Palmitate induces VSMC apoptosis via toll like receptor (TLR)4/ROS/p53 pathway.

BACKGROUND AND AIMS: Toll-like receptor 4 (TLR4) has been implicated in vascular inflammation, as well as in the pathogenesis of atherosclerosis and diabetes. Vascular smooth muscle cell (VSMC) apoptosis has been shown to induce plaque vulnerability in atherosclerosis. Previous studies reported that palmitate induced apoptosis in VSMCs; however, the role of TLR4 in palmitate-induced apoptosis in VSMCs has not yet been defined. In this study, we investigated whether or not palmitate-induced apoptosis depended on the activation of the TLR4 pathway. METHODS: VSMCs were treated with or without palmitate, CRISPR/Cas9z-mediated genome editing methods were used to deplete TLR4 expression, while NADPH oxidase inhibitors were used to inhibit reactive oxygen species (ROS) generation. Cell apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, ROS was measured using the 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) method, the mRNA and protein expression levels of caspase 3, caspase 9, BCL-2 and p53 were studied by real-time polymerase chain reaction (RT-PCR) and ELISA. RESULTS: Palmitate significantly promotes VSMC apoptosis, ROS generation, and expression of caspase 3, caspase 9 and p53; while NADPH oxidase inhibitor pretreatment markedly attenuated these effects. Moreover, knockdown of TLR4 significantly blocked palmitate-induced ROS generation and VSMC apoptosis accompanied by inhibition of caspase 3, caspase 9, p53 expression and restoration of BCL-2 expression. CONCLUSIONS: Our results suggest that palmitate-induced apoptosis depends on the activation of the TLR4/ROS/p53 signaling pathway, and that TLR4 may be a potential therapeutic target for the prevention and treatment of atherosclerosis.

The Association Between Endocan Levels and Subclinical Atherosclerosis in Patients With Type 2 Diabetes Mellitus.

BACKGROUND: Proinflammatory conditions induced by circulating factors in diabetes play a pivotal role in endothelial dysfunction and related vascular complications. Endothelial cell-specific molecule-1 or endocan is a dermatan sulfate proteoglycan secreted primarily by the vascular endothelium. Although endocan has been shown to be a potential biomarker in coronary heart disease, its role in the pathogenesis of atherosclerosis (AS) in diabetes remains unclear. In this study, we investigated the correlation between serum endocan levels and subclinical AS in patients with type 2 diabetes mellitus (T2DM). MATERIALS AND METHODS: Patients (n = 69) with T2DM were included. All the patients were stratified based on the absence (n = 42) or presence (n = 27) of subclinical AS. Healthy subjects (n = 28) served as controls. Serum levels of endocan, fasting blood glucose, glycosylated hemoglobin A1, high-sensitivity C-reactive protein and carotid intima-media thickness (cIMT) were measured. RESULTS: Endocan levels were significantly elevated in both the T2DM (0.89 ± 0.28ng/mL) and T2DM with subclinical AS (1.20 ± 0.33ng/mL) groups relative to the control group (0.68 ± 0.24ng/mL) (P < 0.05 for all). Endocan levels were also positively correlated with glycosylated hemoglobin A1, fasting blood glucose and cIMT (r = 0.292, P = 0.004; r = 0.224, P = 0.027 and r = 0.496, P < 0.001, respectively). In addition, endocan levels were independently associated with cIMT (ß = 0.220, t = 5.816, P = 0.000) and were a significant risk factor for T2DM with subclinical AS (odds ratio = 1.98, 95% CI: 1.43-2.73, P < 0.001). CONCLUSIONS: These findings suggest that serum endocan levels may be a useful biomarker for the early diagnosis of subclinical AS in patients with T2DM.

Reduced serum milk fat globule-epidermal growth factor 8 (MFG-E8) concentrations are associated with an increased risk of microvascular complications in patients with type 2 diabetes.

BACKGROUND: The association between serum milk fat globule-epidermal growth factor 8 (MFG-E8) concentrations and vascular complications in T2DM remains unclear. METHODS: A total of 149 patients with T2DM were included. The serum concentrations of MFG-E8, glycosylated hemoglobin (HbA1c), and high-sensitivity C-reactive protein (hs-CRP) were measured. RESULTS: There was no significant difference in serum MFG-E8 concentrations between the T2DM group and the T2DM with subclinical atherosclerosis (AS) group (615.49±143.54 vs. 596.22±79.46ng/ml, P=0.365), while the serum concentrations of MFG-E8 in the T2DM with microvascular complications group (446.70±61.53ng/ml) and the T2DM with subclinical AS and microvascular complications group (200.87±38.86ng/ml) were significantly lower than those in the T2DM group (P=0.000 for both). In addition, hs-CRP and HbAlc concentrations were independently associated with serum MFG-E8 concentrations (P=0.024 and P=0.01, respectively), and low serum MFG-E8 concentrations were significantly associated with an increased risk of microvascular complications in T2DM patients. CONCLUSIONS: Serum concentrations of MFG-E8 were negatively associated with the risk of microvascular complications in patients with T2DM. Thus, it might be a potential candidate biomarker for diabetic microvascular complications.