[Transfection and expression of human thrombopoietin cDNA for treatment of thrombocytopenia in mice].
OBJECTIVE
To explore that COS-7 cells efficiently transfected with human thrombopoietin (hTPO) cDNA from fetal liver and hTPO potentially treated carboplatin-induced thrombocytopenia in mice.


METHODS
Using molecular cloning technique, expression vectors PCIneo under the control of SV40 early. Applying liprofectin mediating method, recombinant plasmids PCIneo-hTPO were transfected into COS-7 cells in addition to effect of G418. After hTPO cDNA and mRNA in colony COS-7 cells were respectively identified by PCR-Southern and dot blot, hTPO was analysed with sandwich ELISA and administrated intraperitoneally to mice with thrombocytopenia.


RESULTS
COS-7 cells transfected with constructed PCIneo-hTPO expressed and secreted hTPO up to 48.28 ng/ml in the supernatant. We kinetically observed that the number of megakaryocyte-colony-forming unit (CFU-MK) in bone marrow enhanced to threefold (P < 0.01), especially the number of small CFU-MK, accompanied by an increased mean megakaryocyte volume (P < 0.01) and circulating platelets returned to a normal level (80 +/- 26, x 10(9)/L, P < 0.01) in peripheral blood.


CONCLUSIONS
Recombinant PCIneo-hTPO can efficiently be transfected into COS-7 cells and hTPO is an effective cytokine for treating carboplatin-induced thrombocytopenia in mice.
Institute of Hematology, Tongji Medical University, Wuhan.
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D000818:Animals; D016190:Carboplatin; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000096:biosynthesis; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D006801:Humans; D008297:Male; D008533:Megakaryocytes / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics; D013921:Thrombocytopenia / Q000139:chemically induced / Q000628:therapy*; D013926:Thrombopoietin / Q000235:genetics*; D014162:Transfection
D018076:DNA, Complementary; D011994:Recombinant Proteins; D013926:Thrombopoietin; D016190:Carboplatin