Alu 基因定量检测用于胃癌转移模型评估的研究
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作者单位:

1.空军军医大学实验动物中心,西安 710032; 2. 空军军医大学基础医学院,西安 710032


Measurement of Alu gene expression by RT-PCR in the evaluation of models of gastric cancer metastasis
Author:
Affiliation:

1.Laboratory Animal Center, Air Force Medical University, Xi’an 710032, China. 2. Basic Medical College, Air Force Medical University, Xi’an 710032

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    摘要:

    目的 研究人胃癌转移裸鼠模型中人 Alu 基因的表达水平与组织脏器中胃癌转移程度的相关性, 探索建立胃癌转移模型早期评估的分子生物学方法。 方法 分别以人胃癌细胞 SGC-7901、MKN45 和正常胃黏膜细胞 GES1 基因组 DNA 为模板构建标准质粒,通过实时荧光定量 PCR 检测不同比例胃癌细胞 SGC-7901 中 Alu 基因的表达,获取相关性曲线;选取胃癌细胞 SGC-7901 和 MKN45,通过裸鼠皮下接种的方式构建人胃癌细胞异种移植转移模型,实时荧光定量 PCR方法检测模型鼠肝、脾、肺、肾和皮下肿瘤组织中的人 Alu 基因表达,获得 Alu 基因的表达与各器官组织中肿瘤转移程度的相关性曲线;将临床胃癌患者新鲜肿瘤标本皮下接种裸鼠构建异种移植模型,进一步验证模型鼠各组织中人 Alu 基因的表达与器官中肿瘤转移程度的相关性。 结果 胃癌细胞 SGC-7901 的含量与其 Alu 基因的 Ct 值呈负相关(R2 = 0. 9239);人胃癌细胞异种移植(CDX)裸鼠转移模型中,人 Alu 基因的表达在皮下肿瘤中处于较高水平,在肺转移瘤和肝转移瘤中的表达则介于正常裸鼠与皮下肿瘤之间,与组织病理学检查结果相符;胃癌病人肿瘤异种移植(PDX)模型中,已确定发生转移的脏器中虽未形成肉眼可见的转移灶,但其人 Alu 基因的表达(Ct 值 17. 86)与正常裸鼠(Ct 值 22. 18)差异显著(P< 0. 05);而对于肉眼可见的转移灶,其人 Alu 基因的表达(Ct 值 14. 29)则差异极显著(P< 0. 01)。 结论 人胃癌裸鼠转移模型中,人 Alu 基因的表达与胃癌转移程度呈正相关,Alu 基因表达越高,则该组织中转移的肿瘤细胞就越多,形成的转移灶越明显。

    Abstract:

    Objective We aimed to characterize the relationship between the expression of the Alu gene in a model of human gastric cancer metastasis and the degree of gastric cancer metastasis in tissues and organs and establish a molecular biological method for the early evaluation of gastric cancer metastasis. Methods Alu gene expression plasmids were constructed from the genomic DNA of human gastric cancer cell lines SGC- 7901, MKN45 and the normal gastric mucosal cell line GES1. Alu gene expression in various concentration of gastric cancer cells SGC-7901, which was widely mentioned as human gastric metastasis cell line, was measured by real-time PCR. The relationship between different Alu expression of various concentration was measured by cycle threshold (Ct) value to modelling the standard curve. A human gastric cancer cell xenograft metastasis model was established by the subcutaneous inoculation of SGC-7901 and MKN45 cells into nude mice. The expression of Alu in the resulting tumors in the mice (in the liver, spleen, lung, kidney, and subcutaneous tissues) was then measured by real-time PCR, and a curve for the relationship between the expression of Alu and the degree of tumor metastasis in each tissue was constructed. Subsequently, fresh tumor samples from patients with gastric cancer were subcutaneously inoculated into nude mice to construct a xenograft model, and the relationships between Alu expression in the tissues of the mice and the degree of tumor metastasis in the organs of nude mice was similarly evaluated. Results The number of SGC-7901 cells negatively correlated with the cycle threshold (Ct) value for the Alu gene (R2 = 0. 9239). In the xenograft metastasis model, the expression of the Alu gene was higher in subcutaneous tumors than that in lung and liver metastases, and the expression of the Alu gene in lung and liver metastases was higer than normal nude mice. This was consistent with the result of histopathological examination. In the gastric cancer patient xenograft model, if there was no visible metastasis in the organs of nude mice, the expression of Alu gene (Ct 17. 86) was significantly higher(P< 0. 05)than that of normal nude mice (Ct 22. 18), and if there were metastatic lesions that were visible through the naked eye, the expression of Alu (Ct 14. 29) was very significantly higher than that of normal nude mice (P< 0. 01). Conclusions In a model of human gastric cancer metastasis, the expression of the Alu gene positively correlates with the degree of metastasis, namely the higher expression of Alu, the more metastatic tumor cells were present and the more obvious the metastatic foci were in each tissue.

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吴朋朋,柳森森,张彩勤,赵勇,白冰,王洁,师长宏. Alu 基因定量检测用于胃癌转移模型评估的研究[J].中国实验动物学报,2021,29(4):440~447.

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  • 收稿日期:2020-12-23
  • 在线发布日期: 2021-09-10
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