Document Type: Original Article(s)
Center for Advanced facility for Life Sciences, Deogiri College, Aurangabad, India
School of Life Sciences, Jawaharlal Nehru University, New Delhi, India
Background: Calotropis gigantea (family: Asclepiadaceae) has been known to contain cardiac glycosides. The C. gigantea extracts have been reported as cytotoxic to a few cancer cell lines. The present study was designed to examine the effect of Calotropis gigantea methanolic extract (CGME) on the growth and apoptosis in human breast carcinoma cell line (MCF-7 cells).Methods: The study was conducted in Aurangabad (India) from 16 February to 10 June 2015. CGME treated MCF-7 cells were analysed for growth inhibition and apoptosis. The exhibition of phosphatidylserine was analysed with the Annexin-V Fluorescein isothiocyanate flow cytometry (FITC) method. Accumulated poly-caspases were determined with carboxyfluorescein poly-caspase assay, Apo-BrdU™ tunnel assay for DNA fragmentation and pro/anti-apoptotic gene expression with real-time polymerase chain reaction. The high-performance liquid chromatography analysis indicated the presence of two unknown cardenolides along with known cardenolides such as calactin, calatropagenin, usharin, afroside, calatoxin, and gamphoside. The Kruskal-Wallis and Wilcoxon tests were used for statistical analyses. Results: Upon treatment with 40 µg/ml CGME, about 56.9% of the cell population underwent apoptosis. Compared to paclitaxel, the accumulation of active caspases in CGME treated with MCF-7 cells was found to be dose-dependent, whereas the G2/M cell cycle arrest was time dependent. The Apo-BrdU™ tunnel assay confirmed that CGME treatment caused DNA fragmentation and RT-PCR analyses indicated elevated transcription for pro-apoptotic gene expression. Kruskal-Wallis test results were significant; Bcl-2 (P=0.00193), Bak-1 (P=0.00021), and Bax (P=0.0019).Conclusion: CGME treatment caused the accumulation of phosphatidylserine on the cell membrane, recruitment of poly-caspases, DNA fragmentation, and enhanced transcription of pro-apoptotic gene expression.