Gender differences in vascular reactivity to endothelin-1 (1-31) in mesenteric arteries from diabetic mice

Endothelin-1 (1-31) [ET-1 (1-31)], a novel member of the ET family, comprises 31 amino acids and is derived from the selective hydrolysis of big ET-1 by chymase. Although ET-1 (1-31) reportedly exerts biological effects by direct or indirect [via its conversion to ET-1 (1-21)] mechanisms, it is unclear whether in diabetes the vascular effects of ET-1 (1-31) display gender differences. We investigated this question by exposing mesenteric artery rings to ET-1 (1-31), using arteries from mice in the early or chronic phase of diabetes. In the early stage of diabetes, the ET-1 (1-31)-induced contraction was similar between age- and sex-matched control and streptozotocin (STZ)-induced diabetic mice. In the chronic stage of diabetes, the ET-1 (1-31)-induced contraction was enhanced in diabetic female mice, but not in diabetic male mice (vs. both age-matched control and early-stage diabetic mice). This enhancement was largely prevented by Y27632 (Rho kinase inhibitor), PD98059 [inhibitor of extracellular signal related kinases 1 and 2 (ERK1/2)], or SP600125 [C-jun terminal kinase (JNK) inhibitor]. These data indicate that the ET-1 (1-31)-induced vasoconstriction in the mesenteric artery may be specifically enhanced in established diabetic female mice, and that this enhancement may be due to alterations in the activities of Rho/Rho kinase or mitogen-activated protein kinase.     

Enhanced endothelin-1 response and receptor expression in small mesenteric arteries of insulin-resistant rats

Hyperinsulinemia, a primary feature of insulin resistance, is associated with increased endothelin-1 (ET-1) activity. This study determined the vascular response to ET-1 and receptor binding characteristics in small mesenteric arteries of insulin-resistant (IR) rats. Rats were randomized to control (C) (n = 32) or IR (n = 32) groups. The response to ET-1 was assessed (in vitro) in arteries with (Endo+) and without (Endo-) endothelium. In addition, arteries (Endo+) were pretreated with the ET(B) antagonist A-192621 or the ET(A) antagonist A-127722. Finally, binding characteristics of [(125)I]ET-1 were determined. Results showed that in Endo+ arteries the maximal relaxation (E(max)) to ET-1 was similar between C and IR groups; however, the concentration at 50% of maximum relaxation (EC(50)) was decreased in IR arteries. In Endo- arteries, the E(max) to ET-1 was enhanced in both groups. Pretreatment with A-192621 enhanced the E(max) and EC(50) to ET-1 in both groups. In contrast, A-127722 inhibited the ET-1 response in all arteries in a concentration-dependent manner; however, a greater ET-1 response was seen at each concentration in IR arteries. Maximal binding of [(125)I]ET-1 was increased in IR versus C arteries although the dissociation constant values were similar. In conclusion, we found the vasoconstrictor response to ET-1 is enhanced in IR arteries due to an enhanced expression of ET receptors and underlying endothelial dysfunction.     

Cilostazol improves endothelium-derived hyperpolarizing factor-type relaxation in mesenteric arteries from diabetic rats

We previously reported that in mesenteric arteries from streptozotocin (STZ)-induced diabetic rats that 1) endothelium-derived hyperpolarizing factor (EDHF)-type relaxation is impaired, possibly due to a reduced action of cAMP via increased phosphodiesterase 3 (PDE3) activity (Matsumoto T, Kobayashi T, and Kamata K. Am J Physiol Heart Circ Physiol 285: H283-H291, 2003) and that 2) PKA activity is decreased (Matsumoto T, Wakabayashi K, Kobayashi T, and Kamata K. Am J Physiol Heart Circ Physiol 287: H1064-H1071, 2004). Here we investigated whether chronic treatment with cilostazol, a PDE3 inhibitor, improves EDHF-type relaxation in mesenteric arteries isolated from STZ rats. We found that in such arteries 1) cilostazol treatment (2 wk) improved ACh-, A-23187-, and cyclopiazonic acid-induced EDHF-type relaxations; 2) the ACh-induced cAMP accumulation was transient and sustained in arteries from cilostazol-treated STZ rats; 3) the EDHF-type relaxation was significantly decreased by a PKA inhibitor in the cilostazol-treated group, but not in the cilostazol-untreated group; 4) cilostazol treatment improved both the relaxations induced by cAMP analogs and the PKA activity level; and 5) PKA catalytic subunit (Cat-alpha) protein was significantly decreased, but the regulatory subunit RII-beta was increased (and the latter effect was significantly decreased by cilostazol treatment). These results strongly suggest that cilostazol improves EDHF-type relaxations in STZ rats via an increase in cAMP and PKA signaling.     

Alterations in EDHF-type relaxation and phosphodiesterase activity in mesenteric arteries from diabetic rats

In isolated superior mesenteric artery rings from age-matched control rats and streptozotocin (STZ)-induced diabetic rats, we investigated the role of cAMP in endothelium-derived hyperpolarizing factor (EDHF)-type relaxation. The ACh-induced EDHF-type relaxation was significantly weaker in STZ-induced diabetic rats than in control rats, and in both groups of rats it was attenuated by 18alpha-glycyrrhetinic acid (18alpha-GA), an inhibitor of gap junctions, and enhanced by IBMX, a cAMP-phosphodiesterase (PDE) inhibitor. These enhanced EDHF-type responses were very similar in magnitude between diabetic and age-matched control rats. The EDHF-type relaxation was enhanced by cilostamide, a PDE3-selective inhibitor, but not by Ro 20-1724, a PDE4-selective inhibitor. The expression levels of the mRNAs and proteins for two cAMP PDEs (PDE3A, PDE3B) were significantly increased in STZ-induced diabetic rats, but those for PDE4D were not. We conclude that the impairment of EDHF-type relaxations in STZ-induced diabetic rats may be attributed to a reduction in the action of cAMP via increased PDE activity.     

Proteomic analysis of glomeruli from streptozotocin-induced diabetic rats

The kidney glomeruli are the sites of plasma filtration and production of primary urine. However, they are also the locus of kidney diseases, which progress to chronic renal failure. Glomeruli are a major target of injury in diabetic nephropathy (DN). The mechanisms by which glomerular filtration are regulated are poorly understood, and proteomic investigations of isolated glomeruli on the progressive development of DN in animal models have not been determined. To understand the molecular mechanism leading to DN, especially the glomerular injury mechanism, the differences in the glomerular proteomes of streptozotocin (STZ)-induced- and non-diabetic rats at six and 24 weeks were analyzed via two-dimensional electrophoresis (2-DE). To identify the progressive stages of DN, body weight, blood glucose, and proteinuria were measured periodically, and pathological changes were evaluated by periodic acid-Schiff staining. Magnetic beads were used to isolate glomeruli from kidneys and the glomerular proteomes of non-diabetic and STZ-induced diabetic rats were analyzed by 2-DE and nano-LC-ESI-MS/MS. Glutathione peroxidase 3, peroxiredoxin 2, and histone H2A were down-regulated, and annexin A3 was up-regulated, in the STZ-induced group compared with the controls. Glutathione peroxidase 3 and annexin A3, which might help elucidate the mechanism of DN, were verified by Western blotting. These proteins could potentially provide insight into the mechanism of glomerular injury in DN.     

Effect of chronic endothelin blockade on PKC isoform distribution in mesenteric arteries from diabetic rats

Rigid plaques containing protein particles in plasma membrane build on the apical surface of the mammalian urothelium. We have previously shown that dietary fats modified the fatty acid profile as well as the fluorescence anisotropy of rat urothelial plasma membranes. In this study, we have further examined the proportion of phosphatidylcholine, phosphatidylethanolamine, cerebrosides, sulfatides and cholesterol in detergent resistant (DRM) and soluble (DSM) plasma membrane fractions as well as the properties of the particles. Four groups of weaned rats were fed for 12 weeks on a commercial diet (control), or on a formula containing 5% (w/w) of corn oil, fish oil or olein. The control DRM behaved as a distinctive domain since it was enriched in cholesterol and glycosphingolipids. DSM showed higher levels of phosphatidylcholine and phosphatidylethanolamine with respect to DRM. On the other hand, the lipid distributions were affected by the diets. Homogeneous lipid distributions between DSM and DRM were found in olein membranes, suggesting a decreased potential formation of lipid domains. In addition, properties of the uroplakins were altered by dietary treatments. Thus, uroplakins (UP) Ia, Ib, II and III observed by SDS-PAGE, were in lower proportions (mainly olein) than in controls. Moreover, a higher proportion of UPIII was cross-linked to UPIII and UPIb in olein treatment than in control. Meanwhile, only cross-linking to UPIII or UPIb was altered in corn and fish diets, respectively. These results suggest a role of the lipids in the establishment of the uroplakin interactions. Thus, specific dietary fats may have important functional implications. (Mol Cell Biochem 271: 69–75, 2005)     

Mechanisms of endothelin-1-induced contraction in pulmonary arteries from chronically hypoxic rats

Endothelin-1 (ET-1), a potent vasoconstrictor, is believed to contribute to the pathogenesis of hypoxic pulmonary hypertension. Previously we demonstrated that contraction induced by ET-1 in intrapulmonary arteries (IPA) from chronically hypoxic (CH) rats occurred independently of changes in intracellular Ca2+ concentration ([Ca2+]i), suggesting that ET-1 increased Ca2+ sensitivity. The mechanisms underlying this effect are unclear but could involve the activation of myosin light chain kinase, Rho kinase, PKC, or tyrosine kinases (TKs), including those from the Src family. In this study, we examined the effect of pharmacological inhibitors of these kinases on maximum tension generated by IPA from CH rats (10% O2 for 21 days) in response to ET-1. Experiments were conducted in the presence of nifedipine, an L-type Ca2+ channel blocker, to isolate the component of contraction that occurred without a change in [Ca2+]i. The mean change in tension caused by ET-1 (10(-8) M) expressed as a percent of the maximum response to KCl was 184.0+/-39.0%. This response was markedly inhibited by the Rho kinase inhibitors Y-27632 and HA-1077 and the TK inhibitors genistein, tyrphostin A23, and PP2. In contrast, staurosporine and GF-109203X, inhibitors of PKC, had no significant inhibitory effect on the tension generated in response to ET-1. We conclude that the component of ET-1-induced contraction that occurs without a change in [Ca2+]i in IPA from CH rats requires activation of Rho kinase and TKs, but not PKC.     

Aldosterone increases RAMP1 expression in mesenteric arteries from spontaneously hypertensive rats

OBJECTIVE: We analysed the effect of aldosterone on calcitonin gene-related peptide (CGRP) mediated vasodilation in noradrenaline precontracted endothelium denuded mesenteric arteries segments from Wistar Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) and the effect of aldosterone on calcitonin receptor-like receptor (CL receptor) and receptor activity modifying protein 1 (RAMP1) expression in endothelium-denuded mesenteric arteries from SHR rats. RESULTS: CGRP 0.1 nM-0.1 microM induced a concentration-dependent relaxation that was enhanced by aldosterone 1 microM in SHR only. Incubation with RU 486 10 microM significantly reduced the enhancement of CGRP-relaxation produced by aldosterone in SHR. CL receptor expression was not modified in either strain, while RAMP1 expression was enhanced in SHR by aldosterone 1 microM 120 min and 0.1 microM 120 min. This up-regulation of RAMP1 was prevented by RU 486 10 microM. CONCLUSIONS: Aldosterone, through glucocorticoid receptor activation, increases the vasodilatory effect of CGRP in SHR mesenteric arteries, which seems to be mediated by increased RAMP1 expression.     

Mechanisms underlying increased release of endothelin-1 from aorta in diabetic rats

To clarify the mechanism underlying increased endothelin-1 release in diabetic rats, we examined its release from thoracic aortas obtained from streptozotocin-induced diabetic rats. The methoxamine-induced contraction was significantly inhibited by BQ-123 plus BQ-788 (specific antagonists for ET(A) and ET(B) receptors) in diabetic, but not control rats. Preincubation with phosphoramidon also inhibited the methoxamine-induced contraction in diabetic but not control rats. The expression of prepro endothelin-1 mRNA was significantly enhanced in aortas from streptozotocin-induced diabetic rats. These results suggest that the increases in the basal and alpha-agonist-induced release of endothelin-1 in the diabetic state may be due to an overexpression of the mRNA for prepro endothelin-1.     

Polymorphonuclear neutrophil dysfunctions in streptozotocin-induced type 1 diabetic rats

Since conflicting results have been reported on non-specific immune response in type 1 diabetes, this study evaluates polymorphonuclear neutrophil (PMN) functions in the infection free Long Evan diabetic rats (type 1) by using tests that include: polarization assay, phagocytosis of baker\'s yeasts (Saccharomyces cerevisiae) and nitroblue tetrazolium (NBT) dye reduction. Polarization assay showed that neutrophils from diabetic rats were significantly activated at the basal level compared to those from the controls (p < 0.001). After PMN activation with N-formylmethionyl-leucyl-phenylalanine (FMLP), control neutrophils were found to be more polarized than those of the diabetic neutrophils and the highest proportions of polarization were found to be 67 % and 57 % at 10(-7) M FMLP, respectively. In the resting state, neutrophils from the diabetic rats reduced significantly more NBT dye than that of the controls (p < 0.001). The percentages of phagocytosis of opsonized yeast cells by the neutrophils from control and diabetic rats were 87 % and 61 %, respectively and the difference was statistically significant (p < 0.001). Evaluation of the phagocytic efficiency of PMNs revealed that control neutrophils could phagocytose 381 +/- 17 whereas those from the diabetic rats phagocytosed 282 +/- 16 yeast cells, and the efficiency of phagocytosis varied significantly (p < 0.001). Further, both the percentages of phagocytosis and the efficiency of phagocytosis by the diabetic neutrophils were inversely related with the levels of their corresponding plasma glucose (p = 0.02; r = -0.498 and p < 0.05; r = -0.43, respectively), which indicated that increased plasma glucose reduced the phagocytic ability of neutrophils. Such relationship was not observed with the control neutrophils. These data clearly indicate that PMN functions are altered in the streptozotocin (STZ)-induced diabetic rats, and hyperglycemia may be the cause for the impairment of their functions leading to many infectious episodes.     

Impaired arachidonic acid-mediated dilation of small mesenteric arteries in Zucker diabetic fatty rats

Arachidonic acid (AA) is a precursor of important vasoactive metabolites, but the role of AA-mediated vasodilation in Type 2 diabetes is not known. Using Zucker diabetic fatty (ZDF) rats, we examined the effects of AA in small mesenteric arteries preconstricted with endothelin. In ZDF rat mesenteric arteries, 1 microM AA produced only one-third the amount of dilation as in vessels from lean control animals. In lean control rats, the effect of AA was significantly and predominantly inhibited by the lipoxygenase inhibitors baicalein and cinnamyl-3,4-dihydroxy-cyanocinnamate (CDC). However, baicalein and CDC had no effect on AA-mediated dilation in ZDF rat mesenteric arteries. The major [3H]AA metabolite produced by isolated mesenteric arteries in both lean and ZDF rats was 12-hydroxyeicosatetraenoic acid (12-HETE), but the amount of [3H]12-HETE produced by ZDF rat vessels was only 36% of that of control vessels. In addition, 12-HETE produced similar amounts of dilation in lean and ZDF rat mesenteric arteries. Immunoblot analysis showed an 81% reduction in 12-lipoxygenase protein in ZDF rat mesenteric arteries. Immunofluorescence labeling showed strong nitrotyrosine signals in ZDF rat mesenteric arteries that colocalized with 12-lipoxygenase in endothelium, and 12-lipoxygenase coprecipitation with anti-nitrotyrosine antibodies was enhanced in ZDF rat vessels. We conclude that AA-mediated relaxation in ZDF rat small mesenteric arteries is impaired due to reduced 12-lipoxygenase protein and activity. Increased oxidative stress and nitration of 12-lipoxygenase may underlie the impairment of AA-mediated relaxation in small mesenteric arteries of diabetic rats.     

Elevated levels of endothelin-1 and prostaglandin E2 and their effect on nitric oxide generation in placental tissue from neonatal streptozotocin-induced diabetic rats

Endothelin-1 (ET-1), nitric oxide (NO) and prostaglandin E(2) (PGE(2)) are regulators of feto-placental hemodynamics. In this study we explore the inter-regulatory pathways that modulate the levels of these vasoactive agents in control and neonatal streptozotocin-induced (n-stz) diabetic rat placenta. ET-1 levels are increased in diabetic placenta when compared to controls (P<0.001), and are strongly reduced by an NO synthase inhibitor (P<0.001). PGE(2) production is increased in diabetic placenta when compared to controls (P<0.01), but these levels are not modulated by ET-1. NO levels, similar in control and in diabetic placenta, are not influenced by PGE(2), but they are negatively modulated by ET-1 in both control (P<0.05) and diabetic (P<0.01) placenta. We conclude that rat placental ET-1 inhibits NO levels but does not modify PGE(2) concentrations. The elevated levels of ET-1 and PGE(2) in diabetic placenta, potent vasoconstrictors of placental vasculature, are probably related to the induction of placental insufficiency and fetal hypoxia in this pathology.     

Functional changes in adenylyl cyclases and associated decreases in relaxation responses in mesenteric arteries from diabetic rats

To assess the functional change in adenylyl cyclases (AC) associated with the diabetic state, we investigated AC-mediated relaxations and cAMP production in mesenteric arteries from rats with streptozotocin (STZ)-induced diabetes. The relaxations induced by the water-soluble forskolin (FSK) analog NKH477, which is a putative AC5 activator, but not by the beta-adrenoceptor agonist isoproterenol (Iso) and the AC activator FSK, were reduced in intact diabetic mesenteric artery. In diabetic rats, however, Iso-, FSK-, and NKH477-induced relaxations were attenuated in the presence of inhibitors of nitric oxide synthase and cyclooxygenase. To exclude the influence of phosphodiesterase (PDE), we also examined the relaxations induced by several AC activators in the presence of 3-isobutyl-1-methylxanthine (IBMX; a PDE inhibitor). Under these conditions, the relaxation induced by Iso was greatly impaired in STZ-diabetic rats. This Iso-induced relaxation was significantly attenuated by pretreatment with SQ-22536, an AC inhibitor, in mesenteric rings from age-matched controls but not in those from STZ-diabetic rats. Under the same conditions, the relaxations induced by FSK or NKH477 were impaired in STZ-diabetic rats. Neither FSK- nor A-23187 (a Ca2+ ionophore)-induced cAMP production was significantly different between diabetics and controls. However, cAMP production induced by Iso or NKH477 was significantly impaired in diabetic mesenteric arteries. Expression of mRNAs and proteins for AC5/6 was lower in diabetic mesenteric arteries than in controls. These results suggest that AC-mediated relaxation is impaired in the STZ-diabetic rat mesenteric artery, perhaps reflecting a reduction in AC5/6 activity.     

Ovarian dysfunction in streptozotocin-induced diabetic rats

The effect of streptozotocin diabetes on some ovarian functions in adult rats was examined. Diabetic diestrus animals showed reduced ovary weight and lower circulating levels of progesterone. Scatchard plots of binding data derived from ovarian particulate fractions of normal and streptozotocin diabetic rats revealed the presence of one class of binding sites with high affinity for 125I-hCG. The apparent association constant of the hCG receptors of diabetic ovaries was comparable to that of normal gonads. However, a marked decrease (42%) in the number of hCG binding sites was found in diabetic animals. With isolated luteal cells similar results were obtained, and the administration of insulin to streptozotocin diabetic rats restored to normality the number of hCG binding sites. The maximal response of progesterone production by luteal cells from control ovaries was obtained with 10(-10) M hCG. A 100-fold higher concentration of hCG was required for the maximum stimulation of cAMP synthesis. The cAMP response of cells from diabetic rats was significantly higher than that of control cells. However, luteal cells from diabetic rats showed some loss of sensitivity in the synthesis of progesterone during incubation with hCG. Most of the alterations seen in diabetic female rats could be restored with insulin therapy, indicating that insulin plays an important role in the regulation and maintenance of normal reproductive functions. It is suggested that the diminution of the LH receptor population causes the disruption of normal luteal cell function. This fact could be responsible for some of the reproductive alterations in the diabetic female rat.     

Interleukin-2 enhances the release of endothelin-1 from the rat mesenteric artery

We measured the ET-1 concentration in plasma and in the perfusate of the mesenteric arteries of rats treated with a therapeutic dose of IL-2 for 7 days (100000 U/Kg, iv.). The plasma ET-1 concentration in rats given IL-2 was 14.2 +/- 3.2 pg/ml which was significantly greater than that in the controls (2.5 +/- 0.4 pg/ml, P less than 0.05). The mesenteric arteries also released a significantly greater amount of ET-1 (29.5 +/- 1.6 pg/h) than that in controls (16.8 +/- 2.3 pg/h, P less than 0.01). The arterial blood pressure was significantly lower after IL-2 treatment than the pre-dosing level (P less than 0.05). It is concluded that IL-2 induces ET-1 release from the vascular wall, possibly as a result of reversible endothelial dysfunction caused by IL-2.     

Comparison of the liver glycogen synthase from normal and streptozotocin-induced diabetic rats

Glycogen synthase in the liver extracts of short-term (3 days) streptozotocin-induced diabetic rats is poorly activated by the endogenous synthase phosphatase as well as phosphatase(s) from the liver extracts of normal animals. However, synthase in the liver extracts of diabetic rats is readily activated by the 35,000 M_r rabbit liver protein phosphatase (H. Brandt, F. L. Capulong, and E. Y. C. Lee (J. Biol. Chem.250, 8038–8044 (1975)). The purified synthases from normal and diabetic animals respond differently after incubations with three different phosphatases. Both normal and diabetic synthase are activated by the 35,000 M_r protein phosphatase; however, the total activity of diabetic, but not the normal, synthase is significantly increased. Normal, but not the diabetic, synthase is activated by a synthase phosphatase from normal rats; this activation is accompanied by an increase in total synthase activity. Incubation of the diabetic synthase with calf intestinal alkaline phosphatase results in a reduction of the total synthase activity, whereas synthase activity of the normal is not significantly affected. The reduction in total activity of the diabetic synthase by treatment with alkaline phosphatase was prevented by prior dephosphorylation with 35,000 M_r rabbit liver protein phosphatase. In addition to their differences in responses to different phosphatases, the normal and diabetic synthases are also different in their molecular weights as determined by sucrose density gradient centrifugation (154,000 ± 17,000 (n = 6) for the normal and 185,000 ± 15,000 (n = 8) for the diabetic synthase) and their kinetic properties.     

Changes in the kinetic parameters of hepatic gamma-glutamyltransferase from streptozotocin-induced diabetic rats

Previous research has shown that the enzymatic activity of hepatic gamma-glutamyltransferase was increased in streptozotocin-induced diabetic rats with no increase in the expression of the protein. The current work has characterized the differences in the kinetic properties of hepatic gamma-glutamyltransferase from diabetic versus control rats. Hepatic gamma-glutamyltransferase was purified from control male and female rats and from rats made diabetic 30 days previously with streptozotocin. The maximal velocity and the Michaelis constant were determined for the purified enzyme with two separate donors (L-gamma-glutamyl-p-nitroanilide or L-gamma-glutamyl-(7-amido-4-methylcoumarin)) in the presence of one of eight acceptors (L-alanine-glycine, L-glycine-glycine, L-methionine, L-glutamate, L-alanine, L-glutamine, L-phenylalanine or L-aspartate). With both donors, hepatic gamma-glutamyltransferase from diabetic rats had a consistently higher kinetic efficiency than gamma-glutamyltransferase from controls. The kinetic efficiency percent increase of diabetic over control gamma-glutamyltransferase when averaged across all acceptors was higher in males than in females. With L-gamma-glutamyl-p-nitroanilide, the kinetic efficiency increase of diabetic over control gamma-glutamyltransferase was higher with poor acceptors than with highly efficient acceptors. These data indicate that there are differences in the physical properties of hepatic gamma-glutamyltransferase from diabetic versus control rats and from female versus male rats.     

3,5-Diiodo-l-thyronine ameliorates diabetic nephropathy in streptozotocin-induced diabetic rats

3, 5-Diiodothyronine (T2), a natural metabolite of triiodothyronine (T3) from deiodination pathway, can mimic biologic effects of T3 without inducing thyrotoxic effects. Recent studies revealed T3 acted as a protective factor against diabetic nephropathy (DN). Nevertheless, little is known about the effect of T2 on DN. This study was designed to investigate whether and how T2 affects experimental models of DN in vivo and in vitro. Administration of T2 was found to prevent significant decrease in SIRT1 protein expression and activity as well as increases in blood glucose, urine albumin excretion, matrix expansion, transforming growth factor-β1 expression, fibronectin and type IV collagen deposition in the diabetic kidney. Concordantly, similar effects of T2 were exhibited in the cultured rat mesangial cells (RMC) exposed to high glucose and that could be abolished by a known SIRT1 inhibitor, sirtinol. Moreover, enhanced NF-κB acetylation and JNK phosphorylation present in both diabetic rats and high glucose-treated RMC were distinctly dampened by T2. Collectively, these results suggested that T2 was a protective agent against renal damage in diabetic nephropathy, whose action involved regulation of SIRT1.