Chicoric acid ameliorates palmitate-induced sphingosine 1-phosphate signaling pathway in the PBMCs of patients with newly diagnosed type 2 diabetes.

Objective: Sphingosine 1-phosphate (S1P) signaling pathway is involved in the pathogenesis of type 2 diabetes (T2D). So, targeting S1P signaling pathway could be considered as potential therapeutic target for T2D. The aim of this study was to investigate the effects of palmitate and chicoric acid (CA) on S1P signaling pathway in peripheral blood mononuclear cells (PBMCs) from newly diagnosed patients with T2D. Materials and methods: 20 newly diagnosed patients with T2D, aged 40-60 years, were enrolled in the study. PBMCs were isolated and then treated as follows: control groups (untreated, treated with BSA 1% for 12 h), CA groups (treated with 50 µM CA for 6 h), palmitate groups (treated with 500 µM palmitate for 12 h), palmitate + CA groups (treated with 500 µM palmitate for 12 h and then treated with 50 µM CA for 6 h). Finally, sphingosine kinase 1 (SPHK1) and sphingosine 1-phosphate receptor 1 (S1PR1) genes expression were evaluated by real-time PCR and S1PR1 protein levels were quantified using ELISA. Results: Palmitate significantly increased SPHK1 and S1PR1 genes expression and S1PR1 protein levels in PBMCs of patients with diabetes. However, CA ameliorates palmitate-increased SPHK1 and S1PR1 genes expression and S1PR1 levels in these cells. Conclusion: These data indicate that CA could be considered as a novel S1P signaling pathway inhibitor through down regulation of SPHK1 and S1PR1.

Therapeutic effects of Chlorella vulgaris on carbon tetrachloride induced liver fibrosis by targeting Hippo signaling pathway and AMPK/FOXO1 axis.

This study was conducted to present the mechanism of the therapeutic effects of Chlorella vulgaris extract (CV) on the carbon tetrachloride (CCl4) induced liver fibrosis model. Primarily, the mechanism of antioxidant effects of CV were investigated via measuring the expression of forkhead box protein O1 (FOXO1) and phosphorylated 5' adenosine monophosphate-activated protein kinase (p-AMPK) as upstream regulators of superoxide dismutase (SOD) and catalase (CAT). Subsequently, we investigated the regulatory effect of CV treatment on the yes-associated protein (YAP) and transcriptional coactivators with a PDZ-binding motif (TAZ) as fibrogenic factors. Male Wistar rats received CCl4 and olive oil solution 1 ml/kg intraperitoneally for 12 weeks, twice weekly. CV 50 and 100 mg/kg were administered on a daily basis by gavage in the last 4 weeks. Ultimately, liver marker enzymes and hepatic hydroxyproline content were measured. The activity of SOD and CAT and the expression of YAP, TAZ, FOXO1, SOD, and CAT were analyzed. Finally, the protein levels of YAP, TAZ, and p-AMPK were detected. CV administration decreased liver marker enzymes and hydroxyproline content significantly. The expression and protein levels of YAP and TAZ reduced by CV treatment. Furthermore, the augmentation of expression and function of CAT and SOD by CV treatment was followed by an increase in the expression of FOXO1 and protein level of p-AMPK. Our data revealed that the stimulation of expression and function of SOD and CAT by CV treatment could be mediated by FOXO1/p-AMPK axis. Moreover, anti-fibrotic effect of CV might be associated with its inhibitory effect on the hepatic expression of YAP and TAZ. Chlorella vulgaris treatment ameliorates liver fibrosis via two cellular mechanisms. A) Likely, Chlorella vulgaris treatment increases gene expression of enzymatic antioxidants superoxide dismutase (SOD) and catalase (CAT) via upregulating its upstream regulatory elements i.e. phosphorylated 5' adenosine monophosphate-activated protein kinase (p-AMPK) and forkhead box protein O1 (FOXO1). These possible regulatory effects maybe lead to reduce reactive oxygen species level (ROS). B) Chlorella vulgaris treatment decreases hepatic protein level and gene expression of key elements of Hippo signaling pathway i.e. Yes-associated protein (YAP) and Transcriptional coactivators with a PDZ-binding motif (TAZ). Figure created with BioRender ( https://biorender.com ). ROS: Reactive oxygen species, YAP: Yes-associated protein, TAZ: Transcriptional coactivators with a PDZ-binding motif, FOXO1: Fork head Box O1, AMPK: 5' adenosine monophosphate activated protein kinase, SOD: Superoxide dismutase, CAT: Catalase, P: Phosphate group.

Palmitate-induced IL6 expression ameliorated by chicoric acid through AMPK and SIRT1-mediated pathway in the PBMCs of newly diagnosed type 2 diabetes patients and healthy subjects.

Inhibition of inflammation is one of the possible therapeutic approaches for Insulin resistance (IR) during type 2 diabetes mellitus (T2DM). In the current study we investigated the effects of palmitate and chicoric acid (CA) on inflammation in peripheral blood mononuclear cells (PBMCs) of newly diagnosed T2DM patients and healthy subjects and explored the mechanism by which palmitate and CA influence inflammation. 20 newly diagnosed T2DM patients and 20 healthy subjects were recruited in our study. Blood sample were collected and PBMCs were isolated. Interleukin 6 (IL6), silent information regulator type 1 (SIRT1), AMP-activated protein kinase (AMPK) and phospho-AMPK (pAMPK) were evaluated both in vivo and in vitro. PBMCs were treated with palmitate and CA to investigate their effects on inflammation. IL6 and SIRT1 genes expression were evaluated by real-time PCR. The levels of IL6 in culture medium were measured by ELISA. Proteins levels of AMPK and pAMPK in PBMCs were detected by western blotting. IL6 expression was higher and SIRT1 expression and pAMPK levels were lower in PBMCs of diabetic patients and obese subjects compared to healthy subjects and non-obese subjects, respectively. CA significantly prevented against increased IL6 levels as well as its gene expression in PBMCs induced by palmitate. Also, CA returned reduction in SIRT1 expression and pAMPK levels mediated via palmitate to near control level. These findings reveal that CA reduces inflammation in PBMCs probably through upregulation of SIRT1 and pAMPK. Therefore, CA would be suggested as a novel agent for the treatment of T2DM.