|Suspended and adherent tumor cells sense and respond differently to fluid shear stress via YAP-p73 mediated mechanotransduction
|Tan, Youhua (BME)
Yang, Mo (BME)
Cancer -- Research
Hong Kong Polytechnic University -- Dissertations
|Department of Biomedical Engineering
|115 pages : color illustrations
|Tumor cells spread and disseminate to remote organs mainly through the blood circulation system. Within the circulatory system, circulating tumor cells (CTCs) are exposed to considerable levels of fluid shear stress, which is capable of eliminating many CTCs and thus hindering tumor cell dissemination. Even so, a small subset of CTCs survives in blood circulation. However, the response of suspended tumor cells to fluid shear stress is still unclear. Thus, it is pivotal to investigate the underlying mechanotransduction signalling by which CTCs sense and respond to fluid shear stress. Further targeting this mechanotransduction signalling may better eliminate CTCs in the blood circulation, thereby preventing tumor metastasis.
This study took breast cancer as a model of disease and developed in vitro microfluidics-based systems to mimic fluid shear flow in blood circulation in vivo. We found that fluid shear stress eradicated the majority of suspended breast tumor cells (SBTCs), and cell viability depended on the magnitude of fluid shear stress and circulation duration, indicating that SBTCs can sense and respond to fluid shear stress. In particular, fluid shear stress enhanced the expression level of Piezo1 but not Piezo2 in SBTCs. Inhibition/activation of Piezo1 increased/decreased suspended tumor cell survival in shear flow by regulating Piezo1-mediated calcium entry. Further, fluid shear stress enhanced actin polymerization via Piezo1-mediated calcium entry in SBTCs, which led to suspended tumor cell apoptosis. Inhibiting actin polymerization rescued Piezo1-mediated calcium entry and thus tumor cell death, suggesting that fluid shear stress regulates the survival of SBTCs through Piezo1-mediated actin polymerization. Importantly, as the downstream effector of Piezo1-F-actin, fluid shear stress promoted YAP/TAZ nuclear localization. In the nucleus, YAP interacted with p73 but not TEADs via Y357 phosphorylation in SBTCs, which promoted the transcription of the pro-apoptotic gene PUMA and induced cell apoptosis. Activating YAP/TAZ reduced the viability of SBTCs under shear stress, whereas inhibiting YAP/TAZ promoted cell survival. These findings suggest that shear-induced YAP/TAZ activation promotes the apoptosis of suspended tumor cells. YAP/TAZ mediated the effect of the Piezo1-F-actin mechanotransduction signalling on SBTCs, which indicates that fluid shear stress leads to the apoptosis of SBTCs via Piezo1-F-actin mediated YAP/TAZ nuclear translocation. Additionally, fluid shear stress inhibited the activity of LATS1/2 and MST1/2 in Hippo pathway of SBTCs and further promoted YAP nuclear translocation. Silencing/activating LATS1/2 or MST1/2 reduced/increased the viability of SBTCs under fluid shear stress, which could be further reversed by YAP/TAZ inhibition/activation. These findings suggest that Hippo pathway is involved in the response of SBTCs to hemodynamic shear stress.
To confirm the role of YAP in the CTC survival in blood circulation in vivo, we utilized the Luciferase p-Nluc, which was retained in healthy cells while released only from dead and dying ones, to quantify the death of CTCs in blood circulation in vivo. The results showed that inhibiting YAP enhanced the survival of CTCs in the vasculature of mice. Further, YAP activation inhibited the long-term lung metastasis in the mouse model.
In the vasculature, CTCs are arrested and adhere to the endothelium where they are subjected to hemodynamic shear forces. Therefore, we further investigated the influence of fluid shear stress on adherent breast tumor cells (ABTCs). Our results showed that while fluid shear stress decreased the viability of suspended tumor cells, adherent tumor cells evaded shear-induced apoptosis through downregulating Piezo1-F-actin-YAP/TAZ-p73 signalling. Activating Piezo1 or YAP could increase the vulnerability of adherent tumor cells to fluid shear stress by inducing cell apoptosis. These results suggest that breast tumor cells in suspension and adherent exhibit distinct responses to fluid shear stress.
In conclusion, these findings suggest that hemodynamic shear stress has substantial impacts on the survival of suspended but not adherent tumor cells in blood circulation. Our studies have unveiled that suspended/adherent tumor cells sense and respond to fluid shear stress through activating/inactivating Piezo1-F-actin-YAP/TAZ-p73/PUMA signalling, which further leads to the apoptosis of suspended but not adherent tumor cells. Therefore, this study not only has elucidated the mechanotransduction mechanism by which suspended tumor cells sense fluid shear stress but also demonstrated the dependence of mechanosensing on the suspension/adherence state of tumor cells. Further, we have identified Piezo1 and YAP/TAZ as potential therapeutic targets, through which CTCs may be effectively eliminated in the vascular system to prevent tumor metastasis.
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