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PO-6

Anti-proliferation and apoptosis induction activities of Senna siamea

(Lam.) Irwin & Barneby stem extract on KB cell line

1,2 2 1,2
Jiradej Manorsoi , Pongsathorn Dhumtanom , Aranya Manosroi
1
Faculty of Pharmacy, Chiang Mai University, Chiang Mai 50200, Thailand
2
Natural Products Research and Development Center (NPRDC), Institute for Science and Technology Research and Development,
Chiang Mai University, Chiang Mai 50000, Thailand

Rationale: Thai copper pod (Senna siamea (Lam.) Irwin & Barneby) or çKhi Leké in Thai, is the plant in the
family Fabaceae (Leguminosae) subfamily Caesalpinioideae. It has been traditionally used for the treatment of
fever, skin disease, constipation, diabetes, hypertension and insomnia. This plant is widely cultivated in Thailand.
Its leaves and flowers are used as ingredients in Thai curry, while its stem is one of the compositions of many Thai
medicinal recipes for anti-cancer and insomnia. However, anti-proliferation and apoptosis induction activities of the
extract of the stem have never been investigated in order to evaluate for its anti-cancer potential.

Objective: An anti-proliferation activity by sulforhodamine B (SRB) assay and apoptosis induction by morpho-
logical changes and DNA fragmentation of the extract from the stem of Thai copper pod were investigated in KB
(human mouth epidermal carcinoma) cell line.

Methodology: Anti-proliferation (using SRB assay) and apoptosis induction (using cell morphological changes

and DNA fragmentation determination) of the extract from Thai copper pod stem in KB cell line were investigated.
Stems of the plant were dried, chopped and boiled in hot water for 1 h, filtered and then freeze-dried. The extract
at the concentrations of 0.05-2.0 mg/ml were incubated with the KB cells in the 96-well tissue culture plates for
24 h and in 24-well tissue culture plates for 3-24 h, for SRB and apoptosis induction assays, respectively. Cell
morphological changes, apoptotic bodies or single large vesicles, were identified under phase-contrast microscope.

Treated cells stained with acridine orange and ethidium bromide for the determination of the nuclei formation were
identified under fluorescense phase-contrast microscope. DNA fragmentation was identified from the genomic DNA
extracted from the treated cells and electrophoresis on agarose gel.

Results: The Thai copper pod stem extract showed poor anti-proliferation activity against KB cell line. The IC
50
value of the extract was 1.54 mg/ml which was 128.3 times higher than doxorubicin of which the IC was 0.012
50
mg/ml. However, the extract at the concentration of 0.5 mg/ml which was 3 times lower than its IC , the single
50
large vesicles were induced within 24 h. Nuclei condensation (few green spots within the nucleus of the cells)
designated the early apoptosis phase was observed within 6 h when the cells were treated with 0.5 mg/ml of the
extract. Also, DNA fragmentation was induced within 24 h at this same extract concentration.

Conclusions: Although the extract from the stem of Thai copper pod did not show high anti-proliferation activity,

it gave an interesting apoptosis induction activity on KB cell line. At the concentration of the extract 3 times lower
than its IC , it demonstrated single large vesicle induction within 24 h and DNA fragmentation within 6 h. This
50
study has suggested that the extract from the stem of Thai copper pod had the potential for further development
for anti-cancer therapy because of its potent apoptosis induction activity.