Shear Stress Increases V–H + -ATPase and Acidic Vesicle Number Density, and p-mTORC2 Activation in Prostate Cancer Cells

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Abstract

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 cm - 2 wall shear stress for 24 h, followed by immunocytochemistry and fluorescence measurements of β1 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 β1 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.

Original languageEnglish
Pages (from-to)591-604
Number of pages14
JournalCellular and Molecular Bioengineering
Volume13
Issue number6
DOIs
StatePublished - Dec 2020

Keywords

  • Actin cytoskeleton
  • Mechanical stimuli
  • Mechanobiology
  • mTOR pathway

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