论文部分内容阅读
目的研究长链非编码RNA(long non-coding RNA,Lnc RNA)中的同源基因转录反义基因间RNA(HOTAIR)在人胃癌KATO-Ⅲ细胞中的表达及其作用。方法采用免疫磁珠分选技术分选出人胃癌KATO-Ⅲ细胞中的CD133阳性和CD133阴性细胞后,再采用逆转录聚合酶链式反应(reverse transcription polymerase chain reaction,RT-PCR)法检测其中HOTAIR m RNA和CD133 m RNA的表达水平;采用小干扰RNA技术(small interfering RNA,si RNA)干扰CD133阳性细胞中HOTAIR的表达后,再采用RT-PCR法检测细胞中HOTAIR m RNA的表达水平,以筛选出干扰效果最好的si RNA用于后续实验;以筛选出的si RNA干扰序列干扰CD133阳性细胞中HOTAIR的表达后,检测细胞中CD133 m RNA、E-钙黏素(E-cadherin)m RNA及N-钙黏素(N-cadherin)m RNA的表达,并开展Transwell实验以检测细胞迁移和侵袭能力的改变。结果 1 RT-PCR检测结果表明,CD133阳性组细胞中CD133 m RNA和HOTAIR m RNA的表达水平均高于CD133阴性组和未分选组(P<0.05)。2 si HOTAIR干扰细胞中HOTAIR的表达后,si HOTAIR1组、si HOTAIR2组及si HOTAIR3组细胞中HOTAIR m RNA的表达水平均低于空白对照组和阴性对照组(P<0.05),且si HOAIR2组细胞中HOTAIR m RNA的表达水平低于si HOTAIR1组及si HOTAIR3组(P<0.05),表明si HOTAIR2的干扰效果最好。3 si HOTAIR2组细胞中CD133 m RNA和N-cadherin m RNA的表达水平均低于空白对照组和阴性对照组(P<0.05),但E-cadherin m RNA的表达水平却高于空白对照组和阴性对照组(P<0.05);细胞迁移实验和侵袭实验结果均表明,si HOTAIR2组的穿膜细胞数均低于空白对照组和阴性对照组(P<0.05)。结论 HOTAIR m RNA在CD133阳性人胃癌KATO-Ⅲ细胞中的表达水平高于CD133阴性细胞,干扰HOTAIR m RNA的表达可下调CD133阳性人胃癌KATO-Ⅲ细胞中CD133 m RNA的表达,并抑制胃癌细胞的迁移及侵袭。
Objective To study the expression of HOTAIR in human gastric cancer KATO-Ⅲ cells and its role in long non-coding RNA (Lnc RNA). Methods The CD133-positive and CD133-negative cells in human gastric cancer KATO-Ⅲ cells were sorted by immunomagnetic bead sorting technique and then detected by reverse transcription polymerase chain reaction (RT-PCR) HOTAIR m RNA and CD133 m RNA were detected by RT-PCR. The expression of HOTAIR mRNA in CD133 positive cells was detected by small interfering RNA (siRNA) The most effective RNAi was screened out for further experiments. After screening the siRNA against HO13IR in CD133 positive cells, the expression of CD133mRNA, E-cadherin, m RNA and N-cadherin m RNA expression, and carried out Transwell experiments to detect cell migration and invasion ability changes. Results 1 RT-PCR results showed that the expression levels of CD133 m RNA and HOTAIR m RNA in CD133 positive group were higher than those in CD133 negative group and non-sorted group (P <0.05). The expression of HOTAIR m RNA in si HOTAIR1 group, si HOTAIR2 group and si HOTAIR3 group was lower than that in the blank control group and the negative control group (P <0.05) after si HOTAIR interference with HOTAIR. HOTAIR m RNA expression was lower in cells than in si HOTAIR1 and si HOTAIR3 groups (P <0.05), indicating that si HOTAIR2 had the best interference. The expression levels of CD133 m RNA and N-cadherin m RNA in 3 HO HOIR2 group were lower than those in blank control group and negative control group (P <0.05), but the expression of E-cadherin m RNA was higher than that in blank control group and Negative control group (P <0.05). The results of cell migration assay and invasion assay showed that the number of transmembrane cells in si HOTAIR2 group was lower than that in blank control group and negative control group (P <0.05). Conclusion The expression of HOTAIR m RNA in CD133 positive human gastric cancer KATO-Ⅲ cells is higher than that in CD133 negative cells. Interfering HOTAIR m RNA expression can down-regulate the expression of CD133 m RNA in CD133 positive human gastric cancer KATO-Ⅲ cells and inhibit the expression of CD133 m RNA in gastric cancer cells Migration and invasion.