Experimental study on apoptosis of leukemia cell line NB4 transfected with WT1 gene.
- Author:
Hui-Ling SHEN
1
;
Zi-Xing CHEN
;
Wei WANG
;
Jian-Nong CEN
;
Shao-Yan HU
;
Ye ZHAO
Author Information
1. Jiangsu Institute of Hematology, The First Hospital Affiliated to Suzhou University, Suzhou 215006, China.
- Publication Type:Journal Article
- MeSH:
Apoptosis;
genetics;
physiology;
Blotting, Western;
Cell Line, Tumor;
Cyclin-Dependent Kinase Inhibitor p21;
genetics;
Gene Expression Regulation, Neoplastic;
Humans;
Leukemia;
genetics;
metabolism;
pathology;
Proto-Oncogene Proteins c-bcl-2;
genetics;
Proto-Oncogene Proteins c-myc;
genetics;
RNA, Messenger;
biosynthesis;
genetics;
Reverse Transcriptase Polymerase Chain Reaction;
Transfection;
Tumor Suppressor Protein p53;
genetics;
WT1 Proteins;
genetics;
metabolism;
physiology;
bcl-X Protein;
genetics
- From:
Journal of Experimental Hematology
2005;13(6):989-995
- CountryChina
- Language:Chinese
-
Abstract:
In order to study the potential effects of exogenous WT1 gene isoform on apoptosis in leukemia cell line NB4 and its possible molecular mechanisms, the eukaryotic expression recombinant vector (pCB6(+)/WTA) containing full-length human WT1 isoform (WTA: -17aa/-KTS) cDNA and the vacant vector-alone were introduced into the leukemia cell line NB4 respectively by electroporation. The WTA mRNA and protein in cells were detected by RT-PCR and Western blot. Binding of Annexin V were tested by flow cytometry and agarose gel electrophoresis to verify whether exogenous WTA could induce apoptosis of NB4 cells. Expressions of p21, p53, bcl-2, bcl-XL and c-myc genes were determined by semi-quantitative RT-PCR after introducing recombinant vectors into the NB4 cells. The results showed that in exposure to As(2)O(3) at 0.8 micromol/L for 48 hours, the NB4/WTA cells exhibited the morphological hallmarks of apoptosis, the marked DNA ladder shown by gel electrophoresis, and the enhanced apoptosis rate marked by Annexin V. RT-PCR showed an increase in p21 and c-myc genes expression, a decrease in bcl-2 and a relative constant expression of p53, bcl-XL in NB4/WTA cells. It is concluded that the introduction and expression of exogenous WTA gene can lead to apoptosis of NB4/WTA cells by down-regulating the Bcl-2 gene expression and up-regulating the p21 and c-myc genes expression.
