[Reversal of multi-drug resistance in K562/A02 cells by two short hairpin RNAs (shRNA) of mdr1].
OBJECTIVE
To construct two recombinant plasmids of mdr1 and mcl1 shRNA, and to investigate their reversal effect on drug resistance in K562 adriamycin resistant cell lines (K562/A02).


METHODS
Two oligonucleotides of mdr1 and mcl1 gene were designed referring to that of GenBank, double-stranded DNA was derived through annealing, and cloned into pRNAT vector digested by two restricted endoenzymes. K562/A02 cells were transfected with the recombinant plasmids. The mdr1 mRNA expression and its protein product P-glycoprotein (P-gp) were detected by RT-PCR and flow cytometry. The expression of mcl1 gene was detected by RT-PCR. 50% inhibition concentration (IC50) of adriamycin (ADM) on K562/A02 cells was determined by MTT method. Cells apoptosis was analyzed by flow cytometry.


RESULTS
Comparing with K562/A02 cells, the shRNA of mdrl or mcl1 gene in vitro can remarkably increase the sensitivity of K562/A02 to adriamycin, down-regulate mdr1 or mcl1 gene expression, increase the K562/A02 cells apoptosis rates induced by adriamycin. Cotransfection of mdrl and mcl1 genes shRNA can also down-regulate the expression of their gene, more remarkably increase the sensitivity and apoptosis of K562/ A02 to adriamycin.


CONCLUSIONS
Transfection of mdrl or mcl1 gene shRNA can promote the sensitivity of K562/A02 to adriamycin and cotransfection of the two shRNA can more remarkably do so. The mel1 gene might be involved in adriamycin resistant in K562/A02 cells.
Yu
Hai-Qing
HQ
Department of Hematology, Qilu Hospital of Shandong University, Jinan 250012, China.
Ji
Chun-Yan
CY
Ma
Dao-Xin
DX
Zang
Shao-Lei
SL
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000096:biosynthesis / Q000235:genetics*; D017209:Apoptosis; D018432:Drug Resistance, Multiple / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D020014:K562 Cells; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger