Basic Study On The Effects Of Arsenic Trioxide On Molecular Of Hedgehog Signaling Pathway In Chronic Myelogenous Leukemia

Objective:The human chronic myelogenous leukemia(CML)cell line K562 is investigated to research the effect of Arsenic trioxide(ATO)on the apdptosis of K562 cells and expression levels of the mRNAs and proteins of Gli1,Gli2,Smo and Ptch related to Hedgehog(Hh)signaling pathways.Meanwhile,the influence of ATO on the possible mechanism of Hh pathway activity is also analyzed to provide a reliable theoretical basis for applying ATO to CML with abnormal activation of Hh pathway.Methods:(1)K562 cells were treated with different concentrations of ATO for 48 hours.After Annexin V-FITC/PI staining,flow cytometry was used to detect the apoptosis of K562 cells induced by ATO.(2)Western blot(WB)was used to detect the effect of ATO on the protein expression level changes of Glil,Gli2 and Ptch2 proteins in Hh pathway,and the effect on CML key pathogenic molecules P210/Bcr-Abl.(3)Quantitative real-time PCR(qRT-PCR)was used to detect the effect of ATO on the mRNA level of Glil,Gli2,Smo and Ptch in Hh pathway.Results:(1)After 48h ATO treatment with different concentrations,K562 cells showed apoptosis effects in all experimental groups.With the increase of ATO concentration,the apoptosis was more obvious in a dose-dependent manner.(2)ATO significantly inhibited the expression of Gli2 and P210/Bcr-Abl protein in a dose-dependent manner(P<0.05);ATO had a certain promoting effect on Ptch2 protein expression but inhibited at high doses(P>0.05);ATO had no obvious inhibitory effect on Glil protein expression.(3)ATO had a significant inhibitory effect on the mRNA expression of Glil and Gli2 in a dose-dependent manner(P<0.05);ATO could up-regulate the mRNA expression of Ptch(P<0.05);However,the mRNA expression of Smo was not significantly inhibited(P>0.05)in all concentrations of ATO.Conclusion:(1)ATO has obvious apoptosis-inducing effect on K562 cells.(2)ATO may use Gli2 protein as a target to inhibit Hh pathway activity and play a role in degradation of P210/Bcr-Abl protein.(3)ATO may inhibit the abnormal activation of Hh pathway by inhibiting the mRNA expression levels of Glil and Gli2 and up-regulating the mRNA expression level of Ptch.