肠道黏膜 MicroRNAs，miR-219a-5p 和 miR-338-3p 在肠易激综合征中下调，与屏障功能和 MAPK 信号有关。
The Colonic Mucosal MicroRNAs, MicroRNA-219a-5p, and MicroRNA-338-3p Are Downregulated in Irritable Bowel Syndrome and Are Associated With Barrier Function and MAPK Signaling.
机构信息
G. Oppenheimer Center for Neurobiology of Stress and Resilience, Division of Digestive Diseases, Department of Medicine, University of California, Los Angeles, Los Angeles, California.
UCLA Center for Inflammatory Bowel Diseases, Division of Digestive Diseases, Department of Medicine, University of California, Los Angeles, Los Angeles, California.
出版信息
Gastroenterology. 2021 Jun;160(7):2409-2422.e19. doi: 10.1053/j.gastro.2021.02.040. Epub 2021 Feb 20.
BACKGROUND & AIMS: Alterations in microRNA (miRNA) and in the intestinal barrier are putative risk factors for irritable bowel syndrome (IBS). We aimed to identify differentially expressed colonic mucosal miRNAs, their targets in IBS compared to healthy controls (HCs), and putative downstream pathways.
METHODS
Twenty-nine IBS patients (15 IBS with constipation [IBS-C], 14 IBS with diarrhea [IBS-D]), and 15 age-matched HCs underwent sigmoidoscopy with biopsies. A nCounter array was used to assess biopsy specimen-associated miRNA levels. A false discovery rate (FDR) < 10% was considered significant. Real-time polymerase chain reaction (PCR) was used to validate differentially expressed genes. To assess barrier function, trans-epithelial electrical resistance (TEER) and dextran flux assays were performed on Caco-2 intestinal epithelial cells that were transfected with miRNA-inhibitors or control inhibitors. Protein expression of barrier function associated genes was confirmed using western blots.
RESULTS
Four out of 247 miRNAs tested were differentially expressed in IBS compared to HCs (FDR < 10%). Real-time PCR validation suggested decreased levels of miR-219a-5p and miR-338-3p in IBS (P = .026 and P = .004), and IBS-C (P = .02 and P = .06) vs. HCs as the strongest associations. Inhibition of miR-219a-5p resulted in altered expression of proteasome/barrier function genes. Functionally, miR-219a-5p inhibition enhanced the permeability of intestinal epithelial cells as TEER was reduced (25-50%, P < .05) and dextran flux was increased (P < .01). Additionally, inhibition of miR-338-3p in cells caused alterations in the mitogen-activated protein kinase (MAPK) signaling pathway genes.
CONCLUSION
Two microRNAs that potentially affect permeability and visceral nociception were identified to be altered in IBS patients. MiR-219a-5p and miR-338-3p potentially alter barrier function and visceral hypersensitivity via neuronal and MAPK signaling and could be therapeutic targets in IBS.
背景与目的
微小 RNA（miRNA）和肠道屏障的改变被认为是肠易激综合征（IBS）的潜在危险因素。我们旨在鉴定与健康对照（HC）相比 IBS 患者结肠黏膜中差异表达的 miRNA 及其靶标，并确定潜在的下游途径。
方法
29 例 IBS 患者（15 例 IBS 伴便秘[IBS-C]，14 例 IBS 伴腹泻[IBS-D]）和 15 名年龄匹配的 HCs 接受乙状结肠镜检查和活检。使用 nCounter 阵列评估活检标本相关 miRNA 水平。假发现率（FDR）<10%被认为具有统计学意义。实时聚合酶链反应（PCR）用于验证差异表达基因。为了评估屏障功能，用 miRNA 抑制剂或对照抑制剂转染 Caco-2 肠上皮细胞后，进行跨上皮电阻（TEER）和葡聚糖通量测定。使用蛋白质印迹法确认与屏障功能相关的基因的蛋白表达。
结果
在 IBS 与 HCs 相比，247 个测试 miRNA 中有 4 个（FDR<10%）表达差异。实时 PCR 验证提示 miR-219a-5p 和 miR-338-3p 在 IBS（P=0.026 和 P=0.004）和 IBS-C（P=0.02 和 P=0.06）中的水平降低，这是最强的关联。miR-219a-5p 的抑制导致蛋白酶体/屏障功能基因的表达改变。功能上，miR-219a-5p 的抑制降低了肠上皮细胞的通透性（25%-50%，P<0.05），并增加了葡聚糖通量（P<0.01）。此外，细胞中 miR-338-3p 的抑制导致丝裂原激活蛋白激酶（MAPK）信号通路基因的改变。
结论
两种潜在影响通透性和内脏痛觉过敏的 miRNA 在 IBS 患者中发生改变。miR-219a-5p 和 miR-338-3p 可能通过神经元和 MAPK 信号通路改变屏障功能和内脏敏感性，可能成为 IBS 的治疗靶点。
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