天津医科大学学报
天津醫科大學學報
천진의과대학학보
JOURNAL OF TIANJIN MEDICAL UNIVERSITY
2015年
3期
212-216
,共5页
唐培源%戚斌%陈渊%王元国%张鹏
唐培源%慼斌%陳淵%王元國%張鵬
당배원%척빈%진연%왕원국%장붕
胸腺瘤%Wnt4%JNK%干扰%shRNA
胸腺瘤%Wnt4%JNK%榦擾%shRNA
흉선류%Wnt4%JNK%간우%shRNA
Thymoma%Wnt4%JNK%interference%shRNA
目的：研究shRNA干扰Wnt4基因对胸腺瘤细胞JNK基因表达的影响及Wnt4基因发挥作用可能依赖的信号通路。方法：根据shRNA设计原则，设计针对Wnt4基因的4个干扰靶点，构建Wnt4-shRNA干扰质粒，经鉴定片段成功插入后，将空白质粒、TR001质粒和重组的4个干扰质粒分别转染体外培养的胸腺瘤细胞。采用RT-PCR检测转染后Wnt4基因表达，挑选对Wnt4基因抑制效果最好的干扰质粒进行后续实验。将胸腺瘤细胞分3组：空白对照组；转染TR001质粒组；转染干扰质粒组。用LipofectamineTM2000转染，采用RT-PCR及Western blot检测细胞Wnt4和JNK的mRNA及蛋白表达。结果：shRNA-Wnt4-3质粒能显著抑制Wnt4基因表达，抑制率为52.37%（P＜0.05）。 Wnt4基因下调后胸腺瘤细胞内JNK基因表达也显著下调（P＜0.01），差异有统计学意义。结论：采用shRNA干扰Wnt4基因后胸腺瘤细胞内JNK基因表达显著下调，说明Wnt4基因与JNK基因的表达有相关性， Wnt4基因可能依赖于非经典的Wnt/JNK信号通路发挥作用。
目的：研究shRNA榦擾Wnt4基因對胸腺瘤細胞JNK基因錶達的影響及Wnt4基因髮揮作用可能依賴的信號通路。方法：根據shRNA設計原則，設計針對Wnt4基因的4箇榦擾靶點，構建Wnt4-shRNA榦擾質粒，經鑒定片段成功插入後，將空白質粒、TR001質粒和重組的4箇榦擾質粒分彆轉染體外培養的胸腺瘤細胞。採用RT-PCR檢測轉染後Wnt4基因錶達，挑選對Wnt4基因抑製效果最好的榦擾質粒進行後續實驗。將胸腺瘤細胞分3組：空白對照組；轉染TR001質粒組；轉染榦擾質粒組。用LipofectamineTM2000轉染，採用RT-PCR及Western blot檢測細胞Wnt4和JNK的mRNA及蛋白錶達。結果：shRNA-Wnt4-3質粒能顯著抑製Wnt4基因錶達，抑製率為52.37%（P＜0.05）。 Wnt4基因下調後胸腺瘤細胞內JNK基因錶達也顯著下調（P＜0.01），差異有統計學意義。結論：採用shRNA榦擾Wnt4基因後胸腺瘤細胞內JNK基因錶達顯著下調，說明Wnt4基因與JNK基因的錶達有相關性， Wnt4基因可能依賴于非經典的Wnt/JNK信號通路髮揮作用。
목적：연구shRNA간우Wnt4기인대흉선류세포JNK기인표체적영향급Wnt4기인발휘작용가능의뢰적신호통로。방법：근거shRNA설계원칙，설계침대Wnt4기인적4개간우파점，구건Wnt4-shRNA간우질립，경감정편단성공삽입후，장공백질립、TR001질립화중조적4개간우질립분별전염체외배양적흉선류세포。채용RT-PCR검측전염후Wnt4기인표체，도선대Wnt4기인억제효과최호적간우질립진행후속실험。장흉선류세포분3조：공백대조조；전염TR001질립조；전염간우질립조。용LipofectamineTM2000전염，채용RT-PCR급Western blot검측세포Wnt4화JNK적mRNA급단백표체。결과：shRNA-Wnt4-3질립능현저억제Wnt4기인표체，억제솔위52.37%（P＜0.05）。 Wnt4기인하조후흉선류세포내JNK기인표체야현저하조（P＜0.01），차이유통계학의의。결론：채용shRNA간우Wnt4기인후흉선류세포내JNK기인표체현저하조，설명Wnt4기인여JNK기인적표체유상관성， Wnt4기인가능의뢰우비경전적Wnt/JNK신호통로발휘작용。
O bjective: To explore the effect of Wnt4 gene on the expression of JNK gene and the signal pathway in thymoma cells. Methods:Four Wnt4-shRNA interference plasmids were constructed to target directly on Wnt4 gene. The blank plasmid, TR001 and recombinant plasmids were transfected into thymoma cells. The expression of Wnt4 was detected by RT-PCR while the best Wnt4 interference plasmid for subsequent experiments was selected. The thymoma cells were divided into three groups:control group, transfected TR001 plasmid, transfected interference plasmid. The expression of Wnt4 and JNK were detected by RT-PCR and Western blot. Results:Expression of Wnt4 could be significantly inhibited by shRNA-Wnt4-3 plasmid. The ratio of inhibition was 52.37%(P<0.05).With down regulation of Wnt4, the expression of JNK were significantly down-regulated in thymoma cells (P<0.01). Conclusion:After interfering Wnt4 gene by using shRNA , the expression of JNK gene is significantly reduced. This indicates that Wnt4 gene is correlated with the expression of JNK gene in thymoma cells.Which may depend on the non-canonical Wnt/JNK signaling pathway.
