Microvesicles Derived From Human Wharton’s Jelly Mesenchymal Stromal Cells Inhibit Bladder Cancer Cells Growth Via ATM-p53 Pathway

Objective: Bladder cancer is the most common malignant in urinary tumors, especially transitional cell carcinoma, including superficial bladder cancer and current invasive bladder cancer in clinical. And invasive bladder cancer has a highdegree malignancy, easy to transfer, and has a poor prognosis. In our previous study, Microvesicles(MVs) from human umbilical cord Wharton’s jelly mesenchymal stem cells(human umbilical cord Wharton’s jelly mesenchymal stem cells,hWJMSCs) had been proved to inhibit bladder cancer cells proliferation and induce apoptosis. While it had been documented that hWJMSC-MVs may suppress bladder cancer cell proliferation by activating the p53-p21 cascade. However, the regulatory factors ATM(Ataxia telangiectasia mutated kinase) that lie upstream of p53-p21 pathway remain to be identified. There were researchs reported that down-regulation of ATM protein sensitizes cancer cells to stress-induced apoptosis. Herein, we hypothesized that ATM express level before treatment of hWJMSC-MVs may be one of the key factors controling cancer cells fate. To explore the regulatory mechanism of ATM upon hWJMSC-MVs attenuating bladder cancer cells growth, we will alter ATM protein express using pharmaceuticals technique, and then analysize the consequent effect variations of hWJMSC-MVs on bladder cancer cells growth and express or activation of major target proteins of ATM-p53-p21. This study will lay a sound foundation for microvesicles application for cancer therapy in clinical.Methods:1.We used MTT assay to analyze effects of hWJMSC-MVs on cell viability in human bladder cancer cells.2. Flow cytometry was used to estimate the cell cycle and apoptosis of bladder cancer cell.3. Under different conditions, we estimated the expression of ATM/p-ATM, p53, p21, Bax and cleaved Caspase 3 by Western blot technique.Results:1. Increasing the dose of hWJMSC-MVs induce to the decrease of the percentage of viability in examined 3 different bladder cancer cell lines. The highest sensitive cell were T24 cells.2. hWJMSC-MVs induced G0/G1 arrest was mediated by ATM-p53 signaling pathway.3.The hWJMSC-MVs increased the activity of cleaved caspase 3 and affected the Bax protein levels in T24 cells.4. Abrogation of G0/G1 arrest by inhibiting ATM-p53 enhanced apoptosis induced by hWJMSC-MVs.Conclusion:We showed that apoptosis was induced in hWJMSC-MVs treated T24 cells following G0/G1 arrest and this effect was dose dependent. This study for the first time established ATM–p53 pathway was activated in this progression, hWJMSC-MVs induced the up-regulation of p-ATM and p53, p21 protein level. The present study provides a new insight into the relationship between G0/G1 arrest and apoptosis, since the activation of ATM plays a protective role for cell survival. Understanding of the detailed mechanism of G0/G1 arrest has a potential role in the treatment of bladder cancer. Our results suggest that a better understanding of the complex scenario in which cancer cells respond to hWJMSC-MVs may enable more efficient cancer cell-specific killing when hWJMSC-MVs is correctly combined with other chemotherapeutic agents. In the near future, MVs, releasing from mesenchymal stem cells, might become a novel anti-tumor medicine in the clinic.