β1整合素与TUBA1A的相互作用在非霍奇金淋巴瘤CAM-DR中的作用与意义

Cell adhesion mediated drug resistance (CAM-DR) is an important reason for incurable of non-hodgkin's lymphoma (NHL). The expression of integrin β1 in cell membrane and its internalization circulation are closely related with the process of CAM-DR. Our research group has found that TUBA1A may interact with integrin β1 and enhance the expression of integrin β1 in cell membrane. The research also found that the interaction between integrin β1 and TUBA1A can increase the stability of TUBA1A. According to this, we hypothesized that the interaction between integrin β1and TUBA1A was postulated to sustaining the high level of integrin β1 in cell membrane and meanwhile ensuring the stable expression of TUBA1A. And this mechanism may facilitate the formation of CAM-DR. To verify this hypothesis, firstly we intend to investigate the mechanism of interaction between TUBA1A and β1 and identify the role of TUBA1A in regulating β1 circulation. Secondly, the signifiance of this interaction during the formation cell adhesion mediated drug resistance would be detected. Finally, we would prove this hypothesis using clinical tissue and nude mice model. Our research outcome will provide a theoretical basis for clarifying the molecular mechanism of CAM-DR and may provide new targets for lymphoma drug design.

细胞粘附介导的耐药（CAM-DR）是导致非霍奇金淋巴瘤不可治愈的重要原因。整合素β1的胞膜表达及在胞质中的内化循环与CAM-DR形成密切相关。本课题通过质谱分析发现TUBA1A是整合素β1的一个新的相互作用蛋白，并能促进β1的胞膜表达。同时发现整合素β1亦能促进TUBA1A稳定性增加。据此推测，在细胞粘附过程中，整合素β1与TUBA1A的相互作用将形成一个促进β1在胞膜持续表达的正循环，并使TUBA1A在胞膜稳定表达，有利于细胞CAM-DR形成。本课题拟首先在淋巴瘤细胞悬浮及粘附状态分析整合素β1促进TUBA1A稳定性的具体分子机制；其次分析TUBA1A调节β1内化循环的相关机制及对细胞CAM-DR的影响；并利用临床组织标本及裸鼠模型验证上述实验结果。课题的研究结果将从整合素与微管蛋白相互作用的角度阐明淋巴瘤细胞CAM-DR形成的分子机制，研究结果将为克服临床上淋巴瘤化疗耐药提供理论参考。

本课题研究探讨整合素β1与微管蛋白alpha 1A（tubulin alpha 1A，TUBA1A）的相互作用在非霍奇金淋巴瘤细胞粘附介导的耐药（Cell adhesion mediated drug resistance，CAM-DR）中的机制及意义。.首先，通过构建人淋巴瘤细胞OCI-Ly8及Raji与人骨髓基质细胞HS-5或纤连蛋白粘附模型，蛋白免疫印迹实验显示微管蛋白TUBA1A及整合素β1蛋白在淋巴瘤细胞粘附培养时表达较悬浮培养增加。在淋巴瘤细胞中利用干扰质粒沉默整合素β1的表达，显示淋巴瘤细胞与基质细胞发生粘附的数量下降；利用多柔比星诱导淋巴瘤细胞凋亡，整合素β1的表达下降可以增加粘附状态下淋巴瘤细胞对化疗药物的敏感性，并使其细胞活力降低。接下来，在工具细胞及淋巴瘤细胞中均发现整合素β1和TUBA1A的相互作用，并且粘附培养的淋巴瘤细胞可以促进两者相互作用。通过构建整合素β1截短突变体，明确整其通过胞膜内段结构域与TUBA1A发生相互作用。进一步的研究显示，整合素β1与TUBA1A相互作用可以抑制淋巴瘤细胞粘附能力，流式细胞术实验结果显示细胞凋亡数量增加，淋巴瘤细胞对化疗药物敏感性增加，促进淋巴瘤细胞粘附及粘附介导的耐药进程。最后，在淋巴瘤细胞中干扰TUBA1A表达，发现整合素β1的胞膜表达下降，并且整合素β1下游通路活性抑制。淋巴瘤细胞粘附能力及耐药能力均发生显著下降。上述研究结果表明，微管蛋白TUBA1A促进整合素β1的胞膜聚集，并通过调节整合素β1下游信号通路影响非霍奇金淋巴瘤细胞微环境介导的耐药形成。.按照合同约定，课题研究成果以论文形式提交。在该基金资助下，项目负责人在项目实施期间共参与发表 SCI 收录论文 4 篇，其中第一作者2 篇；获得南通市卫计委青年科技项目资助课题 1 项；获得南通市第五期“226高层次人才培养工程”第三层次培养对象；参加学术交流8次。