背景:CircRNAs与许多人类疾病密切相关,然而,它们在痤疮中的作用尚不清楚。本研究旨在确定hsa-circ_0102678在调节痤疮炎症中的作用。
方法:首先,进行微阵列分析以研究circRNAs在痤疮中的表达。随后,利用RNaseR消化测定和FISH测定来确认hsa-circ_0102678的特征。最后,qRT-PCR,蛋白质印迹分析,免疫沉淀,荧光素酶报告基因测定,circRNA探针下拉测定,生物素标记的miRNA下拉测定,利用RNA免疫沉淀(RIP)测定和m6A斑点印迹测定来揭示hsa-circ_0102678在人原代角质形成细胞中由痤疮梭菌生物膜诱导的炎症中的功能作用。
结果:我们的研究表明,在痤疮组织中hsa-circ_0102678的表达显着降低,并且hsa-circ_0102678是一种circRNAs,主要位于原代人角质形成细胞的细胞质中。此外,hsa-circ_0102678显著影响IL-8、IL-6和TNF-α的表达,由痤疮杆菌生物膜诱导。重要的是,机理研究表明,YTHDC1可以直接与hsa-circ_0102678结合,并促进N6-甲基腺苷修饰的hsa-circ_0102678向细胞质的输出。此外,hsa-circ_0102678可以与miR-146a和海绵miR-146a结合以促进IRAK1和TRAF6的表达。
结论:我们的发现揭示了一个先前未知的过程,通过该过程,hsa_circ_0102678通过调节miR-146a/TRAF6和IRAK1轴促进痤疮梭菌生物膜诱导的角质形成细胞炎症。
BACKGROUND: CircRNAs are closely related to many human diseases; however, their role in acne remains unclear. This study aimed to determine the role of hsa_circ_0102678 in regulating inflammation of acne.
METHODS: First, microarray analysis was performed to study the expression of circRNAs in acne. Subsequently, RNase R digestion assay and fluorescence in situ hybridization assay were utilized to confirm the characteristics of hsa_circ_0102678. Finally, qRT-PCR, Western blotting analysis, immunoprecipitation, luciferase reporter assay, circRNA probe pull-down assay, biotin-labeled miRNA pull-down assay, RNA immunoprecipitation assay, and m6A dot blot assay were utilized to reveal the functional roles of hsa_circ_0102678 on inflammation induced by C. acnes biofilm in human primary keratinocytes.
RESULTS: Our investigations showed that the expression of hsa_circ_0102678 was significantly decreased in acne tissues, and hsa_circ_0102678 was a type of circRNAs, which was mainly localized in the cytoplasm of primary human keratinocytes. Moreover, hsa_circ_0102678 remarkably affected the expression of IL-8, IL-6, and TNF-α, which induced by C. acnes biofilm. Importantly, mechanistic studies indicated that the YTHDC1 could bind directly to hsa_circ_0102678 and promote the export of N6-methyladenosine-modified hsa_circ_0102678 to the cytoplasm. Besides, hsa_circ_0102678 could bind to miR-146a and sponge miR-146a to promote the expression of IRAK1 and TRAF6.
CONCLUSIONS: Our findings revealed a previously unknown process by which hsa_circ_0102678 promoted keratinocyte inflammation induced by C. acnes biofilm via regulating miR-146a/TRAF6 and IRAK1 axis.