possible targets for improving survival in diabetic hemodialysis patients. In this prospective, longitudinal, observational study, we enrolled 90 diabetic hemodialysis patients across six dialysis centers in Japan. The median duration of follow-up was 36.0 months (mean follow-up, 29.8 months; range, 3-36 months). There were 11 deaths during the observation period. GA was a significant predictor for mortality (hazard ratio, 1.143 per 1% increase in GA; 95% confidence interval, 1.011-1.292; P = 0.033), whereas HbA1c and random glucose levels were not predictors for mortality. Receiver operating characteristics curve analysis showed that the cutoff value of GA for predicting the risk of mortality was 25%. In the Kaplan-Meier analysis, the cumulative survival rate was significantly greater in patients with GA ≤ 25% than in patients with GA >25%. GA predicted the risk of all-cause and cardiovascular mortality in diabetic hemodialysis patients. Our results suggest that GA ≤ 25% is an appropriate target for improving survival in diabetic hemodialysis patients.

Safety and efficacy of skin patches containing loxoprofen sodium in diabetic patients with overt nephropathy.

BACKGROUND: Because oral nonsteroidal anti-inflammatory drugs (NSAIDs) have adverse effects on kidney function, patients with kidney diseases are administered these drugs as transdermal patches. Little is known about the effects of NSAID patches on renal function. We therefore assessed the effects of topical loxoprofen sodium on kidney function in type 2 diabetic patients with overt nephropathy. METHODS: Twenty patients with type 2 diabetes and overt proteinuria and with knee and/or low back pain were treated with skin patches containing 100 mg loxoprofen on the knee or back for 24 h per day for 5 consecutive days. The degree of pain was assessed using a visual analogue scale (VAS). Blood and 24-h urine samples were obtained at baseline and at the end of the study. Glomerular filtration rate (GFR) was estimated from serum creatinine and cystatin C concentrations. RESULTS: The 20 patients consisted of 11 males and 9 females, of mean age 61.6 ± 13.9 years. Loxoprofen-containing patches significantly reduced VAS pain without affecting blood pressure, GFR or urinary prostaglandin E2 concentration. Serum concentrations of loxoprofen and its active trans-OH metabolite did not correlate with GFR. CONCLUSIONS: Loxoprofen-containing patches do not affect renal function in type 2 diabetic patients with overt nephropathy over a short-term period. Long-term studies are needed to clarify the safety of loxoprofen-containing patches in patients with chronic kidney diseases.

Obesity-mediated autophagy insufficiency exacerbates proteinuria-induced tubulointerstitial lesions.

Obesity is an independent risk factor for renal dysfunction in patients with CKDs, including diabetic nephropathy, but the mechanism underlying this connection remains unclear. Autophagy is an intracellular degradation system that maintains intracellular homeostasis by removing damaged proteins and organelles, and autophagy insufficiency is associated with the pathogenesis of obesity-related diseases. We therefore examined the role of autophagy in obesity-mediated exacerbation of proteinuria-induced proximal tubular epithelial cell damage in mice and in human renal biopsy specimens. In nonobese mice, overt proteinuria, induced by intraperitoneal free fatty acid-albumin overload, led to mild tubular damage and apoptosis, and activated autophagy in proximal tubules reabsorbing urinary albumin. In contrast, diet-induced obesity suppressed proteinuria-induced autophagy and exacerbated proteinuria-induced tubular cell damage. Proximal tubule-specific autophagy-deficient mice, resulting from an Atg5 gene deletion, subjected to intraperitoneal free fatty acid-albumin overload developed severe proteinuria-induced tubular damage, suggesting that proteinuria-induced autophagy is renoprotective. Mammalian target of rapamycin (mTOR), a potent suppressor of autophagy, was activated in proximal tubules of obese mice, and treatment with an mTOR inhibitor ameliorated obesity-mediated autophagy insufficiency. Furthermore, both mTOR hyperactivation and autophagy suppression were observed in tubular cells of specimens obtained from obese patients with proteinuria. Thus, in addition to enhancing the understanding of obesity-related cell vulnerability in the kidneys, these results suggest that restoring the renoprotective action of autophagy in proximal tubules may improve renal outcomes in obese patients.

Altered unfolded protein response is implicated in the age-related exacerbation of proteinuria-induced proximal tubular cell damage.

Aging is a dominant risk factor for end-stage renal disease. We analyzed the mechanism involved in age-related exacerbation of proteinuria-induced proximal tubular cell (PTC) damage by focusing on endoplasmic reticulum-related unfolded protein response (UPR). After equal-degree induction of proteinuria in 24-month-old (aged) and 3-month-old (young) mice by intraperitoneal free fatty acid-bound albumin overload, tubulointerstitial lesions were more severe in aged than in young mice. In aged PTCs, proteinuria-induced cell-adaptive UPR resulting from induction of the molecular chaperone BiP was significantly suppressed, whereas proapoptotic UPR with CHOP overexpression was enhanced. Treatment with the exogenous molecular chaperone tauroursodeoxycholic acid (TUDCA) ameliorated proteinuria-induced tubulointerstitial lesions and PTC apoptosis in aged mice. Among the three UPR branches, alterations in the inositol-requiring 1α (IRE1α) pathway, but not the activating transcription factor 6 or PERK pathway, were associated with impaired BiP induction in aged kidneys. Moreover, siRNA-mediated suppression of BiP and IRE1α exacerbated free fatty acid-bound albumin-induced apoptosis in cultured PTCs, whereas siRNA-mediated CHOP suppression ameliorated apoptosis. Finally, proteinuria-induced BiP induction in PTCs was diminished in kidney specimens from elderly patients. These results indicate that maladaptive UPRs are involved in proteinuria-induced tubulointerstitial lesions exacerbation in aged kidneys, and that supplementation of chaperones may be used to treat elderly patients with persistent proteinuria. These results should improve understanding of cell vulnerability in aged kidneys.

Predictive effects of urinary liver-type fatty acid-binding protein for deteriorating renal function and incidence of cardiovascular disease in type 2 diabetic patients without advanced nephropathy.

OBJECTIVE: To improve prognosis, it is important to predict the incidence of renal failure and cardiovascular disease in type 2 diabetic patients before the progression to advanced nephropathy. We investigated the predictive effects of urinary liver-type fatty acid-binding protein (L-FABP), which is associated with renal tubulointerstitial damage, in renal and cardiovascular prognosis. RESEARCH DESIGN AND METHODS: Japanese type 2 diabetic patients (n = 618) with serum creatinine ≤1.0 mg/dL and without overt proteinuria were enrolled between 1996 and 2000 and followed up until 2011. Baseline urinary L-FABP was measured with an enzyme-linked immunosorbent assay. The primary end points were renal and cardiovascular composites (hemodialysis, myocardial infarction, angina pectoris, stroke, cerebral hemorrhage, and peripheral vascular disease). The secondary renal outcomes were the incidence of a 50% decline in estimated glomerular filtration rate (eGFR), progression to an eGFR <30 mL/min/1.73 m(2), and the annual decline rate in eGFR. RESULTS: During a 12-year median follow-up, 103 primary end points occurred. The incidence rate of the primary end point increased in a stepwise manner with increases in urinary L-FABP. In Cox proportional hazards analysis, the adjusted hazard ratio in patients with the highest tertile of urinary L-FBAP was 1.93 (95% CI 1.13-3.29). This relationship was observed even when analyzed separately in normoalbuminuria and microalbuminuria. Patients with the highest tertile of urinary L-FABP also demonstrated a higher incidence of the secondary renal outcomes. CONCLUSIONS: Our results indicate that urinary L-FABP may be a predictive marker for renal and cardiovascular prognosis in type 2 diabetic patients without advanced nephropathy.

High sodium intake is associated with masked hypertension in Japanese patients with type 2 diabetes and treated hypertension.

BACKGROUND: Knowledge regarding the association between dietary sodium intake and the incidence of masked hypertension is limited. METHODS: A total of 193 Japanese type 2 diabetic outpatients who had been treated with antihypertensive agents and with office blood pressures <140/90 mm Hg were recruited. Masked hypertension was defined as having office blood pressure <140/90 mm Hg and 24-h mean ambulatory blood pressure ≥130/80 mm Hg. The dietary sodium intake was estimated by measuring the 24-h urinary sodium excretion. RESULTS: Masked hypertension was found in 128 (66.3%) patients. An age- and sex-adjusted univariate logistic regression analysis showed that urinary albumin excretion, renin-angiotensin system inhibitor use, office systolic blood pressure, and amount of dietary sodium intake were significantly associated with masked hypertension. A multivariate logistic regression analysis also identified an older age, renin-angiotensin system inhibitor use, an office elevated systolic blood pressure, and high dietary sodium intake to be independently associated with masked hypertension. When compared with those who consumed a low salt diet (sodium <120 mEq/day), the odds ratio for the risk of exhibiting masked hypertension in patients who consumed a medium salt diet (sodium 120 to <200 mEq/day) or a high salt diet (sodium ≥200 mEq/day) were 5.3 (P < 0.001) and 12.6 (P < 0.001), respectively. CONCLUSIONS: Masked hypertension is a common feature in type 2 diabetic patients being treated for hypertension. The observed association with sodium intake raised the hypothesis that excessive sodium intake may play a part in the genesis of masked hypertension in these patients.

Fructose induces tubulointerstitial injury in the kidney of mice.

Fructose induces several kinds of human metabolic disorders; however, information regarding fructose-induced kidney injury is still limited. This study examined fructose-induced kidney injury in mice and clarified the differential susceptibility of three mouse strains: C57Bl/6J, CBA/JN and DBA/2N. In this study all mice were fed with an equal calorie count for sixteen weeks to remove the influence of total energy intake from metabolic effects by fructose-feeding. Only DBA/2N mice, but not C57Bl/6J and CBA/JN mice, fed with fructose displayed tubulointerstitial fibrosis localized on the outer cortex of the kidney together with the increase of mRNA expression of Kim1 and Ngal in the absence of distinct glomerular lesions and albuminuria - decidedly different from diabetic nephropathy. In time-course study of DBA/2N mice fed with fructose diet, the inflammation and fibrosis in the outer cortex of the kidney were enhancing after eight weeks, in parallel with the accumulation of oxidative stress. This progression of renal damage in DBA/2N mice was accompanied with increasing mRNA expression of GLUT5. These results suggest that the responsiveness of GLUT5 expression to fructose at the kidney is one of pivotal roles for the progression of fructose-induced kidney injury.

Role of angiotensin II-mediated AMPK inactivation on obesity-related salt-sensitive hypertension.

Salt-sensitive hypertension is a characteristic of the metabolic syndrome. Given the links to cardiovascular events, the mechanisms underlying sodium metabolism may represent an important therapeutic target for this disorder. Angiotensin II (AII) is a key peptide underlying sodium retention. However, 5'AMP-activated protein kinase (AMPK) has also been reported to participate in the regulation of ion transport. In this study we examined the relationship between AII and AMPK on the development of hypertension in two salt-sensitive mouse models. In the first model, the mice were maintained on a high-fat diet (HFD) for 12 weeks, in order to develop features similar to the metabolic syndrome, including salt-sensitive hypertension. HFD-induced obese mice showed elevated systolic blood pressure and lower sodium excretion in response to salt loading, along with an increase in AII contents and inactivation of AMPK in the kidney, which were significantly improved by the treatment of an angiotensin II antagonist, losartan, for 2 weeks. To clarify the effects of AII, a second group of mice was infused with AII via an osmotic pump, which led to higher systolic blood pressure, and decreases in urinary sodium excretion and the expression of AMPK, in a manner similar to those observed in the HFD mice. However, treatment with an AMPK activator, metformin, improved the changes induced by the AII, suggesting that AII induced sodium retention works by acting on AMPK activity. Finally, we evaluated the changes in salt-sensitivity by performing 2-week salt loading experiments with or without metformin. AII infusion elevated blood pressure by salt loading but metformin prevented it. These findings indicate that AII suppresses AMPK activity in the kidney, leading to sodium retention and enhanced salt-sensitivity, and that AMPK activation may represent a new therapeutic target for obesity-related salt-sensitive hypertension.

The effects of blood pressure control levels on the renoprotection of type 2 diabetic patients without overt proteinuria.

BACKGROUND: There is little evidence regarding the target blood pressure level in patients with type 2 diabetes mellitus without overt proteinuria. METHODS AND RESULTS: We followed 608 Japanese patients with type 2 diabetes without apparent cardiovascular disease and overt proteinuria who underwent cerebral magnetic resonance imaging for a mean of 7.5 years. The patients were categorized according to their mean systolic blood pressure during the follow-up period (strict: <130 mm Hg, moderate: ≥130 and <140 mm Hg, poor: ≥ 140 mm Hg). The risks for the primary composite outcome of death or end-stage renal disease were not different among the three groups. The renal risk of the doubling of serum creatinine for the poor group was significantly higher than those in other groups. In addition, among the patients without silent cerebral infarction (SCI), the renal risk was significantly lower in the strict group than in the moderate group. Further, in both the SCI and non-SCI groups, strict blood pressure control slowed the progression of albuminuria. CONCLUSIONS: In nonproteinuric diabetic patients without SCI, strict blood pressure control was associated with improved renal outcomes. There may be different effects of intensive blood pressure control on the renoprotection of diabetic patients according to their complications.

Arterial stiffness and renal impairment in non-proteinuric type 2 diabetic patients.

UNLABELLED: Aims/Introduction: Although increases in urinary protein excretion generally precede a decline in the glomerular filtration rate, non-proteinuric renal impairment is common in patients with diabetes. In the present study, we examined the relationship between indices of arterial stiffness and renal function in type 2 diabetic patients without proteinuria. METHODS: Blood sampling, 24-h urine collection, brachial-ankle pulse wave velocity, and 24-h ambulatory blood pressure monitoring were performed in type 2 diabetic patients without overt proteinuria. The ambulatory arterial stiffness index was calculated as (1 - the regression slope of diastolic/systolic ambulatory blood pressure). Estimated glomerular filtration rate (eGFR)was calculated using the simplified prediction equation proposed by the Japanese Society of Nephrology. RESULTS: Of 213 non-proteinuric patients with type 2 diabetes, 60 (28.2%) had a reduced eGFR (<60 mL/min per 1.73 m(2)). Although the urinary albumin excretion rate was significantly correlated with the eGFR, 34 of 152 patients with normoalbuminuria (22.4%) had a reduced eGFR. The eGFR was significantly and negatively correlated with the ambulatory arterial stiffness index and brachial-ankle pulse wave velocity, but not with 24-h pulse pressure. Multivariate analysis revealed that increased age and increased urinary albumin excretion were independently associated with decreased eGFR. In addition, the ambulatory arterial stiffness index, but not brachial-ankle pulse wave velocity, were found to be independently and significantly associated with eGFR. CONCLUSIONS: Ambulatory arterial stiffness index is a marker for increased risk of renal failure in non-proteinuric patients with type 2 diabetes. (J Diabetes Invest, doi: 10.1111/j.2040-1124.2011.00146.x, 2012).

Efficacy and tolerability of vildagliptin in type 2 diabetic patients on hemodialysis.

Anti-diabetic agent-related hypoglycemia is a serious complication in type 2 diabetic patients on hemodialysis. Therefore, we assessed the efficacy and tolerability of 24 weeks of monotherapy with vildagliptin, a dipeptidyl peptidase four inhibitor, which is a new class of antidiabetic agent. This open-label, single-arm clinical trial was performed on 26 patients on hemodialysis. The primary assessments were changes in postprandial glucose level and glycated albumin (GA). During the study, three patients dropped out, and data from 23 patients were analyzed. Significant reductions were seen in postprandial glucose (-2.60 ± 3.80 mmol/L, P < 0.001) and GA (-2.59 ± 2.33%, P < 0.001) levels. No serious drug-related adverse events were observed. Vildagliptin monotherapy can be recommended for glycemic control in type 2 diabetic patients on hemodialysis. This trial was registered with the University Hospital Medical Information Network (no. UMIN000003661). (J Diabetes Invest, doi: 10.1111/j.2040-1124.2011.00169.x, 2011).

GW501516, a PPARδ agonist, ameliorates tubulointerstitial inflammation in proteinuric kidney disease via inhibition of TAK1-NFκB pathway in mice.

Peroxisome proliferator-activated receptors (PPARs) are a nuclear receptor family of ligand-inducible transcription factors, which have three different isoforms: PPARα, δ and γ. It has been demonstrated that PPARα and γ agonists have renoprotective effects in proteinuric kidney diseases; however, the role of PPARδ agonists in kidney diseases remains unclear. Thus, we examined the renoprotective effect of GW501516, a PPARδ agonist, in a protein-overload mouse nephropathy model and identified its molecular mechanism. Mice fed with a control diet or GW501516-containing diet were intraperitoneally injected with free fatty acid (FFA)-bound albumin or PBS(-). In the control group, protein overload caused tubular damages, macrophage infiltration and increased mRNA expression of MCP-1 and TNFα. These effects were prevented by GW501516 treatment. In proteinuric kidney diseases, excess exposure of proximal tubular cells to albumin, FFA bound to albumin or cytokines such as TNFα is detrimental. In vitro studies using cultured proximal tubular cells showed that GW501516 attenuated both TNFα- and FFA (palmitate)-induced, but not albumin-induced, MCP-1 expression via direct inhibition of the TGF-ß activated kinase 1 (TAK1)-NFκB pathway, a common downstream signaling pathway to TNFα receptor and toll-like receptor-4. In conclusion, we demonstrate that GW501516 has an anti-inflammatory effect in renal tubular cells and may serve as a therapeutic candidate to attenuate tubulointerstitial lesions in proteinuric kidney diseases.

SIRT3 attenuates palmitate-induced ROS production and inflammation in proximal tubular cells.

Free fatty acid (FFA)-mediated renal lipotoxicity is associated with the progression of tubulointerstitial inflammation in proteinuric kidney disease. SIRT3 is an antiaging molecule regulated by calorie restriction and mitochondria-localized NAD(+)-dependent deacetylase. In this study, we investigated whether SIRT3 reversed renal lipotoxicity-mediated ROS and inflammation. In the kidney of the FFA-bound BSA-overloaded mouse, which is a well-established experimental model of FFA-associated tubulointerstitial inflammation, mRNA expression of SIRT3 was significantly decreased and negatively correlated with mRNA expression of an inflammatory cytokine, monocyte chemoattractant protein-1 (MCP-1). In cultured proximal tubular (mProx) cells, the saturated FFA palmitate stimulated ROS accumulation and expression of MCP-1. These effects were ameliorated by retrovirus-mediated overexpression of SIRT3, whereas they were exacerbated by either overexpression of a dominant-negative form of SIRT3(N87A) lacking deacetylase activity or knockdown of SIRT3 by siRNA transfection. Furthermore, we showed that SIRT3 positively regulated both mitochondrial oxidative capacity and antioxidant gene expression, thereby reducing ROS accumulation in mProx cells, which suggests a mechanism that underlies SIRT3-mediated reversal of palmitate-induced inflammation. In conclusion, these results highlight a new role for SIRT3 in lipotoxicity/ROS-related inflammation, reveal a new molecular mechanism underlying calorie restriction-mediated antioxidant and anti-inflammatory effects, and could aid in the design of new therapies for the prevention of tubulointerstitial lesions in proteinuric kidney disease.