Evaluation of some genes and proteins involved in Apoptosis on human chronic myeloid leukemia cells (K562 cells) by Datura innoxia Leaves Aqueous Extract.

Datura innoxia (D.innoxia) has an extensive usage in traditional medicine and can also be used for intervention therapy in order to treat cancer. Despite of accomplishing some researches on D.innoxia mechanism, still our knowledge is very little about exact D.innoxia apoptotic mechanism on human chronic myeloid leukemia cells (K562 cells). This study purpose was to clarify the molecular mechanism of apoptosis, which was mediated by D.innoxia leaves aqueous extract in K562 cells. MTT assay and flow cytometry was applied in order to assess the viability and apoptosis induction of K562 cells and normal human lymphoid B cells in the D.innoxia presence. Finally, the expression of the apoptotic related genes (p53, BAX, BCL2, Caspases 3, 6, 7 and 9) were evaluated using quantitative Real-Time PCR. Western blot analysis was applied for assessing the protein expression. MTT results indicated that D.innoxia could inhibit the viability of K562 cells in a dose- and time-dependent manner. In parallel, D.innoxia inhibitory effect on normal human lymphoid B cells was lower in comparison with its effect on K562 cells at the same concentrations and same incubation time. Apoptosis induction in K562 cells after D.innoxia exposure was determined by flow cytometry. Apoptosis was activated by D.innoxia in K562 cells throughout increasing the expression of P53, BAX/BCL2 ratio, caspase 9, 3, 6, 7. Western blot analysis demonstrated significant increase in cleaved PARP-1 and cleaved caspase 3 in treated K562 cells with high D.innoxia leaves aqueous extract concentration. D.innoxia leaves trigger apoptosis in K562 cells throughout intrinsic apoptotic pathway.