Overexpression or knock-down of runt-related transcription factor 1 affects BCR-ABL-induced proliferation and migration in vitro and leukemogenesis in vivo in mice.
BACKGROUND
Runt-related transcription factor 1 (Runx1) plays a crucial role in hematogenesis and its dysfunction may contribute to leukemogenesis. However, it is not clear whether or not abnormal expression of Runx1 will induce leukemia and how the change of Runx1 expression level could affect BCR-ABL-induced leukemogenesis. In the present study, we aimed to analyze if abnormal expression of Runx1 in BaF3 cells alone would induce leukemogenesis. And we also wanted to know if abnormal expression of Runx1 in leukemic cells would affect leukemogenesis. Furthermore, we investigated whether overexpression or knock-down of Runx1 in BaF3 cells would induce leukemogenesis.


METHODS
Plasmids containing full-length Runx1 cDNA were transduced into BaF3 cells and BaF3-P185wt cells (BCR-ABL transformed BaF3 cells) by electroporation. Plasmids containing a short hairpin RNA of Runx1 were transduced into BaF3 cells and BaF3-P185wt cells by electroporation. Runx1 expression level was quantified by Western blotting and quantitative real-time PCR. The effects of overexpression or knock-down of Runx1 on proliferation, apoptosis and migration of cells were detected in vitro. Then, using MSCV-P185wt-EGFP as a control, we transplanted MSCV-P185wt-Runx1 cells or MSCV-P185wt-shRNA cells into Balb/c mice through tail vein and observed tumorgenesis of the different phenotypes.


RESULTS
In vitro analysis revealed that overexpression of Runx1 in P185wt cells could inhibit cell proliferation and slow down cell migration; while knock-down of Runx1 could promote cell proliferation and speed up cell migration. In vivo analysis indicated that mice transplanted with MSCV-P185wt-Runx1 survived longer than controls. In contrast, mice transplanted with MSCV-P185wt-shRNA survived shorter than the control group. Gross pathological analysis revealed that the MSCV-P185wt-Runx1 group had less severe splenomegaly and hepatomegaly compared to the control group, and the MSCV-P185wt-shRNA group had more severe splenomegaly and hepatomegaly. No splenomegaly or hepatomegaly was detected in mice transplanted with MSCV-BaF3-Runx1 cells or MSCV-BaF3-shRNA cells. Both the mice of MSCV-BaF3-Runx1 group and MSCV-BaF3-shRNA group were healthy with no sign of leukemia for up to three months.


CONCLUSIONS
Overexpression or knock-down of Runx1 gene in BaF3 cells alone could not induce leukemogenesis. However, in BaF3-P185wt cells, alteration of Runx1 expression could affect BCR-ABL-induced proliferation and migration in vitro and leukemogenesis in vivo.
Yang
Li-jun
LJ
Department of Pediatrics, Peking University People's Hospital, Beijing 100044, China.
Yu
Wei-dong
WD
DU
Jun-bao
JB
Chao
Shuang
S
Chen
Min-xia
MX
Zhao
He-hua
HH
Guo
Jing-zhu
JZ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects*; D050676:Core Binding Factor Alpha 2 Subunit / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D016044:Fusion Proteins, bcr-abl / Q000494:pharmacology*; D007938:Leukemia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D051379:Mice; D008807:Mice, Inbred BALB C; D016133:Polymerase Chain Reaction
D050676:Core Binding Factor Alpha 2 Subunit; D016044:Fusion Proteins, bcr-abl