[In vitro infection of human megakaryocyte precursors by human cytomegalovirus (HCMV) and the antiviral effect of HCMV antisense oligonucleotides].
OBJECTIVE
To explore the suppression effect of human cytomegalovirus (HCMV) on megakaryocytes and their precursors and study the antiviral effect of antisense phosphorothioate deoxyoligonucleotide (ASON) against HCMV.


METHODS
CD34(+) cells were induced to proliferate and differentiate committedly to megakaryocytes in a semi-solid CFU-MK culture system. Cultured cells and ASON pretreated CD34(+) cells were infected by HCMV of AD169 strain. HCMV immediate early protein (IEP) DNA and mRNA and UL36 mRNA were detected by PCR and reverse transcription-polymerase chain reaction (RT-PCR). Cytotoxicity was evaluated by MTT assay.


RESULTS
HCMV AD169 suppressed the proliferation of megakaryocytes significantly. Compared with the mock group, the CFU-MK yields were decreased by 21.6%, 33.8%, and 46.3%, respectively, in 3 different titers of virus infected groups (P < 0.05). The suppression was virus titer dependent. HCMV IEP DNA, HCMV IEP mRNA and UL36 mRNA were detected in the colony cells of viral infection group. Compared with the infected group by HCMV AD169, UL36Anti treatment at 0.08 micromol/L could recover the CFU-MK yields significantly (P < 0.05). In the infected MK, which was pretreated with UL36Anti at 0.08 micromol/L, HCMV UL36 mRNA was undetectable by RT-PCR. The oligonucleotide MM(1) containing a G-to-C substitution in UL36Anti was inactive at 0.08 micromol/L but active at 0.40 micromol/L. The concentration of UL36Anti necessary to significantly affect cell growth was 90.00 micromol/L.


CONCLUSIONS
HCMV AD169 infection inhibits the proliferation and differentiation of megakaryocytes and their precursors. There are early transcriptions of HCMV IE and UL36 protein in infected CFU-MK. The specific ASON has a definite anti-HCMV activity.
Yao
Jun-xia
JX
Institute of Hematology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Cui
Guo-hui
GH
Xia
Ling-hui
LH
Song
Shan-jun
SJ
D000956:Antigens, Viral / Q000235:genetics; D000998:Antiviral Agents / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics* / Q000502:physiology; D005312:Fetal Blood / Q000166:cytology; D054884:Host-Pathogen Interactions; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D007231:Infant, Newborn; D055016:Megakaryocyte Progenitor Cells / Q000166:cytology / Q000187:drug effects* / Q000821:virology; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000956:Antigens, Viral; D000998:Antiviral Agents; D017874:Immediate-Early Proteins; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; C044582:immediate-early proteins, cytomegalovirus