Momordica charantia Linn., commonly known as bitter gourd, has many protective roles due to its medicinal value as it contains bioactive components. However, this extract showed possible toxicity effect on zebrafish embryo. Thus this study was designed to differentiate the toxicity activities in two types of M. charantia sample which are Indian and Chinese M. charantia, as well as to compare between two different aqueous extraction methods, hot and cold aqueous method, using zebrafish embryo assay assessment. It was observed that the survival rate of zebrafish embryo decreased as the concentration of test extract increased for all samples of M. charantia. The LC50 values of hot aqueous Chinese M. charantia, hot aqueous Indian M. charantia, and cold aqueous Chinese M. charantia were 144.54 μg/ml, 199.53 μg/ml, and 251.19 μg/ml, respectively. However, cold aqueous Indian M. charantia has a higher LC50 which was not in the range of the tested concentration. Hatchability of Danio rerio embryo reduced as the concentration of M. charantia extract increased while no hatching was observed in the highest concentration (1000 μg/ml). Scoliosis of zebrafish larvae was only seen in higher concentrations (125-1000 μg/ml) of extract. The heartbeat of zebrafish larvae treated with M. charantia extract was within the normal range, 120-180 bpm, but at higher concentrations (125-1000 μg/ml) the heartbeat differed for all samples of test extract. Hence, although this plant extract was safe to be consumed due to its pharmaceutical effect, it still exhibited mild toxicity effect at higher concentration when it was evaluated on zebrafish embryo.
Lung cancer is the leading cause of cancer related deaths worldwide with about 40% occurring in developing countries. The two varieties of Momordica charantia, which are Chinese and Indian bitter melon, have been subjected to antiproliferative activity in human non-small cell lung cells A549. The A549 cells were treated with hot and cold aqueous extraction for both the bitter melon varieties, and the antiproliferative activity was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The apoptotic mechanism of action on A549 human lung cancer cells was evaluated first morphologically using Hoechst 33358, and cytoskeleton staining using Filamentous-actin (F-actin) cytoskeleton FICT and DAPI followed by caspase-3/7, reactive oxygen species (ROS), and p53 activity. Chinese hot aqueous extraction (CHA) exhibited potent antiproliferative activity against A549 human lung cancer cells. The morphological analysis of mitochondria destruction and the derangement of cytoskeleton showed apoptosis-inducing activity. CHA increased the caspase-3/7 activity by 1.6-fold and the ROS activity by 5-fold. Flow cytometric analysis revealed 34.5% of apoptotic cells significantly (p<0.05) compared to cisplatin-treated A549 human cancer cells. CHA is suggested to induce apoptosis due to their rich bioactive chemical constituents. These findings suggest that the antiproliferative effect of CHA was due to apoptosis via ROS-mediated mitochondria injury.
A therapy which can target apoptosis will be effective for cancer treatment. This preliminary study attempted in vitro and in vivo to target apoptosis capability of Momordica charantia extracts. Here, we describe that the crude hot and cold extraction of two varieties (Chinese and Indian) of M.charantia, induced apoptosis in human lung cancer cell line A549. This was obtained from the cell viability assay where crude extracts were incubated for 24 hours and tested with 3-(4,5-Dimethylthiazol-2-yl)-2,5 Diphenyltetrazolium Bromide (MTT) assay. The inhibitory concentration, IC50 was obtained from the cell viability graph and further used in the upstream assays. The apoptotic morphology was presented by the crude extract-treated cells with a comparison to a positive control cisplatin. The percentage of apoptosis was obtained from a flow cytometric analysis using Hoechst 33342. Induction of apoptosis in vivo was tested using the extracts on dechorionated zebrafish embryo for 24 hours and monitored up to 72-hour post fertilization. Induction of apoptosis in A549 by crude water extracts revealed a decrease in cell viability giving a low IC50. The IC50 of cisplatin was 17.3 µg/ml, Chinese Hot Aqueous was 32.5 µg/ml, Chinese Cold Aqueous was 28.1 µg/ml, Indian Hot Aqueous was 36.9 µg/ml and Indian Cold Aqueous was 26.7 µg/ml significant to the control.The initiation of apoptosis by the crude water extracts was also evidence in vivo in zebrafish embryos with results demonstrating significant changes morphologically. There is a potential for the Indian Cold Aqueous to act as a chemo preventive agent as it has the overall best IC50, percentage of apoptotic cells and changes in the morphology of cell and the zebrafish embryo.
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