摘要
目的·研究gasderminD(GSDMD)抑制剂necrosulfonamide(NSA)通过抑制肺上皮细胞焦亡对脂多糖(lipopolysaccharide,LPS)诱导的新生大鼠支气管肺发育不良(bronchopulmonary dysplasia,BPD)肺泡化阻滞的影响。方法·将孕SD大鼠随机分为对照组、BPD组、BPD+NSA组和NSA组,羊膜腔注射LPS建立新生鼠BPD模型。取各组出生后第1、3、7日新生鼠肺组织,通过苏木精-伊红(H-E)染色观察肺泡化情况;利用免疫荧光法检测各组新生鼠肺部GSDMD-N端蛋白表达情况;荧光定量PCR法检测新生鼠肺组织中炎症因子白介素-1β(interleukin-1β,IL-1β)mRNA水平。体外培养小鼠肺泡上皮细胞系MLE-12,给予LPS以及腺苷三磷酸(adenosine triphosphate,ATP)刺激和NSA干预,CCK-8法检测MLE-12细胞活力,Hoechst 33342和碘化丙啶(propidium iodide,PI)染色法检测细胞焦亡水平,免疫荧光法检测MLE-12细胞表面活性剂蛋白C(surfactant protein C,SFTPC)和GSDMD-N端蛋白的表达情况。结果·体内实验结果显示:羊膜腔内注射LPS可导致肺发育受阻,模拟BPD的病理改变;羊膜腔内注射LPS建立的BPD模型中肺泡上皮细胞GSDMD-N端表达升高;NSA干预明显改善了BPD新生鼠的肺发育阻滞并抑制了IL-1β的mRNA表达(均P<0.05)。体外实验结果显示:LPS/ATP刺激下肺泡上皮细胞MLE-12活力下降,发生焦亡;NSA干预提高了肺泡上皮细胞活力并且抑制了焦亡(均P<0.05);NSA上调了LPS/ATP刺激下肺泡上皮细胞SFTPC的表达,抑制了LPS/ATP刺激下肺泡上皮细胞GSDMD-N端表达的上调(均P<0.05)。结论·抑制肺泡上皮细胞焦亡可改善LPS诱导的BPD新生鼠肺组织病理学改变。
Objective·To study the effect of gasdermin D(GSDMD)inhibitor necrosulfonamide(NSA)on alveolarization arrest in lipopolysaccharide(LPS)-induced bronchopulmonary dysplasia(BPD)newborn rats via inhibiting alveolar epithelial cell pyroptosis.Methods·Pregnant SD rats were randomly assigned to four groups as follows:control,BPD,BPD with NSA and NSA group,and then were prepared to receive intra-amniotic injection of LPS.Lung tissues of newborn rats on the first,third and seventh day after birth were stained by hematoxylin-eosin(H-E)to observe lung development.The expressions of GSDMD-N-terminal in lungs of newborn rats in each group were detected by immunofluorescence.The mRNA levels of interleukin-1β(IL-1β)of newborn rats'lungs was detected by real-time PCR.In vitro,the mouse alveolar epithelial cell line MLE-12 was cultured and treated with LPS/adenosine triphosphate(ATP)and NSA.The cell viability of MLE-12 cells was detected by CCK-8 method,the pyroptosis was detected by Hoechst 33342 and propidium iodide(PI)staining,and the expressions of surfactant protein C(SFTPC)and GSDMD-N protein in MLE-12 cells were detected by immunofluorescence.Results·In vivo,intra-amniotic injection of LPS hindered lung development,resulting in the pathological hallmarks of BPD.The GSDMD-N expression of alveolar epithelial cells increased in the BPD rat model established by intra-amniotic injection of LPS,while NSA treatment significantly improved the lung development of BPD rats and inhibited the IL-1βmRNA expression(both P<0.05).In vitro,the study confirmed that LPS/ATP treatment decreased the viability of alveolar epithelial cells MLE-12 and induced pyroptosis,while NSA treatment increased alveolar epithelial cell viability and inhibited pyroptosis(both P<0.05).In addition,NSA treatment upregulated the SFTPC expression and inhibited the GSDMD-N expression in LPS/ATP-stimulated alveolar epithelial cells(both P<0.05).Conclusion·Inhibiting the alveolar epithelial cell pyroptosis can improve the alveolar development in BPD newborn rats.
作者
郑小雁
王星云
张拥军
ZHENG Xiaoyan;WANG Xingyun;ZHANG Yongjun(Department of Neonatology,Xinhua Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200092,China;Hongqiao International Institute of Medicine,Tongren Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200336,China)
出处
《上海交通大学学报(医学版)》
CAS
CSCD
北大核心
2023年第2期171-179,共9页
Journal of Shanghai Jiao tong University:Medical Science