[Effect of doxorubicin induced GM-CSF gene expression regulated by Egr-1 promoter on recovery of hematopoietic function in bearing-melanoma mice].
This study was purposed to investigate the effect of hematopoietic growth factor expression regulated by Egr-1 promoter on the recovery of hematopoietic function in bearing-melanoma mice after chemotherapy with doxorubicin (ADM). The human GM-CSF cDNA and enhanced green fluorescence protein (GFP) cDNA were linked together with internal ribozyme entry site (IRES) and then were inserted into the expression vector pCI-neo under control of the Egr-1 promoter (Egr-EG). This vector was transduced into human bone marrow stromal cell lines HFCL by lipofectamine and was transfused in severe combined immunodeficiency (SCID) mice. The experimental mice were randomly divided into 4 groups (6 mice in each group): (1) HFCL/EG + ADM group in which the HFCL/EG cells were transplanted intravenously in SCID mice bearing melanoma, ADM was given intraperitoneally after 3 days; (2) HFCL + ADM group in which the HFCL cells were transplanted intravenously, ADM was given intraperitoneally after 3 days; (3) HPCL/EG group in which HFCL/EG cells were transplanted alone; (4) HFCL group in which HFCL cells were transplanted alone. The dynamic change of peripheral blood picture was assayed by hemocytometer; the eGFP(+) human stromal cells were detected by flow cytometry; the expression of GM-CSF mRNA and protein were determined by RT-PCR and Western blot respectively. The results indicated that as compared with HFCL/EG and HFCL groups, the leukocyte count in HFCL/EG + ADM group decreased, but decrease level was weaker than that in HFCL + ADM group, meanwhile the recovery of leukocyte count was earlier than that in HFCL + ADM group. The CFU-GM amount between 4 groups showed no significant difference. The detection results showed that the inhibitory rate of tumor was related to chemotherapy, but not to expression of exogenous gene; the eGFP(+) stromal cells existed in bone marrow of mice treated with ADM. The RT-PCR and Western blot assays revealed enhancement of GM-CSF mRNA and protein. It is concluded that the ADM-inducible GM-CSF gene therapy regulated by Egr-1 promoter may promote the hematopoietic recovery after chemotherapy.
DU
Nan
N
Department of Hematology and Oncology, The First Affiliated Hospital, PLA General Hospital, Beijing, 100037, China. dunan05@yahoo.com.cn
Pei
Xue-Tao
XT
Zhou
Jin-Ming
JM
Sun
Jun-Zhong
JZ
Zhao
Hui
H
Fu
Yan
Y
Hao
Yi-Xin
YX
D000818:Animals; D004317:Doxorubicin / Q000494:pharmacology*; D051766:Early Growth Response Protein 1 / Q000235:genetics*; D005260:Female; D015870:Gene Expression; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D006410:Hematopoiesis / Q000187:drug effects*; D006412:Hematopoietic Stem Cells; D006801:Humans; D051379:Mice; D016513:Mice, SCID; D011401:Promoter Regions, Genetic
D051766:Early Growth Response Protein 1; D004317:Doxorubicin; D016178:Granulocyte-Macrophage Colony-Stimulating Factor