Fig. 2.

Hydrodynamic shear stress (SS) given as oscillatory shear stress (OSS) led to the acquirement of epithelial-mesenchymal transition (EMT) and stemness marker genes in MDA-MB231 cells. a Schematic illustration of in vitro fluid SS, including cone-and-plate viscometer-based laminar shear stress (LSS) or OSS and orbital shaker-based hydrodynamic SS. b (i) Number of suspension cells during LSS (270 rpm corresponds to 20 dyne/cm²) conditions of MDA-MB231 cells in non-coated Petri dishes after 24 h of culture. Cell viability was measured by trypan blue exclusion assay and error bars represent ± SD calculated from at least three independent experiments. Expression level of SS-induced (Egr1, Ap1 and Epcam (ii)), stemness marker (Nanog. Oct4B and Sox2; (iii)), EMT marker (N-Cadherin, Twist and Snail1; (iv)), and epithelial marker (E-Cadherin, Claudin-7, and Cytokeratin-8; (v)) genes in cells in suspension during LSS conditions of MDA-MB231 cells in non-coated Petri dishes, analyzed by quantitative real-time RT-PCR. c (i) Number of suspension cells during OSS (67 rpm corresponds to 5 dyne/cm²) conditions in MDA-MB231 cells in non-coated Petri dishes after 24 h of culture. Expression level of SS-induced (Egr1, Ap1 and Epcam; (ii)), stemness marker (Nanog. Oct4B and Sox2; (iii)), EMT marker (N-Cadherin, Twist and Snail1; (iv)), and epithelial marker (E-Cadherin, Claudin-7, and Cytokeratin-8; v) genes in suspension cells during OSS conditions of MDA-MB231 cells in non-coated Petri dishes, analyzed by quantitative real-time RT-PCR analysis. d Number of suspension cells during +SS (30–240 rpm corresponds to 2.25–18 dyne/cm²) conditions of breast cancer (MDA-MB231, MCF7) cells in non-coated Petri dishes after 24 h of culture. e Ratio of suspension cells in hydrodynamic SS (+SS) conditions (60 rpm corresponds to 4.5 dyne/cm²) of the indicated cells in non-coated Petri dishes were assessed on day 3, 5, 7, and 10. Cell viability was measured by trypan blue exclusion assay and error bars represent ± SD calculated from at least three independent experiments. f Expression of SS-induced (Egr1 and Epcam), stemness marker (Nanog and Oct4B), EMT marker (N-Cadherin and Twist), and epithelial marker (E-Cadherin, Claudin-7, and Cytokeratin-8) genes expressed in the indicated cells over time on the indicated days, analyzed by quantitative real-time RT-PCR; ^#p < 0.05, ^*p < 0.01. Each gene expression in quantitative real-time RT-PCR analysis was normalized to Gapdh. The data presented here are presented as mean ± SEM and are representative of three independent experiments. Statistically significant differences were tested at p < 0.05 significance