[Recombinant adeno-associated virus 2-mediated green fluorescent protein expression in bone marrow mesenchymal stem cells derived from acute myelogenous leukemia patients].
OBJECTIVE
To explore the possibility of using autologous bone marrow mesenchymal stem cells (BMSC) as a vehicle to deliver recombinant adeno-associated virus 2-mediated enhanced green fluorescent protein (rAAV-2-eGFP) in vitro, therefore to find an alternative solution for gene therapy of hematological malignancy.


METHODS
BMSCs isolated from the bone marrow of patients with acute myelogenous leukemia (AML) at the onset of disease were infected by rAAV-2-eGFP at different multiplicity of infection (MOI=10(2), 10(3), 10(4), 10(5), 10(6), and 10(7), respectively). Phase-contrast fluorescent microscope and flow cytometry were employed to evaluate the expression of enhanced green fluorescent protein (eGFP).


RESULTS
Ten to fourteen days after the transfection, eGFP expression began to be detected and the transfection efficiency ranged between 0.3% to 2%, which failed to be increased with the increase of MOI. The transduced eGFP could maintain a long-term stable expression in vitro in the 61 days of observation, and from 12 to 33 days after transfection, eGFP percentage underwent a decrease from the initial 1.16% to 0.5%-0.6% and maintained this expression level till 61 days after transfection.


CONCLUSIONS
rAAV can be used with BMSCs for in vitro gene therapy, but the poor transfection efficiency of these cells remains a significant obstacle for its further application.
Xie
Zheng-Jun
ZJ
Department of Hematology, Nanfang Hospital, Southern Medical University, Guangzhou510515, China. zhj_xie@hotmail.com
Zheng
Wei-Yang
WY
Xu
Bing
B
Song
Llan-Lin
LL
Yi
Zheng-Shan
ZS
Yin
Fang
F
Zhou
Shu-Yun
SY
D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism*; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000378:metabolism* / Q000473:pathology; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins