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. 2013 Nov;16(11):1203-8.

Antiproliferative activity and apoptosis induction of crude extract and fractions of avicennia marina

Amir Abbas Momtazi-Borojeni  1 , Mandana Behbahani  1 , Hojjat Sadeghi-Aliabadi  2
Affiliations

Antiproliferative activity and apoptosis induction of crude extract and fractions of avicennia marina

Amir Abbas Momtazi-Borojeni et al. Iran J Basic Med Sci. 2013 Nov.

Abstract

Objective(s): Regarding the presence of many active biological constituents in Avicennia marina, the present investigation was carried out to study cytotoxic activity of crude methanol leave extract and column chromatographic fractions of A. marina against MDA-MB 231 cell line (human breast cancer cell) and HEK (Human embryonic kidney cell) line.

Materials and methods: The anticancer activity of crude methanol extract and sub-fractions were evaluated, using MTT assay. The induction of apoptosis was determined by analyzing DNA fragmentation in breast cancer cells treated with active fraction of crude methanol extract using agarose gel electrophoresis. To investigate molecular mechanism of apoptosis, gene expression levels of p53 and Bcl-2 were measured using quantitative real time PCR.

Results: Fraction 10 was the most active fraction and was detected with HPLC as luteolin. The 50% cell cytotoxic concentration (CC50) of crude methanol extract and luteolin was 250 and 28 µg/ml, respectively. This fraction was found to be an apoptotic agent against MDA-MB 231 cells, which leads to causing DNA fragmentation. The mRNA expression level of Bcl-2 and p53 was significantly decreased and increased respectively in cancer cells treated by luteolin.

Conclusion: The results suggested that Luteolin isolated from Avicennia marina could probably induce apoptosis on breast cancer cell line by the regulation of p53 and Bcl-2 pathways.

Keywords: Apoptosis; Avicennia marina; Cytotoxic activity; DNA fragmentation; MDA-MB 231 cell line.

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Figures

Figure 1
Figure 1
HPLC analysis of fraction 10 (A) and standard luteolin (B). Chromatographic conditions: RPC18 column, mobile phase: methanol/acetone/water = 70/20/10 (v/v/v) at 1.5 mL min–1
Figure 2
Figure 2
Cytotoxic activity of crude methanol extract ( ) and fraction 10 ( ) from Avicennia marina against MDA- MB 231 and HEK cell lines
Figure 3
Figure 3
Agarose gel electrophoresis (1.8%) of the chromosomal DNA extracted of MDA-MB 231 cells. A: 1kb DNA marker, B: Control cells treated with 0.5 % DMSO, C and D: cells treated with fraction 10 and crude methanol extract respectively, at CC50 concentration for 48 hr
Figure 4
Figure 4
Time dependency effects of the p53 (A), Bcl-2 (B) mRNA levels in human breast cancer cell line, MDA- MB 231, incubated with 28 µg/ml of Luteolin after 2, 6 and 12 hr incubation. GAPDH was used as an endogenous control gene
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