Diabetic KK-Ay Mice Research Articles

Isosteviol increases insulin sensitivity and changes gene expression of key insulin regulatory genes and transcription factors in islets of the diabetic KKAy mouse

Isosteviol (ISV), a diterpene molecule, is an isomer of the backbone structure of a group of substances with recently proven antidiabetic capabilities in both man and rodents. The aim of this study was to investigate if ISV possesses beneficial effects on the metabolism in the diabetic KKAy mouse and to establish the long-term in vivo effects of ISV on the gene expression profile of key insulin regulatory genes in islets. Twenty KKAy mice, aged 5 weeks, were divided into two groups and treated for 9 weeks with either (i) standard chow diet (control) or (ii) chow + 20 mg/kg body weight of ISV. Blood samples were collected before and after intervention and were subsequently analysed. As a non-diabetic control group, 10 normal C57BL mice were fed with standard chow diet. Gene expression was determined in islets by quantitative real-time RT-PCR and Affymetrix microarray. We demonstrated that long-term treatment with ISV improves glucose homeostasis, increases insulin sensitivity, lowers plasma triglycerides and lowers weight in the diabetic KKAy mice. Furthermore, ISV markedly changes the gene expression profile of key insulin regulatory genes GLUT2, Ins1, Ins2, Pdx1/Ipf1, Beta2/Neurod1, Pax6 and 11-beta-HSD-1 and beta-cell transcription factors Nkx2-2, Nkx6-1, C/EBPalpha and FoxA2 in isolated islets of the KKAy mice. The results indicate that ISV improves glucose and insulin sensitivity as well as improving the lipid profile and upregulates the gene expression of key beta-cell genes, including insulin regulatory transcription factors.

Exploitation of the derivatives of Dictyostelium differentiation-inducing factor-1, which promote glucose consumption in mammalian cells

Aims The differentiation-inducing factor-1 (DIF-1) is a signal molecule that induces stalk cell formation in the cellular slime mold Dictyostelium discoideum. DIF-1 has also been shown to possess pharmacological activities, such as the suppression of tumor cell growth and the promotion of glucose uptake in non-transformed mammalian cells. In this study, we tried to develop compounds that possess weaker anti-tumor activity and stronger glucose uptake-promoting activity than DIF-1. Main methods We investigated the in vitro effects of 12 derivatives of DIF-1 on glucose consumption in mouse 3T3-L1 cells and on cell growth in K562 human leukemia cells. We also examined the effect of a good compound on the blood glucose concentration in KK-Ay diabetic mice. Key findings We found that some derivatives at 20 μM promoted glucose consumption more than twice as fast as the control. Of the derivatives, a compound named DIF-1(3M), which has a weaker anti-leukemic effect than DIF-1, promoted glucose consumption as strongly as DIF-1 in confluent 3T3-L1 cells. While DIF-1 at 20 μM was inhibitory to the cell growth of 3T3-L1, DIF-1(3M) at 20 μM exhibited no inhibitory effect on the growing cells. We also found that DIF-1(3M) injected (10–12.5 mg/kg body weight) intraperitoneally in mice tended to lower the blood glucose concentration. Significance The present results open the possibility for the development of new agents that possess strong glucose-uptake-promoting activity but little anti-tumor activity and may have therapeutic potential for the treatment of diabetes and/or obesity.

Increased Insulin Sensitivity and Changes in the Expression Profile of Key Insulin Regulatory Genes and Beta Cell Transcription Factors in Diabetic KKAy-Mice after Feeding with a Soy Bean Protein Rich Diet High in Isoflavone Content

High content isoflavone soy protein (SBP) (Abalon) has been found in animal studies to possess beneficial effects on a number of the characteristic features of the insulin resistance syndrome. The aim of this study was to investigate whether SBP exerts beneficial effects on metabolism in the diabetic KKAy-mouse. Furthermore, we investigated the long-term in vivo effect of SBP on the expression profile in islets of key insulin regulatory genes. Twenty KKAy-mice, aged 5 weeks, were divided into 2 groups and treated for 9 weeks with either (A) standard chow diet (control) or (B) chow + 50% SBP. Twenty normal C57BL-mice fed with standard chow diet served as nondiabetic controls (C). Blood samples were collected and analyzed before and after intervention. Gene expression was determined in islets by quantitative real-time RT-PCR and Affymetrix microarray. It was demonstrated that long-term treatment with SBP improves glucose homeostasis, increases insulin sensitivity, and lowers plasma triglycerides in diabetic KKAy-mice. SBP reduces fasting plasma glucose, insulin, triglycerides, and total cholesterol. Furthermore, SBP markedly changes the gene expression profile of key insulin regulatory genes GLUT2, GLUT3, Ins1, Ins2, IGF1, Beta2/Neurod1, cholecystokinin, and LDLr, and proliferative genes in islets isolated from KKAy-mice. After 9 weeks of treatment with SBP, plasma glucose and insulin homeostasis was normalized compared to start levels. The results indicate that SBP improves glucose and insulin sensitivity and up-regulates the expression of key insulin regulatory genes.

Palmitate impairs and eicosapentaenoate restores insulin secretion through regulation of SREBP-1c in pancreatic islets.

OBJECTIVE—Chronic exposure to fatty acids causes β-cell failure, often referred to as lipotoxicity. We investigated its mechanisms, focusing on contribution of SREBP-1c, a key transcription factor for lipogenesis.RESEARCH DESIGN AND METHODS—We studied in vitro and in vivo effects of saturated and polyunsaturated acids on insulin secretion, insulin signaling, and expression of genes involved in β-cell functions. Pancreatic islets isolated from C57BL/6 control and SREBP-1–null mice and adenoviral gene delivery or knockdown systems of related genes were used.RESULTS—Incubation of C57BL/6 islets with palmitate caused inhibition of both glucose- and potassium-stimulated insulin secretion, but addition of eicosapentaenoate (EPA) restored both inhibitions. Concomitantly, palmitate activated and EPA abolished both mRNA and nuclear protein of SREBP-1c, accompanied by reciprocal changes of SREBP-1c target genes such as insulin receptor substrate-2 (IRS-2) and granuphilin. These palmitate-EPA effects on insulin secretion were abolished in SREBP-1–null islets. Suppression of IRS-2/Akt pathway could be a part of the downstream mechanism for the SREBP-1c–mediated insulin secretion defect because adenoviral constitutively active Akt compensated it. Uncoupling protein-2 (UCP-2) also plays a crucial role in the palmitate inhibition of insulin secretion, as confirmed by knockdown experiments, but SREBP-1c contribution to UCP-2 regulation was partial. The palmitate-EPA regulation of insulin secretion was similarly observed in islets from C57BL/6 mice pretreated with dietary manipulations. Furthermore, administration of EPA to diabetic KK-Ay mice ameliorated impairment of insulin secretion in their islets.CONCLUSIONS—SREBP-1c plays a dominant role in palmitate-mediated insulin secretion defect, and EPA prevents it through SREBP-1c inhibition, implicating a therapeutic potential for treating diabetes related to lipotoxicity.

The kidney expression of matrix metalloproteinase-9 in the diabetic nephropathy of Kkay mice

Background Disorder of the extracellular matrix has been suggested to play a role in the development of diabetic nephropathy. The aim of the present study was to examine the expression of matrix metalloproteinase-9 (MMP-9) in the kidney of Kkay mice, a type 2 diabetic model. Methods Eight Kkay control mice (KA) and eight Kkay diabetic mice (KB) were fed with the same diets for 4 months and sacrificed after glucose levels and weight were measured. The renal pathological changes were examined by periodic acid–Schiff and periodic acid–silver methenamine staining, and the glomerulosclerosis indexes (GIs) were calculated. The levels of MMP-9 protein in the kidney tissue were assessed by immunohistochemistry. The gene expression of MMP-9 was measured by reverse transcription polymerase chain reaction. Results The diabetic group had morphological findings of diabetic nephropathy, and their GI (240.0±17.3) was much higher than that of the control group (118.4±15.1) ( P<.01). The mean positive area of MMP-9 protein in the glomeruli of Kkay mice in the KB group was higher than that in the KA group. The expression of MMP-9 messenger RNA level in the diabetic KB group was up-regulated when compared with the KA group. Conclusion The expression of MMP-9 in the kidney of mice with diabetic nephropathy was enhanced, and MMP-9 might be involved in the development of diabetic nephropathy.

T2384, a Novel Antidiabetic Agent with Unique Peroxisome Proliferator-activated Receptor γ Binding Properties

The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPARgamma) plays central roles in adipogenesis and glucose homeostasis and is the molecular target for the thiazolidinedione (TZD) class of antidiabetic drugs. Activation of PPARgamma by TZDs improves insulin sensitivity; however, this is accompanied by the induction of several undesirable side effects. We have identified a novel synthetic PPARgamma ligand, T2384, to explore the biological activities associated with occupying different regions of the receptor ligand-binding pocket. X-ray crystallography studies revealed that T2384 can adopt two distinct binding modes, which we have termed "U" and "S", interacting with the ligand-binding pocket of PPARgamma primarily via hydrophobic contacts that are distinct from full agonists. The different binding modes occupied by T2384 induced distinct patterns of coregulatory protein interaction with PPARgamma in vitro and displayed unique receptor function in cell-based activity assays. We speculate that these unique biochemical and cellular activities may be responsible for the novel in vivo profile observed in animals treated systemically with T2384. When administered to diabetic KKAy mice, T2384 rapidly improved insulin sensitivity in the absence of weight gain, hemodilution, and anemia characteristics of treatment with rosiglitazone (a TZD). Moreover, upon coadministration with rosiglitazone, T2384 was able to antagonize the side effects induced by rosiglitazone treatment alone while retaining robust effects on glucose disposal. These results are consistent with the hypothesis that interactions between ligands and specific regions of the receptor ligand-binding pocket might selectively trigger a subset of receptor-mediated biological responses leading to the improvement of insulin sensitivity, without eliciting less desirable responses associated with full activation of the receptor. We suggest that T2384 may represent a prototype for a novel class of PPARgamma ligand and, furthermore, that molecules sharing some of these properties would be useful for treatment of type 2 diabetes.

Inhibition of 11βHSD1 with the S-phenylethylaminothiazolone BVT116429 increases adiponectin concentrations and improves glucose homeostasis in diabetic KKAy mice

BackgroundA substantial body of evidence indicates that reduced plasma adiponectin levels may be key in the development of insulin resistance, type 2 diabetes and the metabolic syndrome. Glucocorticoids decrease the levels of adiponectin in animals and humans. Cortisone is transformed to its active form cortisol, via 11β-hydroxysteroid dehydrogenase (HSD) type 1. This study sought to ascertain if inhibition of 11β HSD1 with a new selective inhibitor, BVT116429, affects the concentrations of circulating adiponectin with concomitant effects on glucose homeostasis in diabetic mice.ResultsKKAy mice were treated with BVT116429 (3, 10, 30 mg/kg), rosiglitazone (5 mg/kg) or vehicle once daily for ten days. Plasma adiponectin levels rose in mice treated with BVT116429 and this was found to be both the hexameric and the high molecular weight multimeric forms of adiponectin. Seven days of treatment with the 11β HSD1-inhibitor BVT116429 decreased basal insulin levels but no changes in glucose tolerance were seen. After ten days of treatment, fasting blood glucose level was decreased by BVT116429 comparable to the effects of rosiglitazone. Another 11β HSD1 inhibitor, BVT2733, improved HbA1c but had no effect on adiponectin.ConclusionInhibition of 11β HSD1 can be expected to be beneficial for treating the pathology of type 2 diabetes mellitus. The differences seen in adiponectin between BVT116429 and BVT2733 could be explained by different pharmacodynamics exerted by the compounds in different tissues in the body. Increases in adiponectin concentrations may be an integral component in the mechanism of action of this new11β HSD1 inhibitor and may be a useful marker of efficacy during the clinical development of 11β HSD1 inhibitor compounds.

The effects of β 3-adrenoceptor agonist CL-316,243 on adiponectin, adiponectin receptors and tumor necrosis factor-α expressions in adipose tissues of obese diabetic KKAy mice

We investigated the effects of β 3-adrenoceptor agonist, 5-[(2 R)-2-[[(2 R)-2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1,3-benzodioxole-2,2-dicarboxylate (CL-316,243) in obese diabetic KKAy mice. Two weeks' subcutaneous administration of CL-316,243 reduced serum levels of glucose, insulin, triglyceride, free fatty acid and tumor necrosis factor-α (TNF-α), and increased adiponectin. Adiponectin, adiponectin receptors and β 3-adrenoceptor mRNA expressions were reduced in epididymal white adipose tissue in KKAy mice, and CL-316,243 recovered these mRNA expressions. Meanwhile, CL-316,243 suppressed the overexpressed mRNA level of TNF-α in both epididymal white adipose tissue and brown adipose tissue. These data suggest that the normalization of adiponectin, adiponectin receptors and TNF-α may result in the amelioration of obesity-induced insulin resistance.

Effect of Tangzhiqing on Glucose and Lipid Metabolism in Genetically Type 2 Diabetes KK-Ay Mice

The hypoglycemic and hypolipidemic effects of Tangzhiqing (abbreviated TZ) a mixture of Mulberry Leaf, Lotus Leaf, Danshen Root, and Hawthorn Leaf, were investigated in KK-Ay mice, an animal model of type 2 diabetes. TZ reduced blood glucose, total cholesterol, and triglyceride levels of KK-Ay mice at 4 weeks after oral administration. TZ also improved sucrose or maltose tolerance. Maltase activity in small intestine significantly decreased in the TZ-treated KK-Ay mice. These results support the hypothesis that TZ improves glucose metabolism by reducing α-glycosidase activity. Therefore TZ might have beneficial effects on hyperglycemia and hyperlipidemia in type 2 diabetes.

Astragalus polysaccharide reduces hepatic endoplasmic reticulum stress and restores glucose homeostasis in a diabetic KKAy mouse model

To examine the potential effects of Astragalus polysaccharide (APS) on hepatic endoplasmic reticulum (ER) stress in vivo and in vitro and its link with hypoglycemia activity, thus establishing the mechanism underlying the hypoglycemic action of APS. The obese and type 2 diabetic KKAy mouse model, which is the yellow offspring of the KK mice expressed Ay gene (700 mg/kg-1/d-1, 8 weeks) and a high glucose-induced HepG2 cell model (200 microg/mL, 24 h) were treated with APS. The oral glucose tolerance test was measured to reflex insulin sensitivity with the calculated homeostasis model assessment (HOMA-IR) index. XBP1 (XhoI site-binding protein 1) transcription and splicing, an indicator of ER stress, was analyzed by RT-PCR and real-time PCR. The expression and activation of glycogen synthase kinase 3 beta (GSK3beta), an insulin signaling protein, was measured by Western blotting. APS can alleviate ER stress in cultured cells in vivo. The hyperglycemia status, systemic insulin sensitivity, fatty liver disease, and insulin action in the liver of diabetic mice were partly normalized or improved in response to APS administration. Our results indicate that APS enables insulin-sensitizing and hypoglycemic activity at least in part by enhancing the adaptive capacity of the ER, which can further promote insulin signal transduction. Thus, APS has promising application in the treatment of type 2 diabetes.

Insulin sensitizing pharmacology of thiazolidinediones correlates with mitochondrial gene expression rather than activation of PPAR gamma.

Insulin sensitizing thiazolidinediones (TZDs) are generally considered to work as agonists for the nuclear receptor peroxisome proliferative activated receptor-gamma (PPAR gamma). However, TZDs also have acute, non-genomic metabolic effects and it is unclear which actions are responsible for the beneficial pharmacology of these compounds. We have taken advantage of an analog, based on the metabolism of pioglitazone, which has much reduced ability to activate PPAR gamma. This analog (PNU-91325) was compared to rosiglitazone, the most potent PPAR gamma activator approved for human use, in a variety of studies both in vitro and in vivo. The data demonstrate that PNU-91325 is indeed much less effective than rosiglitazone at activating PPAR gamma both in vitro and in vivo. In contrast, both compounds bound similarly to a mitochondrial binding site and acutely activated PI-3 kinase-directed phosphorylation of AKT, an action that was not affected by elimination of PPAR gamma activation. The two compounds were then compared in vivo in both normal C57 mice and diabetic KKAy mice to determine whether their pharmacology correlated with biomarkers of PPAR gamma activation or with the expression of other gene transcripts. As expected from previous studies, both compounds improved insulin sensitivity in the diabetic mice, and this occurred in spite of the fact that there was little increase in expression of the classic PPAR gamma target biomarker adipocyte binding protein-2 (aP2) with PNU-91325 under these conditions. An examination of transcriptional profiling of key target tissues from mice treated for one week with both compounds demonstrated that the relative pharmacology of the two thiazolidinediones correlated best with an increased expression of an array of mitochondrial proteins and with expression of PPAR gamma coactivator 1-alpha (PGC1 alpha), the master regulator of mitochondrial biogenesis. Thus, important pharmacology of the insulin sensitizing TZDs may involve acute actions, perhaps on the mitochondria, that are independent of direct activation of the nuclear receptor PPAR gamma. These findings suggest a potential alternative route to the discovery of novel insulin sensitizing drugs.

Des-Aspartate-Angiotensin I Exerts Hypoglycemic Action via Glucose Transporter-4 Translocation in Type 2 Diabetic KKAy Mice and GK Rats

The present study investigated the hypoglycemic action of des-aspartate-angiotensin I (DAA-I), a metabolite of angiotensin I, in two animal models of type 2 diabetes. The rationale was based on our earlier studies demonstrating that DAA-I acts on the angiotensin AT(1) receptor and exerts responses opposing those of angiotensin II and on recent reports that curtailment of angiotensin II formation by angiotensin converting enzyme inhibitors and blockade of the AT(1) receptor attenuate hyperglycemia in type 2 diabetics and diabetic animals. Diabetic KKAy mice and GK rats were administered orally (by gavage) one of the following doses of DAA-I: 400, 600, or 800 nmol/kg.d for 4 and 6 wk, respectively. Control diabetic animals were similarly administered water. Blood glucose of each animal was determined fortnightly by oral glucose tolerance test and blood insulin on the last day of treatment. Animals were killed, and the levels of plasma membrane glucose transporter-4 and cytosolic tyrosine-phosphorylated insulin receptor substrate-1 in hind limb skeletal muscles were determined by Western blot in insulin-challenged and nonchallenged animals. Orally administered DAA-I had no effect on blood insulin level but exerted dose-dependent hypoglycemic action in KKAy mice and GK rats after 4 and 6 wk of treatment, respectively. At the maximal effective dose of 600 nmol/kg, insulin induced a significant increase in plasma membrane glucose transporter-4 and cytosolic tyrosine-phosphorylated insulin receptor substrate-1. These findings show that DAA-I is not an insulin secretagogue and exerts hypoglycemic action by attenuating insulin resistance, the first such demonstration indicating that the nonapeptide is involved in glycemic regulation.

Molecular Mechanism for Antidiabetic Activity of [meso-Tetrakis(4-sulfonatophenyl)porphyrinato]oxovanadium(IV) (VO(tpps)) Complex. Studies on Akt Phosphorylation and GLUT4 Translocation in 3T3-L1 Adipocytes

Abstract A [meso-tetrakis(4-sulfonatophenyl)porphyrinato]oxovanadium(IV) (VO(tpps)), has been found to have an antidiabetic activity and thus to be effective for treating diabetic animal models. High hypoglycemic effects have been observed not only in streptozotocin (STZ)-induced type 1 diabetic mice but also type 2 diabetic KKAy mice by oral administration of the complex. However, the molecular mechanism for the activities has not yet been examined. The aim of this study was to investigate the molecular mechanism of VO(tpps) in 3T3-L1 adipocytes with respect to insulin signaling pathway. Addition of VO(tpps) was found to phosphorylate the insulin receptor β (IRβ) subunit, insulin receptor substrate 1 (IRS1) and protein kinase B (PKB)/Akt in 3T3-L1 adipocytes, in which all phosphorylations were time dependent. The VO(tpps)-induced Akt phosphorylation was suppressed by a PI3K inhibitor, wortmannin. VO(tpps) also stimulated GLUT4 protein translocation to the plasma membrane. Based on these results, we conclude that VO(tpps) exhibits both insulinomimetic and antidiabetic activities via tyrosine phosphorylations of IRβ and IRS1, which activate the downstream Akt phosphorylation through PI3K, and this signaling cascade finally causes GLUT4 translocation from the cytosol to the plasma membrane.

Normalization of Prandial Blood Glucose and Improvement of Glucose Tolerance by Liver-Specific Inhibition of SH2 Domain–Containing Inositol Phosphatase 2 (SHIP2) in Diabetic KKAy Mice

Type 2 diabetes is characterized by a progressive resistance of peripheral tissues to insulin. Recent data have established the lipid phosphatase SH2 domain-containing inositol phosphatase 2 (SHIP2) as a critical negative regulator of insulin signal transduction. Mutations in the SHIP2 gene are associated with type 2 diabetes. Here, we used hyperglycemic and hyperinsulinemic KKA(y) mice to gain insight into the signaling events and metabolic changes triggered by SHIP2 inhibition in vivo. Liver-specific expression of a dominant-negative SHIP2 mutant in KKA(y) mice increased basal and insulin-stimulated Akt phosphorylation. Protein levels of glucose-6-phosphatase and phosphoenolpyruvate carboxykinase were significantly reduced, and consequently the liver produced less glucose through gluconeogenesis. Furthermore, SHIP2 inhibition improved hepatic glycogen metabolism by modulating the phosphorylation states of glycogen phosphorylase and glycogen synthase, which ultimately increased hepatic glycogen content. Enhanced glucokinase and reduced pyruvate dehydrogenase kinase 4 expression, together with increased plasma triglycerides, indicate improved glycolysis. As a consequence of the insulin-mimetic effects on glycogen metabolism, gluconeogenesis, and glycolysis, the liver-specific inhibition of SHIP2 improved glucose tolerance and markedly reduced prandial blood glucose levels in KKA(y) mice. These results support the attractiveness of a specific inhibition of SHIP2 for the prevention and/or treatment of type 2 diabetes.

Antidiabetic Activities of Chalcones Isolated from a Japanese Herb, Angelica keiskei

Diabetes mellitus is a chronic disease that is characterized by hyperglycemia caused by insufficient insulin action. We have explored the edible ingredients from folk medicines in Japan that contain substances complementing insulin action, such as the induction of adipocyte differentiation and the enhancement of glucose uptake. We eventually found that the ethanol extract from a Japanese herb "Ashitaba", Angelica keiskei, contained two major chalcones of 4-hydroxyderricin (4-HD) and xanthoangelol that showed strong insulin-like activities via a pathway independent of the peroxisome proliferator-activated receptor-gamma activation. The 4-HD especially showed the preventive effects on the progression of diabetes in genetically diabetic KK-Ay mice.

Antidiabetic effect of the herb Myrcia speciosa in KK-Ay diabetic mice

Antidiabetic effect of the herb Myrcia speciosa in KK-Ay diabetic mice

Expression Analysis of cPLA2 Alpha Interacting TIP60 in Diabetic KKAy and Non-Diabetic C57BL Wild-Type Mice: No Impact of Transient and Stable TIP60 Overexpression on Glucose-Stimulated Insulin Secretion in Pancreatic Beta-Cells

In the present study we investigate the expression levels of cytosolic phospholipase A2 alpha (cPLA2alpha) interacting histone acetyl transferase proteins TIP60alpha and TIP60beta in non-diabetic C57BL wild-type mice and obese type 2 diabetic KKAy model mice. The aim was to test our hypothesis that TIP60 plays a regulatory role in glucose-stimulated insulin secretion from pancreatic beta-cells. Ten obese diabetic KKAy mice and ten non-diabetic C57BL mice were fed a standard chow diet. After nine weeks, islet RNA was purified and used to measure TIP60 expression. We investigated the effect of TIP60alpha and TIP60beta on glucose-stimulated insulin secretion by transient and stable overexpression in the pancreatic mouse beta-cell line MIN6 and the rat beta-cell line INS-1E. We found that non-diabetic C57BL mice and diabetic KKAy mice have the same level of both the alpha and beta splice forms of TIP60. Furthermore, we demonstrated that transient and stable expression of TIP60 in INS-1E cells affects neither glucose-stimulated insulin secretion, insulin output nor cell insulin content. Also susceptibility to developing gluco-toxicity was unaffected. TIP60 over-expression does not affect glucose stimulated insulin secretion, insulin content or abnormal beta-cell function during glucotoxicity.

Bioactive Constituents from Chinese Natural Medicines. XXIV. Hypoglycemic Effects of Sinocrassula indica in Sugar-Loaded Rats and Genetically Diabetic KK-Ay Mice and Structures of New Acylated Flavonol Glycosides, Sinocrassosides A1, A2, B1, and B2

The methanolic extract from the whole plant of Sinocrassula indica (Crassulaceae) was found to inhibit the increase in serum glucose levels in oral administration of sucrose and glucose in rats at a dose of 250 mg/kg (p.o.). However, the extract did not inhibit the increase in serum glucose levels after intraperitoneal administration of glucose in these animals but did partly inhibit the gastric emptying. On the other hand, this extract significantly inhibited the increase in serum glucose levels after administration for 2 weeks in KK-A(y) mice, a genetically type II diabetic mice, at a dose of 250 mg/kg/d (p.o.) without significant changes of the weights of body, liver, and visceral fat. From the extract, four new acylated flavonol glycosides, sinocrassosides A(1), A(2), B(1), and B(2), were isolated together with 11 flavonoids and 2 megastigmanes. The absolute stereostructures of the four new compounds were elucidated on the basis of chemical and physicochemical evidence.

Insulin Sensitizing Pharmacology of Thiazolidinediones Correlates with Mitochondrial Gene Expression rather than Activation of PPARγ

Insulin sensitizing thiazolidinediones (TZDs) are generally considered to work as agonists for the nuclear receptor peroxisome proliferative activated receptor-gamma (PPARγ). However, TZDs also have acute, non-genomic metabolic effects and it is unclear which actions are responsible for the beneficial pharmacology of these compounds. We have taken advantage of an analog, based on the metabolism of pioglitazone, which has much reduced ability to activate PPARγ. This analog (PNU-91325) was compared to rosiglitazone, the most potent PPARγ activator approved for human use, in a variety of studies both in vitro and in vivo. The data demonstrate that PNU-91325 is indeed much less effective than rosiglitazone at activating PPARγ both in vitro and in vivo. In contrast, both compounds bound similarly to a mitochondrial binding site and acutely activated PI-3 kinase-directed phosphorylation of AKT, an action that was not affected by elimination of PPARγ activation. The two compounds were then compared in vivo in both normal C57 mice and diabetic KKAy mice to determine whether their pharmacology correlated with biomarkers of PPARγ activation or with the expression of other gene transcripts. As expected from previous studies, both compounds improved insulin sensitivity in the diabetic mice, and this occurred in spite of the fact that there was little increase in expression of the classic PPARγ target biomarker adipocyte binding protein-2 (aP2) with PNU-91325 under these conditions. An examination of transcriptional profiling of key target tissues from mice treated for one week with both compounds demonstrated that the relative pharmacology of the two thiazolidinediones correlated best with an increased expression of an array of mitochondrial proteins and with expression of PPARγ coactivator 1-alpha (PGC1α), the master regulator of mitochondrial biogenesis. Thus, important pharmacology of the insulin sensitizing TZDs may involve acute actions, perhaps on the mitochondria, that are independent of direct activation of the nuclear receptor PPARγ. These findings suggest a potential alternative route to the discovery of novel insulin sensitizing drugs.

Effects of Ingested Turmeric Oleoresin on Glucose and Lipid Metabolisms in Obese Diabetic Mice: A DNA Microarray Study

Turmeric, the rhizome of Curcuma longa L., has a wide range of effects on human health. Turmeric oleoresin, an extract of turmeric, is often used for flavoring and coloring. Curcuminoids and turmeric essential oil are both contained in turmeric oleoresin, and both of these fractions have hypoglycemic effects. In the present study, we comprehensively assessed the effect of turmeric oleoresin on hepatic gene expression in obese diabetic KK-Ay mice using DNA microarray analysis and quantitative real-time polymerase chain reaction (PCR). Female KK-Ay mice aged 6 weeks (n = 6/group) were fed a high-fat diet containing turmeric oleoresin, curcuminoids, and essential oil for 5 weeks. The same diet without any of these fractions was used as a control diet. Ingestion of turmeric oleoresin and essential oil inhibited the development of increased blood glucose and abdominal fat mass, while curcuminoids only inhibited the increase in blood glucose. DNA microarray analysis indicated that turmeric oleoresin ingestion up-regulated the expression of genes related to glycolysis, beta-oxidation, and cholesterol metabolism in the liver of KK-Ay mice, while expression of gluconeogenesis-related genes was down-regulated. Real-time PCR analysis was conducted to assess the contribution of the curcuminoids and essential oil in turmeric oleoresin to the changes in expression of representative genes selected by DNA microarray analysis. This analysis suggested that curcuminoids regulated turmeric oleoresin ingestion-induced expression of glycolysis-related genes and also that curcuminoids and turmeric essential oil acted synergistically to regulate the peroxisomal beta-oxidation-related gene expression induced by turmeric oleoresin ingestion. These changes in gene expression were considered to be the mechanism by which the turmeric oleoresin affected the control of both blood glucose levels and abdominal adipose tissue masses. All of these results suggest that the use of whole turmeric oleoresin is more effective than the use of either curcuminoids or the essential oil alone.