Pathogenesis of Murine Gammaherpesvirus-68 Infection in Interleukin-6-Deficient Mice

AbstractMurine gammaherpesvirus-68 (MHV-68) induces high levels of interleukin (IL)-6 production in both naive and primed lymphocyte populations. Mice that are homozygous (−/−) for deletion of the IL-6 gene were used to investigate the role of this cytokine in MHV-68 infection. The results showed that IL-6 is not essential for clearance of infectious MHV-68 from the lung or for the establishment, or control, of viral latency. Both IL-6 +/+ and −/− mice eliminated replicating virus from the respiratory tract within 15 days of infection, and their lungs remained clear of infectious virus for ≥150 days. Interestingly, the IL-6 −/− mice had both increased numbers of natural killer (NK)1.1+ cells and higher levels of NK cell activity than the +/+ controls at 10–15 days after infection. However, there was no difference in the cytotoxic T cell activity between the two groups of mice. Levels of latent virus were comparable in IL-6 +/+ and −/− mice over the time course studied. Furthermore, analysis of the numbers, types, and activation status of the various leukocyte subsets (other than NK cells) in the bronchoalveolar lavage population, lymph nodes, and spleens of +/+ and −/− mice revealed no striking differences. Apart from the expected lack of IL-6, cytokine profiles were not dramatically altered in IL-6 −/− mice. Thus, there is no evidence for an obligatory role for IL-6 in T cell activation during infection with MHV-68

Amphipathic DNA polymers exhibit antiviral activity against systemic Murine Cytomegalovirus infection

<p>Abstract</p> <p>Background</p> <p>Phosphorothioated oligonucleotides (PS-ONs) have a sequence-independent, broad spectrum antiviral activity as amphipathic polymers (APs) and exhibit potent in vitro antiviral activity against a broad spectrum of herpesviruses: HSV-1, HSV-2, HCMV, VZV, EBV, and HHV-6A/B, and in vivo activity in a murine microbiocide model of genital HSV-2 infection. The activity of these agents against animal cytomegalovirus (CMV) infections in vitro and in vivo was therefore investigated.</p> <p>Results</p> <p>In vitro, a 40 mer degenerate AP (REP 9) inhibited both murine CMV (MCMV) and guinea pig CMV (GPCMV) with an IC₅₀of 0.045 μM and 0.16 μM, respectively, and a 40 mer poly C AP (REP 9C) inhibited MCMV with an IC₅₀of 0.05 μM. Addition of REP 9 to plaque assays during the first two hours of infection inhibited 78% of plaque formation whereas addition of REP 9 after 10 hours of infection did not significantly reduce the number of plaques, indicating that REP 9 antiviral activity against MCMV occurs at early times after infection. In a murine model of CMV infection, systemic treatment for 5 days significantly reduced virus replication in the spleens and livers of infected mice compared to saline-treated control mice. REP 9 and REP 9C were administered intraperitoneally for 5 consecutive days at 10 mg/kg, starting 2 days prior to MCMV infection. Splenomegaly was observed in infected mice treated with REP 9 but not in control mice or in REP 9 treated, uninfected mice, consistent with mild CpG-like activity. When REP 9C (which lacks CpG motifs) was compared to REP 9, it exhibited comparable antiviral activity as REP 9 but was not associated with splenomegaly. This suggests that the direct antiviral activity of APs is the predominant therapeutic mechanism <it>in vivo</it>. Moreover, REP 9C, which is acid stable, was effective when administered orally in combination with known permeation enhancers.</p> <p>Conclusion</p> <p>These studies indicate that APs exhibit potent, well tolerated antiviral activity against CMV infection in vivo and represent a new class of broad spectrum anti-herpetic agents.</p

Heart Dysfunction Following Long-Term Murine Cytomegalovirus Infection: Fibrosis, Hypertrophy, and Tachycardia

Human cytomegalovirus (HCMV) is associated with increased risk of chronic diseases of the heart and vasculature, including myocarditis, atherosclerosis, and transplant vasculopathy. To investigate CMV infection of the heart, murine cytomegalovirus (MCMV) was used to evaluate both acute and latent infection and the subsequent phenotypic and functional consequences of infection. Female BALB/c mice were intraperitoneally (i.p.) inoculated with 1 × 10 pfu of MCMV and evaluated at 14 and 50 days postinfection (dpi). At each time point, echocardiography was used to evaluate cardiac function and histology was conducted for phenotypic evaluation. MCMV replication in the heart was detected as early as 3 dpi and was no longer detectable at 14 dpi. Infected animals had significant cardiac pathology at 14 and 50 dpi when compared to uninfected controls. Histology revealed fibrosis of the heart as early as 14 dpi and the presence of white fibrous deposits on the surface of the heart. Functional evaluation showed significantly increased heart rate and muscle thickening in the latently infected animals when compared to the control animals. At 50 dpi, latent virus was measured by explant reactivation assay, demonstrating that MCMV establishes latency and is capable of reactivation from the heart, similar to other tissues such as spleen and salivary glands. Collectively, these studies illustrate that MCMV infection results in phenotypic alterations within the heart as early as 14 dpi, which progress to functional abnormalities during latency. These findings are similar to sinus tachycardia and hypertrophy of the heart muscle observed in cases of HCMV-induced acute myocarditis

Murine Cytomegalovirus IE2, an Activator of Gene Expression, Is Dispensable for Growth and Latency in Mice

AbstractThe murine cytomegalovirus α (immediate-early) gene product, IE2391aa, a protein that is related to the human cytomegalovirus US22 protein family, had previously been shown to be dispensable for viral growth in cell culture. In transient assays, however, this protein was found to transactivate the murine CMV ie1/ie3 and ie2 promoters, as well as a number of other promoters. Transactivation was mediated via promoter-proximal elements rather than through elements located upstream in the enhancer region. This activation predicted that ie2 would play a role in regulating gene expression; however, ie2 mutants did not exhibit altered growth or latency in the mouse. ie2-deficient viruses reached peak titers in spleen, salivary glands, lungs, liver, kidneys, pancreas, peripheral blood leukocytes, and adrenal glands that were comparable to wild-type virus. When assayed by spleen explant culture, ie2-deficient viruses yielded reactivation levels similar to wild type. Thus, the murine CMV ie2 gene encodes a regulatory protein that is dispensable for viral infection of cells in culture as well as for interaction with tissues in the infected BALB/c mouse

Characterization of murine cytomegalovirus infection and induction of calcification in Murine Aortic Vascular Smooth Muscle Cells (MOVAS)

Human cytomegalovirus (HCMV) is a widespread pathogen that causes lifelong latent infection in the majority of the world population. HCMV is associated with increased incidence and severity of many cardiovascular diseases including myocarditis, atherosclerosis, and transplant vasculopathy. Due to the species-restricted nature of cytomegalovirus infection, murine cytomegalovirus (MCMV) is a useful model that recapitulates many of the features of HCMV infection of the cardiovascular system. While in vivo MCMV studies are able to answer many questions regarding pathogenesis of infection, in vitro experiments using cell lines are useful tools to further understand the potential underlying mechanisms. In this study, we characterize MCMV infection of the murine aortic smooth muscle cell line (MOVAS). Our findings demonstrate that MOVAS cells are permissive for MCMV infection, form plaques under carboxymethyl cellulose overlay, and produce progeny virus similar to NIH 3T3 murine embryonic fibroblasts. In addition, MCMV infection induces calcification in MOVAS cells similar to that seen in the epicardium of MCMV-infected hearts. We conclude that MOVAS cells are a useful in vitro tool for studying CMV-mediated cardiac calcification