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Substrate Stiffness Modulates the Growth, Phenotype, and Chemoresistance of Ovarian Cancer Cells.

ABSTRACT: Mechanical factors in the tumor microenvironment play an important role in response to a variety of cellular activities in cancer cells. Here, we utilized polyacrylamide hydrogels with varying physical parameters simulating tumor and metastatic target tissues to investigate the effect of substrate stiffness on the growth, phenotype, and chemotherapeutic response of ovarian cancer cells (OCCs). We found that increasing the substrate stiffness promoted the proliferation of SKOV-3 cells, an OCC cell line. This proliferation coincided with the nuclear translocation of the oncogene Yes-associated protein. Additionally, we found that substrate softening promoted elements of epithelial-mesenchymal transition (EMT), including mesenchymal cell shape changes, increase in vimentin expression, and decrease in E-cadherin and β-catenin expression. Growing evidence demonstrates that apart from contributing to cancer initiation and progression, EMT can promote chemotherapy resistance in ovarian cancer cells. Furthermore, we evaluated tumor response to standard chemotherapeutic drugs (cisplatin and paclitaxel) and found antiproliferation effects to be directly proportional to the stiffness of the substrate. Nanomechanical studies based on atomic force microscopy (AFM) have revealed that chemosensitivity and chemoresistance are related to cellular mechanical properties. The results of cellular elastic modulus measurements determined by AFM demonstrated that Young's modulus of SKOV-3 cells grown on soft substrates was less than that of cells grown on stiff substrates. Gene expression analysis of SKOV-3 cells showed that mRNA expression can be greatly affected by substrate stiffness. Finally, immunocytochemistry analyses revealed an increase in multidrug resistance proteins, namely, ATP binding cassette subfamily B member 1 and member 4 (ABCB1 and ABCB4), in the cells grown on the soft gel resulting in resistance to chemotherapeutic drugs. In conclusion, our study may help in identification of effective targets for cancer therapy and improve our understanding of the mechanisms of cancer progression and chemoresistance.

SUBMITTER: Fan Y

PROVIDER: S-EPMC8421636 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:In vitro cultures of primary cardiac fibroblasts (CFs), the major extracellular matrix (ECM)-producing cells of the heart, are used to determine molecular mechanisms of cardiac fibrosis. However, the supraphysiologic stiffness of tissue culture polystyrene (TCPS) automatically triggers the conversion of CFs into an activated myofibroblast-like state, and serial passage of the cells results in the induction of replicative senescence. These dramatic phenotypic switches confound interpretation of experimental data obtained with cultured CFs. In an attempt to circumvent TCPS-induced activation and senescence of CFs, we utilized poly (ethylene glycol) (PEG) hydrogels as cell culture platforms with low and high stiffness formulations to mimic healthy and fibrotic cardiac ECM, respectively. As hypothesized, low hydrogel stiffness converted activated CFs into a quiescent state with reduced abundance of a-smooth muscle actin (a-SMA)-containing stress fibers. Unexpectedly, lower substrate stiffness concomitantly augmented CF senescence, marked by elevated senescence-associated b-galactosidase (SA-b-Gal) activity and increased expression of p16 and p21, which are cyclin-dependent kinase (CDK) inhibitors and markers of senescence. Using dynamically stiffening hydrogels with phototunable crosslinking capabilities, we demonstrate that substrate-induced CF senescence is partially reversible. RNA-sequencing analysis revealed widespread transcriptional reprogramming of CFs cultured on low stiffness hydrogels, with a dramatic reduction in the expression of profibrotic genes encoding ECM proteins, and an attendant increase in expression of NF-kB-responsive inflammatory genes that typify the senescence-associated secretory phenotype (SASP). Our findings further demonstrate that alterations in matrix stiffness profoundly impact CF cell state transitions, and suggest mechanisms by which CFs change phenotype in vivo depending on the stiffness of the myocardial microenvironment to which they are exposed.

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Substrate Stiffness Modulates the Growth, Phenotype, and Chemoresistance of Ovarian Cancer Cells.

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