In vitro phenotypic correction of hematopoietic progenitors from Fanconi anemia group A knockout mice.
Fanconi anemia (FA) is a rare autosomal recessive disease, characterized by bone marrow failure and cancer predisposition. So far, 8 complementation groups have been identified, although mutations in FANCA account for the disease in the majority of FA patients. In this study we characterized the hematopoietic phenotype of a Fanca knockout mouse model and corrected the main phenotypic characteristics of the bone marrow (BM) progenitors using retroviral vectors. The hematopoiesis of these animals was characterized by a modest though significant thrombocytopenia, consistent with reduced numbers of BM megakaryocyte progenitors. As observed in other FA models, the hematopoietic progenitors from Fanca(-/-) mice were highly sensitive to mitomycin C (MMC). In addition, we observed for the first time in a FA mouse model a marked in vitro growth defect of Fanca(-/-) progenitors, either when total BM or when purified Lin(-)Sca-1(+) cells were subjected to in vitro stimulation. Liquid cultures of Fanca(-/-) BM that were stimulated with stem cell factor plus interleukin-11 produced low numbers of granulocyte macrophage colony-forming units, contained a high proportion of apoptotic cells, and generated a decreased proportion of granulocyte versus macrophage cells, compared to normal BM cultures. Aiming to correct the phenotype of Fanca(-/-) progenitors, purified Lin(-)Sca-1(+) cells were transduced with retroviral vectors encoding the enhanced green fluorescent protein (EGFP) gene and human FANCA genes. Lin(-)Sca-1(+) cells from Fanca(-/-) mice were transduced with an efficiency similar to that of samples from wild-type mice. More significantly, transductions with FANCA vectors corrected both the MMC hypersensitivity as well as the impaired ex vivo expansion ability that characterized the BM progenitors of Fanca(-/-) mice.
Río
Paula
P
Hematopoietic Gene Therapy Program, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT)/Marcelino Botín Foundation, 22840 Madrid, Spain.
Segovia
José Carlos
JC
Hanenberg
Helmut
H
Casado
José Antonio
JA
Martínez
Jesús
J
Göttsche
Kerstin
K
Cheng
Ngan Ching
NC
Van de Vrugt
Henri J
HJ
Arwert
Fré
F
Joenje
Hans
H
Bueren
Juan A
JA
D000818:Animals; D017209:Apoptosis; D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism; D018929:Cell Culture Techniques; D004268:DNA-Binding Proteins; D005199:Fanconi Anemia / Q000473:pathology* / Q000628:therapy; D052217:Fanconi Anemia Complementation Group A Protein; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000627:therapeutic use; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D016685:Mitomycin / Q000494:pharmacology; D010641:Phenotype; D011506:Proteins / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic
D004268:DNA-Binding Proteins; C498471:FANCA protein, human; C120277:Fanca protein, mouse; D052217:Fanconi Anemia Complementation Group A Protein; D011506:Proteins; D016685:Mitomycin