Effects of metoprolol on phosphorylated connexin 43 expression in myocardial tissues and apoptosis of myocardial cells in heart failure rats

AIM：To explore the in vivo effects of metoprolol on the expression of phosphorylated connexin 43 (p-Cx43) in myocardial tissues and the apoptosis of myocardial cells in rats with heart failure (HF).METHODS：One hundred Sprague-Dawley rats were randomly divided into 5 groups (each n=20): sham group, HF group, low-dose (1.25 mg·kg-1·d-1) metoprolol treatment (MetoA) group, middle-dose (5 mg·kg-1·d-1) metoprolol treatment (MetoB) group and high-dose (20 mg·kg-1·d-1) metoprolol treatment (MetoC) group.The rats in HF group and metoprolol treatment groups were subject to abdominal aortic ligation, and different doses of metoprolol were given 4 weeks later till 8 weeks after operation.Echocardiography was conducted to monitor the hemodynamic parameters at the 4th and 8th weeks, and the rat hearts were taken at the 8th week after operation.The morphological changes and the proliferation of collagen fibers in myocardial tissues were observed by HE and Masson staining, respectively.The expression level of p-Cx43 was detected by Western blotting and the apoptosis of myocardial cells was assessed by TUNEL method.The relationship between p-Cx43 expression level and apoptotic index was analyzed by Pearson’s correlation.RESULTS：(1) Echocardiography showed that metoprolol could effectively improved cardiac hemodynamics in HF rats, and pathological findings suggested that metoprolol could effectively reverse HF-induced cardiac remodeling in a dose-dependent manner within the therapeutic dose range.(2) Western blotting showed that p-Cx43 expression in HF group was significantly higher than that in sham group (P<001), and that in all metoprolol treatment groups was significantly decreased compared with HF group (P<005 or P<001), among which pairwise comparisons also showed significant differences (P<001).(3) The myocardial apoptotic index in HF group ［(51.17±6.94)%］ was significantly increased compared with sham group ［(4.62±160)%, P<001］.Compared with HF group, myocardial apoptotic indexes in MetoA group ［(40.60±4.15)%］, MetoB group ［(30.66±4.00)%］ and MetoC group ［(22.24±5.69)%］ were significantly decreased (P<001), among which pairwise comparisons also showed significant differences (P<001).(4) The expression level of p-Cx43 was positively correlated with the apoptotic index (r=0.905, P<001).CONCLUSION: The mechanism of metoprolol against HF-induced myocardial apoptosis may be related to inhibition of p-Cx43 expression.     

Down-regulation of PPAR-γ expression by p300-mediated histone acetylation induces glucose-lipid metabolism disorder in cardiomyocytes of GDM offspring mice

AIM: To investigate the effect of gestational diabetes mellitus (GDM) on glucose-lipid metabolism in the offspring mice and the underlying mechanisms. METHODS: Wild-type female mice were intraperitoneally injected with streptozotocin at 30 mg/kg in the second trimester of pregnancy to establish GDM model. Normal saline was used as control. F1 offspring mice were fed for 8 weeks after birth. The blood glucose and lipid levels were detected randomly. The mRNA levels of p300 and p300/CBP-associated factor (PCAF) were detected by qPCR. The expression of peroxisome proliferator-activated receptor-γ (PPAR-γ), glucose transporter typer 4 (GLUT-4) and medium-chain acyl-CoA dehydroge-nase (MCAD) at mRNA and protein levels was determined by qPCR and Western blot. ChIP-qPCR was employed to analyze the binding status of p300 with the promoter of PPAR-γ and the acetylation level of histone H3 in the promoter region of PPAR-γ. RESULTS: Blood glucose and total cholesterol levels were significant increased in the offspring mice (P<0.05). The expression levels of p300, PPAR-γ, GLUT-4 and MCAD were decreased compared with the control group (P<0.05). Binding affinity of p300 with the promoter of PPAR-γ was reduced (P<0.05). The level of acetylated histone H3 in the promoter region of PPAR-γ was decreased significantly (P<0.05). CONCLUSION: Regulation of PPAR-γ expression by p300 may induce glucose-lipid metabolism disorder in the cardiomyocytes of GDM offspring mice.     

Effects of liraglutide on adiponectin and insulin resistance in rats with non-alcoholic fatty liver disease

AIM：To investigate the effects of glucagon-like peptide 1 analog, liraglutide, on adiponectin and insulin resistance in the rats with diet-induced non-alcoholic fatty liver disease (NAFLD). METHODS：Male rats were randomly divided into 3 groups:normal diet (ND) group (n=10), high-fat diet (HFD) group (n=10), and HFD with intraperitoneal injection of liraglutide group (n=10, first 12 weeks with HFD, later 4 weeks with liraglutide). All treatments continued for 16 weeks, and then the rats were killed ethically and the blood samples and liver tissues were collected. The levels of alanine aminotransferase (ALT), fasting blood glucose (FBG), total cholesterol (TC) and triglyceride (TG) were detected by a biochemical automatic analyzer. The levels of free fatty acids (FFAs), fasting insulin (FINS) and adiponectin were measured by RIA and ELISA. RESULTS：Compared with HFD group, the body weight, liver index, homeostasis assessment-insulin resistance (HOMA-IR), the serum levels of TG, TC, ALT and FBG, and the liver levels of TG, TC and FFAs in the rats in liraglutide group were apparently lower, the degree of hepatic steatosis and inflammatory activity significantly decreased (P<0.05), and the level of adiponectin in the serum and liver homogenate increased ob-viously (P<0.05). The level of adiponectin in the liver homogenate was negatively correlated with the levels of FFAs in the liver homogenate. CONCLUSION:Liraglutide is beneficial for NAFLD rats to improve insulin resistance and reduce hepatic steatosis by increasing the level of adiponectin in the serum and liver tissues.     

Effects of FK506 on anti-glomerular basement membrane nephritis in rats

AIM:To investigate the effects of FK506 on anti-glomerular basement membrane(GBM) nephritis in rats. METHODS: Anti-GBM nephritis model was elaborated by rabbit anti-rat GBM antibody injection in SD rats in this study. The rats were divided into three groups: FK506 treated group(0.5 mg·kg^-1·d^-1, sc), untreated nephritis control group and normal control group. FK506 was administered daily six hours after injection of anti-GBM IgG. All the rats were observed urinary protein at the 4th day, the 14th day and the 21st day. At the same time, the kidney specimens were collected, and T cell transforming function was also monitored.RESULTS:Rats injected with rabbit anti-GBM Ab developed heavy proteinuria by 4 days, and serum creatinine and serum urea appeared which kept on the rising. Glmerular hypercellularity, crescents, and protein casts were observed in nephritic rats. By electron microscopy, the thickening of GBM and loss of foot processes were seen. T cell transforming function was higher than normal. But, all pathological changes obviously turned for the better in FK506 treated group. CONCLUSION:FK506 could inhibit the progression of rat anti-GBM nephritis.     

Effects of Shan He Jian Fei Granule on the expression of leptin,adiponectin and resistin in obese rats induced by high-fat feeding

AIM: To study the effects of Shan He Jian Fei Granule (SHJFG), a Chinese medicine, on weight-reduction and fat-decrease in adiposity rats, and to observe the changes of leptin, adiponectin and resistin. METHODS: After the models were prepared successfully, the rats were randomly divided into four groups: normal group, low dosage, middle dosage, high dosage and obesity control group. After 8 weeks interference with SHJFG, the weight and the naso-anal length of each rat was measured and Lees index were calculated. The levels of TG, TC, LDL, HDL and leptin in serum were carefully determined. The gene expressions of adiponectin and resistin in adipose tissues of rats were determined by RT-PCR. RESULTS: Compared to high-fat diet group, the body weights, the Lees indexes, the weight of fat tissues and levels of TG, TC, LDL and leptin in SHJFG groups significantly decreased (P<0.05). Adiponectin mRNA expression in SHJFG group significantly increased (P<0.05), and the resistin mRNA expression also significantly increased. CONCLUSION: SHJFG significantly decreases the body weight and the serum levels of TG, TC and LDL in obese rats. The effects of SHJFG in raising the mRNA expressions of adiponectin and resistin in fat tissues may be one of the main role that results in lowering body weight in obese rats.     

Correlation among plasma leptin,resistin an type 2 diabetes mellitus

AIM：To assess the association of resistin,obesity,serum lipid levels and insulin resistance with plasma leptin.METHODS：The concentrations of fasting serum glucose,insulin,lipid profiles,plasma resistin and leptin were assayed in 80 cases (including 37 controls with normal glucose tolerance and 43 patients with type 2 diabetes mellitus).RESULTS：Fasting plasma leptin level was positively correlated with sex,body mass index (BMI),waist circumference (WC),waist-hip ratio and fasting serum insulin (F Ins) （P<0.01）.Fasting plasma leptin level was negatively correlated with insulin sensitivity index (r=-0.373,P<0.01).There was no correlation between the concentrations of plasma leptin and FPG,TG,TC and resistin （P>0.05）.CONCLUSION：Fasting plasma leptin level is positively correlated with obesity and insulin resistance,not resistin.Leptin may play a role in the pathogenesis of type 2 diabetes mellitus.     

High-fat diet affects plasma membrane GLUT4 content in skeletal muscle from Sprague-Dawley rats

AIM: To explore the change of the amount of GLUT4 protein at the plasma membrane of the rat skeletal muscle after high-fat feeding. METHODS: The animals were divided into three groups (ten for each): group I: control; group II: high-fat feeding; group III: high-fat feeding + dietary treatment. The rat model of insulin resistance (IR) was made by feeding high-fat diet for eight weeks. And then insulin-resistant rats were fed with chow diet for 4 weeks. Fasting plasma glucose and fasting serum insulin levels were measured before and after dietary treatment, respectively. Insulin treatment was achieved by intraperitoneal injection of insulin (10 unit insulin per kg body weight) 15 minutes before killing the animals. The right hindlimb skeletal muscle was rapidly dissected. Then the expression of GLUT4 protein at the plasma membrane in all the animals was assessed with Western bloting. RESULTS: The GLUT4 content at the plasma membrane in high-fat-fed rat skeletal muscle was significantly lower (about 31%) than that in controls (P<0.01). Dietary treatment partly corrected fasting blood glucose [from(6.20±0.39)mmol/L to(5.78±0.74)mmol/L]and fasting serum insulin levels [from(17.19±1.93)mU/L to(11.68±1.28)mU/L] and increased the GLUT4 content at the plasma membrane by 1.14-fold in insulin-resistant rat skeletal muscle. CONCLUSION: High-fat feeding induces IR in Sprague-Dawley rats. The mechanism may be involved in decreased cell-surface level of GLUT4 through affecting intracellular insulin signaling and then decreasing GLUT4 trafficking.     

“2006中国蔬菜产业可持续发展研讨会”在武汉召开

由中国园艺学会主办，湖北省园艺学会、国家蔬菜改良中心华中分中心、华中农业大学园艺林学学院共同承办的“2006中国蔬菜产业可持续发展研讨会”于2006年4月13-15日在湖北武汉华中农业大学举行，来自全国17个省（市）和自治区47个单位的近120名代表出席这次全国性会议。会议开幕式由中国园艺学会蔬菜专业委员会主任、中国农业科学院蔬菜花卉研究所副所长孙日飞研究员主持。华中农业大学副校长李名家教授、武汉市张学忙副市长、科技部农村司魏勤芳处长、农业部科技司刘艳处长、武汉市科协黄和平主席和中国园艺学会理事长方智远院士先后致辞。     

Association of rs2162459 polymorphism in CYP7A1 gene with effectiveness of atorvastatin in northern Chinese Han population

AIM：To investigate the association of the polymorphism of rs2162459 locus in cytochrome P-450, family 7, subfamily A, polypeptide 1 (CYP7A1) gene, encoding cholesterol 7α-hydroxylase, with the effectiveness of atorvastatin in northern Chinese Han population. METHODS：Clinical data and blood samples of 200 cases of hyperlipidemia patients were collected. The variants of rs2162459 in CYP7A1 gene were detected by multiplex SNaPshot technology. Several genetic models were constructed to analyse the association of gene polymorphism with the effectiveness of atorvastatin by logistic regression method. RESULTS：The polymorphism of rs2162459 was consistent with Hardy-Weinberg equilibrium. The baseline high-density lipoprotein cholesterol (HDL-C) concentrations in three genotypes of AA, GA and GG were (136±0.94) mmol/L, (1.16±0.38) mmol/L and (1.07±0.28) mmol/L, respectively (P<0.05). There were differences in the genotypic frequency and allelic frequency between atorvastatin effective and ineffective groups (both P<005). Significant differences in regulating HDL-C level among the three genotypes of rs2162459 were found after logistic regression. The results of additive model, generalized model and dominant model, presented as OR (95% CI), were 1.74 (1.09~2.77), 2.86 (1.13~7.25) and 2.21 (1.12~4.33), respectively. CONCLUSION: The baseline HDL-C level in the carriers of GG genotype is lower than that in the carriers of the other two genotypes. The HDL-C-elevating effect of atorvastatin on GG genotype carriers is more significant than that on AA genotype carriers.     

Effects of transplantation of adipose stromal vascular fraction on cardiac function in adriamycin-induced heart failure rats

AIM: To investigate the effects of transplantation of adipose stromal vascular fraction (SVF) on the cardiac function in adriamycin-induced heart failure rats. METHODS: SVF was isolated from adipose tissue of a Sprague-Dawley (SD) rat by collagenase digestion and marked with green fluorescent protein (GFP) in vitro. Twenty-eight SD rats were randomized into normal control group (n=8), adriamycin control group (n=10) and SVF treatment group (n=10). Adriamycin at dose of 15 mg/kg was intraperitoneally injected into the rats twice a week for 4 weeks to induce heart failure. SVF cells (05 mL, 1×107/L) were injected via penis vein, and PBS vehicle was injected into the control animals in the same way. Four weeks later, the cardiac function was determined by multichannel physiologic recorder via cardiac catheterization. SVF was demonstrated in the myocardium by frozen section fluorescence microscopy. The CD31 expression was determined by an immunohistochemical test. RESULTS: Compared with adriamycin control group, SVF transplantation increased left ventricular peak systolic pressure ［LVSP, （13565±21.58) mmHg vs (10558±2262) mmHg, P<005］, left ventricular pressure maximal rise rate ［+dp/dtmax, (4 81565±56624) mmHg/s vs (3 53550±46528) mmHg/s, P<005］, and left ventricular pressure maximal decline rate ［-dp/dtmax, (3 67756±46775) mmHg/s vs (2 73865±51251) mmHg/s, P<005］. The results of the CD31 immunohistochemical test showed that the nuclear staining and granule distribution were more uniform, and the number of blood vessels per visual field increased in SVF treatment group as compared with adriamycin control group (P<005). CONCLUSION: SVF transplantation improves the cardiac function in the rat model of heart failure, possibly and partly through the promotion of myocardial neovascularization.     

Apoptosis in diabetic foot ulcers and human fibroblast cells treated with AGEs

AIM：To investigate cell apoptosis in diabetic foot ulcers and the effect of advanced glycosylation end products (AGEs) on apoptosis in human fibroblast cells. METHODS：Diabetic foot patients (n=18) and 18 age-matched non-diabetic controls were recruited. The clinical and biochemical features were compared by statistics methods. Skin biopsies were obtained from foot. Cleaved caspase-3 was measured by immunohistochemistry using the technique of streptavidin-biotin complex (SABC) staining. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) technique was used to detect apoptosis of the skin tissues. Human primary foreskin fibroblasts were isolated and cultured in the presence of 5.6 mmo/L glucose, 25 mmo/L glucose, fluctuant glucose (changing the glucose from 5.6 mmo/L to 25 mmo/L every 8 h) or AGEs (150 mg/L, containing 5.6 mmo/L glucose). After 72 h treatment, Western blotting was used to determine the levels of the apoptotic protein cleaved-caspase-3. Other cells were trypsinized, washed with cold PBS and incubated with PI and Annexin V-FITC, then analyzed by flow cytometry to detect cell apoptosis. RESULTS：Diabetic patients had higher levels of fasting blood glucose (FBG), 2-hour postprandial blood glucose (2 h PBG) and glycosylated hemoglobin A1c (HbA1c), and longer wound duration. The protein level of cleaved caspase-3 was significantly higher in diabetic group, suggesting that apoptosis was increased in diabetic skin tissues. TUNEL analysis showed that apoptotic index was higher in diabetic group compared with that in non-diabetic group (8.4%±1.5% vs 3.8%±08%), which further confirmed that cell apoptosis was increased in diabetic foot tissues. In human fibroblasts, the levels of cleaved caspase-3 in normal group, sustained high glucose group, fluctuant high glucose group and AGEs group were 080±0.13, 1.22±0.18, 1.46±0.32 and 1.83±0.25, respectively. The apoptotic rates detected by flow cytometry were 2.43%±0.19%, 2.89%±0.51%, 3.99%±0.24% and 6.83%±0.36%, respectively. Both the level of cleaved caspase-3 and the apoptotic rate in AGEs group were higher than those in normal glucose group and sustained high glucose group. CONCLUSION：Increased apoptosis in diabetic foot ulcers is one of the most important reasons for impaired wound healing. As compared to sustained high glucose and glucose fluctuations, AGEs induce greater apoptosis in human fibroblast cells.     

Effects of stretch on transient outward potassium and inward rectifier potassium current in cultured neonatal rat atrial myocytes

AIM：To investigate the effects of mechanical stretch on transient outward potassium current (Ito), inward rectifier potassium current (IK1) and action potential duration(APD) of cultured neonatal rat atrial myocytes. METHODS：Neonatal rat atrial myocytes were isolated and cultured on silicone sheeting with or without stretch for 24 h. The silicone membrane area was increased by 12% in stretched group. The cells without stretch served as control. Ito, IK1 and APD were recorded by the technique of whole-cell patch clamp. RESULTS：Compared with control group, Ito density in stretched myocytes was significantly reduced ［(16±04) pA/pF vs (121±29) pA/pF, P<001］, whereas IK1 density was increased ［(-108±08) pA/pF vs (-88±09) pA/pF, P<001］. The APDs at 50% and 90% levels of repolarization (APD50 and APD90) in the stretched cells were obviously decreased than those in non-stretched cells ［(105±14) ms vs (155±24) ms, (300±28) ms vs (563±36) ms, P<001］. CONCLUSION：Stretch stimulation leads to the reduction of Ito density, the increase in IK1 density and the shortness of APD in cultured rat atrial neonatal myocytes, which may contribute to atrial electrical remodeling induced by pressure overload.     

Panax quinquefolium saponins attenuate hypoxia/reoxygenation-induced injury in rat cardiomyocytes

AIM： To investigate the effects of Panax quinquefolium saponins (PQS) and calcineurin (CaN) signal pathway on cardiomyocyte injury induced by myocardial hypoxia/reoxygenation(H/R). METHODS：Cultured cardiomyocytes isolated from neonatal Sprague-Dawley rats were used to establish the H/R model. The cells were transfected with pCDB-CaN plasmid to overexpress CaN, or exposed to the CaN inhibitor FK506 to interfere the CaN expression. The cardiomyocytes were divided into control group, H/R group, PQS+H/R group, CaN+PQS+H/R group, pCDB+PQS+H/R group and FK506+PQS+H/R group. The apoptosis was analyzed by flow cytometry. The activity of CaN in the cardiomyocytes was detected. The protein expression of CaN was determined by Western blotting. RESULTS：Compared with control group, the apoptosis of the cardiomyocytes in CaN group was significantly increased. Compared with PQS+H/R group, the cell apoptosis, the expression of Bcl-2 and Bax, the activity of CaN and its protein expression in FK506 group were not significantly different. CONCLUSION：Inhibition of CaN activity reduces the H/R injury in cardiomyocytes. However, the mechanism of PQS protecting cardiomyocytes from H/R injury may not be associated with the CaN signaling pathway.     

Correlation of serum omentin-1 level and insulin resistance in rats

AIM：To establish the insulin resistance rat model for evaluating the correlation of omentin-1 level and insulin resistance. METHODS：SPF male Wistar rats (n=30) were randomly divided into normal control group (NC, n=15) and high-fat diet group (HF, n=15). The rats in NC group were fed with basic diet. The insulin resistant model was established by feeding the rats with high-fat diet in HF group. After 10 weeks, 5 rats in each group were assessed by the technique of hyperinsulinaemic-euglycaemic clamp. After the insulin resistant model was successfully established, the body weight and fasting blood glucose were detected. The concentration of fasting serum omentin-1 was analyzed by ELISA. Fasting serum insulin was measured by radioimmunoassay. RESULTS：No difference of fasting blood glucose between the 2 groups was observed. The level of fasting serum insulin in HF group was significantly higher than that in NC group (P<0.05). The level of serum omentin-1 in HF group were significantly decreased compared with NC group (P<0.01). Pearson’s correlation analysis showed that negative correlations between serum omentin-1 and fasting serum insulin (r=-0.654,P<0.01), serum omentin-1 and free fatty acid (r=-0.446, P<0.05) was found. CONCLUSION：In rats, serum omentin-1 level began to decrease at insulin resistance stage. As serum omentin-1 level decreased, the basal insulin level increased, indicating that decreased serum omentin-1 level may be an early factor of IR, diabetes and cardiovascular diseases.     

Role of asiatic acid in treatment of insulin resistance in mouse adipocytes

AIM：To investigate the effects of asiatic acid, one of triterpenoids from Psidium guajava leaves, on the proliferation and differentiation of 3T3-L1 preadipocytes, and glucose and lipid metabolism of insulin-resistant adipocytes. METHODS：The proliferation of 3T3-L1 preadipocytes was tested by MTT assay, and the accumulation of lipid droplets in differentiated preadipocytes was measured by oil red O staining. The insulin-resistant cell model was established by exposure of the cells to dexamethasone. The cellular glucose uptake was determined by glucose oxidase-peroxidase assay. The free fat acid (FFA) concentration was detected by colorimetric method. Secreted adiponectin were measured by ELISA. The protein levels of peroxisome proliferator-activated receptor γ (PPARγ) and protein tyrosine phosphatase 1B (PTP1B) in insulin-resistant adipocytes were analyzed by Western blotting. RESULTS：Compared with medium group, asiatic acid increased the proliferation of 3T3-L1 preadipocytes and inhibited their differentiation at a concentration range of 10~100 μmol/L (P<0.05 or P<0.01). At concentrations of 30 μmol/L and 100 μmol/L, asiatic acid enhanced cellular glucose uptake in the insulin-resistant adipocytes both in basic and insulin-stimulation states. Asiatic acid decreased FFA production (P<0.05), and down-regulated the protein expression of PTP1B (P<0.05, or P<0.01). However, no effect on the secretion of adiponectin and the protein expression of PPARγ was observed (P>0.05). CONCLUSION：Asiatic acid enhances glucose uptake and inhibits FFA production in insulin-resistant adipocytes via down-regulating the protein expression of PTP1B, all of which play the roles of increasing insulin signaling sensitivity to improve insulin resistance.     

Hypolipemic and hypoglycemic effects of total triterpenoids from Psidium guajava leaves on type 2 diabetic rats

AIM: To investigate the effects of total triterpenoids from Psidium guajava leaves (TTPGL) on blood glucose and lipids in type 2 diabetic rats. METHODS: The diabetic rats were induced by intraperitoneal injection of streptozotocin at dose of 35 mg/kg and feeding with high-fat diet. The animals were divided into 5 groups: diabetic model control group (model), TTPGL treatment groups (with the doses of 60, 120 and 240 mg/kg, respectively) and rosiglitazone treatment group (3 mg/kg). Another 12 normal SD rats were used as the normal controls. The rats received daily treatment for 6 weeks, and then the levels of fasting blood glucose (FBG), fasting insulin (FINS), triglyceride (TG), total cholesterol (TCH), free fatty acid (FFA), glycosylated hemoglobin (GHb) and glycosylated serum proteins (GSP) were measured. The protein expression of peroxisome proliferator-activated receptor γ (PPARγ) in adipose tissues was detected by Western blotting. RESULTS: Compared with normal control group, the levels of FBG, GHb and blood lipids were increased in type 2 diabetic rats. The FINS, insulin sensitivity index, and the protein expression of PPARγ in adipose tissues were decreased. Compared with model group, the levels of FBG and GSP were decreased,and the FINS, insulin sensitivity index, and the protein expression of PPARγ in adipose tissues significantly increased in TTPGL treatment groups (with the doses of 120 and 240 mg/kg). The levels of serum TG,TCH and FFA were significantly lower in TTPGL treatment groups (P<0.01 or P<0.05) as compared with the model controls. CONCLUSION: TTPGL decreases the levels of blood glucose and lipids in diabetic rats. TTPGL also increases serum insulin level and improves insulin sensitivity. The action mechanism of TTPGL may be related to the increase in the protein expression of PPARγ.     

Valproate enhances imatinib-induced apoptosis and down-regulates Bcr/Abl mRNA and phosphorylated protein kinase B in chronic myeloid leukemia cell line K562

AIM：To investigate the effects of valproate and imatinib on the apoptosis of chronic myeloid leukemic cell line K562. METHODS：K562 cells were divided into 3 groups and treated with valproate, imatinib and cotreatment, respectively. Cell cycle, apoptosis, the mRNA expression of Bcr/Abl, total protein kinase B (PKB) and phosphorylated PKB (p-PKB) were analyzed. RESULTS：The apoptotic rates in valproate group, imatinib group and cotreatment group were (11.47±0.25)%, (28.43±1.70)% and (57.73±4.38)%, respectively (P<0.05). No obvious difference was observed in cell cycle between cotreatment group and monodrug group. Bcr/Abl mRNA and p-PKB in the above 3 groups were (0.00±0.00), (64.17±12.27), and (0.00±0.00) ×10 9 copies/(g total mRNA), respectively (P<005), and 0.25±0.02, 0.17±0.01 and 0.08±0.01, respectively (P<0.05). No apparent difference of PKB was found in the 3 groups. CONCLUSION：Valproate enhances imatinib-induced apoptosis and may link to the down-regulation of Bcr/Abl mRNA and p-PKB in chronic myeloid leukemic cell line K562.     

Protective effects of Zhenwu decoction on kidney injury of diabetic rats induced by streptozotocin

AIM：To investigate the effects of Zhenwu decoction (ZWD) on the expression of α-smooth muscle actin (α-SMA) and NF-κB in diabetic nephropathy (DN) rats. METHODS：Diabetic rat model was induced by intrape-ritoneal injection of streptozotocin (STZ), and the animals were randomly divided into STZ group (n=22) and STZ+ZWD group (n=23). The normal rats served as control (n=16). All rats were sacrificed on 8 weeks after modeling. Biochemical assay and pathological observation (HE staining and transmission electron microscopy) were used to evaluate the effects of Zhenwu decoction on the renal function and pathological morphology. The body weight, renal index, blood glucose, total urinary protein in 24 h, and superoxide dismutase (SOD), malondialdehyde (MDA),inducible nitric oxide synthase (iNOS) were determined as well. Western blotting was used to observe the effects of Zhenwu decoction on the expression of α-SMA and NF-κB in diabetic nephropathy (DN) rats. RESULTS：Compared with normal group, the renal index, blood glucose concentration, total urinary protein in 24 h, blood urea nitrogen (BUN), serum creatinine (SCr) and MDA were significantly higher and body weight was lower in DN rats (P<0. 05). Pathological examination of the kidneys in DN group showed glomerular hypertrophy, glomerular basement membrane thickening, tubular epithelial cell degeneration, mesangial matrix proliferation, protein cast formation in some renal tubules. The protein expression levels of α-SMA and NF-κB were markedly increased (P<0.05). After ZWD treatment, the level of renal index, total urinary protein in 24 h, BUN, SCr and the expression of α-SMA and NF-κB at the protein level were significantly decreased (P<0.05). The renal histological injury in ZWD group was significantly ameliorated. CONCLUSION：Zhenwu decoction might protect kidney against STZ-induced injury via decreasing the expression of α-SMA and NF-κB.     

Regulatory effect of RXR/VDR agonists on atherosclerosis and NF-κB expression in diabetic ApoE knockout mice

AIM：To explore the effect of retinoid X receptor (RXR) agonist bexarotene (Bex) and vitamin D receptor (VDR) agonist calcitriol (Cal) on the expression of nuclear factor-kappa B (NF-κB) and the development of atherosclerosis in streptozotocin-induced diabetic apolipoprotein E knockout (STZ-ApoE-/-) mice. METHODS：Male mice were treated for 12 weeks as follows: (1) C57+vehicle; (2) ApoE-/-+vehicle; (3) STZ-ApoE-/-+vehicle; (4) STZ-ApoE-/-+Bex (10 mg·kg-1·d-1); (5) STZ-ApoE-/-+Cal (10 μg/kg, twice a week); (6) STZ-ApoE-/-+Bex (10 mg·kg-1·d-1)+Cal (10 μg/kg, twice a week). Intraperitoneal injection of STZ was performed to establish the diabetic animal model. Western blotting and immunohistochemical method was used to detect NF-κB level in the thoracic aorta. Plaque area in the thoracic aorta was measured using HE staining. RESULTS：Compared with the C57 mice, the fasting blood glucose in the ApoE-/- mice was not remarkably changed. The levels of total cholesterol (TC) and low-density lipoprotein (LDL) were greatly increased. The fasting blood glucose and lipid levels in STZ-ApoE-/-group were much higher than those in ApoE-/- group. Compared with STZ-ApoE-/- group, the fasting blood glucose and lipid levels in Bex group and Cal group were not significantly changed. Compared with the C57 mice, the protein expression of NF-κB in the ApoE-/- mice and the STZ-ApoE-/- mice was remarkably increased. Compared with STZ-ApoE-/- group, the levels of NF-κB in Bex group, Cal group and combination group were greatly decreased.Compared with STZ-ApoE-/- group, the thoracic artery plaque areas in Bex group and Cal group were inhibited (both P<005). Compared with Bex group, the plaque area of the thoracic artery in combination group was significantly decreased (P<005). CONCLUSION：Bexarotene or calcitriol decreases the development of atherosclerosis in streptozotocin-induced diabetic ApoE-/- mice. Bexarotene combined with calcitriol affords greater protection than monotherapy. The mechanism may be involved in down-regulating the expression of NF-κB.     

Role of calcineurin in cTnI Asp128Tyr mutation-induced myocardial hypertrophy

AIM：To investigate the role of calcineurin (CaN) signaling pathway in myocardial hypertrophy induced by cardiac troponin I (cTnI) Asp128Tyr mutation. METHODS：The adenovirus containing cTnI Asp128Tyr mutation was transfected into the cultured neonatal rat cardiomyocytes. Cardiac hypertrophy was evaluated by determining the mRNA expression of atrial natriuretic factor (ANF) and brain natriuretic peptide (BNP) in the transfected cells. The effects of CaN inhibitors cyclosporine A (CsA) and FK506 on cardiac hypertrophy were also observed. The mRNA expression of ANF, BNP and CaN was measured by real-time quantitative PCR. The activity of CaN was also detected. RESULTS：Compared with blank control group, negative control virus group or virus containing wildtype cTnI gene group, the mRNA expression of ANF and BNP in mutation group (transfected with the adenovirus containing cTnI Asp128Tyr mutation) significantly increased, and decreased after treatment with CsA or FK506. The mRNA expression and activity of CaN increased in the mutation group compared with blank control group, negative control virus group or virus containing wild-type cTnI gene group. At the same time, the mRNA expression and the activity of CaN decreased after treatment with CsA or FK506. CONCLUSION：Calcineurin signaling pathway is involved in the cardiac hypertrophy induced by cTnI Asp128Tyr mutation.