Leucine zipper protein 1 attenuates pressure overload-induced cardiac hypertrophy through inhibiting Stat3 signaling.

INTRODUCTION: Cardiac hypertrophy is an important contributor of heart failure, and the mechanisms remain unclear. Leucine zipper protein 1 (LUZP1) is essential for the development and function of cardiovascular system; however, its role in cardiac hypertrophy is elusive. OBJECTIVES: This study aims to investigate the molecular basis of LUZP1 in cardiac hypertrophy and to provide a rational therapeutic approach. METHODS: Cardiac-specific Luzp1 knockout (cKO) and transgenic mice were established, and transverse aortic constriction (TAC) was used to induce pressure overload-induced cardiac hypertrophy. The possible molecular basis of LUZP1 in regulating cardiac hypertrophy was determined by transcriptome analysis. Neonatal rat cardiomyocytes were cultured to elucidate the role and mechanism of LUZP1 in vitro. RESULTS: LUZP1 expression was progressively increased in hypertrophic hearts after TAC surgery. Gain- and loss-of-function methods revealed that cardiac-specific LUZP1 deficiency aggravated, while cardiac-specific LUZP1 overexpression attenuated pressure overload-elicited hypertrophic growth and cardiac dysfunction in vivo and in vitro. Mechanistically, the transcriptome data identified Stat3 pathway as a key downstream target of LUZP1 in regulating pathological cardiac hypertrophy. Cardiac-specific Stat3 deletion abolished the pro-hypertrophic role in LUZP1 cKO mice after TAC surgery. Further findings suggested that LUZP1 elevated the expression of Src homology region 2 domain-containing phosphatase 1 (SHP1) to inactivate Stat3 pathway, and SHP1 silence blocked the anti-hypertrophic effects of LUZP1 in vivo and in vitro. CONCLUSION: We demonstrate that LUZP1 attenuates pressure overload-induced cardiac hypertrophy through inhibiting Stat3 signaling, and targeting LUZP1 may develop novel approaches to treat pathological cardiac hypertrophy.

PBEF promotes the apoptosis of pulmonary microvascular endothelial cells and regulates the expression of inflammatory factors and AQP1 through the MAPK pathways.

Pre-B cell colony-enhancing factor (PBEF) has been shown to have a variety of biological functions. Studies have proven that PBEF plays a functional role in acute lung injury (ALI). Therefore, in this study, we aimed to confirm the importance of PBEF in ALI. The effects of PBEF overexpression on the apoptosis of human pulmonary microvascular endothelial cells (HPMECs) were analyzed by flow cytometry, and the results indicated that PBEF promoted the apoptosis of HPMECs, which aggravated the development of ALI. Comparative experiments involving increasing and decreasing PBEF expression demonstrated that PBEF promoted the expression of inflammatory factors, such as interleukin (IL)­1ß, IL­6 and IL­8 in the HPMECs , thus intensifying the inflammatory response. PBEF also inhibited the expression of aquaporin 1 (AQP1), which caused a dysfunction and imbalance in water transport. Moreover, we also found that tumor necrosis factor (TNF)­α promoted the expression of PBEF in the HPMECs. After blocking the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K) pathways, we found that PBEF regulated the expression of inflammatory factors and AQP1, mainly through the MAPK pathways. Taken together, these results demonstrate that the increase in intracellular PBEF expression promoted the apoptosis of HPMECs and the expression of inflammatory factors and thus enhanced the inflammatory response and inhibited the expression of AQP1, which resulted in abnormal water transport, diminishing the regulatory effects of AQP1 on water transport.

[Effects of hydrogen sulfide on ouabain-induced delayed after-depolarization and triggered activity in guinea pig papillary muscles].

OBJECTIVE: To explore the effects of hydrogen sulfide (H(2)S) on delayed after-depolarization (DAD) and triggered activity induced by ouabain in male guinea pig papillary muscles and to elucidate the underlying mechanisms. METHODS: An intracellular microelectrode was used to record the patterns of DAD and triggered activity by K-H solution containing ouabain and a high concentration of calcium ion. The latent period, amplitude, duration of DAD and incidence of triggered activity were observed under a pre-treatment with different concentrations of NaHS (donor of H(2)S). The effects of glibenclamide, Bay K8644 and NG-nitro-L-arginine methyl ester (L-NAME) pretreatment on the actions of H(2)S were also studied. RESULTS: NaHS (100, 200 µmol/L) prolonged the latent period of DAD from (12.0 ± 1.0) min to (19.9 ± 1.6) min (P < 0.05), (23.7 ± 1.3) min (P < 0.01), decreased the altitude of DAD from (11.47 ± 0.74) mV to (6.47 ± 0.33) mV, (5.65 ± 0.26) mV (both P < 0.01), shortened the duration of DAD from (205 ± 11) ms to (173 ± 10) ms and (134 ± 7) ms (both P < 0.05). The occurrence of triggered activity was inhibited from 5 samples to 4, 2 and 1 sample in 6 samples. A pretreatment of adenosine triphosphate (ATP)-sensitive potassium channel (K(ATP)) blocker glibenclamide partially blocked the preventive effects of H(2)S on ouabain-induced DAD and triggered activity. The effects of H(2)S were completely blocked by L-type calcium channel agonist Bay K8644 (0.25 µmol/L). However a pretreatment of L-NAME (1 mmol/L), a nitric oxide (NO) synthase inhibitor, showed no effects on H(2)S. CONCLUSION: H(2)S inhibits the ouabain-induced DAD and triggered activity in guinea pig papillary muscles. The opening of K(ATP) channel with a reduced influx of calcium ion may be involved in the protective effects of H(2)S.

[Role of JNK signal transduction pathway in nonalcoholic fatty liver disease].

OBJECTIVE: To investigate the role of c-Jun N-terminal kinase (JNK) signal transduction pathway in the rats of nonalcoholic fatty liver disease (NAFLD). METHODS: Sixty four Sprague-Dawley rats were randomly divided into four groups: 8-week control group (NG8w), 12-week control group (NG12 w), 8-week high-fat diet (HG8w), and 12-week high-fat diet group (HG12w), with 16 rats in each group. Glucose infusion rate (GIR) was tested by euglycemic hyperinsulinemic clamp; aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), free fatty acid (FFAs), fast insulin (FIns), tumor necrosis factor alpha (TNF alpha), superoxide dismutase (SOD) and malondialdehyde (MDA) were tested by biochemistry automatic analyzer or RIA; The expression of JNK1, insulin receptor substrate-1 (IRS-1), phospho-IRS-1 Ser307 (p-IRS-1 Ser307), Protein kinase B (PKB) and phospho-PKB Ser473 (p- PKB Ser473) were detected by Western blot. RESULTS: Compared to control group, body weight, liver index, serum levels of ALT, AST, TG, TC, FIns, FFAs, TNF alpha, and TC, TG FFAs, MDA in liver homogenates were increased, while the level of SOD, and GIR were decreased. The expression of JNK1 protein and p-IRS-1 Ser307 in liver tissue was up-regulated, while expression of p-PKB Ser473 was decreased (P < 0.05). A positive correlation was found between the expression intensity of JNK1 and IR (Pearson correlation: 0.718, P < 0.01). CONCLUSION: The high-fat could induce the expression of JNK1, which in turn modulates the phosphorylation of proteins in the insulin signaling pathway, and induces insulin resistant.

High glucose and insulin in combination cause insulin receptor substrate-1 and -2 depletion and protein kinase B desensitisation in primary cultured rat adipocytes: possible implications for insulin resistance in type 2 diabetes.

OBJECTIVE: The purpose of this study was to investigate the cellular effects of long-term exposure to high insulin and glucose levels on glucose transport and insulin signalling proteins. DESIGN AND METHODS: Rat adipocytes were cultured for 24 h in different glucose concentrations with 10(4) microU/ml of insulin or without insulin. After washing, (125)I-insulin binding, basal and acutely insulin-stimulated d-[(14)C]glucose uptake, and insulin signalling proteins and glucose transporter 4 (GLUT4) were assessed. RESULTS: High glucose (15 and 25 mmol/l) for 24 h induced a decrease in basal and insulin-stimulated glucose uptake compared with control cells incubated in low glucose (5 or 10 mmol/l). Twenty-four hours of insulin treatment decreased insulin binding capacity by approximately 40%, and shifted the dose-response curve for insulin's acute effect on glucose uptake 2- to 3-fold to the right. Twenty-four hours of insulin treatment reduced basal and insulin-stimulated glucose uptake only in the presence of high glucose (by approximately 30-50%). At high glucose, insulin receptor substrate-1 (IRS-1) expression was downregulated by approximately 20-50%, whereas IRS-2 was strongly upregulated by glucose levels of 10 mmol/l or more (by 100-400%). Insulin treatment amplified the suppression of IRS-1 when combined with high glucose and also IRS-2 expression was almost abolished. Twenty-four hours of treatment with high glucose or insulin, alone or in combination, shifted the dose-response curve for insulin's effect to acutely phosphorylate protein kinase B (PKB) to the right. Fifteen mmol/l glucose increased GLUT4 in cellular membranes (by approximately 140%) compared with 5 mmol/l but this was prevented by a high insulin concentration. CONCLUSIONS: Long-term exposure to high glucose per se decreases IRS-1 but increases IRS-2 content in rat adipocytes and it impairs glucose transport capacity. Treatment with high insulin downregulates insulin binding capacity and, when combined with high glucose, it produces a marked depletion of IRS-1 and -2 content together with an impaired sensitivity to insulin stimulation of PKB activity. These mechanisms may potentially contribute to insulin resistance in type 2 diabetes.

Dexamethasone impairs insulin signalling and glucose transport by depletion of insulin receptor substrate-1, phosphatidylinositol 3-kinase and protein kinase B in primary cultured rat adipocytes.

OBJECTIVE: Glucocorticoid excess leads to insulin resistance. This study explores the effects of glucocorticoids on the glucose transport system and insulin signalling in rat adipocytes. The interaction between glucocorticoids and high levels of insulin and glucose is also addressed. DESIGN AND METHODS: Isolated rat adipocytes were cultured for 24 h at different glucose concentrations (5 and 15 mmol/l) with or without the glucocorticoid analogue dexamethasone (0.3 micromol/l) and insulin (10(4) microU/ml). After the culture period, the cells were washed and then basal and insulin-stimulated glucose uptake, insulin binding and lipolysis as well as cellular content of insulin signalling proteins (insulin receptor substrate-1 (IRS-1), IRS-2, phosphatidylinositol 3-kinase (PI3-K) and protein kinase B (PKB)) and glucose transporter isoform GLUT4 were measured. RESULTS: Dexamethasone in the medium markedly decreased both basal and insulin-stimulated glucose uptake at both 5 and 15 mmol/l glucose (by approximately 40-50%, P<0.001 and P<0.05 respectively). Combined long-term treatment with insulin and dexamethasone exerted additive effects in decreasing basal, and to a lesser extent insulin-stimulated, glucose uptake capacity (P<0.05) compared with dexamethasone alone, but this was seen only at high glucose (15 mmol/l). Insulin binding was decreased (by approximately 40%, P<0.05) in dexamethasone-treated cells independently of surrounding glucose concentration. Following dexamethasone treatment a approximately 75% decrease (P<0.001) in IRS-1 expression and an increase in IRS-2 (by approximately 150%, P<0.001) was shown. Dexamethasone also induced a subtle decrease in PI3-K (by approximately 20%, P<0.01) and a substantial decrease in PKB content (by approximately 45%, P<0.001). Insulin-stimulated PKB phosphorylation was decreased (by approximately 40%, P<0.01) in dexamethasone-treated cells. Dexamethasone did not alter the amount of total cellular membrane-associated GLUT4 protein. The effects of dexamethasone per se on glucose transport and insulin signalling proteins were mainly unaffected by the surrounding glucose and insulin levels. Dexamethasone increased the basal lipolytic rate (approximately 4-fold, P<0.05), but did not alter the antilipolytic effect of insulin. CONCLUSIONS: These results suggest that glucocorticoids, independently of the surrounding glucose and insulin concentration, impair glucose transport capacity in fat cells. This is not due to alterations in GLUT4 abundance. Instead dexamethasone-induced insulin resistance may be mediated via reduced cellular content of IRS-1 and PKB accompanied by a parallel reduction in insulin-stimulated activation of PKB.

[High glucose impaires glucose transport and PKB activity in adipocytes of rats].

OBJECTIVE: To explore the effect of high glucose on glucose transport activity, protein kinase B (PKB) activity and glucose transporter 4 (GLUT4) in primary cultured rat adipocytes. METHODS: Isolated rat adipocytes were cultured for 24 h at different glucose concentrations (5, 10, 15 and 25 mmol.L-1). The glucose uptake, cellular and membrane GLUT4 expression, PKB protein expression, and PKB serine phosphorylation and activity were measured. RESULTS: These adipocytes treated with glucose of different concentrations showed that high glucose impaired glucose uptake, PKB phosphorylation and activity, and up-regulated GLUT4 translocation, but didn't affect protein expression of PKB and GLUT4. CONCLUSION: High glucose can induce insulin resistance; the mechanism may be involved in the effect of high glucose on PKB serine phosphorylation and activity as well as GLUT4 function.