Honghui Guo

Cyanidin 3-glucoside attenuates obesity-associated insulin resistance and hepatic steatosis in high-fat diet-fed and db/db mice via the transcription factor FoxO1

Obesity is a major risk factor for the development of type 2 diabetes, and both conditions are now recognized to possess significant inflammatory components underlying their pathophysiologies. Here, we hypothesized that cyanidin 3-glucoside (C3G), a typical anthocyanin reported to possess potent anti-inflammatory properties, would ameliorate obesity-associated inflammation and metabolic disorders, such as insulin resistance and hepatic steatosis in mouse models of diabesity. Male C57BL/6J obese mice fed a high-fat diet for 12 weeks and genetically diabetic db/db mice at an age of 6 weeks received dietary C3G supplementation (0.2%) for 5 weeks. We found that dietary C3G lowered fasting glucose levels and markedly improved the insulin sensitivity in both high-fat diet fed and db/db mice as compared with unsupplemented controls. White adipose tissue messenger RNA levels and serum concentrations of inflammatory cytokines (tumor necrosis factor-α, interleukin-6, and monocyte chemoattractant protein-1) were reduced by C3G, as did macrophage infiltration in adipose tissue. Concomitantly, hepatic triglyceride content and steatosis were alleviated by C3G. Moreover, C3G treatment decreased c-Jun N-terminal kinase activation and promoted phosphorylation and nuclear exclusion of forkhead box O1 after refeeding. These findings clearly indicate that C3G has significant potency in antidiabetic effects by modulating the c-Jun N-terminal kinase/forkhead box O1 signaling pathway and the related inflammatory adipocytokines.

Cyanidin 3-glucoside protects 3T3-L1 adipocytes against H2O2-or TNF-α-induced insulin resistance by inhibiting c-Jun NH2-terminal kinase activation

Anthocyanins are naturally occurring plant pigments and exhibit an array of pharmacological properties. Our previous study showed that black rice pigment extract rich in anthocyanin prevents and ameliorates high-fructose-induced insulin resistance in rats. In present study, cyanidin 3-glucoside (Cy-3-G), a typical anthocyanin most abundant in black rice was used to examine its protective effect on insulin sensitivity in 3T3-L1 adipocytes exposed to H(2)O(2) (generated by adding glucose oxidase to the medium) or tumor necrosis factor alpha (TNF-alpha). Twelve-hour exposure of 3T3-L1 adipocytes to H(2)O(2) or TNF-alpha resulted in the increase of c-Jun NH(2)-terminal kinase (JNK) activation and insulin receptor substrate 1 (IRS1) serine 307 phosphorylation, concomitantly with the decrease in insulin-stimulated IRS1 tyrosine phosphorylation and cellular glucose uptake. Blocking JNK expression using RNA interference efficiently prevented the H(2)O(2)- or TNF-alpha-induced defects in insulin action. Pretreatment of cells with Cy-3-G reduced the intracellular production of reactive oxygen species, the activation of JNK, and attenuated H(2)O(2)- or TNF-alpha-induced insulin resistance in a dose-dependent manner. In parallel, N-acetyl-cysteine, an antioxidant compound, did not exhibit an attenuation of TNF-alpha-induced insulin resistance. Taken together, these results indicated that Cy-3-G exerts a protective role against H(2)O(2)- or TNF-alpha-induced insulin resistance in 3T3-L1 adipocytes by inhibiting the JNK signal pathway.

Cyanidin-3-O-β-glucoside inhibits iNOS and COX-2 expression by inducing liver X receptor alpha activation in THP-1 macrophages

Anthocyanins belong to a large and widespread group of water-soluble phytochemicals and exhibit potent antioxidative and anti-inflammatory properties; however, the molecular mechanisms of these biochemical actions mediated by anthocyanins remain unclear. In this study, our data show that pretreatment of THP-1 macrophages with Cyanidin-3-O-beta-glucoside (C3G) for 12 h can enhance the expression and transcriptional activities of the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) and liver X receptor alpha (LXRalpha). Furthermore, pretreatment of these cells with C3G for 12 h causes dose-dependent inhibition of lipopolysaccharide (LPS)-induced nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at both the mRNA and protein levels together with a decrease in nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production. Consequently, addition of geranylgeranyl pyrophosphate ammonium salt (GGPP), an LXRalpha antagonist, significantly downregulates the inhibitory effect of C3G on LPS-induced iNOS and COX-2 expression in THP-1 macrophages, whereas the PPARgamma antagonist GW9662 has no effect. Further investigation revealed that LXRalpha might interfere with LPS-induced iNOS and COX-2 expression by suppressing the functional activation of nuclear factor-kappaB (NF-kappaB), not - as was previously proposed - by reducing NF-kappaB nuclear translocation. Taken together, these results indicate that LXRalpha activation has an essential role in the anti-inflammatory property of C3G. Moreover, they provide new insight into the molecular basis for the anti-inflammatory property of anthocyanins.

Cyanidin-3-O-β-glucoside regulates fatty acid metabolism via an AMP-activated protein kinase-dependent signaling pathway in human HepG2 cells

BackgroundHepatic metabolic derangements are key components in the development of fatty liver disease. AMP-activated protein kinase (AMPK) plays a central role in controlling hepatic lipid metabolism through modulating the downstream acetyl CoA carboxylase (ACC) and carnitine palmitoyl transferase 1 (CPT-1) pathway. In this study, cyanidin-3-O-β-glucoside (Cy-3-g), a typical anthocyanin pigment was used to examine its effects on AMPK activation and fatty acid metabolism in human HepG2 hepatocytes.ResultsAnthocyanin Cy-3-g increased cellular AMPK activity in a calmodulin kinase kinase dependent manner. Furthermore, Cy-3-g substantially induced AMPK downstream target ACC phosphorylation and inactivation, and then decreased malonyl CoA contents, leading to stimulation of CPT-1 expression and significant increase of fatty acid oxidation in HepG2 cells. These effects of Cy-3-g are largely abolished by pharmacological and genetic inhibition of AMPK.ConclusionThis study demonstrates that Cy-3-g regulates hepatic lipid homeostasis via an AMPK-dependent signaling pathway. Targeting AMPK activation by anthocyanin may represent a promising approach for the prevention and treatment of obesity-related nonalcoholic fatty liver disease.

Anthocyanin attenuates CD40-mediated endothelial cell activation and apoptosis by inhibiting CD40-induced MAPK activation

CD40-mediated inflammatory signaling is a potent activator of endothelial cells (ECs) and effective in triggering the pathogenesis of atherosclerosis, a chronic inflammatory disease. Anthocyanin is considered to exert potent cardiovascular-protective effect partially through its anti-inflammatory property, however, the precise mechanism is still unknown. Here we chose cultured human umbilical vein endothelial cells (HUVECs) to explore the influence of anthocyanin on CD40-mediated endothelial activation and apoptosis and the underlying mechanism. Stimulation of human primary HUVECs by CD40 with its physiological ligand CD40L not only augmented MMP-1, -9 secretion and promoted MMP-1, -9 activities, but also induced endothelial cell apoptosis and death. Treatment of ECs with anthocyanins cyanidin-3-O-beta-glucoside (Cy-3-g) and peonidin-3-O-beta-glucoside (Pn-3-g) prevents CD40-induced endothelial activation by inhibiting production of proinflammatory cytokines and matrix metalloproteinases (MMPs). In addition, exposure to anthocyanins inhibits CD40-induced endothelial apoptosis. Anthocyanins also decreased activation of JNK and p38 induced by CD40. Collectively, our findings suggested that the inhibition of JNK and p38 activation interrupts CD40 induced endothelial cell activation and apoptosis, which thereby may represent a mechanism that would explain the anti-inflammatory response of anthocyanin and its athero-protective function.

Optimization of Microwave-Assisted Extraction of Anthocyanins from Mulberry and Identification of Anthocyanins in Extract Using HPLC-ESI-MS

Anthocyanins are naturally occurring compounds that impart color to fruits, vegetables, and plants. This study aims to optimize the microwave-assisted extraction (MAE) conditions of anthocyanins from mulberry (M. atropurpurea Roxb.) using response surface methodology (RSM). A Box-Behnken experiment was employed in this regard. Methanol concentration, microwave power, and extraction time were chosen as independent variables. The optimized conditions of MAE were as follows: 59.6% acidified methanol, 425 W power, 25 (v/w) liquid-to-solid ratio, and 132 s time. Under these conditions, 54.72 mg anthocyanins were obtained from 1.0 g mulberry powder. Furthermore, 8 anthocyanins were identified by high-performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS) in mulberry extract. The results showed that cyanidin-3-glucoside and cyanidin-3-rutinoside are the major anthocyanins in mulberry. In addition, in comparison with conventional extraction, MAE is more rapid and efficient for extracting anthocyanins from mulberry.

Retinol binding protein 4 stimulates hepatic sterol regulatory element-binding protein 1 and increases lipogenesis through the peroxisome proliferator-activated receptor-γ coactivator 1β-dependent pathway

Recent studies have revealed the essential role of retinol binding protein 4 (RBP4) in insulin resistance. However, the impact of RBP4 on aberrant lipogenesis, the common hepatic manifestation in insulin resistance states, and the underlying mechanism remain elusive. The present study was designed to examine the effect of RBP4 on sterol regulatory element‐binding protein (SREBP‐1) and hepatic lipogenesis. Treatment with human retinol‐bound RBP4 (holo‐RBP4) significantly induced intracellular triglyceride (TAG) synthesis in HepG2 cells and this effect is retinol‐independent. Furthermore, RBP4 treatment enhanced the levels of mature SREBP‐1 and its nuclear translocation, thereby increasing the expression of lipogenic genes, including fatty acid synthase (FAS), acetyl coenzyme A carboxylase‐1 (ACC‐1), and diacylglycerol O‐acyltransferase 2 (DGAT‐2). Stimulation of HepG2 cells with RBP4 strongly up‐regulated the expression of transcriptional coactivator peroxisome proliferator‐activated receptor‐γ coactivator 1β (PGC‐1β) at both the messenger RNA (mRNA) and protein levels. The transcriptional activation of PGC‐1β is necessary and sufficient for the transcriptional activation of SREBP‐1 in response to RBP4. The cyclic adenosine monophosphate (cAMP)‐response element binding protein (CREB) was identified as the target transcription factor involved in the RBP4‐mediated up‐regulation of PGC‐1β transcription as a result of phosphorylation on Ser133. Furthermore, in vivo RBP4 infusion induced SREBP‐1c activation and consequently accelerated hepatic lipogenesis and plasma TAG in C57BL/6J mice, a phenomenon not observed in Ppargc1b knockout mice. Conclusion: These findings reveal a novel mechanism by which RBP4 achieves its effects on hepatic lipid metabolism. (HEPATOLOGY 2013;8:564‐575)

Anthocyanin inhibits high glucose-induced hepatic mtGPAT1 activation and prevents fatty acid synthesis through PKCζ

Mitochondrial acyl-CoA:glycerol-sn-3-phosphate acyltransferase 1 (mtGPAT1) controls the first step of triacylglycerol (TAG) synthesis and is critical to the understanding of chronic metabolic disorders such as primary nonalcoholic fatty liver disease (NAFLD). Anthocyanin, a large group of polyphenols, was negatively correlated with hepatic lipid accumulation, but its impact on mtGPAT1 activity and NAFLD has yet to be determined. Hepatoma cell lines and KKAy mice were used to investigate the impact of anthocyanin on high glucose-induced mtGPAT1 activation and hepatic steatosis. Treatment with anthocyanin cyanidin-3-O-β-glucoside (Cy-3-g) reduced high glucose-induced GPAT1 activity through the prevention of mtGPAT1 translocation from the endoplasmic reticulum to the outer mitochondrial membrane (OMM), thereby suppressing intracellular de novo lipid synthesis. Cy-3-g treatment also increased protein kinase C ζ phosphorylation and membrane translocation in order to phosphorylate the mtF0F1-ATPase β-subunit, reducing its enzymatic activity and thus inhibiting mtGPAT1 activation. In vivo studies further showed that Cy-3-g treatment significantly decreases hepatic mtGPAT1 activity and its presence in OMM isolated from livers, thus ameliorating hepatic steatosis in diabetic KKAy mice. Our findings reveal a novel mechanism by which anthocyanin regulates lipogenesis and thereby inhibits hepatic steatosis, suggesting its potential therapeutic application in diabetes and related steatotic liver diseases.

Lack of association between four SNPs in the SLC22A3-LPAL2-LPA gene cluster and coronary artery disease in a Chinese Han population: a case control study

BackgroundLipoprotein (a) (Lp [a]) is known being correlated with coronary artery disease (CAD). The SLC22A3-LPAL2-LPA gene cluster, relating with modulating the level of plasma Lp (a), has recently been reported to be associated with CAD in Caucasians. The purpose of this study was to verify whether this finding can be expanded to the Chinese Han population.Methods and ResultsUsing a Chinese Han sample, which consisted of 1012 well-characterized CAD patients and 889 healthy controls, we tested the associations of four SNPs (rs2048327, rs3127599, rs7767084 and rs10755578) in the SLC22A3-LPAL2-LPA gene cluster, and their inferred haplotypes with the risk of CAD. Allelic, genotypic and haplotype association analyses all showed that the gene cluster was not associated with CAD in this Chinese Han sample.ConclusionsWe for the first time explored the association of the four SNPs in the SLC22A3-LPAL2-LPA gene cluster with CAD in a large Chinese Han sample. Nevertheless, this study did not reveal any significant evidence of this gene cluster to increase the risk of CAD in this population.

Association between rs10118757(A/G) in methylthioadenosine phosphorylase gene and coronary artery disease in Chinese Hans

Studies focusing on the association of gene methylthioadenosine phosphorylase (MTAP) with the risk of coronary artery disease (CAD) and myocardial infarction (MI) are limited. In this study, we explored the effects of rs10118757 in MTAP gene on CAD and MI by performing association analysis in a Chinese Han population. rs10118757 was genotyped in 1007 CAD patients (including 338 MI patients) and 885 healthy controls. Allelic analysis showed that allele A of rs10118757 was associated with increased risk of CAD, with OR (95%CI)=1.193 (1.035-1.376), and P=0.015. After adjusted for age, BMI, gender, hypertension and smoking, rs10118757 was still significantly associated with CAD under additive and dominant models, with OR (95%CI)=1.252 (1.070-1.465), P=0.005, and OR (95%CI)=1.698 (1.168-2.467), P=0.006, respectively. Compared to additive model, dominant model may be the best-fitting model (P=6.63E-10 vs P=6.70E-10). As reported previously, rs10118757 was not associated with MI in the current study. Our study firstly reported that SNP rs10118757 was associated with CAD risk in a Chinese Han population, indicating that MTAP gene may play a potential role in the pathophysiological process of CAD.