关键词: RNA, 环状, 微RNAs, 竞争性内源性RNA, 痤疮丙酸杆菌, HaCaT细胞, 白细胞介素类, 肿瘤坏死因子α

Abstract: 【Abstract】 Objective To investigate the effects of circular RNA circ_0086376 on the expression of acne-related inflammatory factors by targeting microRNA （miR）-5195-3p. Methods Circ_0086376-overexpressing or -interferred stable HaCaT cell lines were constructed and co-cultured with Propionibacterium acnes （P.acne）. Quantitative real-time PCR was used to detect the effect of co-culture on circ_0086376 expression, while enzyme-linked immunosorbent assay （ELISA） was performed to measure the expression of inflammatory factors in the cell culture supernatant. The overexpression or interference of circ_0086376 plasmid and lentivirus mimicking or inhibiting miR-5195-3p fragment were transfected into HaCaT cells, and ELISA was used to detect the expression of inflammatory factors in the culture supernatant of HaCaT cells after overexpression or interference of circ_0086376 cultured alone or with P.acne. The luciferase reporter assay was conducted to verify the targeting relationship between circ_0086376 and miR-5195-3p. Additionally, ELISA was used to detect the effects of overexpression/interference of circ_0086376 and mimic/inhibition of miR-5195-3p on the expression of inflammatory factors in the culture supernatant of HaCaT cells co-cultured with P.acne. Results After co-culture with P.acne, the levels of inflammatory factors in the culture supernatant were significantly higher than those in the HaCaT cells cultured alone, including Interleukins （IL）-1β （［355.80 ± 23.20］ vs. ［260.50 ± 16.58］ pg/ml, t = 5.79, P < 0.01）, IL-6 （［38.04 ± 2.69］ vs. ［14.33 ± 0.75］ pg/ml, t = 14.65, P < 0.01）, IL-12 （［10.87 ± 0.78］ vs. ［6.52 ± 0.77］ pg/ml, t = 6.89, P < 0.01）, IL-18 （［222.60 ± 21.07］ vs. ［146.10 ± 9.14］ pg/ml, t = 5.77, P < 0.01）, and Tumor necrosis factor （TNF）-α （［50.39 ± 1.29］ vs. ［20.46 ± 0.83］ pg/ml, t = 33.83, P < 0.01）. The expression levels of IL-1β, IL-6, IL-12, IL-18 and TNF-α in the culture supernatant of HaCaT cells in the circ-empty vector + P.acne group were higher than those in the circ-empty vector overexpression group. The levels of inflammatory factors mentioned above in circ-overexpression + P.acne group were lower than those in circ-overexpression empty vector + P.acne group （P < 0.01）. The expression levels of inflammatory factors mentioned above in the culture supernatant of HaCaT cells in the interference circ-empty vector + P.acne group were higher than those in the interference circ-empty vector group. Compared with the interference circ-empty vector + P.acne group, the expression of circ in the interference circ + P.acne group was higher （P < 0.01）. Luciferase reporter assay confirmed that circ_0086376 could bind to miR-5195-3p. The expression levels of IL-1β, IL-6, IL-12, IL-18 and TNF-α in the circ-overexpression group were lower than those in the empty vector group （P < 0.05）, and the levels of these inflammatory factors in the circ-overexpression + miR mimic group were higher than those in the circ overexpression group （P < 0.05）. The expression levels of inflammatory factors in the interference circ group were higher than those in the empty vector group, levels of inflammatory factors in the circ + miR-inhibiting group were lower than those in the circ interference group （P < 0.05）. Conclusion Circ_0086376 can inhibit the expression of acne-related inflammatory factors by targeting and downregulating miR-5195-3p.

Key words: RNA, circular, MicroRNAs, Competitive endogenous RNA, Propionibacterium acnes, HaCaT cells, Interleukins, Tumor necrosis factor-alpha