Kaposi's sarcoma-associated herpesvirus can productively infect primary human keratinocytes and alter their growth properties.
Previous studies have shown the presence of Kaposi's sarcoma-associated herpesvirus (KSHV/HHV8) DNA in endothelial cells, in keratinocytes in the basal layer of the epidermis overlying plaque-stage nodular lesions of cutaneous Kaposi's sarcoma (KS), and in the epithelial cells of eccrine glands within KS lesions. We infected primary cell cultures of human keratinocytes with KSHV/HHV8. At 6 days post infection, transcription of viral genes was detected by reverse transcriptase PCR (RT-PCR), and protein expression was documented by an immunofluorescence assay with an anti-LANA monoclonal antibody. To determine whether the viral lytic cycle was inducible by chemical treatment, KSHV/HHV8-infected keratinocytes were treated with 12-O-tetradecanoylphorbol-13-acetate (TPA) and RT-PCR was performed to confirm the transcription of lytic genes such as open reading frame 26, (which encodes a capsid protein). Finally, to assess infectious viral production, other primary human cells (human umbilical vein endothelial cells), were infected with concentrated supernatant of KSHV-infected, TPA-induced keratinocytes and the presence of viral transcripts was confirmed by RT-PCR. The uninfected keratinocytes senesced 3 to 5 weeks after mock infection, while the KSHV/HHV8-infected keratinocytes continued to proliferate and to date are still in culture. However, 8 weeks after infection, viral genomes were no longer detectable by nested PCR. Although the previously KSHV/HHV8-infected keratinocytes still expressed epithelial markers, they acquired new characteristics such as contact inhibition loss, telomerase activity, anchorage-independent growth, and changes in cytokine production. These results show that KSHV/HHV8, like other herpesviruses, can infect and replicate in epithelial cells in vitro and suggest that in vivo these cells may play a significant role in the establishment of KSHV/HHV8 infection and viral transmission.
Cerimele
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Department of Microbiology, New York University School of Medicine, New York, New York 10016, USA.
Curreli
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Ely
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Friedman-Kien
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Cesarman
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Flore
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D000956:Antigens, Viral / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis; D016228:Endothelial Growth Factors / Q000235:genetics; D016222:Fibroblast Growth Factor 2 / Q000235:genetics; D006566:Herpesviridae Infections / Q000821:virology*; D019288:Herpesvirus 8, Human / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D015603:Keratinocytes / Q000502:physiology / Q000821:virology*; D008222:Lymphokines / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010750:Phosphoproteins; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D014779:Virus Replication
D000956:Antigens, Viral; D016207:Cytokines; D016228:Endothelial Growth Factors; D008222:Lymphokines; D009687:Nuclear Proteins; D010750:Phosphoproteins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; C118485:latent nuclear antigen (LNA); D016222:Fibroblast Growth Factor 2