Introduction

Cells in the tumor microenvironment experience mechanical stresses, such as compression generated by uncontrolled cell growth within a tissue, increased substrate stiffness due to tumor cell extracellular matrix (ECM) remodeling, and leaky angiogenic vessels which involve low fluid shear stress. With our hypothesis that shear stress increases V–H\(^+\)-ATPase number density in prostate cancer cells via activation of the mTORC1 and mTORC2 pathways, we demonstrated and quantified such a mechanism in prostate cancer cells.

Methods

Moderately metastatic DU145 and highly metastatic PC3 prostate cancer cells were subjected to 0.05 dynes \(\hbox {cm}^{-2}\) wall shear stress for 24 h, followed by immunocytochemistry and fluorescence measurements of \(\beta\)1 integrin, endosome, lysosome, V–H\(^{+}\)-ATPase proton pump, mTORC1, and p-mTORC2 antibodies. Post shear stress migration assays, and the effects of vacuolar proton pump inhibitor Bafilomycin A1 (60 nM, 24 h) as well as shear stress on the ICC fluorescence intensity of the proteins of interest were conducted with DU145 cells.

Results

Low fluid shear stress increases the fluorescence intensity of \(\beta\)1 integrin, endosome, lysosome, V–H\(^{+}\)-ATPase, mTORC1, and p-mTORC2 antibodies in PC3 and DU145 cells, and also increased cell migration. However, Bafilomycin A1 decreased fluorescence intensity of all of these proteins in DU145 cells exposed to shear stress, revealing that V–H\(^+\)-ATPase controls the expression of these proteins.

Conclusions

Prostate cancer cell mechanotransduction increases endosomes, lysosomes, and proton pumps—where increases have been associated with enhanced cancer aggressiveness. We also show that the prostate cancer cell’s response to force promotes the cancer drivers mTORC1 and mTORC2.

中文翻译：

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剪切应力增加 V–H $$^{+}$$ + -ATPase 和酸性囊泡数密度，以及前列腺癌细胞中 p-mTORC2 的激活

介绍

肿瘤微环境中的细胞会经历机械应力，例如组织内不受控制的细胞生长产生的压缩、肿瘤细胞细胞外基质 (ECM) 重塑导致的基质硬度增加以及涉及低流体剪切应力的血管生成血管渗漏。我们假设剪切应力通过激活 mTORC1 和 mTORC2 途径增加前列腺癌细胞中的 V-H \(^+\) -ATPase 数量密度，我们证明并量化了前列腺癌细胞中的这种机制。

方法

对中度转移性 DU145 和高度转移性 PC3 前列腺癌细胞施加 0.05 达因\(\hbox {cm}^{-2}\)壁剪切应力 24 小时，然后对\(\beta\) 1进行免疫细胞化学和荧光测量整合素、内体、溶酶体、V-H \(^{+}\) -ATPase 质子泵、mTORC1 和 p-mTORC2 抗体。用 DU145 细胞进行后剪切应力迁移测定，液泡质子泵抑制剂 Bafilomycin A1（60 nM，24 小时）以及剪切应力对感兴趣蛋白质的 ICC 荧光强度的影响。

结果

低流体剪切应力会增加 PC3 和 DU145 细胞中\(\beta\) 1 整合素、内体、溶酶体、V-H \(^{+}\) -ATPase、mTORC1 和 p-mTORC2 抗体的荧光强度，以及也增加了细胞迁移。然而，巴弗洛霉素 A1 降低了暴露于剪切应力的 DU145 细胞中所有这些蛋白质的荧光强度，表明 V-H \(^+\) -ATPase 控制这些蛋白质的表达。

结论

前列腺癌细胞机械转导增加了内体、溶酶体和质子泵——这些增加与增强的癌症侵袭性有关。我们还表明，前列腺癌细胞对力的反应促进了癌症驱动因子 mTORC1 和 mTORC2。