中华皮肤科杂志
中華皮膚科雜誌
중화피부과잡지
Chinese Journal of Dermatology
2010年
4期
269-272
,共4页
林福全%关翠萍%刘东银%洪为松%周妙妮%许爱娥
林福全%關翠萍%劉東銀%洪為鬆%週妙妮%許愛娥
림복전%관취평%류동은%홍위송%주묘니%허애아
InnVit基因%白癜风%细胞增殖%细胞凋亡%细胞周期%细胞,培养的
InnVit基因%白癜風%細胞增殖%細胞凋亡%細胞週期%細胞,培養的
InnVit기인%백전풍%세포증식%세포조망%세포주기%세포,배양적
InnVit gene%Vitiligo%Cell proliferation%Apoptosis%Cell cycle%Cells,cultured
目的 分析InnVit基因的抑制和过表达对永生化黑素细胞株B10BR细胞增殖、周期及凋亡的影响,探讨InnVit基因的生物学功能及其对白癜风中黑素细胞缺失的影响.方法 化学合成InnVit基因特异性siRNA片段以及构建过表达质粒载体P3XF-P120,lipofectamine~(TM) 2000脂质体方法转染BIOBR细胞.半定量RT-PCR法检测InnVit基因mRNA水平;Western印迹检测该蛋白水平变化;MTT法测定细胞增殖情况;流式细胞技术分析细胞周期和凋亡的变化.结果 InnVit基因抑制后mRNA和蛋白表达水平明显低于对照组,InnVit基因过表达质粒组mRNA和蛋白表达水平显著高于对照组;转染后24、48、72 h抑制组的细胞存活量极显著下降(P＜0.01),过表达质粒组的细胞存活量在48和72 h极显著增加(P＜0.01).转染后48 h基因抑制组的早期凋亡率从(10.24±1.00)%升至(37.34±3.26)%,过表达质粒组早期凋亡率从(14.58±1.49)%降至(8.43±0.86)%,与对照组比较,差异均有统计学意义(P＜0.01);转染48 h基因抑制组的G_1期细胞由(56.11±5.46)%增至(69.76.±6.08)%(P＜0.05),过表达质粒组的G_1期细胞由(55.14±5.65)%降至(29.33±3.01)%(P＜0.01).结论 InnVit基因抑制使细胞增殖能力下降,促进细胞凋亡;InnVit基因的过表达增强了细胞的增殖能力,抑制细胞凋亡.其增殖能力的改变可能是通过细胞周期的调控调节的.
目的 分析InnVit基因的抑製和過錶達對永生化黑素細胞株B10BR細胞增殖、週期及凋亡的影響,探討InnVit基因的生物學功能及其對白癜風中黑素細胞缺失的影響.方法 化學閤成InnVit基因特異性siRNA片段以及構建過錶達質粒載體P3XF-P120,lipofectamine~(TM) 2000脂質體方法轉染BIOBR細胞.半定量RT-PCR法檢測InnVit基因mRNA水平;Western印跡檢測該蛋白水平變化;MTT法測定細胞增殖情況;流式細胞技術分析細胞週期和凋亡的變化.結果 InnVit基因抑製後mRNA和蛋白錶達水平明顯低于對照組,InnVit基因過錶達質粒組mRNA和蛋白錶達水平顯著高于對照組;轉染後24、48、72 h抑製組的細胞存活量極顯著下降(P＜0.01),過錶達質粒組的細胞存活量在48和72 h極顯著增加(P＜0.01).轉染後48 h基因抑製組的早期凋亡率從(10.24±1.00)%升至(37.34±3.26)%,過錶達質粒組早期凋亡率從(14.58±1.49)%降至(8.43±0.86)%,與對照組比較,差異均有統計學意義(P＜0.01);轉染48 h基因抑製組的G_1期細胞由(56.11±5.46)%增至(69.76.±6.08)%(P＜0.05),過錶達質粒組的G_1期細胞由(55.14±5.65)%降至(29.33±3.01)%(P＜0.01).結論 InnVit基因抑製使細胞增殖能力下降,促進細胞凋亡;InnVit基因的過錶達增彊瞭細胞的增殖能力,抑製細胞凋亡.其增殖能力的改變可能是通過細胞週期的調控調節的.
목적 분석InnVit기인적억제화과표체대영생화흑소세포주B10BR세포증식、주기급조망적영향,탐토InnVit기인적생물학공능급기대백전풍중흑소세포결실적영향.방법 화학합성InnVit기인특이성siRNA편단이급구건과표체질립재체P3XF-P120,lipofectamine~(TM) 2000지질체방법전염BIOBR세포.반정량RT-PCR법검측InnVit기인mRNA수평;Western인적검측해단백수평변화;MTT법측정세포증식정황;류식세포기술분석세포주기화조망적변화.결과 InnVit기인억제후mRNA화단백표체수평명현저우대조조,InnVit기인과표체질립조mRNA화단백표체수평현저고우대조조;전염후24、48、72 h억제조적세포존활량겁현저하강(P＜0.01),과표체질립조적세포존활량재48화72 h겁현저증가(P＜0.01).전염후48 h기인억제조적조기조망솔종(10.24±1.00)%승지(37.34±3.26)%,과표체질립조조기조망솔종(14.58±1.49)%강지(8.43±0.86)%,여대조조비교,차이균유통계학의의(P＜0.01);전염48 h기인억제조적G_1기세포유(56.11±5.46)%증지(69.76.±6.08)%(P＜0.05),과표체질립조적G_1기세포유(55.14±5.65)%강지(29.33±3.01)%(P＜0.01).결론 InnVit기인억제사세포증식능력하강,촉진세포조망;InnVit기인적과표체증강료세포적증식능력,억제세포조망.기증식능력적개변가능시통과세포주기적조공조절적.
Objective To assess the effect of InnVit gene expression on cell proliferation, apoptosis and cell cycle of B10BR cells, and to explore the biological function of InnVit gene and its influence on the loss of melanocytes in vitiligo.Methods InnVit gene-targeting siRNA was synthesized, and overexpression plasmid P3XF-P120 was constructed: B10BR cells were classified into 4 groups to be transfected with control siRNA(negative control), siRNA targeting InnVit gene(siRNA group), empty plasmid(plasmid control group), and P3XF-P120 plasmid(P3XF-P120 group), respectively.After additional culture for various periods,the mRNA and protein levels of InnVit gene were detected by RT-PCR and Western blot, respectively, cell surviva1 by MTT assay, cell cycle and apoptosis by flow cytometry.Results The expression of lnnVit mRNA and protein significantly increased in P3XF-P120 group compared with the plasmid control group, but decreased in siRNA group compared with the negative control group(all P ＜ 0.01).After the transfection, the survival rate of cells was significantly lower in siRNA group than in negative control group at 24, 48 and 72 hours, but higher in P3XF-P120 group than in plasmid control group at 48 and 72 hours(all P＜ 0.01).At 48 hours after the transfection, the early apoptosis rate and proportion of cells in G_1 phase increased from(10.24 ± 1.00)% and(56.11 ± 5.46)% in negative control group to(37.34 ± 3.26)% and(69.76 ± 6.08)%, in siRNA group,respectively(P＜ 0.01 or 0.05), from(14.58 ± 1.49)% and(55.14 ± 5.65)% in plasmid control group to (8.43 ± 0.86)% and(29.33 ± 3.01)%, in P3XF-P120 group, respectively(both P ＜ 0.01).Conclusions The suppression of InnVit gene efficiently inhibits the proliferation and induces apoptosis of B10BR cells, and vice versa.Therefore, the regulation of cell cycle may affect the proliferation of B10BR cells.
