Smuczek Basilio, Santos Emerson de S, Siqueira Adriane S, Pinheiro Joao J V, Freitas Vanessa M, Jaeger Ruy G
Department of Cell and Developmental Biology, Institute of Biomedical Sciences, University of Sao Paulo, Sao Paulo, Brazil.
Department of Cell and Developmental Biology, Institute of Biomedical Sciences, University of Sao Paulo, Sao Paulo, Brazil; School of Pharmaceutical Sciences of Ribeirao Preto, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil.
Exp Cell Res. 2017 Sep 15;358(2):323-334. doi: 10.1016/j.yexcr.2017.07.005. Epub 2017 Jul 5.
Breast cancer is an important public health problem, and its progression may be related to the extracellular matrix (ECM), which acts as a structural scaffold and instruction source for neoplastic cells. Laminins are ECM proteins regulating tumor biology. The laminin-derived peptide C16 regulates different properties of tumor cells. Here we analyzed C16-induced differential gene expression in MDA-MB-231 breast cancer cells. MCF-10A normal-like breast cells served as control. Among different cancer-related genes, C16 induced overexpression of GPNMB. This gene encodes a transmembrane protein GPNMB (glycoprotein non-metastatic B), involved with malignant phenotype of breast cancer cells. Immunoblot validated microarray results. To correlate gene and protein expression with cellular function, we investigated whether C16 would regulate invasion in breast cancer cells. siRNA experiments strongly suggested that C16 and GPNMB cooperate to regulate invasion of highly aggressive MDA-MB-231 cancer cells. We addressed regulatory mechanisms involved in C16-mediated increase of GPNMB protein levels in MDA-MB-231 cells, and observed that C16 stimulates β1 integrin and Src phosphorylation. Furthermore, Src inhibition decreases peptide-induced GPNMB expression levels. To contextualize in vivo our results in vitro, we addressed GPNMB immunostaining in breast cancer human tissue microarrays. Quantitative immunohistochemistry showed that GPNMB is significantly more expressed in breast cancer compared to normal tissue. We concluded that laminin-derived peptide C16 regulates gene and protein expression of GPNMB in breast cancer cells. C16 and GPNMB may cooperate to regulate invasion of highly aggressive MDA-MB-231 cells, probably through Src signaling. GPNMB presented increased expression in breast cancer in vivo compared to normal breast tissue.
乳腺癌是一个重要的公共卫生问题，其进展可能与细胞外基质（ECM）有关，细胞外基质作为肿瘤细胞的结构支架和指令来源。层粘连蛋白是调节肿瘤生物学的细胞外基质蛋白。层粘连蛋白衍生肽C16调节肿瘤细胞的不同特性。在此，我们分析了C16诱导的MDA-MB-231乳腺癌细胞中的差异基因表达。MCF-10A正常样乳腺细胞作为对照。在不同的癌症相关基因中，C16诱导了GPNMB的过表达。该基因编码一种跨膜蛋白GPNMB（糖蛋白非转移性B），与乳腺癌细胞的恶性表型有关。免疫印迹验证了微阵列结果。为了将基因和蛋白质表达与细胞功能相关联，我们研究了C16是否会调节乳腺癌细胞的侵袭。小干扰RNA实验强烈表明，C16和GPNMB协同调节高侵袭性MDA-MB-231癌细胞的侵袭。我们探讨了MDA-MB-231细胞中C16介导的GPNMB蛋白水平增加所涉及的调节机制，并观察到C16刺激β1整合素和Src磷酸化。此外，Src抑制降低了肽诱导的GPNMB表达水平。为了将我们的体外结果与体内情况相结合，我们对乳腺癌人组织微阵列进行了GPNMB免疫染色。定量免疫组织化学显示，与正常组织相比，GPNMB在乳腺癌中的表达明显更高。我们得出结论，层粘连蛋白衍生肽C16调节乳腺癌细胞中GPNMB的基因和蛋白质表达。C16和GPNMB可能协同调节高侵袭性MDA-MB-231细胞的侵袭，可能是通过Src信号传导。与正常乳腺组织相比，GPNMB在体内乳腺癌中的表达增加。
