中华普通外科杂志
中華普通外科雜誌
중화보통외과잡지
CHINESE JOURNAL OF GENERAL SURGERY
2009年
7期
568-572
,共5页
胰腺肿瘤%乳酸脱氢酶类%糖酵解%RNA干扰
胰腺腫瘤%乳痠脫氫酶類%糖酵解%RNA榦擾
이선종류%유산탈경매류%당효해%RNA간우
Pancreatic neoplasms%Lactate dehydrogenases%Glycolysis%RNA interference
目的 通过构建乳酸脱氢酶A(lactate dehydrogenase,LDH-A)shRNA并转染PANC-1细胞,分析其对胰腺癌细胞生物学特性的影响.方法 构建3条LDH-A shRNA质粒,脂质体转染质粒至PANC-1细胞中,实时荧光定量PCR法检测不同质粒转染后LDH-A mRNA的表达变化.将抑制效率最高的shRNA质粒-3转染PANC-1细胞,MTT法检测转染前后细胞增殖的变化,流式细胞仪检测细胞凋亡的变化.RT-PCR检测LDH-A表达以及酶化学染色检测转染前后LDH活性的变化.结果 3种LDH-A shRNA质粒转染胰腺癌细胞后,LDH-A shRNA质粒.3的2-△△Ct值为(0.47±0.02),较正常细胞(0.71±0.01)小,存在抑制作用,且抑制效率较高.PANC-1细胞在转染shRNA质粒-3后12 h转染组吸光度值显著低于对照组,细胞出现增殖抑制,24、36、48和72 h,转染组的吸光度值均显著低于对照组(P<0.01).转染质粒组细胞凋亡也明显增高,其凋亡率达到61.74%;转染shRNA质粒-3的胰腺癌细胞株,其LDH-A mRNA的表达明显抑制,酶化学染色显示LDH活性明显减弱.结论 LDH-A shRNA通过抑制胰腺癌细胞LDH-A mRNA的表达抑制其增殖及诱导细胞凋亡.
目的 通過構建乳痠脫氫酶A(lactate dehydrogenase,LDH-A)shRNA併轉染PANC-1細胞,分析其對胰腺癌細胞生物學特性的影響.方法 構建3條LDH-A shRNA質粒,脂質體轉染質粒至PANC-1細胞中,實時熒光定量PCR法檢測不同質粒轉染後LDH-A mRNA的錶達變化.將抑製效率最高的shRNA質粒-3轉染PANC-1細胞,MTT法檢測轉染前後細胞增殖的變化,流式細胞儀檢測細胞凋亡的變化.RT-PCR檢測LDH-A錶達以及酶化學染色檢測轉染前後LDH活性的變化.結果 3種LDH-A shRNA質粒轉染胰腺癌細胞後,LDH-A shRNA質粒.3的2-△△Ct值為(0.47±0.02),較正常細胞(0.71±0.01)小,存在抑製作用,且抑製效率較高.PANC-1細胞在轉染shRNA質粒-3後12 h轉染組吸光度值顯著低于對照組,細胞齣現增殖抑製,24、36、48和72 h,轉染組的吸光度值均顯著低于對照組(P<0.01).轉染質粒組細胞凋亡也明顯增高,其凋亡率達到61.74%;轉染shRNA質粒-3的胰腺癌細胞株,其LDH-A mRNA的錶達明顯抑製,酶化學染色顯示LDH活性明顯減弱.結論 LDH-A shRNA通過抑製胰腺癌細胞LDH-A mRNA的錶達抑製其增殖及誘導細胞凋亡.
목적 통과구건유산탈경매A(lactate dehydrogenase,LDH-A)shRNA병전염PANC-1세포,분석기대이선암세포생물학특성적영향.방법 구건3조LDH-A shRNA질립,지질체전염질립지PANC-1세포중,실시형광정량PCR법검측불동질립전염후LDH-A mRNA적표체변화.장억제효솔최고적shRNA질립-3전염PANC-1세포,MTT법검측전염전후세포증식적변화,류식세포의검측세포조망적변화.RT-PCR검측LDH-A표체이급매화학염색검측전염전후LDH활성적변화.결과 3충LDH-A shRNA질립전염이선암세포후,LDH-A shRNA질립.3적2-△△Ct치위(0.47±0.02),교정상세포(0.71±0.01)소,존재억제작용,차억제효솔교고.PANC-1세포재전염shRNA질립-3후12 h전염조흡광도치현저저우대조조,세포출현증식억제,24、36、48화72 h,전염조적흡광도치균현저저우대조조(P<0.01).전염질립조세포조망야명현증고,기조망솔체도61.74%;전염shRNA질립-3적이선암세포주,기LDH-A mRNA적표체명현억제,매화학염색현시LDH활성명현감약.결론 LDH-A shRNA통과억제이선암세포LDH-A mRNA적표체억제기증식급유도세포조망.
Objective To construct a short hairpin RNA (shRNA) targeting LDH-A, and evaluate the effects on growth, apoptosis and the expression of LDH-A in PANC-1 cells. Method Three shRNAs targeting LDH-A were combined to pGCsilencer vector, and transfected into PANC-1 cells. The expression of LDH-A after transfected by the three shRNAs in pancreatic cancer cell lines PANC-1 was detected by quantitative real time PCR. After tranfected by LDH shRNA-3 with the highest inhibited rate, cell growth was analyzed by MTT assay, apoptosis was detected by flow cytometry. The LDH-A expression was detected by reverse transcription polymerase chain reaction, LDH activity was observed with enzyme cytochemical method. Results The 2-AACt of LDH-A shRNA-3 was (0. 47 ± 0. 02), less than the untransfected pancreatic cancer cell(0. 71 ± 0. 01), the LDH-A shRNA-3 could inhibit the expression of the LDH-A most effectively. The growth of the pancreatic cancer cell was inhibited after 12 h transfected by LDH-A shRNA-3, all the absorbance value of transfected cell in 24 h,36 h,48 h,72 h decreased obviously compared to the untreated pancreatic cancer cell(P <0. 01). The apoptosis rate of the transfected cell increased to 61.74%. The inhibition of LDH-A expression in PANC-1 cells transfected by shRNA-3 was significantly and the activity of LDH reduced. Conclusion LDH-A shRNA inhibits the expression of LDH-A, the proliferation of cancer cells inducing the apoptosis of PANC-1 cells.
