Kayamori Kou, Katsube Ken-Ichi, Hirai Hideaki, Harada Hiroyuki, Ikeda Tohru
Department of Oral Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
Department of Oral Pathology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan; Faculty of Human Care, Tohto University, Saitama, Japan.
Lab Invest. 2023 Oct;103(10):100228. doi: 10.1016/j.labinv.2023.100228. Epub 2023 Aug 2.
Wnt signaling plays a crucial role in the progression of various cancers, including oral squamous cell carcinoma (OSCC). However, the tumor microenvironment (TME) regulating Wnt signaling has not yet been fully elucidated. In this study, we investigated whether cancer-associated fibroblasts (CAFs), the primary components of the TME, activate Wnt signaling and promote tumor progression in OSCC. We conducted a Transwell coculture assay using human OSCC cell lines and normal human dermal fibroblasts (NHDFs). NHDFs stimulated WNT7A expression in several OSCC cell lines, especially HO-1-N-1 and HSC-5. An immunohistochemical study using 122 human OSCC samples indicated that high WNT7A expression in tumor cells was significantly associated with invasion depth and poor prognosis. Moreover, WNT7A expression in OSCC cells was positively correlated with α-smooth muscle actin expression in CAFs. WNT7A knockdown in OSCC cells demonstrated that OSCC cells cocultured with NHDFs significantly promoted tumor cell migration and invasion, which was dependent on WNT7A expression in OSCC cells. We also isolated HSC-5 cells from the coculture and conducted microarray analysis to investigate the factors that promote tumor progression induced by WNT7A. Among the various differentially expressed genes, we identified a downregulated gene encoding CLDN1 and confirmed that WNT7A negatively regulated CLDN1 expression in OSCC cells and CLDN1 knockdown in OSCC cells promoted their migration. Phosphokinase array analysis showed that WNT7A activates protein kinase B (AKT) phosphorylation. Activating AKT signaling using the SC79 agonist induced CLDN1 downregulation in OSCC cells. In the coculture assay, the AKT inhibitor MK2206 significantly recovered CLDN1 expression downregulated by WNT7A, resulting in OSCC cell migration suppression. These results suggest that CAFs stimulate OSCC cells to produce WNT7A, following CLDN1 expression downregulation by activating AKT signaling, promoting cancer cell migration. These findings highlight the importance of molecular therapies targeting the TME in OSCC.
Wnt信号通路在包括口腔鳞状细胞癌（OSCC）在内的多种癌症进展中起着关键作用。然而，调节Wnt信号通路的肿瘤微环境（TME）尚未完全阐明。在本研究中，我们调查了作为TME主要成分的癌症相关成纤维细胞（CAFs）是否激活Wnt信号通路并促进OSCC中的肿瘤进展。我们使用人OSCC细胞系和正常人皮肤成纤维细胞（NHDFs）进行了Transwell共培养试验。NHDFs刺激了几种OSCC细胞系中WNT7A的表达，尤其是HO-1-N-1和HSC-5。一项使用122例人OSCC样本的免疫组织化学研究表明，肿瘤细胞中高WNT7A表达与侵袭深度和不良预后显著相关。此外，OSCC细胞中WNT7A表达与CAFs中α平滑肌肌动蛋白表达呈正相关。OSCC细胞中WNT7A基因敲低表明，与NHDFs共培养的OSCC细胞显著促进肿瘤细胞迁移和侵袭，这取决于OSCC细胞中WNT7A的表达。我们还从共培养物中分离出HSC-5细胞并进行微阵列分析，以研究促进WNT7A诱导的肿瘤进展的因素。在各种差异表达基因中，我们鉴定出一个编码CLDN1的下调基因，并证实WNT7A在OSCC细胞中负调节CLDN1表达，且OSCC细胞中CLDN1基因敲低促进其迁移。磷酸激酶阵列分析表明，WNT7A激活蛋白激酶B（AKT）磷酸化。使用SC79激动剂激活AKT信号通路可诱导OSCC细胞中CLDN1下调。在共培养试验中，AKT抑制剂MK2206显著恢复了WNT7A下调的CLDN1表达，从而抑制了OSCC细胞迁移。这些结果表明，CAFs刺激OSCC细胞产生WNT7A，随后通过激活AKT信号通路下调CLDN1表达，促进癌细胞迁移。这些发现突出了针对OSCC中TME的分子疗法的重要性。
