Abstract

Essential thrombocythemia (ET) is a clonal myeloproliferative disease that involves primarily the megakaryocytic lineage. ET is characterized by sustained thrombocytosis in the blood and increased in numbers of large, mature megakaryocytes in the marrow and, occasionally, in the extramedullary organs. Currently, there is no known genetic or biologic marker specific for ET. The etiology and pathogenesis of ET remain largely unclear, partially due to a lack of suitable animal model for the disease. We reported here the development of a transgenic (TG) mouse model, in which most aged mice presented with thrombocytosis, marrow megakaryocytic and myeloid hyperplasia, splenomegaly with marked extramedullary hematopoiesis, features that closely mimic ET in humans. TG mice were generated via microinjection of a mammalian construct consisting of a mitochondrial isoform of human 8-oxoguanine-DNA glycosylase (hOGG) under the control of a mouse metallothionein-1 (mMT-1) promoter. A total of 11 founder mice were obtained and shown to successfully integrate the transgene in their genome, as verified by PCR. Two of the male founder mice successfully transmitted the transgene to their offspring at an expected frequency of 50%. Three founder mice at the age of 12 months and 5 F1 offspring at the age of 4 months were examined. All aged hOGG TG founder mice and their offspring expressed high levels of hOGG mRNA in their liver by RT-PCR and direct DNA sequencing. Upon histologic examination, 3 TG founder mice displayed moderate to severe splenomegaly. The spleen weights were 2-, 4- and 10 times respectively in the 3 TG founder mice as compared to wild type, age-matched mice. Microscopically, the red pulp in the enlarged spleens was markedly expanded with aggregates of large, mature but dysplastic-appearing megakaryocytes, focally disrupting the normal splenic structure. Immunostaining for myeloperoxidase highlighted multiple clusters of myeloid precursors in the spleen of aged hOGG TG mice but none in the spleen of wild type aged mice. Peripheral blood from these aged TG founder mice showed marked thrombocytosis and platelet clumping. In bone marrow, the aged TG founder mice displayed marked myeloid and megakaryocytic hyperplasia without marrow fibrosis, indicative of myeloid and megakaryocytic proliferation. Of note, none of the above phenotype was seen in younger hOGG TG mice (4 months), indicating that the full development of this pathologic process was dependent on age. In summary, TG mice overexpressing the mitochondrial isoform of human OGG gene developed a phenotype, closely mimicking ET in humans and the full manifestation of this phenotype was age-dependent. The molecular basis for this process is currently unclear and remains speculative. One plausible and attractive hypothesis is related to mitochondrial DNA damage with resultant mitochondrial dysfunction, in which overexpression of hOGG gene causes more active repair of free radical-induced 8-oxoguanine from DNA, leaving an increased number of abasic sites, which may generate mutation, inhibit transcription and, ultimately, leading to mitochondrial dysfunction and development of this myeloproliferative disorder. Extensive molecular characterization is currently underway to explore this possibility.

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