Yang L, Zhou Z-G, Zheng X-L, Wang L, Yu Y-Y, Zhou B, Gu J, Li Y
Institute of Digestive Surgery and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, 1 Keyuan Road 4, Gaopengdadao, Chengdu, 610041, Sichuan Province, China.
Dis Colon Rectum. 2008 Mar;51(3):318-26; discussion 326-8. doi: 10.1007/s10350-007-9145-8. Epub 2008 Jan 24.
This study was designed to investigate the effects of peroxisome proliferator-activated receptor delta (PPAR delta) on the proliferation and apoptosis of human colorectal cancer cells.
For RNA interfering (RNAi), HCT-116 cells were transfected with short hairpin RNA (shRNA)-expressing plasmids against PPAR delta or negative control vectors, and the stably transfected cells were selected with G418. The efficacy of RNAi was assessed by real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blotting analysis. The proliferation, cell cycle, and apoptosis of HCT-116 cells treated by RNAi, compared with those containing control vectors or untreated, were analyzed respectively by using MTT (methyl thiazolyl tetrazolium), flow cytometry, and TdT (terminal deoxynucleotidyl transferase)-mediated dUTP nick end-labeling (TUNEL) assay.
RNAi targeting PPAR delta resulted in substantial suppression of PPAR delta expression and significantly promoted the proliferation of HCT-116 cells relative to those with control vectors or untreated, obviously decreasing the frequency of G1-phase cells but had no effect on cell apoptosis.
PPAR delta may inhibit the proliferation of CRC cells and increase the number of cells in G1 phase, without any function in cell apoptosis.
本研究旨在探讨过氧化物酶体增殖物激活受体δ(PPARδ)对人结肠癌细胞增殖和凋亡的影响。
对于RNA干扰(RNAi),将针对PPARδ的短发夹RNA(shRNA)表达质粒或阴性对照载体转染至HCT-116细胞,并用G418筛选稳定转染的细胞。通过实时逆转录聚合酶链反应(RT-PCR)和蛋白质免疫印迹分析评估RNAi的效果。分别使用MTT(噻唑蓝)、流式细胞术和TdT(末端脱氧核苷酸转移酶)介导的dUTP缺口末端标记(TUNEL)法分析经RNAi处理的HCT-116细胞与含有对照载体的细胞或未处理细胞相比的增殖、细胞周期和凋亡情况。
靶向PPARδ的RNAi导致PPARδ表达大幅抑制,相对于含有对照载体的细胞或未处理细胞,显著促进了HCT-116细胞的增殖,明显降低了G1期细胞的频率,但对细胞凋亡无影响。
PPARδ可能抑制结肠癌细胞的增殖并增加G1期细胞数量,而对细胞凋亡无作用。