Mechanism of Circadian Clock Gene Bmal1 Regulating Clock-controlled Gene Piezo1 Involved in TPH1-5-HT Signaling Pathway in Enterochromaffin Cells
10.3969/j.issn.1008-7125.2024.09.001
- VernacularTitle:生物钟基因Bmal1调控肠嗜铬细胞钟控基因Piezo1影响TPH1-5-HT通路的分子机制
- Author:
Lu ZOU
1
;
Yi LI
;
Yuanfeng REN
;
Wenbo LI
;
Zongsheng HE
;
Dongfeng CHEN
;
Huaping LIANG
;
Min YANG
Author Information
1. 陆军军医大学陆军特色医学中心(大坪医院)消化内科(400042)
- Publication Type:Journal Article
- Keywords:
Enterochromaffin Cell Model;
Biological Clock Gene;
Mechanosensitive Ion Channel Piezo1;
5-Hydroxytryptamine
- From:
Chinese Journal of Gastroenterology
2024;29(9):513-520
- CountryChina
- Language:Chinese
-
Abstract:
Background:The core circadian clock gene Bmal1 has been shown to be involved in the formation of visceral sensitization in irritable bowel syndrome(IBS)by affecting the tryptophan hydroxylase 1(TPH1)-5-hydroxytryptamine(5-HT)pathway,but the exact mechanism of its regulation is unknown.Aims:To investigate the molecular mechanism by which Bmal1 regulates the TPH1-5-HT pathway through the clock-controlled gene Piezo1.Methods:Dexamethasone was used to synchronize the expression of the circadian clock genes in RIN-14B cells.Bmal1 expression was up-regulated or down-regulated in RIN-14B cells by plasmid and siRNA transfection of the enterochromaffin cell(EC)model.The expression levels of target genes and proteins were detected by immunofluorescence staining,RT-qPCR,and Western blotting.5-HT content was detected by ELISA method.Results:(1)This study was the first to report the oscillation characteristics of RIN-14B circadian clock genes in EC model,among which the oscillation of Bmal1 was the most significant.Immunofluorescence showed that RIN-14B cells expressed CGA,Bmal1 and Piezo1.(2)After transfected with the Bmal1 overexpression plasmid,the mRNA and protein expression of Bmal1 were significantly up-regulated in RIN-14B cells compared with the negative control group(all P<0.001);while transfected with Bmal1 siRNA significantly decreased the mRNA and protein expression of Bmal1 in RIN-14B cells compared with the negative control group(all P<0.05).(3)After transfected with Bmal1 overexpression plasmid,the mRNA and protein expression of Piezo1,the protein expression of TPH1,and the intracellular content of 5-HT were significantly increased(all P<0.051).(4)After transfected with Bmal siRNA,mRNA expression of Piezo1 and TPH1 in RIN-14B cells was significantly down-regulated(all P<0.05),and the protein expression of Piezo1,TPH1,the intracellular 5-HT content tended to be decreased by 21%,31%,and 10%,respectively.Conclusions:RIN-14B cells have the characteristics of rhythmic oscillation of circadian clock genes,and Bmal1 overexpression and underexpression EC models can be successfully established by using RIN-14B cells.Overexpression and underexpression of Bmal1 can lead to significant changes in the clock-controlled Piezo1-TPH1-5-HT signaling pathway,suggesting that Bmal1 can be expressed by clock-control signals through the Piezo1-TPH1 pathway.This suggests that Bmal1 may be involved in the development of visceral hypersensitivity in IBS through regulation of 5-HT synthesis by the clock-controlled gene Piezo1.
