Claudin-7作为细胞基底蛋白在肺癌侵袭转移过程中的生物学功能及机制研究

Lung cancer is the leading cause of cancer-related death in China. It is most likely to become fatal when cancerous cells metastasize, or spread, to other parts of the body. However,the underlying molecular mechanisms of lung cancer invasion and metastasis are still largely unknown. Tight junctions are the most apical component of the junctional complex and provide one form of cell-cell adhesion in epithelial or endothelial cells. Tight junctions serve as selective channels, regulating ions and small molecules passing through the paracellular pathway and protecting the organism from the external environment. Disruption of the tight junction characteristics could potentially change the normal equilibrium maintained, resulting in abnormal cellular physiology including cancer..Claudins are a family of tight junctions with at least 24 members in mammals. We have previously reported that claudin-7 is closely associated with lung cancer. We also found that claudin-7 is a unique tight junction protein in that it is not only localized at the apical tight junctions of lung cancer cells, but also has a strong expression at the basolateral membrane. Based on our previous results, we will investigate the biological roles and regulatory mechanisms of claudin-7 as a basal protein in lung cancer invasion and metastasis both in vitro and in vivo. We will overexpress claudin-7 in NCI-H522 lung adenocarcinoma cell line and knockdown claudin-7 in HCC827 lung adenocarcinoma cell line to investigate whether alterations of claudin-7 expression affect the cell-matrix adhesions and cell invasion ability examined by cell attachment assay and cell invasion assay, respectively. We will then use molecular and cellular technologies to examine the localization of integrin subunits, expression levels of integrin subunits at both mRNA and protein levels,and integrin regulated downstream signaling pathways, such as ERK/MAPK and PI3K/Akt pathways. In addition, we will induce lung tumorigenesis by vinyl carbamate in claudin-7 wild type (Cldn7+/+) and heterozygous (Cldn7+/-) mice to investigate the role of claudin-7 in lung cancer formation and regulating integrin subunits and ERK/MAPK and PI3K/Akt signaling pathways in vivo. Our study will provide a molecular basis for the novel non-tight junction function of claudin-7 in regulating cell-matrix adhesions and cell invasion ability in lung cancer.

肺癌的侵袭、转移机制是肺癌研究领域中的热点和难点。紧密连接位于上皮细胞和内皮细胞细胞膜顶端,是维持两个邻近细胞紧密相连的结构，其结构功能的改变可以引起肿瘤细胞的快速增殖和转移。Claudin家族蛋白是紧密连接的重要组成蛋白之一。我们前期研究发现，claudin-7蛋白不仅在肺癌细胞顶端紧密连接处有表达，而且在基底端有着更高水平的表达。Claudin-7作为一个基底蛋白在肺癌中的作用，国内外至今未有报导。本项目是以H522与HCC827肺癌细胞株和claudin-7基因杂合小鼠为研究对象，运用细胞分子生物学方法研究claudin-7在肺癌细胞与细胞外基质连接及肺癌细胞侵袭转移过程中发挥的生物学功能及其可能的通过调节整合素的表达及整合素下游信号通路而产生作用的机制。本项目将探究紧密连接蛋白对整合素调节的一个新领域，为claudin-7作为非紧密连接蛋白在肺癌侵袭转移过程中的作用提供理论依据。

肺癌是中国恶性肿瘤第一位死亡原因，肺癌细胞的侵袭转移是肺癌致死的主要原因，70%的肺癌患者死于肿瘤转移。因此肺癌的侵袭、转移机制是肺癌研究领域中的热点和难点，深入研究肺癌侵袭转移的分子生物学基础具有重要的应用价值。紧密连接位于上皮细胞和内皮细胞细胞膜顶端，是维持两个邻近细胞紧密连接的结构，其结构功能的改变可以引起肿瘤细胞的快速增殖和转移。Claudin家族蛋白是紧密连接的重要组成蛋白之一。我们前期研究发现，claudin-7蛋白不仅在肺癌细胞顶端紧密连接处有表达，而且在基底端有更高水平的表达。本项目探究了claudin-7作为一个基底蛋白在肺癌中的作用。历时三年研究，本项目明确了claudin-7可以通过调节integrin β1的表达调节细胞与细胞外基质之间的连接，具体地，本项目在细胞、动物、临床三个水平上：1. 明确了claudin-7 在肺癌细胞侵袭转移及小鼠肺肿瘤生成转移过程中发挥的生物学功能；2. 明确了claudin-7对整合素表达水平、表达位置的影响，对整合素下游细胞信号通路ERK/MAPK的调节作用，为肺癌细胞的侵袭转移分子机制提供理论依据；3. 对临床肺癌标本进行研究，明确了claudin-7表达破碎度与肺癌之间的关系。本项目为claudin-7作为非紧密连接蛋白在肺癌侵袭过程中的作用提供了理论依据，为挖掘更多的分子靶标提供理论基础，同时对临床上寻找有效的药物治疗或干预措施降低肺癌的死亡率，具有重要的参考和应用价值。