The Mechanisms Of Matrine Alleviating Retinoic Acid Resistance In Acute Promyelocytic Leukemia

Objective:To explore the role of Telomerase activity (TA) and cAMP/PKA signaling pathway in alleviating all trans retinoic acid (ATRA) resistance in acute promyelocytic leukemia (APL) with matrine(MAT).Methods:ATRA sensitive cell line of APL (NB4) and resistant cell line (NB4-R1) were used in this study. The influence of1μmol/L ATRA,0.1mmol/L MAT,1μmol/L ATRA combined with0.1mmol/L MAT on the differentiation of the two cell lines was tested by nitro blue tetrazolium chioride(NBT) staining. PCR-ELISA was used to detect the telomerase activity (TA) in different groups. The expression of cell surface differentiation markers, CD11b, after treatment with protein kinaseA inhibitor, H89, was determined by flow cytometry. In addition, RT-PCR and Western blotting were used to evaluate the effects of H89on the mRNA expression and the protein level of PML/RARa and PLSCR1in the two cell lines.Results:(1) ATRA alone or combined with MAT significantly increased NBT-reducing activity in NB4cells (P<0.01), but there was no statistical difference between the two groups (P>0.05).In NB4-R1cells, only combination therapy can improved the resistance of ATRA in increasing NBT-reducing activity (P<0.05).(2) ATRA treatment significantly inhibited TA in NB4cells (P<0.01), but has no obvious effect on NB4-R1cells (P>0.05). In conjunction with MAT has strong suppression of TA in two cell lines (NB4:P<0.01, NB4-R1:P<0.05). (3) ATRA alone can promote the CD11b expression in NB4(P<0.01) and NB4-R1cells (P<0.05), but further elevated when MAT was added in NB4-R1cells (P<0.01). The effects of ATRA (NB4:P<0.01, NB4-R1:P<0.05) or combined with MAT (NB4:P<0.01, NB4-R1:P<0.01) on the expression of CD11b were abolished by H89.(4) ATRA significantly inhibited the PML/RARa and promoted the PLSCR1expression at the mRNA and protein level in NB4, but for NB4-R1, ATRA alone can only enhance the protein level of PML/RARa and PLSCR1, evident effects can be seen in the combination therapy groups.(5) The effects of ATRA or MAT on the protein and transcriptional level of PML/RARα and PLSCR1can be blockd by H89in both cell lines.Conclusion:MAT alleviated the ATRA resistance in APL by regulating the expression of PML/RARα and PLSCR1at the genetic and protein level through the cAMP/PKA signaling pathway and inhibiting the telomerase activity.