RIP3 overexpression sensitizes human breast cancer cells to parthenolide in vitro via intracellular ROS accumulation
Abstract
Can LU1, #, Li-yan ZHOU1, #, Hui-jun XU1, Xing-yu CHEN1, Zhong-sheng TONG1, *, Xiao-dong LIU1, Yong-sheng JIA1, Yue CHEN2
1Department of Breast Oncology, Tianjin Medical University Cancer Institute and Hospital, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China; 2The State Key Laboratory of Medicinal Chemical Biology, Synergetic Innovation Center of Chemical Science and Engineering (Tianjin), College of Pharmacy, and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300060, China
Aim: Receptor-interacting protein 3 (RIP3) is involved in tumor necrosis factor receptor signaling, and results in NF-κB-mediated prosurvival signaling and programmed cell death. The aim of this study was to determine whether overexpression of the RIP3 gene could sensitize human breast cancer cells to parthenolide in vitro.
Methods: The expression of RIP3 mRNA in human breast cancer cell lines (MCF-7, MDA-MB-231, MDA-MB-435 and T47D) was detected using RT-PCR. Both MDA-MB-231 and MCF-7 cells were transfected with RIP3 expression or blank vectors via lentivirus. Cell viability was measured with MTT assay; intracellular ROS level and cell apoptosis were analyzed using flow cytometry.
Results: RIP3 mRNA expression was not detected in the four human breast cancer cell lines tested. However, the transfection induced higher levels of RIP3 protein in MCF-7 and MDA-MB-231 cells. Furthermore, overexpression of RIP3 decreased the IC50 values of parthenolide from 17.6 to 12.6 μmol/L in MCF-7 cells, and from 16.6 to 9.9 μmol/L in MDA-MB-231 cells. Moreover, overexpression of RIP3 significantly increased parthenolide-induced apoptosis and ROS accumulation in MCF-7 and MDA-MB-231 cells. Pretreatment with N-acetyl-cysteine abrogated the increased sensitivity of RIP3-transfected MCF-7 and MDA-MB-231 cells to parthenolide.
Conclusion: Overexpression of RIP3 sensitizes MCF-7 and MDA-MB-231 breast cancer cells to parthenolide in vitro via intracellular ROS accumulation.
Keywords: breast cancer; receptor-interacting protein 3; parthenolide; reactive oxygen species; apoptosis; N-acetyl-cysteine
This work was supported by the Anticancer Key Technologies R & D Program of Tianjin, China (No 12ZCDZSY16200).
# These authors contributed equally to this work.
* To whom correspondence should be addressed.
E-mail 18622221181@163.com
Received 2013-06-20 Accepted 2014-01-03
Keywords:
1Department of Breast Oncology, Tianjin Medical University Cancer Institute and Hospital, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China; 2The State Key Laboratory of Medicinal Chemical Biology, Synergetic Innovation Center of Chemical Science and Engineering (Tianjin), College of Pharmacy, and Tianjin Key Laboratory of Molecular Drug Research, Nankai University, Tianjin 300060, China
Aim: Receptor-interacting protein 3 (RIP3) is involved in tumor necrosis factor receptor signaling, and results in NF-κB-mediated prosurvival signaling and programmed cell death. The aim of this study was to determine whether overexpression of the RIP3 gene could sensitize human breast cancer cells to parthenolide in vitro.
Methods: The expression of RIP3 mRNA in human breast cancer cell lines (MCF-7, MDA-MB-231, MDA-MB-435 and T47D) was detected using RT-PCR. Both MDA-MB-231 and MCF-7 cells were transfected with RIP3 expression or blank vectors via lentivirus. Cell viability was measured with MTT assay; intracellular ROS level and cell apoptosis were analyzed using flow cytometry.
Results: RIP3 mRNA expression was not detected in the four human breast cancer cell lines tested. However, the transfection induced higher levels of RIP3 protein in MCF-7 and MDA-MB-231 cells. Furthermore, overexpression of RIP3 decreased the IC50 values of parthenolide from 17.6 to 12.6 μmol/L in MCF-7 cells, and from 16.6 to 9.9 μmol/L in MDA-MB-231 cells. Moreover, overexpression of RIP3 significantly increased parthenolide-induced apoptosis and ROS accumulation in MCF-7 and MDA-MB-231 cells. Pretreatment with N-acetyl-cysteine abrogated the increased sensitivity of RIP3-transfected MCF-7 and MDA-MB-231 cells to parthenolide.
Conclusion: Overexpression of RIP3 sensitizes MCF-7 and MDA-MB-231 breast cancer cells to parthenolide in vitro via intracellular ROS accumulation.
Keywords: breast cancer; receptor-interacting protein 3; parthenolide; reactive oxygen species; apoptosis; N-acetyl-cysteine
This work was supported by the Anticancer Key Technologies R & D Program of Tianjin, China (No 12ZCDZSY16200).
# These authors contributed equally to this work.
* To whom correspondence should be addressed.
E-mail 18622221181@163.com
Received 2013-06-20 Accepted 2014-01-03