Expression and effect of microRNA-146a gene mediated by ultrasound microbubble in rats with ischemia/reperfusion liver injury
10.3969/j.issn.1008-9691.2019.01.005
- VernacularTitle:超声微泡介导微小RNA-146a基因在缺血/再灌注肝损伤大鼠中的表达与作用
- Author:
Chungen YAN
1
;
Dongfang ZHU
;
Guoyuan XIA
Author Information
1. 绍兴文理学院附属医院消化内科
- Keywords:
MicroRNA-146a;
Ischemia/reperfusion,liver;
Ultrasound microbubble;
Gene transfection
- From:
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care
2019;26(1):21-25
- CountryChina
- Language:Chinese
-
Abstract:
Objective To evaluate the relationship between the expression of microRNA-146a (miR-146a) in liver tissue and the inflammatory hepatic injury induced by ischemia/reperfusion (I/R) in rats. Methods One hundred and forty-four Sprague-Dawley (SD) rats were randomly divided into three groups: control (group N), sham operation (group S) and group I/R. Each group was subdivided into four subgroups (n = 12), and different substances were respectively injected intravenously to rats in different subgroups at 1 hour before the experiment: 220 μL physiological saline (group A), 20 μL miR-146a mimic + 200 μL physiological saline (group B), 20 μL miR-146a mimic + 200 μL ultrasound microbubble contrast agent (group C) and 20 μL miR-146a inhibitor + 200 μL ultrasound microbubble contrast agent (group D). Before the experiment and after experiment for 24 hours, the plasma concentrations of alanine aminotransferase (ALT), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were detected, the reverse transcription-polymerase chain reaction (RT-PCR) was used to measure the expression of miR-146a in liver tissue, and Western Blot was applied to detect protein expressions of Toll-like receptor 4 (TLR4), IL-1 receptor associated kinase 1 (IRAK-1), IL-6 and TNF-α, and the pathological hepatic cell injury was observed. Results Before the experiment and 24 hours after experiment in various subgroups of N and S groups, there were no statistical significant differences in the plasma concentrations of ALT, IL-6 and TNF-α, and the expression of miR-146a level and the protein expressions of TLR4, IRAK-1, IL-6 and TNF-α in liver tissues; the pathological examination also did not show any obvious hepatic cell injury. After the experiment for 24 hours: compared to the group S, the liver tissue miR-146a expression was significantly decreased in the subgroups A and D of group I/R (miR-146a/U6nsRNA: 0.51±0.13, 0.22±0.09 vs. 1.01±0.02, both P < 0.01), and the plasma concentrations of ALT, IL-6 and TNF-α and the protein expressions of TLR4, IRAK-1, IL-6 and TNF-α in liver tissues were significantly increased [ALT (U/L): 103.23±26.64 vs. 44.16±18.55, 176.46±7.26 vs. 49.74±6.83, IL-6 (μg/L): 64.28±16.19 vs. 17.68±7.54, 88.49±3.23 vs. 15.58±2.38; TNF-α (μg/L): 31.28±2.57 vs. 5.58±3.35, 59.12±8.74 vs. 5.27±1.37; TLR4/GAPDH: 2.43±0.36, 3.23±0.71 vs. 0.96±0.24, IRAK-1/GAPDH: 2.34±0.52, 3.14±0.63 vs. 0.76±0.21, IL-6/GAPDH: 1.01±0.22, 1.11±0.16 vs. 0.98±0.37, TNF-α/GAPDH: 2.05±0.48, 2.86±0.27 vs. 0.59±0.16, all P < 0.01], moreover, the hepatic pathological lesions were obvious; the liver tissue expression of miR-146a was significantly increased after being transfected with miR-146a mimic in subgroups B and C of group I/R (miR-146a/U6nsRNA: 1.56±0.31, 2.40±0.53 vs. 1.01±0.02, both P < 0.01), especially in group C combined with ultrasound microbubble (P < 0.01). However, the protein expressions of TLR4, IRAK-1, IL-6 and TNF-α in liver tissues were significantly decreased (TLR4/GAPDH:0.77±0.18, 0.65±0.27 vs. 0.96±0.24, IRAK-1/GAPDH: 0.61±0.14, 0.47±0.20 vs. 0.76±0.21, IL-6/GAPDH:0.80±0.13, 0.54±0.22 vs. 0.98±0.37, TNF-α/GAPDH: 0.41±0.14, 0.16±0.03 vs. 0.59±0.16; all P < 0.01), and the expressions were more significant in the group C combined with ultrasound microbubbles (P < 0.01), and the hepatic pathological damage was mild, however, the plasma concentrations of ALT, IL-6 and TNF-α were of no statistical significant differences. Conclusion Ultrasound microbubble can efficiently transfect miR-146a mimic and inhibitor into the liver tissue, and miR-146a may negatively regulate the I/R inflammatory liver injury mediated by TLR signaling pathway.
