Comparison of area under the curve in various models of diabetic rats receiving chronic medication.

Introduction: The oral glucose tolerance test (OGTT) is widely used as a diagnostic tool for impaired glucose tolerance (IGT) in clinical settings and animal experiments. The area under the curve (AUC) is then developed to quantify the total increase in blood glucose during the OGTT. Similarly, attenuation of the increased AUC indicates the improvement of IGT in animals. Variations in fasting plasma glucose between individuals stimulate the development of incremental area under the curve (iAUC). However, the iAUC determined from subtracting the baseline value of fasting plasma glucose (similar to ΔAUC) has been challenged as problematic without evidence. Material and methods: We developed four different diabetic animal models. In each model, rats were treated with metformin, dapagliflozin, and insulin respectively for 1 week. OGTTs were performed after 7 days of the drug treatment. The acute blood glucose changes induced by one-time treatment of drugs were also compared. Results: After a daily application of each drug at an effective dose for 7 days, results indicated potency in the following order: insulin > dapagliflozin > metformin. This was determined by calculation using the AUC in all diabetic models. However, the order changed when using the calculation with iAUC. Additionally, signals were changed before the OGTT in each model that received repeated treatment of each drug. Notably, drug potency was shown to be the same in OGTT calculated from iAUC and AUC in diabetic rats receiving acute treatment. Conclusions: iAUC seems unsuitable for application in cases where subjects are receiving chronic medication(s).

Myricetin Increases Circulating Adropin Level after Activation of Glucagon-like Peptide 1 (GLP-1) Receptor in Type-1 Diabetic Rats.

Myricetin is a common plant-derived flavonoid, considered an agonist of glucagon-like peptide 1 (GLP-1) receptor. It improves glycemic control and helps reduce body weight in diabetic subjects. The potential mechanisms of action of myricetin in this context might be enhancing the secretion of ß-endorphin (BER) to activate peripheral µ-opioid receptors. Moreover, adropin is a nutritionally regulated peptide hormone, which regulates energy metabolism, and plays a role in ameliorating diabetes. Because their mechanisms of insulin sensitivity are closely related, we hypothesized that myricetin may interact with adropin and plasma BER. The present study investigated the glucose-lowering effect of acute and chronic treatments of myricetin in type-1 diabetic rats. Plasma BER and adropin levels were determined by enzyme-linked immunosorbent assay (ELISA). The secretion of BER was measured in rats who received adrenalectomy. The changes in adropin gene (Enho) or mRNA level of GLP-1 receptor were measured using qPCR analysis. The results showed that myricetin dose-dependently increased plasma BER and adropin levels like the reduction of hyperglycemia after bolus injection as acute treatment. In addition, these effects of myricetin were inhibited by the antagonist of GLP-1 receptor. Moreover, in HepG2 cell line, myricetin induced GLP-1 receptor activation, which modulated the expression of adropin. In diabetic rats, the plasma adropin increased by myricetin is mainly through endogenous ß-endorphin after activation of GLP-1 receptor via bolus injection as acute treatment. Additionally, chronic treatment with myricetin increased adropin secretion in diabetic rats. In conclusion, our results provide a new finding that activation of opioid µ-receptor in the liver may enhance circulating adropin in animals.

TGR5 Expression Is Associated with Changes in the Heart and Urinary Bladder of Rats with Metabolic Syndrome.

Adipose-derived cytokines may contribute to the inflammation that occurs in metabolic syndrome (MetS). The Takeda G protein-coupled receptor (TGR5) regulates energy expenditure and affects the production of pro-inflammatory biomarkers in metabolic diseases. Etanercept, which acts as a tumor necrosis factor (TNF)-α antagonist, can also block the inflammatory response. Therefore, the interaction between TNF-α and TGR5 expression was investigated in rats with high-fat diet (HFD)-induced obesity. Heart tissues isolated from the HFD-induced MetS rats were analyzed. Changes in TGR5 expression were investigated with lithocholic acid (LCA) as the agonist. Betulinic acid (BA) was used to activate TGR5 in urinary bladders. LCA was more effective in the heart tissues of HFD-fed rats, although etanercept alleviated the function of LCA. STAT3 activation and higher TGR5 expression were observed in the heart tissues collected from HFD-fed rats. Thus, cardiac TGR5 expression is promoted by HFD through STAT3 activation in rats. Moreover, the urinary bladders of female rats fed a HFD showed a low response, which was reversed by etanercept. Relaxation by BA in the bladders was more marked in HFD-fed rats. The high TGR5 expression in HFD-fed rats was characterized using a mRNA assay, and the increased cAMP levels were found to be stimulated by BA in the isolated bladders. Therefore, TGR5 expression increases with a HFD in both the hearts and urinary bladders. Collectively, cytokine-medicated TGR5 activation was observed in the hearts and urinary bladders of rats.

Connective tissue growth factor in hepatocytes is elevated by carbon tetrachloride via STAT3 activation.

Carbon tetrachloride (CCl4) is widely used to induce hepatic fibrosis. Therapeutic agents alleviate hepatic fibrosis by inhibiting signal transducer and activator of transcription 3 (STAT3) activation. To understand the direct effects of CCl4 on STAT3 expression in the liver, the present study incubated cultured hepatocytes expressing connective tissue growth factor (CTGF) with CCl4. Rats exposed to CCl4 for 8 weeks exhibited hepatic fibrosis, which was confirmed through the assessment of plasma biomarkers. Isolated liver samples were used to determine the protein levels of CTGF and STAT3 using western blotting. In addition, STAT3 expression was silenced in α mouse liver 12 (AML­12) cells using small interfering RNA transfection. In addition, a pharmacological inhibitor, stattic, was used to inhibit STAT3 expression. The incubation of AML­12 cells with CCl4 induced a dose­dependent increase in CTGF expression and STAT3 activation. Notably, silymarin, an extract from milk thistle, inhibited these changes in AML­12 cells and the antioxidant tiron produced similar effects. Silencing of STAT3 reduced the CTGF expression promoted by CCl4 in the hepatocytes. Additionally, similar to tiron, stattic inhibited CTGF expression induced by CCl4. In conclusion, CCl4 may activate STAT3 through oxidative stress to promote CTGF expression, which is one of the main factors contributing to the risk of hepatic fibrosis.

Induction of apoptosis in prostate cancer by ginsenoside Rh2.

The therapeutic action of ginsenoside Rh2 on several cancer models has been reported. This study aimed to evaluate its apoptotic effect on prostate cancer and the underlying mechanism. Cultured DU145 cells were treated with Rh2 (5 × 10-5 to 1 × 10-4 M), peroxisome proliferator-activated receptor-delta (PPAR-delta) antagonist GSK0660 (1 × 10-6 to 5 × 10-6 M); or small interfering RNA (siRNA) of PPAR-delta. The treatment effects were evaluated with cell viability assay, life/death staining and flow cytometry for apoptosis. Immunostaining was used for reactive oxygen species (ROS) and superoxide detection. Western blot analysis for PPAR-delta and signal transducer and activator of transcription 3 (STAT3) protein expression were performed. The results showed that Rh2 significantly decreased DU145 cell survival and increased cell apoptosis. ROS and superoxide induction, PPAR-delta up-regulation and phosphorylated STAT3 (p-STAT3) down-regulation by Rh2 were demonstrated. GSK0660 partially but significantly inhibited the Rh2-induced apoptosis and restored cell viability. Treatment with siRNA reversed the Rh2-induced apoptosis as well as changes in PPAR-delta and p-STAT3 expression. In conclusion, our findings have demonstrated that ginsenoside Rh2 induces prostate cancer DU145 cells apoptosis through up-regulation of PPAR-delta expression which is associated with p-STAT3 up-regulation and ROS/superoxide induction. Rh2 may be potentially useful in the treatment of prostate cancer.

Investigation of the pronounced erythropoietin-induced reduction in hyperglycemia in type 1-like diabetic rats.

Erythropoietin (EPO) is known to stimulate erythropoiesis after binding with its specific receptor. In clinics, EPO is widely used in hemodialyzed patients with diabetes. However, changes in the expression of the erythropoietin receptor (EPOR) under diabetic conditions are still unclear. Therefore, we investigated EPOR expression both in vivo and in vitro. Streptozotocin-induced type 1-like diabetic rats (STZ rats) were used to evaluate the blood glucose-lowering effects of EPO. The expression and activity of the transducer and activator of transcription 3 (STAT3), the potential signaling molecule, was investigated in cultured rat skeletal myoblast (L6) cells incubated in high-glucose (HG) medium to mimic the in vivo changes. The EPO-induced reduction in hyperglycemia was more pronounced in diabetic rats. The increased EPOR expression in the soleus muscle of diabetic rats was reversed by the reduction in hyperglycemia. Glucose uptake was also increased in high-glucose (HG)-treated L6 cells. Western blotting results indicated that the EPO-induced hyperglycemic activity was enhanced mainly through an increase in EPOR expression. Increased EPOR expression was associated with the enhanced nuclear expression of STAT3 in HG-exposed L6 cells. In addition, treatment with siRNA specific to STAT3 reversed the increased expression of EPOR observed in these cells. Treatment with Stattic at a dose sufficient to inhibit STAT3 reduced the expression level of EPOR in STZ rats. In conclusion, the increased expression of EPOR by hyperglycemia is mainly associated with an augmented expression of nuclear STAT3, which was identified both in vivo and in vitro in the present study.

Phosphorylation of signal transducer and activator of transcription 3 induced by hyperglycemia is different with that induced by lipopolysaccharide or erythropoietin via receptor­coupled signaling in cardiac cells.

The signal transducer and activator of transcription 3 (STAT3) is known to be involved in hypertrophy and fibrosis in cardiac dysfunction. The activation of STAT3 via the phosphorylation of STAT3 is required for the production of functional activity. It has been established that lipopolysaccharide (LPS)­induced phosphorylation of STAT3 in cardiomyocytes primarily occurs through a direct receptor­mediated action. This effect is demonstrated to be produced rapidly. STAT3 in cardiac fibrosis of diabetes is induced by high glucose through promotion of the STAT3­associated signaling pathway. However, the time schedule for STAT3 activation between LPS and high glucose appears to be different. Therefore, the difference in STAT3 activation between LPS and hyperglycemia in cardiomyocytes requires elucidation. The present study investigated the phosphorylation of STAT3 induced by LPS and hyperglycemia in the rat cardiac cell line H9c2. Additionally, phosphorylation of STAT3 induced by erythropoietin (EPO) via receptor activation was compared. Then, the downstream signals for fibrosis, including the connective tissue growth factor (CTGF) and matrix metalloproteinase (MMP)­9, were determined using western blotting, while the mRNA levels were quantified. LPS induced a rapid elevation of STAT3 phosphorylation in H9c2 cells within 30 min, similar to that produced by EPO. However, LPS or EPO failed to modify the mRNA level of STAT3, and/or the downstream signals for fibrosis. High glucose increased STAT3 phosphorylation to be stable after a long period of incubation. Glucose incubation for 24 h may augment the STAT3 expression in a dose­dependent manner. Consequently, fibrosis­associated signals, including CTGF and MMP­9 protein, were raised in parallel. In the presence of tiron, an antioxidant, these changes by hyperglycemia were markedly reduced, demonstrating the mediation of oxidative stress. Therefore, LPS­ or EPO­induced STAT3 phosphorylation is different compared with that caused by high glucose in H9c2 cells. Sustained activation of STAT3 by hyperglycemia may promote the expression of fibrosis­associated signals, including CTGF and MMP­9, in H9c2 cells. Therefore, regarding the cardiac dysfunctions associated with diabetes and/or hyperglycemia, the identification of nuclear STAT3 may be more reliable compared with the assay of phosphorylated STAT3 in cardiac cells.

Increase in renal erythropoietin receptors in diabetic rats is mainly mediated by hyperglycemia associated with the STAT3/GATA-1 signaling pathway.

Erythropoietin (EPO) produces cytoprotection in many tissues by activating the EPO receptor (EPOR); these effects include anti-oxidative stress and the inhibition of apoptosis in renal tubular cells. Moreover, EPO is clinically used in diabetic patients who suffer from chronic renal disease. However, the effect of hyperglycemia on renal EPOR expression remains unknown. Therefore, we determined the changes in renal EPOR expression in diabetic rats and investigated the role of potential factors using cultured cells. Streptozotocin-induced hyperglycemic rats (STZ rats) treated with insulin or phloridzin to correct hyperglycemia were used to investigate treated with insulin or phloridzin to correct hyperglycemia were used to investigate the EPOR changes. Potential factors, including the transducer and activator of transcription 3 (STAT3) and GATA binding protein 1 (GATA1), were identified in cultured NRK-49F cells after incubation with high glucose (HG) levels to mimic diabetic animals. Renal EPOR expression was reduced by insulin and phloridzin in STZ rats, and hyperglycemia recovered. An increase in EPORs was also reproduced in hyperglycemia-exposed NRK-49F cells and HK-2 cells, which showed a higher expression of STAT3 or GATA1. Furthermore, the application of siRNA specific to STAT3 or GATA1 attenuated the higher expression of EPORs in HG-incubated NRK-49F cells. Moreover, stattic administered at a dose that was sufficient to inhibit STAT3 restored the level of renal EPORs in diabetic rats. Taken together, the expression of renal EPORs is increased by hyperglycemia via the STAT3/GATA1 pathway and has been characterized in both diabetic rats and cultured cells.

Telmisartan Activates PPARδ to Improve Symptoms of Unpredictable Chronic Mild Stress-Induced Depression in Mice.

Major depression is a common mental disorder that has been established to be associated with a decrease in serotonin and/or serotonin transporters in the brain. Peroxisome proliferator-activated receptor δ (PPARδ) has been introduced as a potential target for depression treatment. Telmisartan was recently shown to activate PPARδ expression; therefore, the effectiveness of telmisartan in treating depression was investigated. In unpredictable chronic mild stress (UCMS) model, treatment with telmisartan for five weeks notably decrease in the time spent in the central and the reduced frequency of grooming and rearing in open filed test (OFT) and the decreased sucrose consumption in sucrose preference test (SPT) compared with the paradigms. Telmisartan also reversed the decrease in PPARδ and 5-HTT levels in the hippocampus of depression-like mice. Administration of PPARδ antagonist GSK0660 and direct infusion of sh-PPARδ into the brain blocked the effects of telmisartan on the improvement of depression-like behavior in these mice. Moreover, telmisartan enhanced the expression of PPARδ and 5HTT in H19-7 cells. In conclusion, the obtained results suggest that telmisartan improves symptoms of stress-induced depression in animals under chronic stress through activation of PPARδ. Therefore, telmisartan may be developed as a potential anti-depressant in the future.

The Dietary Furocoumarin Imperatorin Increases Plasma GLP-1 Levels in Type 1-Like Diabetic Rats.

Imperatorin, a dietary furocoumarin, is found not only in medicinal plants, but also in popular culinary herbs, such as parsley and fennel. Recently, imperatorin has been shown to activate GPR119 in cells. Another GPR, GPR131, also called TGR5 or G-protein-coupled bile acid receptor 1 (GPBAR1), is known to regulate glucose metabolism. Additionally, TGR5 activation increases glucagon-like peptide (GLP-1) secretion to lower blood sugar levels in animals. Therefore, the present study aims to determine whether the effects of imperatorin on GLP-1 secretion are mediated by TGR5. First, we transfected cultured Chinese hamster ovary cells (CHO-K1 cells) with the TGR5 gene. Glucose uptake was confirmed in the transfected cells using a fluorescent indicator. Moreover, NCI-H716 cells, which secrete GLP-1, were used to investigate the changes in calcium concentrations and GLP-1 levels. In addition, streptozotocin (STZ)-induced type 1-like diabetic rats were used to identify the effects of imperatorin in vivo. Imperatorin dose-dependently increased glucose uptake in CHO-K1 cells expressing TGR5. In STZ diabetic rats, similar to the results in NCI-H716 cells, imperatorin induced a marked increase of GLP-1 secretion that was reduced, but not totally abolished, by a dose of triamterene that inhibited TGR5. Moreover, increases in GLP-1 secretion induced by imperatorin and GPR119 activation were shown in NCI-H716 cells. We demonstrated that imperatorin induced GLP-1 secretion via activating TGR5 and GPR119. Therefore, imperatorin shall be considered as a TGR5 and GPR119 agonist.

Glycyrrhizic acid increases glucagon like peptide-1 secretion via TGR5 activation in type 1-like diabetic rats.

Glycyrrhizic acid (GA) is belonged to triterpenoid saponin that is contained in the root of licorice and is known to affect metabolic regulation. Recently, glucagon like peptide-1 (GLP-1) has widely been applied in diabetes therapeutics. However, the role of GLP-1 in GA-induced anti-diabetic effects is still unknown. Therefore, we are interested in understanding the association of GLP-1 with GA-induced effects. In type 1-like diabetic rats induced by streptozotocin (STZ-treated rats), GA increased the level of plasma GLP-1, which was blocked by triamterene at a dose sufficient to inhibit Takeda G-protein-coupled receptor 5 (TGR5). The direct effect of GA on TGR5 has been identified using the cultured Chinese hamster ovary cells (CHO-K1 cells) transfected TGR5 gene. Moreover, in intestinal NCI-H716 cells that secreted GLP-1, GA promoted GLP-1 secretion with a marked elevation of calcium levels. However, both effects of GA were reduced by ablation of TGR5 with siRNA in NCI-H716 cells. Therefore, we demonstrated that GA can enhance GLP-1 secretion through TGR5 activation.

Ginsenoside Rh2 Improves Cardiac Fibrosis via PPARδ-STAT3 Signaling in Type 1-Like Diabetic Rats.

Ginsenoside Rh2 (Rh2) is an active principal ingredient contained in ginseng (Panax ginseng Meyer), a medicinal herb used to enhance health worldwide. The present study is designed to investigate the effect of Rh2 on myocardial fibrosis in diabetic rats. In a streptozotocin-induced model of type-1 diabetic rats (STZ-diabetic rats), the increased fasting blood glucose levels and heart weight/body weight (HW/BW) ratio were substantially alleviated by Rh2. Moreover, Rh2 improved cardiac performance in STZ-diabetic rats. Histological results from Masson staining showed that Rh2 attenuated cardiac fibrosis in STZ-diabetic rats. The effects of Rh2 were reversed by GSK0660 at a dose sufficient to inhibit peroxisome proliferator-activated receptor δ (PPARδ) in STZ-diabetic rats. The role of PPARδ was subsequently investigated in vitro. Rh2 restored the decreased PPARδ expression level in high glucose-cultured cardiomyocytes. Moreover, increased protein levels of fibrotic signals, including signal transducer and activator of transcription 3 (STAT3), connective tissue growth factor (CCN2) and fibronectin, were reduced by Rh2 in high glucose-cultured cardiomyocytes. These effects of Rh2 were reversed by GSK0660 or siRNA specific for PPARδ Taken together, PPARδ activation may inhibit STAT3 activation to reduce CCN2 and fibronectin expression in diabetic rats with cardiac fibrosis. Moreover, Rh2 improves cardiac function and fibrosis by increasing PPARδ signaling. Therefore, Rh2 is suitable to develop as an alternative remedy for cardiac fibrosis.

Ursolic acid activates the TGR5 receptor to enhance GLP-1 secretion in type 1-like diabetic rats.

Endogenous Takeda G-protein-coupled receptor 5 (TGR5), G-protein-coupled bile acid receptor 1 (GPBAR1), regulates glucose metabolism. In animals, TGR5 activation by a chemical agonist may increase incretin secretion and reduce the blood sugar level. Recently, betulinic acid has been suggested to activate TGR5. Ursolic acid is a well-known pentacyclic triterpenoid that is similar to betulinic acid. It is of special interest to determine the potential effect of ursolic acid on TGR5. Therefore, we transfected cultured Chinese hamster ovary (CHO-K1) cells with the TGR5 gene. The functions of the transfected cells were confirmed via glucose uptake using a fluorescent indicator. Moreover, NCI-H716 cells that secreted incretin were also investigated, and the glucagon-like peptide (GLP-1) levels were quantified using ELISA kits. In addition, streptozotocin (STZ)-induced type 1-like diabetic rats were used to identify the effect of ursolic acid in vivo. Ursolic acid concentration dependently increased glucose uptake in CHO-K1 cells expressing TGR5. In NCI-H716 cells, ursolic acid induced a concentration-dependent elevation in GLP-1 secretion, which was inhibited by triamterene at the effective concentrations to block TGR5. Ursolic acid also increased the plasma GLP-1 level via TGR5 activation, which was further characterized in vivo with type 1-like diabetic rats. Moreover, ursolic acid is more effective than betulinic acid in reduction of hyperglycemia and increase of GLP-1 secretion. Therefore, we demonstrated that ursolic acid can activate TGR5, enhancing GLP-1 secretion in vitro and in vivo. Therefore, ursolic acid is suitable for use in TGR5 activation.

Loperamide-induced Cardiac Depression Is Enhanced by Hyperglycemia: Evidence Relevant to Loperamide Abuse.

BACKGROUND AND AIMS: Cardiac dysryhthmias and death are reported after loperamide abuse. The mechanism of death is not clear and cardiac depression may play a role in this mechanism. Loperamide is widely used as an agonist of the µ-opioid receptor (MOR) in clinical practice. In skeletal muscle, an increase in MOR in response to hyperglycemia is largely attributable to higher expression of the transducer and activator of transcription 3 (STAT3), which binds to the promoter of the MOR genes. Therefore, we investigated the changes in cardiac MOR caused by hyperglycemia both in vivo and in vitro. METHODS: Streptozotocin-induced type 1-like diabetic rats (STZ rats) were used to estimate cardiac performance and changes in cardiac MOR under the influence of loperamide. STAT3 was measured in cultured cardiomyocytes under high glucose (HG) to mimic the in vivo changes. RESULTS: Loperamide-induced reduction of cardiac performance was more marked in STZ rats than in normal rats. The increased MOR in the hearts of STZ rats was reversed by the reduction of hyperglycemia. Higher MOR expression paralleled the increase in STAT3 in cardiomyocytes under HG and was reversed by siRNA of STAT3. Stattic at a dose sufficient to inhibit STAT3 reduced MOR both in vivo and in vitro. CONCLUSION: Cardiac depression induced by loperamide is enhanced by hyperglycemia due to higher MOR expression, which is associated with higher expression of STAT3 in the heart. These results suggest that loperamide abuse is particularly dangerous for individuals with hyperglycemia.

Investigation of triamterene as an inhibitor of the TGR5 receptor: identification in cells and animals.

BACKGROUND: G-protein-coupled bile acid receptor 1 (GPBAR1, also known as TGR5) has been shown to participate in glucose homeostasis. In animal models, a TGR5 agonist increases incretin secretion to reduce hyperglycemia. Many agonists have been developed for clinical use. However, the effects of TGR5 blockade have not been studied extensively, with the exception of studies using TGR5 knockout mice. Therefore, we investigated the potential effect of triamterene on TGR5. METHODS: We transfected the TGR5 gene into cultured Chinese hamster ovary cells (CHO-K1 cells) to express TGR5. Then, we applied a fluorescent indicator to examine the glucose uptake of these transfected cells. In addition, NCI-H716 cells that secrete incretin were also evaluated. Fura-2, a fluorescence indicator, was applied to determine the changes in calcium concentrations. The levels of cyclic adenosine monophosphate (cAMP) and glucagon-like peptide (GLP-1) were estimated using enzyme-linked immunosorbent assay kits. Moreover, rats with streptozotocin (STZ)-induced type 1-like diabetes were used to investigate the effects in vivo. RESULTS: Triamterene dose dependently inhibits the increase in glucose uptake induced by TGR5 agonists in CHO-K1 cells expressing the TGR5 gene. In cultured NCI-H716 cells, TGR5 activation also increases GLP-1 secretion by increasing calcium levels. Triamterene inhibits the increased calcium levels by TGR5 activation through competitive antagonism. Moreover, the GLP-1 secretion and increased cAMP levels induced by TGR5 activation are both dose dependently reduced by triamterene. However, treatment with KB-R7943 at a dose sufficient to block the Na+/Ca2+ exchanger (NCX) failed to modify the responses to TGR5 activation in NCI-H716 cells or CHO-K1 cells expressing TGR5. Therefore, the inhibitory effects of triamterene on TGR5 activation do not appear to be related to NCX inhibition. Blockade of TGR5 activation by triamterene was further characterized in vivo using the STZ-induced diabetic rats. CONCLUSION: Based on the obtained data, we identified triamterene as a reliable inhibitor of TGR5. Therefore, triamterene can be developed as a clinical inhibitor of TGR5 activation in future studies.

Cryptotanshinone Inhibits STAT3 Signaling to Alleviate Cardiac Fibrosis in Type 1-like Diabetic Rats.

Cryptotanshinone is an active principal ingredient isolated from Salvia miltiorrhiza (Danshen), a medicinal plant used in China to treat cardiac disorders. The objective of this study was to investigate the effect of cryptotanshinone on myocardial fibrosis in diabetic rats. In streptozotocin-induced type 1 diabetic model hyperglycemic rats (STZ-treated rats), fasting blood glucose levels and heart weight/body weight ratio were markedly increased but both were not modified by cryptotanshinone. Additionally, cardiac performance in catheterized STZ-treated rats was improved. The histological results from Masson staining showed that cryptotanshinone attenuated cardiac fibrosis in STZ-treated rats. Moreover, both the mRNA and protein levels of the signal transducer and activator of transcription 3 (STAT3), matrix metalloproteinase-9, and connective tissue growth factor were reduced by cryptotanshinone in high glucose-cultured cardiomyocytes, similar to the reductions observed in the hearts of STZ-treated rats. In conclusion, while STAT3 regulates matrix metalloproteinase-9 and connective tissue growth factor expression in diabetic rats with cardiac fibrosis, cryptotanshinone inhibited fibrosis to improve cardiac function by suppressing the STAT3 pathway. Cryptotanshinone is suitable as an alternative remedy for therapy of cardiac fibrosis. Copyright © 2017 John Wiley & Sons, Ltd.

Lung damage induced by hyperglycemia in diabetic rats: The role of signal transducer and activator of transcription 3 (STAT3).

Increased evidence has shown that diabetes can be a risk factor for pulmonary fibrosis. The objective of this study was to use streptozotocin-induced diabetic rats (STZ rats) to assess the possible signals associated with lung damage in diabetic disorders. The expression levels of signal transducer and activator of transcription 3 (STAT3) and connective tissue growth factor (CTGF) in lung tissues were measured through Western blot analysis and real-time PCR. Additionally, the potential mechanisms were confirmed in cultured rat lung cell line (L2) incubated in high-glucose (HG) medium to mimic the in vivo changes. The pathological changes in the lung tissues of STZ rats were characterized using the bleomycin-treated tissues as reference. Moreover, the higher expression levels of STAT3 and CTGF in the lung tissues of STZ rats were reversed by treating the hyperglycemia. CTGF expression increased following the higher expression of STAT3 in the cultured L2 cells exposed to HG, and this change was reversed by siRNA treatment specific for STAT3. Stattic, at a dose sufficient to inhibit STAT3, reduced the CTGF levels in the lungs of STZ rats. In conclusion, STAT3 enhanced CTGF expression in a type-1 diabetes model associated with lung damage. Thus, STAT3 inhibitors may be developed to improve diabetes-induced lung damage in the future.

Amarogentin ameliorates diabetic disorders in animal models.

Amarogentin is a bitter-tasting secoiridoid glycoside isolated from an herb. Inhibition of aldose reductase by amarogentin has been documented as an antidiabetic action. However, the mechanisms of action of amarogentin in diabetic disorders remain unknown. The present study employed streptozotocin-induced type 1 diabetic (T1DM) rats to investigate the antihyperglycemic action of amarogentin. Changes in the protein expression of glucose transporter 4 (GLUT4) and phosphoenolpyruvate carboxykinase (PEPCK) in skeletal muscle and liver, respectively, were also detected by Western blotting. Additionally, a type 2 diabetes (T2DM) animal model induced using a fructose-rich diet was also applied to assess the effect of amarogentin on insulin resistance according to the homeostasis model assessment-insulin resistance (HOMA-IR). Amarogentin dose-dependently attenuated hyperglycemia in the T1DM rats lacking insulin. The action of amarogentin was further supported in rats administered the oral glucose tolerance test. Western blotting showed that amarogentin reversed the decreased GLUT4 level in skeletal muscle and reduced the elevated PEPCK expression in livers isolated from the T1DM rats. Moreover, amarogentin decreased the HOMA-IR and increased insulin sensitivity in the T2DM rats. These data show that amarogentin may ameliorate glucose homeostasis in diabetic rats, indicating its potential for future development as an antidiabetic drug.

Erythropoietin ameliorates hyperglycemia in type 1-like diabetic rats.

BACKGROUND: Erythropoietin (EPO) is widely used in diabetic patients receiving hemodialysis. The role of EPO in glucose homeostasis remains unclear. Therefore, we investigated the effect of EPO on hyperglycemia in rats with type 1-like diabetes. METHODS: Rats with streptozotocin-induced type 1-like diabetes (STZ rats) were used to estimate the blood glucose-lowering effects of EPO, and changes in the expression levels of glucose transporter 4 (GLUT4) and the hepatic enzyme phosphoenolpyruvate carboxykinase (PEPCK) were identified by Western blot analysis. RESULTS: EPO attenuated the hyperglycemia in the STZ rats in a dose-dependent manner without altering the hematopoietic parameters, including the hematocrit and number of red blood cells. The involvement of the EPO receptor (EPOR) was identified using EPOR-specific antibodies. In addition, injection of EPO enhanced the glucose utilization, which was assessed using an intravenous glucose tolerance test in rats. However, blood insulin was not changed by EPO in this assay, showing the insulinotropic action of EPO. Moreover, EPO treatment increased the insulin sensitivity. Western blots indicated that the phosphorylation of AMP-activated protein kinase was enhanced by EPO to support the signaling caused by EPOR activation. Furthermore, the decrease in the GLUT4 level in skeletal muscle was reversed by EPO, and the increase in the PEPCK expression in liver was reduced by EPO, as shown in STZ rats. CONCLUSION: Taken together, the results show that EPO injection may reduce hyperglycemia in diabetic rats through activation of EPO receptors. Therefore, EPO is useful for managing diabetic disorders, particularly hyperglycemia-associated changes. In addition, EPO receptor will be a good target for the development of antihyperglycemic agent(s) in the future.