Cytotoxicity and cell death mechanisms against cancer cell lines elicited by the extracts of Physalis Minima L.
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Date
2009
Authors
Ooi, Kheng Leong
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Abstract
Herbal therapy is fast becoming an important alternative medicine for cancer treatment. Therefore, the aims of this study were to determine cytotoxicity of some medicinal herbs and to investigate cell death mechanism elicited by the most potent extract as well as its fractions. A total of 20 extracts form four anticancer plants (Pereskia grandifolia, Vernonia cinerea, Elephantopus scaber and Physalis minima) were screened against three different cancer cell lines. Physalis minima (Leletup-direct translation from Malay) chloroform extract was shown to exhibit remarkable cytotoxic effects on NCI-H23 (human lung adenocarcinoma), T-47D (human breast carcinoma) and Caov-3 (human ovarian carcinoma) cell lines, with EC50 derived at 2.80μg/ml, 3.80μg/ml and 5.10μg/ml, respectively. Different cell death mechanisms exerted by this extract on different cell lines. It was found that DNA fragmentation level of the extract-treated NCI-H23 and T-47D cells was higher than Caov-3 cells. Acute exposure to the extract produced a significant regulation of c-myc, caspase-3 and p53 mRNA expression in all cell lines. Ultrastructural analysis and annexin V staining also demonstrated the presence of apoptotic programmed cell death in the extract-treated cell lines. Furthermore, the appearance of non-apoptotic (vacuolar) morphology was observed in some treated Caov-3 cells and in minority of treated NCI-H23 and T-47D cells. Trypan blue exclusion assay ruled out necrosis as the main cause of death. Thus, cell death mechanism elicited by the Physalis minima chloroform extract appeared to be a mixture of programmed cell death in Caov-3 cells and a major apoptotic cell death in both NCI-H23 and T-47D cells.
Fractionation of Physalis minima chloroform extract revealed 16 different fractions. Only some of the fractions exhibited cytotoxic activities against T-47D cells, with F11 being the most potent, exhibiting the lowest EC50 (3.60μg/ml). Fractions F10, F11 and F13 exhibited typical DNA fragmentation associated with apoptosis in T-47D cells. Trypan blue exclusion assay demonstrated that necrosis did not play a major role in eliciting T-47D cell death of these fractions. Further investigations revealed both fraction F11 and chloroform extract induced major apoptotic cell death in T-47D cells, which was based on majority of weakly diffused annexin V stained cells and significant regulation of c-myc, caspase-3 and p53 mRNA expression levels. Meanwhile, a considerable number of T-47D cells treated with fractions F10 and F13 were stained with annexin V and propidium iodide, which further suggested the presence of a mixture programmed cell death. Physalins B, F and K were identified as major constituents in fraction F11. Combination of these physalin compounds in the extract and fraction may have contributed to the cytotoxic activities and programmed cell death of T-47D cells. These findings warrant further research on Physalis minima plant for use in cancer therapy due to its apoptosis- and autophagy-dependent anticancer effect on cancer cell lines.
Description
PhD
Keywords
Biological Science , Cytotoxicity , Cell death , Cancer cell lines