Role of cholinergic anti-inflammatory pathway in Ghrelin regulation of peptide transporter 1 expression in small intestinal epithelium of septic rats
10.3760/cma.j.cn121430-20210817-01197
- VernacularTitle:胆碱能抗炎通路在Ghrelin调控脓毒症大鼠小肠上皮PepT1表达中的作用
- Author:
Ziqiang SHAO
1
;
Jun HONG
;
Minhua CHEN
;
Yang ZHENG
;
Zongbin LIN
;
Xianghong YANG
;
Renhua SUN
;
Jingquan LIU
Author Information
1. 浙江省人民医院（杭州医学院附属人民医院）急危重症中心重症医学科，杭州 310014
- Keywords:
Sepsis;
Cholinergic anti-inflammatory pathway;
Ghrelin;
Intestinal epithelial cell
- From:
Chinese Critical Care Medicine
2022;34(11):1132-1137
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the role of cholinergic anti-inflammatory pathway in the regulation of peptide transporter 1 (PepT1) expression in small intestinal epithelium of septic rats by Ghrelin.Methods:One hundred adult male Sprague-Dawley (SD) rats were randomly divided into sham operation group, sepsis group, sepsis+vagotomy group, sepsis+Ghrelin group, and sepsis+vagotomy+Ghrelin group, with 20 rats in each group. In the sham operation group, the cecum was separated after laparotomy, without ligation and perforation. In the sepsis group, the rats received cecal ligation puncture (CLP). In the sepsis+vagotomy group, the rats received CLP and vagotomy after laparotomy. In the sepsis+Ghrelin group, 100 μmol/L Ghrelin was intravenously injected after CLP immediately. The rats in the sepsis+vagotomy+Ghrelin group received CLP and vagotomy at the same time, then the Ghrelin was intravenously injected immediately with the same dose as the sepsis+Ghrelin group. Ten rats in each group were taken to observe their survival within 7 days. The remaining 10 rats were sacrificed 20 hours after the operation to obtain venous blood and small intestinal tissue. The condition of the abdominal intestine was observed. The injury of intestinal epithelial cells was observed with transmission electron microscopy. The contents of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in serum and small intestinal tissue were detected by enzyme-linked immunosorbent assay (ELISA). The brush border membrane vesicle (BBMV) was prepared, the levels of mRNA and protein expression of PepT1 in the small intestinal epithelium were detected by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) and Western blotting.Results:All rats in the sham operation group survived at 7 days after operation. The 7-day cumulative survival rate of rats in the sepsis group was significantly lower than that in the sham operation group (20% vs. 100%, P < 0.05). The cumulative survival rate of rats after Ghrelin intervention was improved (compared with sepsis group: 40% vs. 20%, P < 0.05), but the protective effect of Ghrelin was weakened after vagotomy (compared with sepsis+Ghrelin group: 10% vs. 40%, P < 0.05). Compared with the sham operation group, in the sepsis group, the small intestine and cecum were dull red, the intestinal tubules were swollen and filled with gas, the intestinal epithelial cells were seriously injured under transmission electron microscopy, the levels of TNF-α and IL-1β in serum and small intestinal were significantly increased, and the expression levels of PepT1 mRNA and protein in the small intestinal epithelium were significantly decreased. It indicated that the sepsis rat model was successfully prepared. After vagotomy, the intestinal swelling and gas accumulation became worse in septic rats, leading to the death of all rats. Compared with the sepsis group, the abdominal situation in the sepsis+Ghrelin group was improved, the injury of intestinal epithelial cells was alleviated, the serum and small intestinal TNF-α and IL-1β were significantly decreased [serum TNF-α (ng/L): 253.27±23.32 vs. 287.90±19.48, small intestinal TNF-α (ng/L): 95.27±11.47 vs. 153.89±18.15, serum IL-1β (ng/L): 39.16±4.47 vs. 54.26±7.27, small intestinal IL-1β (ng/L): 28.47±4.13 vs. 42.26±2.59, all P < 0.05], and the expressions of PepT1 mRNA and protein in the small intestinal epithelium were significantly increased [PepT1 mRNA (2 -ΔΔCt): 0.66±0.05 vs. 0.53±0.06, PepT1 protein (PepT1/GAPDH): 0.80±0.04 vs. 0.60±0.05, both P < 0.05]. Compared with the sepsis+Ghrelin group, after vagotomy in the sepsis+vagotomy+Ghrelin group, the effect of Ghrelin on reducing the release of inflammatory factors in sepsis rats was significantly reduced [serum TNF-α (ng/L): 276.58±19.88 vs. 253.27±23.32, small intestinal TNF-α (ng/L): 144.28±12.99 vs. 95.27±11.47, serum IL-1β (ng/L): 48.15±3.21 vs. 39.16±4.47, small intestinal IL-1β (ng/L): 38.75±4.49 vs. 28.47±4.13, all P < 0.05], the up-regulated effect on the expression of PepT1 in small intestinal epithelium was lost [PepT1 mRNA (2 -ΔΔCt): 0.58±0.03 vs. 0.66±0.05, PepT1 protein (PepT1/GAPDH): 0.70±0.02 vs. 0.80±0.04, both P < 0.05], and the injury of small intestinal epithelial cells was worse. Conclusion:Ghrelin plays a protective role in sepsis by promoting cholinergic neurons to inhibit the release of inflammatory factors, thereby promoting the transcription and translation of PepT1.
