Regulation of hepatic insulin signaling and glucose homeostasis by sphingosine kinase 2.

Sphingolipid dysregulation is often associated with insulin resistance, while the enzymes controlling sphingolipid metabolism are emerging as therapeutic targets for improving insulin sensitivity. We report herein that sphingosine kinase 2 (SphK2), a key enzyme in sphingolipid catabolism, plays a critical role in the regulation of hepatic insulin signaling and glucose homeostasis both in vitro and in vivo. Hepatocyte-specific Sphk2 knockout mice exhibit pronounced insulin resistance and glucose intolerance. Likewise, SphK2-deficient hepatocytes are resistant to insulin-induced activation of the phosphoinositide 3-kinase (PI3K)-Akt-FoxO1 pathway and elevated hepatic glucose production. Mechanistically, SphK2 deficiency leads to the accumulation of sphingosine that, in turn, suppresses hepatic insulin signaling by inhibiting PI3K activation in hepatocytes. Either reexpressing functional SphK2 or pharmacologically inhibiting sphingosine production restores insulin sensitivity in SphK2-deficient hepatocytes. In conclusion, the current study provides both experimental findings and mechanistic data showing that SphK2 and sphingosine in the liver are critical regulators of insulin sensitivity and glucose homeostasis.

Sphingosine kinase 2 promotes lipotoxicity in pancreatic ß-cells and the progression of diabetes.

Loss of functional ß-cell mass caused by lipotoxicity is a key pathogenic factor in the development of type 2 diabetes mellitus (T2DM). We have previously reported that sphingosine kinase (SK)1 is an endogenous protector of ß-cells against lipotoxicity. The current study reports that SK2, another isoform of SK, is a crucial mediator of lipotoxicity in ß-cells. Exposure of ß-cells to palmitatic acid (PA), a saturated free fatty acid, resulted in a nearly 2-fold increase in SK2 expression, which paralleled the induction of cell death in a similar dose- and time-dependent fashion. Silencing SK2 expression by its specific small interfering RNAs significantly inhibited PA-induced cell death and caspase-3 activation, whereas overexpression of SK2 promoted lipotoxicity in ß-cells. Mechanistically, upon exposure to PA, endogenous SK2 was shuttled from the nucleus to the cytoplasm, where it interacted with B-cell lymphoma-extra-large (Bcl-xL), leading to mitochondrial apoptotic pathway activation and cell death. By blocking SK2 translocation and its interaction with Bcl-xL, either the nuclear export signal mutant (L423A/L425A) or the BH3 domain mutant (L219A) of SK2 significantly attenuated ß-cell lipotoxicity. Furthermore, SK2 deficiency in mice significantly prevented the loss of ß-cell mass, preserved insulin production, and ameliorated the diabetic phenotype in an established T2DM model induced by feeding a high-fat diet accompanied by administration of streptozotocin. These findings provide the first evidence, in vitro and in vivo, of a critical role for SK2 in mediating ß-cell lipotoxicity and the progression of diabetes.-Song, Z., Wang, W., Li, N., Yan, S., Rong, K., Lan, T., Xia, P. Sphingosine kinase 2 promotes lipotoxicity in pancreatic ß-cells and the progression of diabetes.

Slit2 signaling contributes to cholestatic fibrosis in mice by activation of hepatic stellate cells.

Liver Cholestasis is a widespread disease of broad etiologies and ultimately results in fibrosis, which is still lacking effective therapeutic strategies. Activation of hepatic stellate cells (HSCs) is the key event of liver fibrosis. Here, we aimed to investigate the effect and mechanism of the Slit2 signaling in cholestasis-induced liver fibrosis. Our findings revealed that the serum levels and hepatic expression of Slit2 were significantly increased in patients with primary biliary cirrhosis (PBC). Additionally, Slit2-Tg mice were much more vulnerable to BDL-induced liver injury and fibrosis compared to WT control. Slit2 up-regulation by Slit2 recombinant protein induced proliferation, and inhibited apoptosis of human HSCs cell line LX-2 via p38 and ERK signaling pathway, resulting in the activation of HSCs. In contrast, Slit2 down-regulation by siRNA silencing inhibit the activation of HSCs. In conclusion, Slit2 is involved in the activation of HSCs and liver fibrogenesis, highlighting Slit2 as a potential therapeutic target for liver fibrosis.

Effects of andrographolide on renal tubulointersticial injury and fibrosis. Evidence of its mechanism of action.

BACKGROUND: Diabetic nephropathy (DN) is associated with renal interstitial injury and fibrosis. Our previous study showed that andrographolide protected against the progression of DN and high glucose (HG)-induced mesangial dysfunction. However, the protective effects of andrographolide on renal tubular epithelial cells have not been fully elucidated. PURPOSE: To determine the protective effects of andrographolide on renal tubular damage and explore the underlying mechanism. STUDY DESIGN: Human tubular epithelial cells (HK-2 cells) were treated with andrographolide (5 and 10 µM) under HG conditions. Diabetic mice were treated with andrographolide (i.p. 2 and 4 mg/kg, twice per week). METHODS: Western blotting, reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence and flow cytometry were used to analyze the effects of andrographolide on renal tubular injury and fibrotic mechanisms in HK-2 cells. The protective effects of andrographolide against renal tubulointerstitial injury and fibrosis were investigated in diabetic mice fed a high-fat diet (HFD). Renal interstitial tissue was collected at sacrifice for immunohistochemistry, immunofluorescence analysis, RT-PCR and Western blotting to analyze the effects of andrographolide on renal tubular injury and fibrosis. RESULTS: In vitro assay results indicated that andrographolide (5 and 10 µM) effectively inhibited HG-induced apoptosis, epithelial-mesenchymal transition (EMT) and collagen deposition in HK-2 cells. Mechanistically, HG stimulated mitochondrial reactive oxygen species (mtROS)-mediated NOD-like receptor family and pyrin domain-containing protein 3 (NLRP3) inflammasome activation and EMT in tubular epithelial cells, and andrographolide (5 and 10 µM) inhibited these effects by ameliorating mitochondrial dysfunction. In vivo, treatment with andrographolide (2 and 4 mg/kg) inhibited renal tubular cell apoptosis, EMT and tubulointerstitial fibrosis, mitochondrial dysfunction and NLRP3 inflammasome activation in diabetic mice. CONCLUSION: Andrographolide (5 and 10 µM) prevents HG-induced renal tubular cell damage, and andrographolide (2 and 4 mg/kg) protects against the progression of diabetic tubular injury and fibrosis in mice by suppressing mitochondrial dysfunction and NLRP3 inflammasome activation.

[Lipid-lowering effect of propolis in mice with Triton-WR1339-induced hyperlipidemia and its mechanism for regulating lipid metabolism].

OBJECTIVE: To evaluate the therapeutic effect of propolis against Triton-WR1339-induced hyperlipidemia in mice and explore the underlying mechanism. METHODS: C57BL/6 mice were randomly divided into 7 groups (n=10), including the control group, hyperlipidemia model group, fenofibrate (30 mg/kg) treatment group, and 4 treatment groups treated with low- (30 mg/kg) or high-dose (60 mg/kg) propolis HB01 or HB02. In all but the control group, acute hyperlipidemia models were established by intramuscular injection of Triton WR-1339, and corresponding treatments were administered via gastric lavage for 7 days. After the treatments, blood samples were collected for testing the levels of total cholesterol (TC), triglycerides (TG), highdensity lipoprotein-cholesterol (HDL-C), low-density lipoprotein-cholesterol (LDL-C), malondialdehyde (MDA), superoxide dismutase (SOD), alanine aminotransferase (GPT), and aspartate aminotransferase (GOT); Western blotting was used to detect the expressions of the proteins involved in lipid metabolism in the liver tissues including ABCA1, ABCG8, LDLR, and SR-B1. RESULTS: Compared with the normal control group, the mice with Triton-WR1339-induced hyperlipidemia showed significantly increased levels of TC, TG, LDL, MDA, GPT, and GOT and lowered HDL-C levels and SOD activity (P < 0.05). Treatments with fenofibrate and the 2 propolis at either low or high dose significantly reversed Triton-WR1339-induced changes in blood lipids (P < 0.05), and the effects of propolis were more potent. Triton-WR1339 injection also significantly decreased the expressions levels of ABCA1, ABCG8, LDLR, and SR-B1 in the liver (P < 0.05), and these changes were obviously reversed by treatments with fenofibrate and propolis (P < 0.05), especially by the latter. CONCLUSIONS: The lipid-lowering effects of propolis are mediated by improving lipid metabolism and regulating the expressions of lipid transport proteins in the liver tissue.

Andrographolide Ameliorates Atherosclerosis by Suppressing Pro-Inflammation and ROS Generation-Mediated Foam Cell Formation.

Inflammation, oxidative stress, and dyslipidemia are major factors in the pathogenesis of atherosclerosis. Andrographolide, a bioactive component of Andrographis paniculata, has several biological activities, including anti-inflammatory, antioxidant, and anticancer effects. This study shows that andrographolide downregulates the oxidized low-density lipoprotein (oxLDL)-induced expression of the pro-inflammatory molecules monocyte chemotactic protein (MCP)-1 and interleukin (IL)-6 and blocks the nuclear factor-κB signaling pathway in macrophages. Additionally, andrographolide treatment decreased reactive oxygen species (ROS) generation in oxLDL-induced macrophages, indicating that the compound can decrease oxidative stress. The results also suggest that andrographolide suppresses oxLDL-induced foam cell formation and inhibits oxLDL-induced CD36 expression in vitro. Furthermore, in vivo studies have indicated that andrographolide treatment ameliorates atherosclerosis pathogenesis in apolipoprotein E knockout mice. Therefore, by suppressing inflammation, ROS generation, and foam cell formation, andrographolide may ameliorate the progression of atherosclerosis, suggesting its potential as a therapeutic drug for the prevention and/or treatment of this disease.

Polydatin attenuates diet-induced nonalcoholic steatohepatitis and fibrosis in mice.

Scope: Non-alcoholic steatohepatitis (NASH) is characterized by lipid accumulation in hepatocytes and inflammatory cell infiltration. In view of the anti-oxidative and anti-inflammatory effects of polydatin, the current study aimed to investigate the pharmacological effects of polydatin on NASH and its related fibrosis. Methods: C57BL/6 mice were fed with methionine-choline deficient (MCD) diet to induce NASH and liver fibrosis, and treated with or without polydatin (5 mg/kg, every other day, i.p) for 4 weeks. HepG2 cells induced by palmitic acid (PA) were treated with polydatin. Results: The elevations of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST), active caspase-3, TUNEL-positive cells, and triglyceride content were decreased by polydatin treatment. In addition, administration of polydatin to MCD-fed mice reduced oxidative stress by down-regulating NOX4 enzymes. Furthermore, the reduction in inflammation and CD68 macrophage activation correlated with inhibition of toll-like receptor (TLR)-4/NF-κB p65 signaling pathway by polydatin treatment. Polydatin also attenuated lipid accumulation, inflammation and apoptosis in HepG2 cells challenged by palmitic acid (PA) combined with or without lipopolysaccharide (LPS). Finally, the reduction of hepatic fibrosis by polydatin treatment corresponded to a reduction in hepatic gene expression of fibrosis markers. Conclusions: These results suggest that polydatin prevents NASH and fibrosis via inhibition of oxidative stress and inflammation, highlighting polydatin as a potential therapeutic agent for prevention and treatment of NASH.

Protective Effects of Chinese Traditional Medicine Longhu Rendan against Atherosclerosis via Negative Regulation of LOX-1.

Longhu Rendan (LHRD), a Chinese traditional compound medicine, has a remarkable treatment effect on motion sickness for about half a century. However, the role of LHRD in atherosclerosis treatment is still unclear. In this study, LHRD treatment significantly diminished total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) levels in apolipoprotein E gene-knockout (ApoE-/-) mice fed with high fat and high cholesterol diet (western diet). Besides, LHRD treatment significantly reduced atherosclerotic lesion and plaques formation in both aortic roots and aortic trees. Furthermore, immunofluorescence staining in aortic roots demonstrated that LHRD treatment inhibited lectin-like oxidized low-density-lipoprotein receptor-1 (LOX-1) expression in atherosclerotic plaques. These results indicated that LHRD ameliorated atherosclerosis via reducing serum levels of TC, TG, and LDL-C as well as LOX-1 expression, subsequently attenuating atherosclerotic lesion and lipid deposition. In conclusion, LHRD could significantly attenuate experimental atherosclerosis and might be a novel potential drug for the prevention and treatment of atherosclerosis.