MiR-146a protects small intestine against ischemia/reperfusion injury by down-regulating TLR4/TRAF6/NF-κB pathway.

Previous studies reported that miR-146a was involved in small intestine ischemia-reperfusion (I/R) injury, but the mechanism is largely vague. Here, we aimed to identify the change of miR-146a in patients with mesenteric ischemia and explore the potential regulatory mechanism of miR-146a in intestine epithelial cells survival under ischemia and I/R injury. The plasma of 20 patients with mesenteric ischemia and 25 controls was collected to examine the miR-146a expression by qPCR. Rat intestinal epithelial cells (IEC-6) and 24 male Sprague-Dawley rats were included to build ischemia and I/R model in vitro and in vivo. The qPCR results showed that miR-146a decreased both in the plasma of patients with mesenteric ischemia and in IEC-6 cells and rat small intestine tissues in ischemia and I/R model compared to controls. Both the in vitro and in vivo results showed that I/R resulted in more severe apoptotic injury than ischemia. Cleaved-caspase 3, TLR4, TRAF6, and nuclear NF-κB p65 were up-regulated accompanying reduced XIAP and SOCS3 expression in intestinal ischemia and I/R injury. After up-regulation of miR-146a in IEC-6 cells, increased cell survival and decreased cell apoptosis were observed, concomitant with decreased cleaved-caspase 3 and down-regulated TLR4/TRAF6/NF-κB pathway. What is more, this protective effect was blocked by TRAF6 overexpression and increased nuclear NF-κB p65 nuclear. Taken together, this study revealed that miR-146a expression was decreased in small intestine ischemia and I/R injury. And miR-146a improves intestine epithelial cells survival under ischemia and I/R injury through inhibition TLR4, TRAF6, and p-IκBα, subsequently leading to decreased NF-κB p65 nuclear translocation.

Altered Plasma MicroRNAs as Novel Biomarkers for Arteriosclerosis Obliterans.

AIM: Arteriosclerosis obliterans (ASO) of the lower extremities is a major cause of adult limb loss worldwide. A timely diagnosis in the early stages of the disease determines the clinical outcomes, however lacking of palpable symptoms remains the biggest obstacle. This study aimed to screen a cluster of microRNAs (miRNAs) that can be used as biomarker for the ASO in the earlier stages. METHODS: Plasma from 3 patients with ASO and 3 healthy controls were profiled to screen altered miRNAs by microarray, then Real time PCR was further used to confirm the changes in 55 ASO patients and 54 controls.We also analyzed the correlation of miRNAs level with Fontaine stages and the influence of T2DM which is a common complication with ASO on the level of miRNAs. RESULT: Twenty-four aberrantly expressed miRNAs were screened in the plasma of ASO patients. Real time PCR verified that the level of miR-4284 was significantly increased, while levels of miR-4463, miR-4306 and miR-221-3p were significantly decreased both in the plasma and in the sclerotic samples compared with the controls. Interestingly, we revealed a time and stage specific expression manner, as shown that expression of miR-4284 increased at the stage I of ASO and maintained the tendency to stage IV, while miR-4463 expression decreased at every stage of ASO; however, the expression of miR-4463 showed opposite changes in ASO patients with or without T2DM. CONCLUSION: Altered expressions of miR-4284 and miR-4463 are novel characteristics and may serve as potential biomarkers for the early diagnosis of ASO.

Experimental study on apoptosis of TNFR1 receptor pro-endothelial progenitor cells activated by high glucose induced oxidative stress.

OBJECTIVE: To investigate whether high glucose in vitro activating TNFR1 and further promote rat marrow endothelial progenitor cells (EPCs) apoptosis. METHODS: Rat morrow endothelial progenitor cells were cultured and identified by Confocal Microscopy; then were treated with high glucose (5.5, 15, 30, 60 mmol/L), mannitol (15, 30, 60, 90 mmol/L), high glucose + Tempol and high glucose+ MAB430. Apoptosis rate of the above cells were detected by flow cytometry. ROS and MDA level and anti-O2- were detected by colorimetric technique; the expression level of TNFR1 induced signal pathway related proteins were detected by Western blotting. RESULTS: High glucose can induce endothelial progenitor cells apoptosis, which is mostly in the later stage (72 h-96 h) instead of the earlier stage (24 h-48 h); high glucose can also induce oxidative stress reaction and the produces ROS and MDA increase significantly in the later stage (after 72 h), but anti-O2- decrease significantly. TNF apoptosis signal pathway related protein expression level not increase in the earlier stage (before 24 h) but increase significantly in the later stage (after 72 h). Tempol and MAB430 down-regulate TNF apoptosis signal pathway related protein expression and reduce EPCs apoptosis. CONCLUSION: High glucose activates the TNFR1 of TPCs through oxidative stress reaction and further induces cell apoptosis.