CAJ | 학술논문

目的研究A549细胞呼吸道合胞病毒（ RSV）感染12 h、24 h、48 h后调节激活正常T细胞表达和分泌细胞因子（RANTES）、分形趋化因子（FKN）、干扰素诱导蛋白-10（IP-10） mRNA和蛋白水平的变化及不同剂量过氧化物酶体增殖物激活受体γ（PPARγ）激动剂15-脱氧前列腺素J2（15d-PGJ2）和罗格列酮及其拮抗剂（ GW9662）的干预作用。方法建立RSV感染A549细胞的体外模型，将传代培养的细胞随机分成5组：A组（15d-PGJ2＋RSV组），B组（罗格列酮＋RSV组）、C组（DMSO＋RSV组）、D组（ GW9662＋罗格列酮＋RSV组）、E组（细胞对照组）。各组分别在培养12 h、24 h、48 h收获细胞及上清液待测。应用ELISA检测各组上清液RANTES、FKN、IP-10蛋白水平，实时荧光定量RT-PCR检测各组RANTES、FKN、IP-10 mRNA表达。结果与细胞对照组相比， RSV感染组RAN-TES、FKN、IP-10 mRNA和蛋白的表达在12 h、24 h和48 h均明显升高（P均＜0．05），其中RANTES、FKN、IP-10 mRNA的表达量在24 h达高峰，48 h有所下降，与12 h的表达量比较差异无统计学意义（P均＞0．05）；而3种趋化因子蛋白的表达量均在48 h达高峰，与12 h、24 h比较差异有统计学意义（P均＜0．05）。在同一作用时间点上，随着15d-PGJ2和罗格列酮药物浓度的增加，RANTES、FKN、IP-10 mRNA和蛋白的表达量均呈剂量依赖性下降，与 RSV 感染组相比差异有统计学意义（ P 均＜0．05），其中以20μmol/L 15d-PGJ2和30μmol/L罗格列酮干预后RANTES、FKN和IP-10 mRNA和蛋白的表达量最低。结论 RSV感染可导致RANTES、FKN和IP-10 mRNA及蛋白表达升高，其中mRNA水平在感染后24 h达到高峰，蛋白的表达自感染后48 h达到高峰；而PPARγ激动剂能以剂量依赖性方式抑制上述趋化因子mRNA和蛋白的表达，从而起到减轻炎症的作用。
목적연구A549세포호흡도합포병독（ RSV）감염12 h、24 h、48 h후조절격활정상T세포표체화분비세포인자（RANTES）、분형추화인자（FKN）、간우소유도단백-10（IP-10） mRNA화단백수평적변화급불동제량과양화물매체증식물격활수체γ（PPARγ）격동제15-탈양전렬선소J2（15d-PGJ2）화라격렬동급기길항제（ GW9662）적간예작용。방법건립RSV감염A549세포적체외모형，장전대배양적세포수궤분성5조：A조（15d-PGJ2＋RSV조），B조（라격렬동＋RSV조）、C조（DMSO＋RSV조）、D조（ GW9662＋라격렬동＋RSV조）、E조（세포대조조）。각조분별재배양12 h、24 h、48 h수획세포급상청액대측。응용ELISA검측각조상청액RANTES、FKN、IP-10단백수평，실시형광정량RT-PCR검측각조RANTES、FKN、IP-10 mRNA표체。결과여세포대조조상비， RSV감염조RAN-TES、FKN、IP-10 mRNA화단백적표체재12 h、24 h화48 h균명현승고（P균＜0．05），기중RANTES、FKN、IP-10 mRNA적표체량재24 h체고봉，48 h유소하강，여12 h적표체량비교차이무통계학의의（P균＞0．05）；이3충추화인자단백적표체량균재48 h체고봉，여12 h、24 h비교차이유통계학의의（P균＜0．05）。재동일작용시간점상，수착15d-PGJ2화라격렬동약물농도적증가，RANTES、FKN、IP-10 mRNA화단백적표체량균정제량의뢰성하강，여 RSV 감염조상비차이유통계학의의（ P 균＜0．05），기중이20μmol/L 15d-PGJ2화30μmol/L라격렬동간예후RANTES、FKN화IP-10 mRNA화단백적표체량최저。결론 RSV감염가도치RANTES、FKN화IP-10 mRNA급단백표체승고，기중mRNA수평재감염후24 h체도고봉，단백적표체자감염후48 h체도고봉；이PPARγ격동제능이제량의뢰성방식억제상술추화인자mRNA화단백적표체，종이기도감경염증적작용。
Objective To observe the expressions of RANTES , FKN and IP-10 at mRNA and pro-tein levels in human lung epithelial cells (A549) infected by respiratory syncytial virus (RSV), and to eval-uate the changes of them interfered with 15-deoxy-delta12,14prostaglandin J2(15d-PGJ2), rosiglitazone or 2-chloro-5-nitrobenzanilide ( GW9662 ) .Methods A549 cells were seeded in 6-well culture plates and cul-tured overnight in F12K culture solution.Then they were randomly divided into five groups , including group A (15d-PGJ2+RSV group), group B (rosiglitazone+RSV group), group C (DMSO+RSV group), group D (GW9662+rosiglitazone+RSV group) and group E (cell control group).Cells and supernatants were harves-ted from each group at different time points (12 h, 24 h and 48 h) of culture.The expressions of RANTES , FKN and IP-10 at mRNA and protein levels were measured by ELISA and real-time quantitative RT-PCR analysis.Results The expressions of RANTES , FKN and IP-10 at mRNA and protein levels in group C were significantly higher than those in group E at time points of 12 h, 24 h and 48 h (all P<0.05).In group C, the expressions of the three chemokines at mRNA level reached a peak at 24 h, but began to de-crease at 48 h, which showed no statistical significance compared with those at 12 h (all P>0.05).Moreo-ver, the expressions at protein level were peaked at 48h, and had significant difference with those expressed at 12 h and 24 h (all P<0.05).Compared with group C, the expressions of the three chemokines both at mRNA level and protein level were decreased in group A and B as the dose was increased (all P<0.05), and the lowest levels were observed with the intervention of 20 μmol/L of 15d-PGJ2 in group A and 30μmol/L of rosiglitazone in group B .Conclusion The expressions of RANTES , FKN and IP-10 at mRNA and protein levels were increased with RSV infection , and the peaks of mRNA level and protein level were respectively achieved at 24 h and 48 h after infection.PPARγagonists played an anti-inflammatory role through inhibiting the expressions of the three chemokines both at mRNA level and protein level in a dose -de-pendent manner .