Immunosuppression induces transcription of murine cytomegalovirus glycoprotein H in the eye and at non-ocular sites.
In these studies, DNA PCR was used to identify sites of murine cytomegalovirus (MCMV) latency after inoculation of virus into the supraciliary space of the eye. Reverse transcription (RT) PCR for an immediate early gene and a late gene was used to identify putative sites of virus reactivation after methylprednisolone (steroid)-induced immunosuppression. Ten weeks after inoculation of 5 x 10(2) PFU of MCMV, BALB/c mice were immunosuppressed by intramuscular injection of steroid. Control mice were infected but not immunosuppressed. Two weeks after initiation of immunosuppression, mice were sacrificed. DNA and RNA extracted from homogenized tissues were amplified for immediate early gene 1 (IE1) and late gene, glycoprotein H (gH), DNA and mRNA by PCR and RT-PCR, respectively. Replicating virus was detected in homogenized ocular and non-ocular tissues by plaque assay. In the latently infected PBS-treated control group, viral DNA was detected in the inoculated eye and in several non-ocular tissues; IE1 mRNA was found in most of the DNA-positive tissues, while gH mRNA was amplified only in a few of the MCMV DNA-positive tissues from a single mouse. After immunosuppression, viral DNA and IE1 mRNA were detected at a higher frequency in various tissues of steroid-treated mice. gH mRNA was detected in a significantly higher number of the inoculated eyes, salivary glands and other non-ocular tissues of steroid-treated mice. After immunosuppression, low titers of infectious virus were recovered from the salivary glands of steroid-treated mice, but infectious virus was not recovered from the inoculated eye of either steroid-treated of non-immunosuppressed mice. The DNA PCR results suggest that after inoculation of 5 x 10(2) PFU of MCMV into the supraciliary space of euthymic BALB/c mice, virus becomes latent in the inoculated eye, salivary gland and other extraocular tissues. The RT-PCR results suggest that latent MCMV can be reactivated in multiple tissues by immunosuppression.
Duan
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Y
Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, USA.
Atherton
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SS
D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis; D005123:Eye / Q000821:virology; D005260:Female; D006801:Humans; D017874:Immediate-Early Proteins / Q000096:biosynthesis / Q000235:genetics; D007165:Immunosuppression Therapy; D008775:Methylprednisolone / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D018146:Muromegalovirus / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D012367:RNA, Viral / Q000032:analysis; D051381:Rats; D014158:Transcription, Genetic; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000235:genetics; D014764:Viral Proteins; D014775:Virus Activation / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D017735:Virus Latency
D017931:DNA Primers; D004279:DNA, Viral; C088645:IE1 protein, cytomegalovirus; D017874:Immediate-Early Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C071440:glycoprotein H, Cytomegalovirus; C499358:glycoprotein H, Human cytomegalovirus; D008775:Methylprednisolone