Trejo Vazquez Jessica Angelina, Towle Rebecca, Farnsworth Dylan Andrew, Sarafan Masih, Lockwood William Wallace, Garnis Cathie
Department of Integrative Oncology, British Columbia Cancer Research Center, Vancouver, BC V5Z1L3, Canada.
Interdisciplinary Oncology Program, University of British Columbia, Vancouver, BC V5Z1L3, Canada.
Biomedicines. 2024 Nov 4;12(11):2523. doi: 10.3390/biomedicines12112523.
Lung cancer, including the major subtype lung adenocarcinoma (LUAD), is the leading cause of cancer deaths worldwide, largely due to metastasis. Improving survival rates requires new treatment strategies and a deeper understanding of the mechanisms that drive tumor progression within the tumor microenvironment (TME). This study investigated the impact of extracellular vesicles (EVs) derived from LUAD cells on lung fibroblasts. EVs were isolated from LUAD cell lines via ultracentrifugation and characterized using nanoparticle tracking analysis and Western blotting. Lung fibroblasts were treated with PBS, TGFβ, or EVs, and their activation was assessed through protein (Western blotting) and RNA analyses (RNA seq and RT-qPCR). The results confirmed the TGFβ induced activation and showed that LUAD EVs could also activate fibroblasts, increasing cancer-associated fibroblast (CAF) markers. While EV-induced CAF activation displayed unique features, like an increase in proliferation-related genes, the EV and TGFβ treatments also shared some differentially expressed genes. The EV groups induced a higher expression of ECM remodeling and EMT-associated genes, but some of those genes were also upregulated in the TGFβ group. Mesenchymal genes and were significantly upregulated in TGFβ- and EV-treated fibroblasts. Their secretion as proteins from the TGFβ- and EV-induced CAFs was not significant, confirmed through ELISA. These findings suggest that LUAD EVs play a role in CAF activation through both shared and distinct pathways compared to canonical TGFβ activation, potentially identifying novel gene expressions involved in CAF activation. Additionally, optimal protein secretion conditions of confirmed CAF-upregulated genes need to be established to determine their contribution to the TME.
肺癌，包括主要亚型肺腺癌（LUAD），是全球癌症死亡的主要原因，主要是由于转移。提高生存率需要新的治疗策略以及对肿瘤微环境（TME）中驱动肿瘤进展的机制有更深入的了解。本研究调查了源自LUAD细胞的细胞外囊泡（EVs）对肺成纤维细胞的影响。通过超速离心从LUAD细胞系中分离出EVs，并使用纳米颗粒跟踪分析和蛋白质印迹法对其进行表征。用磷酸盐缓冲液（PBS）、转化生长因子β（TGFβ）或EVs处理肺成纤维细胞，并通过蛋白质分析（蛋白质印迹法）和RNA分析（RNA测序和逆转录定量聚合酶链反应（RT-qPCR））评估其活化情况。结果证实了TGFβ诱导的活化，并表明LUAD EVs也可以激活成纤维细胞，增加癌症相关成纤维细胞（CAF）标志物。虽然EV诱导的CAF活化表现出独特的特征，如增殖相关基因增加，但EV和TGFβ处理也有一些差异表达基因。EV组诱导了更高的细胞外基质重塑和上皮-间质转化（EMT）相关基因表达，但其中一些基因在TGFβ组中也上调。间充质基因在TGFβ和EV处理的成纤维细胞中显著上调。通过酶联免疫吸附测定（ELISA）证实，它们作为蛋白质从TGFβ和EV诱导的CAF中分泌并不显著。这些发现表明，与经典的TGFβ活化相比，LUAD EVs通过共同和不同的途径在CAF活化中发挥作用，可能识别出参与CAF活化的新基因表达。此外，需要建立已确认的CAF上调基因的最佳蛋白质分泌条件，以确定它们对TME的贡献。
