甲醛对嗜酸性粒细胞EOL-1的急性损伤作用机制  

Mechanism of acute injury of eosinophil EOL-1 induced by formaldehyde

作　　者：李娜[1] 郭增丽 迟令懿[3] 杨立卓 马志勇[1] 付志婕 LI Na;GUO Zengli;CHI Lingyi;YANG Lizhuo;MA Zhiyong;FU Zhijie(Department of Cardiology,Qilu Hospital of Shandong University,The Key Laboratory of Cardiovascular Remodeling and Function Research,Chinese Ministry of Education,Chinese National Health Commission and Chinese Academy of Medical Sciences,Jinan 250012,Shandong,China;Department of Otorhinolaryngology,The First Affiliated Hospital of Shandong First Medical University,Jinan 250014,Shandong,China;Department of Neurosurgery,Qilu Hospital of Shandong University,Jinan 250012,Shandong,China)

机构地区：[1]山东大学齐鲁医院心内科,教育部和国家卫健委心血管重构与功能研究重点实验室,山东济南250012 [2]山东第一医科大学第一附属医院耳鼻咽喉科,山东济南250014 [3]山东大学齐鲁医院神经外科,山东济南250012

出　　处：《山东大学学报（医学版）》2022年第11期54-62,101,共10页Journal of Shandong University:Health Sciences

基　　金：国家自然科学基金(81700891);山东省自然科学基金(ZR2020MH038);山东第一医科大学学术提升计划(2019QL015)

摘　　要：目的 探讨甲醛对人嗜酸性粒细胞(EOL-1)的急性损伤作用及相关机制。方法 体外培养EOL-1细胞,将甲醛室温处理2 h的EOL-1细胞设置为0 mmol组、5 mmol组、10 mmol组、25 mmol组及50 mmol组;同时将3种活性氧(ROS)通路抑制剂(NADPH氧化酶抑制剂DPI、细胞透性超氧化物清除剂Tiron、谷胱甘肽稳定前体NAC)和25 mmol甲醛共同处理后的细胞设置为对照组、甲醛组、甲醛+DPI组、甲醛+Trion组和甲醛+NAC组;依据NAC浓度的不同,设置为对照组、甲醛组、甲醛+0.01 mmol NAC组、甲醛+0.1 mmol NAC组和甲醛+1 mmol NAC组。采用碘化丙啶(PI)和Hoechst荧光染色法检测各组细胞凋亡和死亡的表达,采用罗丹明123 (Rho-123)荧光标记法检测细胞线粒体功能损伤,采用Western blotting法检测损伤信号通路蛋白Bax、Bcl-2的表达。结果 不同浓度甲醛组(0、5、10、25、50 mmol)细胞死亡率分别为(3.313±2.395)%、(8.205±5.719)%、(20.335±5.167)%、(19.387±6.056)%、(28.043±8.851)%,与0 mmol组比较,5 mmol组细胞凋亡和死亡无增加(P=1.00),但是10 mmol组(P=0.030)、25 mmol组(P=0.033)和50 mmol组(P=0.001)甲醛处理细胞死亡显著增加,其半数效应浓度EC50=25 mmol;甲醛组、甲醛+DPI组、甲醛+Trion组、甲醛+NAC组细胞死亡率分别为(61.430±9.885)%、(57.907±13.619)%、(55.700±18.487)%、(21.837±6.674)%,与甲醛组比较,甲醛+DPI组(P=1.00)、甲醛+Trion组(P=1.00)对甲醛诱导的细胞损伤无影响,但是甲醛+NAC组逆转甲醛诱导的细胞死亡(P=0.01);与甲醛组(52.853±11.338)%对比,随着NAC浓度的不同,细胞死亡率不同[甲醛+0.01 mmol NAC组(10.620±4.483)%,甲醛+0.1 mmol NAC组(6.257±6.265)%,甲醛+1 mmol NAC组(4.002±2.50)%],NAC对甲醛诱导的细胞死亡呈浓度依赖性的逆转作用。Rho-123荧光标记结果显示,与0 mmol组相比,10 mmol组、25 mmol组、50 mmol组可以降低线粒体功能(P<0.001),而甲醛+NAC组逆转甲醛诱导的线粒体功能损伤(809.339±163.210 vs 675.552±Objective To explore the acute injury effect of formaldehyde on human eosinophils(EOL-1) and the mechanism. Methods EOL-1 cells were cultured in vitro. Eol-1 cells treated with formaldehyde for 2 hours at room temperature were set as the 0 mmol, 5 mmol, 10 mmol, 25 mmol and 50 mmol groups. Cells co-treated with DPI, Tiron and NAC and 25 mmol formaldehyde were divided into control group, formaldehyde group, formaldehyde + DPI group, formaldehyde + Trion group, and formaldehyde + NAC group. According to the different concentrations of NAC, the cells were set as the control group, formaldehyde group, formaldehyde + 0.01 mmol NAC group, formaldehyde + 0.1 mmol NAC group, and formaldehyde +1 mmol NAC group. Cell apoptosis and death were detected with propidium iodide(PI) and Hoechst fluorescent staining. The mitochondrial function damage was detected with rhodamine 123(Rho-123) fluorescent labeling method. The expressions of Bax and Bcl-2 were detected with Western blotting. Results The cell death rates of different concentrations of formaldehyde(0, 5, 10, 25, 50 mmol) were(3.313±2.395)%,(8.205±5.719)%,(20.335±5.167)%,(19.387±6.056)%, and(28.043±8.851)%, respectively. Compared with the 0 mmol group, the 5 mmol group had unchanged cell apoptosis and death(P=1.00), but the 10 mmol(P=0.030), 25 mmol(P=0.033) and 50 mmol groups(P=0.001) had significantly increased cell death, with EC50=25 mmol. The cell mortality rates of formaldehyde group, formaldehyde + DPI group, formaldehyde + Trion group and formaldehyde + NAC group were(61.430±9.885)%,(57.907±13.619)%,(55.700±18.487)% and(21.837±6.674)%, respectively. Formaldehyde + DPI group(P=1.00) and formaldehyde + Trion group(P=1.00) had no effect on formaldehyde-induced cell damage, but formaldehyde + NAC group reversed the formaldehyde-induced cell death(P=0.01). Compared with formaldehyde group(52.853±11.338)%, with different concentrations of NAC, the cell mortality was different [formaldehyde + 0.01 mmol NAC(10.620±4.483)%, formaldehyde + 0.1 mmol NAC(6.257±6.

关 键 词：甲醛 EOL-1细胞 细胞死亡 线粒体功能 信号通路 

分 类 号：R114[医药卫生—卫生毒理学]