metallothionein and Diabetic-Nephropathies

Impaired zinc homeostasis is observed in diabetes mellitus (DM2) and its complications. Zinc has a specific role in pancreatic β-cells via insulin synthesis, storage, and secretion. Intracellular zinc homeostasis is tightly controlled by zinc transporters (ZnT and Zip families) and metallothioneins (MT) which modulate the uptake, storage, and distribution of zinc. Several investigations in animal models demonstrate the protective role of MT in DM2 and its cardiovascular or renal complications, while a copious literature shows that a common polymorphism (R325W) in ZnT8, which affects the protein's zinc transport activity, is associated with increased DM2 risk. Emerging studies highlight a role of other zinc transporters in β-cell function, suggesting that targeting them could make a possible contribution in managing the hyperglycemia in diabetic patients. This article summarizes the current findings concerning the role of zinc homeostasis in DM2 pathogenesis and development of diabetic cardiomyopathy and nephropathy and suggests novel therapeutic targets. © 2017 BioFactors, 43(6):770-784, 2017.

Topics: Animals; Biological Transport; Diabetes Mellitus, Type 2; Diabetic Cardiomyopathies; Diabetic Nephropathies; Dietary Supplements; Disease Models, Animal; Gene Expression Regulation; Homeostasis; Humans; Insulin; Insulin-Secreting Cells; Metallothionein; Zinc; Zinc Transporter 8

The present study was conducted to explore the toxic effects of lead (Pb) on the physiological responses of Japanese quails and to investigate the potential modulatory role of Yucca schidigera extract (YSE) against these effects. 360 mature Japanese quails (at 2 months of age) were used and the experiment was lasted for 8 weeks. The birds were divided into six equal groups as follow: control (basal diet, BD), BD+Pb (100 mg/kg diet), BD+YSE (100 mg/kg diet), BD+YSE (200 mg/kg diet), BD+Pb (100 mg/kg diet) +YSE (100 mg/kg diet) and BD+ Pb (100 mg/kg diet) + YSE (200 mg/kg diet). Pb induced a significant reduction in superoxide dismutase (SOD) and catalase (CAT) activities and reduced glutathione (GSH) level. While, increased protein carbonyl (PC) and malondialdehyde (MDA) content in tissues of exposed birds. Pb increased level of 8-hydroxy-2-deoxyguanosine (8-OHdG) and lactate dehydrogenase (LDH) activity in serum. YSE significantly reduced the Pb -induced oxidative stress in co-treated groups especially at 200 mg/kg diet. YSE could modulate the Pb -induced decreased urea, creatinine and beta-2 microglobulin (B2M) levels. YSE200 was found to be better than the YSE100 in decreasing levels of inflammatory markers including tumor necrosis factor (TNF-α), nitric oxide (NO), transforming growth factor-β1 (TGF-β1) and vascular endothelial growth factor (VEGF). Furthermore, YSE significantly regulates glucose homeostasis in co-exposed quails. Pb residues were found to be significantly higher in kidney and pancreas tissues of Pb group compared to other groups. YES decreased the expression of metallothionein-1 in the renal and pancreatic tissues, while elevated insulin expression in the pancreatic cells by immunostaining in co-exposed groups. In conclusion, the present results conclusively demonstrate the potential modulatory effect of YSE against the Pb-induced toxic effects in different organs of Japanese quails.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Antioxidants; Biomarkers; Blood Glucose; Catalase; Coturnix; Deoxyguanosine; Diabetic Nephropathies; Glomerular Filtration Rate; Glucose Transporter Type 2; Glutathione; Homeostasis; Insulin; Kidney; Lead; Malondialdehyde; Metallothionein; Oxidative Stress; Pancreas; Plant Extracts; Superoxide Dismutase; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A; Yucca

We previously demonstrated that OVE transgenic diabetic mice are susceptible to chronic complications of diabetic nephropathy (DN) including substantial oxidative damage to the renal glomerular filtration barrier (GFB). Importantly, the damage was mitigated significantly by overexpression of the powerful antioxidant, metallothionein (MT) in podocytes. To test our hypothesis that GFB damage in OVE mice is the result of endothelial oxidative insult, a new JTMT transgenic mouse was designed in which MT overexpression was targeted specifically to endothelial cells. At 60 days of age, JTMT mice were crossed with age-matched OVE diabetic mice to produce bi-transgenic OVE-JTMT diabetic progeny that carried the endothelial targeted JTMT transgene. Renal tissues from the OVE-JTMT progeny were examined by unbiased TEM stereometry for possible GFB damage and other alterations from chronic complications of DN. In 150 day-old OVE-JTMT mice, blood glucose and HbA1c were indistinguishable from age-matched OVE mice. However, endothelial-specific MT overexpression in OVE-JTMT mice mitigated several DN complications including significantly increased non-fenestrated glomerular endothelial area, and elimination of glomerular basement membrane thickening. Significant renoprotection was also observed outside of endothelial cells, including reduced podocyte effacement, and increased podocyte and total glomerular cell densities. Moreover, when compared to OVE diabetic animals, OVE-JTMT mice showed significant mitigation of nephromegaly, glomerular hypertrophy, increased mesangial cell numbers and increased total glomerular cell numbers. These results confirm the importance of oxidative stress to glomerular damage in DN, and show the central role of endothelial cell injury to the pathogenesis of chronic complications of diabetes. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:560-576, 2017. © 2016 Wiley Periodicals, Inc.

Topics: Animals; Blood Glucose; Diabetic Nephropathies; Disease Models, Animal; Endothelial Cells; Kidney Glomerulus; Metallothionein; Mice; Mice, Transgenic; Oxidative Stress; Podocytes

Hyperglycemia, a major metabolic disturbance present in diabetes, promotes oxidative stress. Activation of antioxidant defense is an important mechanism to prevent cell damage. Levels of heavy metals and their binding proteins can contribute to oxidative stress. Antiradical capacity and levels of metallothionein (MT), metals (zinc and copper), and selected antioxidants (bilirubin, cysteine, and glutathione) were determined in 70 type 2 diabetes mellitus (T2DM) subjects and 80 healthy subjects of Caucasian origin. Single nucleotide polymorphism (rs28366003) in MT gene was detected. Antiradical capacity, conjugated bilirubin, and copper were significantly increased in diabetics, whereas MT and glutathione were decreased. Genotype AA of rs28366003 was associated with higher zinc levels in the diabetic group. The studied parameters were not influenced by renal function. This is the first study comprehensively investigating differences in MT and metals relevant to oxidative stress in T2DM. Ascertained differences indicate increased oxidative stress in T2DM accompanied by abnormalities in non-enzymatic antioxidant defense systems.

Topics: Aged; Aged, 80 and over; Biomarkers; Case-Control Studies; Copper; Czech Republic; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Female; Genetic Association Studies; Humans; Kidney; Male; Metallothionein; Middle Aged; Oxidative Stress; Polymorphism, Single Nucleotide; Renal Insufficiency, Chronic; Severity of Illness Index; Zinc

Diabetic nephropathy (DN) is thought to be partially due to the injury of renal cells and the renal micro-environment by free radicals. Free radial scavenging agents that inhibit free radical damage may well prevent the development of underlying conditions such as mesangial expansion (by inhibiting extracellular matrix expression) in these patients.. Using techniques for intra-cellular delivery of peptides, we made metallothionein (MT) and superoxide dismutase (SOD), potent endogenous antioxidants, readily transducible into cell membrane and tested their protective effect against the development of DN in OLETF rats. Herein, we study antioxidant peptides for their ability to prevent oxidative damage to primary rat mesangial cells (MCs), which are important constituents of renal glomeruli.. Intraperitoneal administration of these antioxidants resulted in delivery to the kidney and decreased ROS and the expression of downstream signals in renal cells and postponed the usual progression to DN. In in vitro experiments, MT and SOD were efficiently transferred to MCs, and the increased removal of ROS by MT and SOD was proportional to the degree of scavenging enzymes delivered. MT and SOD decreased three major oxidative injuries (hyperglycemia, AGE and ROS exposure) and also injuries directly mediated by angiotensin II in MCs while changing downstream signal transduction.. The protective effects of MT and SOD for the progression of DN in experimental animals may be associated with the scavenging of ROS by MT and SOD and correlated changes in signal transduction downstream. Concomitant administration of these antioxidant peptides may prove to be a new approach for the prevention and therapy of DN.

Topics: Animals; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Free Radical Scavengers; Male; Mesangial Cells; Metallothionein; Peptides; Rats; Rats, Long-Evans; Superoxide Dismutase

Sulforaphane (SFN) prevents diabetic nephropathy (DN) in type 1 diabetes via up-regulation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2). However, it has not been addressed whether SFN also prevents DN from type 2 diabetes or which Nrf2 downstream gene(s) play(s) the key role in SFN renal protection. Here we investigated whether Nrf2 is required for SFN protection against type 2 diabetes-induced DN and whether metallothionein (MT) is an Nrf2 downstream antioxidant using Nrf2 knockout (Nrf2-null) mice. In addition, MT knockout mice were used to further verify if MT is indispensable for SFN protection against DN. Diabetes-increased albuminuria, renal fibrosis, and inflammation were significantly prevented by SFN, and Nrf2 and MT expression was increased. However, SFN renal protection was completely lost in Nrf2-null diabetic mice, confirming the pivotal role of Nrf2 in SFN protection from type 2 diabetes-induced DN. Moreover, SFN failed to up-regulate MT in the absence of Nrf2, suggesting that MT is an Nrf2 downstream antioxidant. MT deletion resulted in a partial, but significant attenuation of SFN renal protection from type 2 diabetes, demonstrating a partial requirement for MT for SFN renal protection. Therefore, the present study demonstrates for the first time that as an Nrf2 downstream antioxidant, MT plays an important, though partial, role in mediating SFN renal protection from type 2 diabetes.

Topics: Animals; Anticarcinogenic Agents; Blotting, Western; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Immunoenzyme Techniques; Isothiocyanates; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-E2-Related Factor 2; Oxidative Stress; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sulfoxides

Oxidative stress and inflammation play important roles in diabetic complications, including diabetic nephropathy. Metallothionein (MT) is induced in proximal tubular epithelial cells as an antioxidant in the diabetic kidney; however, the role of MT in renal function remains unclear. We therefore investigated whether MT deficiency accelerates diabetic nephropathy through oxidative stress and inflammation. Diabetes was induced by streptozotocin injection in MT-deficient (MT(-/-)) and MT(+/+) mice. Urinary albumin excretion, histological changes, markers for reactive oxygen species (ROS), and kidney inflammation were measured. Murine proximal tubular epithelial (mProx24) cells were used to further elucidate the role of MT under high-glucose conditions. Parameters of diabetic nephropathy and markers of ROS and inflammation were accelerated in diabetic MT(-/-) mice compared with diabetic MT(+/+) mice, despite equivalent levels of hyperglycemia. MT deficiency accelerated interstitial fibrosis and macrophage infiltration into the interstitium in the diabetic kidney. Electron microscopy revealed abnormal mitochondrial morphology in proximal tubular epithelial cells in diabetic MT(-/-) mice. In vitro studies demonstrated that knockdown of MT by small interfering RNA enhanced mitochondrial ROS generation and inflammation-related gene expression in mProx24 cells cultured under high-glucose conditions. The results of this study suggest that MT may play a key role in protecting the kidney against high glucose-induced ROS and subsequent inflammation in diabetic nephropathy.

Topics: Animals; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Gene Expression Regulation; Kidney; Macrophages; Male; Metallothionein; Mice; Mice, Knockout; Reactive Oxygen Species

Human epidemiological and animal studies have shown the beneficial effect of zinc supplementation on mitigating diabetic nephropathy. However, the mechanism by which zinc protects the kidney from diabetes remains unknown. Here we demonstrate the therapeutic effects of zinc on diabetes-induced renal pathological and functional changes. These abnormalities were found in both transgenic OVE26 and Akt2-KO diabetic mouse models, accompanied by significant changes in glucose-metabolism-related regulators. The changes included significantly decreased phosphorylation of Akt and GSK-3β, increased phosphorylation of renal glycogen synthase, decreased expression of hexokinase II and PGC-1α, and increased expression of the Akt negative regulators PTEN, PTP1B, and TRB3. All of these were significantly prevented by zinc treatment for 3 months. Furthermore, zinc-stimulated changes in glucose metabolism mediated by Akt were actually found to be metallothionein dependent, but not Akt2 dependent. These results suggest that the therapeutic effects of zinc in diabetic nephropathy are mediated, in part, by the preservation of glucose-metabolism-related pathways via the prevention of diabetes-induced upregulation of Akt negative regulators. Given that zinc deficiency is very common in diabetics, this finding implies that regularly monitoring zinc levels in diabetic patients, as well as supplementing if low, is important in mitigating the development of diabetic nephropathy.

Topics: Animals; Diabetic Nephropathies; Disease Models, Animal; Glucose; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Metallothionein; Mice; Mice, Inbred NOD; Mice, Transgenic; Proto-Oncogene Proteins c-akt; Signal Transduction; Zinc

We previously demonstrated that cellular and extracellular components of the blood-urine barrier in renal glomeruli are susceptible to damage in OVE transgenic mice, a valuable model of human diabetic nephropathy that expresses profound albuminuria.. To test our hypothesis that glomerular filtration barrier damage in OVE mice may be the result of oxidative insult to podocytes, 150-day-old bi-transgenic OVENmt diabetic mice that overexpress the antioxidant metallothionein specifically in podocytes were examined by enzyme-linked immunosorbent assay for albuminuria mitigation and by unbiased transmission electron microscopy (TEM) stereometry for protection from chronic structural diabetic complications.. Although blood glucose and HbA(1c) levels were indistinguishable in OVE and OVENmt animals, albuminuria was significantly reduced (average >7-fold) in OVENmt mice through 8 months of age. Interestingly, the Nmt transgene provided significant glomerular protection against diabetic nephropathic complications outside of the podocyte. Glomerular filtration barrier damage was reduced in OVENmt mice, including significantly increased area occupied by endothelial luminal fenestrations (~13%), significantly reduced glomerular basement membrane (GBM) thickening (~17%) and significantly less podocyte effacement (~18%). In addition, OVENmt mice exhibited significantly reduced glomerular volume (~50%), fewer glomerular endothelial cells (~33%), fewer mesangial cells (~57%) and fewer total glomerular cells (~40%).. These results provide evidence of oxidative damage to podocytes induces primary diabetic nephropathic features including severe and sustained albuminuria, specific glomerular filtration barrier damage and alterations in glomerular endothelial and mesangial cell number. Importantly, these diabetic complications are significantly mitigated by podocyte targeted metallothionein overexpression.

Topics: Albuminuria; Animals; Diabetic Nephropathies; Glomerular Basement Membrane; Glomerular Filtration Barrier; Kidney Glomerulus; Metallothionein; Mice; Mice, Transgenic; Microscopy, Electron, Transmission; Podocytes

Zinc is an element that under physiological conditions preferentially binds to and is a potent inducer of metallothionein under physiological conditions. The present study was conducted to explore whether zinc supplementation morphologically and biochemically protects against diabetic nephropathy through modulation of kidney metallothionein induction and oxidative stress in streptozotocin-induced diabetic rats. Thirty-two Wistar albino male rats were equally divided into four groups. The first group was used as untreated controls and the second group was supplemented with 30 mg/kg/day zinc as zinc sulfate. The third group was treated with streptozotocin to induce diabetes and the fourth group was treated with streptozotocin and supplemented with zinc as described for group 2. The blood glucose and micro-albuminuria levels, body and kidney weights were measured during the 42-day experimental period. At the end of the experiment, the kidneys were removed from all animals from the four groups. Diabetes resulted in degenerative kidney morphological changes. The metallothionein immunoreactivity level was lower and the kidney lipid peroxidation levels were higher in the diabetes group than in the controls. The metallothionein immunoreactivity levels were higher in the tubules of the zinc-supplemented diabetic rats as compared to the non-supplemented diabetic group. The zinc and metallothionein concentrations in kidney tissue were higher in the supplemented diabetic group compared to the non-supplemented diabetes group. The activity of glutathione peroxidase did not change in any of the four groups. In conclusion, the present study shows that zinc has a protective effect against diabetic damage of kidney tissue through stimulation of metallothionein synthesis and regulation of the oxidative stress.

Topics: Animals; Antioxidants; Diabetes Mellitus, Type 1; Diabetic Nephropathies; Dietary Supplements; Glutathione Peroxidase; Hypoglycemic Agents; Kidney; Kidney Tubules; Lipid Peroxidation; Male; Metallothionein; Oxidative Stress; Rats; Rats, Wistar; Streptozocin; Tissue Distribution; Up-Regulation; Zinc; Zinc Sulfate

Metallothionein (MT) is an intracellular metal-binding, cysteine-rich protein, and is a potent antioxidant that protects cells and tissues from oxidative stress. Although the major isoforms MT-1 and -2 (MT-1/-2) are highly inducible in many tissues, the distribution and role of MT-1/-2 in diabetic nephropathy are poorly understood. In this study, diabetes was induced in adult male rats by streptozotocin, and renal tissues were stained with antibodies for MT-1/-2. MT-1/-2 expression was also evaluated in mProx24 cells, a mouse renal proximal tubular epithelial cell line, stimulated with high glucose medium and pretreated with the antioxidant vitamin E. MT-1/-2 expression was gradually and dramatically increased, mainly in the proximal tubular epithelial cells and to a lesser extent in the podocytes in diabetic rats, but was hardly observed in control rats. MT-1/-2 expression was also increased by high glucose stimulation in mProx24 cells. Because the induction of MT was suppressed by pretreatment with vitamin E, the expression of MT-1/-2 is induced, at least in part, by high glucose-induced oxidative stress. These observations suggest that MT-1/-2 is induced in renal proximal tubular epithelial cells as an antioxidant to protect the kidney from oxidative stress, and may offer a novel therapeutic target against diabetic nephropathy.

Topics: Animals; Antioxidants; Cell Line; Diabetic Nephropathies; Epithelial Cells; Gene Expression; Glucose; Kidney Tubules, Proximal; Male; Metallothionein; Mice; Oxidative Stress; Rats; Rats, Sprague-Dawley; Vitamin E

We have demonstrated that Zn supplementation mediated up-regulation of cardiac metallothionein (MT) as a potent antioxidant prevented the development of diabetic cardiomyopathy. The present study was undertaken to test whether induction of renal MT synthesis by Zn supplementation protects the kidney from diabetes-induced damage. Streptozotocin-induced diabetic rats were treated with and without Zn supplementation at 5 mg/kg in drinking water for 3 months. Diabetic renal damage was detected by examining renal pathological alterations and 24-h urinary protein levels. Three-month Zn supplementation immediately after the onset of diabetes, partially but significantly, prevented the kidney from diabetes-induced increases in 24-h urinary proteins and pathological alterations. Diabetes-induced renal oxidative damage, inflammation and up-regulated expression of profibrosis mediator connective tissue growth factor (CTGF) were also markedly attenuated by Zn supplementation, along with significant increases in Zn levels concomitant with MT expression in renal tubular cells. Direct exposure of renal tubular (HK11) cells to high levels of glucose (HG) induced CTGF up-regulation predominantly through ERK (extracellular signal-regulated kinase)1/2-dependent, and partially through p38 mitogen-activated protein kinase (MAPK)-dependent pathways. Pretreatment of HK11 cells with Zn or cadmium induced MT expression and also significantly suppressed HG-induced CTGF expression. These results provide the first evidence for Zn supplementation to attenuate diabetes-induced renal pathological changes, likely through prevention of hyperglycemia-induced CTGF expression by Zn-induced MT in renal tubular cells.

Topics: Animals; Blood Glucose; Cell Line; Connective Tissue Growth Factor; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Gene Expression Regulation; Glucose; Humans; Kidney; Kidney Tubules, Proximal; Lipid Peroxidation; Male; Metallothionein; Organ Specificity; Plasminogen Activator Inhibitor 1; Protein Isoforms; Proteinuria; Random Allocation; Rats; Rats, Wistar; RNA, Messenger; Superoxide Dismutase; Zinc

Podocytes are critical components of the selective filtration barrier of the glomerulus and are susceptible to oxidative damage. For investigation of the role of oxidative stress and podocyte damage in diabetic nephropathy, transgenic mice that overexpress the antioxidant protein metallothionein (MT) specifically in podocytes (Nmt mice) were produced. MT expression was increased six- and 18-fold in glomeruli of two independent lines of Nmt mice, and podocyte-specific overexpression was confirmed. Glomerular morphology and urinary albumin excretion were normal in Nmt mice. OVE26 transgenic mice, a previously reported model of diabetic nephropathy, were crossed with Nmt mice to determine whether an antioxidant transgene targeted to podocytes could reduce diabetic nephropathy. Double-transgenic OVE26Nmt mice developed diabetes similar to OVE26 mice, but MT overexpression reduced podocyte damage, indicated by more podocytes, less glomerular cell death, and higher density of podocyte foot processes. In addition, expansion of glomerular and mesangial volume were significantly less in OVE26Nmt mice compared with OVE26 mice. Four-month-old OVE26Nmt mice had a 70 to 90% reduction in 24-h albumin excretion, but this protection does not seem to be permanent. These results provide evidence for the role of oxidative damage to the podocyte in diabetic mice and show that protection of the podocyte can reduce or delay primary features of diabetic nephropathy.

Topics: Albuminuria; Animals; Antioxidants; Apoptosis; Diabetic Nephropathies; Gene Expression; Humans; Kidney Glomerulus; Metallothionein; Mice; Mice, Transgenic; Oxidative Stress; Podocytes; Recombinant Proteins

Topics: Animals; Apoptosis; Diabetic Nephropathies; Gene Expression; Humans; Metallothionein; Mice; Oxidative Stress; Podocytes; Reactive Oxygen Species

It has been reported that diabetes may increase the risk of cadmium-induced kidney damage. The presence of metallothionein antibody (MT-Ab) increased the susceptibility for tubular damage among cadmium workers. This study focused on the relationships between levels of MT-Ab, urinary cadmium, and kidney function in a Chinese type 2 diabetic population.. A cross-sectional study was performed on 229 type 2 diabetic patients (92 men and 137 women) who were recruited from two community centers in one district of Shanghai City in China. Information was obtained from interviews, health records, and blood and urine samples.. Levels of the tubular biomarker beta2-microglobulin increased significantly when the levels of MT-Ab and urinary cadmium were elevated in male and female subjects; in contrast, the levels of urinary albumin, a glomerular biomarker, did not display such a pattern. After adjusting for potential confounding covariates, logistic regression showed that the odds ratios (ORs) of tubular dysfunction increased upon 1) increasing the MT-Ab concentration from a low to high level (OR 5.56 [95% CI 2.25-13.73]) and 2) increasing the level of urinary cadmium from <1 to >or=1 microg/g creatinine (3.34 [1.17-9.53]); the OR of patients currently smoking was 3.51 (1.14-10.80) relative to that of those who had never smoked.. This study proves that the presence of MT-Ab can potentiate tubular dysfunction among diabetic subjects and that patients with high MT-Ab levels are more prone to development of tubular damage.

Topics: Aged; Albuminuria; Alcohol Drinking; Antibodies; Cadmium; China; Cross-Sectional Studies; Diabetes Mellitus, Type 2; Diabetic Nephropathies; Female; Humans; Male; Metallothionein; Middle Aged; Smoking