CAJ | 학술논문

目的探讨氯化镧对宫颈癌HeLa细胞增殖和迁移能力的影响,为寻找新的有效治疗宫颈癌的药物提供实验依据.方法宫颈癌细胞株HeLa细胞经培养、传代后分为两组,即实验组(加入5、50、100 μmol/L的氯化镧)和对照组(未加入氯化镧).倒置显微镜下观察两组细胞的生长情况,激光共聚焦显微镜观察两组细胞核的形态变化.采用四甲基偶氮唑蓝(MTF)比色法检测两组细胞的增殖情况,双染色流式细胞仪检测两组细胞的凋亡率,体外迁移实验检测两组细胞迁移能力的变化,逆转录(RT)-PCR技术检测两组细胞中增殖、抗凋亡和迁移相关基因细胞周期蛋白(cyclinD1)、锌指蛋白(A20)及基质金属蛋白酶9(MMP-9)mRNA的表达.结果倒置显微镜下观察:实验组细胞随着氯化镧浓度的升高,细胞密度逐渐降低,细胞质内颗粒逐渐增多,颜色加深,细胞间隙增大,少量细胞变圆漂浮,脱落细胞也逐渐增多;而对照组细胞贴壁生长密集,形态清晰,胞质饱满,相邻细胞生长融合成片.激光共聚焦显微镜观察:实验组细胞核染色质浓聚边集,核体积变小,随着氯化镧浓度的升高,核染色质崩解,核膜破裂、核碎裂,直至细胞核完全碎裂;而对照组细胞核饱满,核膜完整.实验组细胞经不同浓度(5、50、100 μmol/L)的氯化镧作用后,细胞生长抑制率分别为24%、51%、78%,高于对照组(0),差异有统计学意义(P<0.05);细胞凋亡率分别为(4.91±0.39)%、(7.30±0.71)%、(13.48±0.92)%,高于对照组的(0.89±0.11)%,差异有统计学意义(P<0.01);穿膜细胞数分别为(22.2±4.3)、(12.0±3.2)、(7.8±2.6)个,低于对照组的(41.2±5.4)个,差异有统计学意义(P<0.01).实验组细胞中cyclinD1、A20及MMP-9 mRNA的表达强度随氯化镧浓度(5、50、100 μmol/L)的升高逐渐减弱.结论氯化镧通过下调宫颈癌细胞中增殖、抗凋亡和迁移相关基因cyclinD1、A20及MMP-9 mRNA的表达,抑制宫颈癌细胞的增殖和迁移并诱导其凋亡. 目的探討氯化鑭對宮頸癌HeLa細胞增殖和遷移能力的影響,為尋找新的有效治療宮頸癌的藥物提供實驗依據.方法宮頸癌細胞株HeLa細胞經培養、傳代後分為兩組,即實驗組(加入5、50、100 μmol/L的氯化鑭)和對照組(未加入氯化鑭).倒置顯微鏡下觀察兩組細胞的生長情況,激光共聚焦顯微鏡觀察兩組細胞覈的形態變化.採用四甲基偶氮唑藍(MTF)比色法檢測兩組細胞的增殖情況,雙染色流式細胞儀檢測兩組細胞的凋亡率,體外遷移實驗檢測兩組細胞遷移能力的變化,逆轉錄(RT)-PCR技術檢測兩組細胞中增殖、抗凋亡和遷移相關基因細胞週期蛋白(cyclinD1)、鋅指蛋白(A20)及基質金屬蛋白酶9(MMP-9)mRNA的錶達.結果倒置顯微鏡下觀察:實驗組細胞隨著氯化鑭濃度的升高,細胞密度逐漸降低,細胞質內顆粒逐漸增多,顏色加深,細胞間隙增大,少量細胞變圓漂浮,脫落細胞也逐漸增多;而對照組細胞貼壁生長密集,形態清晰,胞質飽滿,相鄰細胞生長融閤成片.激光共聚焦顯微鏡觀察:實驗組細胞覈染色質濃聚邊集,覈體積變小,隨著氯化鑭濃度的升高,覈染色質崩解,覈膜破裂、覈碎裂,直至細胞覈完全碎裂;而對照組細胞覈飽滿,覈膜完整.實驗組細胞經不同濃度(5、50、100 μmol/L)的氯化鑭作用後,細胞生長抑製率分彆為24%、51%、78%,高于對照組(0),差異有統計學意義(P<0.05);細胞凋亡率分彆為(4.91±0.39)%、(7.30±0.71)%、(13.48±0.92)%,高于對照組的(0.89±0.11)%,差異有統計學意義(P<0.01);穿膜細胞數分彆為(22.2±4.3)、(12.0±3.2)、(7.8±2.6)箇,低于對照組的(41.2±5.4)箇,差異有統計學意義(P<0.01).實驗組細胞中cyclinD1、A20及MMP-9 mRNA的錶達彊度隨氯化鑭濃度(5、50、100 μmol/L)的升高逐漸減弱.結論氯化鑭通過下調宮頸癌細胞中增殖、抗凋亡和遷移相關基因cyclinD1、A20及MMP-9 mRNA的錶達,抑製宮頸癌細胞的增殖和遷移併誘導其凋亡.
목적 탐토록화란대궁경암HeLa세포증식화천이능력적영향,위심조신적유효치료궁경암적약물제공실험의거.방법 궁경암세포주HeLa세포경배양、전대후분위량조,즉실험조(가입5、50、100 μmol/L적록화란)화대조조(미가입록화란).도치현미경하관찰량조세포적생장정황,격광공취초현미경관찰량조세포핵적형태변화.채용사갑기우담서람(MTF)비색법검측량조세포적증식정황,쌍염색류식세포의검측량조세포적조망솔,체외천이실험검측량조세포천이능력적변화,역전록(RT)-PCR기술검측량조세포중증식、항조망화천이상관기인세포주기단백(cyclinD1)、자지단백(A20)급기질금속단백매9(MMP-9)mRNA적표체.결과 도치현미경하관찰:실험조세포수착록화란농도적승고,세포밀도축점강저,세포질내과립축점증다,안색가심,세포간극증대,소량세포변원표부,탈락세포야축점증다;이대조조세포첩벽생장밀집,형태청석,포질포만,상린세포생장융합성편.격광공취초현미경관찰:실험조세포핵염색질농취변집,핵체적변소,수착록화란농도적승고,핵염색질붕해,핵막파렬、핵쇄렬,직지세포핵완전쇄렬;이대조조세포핵포만,핵막완정.실험조세포경불동농도(5、50、100 μmol/L)적록화란작용후,세포생장억제솔분별위24%、51%、78%,고우대조조(0),차이유통계학의의(P<0.05);세포조망솔분별위(4.91±0.39)%、(7.30±0.71)%、(13.48±0.92)%,고우대조조적(0.89±0.11)%,차이유통계학의의(P<0.01);천막세포수분별위(22.2±4.3)、(12.0±3.2)、(7.8±2.6)개,저우대조조적(41.2±5.4)개,차이유통계학의의(P<0.01).실험조세포중cyclinD1、A20급MMP-9 mRNA적표체강도수록화란농도(5、50、100 μmol/L)적승고축점감약.결론 록화란통과하조궁경암세포중증식、항조망화천이상관기인cyclinD1、A20급MMP-9 mRNA적표체,억제궁경암세포적증식화천이병유도기조망.
Objective To investigate the effects of lanthanum chloride on proliferation and migration activity of human cervical cancer cells in vitro which may be a new anti-cervical cancer drug and provide experimental data for cervical cancer treatment. Methods HeLa cells cultured in vitro were divided into two groups: experimental group and control group. In experimental group, the cells were respectively treated with lanthanum chloride at different concentrations, 5, 50 and 100 μmol/L, while the cells in the control group were not treated with lanthanum chloride. The cell growth was observed by inverted microscope and the morphology changes of the cells were observed by the laser scanning confocal microscope (LSCM).Proliferation of HeLa cells in the two groups was detected by methyl thiazolyl tetrazolium (MTT) test;apoptosis rate was analyzed by flow cytometry (FCM). Cell migration test was applied to observe the effect of lanthanum chloride on migration. Reverse transcription (RT)-PCR was employed to evaluate the effects of lanthanum chloride on proliferation gene (cyclinD1), anti-apoptosis gene (zinc finger protein A20) and migration-related gene (matrix metalloproteinase 9, MMP-9). Results The status of cell growth was observed under the inverted microscope: with the increased of the lanthanum chloride concentrations, the cell density of reduced, the granule in cytoplasm increased, color intensifying and intercellular space enlarged; some cells became rounding and dead, floating in the culture media; the exfoliated cells increased gradually in the experimental groups. While In the control group, the cells grew adherently, with clear morphology and plump cytoplasm, and adjacent cell grew in lamellar. Observed with LSCM: the nuclear chromatin condensated and marginated with the volume of nuclear decreased in experimental groups. With the increase of the lanthanum chloride concentrations, nuclei in the experimental groups became pyknotic and then underwent karyorrhexis. However, the nuclear of the cells in control group were inact. The growth inhibition rates of lanthanum chloride groups (5, 50, 100 μmol/L) were 24%, 51% and 78%,respectively, in which each was significantly higher than that of the control group (P < 0. 05); the apoptosis rates of lanthanum chloride group were (4. 91 + 0. 39) %, (7. 30 + 0. 71) % and (13.48 + 0. 92) %,respectively, which were all significantly higher than that of the control group [(0. 89 + 0. 11) %, P <0.01]. The migration ability of the cells was also decreased by the treatment of lanthanum chloride, the number of migrated cells in lanthanum chloride groups were 22.2±4. 3, 12. 0±3.2 and 7. 8±2. 6 respectively, which were all significantly lower than that of the control group (41.2±5.4, P < 0. 01). The expression of genes of cyclinD1, A20 and MMP-9, were all decreased by the treatment of lanthanum chloride in a dose-dependent manner. Conclusion Lanthanum chloride can inhibit the proliferation and migration of cervical cancer cells, and induce apoptosis by down-regulating cyclinD1, A20, and MMP-9 expressions in vitro.