中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2015年
5期
1062-1065
,共4页
吴世乐%宋世铎%张逸%张立峰%张子祥%朱东明%周健%李德春
吳世樂%宋世鐸%張逸%張立峰%張子祥%硃東明%週健%李德春
오세악%송세탁%장일%장립봉%장자상%주동명%주건%리덕춘
诱骗受体3%胰腺癌%T淋巴细胞%RNA干扰
誘騙受體3%胰腺癌%T淋巴細胞%RNA榦擾
유편수체3%이선암%T림파세포%RNA간우
Decoy receptor 3%Pancreatic cancer%T lymphocyte%RNA interference
目的 观察胰腺癌Panc-1细胞中诱骗受体3(DcR3)基因沉默对T淋巴细胞功能的影响.方法 构建针对DcR3基因的小干扰RNA真核表达载体,脂质体LipofectamineTM 2000介导DcR3干扰质粒转染人胰腺癌Panc-1细胞,转染48 h后采用反转录-聚合酶链反应(RT-PCR)和酶联免疫吸附试验(ELISA)检测DcR3的表达变化;与淋巴细胞混合培养72 h后采用流式细胞术检测T细胞亚群的变化及Th1、Th2细胞的偏移,ELISA法检测干扰素(IFN)-γ和IL-4水平.结果 与空载体转染组比较,转染靶向DcR3的小干扰RNA(DcR3-siRNA) 48 h后的Panc-1细胞中DcR3 mRNA和蛋白表达下降(P＜0.05);沉默DcR3的胰腺癌Panc-1细胞与淋巴细胞混合培养后CD3+ CD4+T细胞增多(P＜0.05),而CD8+T细胞无明显变化(P＞0.05);Th1细胞增多[(7.87±1.07)％比(5.57±0.70)％,P＜0.05],Th2细胞细胞减少[(0.91±0.15)％比(2.83±0.21)％,P＜0.05];细胞上清液中Th1型细胞因子的IFN-γ浓度升高[(20.88±1.32) μg/L比(15.61±1.26) μg/L,P＜0.01],而Th2型细胞因子的IL-4水平下降[(3.27±0.34) μg/L比(5.31±0.37) μg/L,P＜0.01].结论 在胰腺癌细胞中靶向沉默DcR3的表达,可以有效恢复T淋巴细胞的免疫调节功能,纠正失衡的Th1/Th2细胞因子网络.
目的 觀察胰腺癌Panc-1細胞中誘騙受體3(DcR3)基因沉默對T淋巴細胞功能的影響.方法 構建針對DcR3基因的小榦擾RNA真覈錶達載體,脂質體LipofectamineTM 2000介導DcR3榦擾質粒轉染人胰腺癌Panc-1細胞,轉染48 h後採用反轉錄-聚閤酶鏈反應(RT-PCR)和酶聯免疫吸附試驗(ELISA)檢測DcR3的錶達變化;與淋巴細胞混閤培養72 h後採用流式細胞術檢測T細胞亞群的變化及Th1、Th2細胞的偏移,ELISA法檢測榦擾素(IFN)-γ和IL-4水平.結果 與空載體轉染組比較,轉染靶嚮DcR3的小榦擾RNA(DcR3-siRNA) 48 h後的Panc-1細胞中DcR3 mRNA和蛋白錶達下降(P＜0.05);沉默DcR3的胰腺癌Panc-1細胞與淋巴細胞混閤培養後CD3+ CD4+T細胞增多(P＜0.05),而CD8+T細胞無明顯變化(P＞0.05);Th1細胞增多[(7.87±1.07)％比(5.57±0.70)％,P＜0.05],Th2細胞細胞減少[(0.91±0.15)％比(2.83±0.21)％,P＜0.05];細胞上清液中Th1型細胞因子的IFN-γ濃度升高[(20.88±1.32) μg/L比(15.61±1.26) μg/L,P＜0.01],而Th2型細胞因子的IL-4水平下降[(3.27±0.34) μg/L比(5.31±0.37) μg/L,P＜0.01].結論 在胰腺癌細胞中靶嚮沉默DcR3的錶達,可以有效恢複T淋巴細胞的免疫調節功能,糾正失衡的Th1/Th2細胞因子網絡.
목적 관찰이선암Panc-1세포중유편수체3(DcR3)기인침묵대T림파세포공능적영향.방법 구건침대DcR3기인적소간우RNA진핵표체재체,지질체LipofectamineTM 2000개도DcR3간우질립전염인이선암Panc-1세포,전염48 h후채용반전록-취합매련반응(RT-PCR)화매련면역흡부시험(ELISA)검측DcR3적표체변화;여림파세포혼합배양72 h후채용류식세포술검측T세포아군적변화급Th1、Th2세포적편이,ELISA법검측간우소(IFN)-γ화IL-4수평.결과 여공재체전염조비교,전염파향DcR3적소간우RNA(DcR3-siRNA) 48 h후적Panc-1세포중DcR3 mRNA화단백표체하강(P＜0.05);침묵DcR3적이선암Panc-1세포여림파세포혼합배양후CD3+ CD4+T세포증다(P＜0.05),이CD8+T세포무명현변화(P＞0.05);Th1세포증다[(7.87±1.07)％비(5.57±0.70)％,P＜0.05],Th2세포세포감소[(0.91±0.15)％비(2.83±0.21)％,P＜0.05];세포상청액중Th1형세포인자적IFN-γ농도승고[(20.88±1.32) μg/L비(15.61±1.26) μg/L,P＜0.01],이Th2형세포인자적IL-4수평하강[(3.27±0.34) μg/L비(5.31±0.37) μg/L,P＜0.01].결론 재이선암세포중파향침묵DcR3적표체,가이유효회복T림파세포적면역조절공능,규정실형적Th1/Th2세포인자망락.
Objective To investigate the effect of T lymphocyte function by silencing decoy receptor 3 (DcR3) in human Pancreatic cancer cell line Panc-1.Methods Small interfering RNA (siRNA)eukaryotic expression vector targeting DcR3 gene was transfected into Panc-1 cells by Lipofectamine 2000.The expression of DcR3 was detected by using reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme linked immunosorbent assay (ELISA) after transfection 48 h.After mixed lymphocyte tumor cell cultured for 72 h,lymphocyte subsets were examined by flow cytometry analysis and the level of interferon (IFN)-γ,interleukin (IL)-4 was determined by ELISA.Results In the DcR3-siRNA transfection group,the expression of DcR3 was decreased in Panc-1 cells remarkably.After Panc-1 cells silencing DcR3 in mixed lymphocyte tumor cell cultured,the number of CD3 +,CD4 + T lymphocyte was increased (P ＜ 0.05) and the number of CD8 + T lymphocyte was unchanged statistically (P ＞ 0.05).Additionally,the number of Th1 cells was increased [(7.87 ± 1.07) ％ vs.(5.57 ± 0.70) ％,P ＜ 0.05],while the number of Th2 cells was decreased [(0.91 ± 0.15) ％ vs.(2.83 ± 0.21) ％,P ＜ 0.05] as compared with negative control group (NC-siRNA).Conclusion The silencing DcR3 may reverse immune regulating of T lymphocyte and Correct the imbalance of Th1/Th2.
