长链非编码 RNA CDKN2B-AS1 通过海绵吸附 miR-16 和 miR-195 缓解溃疡性结肠炎的炎症。

LncRNA CDKN2B-AS1 relieved inflammation of ulcerative colitis via sponging miR-16 and miR-195.

机构信息

Department of Gastroenterology, Hainan General Hospital & Hainan Affiliated Hospital of Hainan Medical University, Haikou City, Hainan Province 570311, China.

Department of Pathology, Haikou Hospital of Traditional Chinese Medicine, Haikou City, Hainan Province 570216, China.

出版信息

Int Immunopharmacol. 2020 Nov;88:106970. doi: 10.1016/j.intimp.2020.106970. Epub 2020 Oct 14.

DOI:10.1016/j.intimp.2020.106970

PMID:33182065

Abstract

BACKGROUND

This study was aimed to explore the differential expression of lncRNA CDKN2B-AS1-miR-195-5p/miR-16-5p axis in ulcerative colitis (UC) and its role in regulating UC pathogenesis.

METHODS

One hundred and eighty-seven UC patients and one hundred and fifty-two healthy volunteers were recruited, and their blood samples were collected. Inflammatory cytokines in serum were determined with ELISA, and lncRNA CDKN2B-AS1, miR-195-5p and miR-16-5p levels were detected with RT-PCR. Then pcDNA3.1-CDKN2B-AS1, si-CDKN2B-AS1, miR-195-5p mimic, miR-195-5p inhibitor, miR-16-5p mimic and miR-16-5p inhibitor were transfected into HT29 cells, and proliferation and apoptosis of the cells were assessed. Dual-luciferase reporter gene assay was implemented to identify the sponging relationship between lncRNA CDKN2B-AS1 and miR-195-5p/miR-16-5p.

RESULTS

CDKN2B-AS1 level was negatively correlated with levels of inflammatory cytokines, including TNF-α, IL-6 and sIL-2R, yet miR-16-5p and miR-195-5p levels were negatively correlated with the CDKN2B-AS1 level. The CDKN2B-AS1 combined with miR-16-5p and miR-195-5p also achieved an optimum efficacy in differentiating between light and medium UC, light and severe UC, as well as medium and heavy UC. Furthermore, pcDNA3.1-CDKN2B-AS1 depressed expressions of IFN-γ, IL-8, IL-1β and TNF-α in HT29 cells (P < 0.05), and strengthened proliferation of the cells (P < 0.05). CDKN2B-AS1 also sponged and regulated miR-16-5p and miR-195-5p in HT29 cells, and miR-16-5p and miR-195-5p could reverse the effect of CDKN2B-AS1 on inflammatory cytokine production, barrier function and apoptosis of HT29 cells (P < 0.05).

CONCLUSION

LncRNA CDKN2B-AS1 regulated inflammation of UC by sponging miR-195-5p and miR-16-5p, providing an alternative for diagnosis and treatment of UC.

摘要

背景

本研究旨在探讨长链非编码 RNA CDKN2B-AS1-miR-195-5p/miR-16-5p 轴在溃疡性结肠炎（UC）中的差异表达及其在调节 UC 发病机制中的作用。

方法

招募 187 例 UC 患者和 152 例健康志愿者，采集其血样。采用 ELISA 法测定血清中炎症细胞因子水平，采用 RT-PCR 法检测 lncRNA CDKN2B-AS1、miR-195-5p 和 miR-16-5p 水平。然后将 pcDNA3.1-CDKN2B-AS1、si-CDKN2B-AS1、miR-195-5p 模拟物、miR-195-5p 抑制剂、miR-16-5p 模拟物和 miR-16-5p 抑制剂转染 HT29 细胞，评估细胞增殖和凋亡。实施双荧光素酶报告基因检测鉴定 lncRNA CDKN2B-AS1 与 miR-195-5p/miR-16-5p 的海绵关系。

结果

CDKN2B-AS1 水平与 TNF-α、IL-6 和 sIL-2R 等炎症细胞因子水平呈负相关，而 miR-16-5p 和 miR-195-5p 水平与 CDKN2B-AS1 水平呈负相关。CDKN2B-AS1 与 miR-16-5p 和 miR-195-5p 的联合也能最佳地区分轻中度 UC、轻重度 UC 和中重度 UC。此外，pcDNA3.1-CDKN2B-AS1 下调 HT29 细胞 IFN-γ、IL-8、IL-1β 和 TNF-α 的表达（P<0.05），增强细胞增殖（P<0.05）。CDKN2B-AS1 还能在 HT29 细胞中海绵吸附和调节 miR-16-5p 和 miR-195-5p，miR-16-5p 和 miR-195-5p 能逆转 CDKN2B-AS1 对 HT29 细胞炎症细胞因子产生、屏障功能和凋亡的影响（P<0.05）。