长链非编码RNA HULC对胶质母细胞瘤原位移植瘤模型肿瘤生长的促进作用

Long Non-Coding RNA HULC Enhances Tumor Growth in Orthotopic Xenograft Model of Glioblastoma

[目的]初步探讨LncRNAHULC对胶质母细胞瘤U87细胞株体内体外生长的作用机制。[方法]实时荧光定量PCR(qRT-PCR)检测胶质母细胞瘤U87细胞HULC过表达组(H ULC-over)及其对照组(V EC)和沉默表达组(H ULC-siRNA)及其对照组(N C)中HULC表达水平。CCK8增殖实验和克隆形成实验检测U87细胞增殖能力。流式细胞学技术检测U87细胞凋亡能力。小鼠异种原位移植模型按照注射U87细胞对应分为HULC-over组(n=10)和VEC组(n=10)及HULC-siRNA组(n=10)和NC组(n=10),观察各组小鼠生存期。免疫组化检测Ki67的表达。[结果]HULC-over组较VEC组HULC表达量显著升高,HULC-siRNA组较NC组HULC表达量显著降低(均P<0.01)。CCK8增殖实验显示HULC-over组较VEC组细胞增殖率显著升高;HULC-siRNA组较NC组细胞增殖率显著降低(第2、3、4天,均P<0.01)。细胞克隆实验显示VEC组和HULC-over组克隆形成率分别为(34.47±1.56)%和(95.4±2.74)%;NC组和HULC-siRNA组克隆形成率分别为(23.83±0.92)%和(10.23±0.61)%,过表达HULC克隆形成率升高,沉默表达HULC克隆形成率降低(均P<0.01)。细胞凋亡实验显示VEC组和HULC-over组早期凋亡率分别为(3.55±0.56)%和(0.09±0.01)%;NC组和HULC-siRNA组早期凋亡率分别为(2.89±0.67)%和(7.13±0.14)%,过表达HULC细胞早期凋亡率降低,沉默表达HULC细胞早期凋亡率升高(P<0.01)。小鼠生存期数据表明HULC-over组比VEC组存活时间更短,而HULC-siRNA组比NC组存活时间更长(P<0.05)。与VEC组相比,Ki67蛋白在HULC-over组中表达上调;与NC组相比,Ki67蛋白在HULC-siRNA组中表达下调(均P<0.05)。[结论]LncRNAHULC对胶质母细胞瘤U87细胞体内体外生长具有促进作用。

【Objective】To investigate the mechanism of action of long non-coding RNA highly up-regulated in liver cancer(LncRNA HULC)on the growth of glioblastoma U87 cells in vitro and in vivo.【Methods】The cultured glioblastoma U87 cells were divided into four groups:overexpression group(HULC-over)and its vector control group(VEC),silent HULCexpression group(HULC-siRNA)and its negative control group(NC).Quantitative real-time polymerase chain reaction PCR(qRT-PCR)was used to verify the expression levels of HULC.CCK8 proliferation assay and colony formation assay were adopted to monitor the proliferation of glioblastoma U87 cells.Flow cytometry was utilized to detect the apoptosis of glioblastoma U87 cells.By injecting U87 cells,we divided the orthotopic xenograft mouse modelinto HULC-over group(n=10),VEC group(n=10),HULC-siRNA group(n=10)and NC group(n=10)accordingly.The survival of the mice in each group was observed.The expression of Ki67 was analyzed by immunohistochemistry.【Results】The expression level of HULC was significantly higher in HULC-over group than that in VEC group and significantly lower in HULC-siRNA group than that in NC group(P<0.01).The cell proliferation ability was significantly increased in HULC-over group compared with that in VEC group and significantly decreased in HULC-siRNA group compared with that in NC group(P<0.01 on days2,3and4).The colony formation rates in VEC group,HULC-over group,NC group and HULC-siRNA group were,respectively,(34.47±1.56)%,(95.4±2.74)%,(23.83±0.92)%and(10.23±0.61)%,which revealed that overexpression of HULC elevated the colony formation rate and silencing expression of HULC reduced the colony forma⁃tion rate(P<0.01).The early apoptosis rates in VEC group,HULC-over group,NC group and HULC-siRNA group were,respectively,(3.55±0.56)%,(0.09±0.01)%,(2.89±0.67)%,and(7.13±0.14)%,which showed that overex⁃pression of HULC elevated the early apoptosis rate and silencing expression of HULC reduced the early apoptosis rate(P<0.01).The survival curve of nude mouse indicated shorter survival time in HULC-over group than that in VEC group and longer survival time in HULC-siRNA group than that in NC group(P<0.05).Ki67 protein expression was up-regulated in the HULC-over group compared with that in VEC group and down-regulated in the HULC-siRNA group compared with that in NC group(P<0.05).【Conclusion】LncRNA HULC can enhance the growth of glioblastoma U87 cells in vitro and in vivo.