兰州大学机构库 >生命科学学院
CARD9在乳腺癌组织中的表达及其对乳腺癌细胞生物学行为的影响
Alternative TitleThe expression of CARD9 in breast cancer tissues and its functions on the biological behavior of breast cancer cells
孙振
Thesis Advisor陈学忠
2017-03-25
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name硕士
KeywordCARD9:乳腺癌:细胞增殖:NF-κB:信号通路
Abstract

乳腺癌是女性常见的恶性肿瘤,发病率呈上升趋势,且趋向于年轻化。虽然,近年来,乳腺癌的早期诊断、手术方式、放射治疗以及靶向治疗方面已取得很大进展,但预后仍然较差,靶向药物效率低且价格昂贵。研究乳腺癌发生和发展的分子机制,探索新的分子靶标仍然是乳腺癌研究和临床中需要解决的难题之一。

胱天蛋白酶募集域蛋白9(caspase recruitment domain containing protein 9,CARD9)是一个重要的衔接蛋白,通过蛋白间的相互作用调节NF-κB等信号通路,在胃B细胞淋巴瘤、肾癌、丙型肝炎病毒相关的肝细胞癌及结肠癌的发生和发展中扮演着重要角色,但其在乳腺癌中的表达及对乳腺癌发生发展的影响尚未见相关报道。

本实验利用实时荧光定量PCR和Western-Blotting技术分别检测乳腺癌癌组织、癌旁组织、乳腺癌细胞、正常乳腺细胞中CARD9 mRNA和蛋白的表达情况,分析CARD9表达与乳腺癌患者临床病理特征间的关系。通过WST-1法、平板克隆形成实验、流式细胞术、划痕实验研究CARD9高表达对乳腺癌细胞增殖、凋亡、周期、侵袭转移等生物学行为的影响。分析CARD9表达和NF-κB信号通路激活的相关性,初步探讨CARD9在乳腺癌中的作用机制。

结果表明:乳腺癌组织中CARD9 mRNA(p<0.01)和蛋白(p=0.012)表达水平显著高于癌旁组织;乳腺癌T-47D细胞(p=0.021)和MCF-7细胞(p=0.014)中CARD9 mRNA表达水平高于正常乳腺细胞Hs 578Bst;乳腺癌ZR-75-1细胞中CARD9 mRNA(p=0.003)和蛋白(p=0.015)表达水平均高于正常乳腺细胞Hs 578Bst。CARD9蛋白表达与肿瘤大小和雌激素受体(ER)表达有关(p值均<0.05)。CARD9高表达可促进乳腺癌细胞的增殖(p<0.05),但对细胞凋亡、周期、侵袭和转移未见显著性影响。乳腺癌组织中P50(p<0.05)和p-IKKα(p<0.01)蛋白的表达水平高于癌旁组织,而且乳腺癌组织中CARD9 mRNA表达与P50(r=0.622, p<0.001)、P65(r=0.392, p=0.005)、IL-1β(r=0.685, p<0.001)和IL-12(r=0.556, p<0.001)mRNA表达间呈正相关。本实验的研究结果揭示了CARD9在乳腺癌组织中的表达情况及其与乳腺癌临床病理学的关系。同时发现,CRAD9很可能通过激活NF-κB信号通路促进乳腺癌细胞增殖从而参与乳腺癌发生发展进程。这些研究结果有助于深入了解CRAD9在乳腺癌发生发展及治疗中的潜在价值,为CARD9成为乳腺癌新的治疗靶点提供了理论依据。

Other Abstract

Breast cancer is one of the most common malignant tumor in women, and tends to be more likely to recur or spread in younger women. Although, the progresses have been made in early diagnosis, surgical treatment, radiotherapy and targeted therapy for the treatment of breast cancer recently years, but the curative efficacy is less satisfactory. The efficiency of target therapy on breast cancer is also required to be improved. Therefore, the detection of the molecular mechanism on the pathologenesis of breast cancer and the new therapeutic targets are urgently things that need to be settled in breast cancer curing.

CARD9 protein is an important adaptor protein, which binds to Bcl10, activates and regulates NF-κB and other signal pathway through the interaction between proteins, plays an important role in the occurrence and development of gastric B cell lymphoma, renal cell carcinoma, hepatitis C virus related hepatocellular carcinoma and colon cancer. However, its expression in breast cancer and its influence on the development of breast cancer has been rarely reported.

The expressions of CARD9 mRNA and protein in breast cancer cell line T-47D and MCF-7, normal breast cell line Hs578Bst, breast cancer tissues and the adjacent normal tissues were detected by real-time fluorescent quantitative PCR and Western blotting, respectively. The correlation between CARD9 expression and clinicopathological features was analyzed. WST-1 assay, plate clone formation assay, flow cytometry and scarification assay were conducted to test the proliferation, cell apoptosis, cell cycle, invasion and metastasis of breast cancer cells respectively after enhancing the expression level of CARD9 via transient transfection.Finally,we analysed the correlation between CARD9 expression and activation of NF-κB signaling pathway, and explored the possible mechanismconcerned.The results showed that the expressions of CARD9 mRNA (P < 0.01) and protein (P = 0.012) in breast cancer tissues were higher than those in the adjacent normal tissues. The expression of CARD9 mRNA in breast cancer T-47D cells (P = 0.021) and MCF-7 cells (P = 0.014) was higher than that in normal breast Hs578Bst cells. The expressions of CARD9 mRNA and protein in breast cancer ZR-75-1 cells were higher than those in normal breast Hs578Bst cells (P = 0.003 and P = 0.01). Expression of CARD9 protein was positively correlated with the tumor size and ER (both P < 0.015). Enhancing the expression of CARD9 could promote the proliferation of breast cancer cells (P < 0.05), while had no effect on cell apoptosis, cell cycle, Invasion and metastasis of breast cancer cells. The expressions of P50 (P < 0.05), P65 (P = 0.098), p-IKKα (P < 0.01) in breast cancer tissues were higher than those in the adjacent normal tissues. The mRNAs expressions of p50 (r = 0.622, P < 0.01), p65 (r = 0.392, P = 0.005), IL-1β (r = 0.685, P < 0.01), IL-12 (r = 0.556,P < 0.01) were positive correlated with the expression of CARD9 mRNA.The results of this study revealed the expression of CARD9 in breast cancer and its relationship with the clinicopathological features of breast cancer .Meanwhile, it was found that CRAD9 could promote the proliferation of breast cancer cells by activating the NF-κB signaling pathway.In this way CARD9 participate in the development of breast cancer. These findings contribute to a better understanding of the potential value of CRAD9 in the development and treatment of breast cancer, and make CARD9 a new key risk indicator for breast cancer. It can provide a theoretical evidence for CARD9 to become a new therapeutic target for breast cancer and provide new ideas for the diagnosis and treatment of breast cancer.

URL查看原文
Language中文
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/222045
Collection生命科学学院
Recommended Citation
GB/T 7714
孙振. CARD9在乳腺癌组织中的表达及其对乳腺癌细胞生物学行为的影响[D]. 兰州. 兰州大学,2017.
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