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目的 :观察过表达肠凝集素1(intelectin 1,ITLN-1)对子人宫内膜腺癌Ishikawa细胞凋亡和增殖的影响。方法:采用脂质体法将重组质粒pCMV6-ITLN-1转入Ishikawa细胞(ITLN-1过表达组),同时设未做任何处理的细胞作为空白对照组和转入空载体pCMV6-N1的空载体组。48 h后,应用实时荧光定量PCR法检测ITLN-1、p53、环氧合酶2(cyclooxygenase-2,Cox-2)和第10号染色体缺失的磷酸酶及张力蛋白同源基因(phosphate and tension homology deleted on chromsome ten,PTEN)mRNA的表达水平;蛋白质印迹法检测ITLN-1、p53、Cox-2、Bcl-2及cleaved-caspase 3蛋白的表达水平。采用MTS法检测细胞的增殖活性;FCM法检测细胞的凋亡率,倒置相差显微镜下观察凋亡细胞形态的变化。结果:ITLN-1、p53和PTEN mRNA的表达水平均较空白对照组和空载体组明显上调(P<0.01、P<0.05和P<0.01),而Cox-2 mRNA的表达水平则明显下调(P<0.01)。p53和cleaved-caspase 3蛋白的表达水平均明显上调(P值均<0.01),Bcl-2和Cox-2蛋白的表达水平均明显下调(P值均<0.01)。与2个对照组相比,24和48 h时ITLN-1过表达组Ishikawa细胞的生长受到明显抑制(P<0.05和P<0.01),细胞凋亡率明显提高(P<0.01),倒置相差显微镜下细胞形态呈凋亡改变。结论 :ITLN-1表达上调能抑制Ishikawa细胞的增殖,并诱导其凋亡,这可能与上调Ishikawa细胞中凋亡相关蛋白的表达有关。
Objective: To observe the effect of overexpression of intelectin 1 (ITLN-1) on apoptosis and proliferation of endometrial adenocarcinoma Ishikawa cells. Methods: The recombinant plasmid pCMV6-ITLN-1 was transfected into Ishikawa cells (ITLN-1 overexpression group) by liposome method. Meanwhile, untreated cells were used as blank control group and empty vector pCMV6-N1 Vector group. After 48 h, the expression of ITLN-1, p53, cyclooxygenase-2 (Cox-2) and phosphatase and tensin homology genes deleted on chromosome 10 were detected by real-time fluorescence quantitative PCR homology deleted on chromsome ten (PTEN) mRNA expression level. Western blotting was used to detect the expression of ITLN-1, p53, Cox-2, Bcl-2 and cleaved-caspase 3 protein. MTS assay was used to detect the proliferation activity of the cells. FCM was used to detect the apoptosis rate of the cells. The morphological changes of apoptotic cells were observed under inverted phase contrast microscope. Results: The expressions of ITLN-1, p53 and PTEN mRNA were significantly up-regulated compared with the blank control group and empty vector group (P <0.01, P <0.05 and P <0.01) P <0.01). The expressions of p53 and cleaved-caspase 3 were significantly up-regulated (P <0.01), while the expressions of Bcl-2 and Cox-2 were significantly down-regulated (all P <0.01). Compared with the two control groups, the growth of Ishikawa cells in ITLN-1 over-expression group was significantly inhibited at 24 and 48 h (P <0.05 and P <0.01), and the apoptosis rate was significantly increased (P <0.01) The morphology of the cells under the microscope showed apoptosis. CONCLUSION: Upregulation of ITLN-1 can inhibit Ishikawa cell proliferation and induce its apoptosis, which may be related to the up-regulation of the expression of apoptosis-related proteins in Ishikawa cells.