Basic Study
Copyright ©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Feb 7, 2020; 26(5): 478-498
Published online Feb 7, 2020. doi: 10.3748/wjg.v26.i5.478
lncRNACNN3-206 activates intestinal epithelial cell apoptosis and invasion by sponging miR-212, an implication for Crohn's disease
Na Li, Rui-Hua Shi
Na Li, Rui-Hua Shi, Medical School of Southeast University, Nanjing 210009, Jiangsu Province, China
Na Li, Rui-Hua Shi, Department of Gastroenterology, Zhongda Hospital, Affiliated Hospital of Southeast University, Nanjing 210009, Jiangsu Province, China
Na Li, Department of Biomedical Science, Mercer University School of Medicine, Savannah, GA 31404, United States
Author contributions: Shi RH supervised the projects and designed experiments; Li N performed all the experiments; Li N and Shi RH collected normal ileal epithelial tissues and ileal epithelial tissues from patients with Crohn's disease; Li N summed up the experimental results and prepared the first manuscript; and Shi RH made revision.
Supported by Postgraduate Research and Practice Innovation Program of Jiangsu Province, No. KYCX18_0174.
Institutional review board statement: This research was approved by the Ethics Committee of Zhongda Hospital, Affiliated to Southeast University.
Institutional animal care and use committee statement: This research was approved by Animal Experimental Ethical Inspection of Southeast University, NO. 20190227011.
Conflict-of-interest statement: The authors do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Corresponding author: Rui-Hua Shi, PhD, Professor, Chief Physician, Department of Gastroenterology, Zhongda Hospital, Affiliated Hospital of Southeast University, Dingjiaqiao Road 87, Nanjing 210009, Jiangsu Province, China.
Received: October 9, 2019
Peer-review started: October 9, 2019
First decision: December 5, 2019
Revised: December 20, 2019
Accepted: January 8, 2020
Article in press: January 8, 2020
Published online: February 7, 2020
Research background

Crohn's disease (CD) is a chronic, recurrent, inflammatory disorder of the digestive tract, with clinical manifestations of abdominal pain, diarrhea, enterelcosis and intestinal obstruction. Statistics showed that CD incidence rates are climbing in Europe and the United States, as well as in Asian countries.

Research motivation

Due to a poor understanding of the etiology and pathogenesis of this disease, current therapy is limited to anti-symptomatic and surgical treatments. The recurrence rate within 3 years after surgery is as high as 80-100%. Exploration of the cellular and molecular mechanisms is urgently required to develop new therapeutic modalities against this disease. While many studies have indicated that long noncoding RNAs (lncRNAs) might participate in the pathogenesis of autoimmune diseases, very few studies focused on whether lncRNAs may contribute to CD occurrence. Previous studies using high-throughput gene sequencing techniques have shown that the serum of CD patients contained multiple differentially expressed lncRNAs. While these differentially expressed lncRNAs could be used as biomarkers for the diagnosis and/or prognosis of CD, their sources remain unclear.

Research objectives

Our goal is to identify the differentially expressed lncRNAs in the intestinal mucosa associated with CD, and to characterize their pathogenic role(s) and related mechanisms.

Research methods

In this study, we collected tissue samples of the ileal lesions from CD patients and normal intestinal mucosal tissue samples isolated during physical examinations of healthy patients. The expression patterns of lncRNAs and mRNAs were compared between the lesions and normal ileal tissue samples by microarray hybridization. Bioinformatic analysis was performed to predict the gene regulatory network of lncRNAs-miRNAs-mRNAs. The biological functions of the lncRNAs, relevant miRNAs and target genes, as well as the related regulatory mechanisms, were comprehensively investigated using cell culture and animal models.

Research results

Our studies identified lncRNACNN3-206 as a highly upregulated non-coding RNA in the intestinal lesions of CD patients. Manipulation of lncRNACNN3-206 in cell culture led to significant changes in Caspase10 expression, cell apoptosis and invasion activity. Experiments using a CD mouse model demonstrated that lncRNACNN3-206 knockdown could alleviate some CD manifestations, most likely through inhibition of intestinal epithelial cell apoptosis and inflammatory responses.

Research conclusions

Overexpression of lncRNACNN3-206 in intestinal epithelial cells led to increased cell apoptosis and invasion. These changes in cell function may play important roles in CD pathogenesis. These findings do not only facilitate our understanding of the pathological mechanisms but also have strong clinical implications. The upregulated lncRNACNN3-206 gene could be a new therapeutic target for the prevention and treatment of CD.

Research perspectives

The top 10 lncRNAs with differential expression were screened out by high-throughput sequencing, but only six lncRNAs with true differential expression were identified by PCR validation in expanded sample tissues. All of the subjects had to therefore be definitively confirmed using a large number of clinical tissue samples. Moreover, experimental research insight is given through the signaling pathways predicted by bioinformatics websites, as well as the knowledge of signaling pathways predicted by GO/Pathway analysis, which requires the detection of specific biological gene functions through cell experiments. The predicted function is sometimes experimentally shown to be absent in the cell. In this study, due to limited conditions, the targeted binding of lncRNACNN3-206 and miR212 was demonstrated by luciferase reporter assays. Nevertheless, the better method is RIP technology, which can directly observe the adsorption of miR212 by lncRNA as a sponge.