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Objective We investigated the anti-obesity effect and underlying action mechanism of INDUS810 isolated from Trigonella foenum-graecum L. (Fabaceae), an annual herb commonly known as fenugreek and reported to have hypocholesterolemic, antidiabetic, anticancer and gastroprotective properties. Methods For obese animal study, 4-week old mice were fed with normal diet or high-fat diet together with or without intraperitoneal injection of INDUS810 (200 mg/kg) twice per week for 15 weeks. 3T3-L1 cells were used to study action mechanism of INDUS810 in adipocyte differentiation and lipid metabolism. Results We found that INDUS810 can reduce high-fat diet-induced weight increase in epididymal white adipose tissue, interscapular brown adipose tissue and liver, as well as serum levels of total cholesterol and low-density lipoprotein cholesterol. Moreover, the insulin sensitivity was significantly improved in INDUS810-treated obese mice. In 3T3-L1 adipocytes, we found that INDUS810 could inhibit lipid accumulation at either differentiating or mature stages, and increase lipolysis activity in mature adipocytes. Additionally, INDUS810 has no effects on cell viability nor the expressions of adipocyte differentiation markers like fatty acid synthase, peroxisome proliferator-activated receptor γ and CCAAT/enhancer-binding protein α. In contrast, INDUS810 can increase protein levels of peroxisome proliferator-activated receptor α, peroxisome-proliferator-activated receptor-γ co-activator 1β, sirtuin 1 and sirtuin 3. Of note, INDUS810 can activate adenosine monophosphate-activated protein kinase, which leads to the reduction of lipid contents in adipocytes. Conclusion Our in vitro and in vivo studies suggest that INDUS810 is a potential anti-obesity agent, and this action depends on activate adenosine monophosphate-activated protein kinase activation.

Andrographolide (Ap) is a bioactive compound in Andrographis paniculata that is a Chinese herb. The pi class of glutathione S-transferase (GSTP) is one kind of phase II detoxification enzyme. Here we show that induction of GSTP protein and mRNA expression in rat primary hepatocytes by Ap was inhibited by forskolin and a variety of cAMP analogues. The inhibitory effect of the cAMP analogues was partially blocked by pretreatment with H89. In the presence of Ap, forskolin, or both, the expression of phospho-cAMP response element-binding protein (CREB) was increased. Ap alone had no effect on inducible cAMP early repressor (ICER) mRNA expression; however, Ap played a potentiating role in forskolin-induced ICER mRNA expression. An EMSA and immunoprecipitation assay showed that ICER binding to cAMP-response element (CRE) was increased in cells cotreated with Ap and forskolin for 3 and 8 h. Taken together, these results suggest that ICER is likely to be involved in the suppression of Ap-induced GSTP expression caused by the increase of cAMP in rat primary hepatocytes.

Andrographis paniculata (Ap) is a commonly used herb for traditional medicine in many Southeast Asian countries. In the present study, we investigated the effect of Ap on the expression of the pi class of glutathione S-transferase (GSTP) in rat primary hepatocytes. Hepatocytes were treated with 25 or 50 microg/mL of ethanol or ethyl acetate extracts of Ap (ApEE or ApEAE) or 10 or 20 microM andrographolide, which is the major active diterpene lactone of Ap, for 48 h. ApEE, ApEAE, and andrographolide dose-dependently induced GSTP protein and mRNA expression. In a GST activity assay, GST activity was significantly higher in cells treated with the maximum concentrations of ApEE, ApEAE, and andrographolide than in control cells (P0.05). The pTA-2713 luciferase reporter construct containing rat GSTP enhancer 1 (GPE1) was transiently transfected into Clone 9 liver cells. Cells treated with ApEE, ApEAE, and andrographolide showed a dose-dependent increase in luciferase activity. GPE1 deletion abolished the induction efficiency of Ap. Also, the induction of GSTP expression by Ap was inhibited by wortmannin, which is an inhibitor of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. These results indicate that ApEE, ApEAE, and andrographolide induce GSTP expression. This induction is likely related to the PI3K/Akt pathway, and GPE1, an enhancer element in GSTP promoter, is essential for the induction.

Statin-induced caspase-1 activation and IL-1β release are due to unmask the anti-inflammatory action of isoprenoids by activating P2X7. Recent studies have demonstrated the stimulatory effects of HMG-CoA reductase inhibitors, statins, on IL-1β secretion in monocytes and suggest a crucial role for isoprenoids in the inhibition of caspase-1 activity. In this study, we further elucidated the molecular mechanisms underlying the stimulatory effects of statins on caspase-1. Three commonly recognized mechanistic models for NLRP3 inflammasome activation (i.e., ATP/P2X7/K+ efflux, ROS production, and lysosomal rupture) were investigated in statin-stimulated human THP-1 monocytes. We found that fluvastatin and lovastatin can synergize with LPS to trigger inflammasome activation. Moreover, statin-induced caspase-1 activation and IL-1β production in LPS-primed THP-1 cells are dependent on GGPP deficiency and P2X7 activation. In particular, increased ATP release accounts for the action of statins in P2X7 activation. We also provide evidence that statin-induced moderate ROS elevation is involved in this event. Moreover, the cathepsin B inhibitor was shown to reduce statin-induced IL-1β secretion. Consistently statins can induce cathepsin B activation and lysosomal rupture, as evidenced by LysoTracker staining. Statins also increase intracellular ATP secretion and IL-1β release in primary human monocytes and murine macrophages. Notably, exogenous ATP-elicited P2X7 activation and consequent IL-1β release, an index of direct NLRP3 inflammasome activation, were not altered by statins. Taken together, statin-induced enhancement of inflammasome activation in monocytes and macrophages covers multiple mechanisms, including increases in ATP release, ROS production, and lysosomal rupture. These data not only shed new insight into isoprenylation-dependent regulation of caspase-1 but also unmask mechanisms for statin-elicited inflammasome activation.

Ethnopharmacological relevance The Ger-Gen-Chyn-Lian-Tang (GGCLT), an officially standardized mixture of Chinese herbal medicines, consists of Puerariae Radix, Scutellariae Radix, Coptidis Rhizoma and Glycyrrhizae Radix in a ratio of 8:3:3:2. In this study, we evaluated the benefits of GGCLT in atherosclerotic progression. Methods The major constituents of GGCLT were analyzed by HPLC. ApoE−/− mice taken 0.15% cholesterol diet were orally given vehicle or GGCLT (2 g/kg/day) for 12 weeks. Serum levels of lipid and glucose were analyzed, and atherosclerosis was examined by histological analyses. Cultures of vascular smooth muscle cells, hepatocytes and bone marrow-derived macrophages were used to investigate the action mechanisms of GGCLT. Results Our quantitation results indicated that GGCLT contains puerarin, daidzin, daidzein, baicalin, baicalein, wogonin, palmatine, coptisine, berberine and glycyrrhizin. GGCLT decreased serum levels of total cholesterol and LDL, but not TG and HDL in ApoE−/− mice. In parallel, GGCLT treatment reduced atherosclerotic lesions and collagen expression in atheroma plaques. In vascular smooth muscle cells, GGCLT could reduce cell migration, but failed to affect cell viability and proliferation. In hepatocytes, GGCLT can reduce lipid accumulation, and this action was accompanied by the activation of AMPK, upregulation of PPARs, and downregulation of FAS. Pharmacological approach indicated that the latter two events contributing to the anti-lipogenesis is resulting from AMPK pathway, and the lipid lowering effect of GGCLT in hepatocytes is mediated by AMPK and PPARα pathways. Meanwhile, two of the major components of GGCLT, berberine and puerarin, also activated AMPK and decreased lipid accumulation in hepatocytes with berberine of higher efficacy. Besides in hepatocytes, AMPK signaling was also activated by GGCLT in vascular smooth muscle cells and macrophages. Conclusions These results demonstrate the anti-atherosclerotic action of Chinese medicine mixture GGCLT in ApoE−/− atherosclerotic mouse model. Mechanistic study suggests that activation of AMPK and PPARα in hepatocytes leading to a decrease of lipid formation contributes to the beneficial action of GGCLT in atherosclerosis treatment.