Connexins act as tumor suppressors in three-dimensional mammary cell organoids by regulating differentiation and angiogenesis
Connexins act as tumor suppressors in three-dimensional mammary cell organoids by regulating differentiation and angiogenesis
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DOI：
10.1158/0008-5472.can-05-4302
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发表时间：
2006-10-15
期刊：
影响因子：
11.2
通讯作者：
Laird, Dale W.
中科院分区：
文献类型：
--
作者：
McLachlan, Elizabeth;Shao, Qing;Laird, Dale W.
连接蛋白是肿瘤抑制因子，并且人乳腺连接蛋白26（Cx 26）和连接蛋白43（Cx43）间隙连接在乳腺癌中经常下调。我们以前的研究表明，在MDA-MB-231乳腺癌细胞中过表达的Cx 26和Cx43在体内抑制肿瘤生长，但在二维培养中没有。在目前的研究中，我们表明Cx 26或Cx43的过表达在三维环境中具有肿瘤抑制特性，从而减少了锚定非依赖性细胞生长并诱导了MDA-MB-231细胞三维类器官的部分再分化。重要的是，大多数外源性连接蛋白没有定位到细胞-细胞界面或救援间隙连接细胞间通讯（GJIC）的染料转移评估，提供了一个GJIC独立的乳腺肿瘤抑制机制的证据。为了进一步阐明连接蛋白诱导的肿瘤细胞三维再分化的机制，我们研究了连接蛋白表达是否在上皮间质转化（EMT）中起作用。Cx 26和Cx43减少细胞迁移，增加细胞角蛋白18的表达，并降低波形蛋白水平，表明从间充质向上皮表型的转变。此外，我们研究了连接蛋白在血管生成中的作用，通过探测血管生成抗体阵列与条件培养基从三维MDA-MB-231文化。这揭示了连接蛋白过表达调节多种血管生成相关蛋白。此外，从连接蛋白过表达细胞分泌的因子抑制内皮细胞小管形成和迁移，Cx43过表达MDA-MB-231细胞异种移植物显示减少肿瘤血管生成。总之，Cx 26和Cx43通过GJIC非依赖性机制抑制MDA-MB-231细胞的恶性特性，包括调节EMT和血管生成。
Connexins are tumor suppressors, and human breast connexin 26 (Cx26) and connexin 43 (Cx43) gap junctions are often down-regulated in breast cancer. We previously showed that Cx26 and Cx43 overexpressed in MDA-MB-231 breast cancer cells inhibited tumor growth in vivo but not in two-dimensional cultures. In the current study, we show that overexpression of Cx26 or Cx43 has tumor-suppressive properties in a three-dimensional environment such that they reduced anchorage-independent cell growth and induced partial redifferentiation of three-dimensional organoids of MDA-MB-231 cells. Importantly, the majority of exogenous connexins did not localize to the cell-cell interface or rescue gap junctional intercellular communication (GJIC) as assessed by dye transfer, providing evidence of a GJIC-independent mechanism of mammary tumor suppression. To further elucidate the mechanisms involved in connexin-induced three-dimensional redifferentiation of tumor cells, we examined whether connexin expression has a role in epithelial to mesenchymal transition (EMT). Cx26 and Cx43 reduced cell migration, increased cytokeratin 18 expression, and decreased vimentin levels, indicating a shift from a mesenchymal towards an epithelial phenotype. In addition, we examined the role of connexins in angiogenesis by probing an angiogenesis antibody array with conditioned media from three-dimensional MDA-MB-231 cultures. This revealed that connexin overexpression regulated various angiogenesis-linked proteins. Furthermore, secreted factors from connexin overexpressing cells inhibited endothelial cell tubulogenesis and migration, and xenografts of Cx43 overexpressing MDA-MB-231 cells showed reduced tumor angiogenesis. In summary, Cx26 and Cx43 inhibit the malignant properties of MDA-MB-231 cells via GJIC-independent mechanisms, including regulation of EMT and angiogenesis.