NETs在促进肺泡Ⅱ型上皮细胞分泌炎性介质中的作用
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江苏省妇幼健康重点人才项目(FRC201735);


Relevant studies that NETs promote the secretion of inflammatory media-tors by alveolar-type epithelial cells
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    摘要:

    目的:探讨中性粒细胞胞外诱捕网(NETs)在促进肺泡Ⅱ型上皮细胞(AECII)分泌炎性介质中的作用。方法:选取出生7 d的30只SD大鼠为研究对象,抽取心脏血液,提取中性粒细胞,按处理方法的不同分成对照组(Control组)、LPS组及LPS+DNase组,使用免疫荧光法检测各组中性粒细胞体外生成NETs情况。对NETs与AECII进行共培养,按细胞悬液孵育方法的不同分为对照组(Control组)、NETs组及NETs+DNase组,使用流式细胞术检测AECII凋亡情况;使用荧光酶标仪检测各组共培养上清液中NETs游离脱氧核糖核酸(cf-DNA)水平;使用ELISA试剂盒测定各组共培养上清液中白细胞介素6(IL-6)、白细胞介素8(IL-8)、肿瘤坏死因子α(TNF-α)浓度,并使用Pearson相关性分析评价cf-DNA与IL-6、IL-8、TNF-α间的相关性。结果:经免疫荧光检测,LPS组与LPS+DNase组NE中均发现小鼠瓜氨酸组蛋白H3(CitH3)与髓过氧化物酶(MPO),LPS组与LPS+DNase组CitH3、MPO水平均高于Control组(P<0.05),LPS+DNase组CitH3、MPO较LPS组更低(P<0.05)。与NE、NETs共培养后AECII凋亡情况,三组早期凋亡与晚期凋亡比例无统计学差异(P>0.05)。NETs组与NETs+DNase组的细胞共培养上清液中cf-DNA、IL-6、IL-8、TNF-α水平均高于Control组(P<0.05);且NETs+DNase组低于NETs组(P<0.05)。细胞培养上清液中cf-DNA水平与IL-6、IL-8、TNF-α水平正相关(r=0.432、0.387、0.326,P<0.05)。结论:AECII与中性粒细胞共培养上清液中存在NETs, NETs可促进AECII分泌IL-6、IL-8、TNF-α等炎性介质,且NETs生成越多,IL-6、IL-8、TNF-α等炎性介质分泌越多。

    Abstract:

    Objective:To explore the role of neutrophil extracellular traps(NETs)in promoting the secretion of inflammatory mediators by alveolar type epithelial cells(AECII).Methods:In 30 7 d born SD rats,blood was drawn from neutrophils(NE)and di-vided into control(control group),LPS and LPS+DNase groups according to different treatment methods.NETs formation was detec-ted by immunofluorescence in vitro.NETs and AECII were co-cultured,and divided into control group(control group),NETs group and NETs+DNase group according to cell suspension incubation method.AECII apoptosis was detected by flow method,the content of free deoxyribonucleic acid(cf-DNA)in coculture supernatants in each group was detected by fluorescence microarray,and the coculture su-pernatants of interleukin 6(IL-6),interleukin 8(IL-8)and tumor necrosis factor(TNF-α)in each group were determined using ELISA kit.And,the correlation between cf-DNA and IL-6,IL-8,and TNF-α was evaluated using Pearson correlation analysis.Results:According to immunofluorescence,mouse citrullinated histone H3(CitH3)and myeloperoxidase(MPO)were found in neutrophil in LPS and LPS+DNase group,CitH3 and MPO levels in LPS and LPS+DNase groups were higher than in the control group(P<0.05),and CitH3 and MPO in LPS+DNase group were lower than those in LPS group(P<0.05).After AECII apoptosis with NE and NETs,there was no significant difference between early and late apoptosis in the three groups(P>0.05).The levels of cf-DNA,IL-6,IL-8,and TNF-α in NETs group and NETs+DNase group were higher than those of the control group(P<0.05),NETs+DNase group was lower than NETs group(P<0.05).The content of cf-DNA in cell culture supernatant was positively correlated with the levels of IL-6,IL-8,and TNF-α(r=0.432,0.387,0.326,P<0.05).Conclusion:AECII with NETs in the supernatants with NE,NETs could promote the secretion of inflammatory mediators such as IL-6,IL-8,and TNF-α.Moreover,the more NETs are generated,the more inflammatory mediators such as IL-6,IL-8,and TNF-α are secreted.

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徐苏晴;田佳梅;卢红艳;. NETs在促进肺泡Ⅱ型上皮细胞分泌炎性介质中的作用[J].川北医学院学报,2025,40(5):557-561.

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  • 在线发布日期: 2025-07-09
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