Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer, with increasing incidence worldwide. The molecular basis of cSCC progression to invasive and metastatic disease is still incompletely understood. Here, we show that fibroblasts and transforming growth factor-β (TGF-β) signaling promote laminin-332 synthesis in cancer cells in an activated H-Ras-dependent manner, which in turn promotes cancer cell invasion. Immunohistochemical analysis of sporadic UV-induced invasive human cSCCs (n = 208) revealed prominent cSCC cell specific immunostaining for laminin-332 γ2 chain, located in the majority of cases (90%, n = 173) in the invasive edge of the tumors. To mimic the progression of cSCC we established 3D spheroid cocultures using primary skin fibroblasts and HaCaT/ras-HaCaT human keratinocytes. Our results indicate that in 3D spheroids, unlike in monolayer cultures, TGF-β upregulates laminin-332 production, but only in cells that harbour oncogenic H-Ras. Accumulation of laminin-332 was prevented by both H-Ras knock down and inhibition of TGF-β signaling by SB431542 or RAdKD-ALK5 kinase-defective adenovirus. Furthermore, fibroblasts accelerated the invasion of ras-HaCaT cells through collagen I gels in a Ras/TGF-β signaling dependent manner. In conclusion, we demonstrate the presence of laminin-332 in the invasive front of cSCC tumors and report a new Ras/TGF-β-dependent mechanism that promotes laminin-332 accumulation and cancer cell invasion.

中文翻译：

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H-Ras 激活和成纤维细胞诱导的 TGF-β 信号传导促进皮肤鳞状细胞癌中层粘连蛋白 332 的积累和侵袭。


皮肤鳞状细胞癌（cSCC）是最常见的转移性皮肤癌，全球发病率不断增加。 cSCC 进展为侵袭性和转移性疾病的分子基础仍不完全清楚。在这里，我们发现成纤维细胞和转化生长因子-β (TGF-β) 信号以激活的 H-Ras 依赖性方式促进癌细胞中层粘连蛋白 332 的合成，进而促进癌细胞侵袭。对散发性紫外线诱导的侵袭性人类 cSCC (n = 208) 的免疫组织化学分析显示，层粘连蛋白-332 γ2 链具有显着的 cSCC 细胞特异性免疫染色，该链位于大多数病例 (90%，n = 173) 的肿瘤侵袭边缘。为了模拟 cSCC 的进展，我们使用原代皮肤成纤维细胞和 HaCaT/ras-HaCaT 人角质形成细胞建立了 3D 球体共培养物。我们的结果表明，与单层培养物不同，在 3D 球体中，TGF-β 上调层粘连蛋白 332 的产生，但仅限于含有致癌 H-Ras 的细胞。 H-Ras 敲除和 SB431542 或 RAdKD-ALK5 激酶缺陷型腺病毒对 TGF-β 信号传导的抑制可防止层粘连蛋白-332 的积累。此外，成纤维细胞通过 I 型胶原蛋白凝胶以 Ras/TGF-β 信号传导依赖性方式加速 ras-HaCaT 细胞的侵袭。总之，我们证明了层粘连蛋白 332 在 cSCC 肿瘤的侵袭性前沿的存在，并报告了一种新的 Ras/TGF-β 依赖性机制，可促进层粘连蛋白 332 积累和癌细胞侵袭。