[The influence of different WT1 gene isoforms expression pattern on the differentiation of leukemia cell line NB4].
OBJECTIVE
To study the potential effects of exogenous WT1 gene isoform on the differentiation of leukemia cell line NB4 and its possible molecular mechanisms.


METHODS
The recombinant eukaryotic expression vector (pCB6 + /WTA) containing full-length human WT1 isoform (WTA: -17AA/ -KTS) cDNA and the blank pCB6 + vector were transfected into leukemia cell line NB4 by electroporation. The WTA mRNA and protein in cells were detected by RT-PCR and Western blot. Cell morphology, NBT reduction and CD11b antigen expression in NB4 cells were assayed to evaluate cell differentiation. Expression of PML/RARalpha, p21 and c-myc genes was determined by semi-quantitative RT-PCR after transfection.


RESULTS
Compared with NB4/WTA cells, NB4 and NB4/CMV (NB4 cells transfected with pCB6 + vector) cells had higher morphological differentiation rates and higher CD11b expression levels after exposure to ATRA for 48 hours. The percentage of NBT reduction in NB4/WTA cells was lower than that in control groups. The difference in NBT reduction rate between NB4/WTA and control cells was gradually increased after treated with ATRA for three days. The expression levels of PML/RARalpha, p21 and c-myc genes in NB4/WTA cells were notably increased.


CONCLUSIONS
Overexpression of exogenous WTA gene could partially inhibit the differentiation of NB4 cells by up-regulating the expression of PML/RARalpha, p21 and c-myc genes.
Jiangsu Institute of Hematology, The First Hospital Affiliated to Suzhou University, China.
Hui-ling
HL
Shen
Zi-xing
ZX
Chen
Wei
W
Wang
Jian-nong
JN
Cen
Shao-yan
SY
Hu
Ye
Y
Zhao
D002453:Cell Cycle; D002454:Cell Differentiation / Q000235:genetics*; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D005822:Genetic Vectors; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D014162:Transfection; D025721:WT1 Proteins / Q000235:genetics* / Q000378:metabolism
D050759:Cyclin-Dependent Kinase Inhibitor p21; D016271:Proto-Oncogene Proteins c-myc; D012333:RNA, Messenger; D025721:WT1 Proteins