Janet K. Chantler

Persistent Rubella Virus Infection Associated with Chronic Arthritis in Children

Abstract We isolated rubella virus from lymphoreticular cells in 7 of 19 children with chronic rheumatic disease, including patients with systemic-onset juvenile rheumatoid arthritis (Stills disease) (1 of 5), polyarticular juvenile rheumatoid arthritis (2 of 2), pauciarticular juvenile rheumatoid arthritis (2 of 6), and seronegative spondyloarthritis (2 of 6). In contrast, rubella virus was not isolated from the control group, which included eight normal subjects and eight patients with other connective tissue diseases or traumatic joint effusion. In most members of the study group, mononuclear cells from both synovial fluid and peripheral blood were examined. Rubella virus was isolated from both cell populations in three patients, from only peripheral blood in one, and from only synovial fluid in two. In the children with systemic-onset juvenile rheumatoid arthritis, only peripheral blood was examined, and of the five samples analyzed, one was shown to have rubella virus. Virus was isolated on more tha...

PERSISTENT RUBELLA INFECTION AND RUBELLA-ASSOCIATED ARTHRITIS

Rubella virus has been isolated from peripheral blood lymphocytes in six out of seven women with rubella-associated arthritis. The arthritis occurred after natural infection (one case) or immunisation with HPV 77 DES vaccine (six cases) and had been present for up to 6 years. The identification of rubella virus was confirmed by plaque/microfocus reduction in the presence of anti-rubella antiserum, and by immunoprecipitation of rubella virus antigens and their analysis on polyacrylamide gels. These women with rubella virus in their lymphocytes did not have abnormal serum antibody levels, but a standard lymphoproliferative assay showed that they had strong cell-mediated immune responses to rubella virus antigens.

Replication and expression of rubella virus in human lymphocyte populations.

Human mononuclear cells (lymphocytes and monocytes) from peripheral blood were examined for their ability to support the replication of rubella virus (RV) after infection in vitro. Replication was shown to occur in mixed lymphocytes and to be enhanced by stimulation with phytohaemagglutinin or pokeweek mitogen. A comparison of RV polypeptide synthesis in lymphocytes and RK13 cells showed no major differences in the polypeptides induced by infection. However, cellular translation was inhibited in the lymphocytes facilitating identification of virus polypeptides and eliminating the need for hypertonic labelling conditions used with RK13 cells. RV replication was also shown to occur in sub-populations of T-cells but not in B-cells. However, RV could be rescued from the B and monocyte population by co-cultivation with RK13 cells.

Proteins of murine cytomegalovirus: Identification of structural and nonstructural antigens in infected cells

Abstract The synthesis of viral proteins in tertiary mouse embryo cells infected with murine cytomegalovirus (MCMV) has been studied by polyacrylamide gel electrophoresis. Three immediate-early proteins were detected by 4 hr postinfection (p.i.), but between 4 hr and the onset of viral DNA synthesis at 8–10 hr, no further viral proteins could be distinguished. The major structural proteins appeared after viral DNA synthesis had commenced and continued to be made until 36 hr p.i. or later. Host translation occurred until 24 hr p.i. but was inhibited between then and 30 hr p.i. at which time viral protein synthesis predominated. At this time 52 proteins could be labeled in infected cells and, of these, 30 could be precipitated with viral-specific antisera. The proteins precipitated included 22 with electrophoretic mobilities similar to structural viral proteins and a further 8 which were not present in purified MCMV. These, together with the three immediate-early proteins, were the only nonstructural proteins which were detected in the infected cell.

Vertical transmission of murine cytomegalovirus.

Congenital abnormalities induced by murine cytomegalovirus have previously been suggested to be an indirect effect of maternal illness as no infectious virus has been isolated from the foetus. However, in this article we report that latent virus detectable by immunofluorescence and in situ hybridization, may frequently be present in foetal tissues.

Congenital rubella syndrome associated with calcific epiphyseal stippling and peroxisomal dysfunction

An infant girl had the clinical and immunologic findings of congenital rubella syndrome but also had arthrogryposis multiplex and calcific epiphyseal stippling. Spastic quadriparesis developed, and both physical and behavioral development were slow. Increased spasticity of the legs at 5 1/2 years was related not to progressive rubella encephalomyelopathy but to spinal cord compression by abnormal cartilaginous tissue. The presence of a peroxisomal disorder was demonstrated by a greatly increased level of phytanic acid and slightly increased levels of hexacosanoate in serum and by reduced activity of peroxisomal dihydroxyacetone phosphate acyltransferase and a slightly increased ratio of cytosolic to peroxisomal catalase activity in cultured fibroblasts. A reduction in the number and size of peroxisomes was demonstrated in cultured fibroblasts, and a needle biopsy specimen of the liver also showed the peroxisomes to have a smaller diameter than usual. We recommend that any child with epiphyseal stippling be assessed for peroxisomal disease and that the potential for spinal cord compression by dysplastic bone or cartilage be recognized. The association of peroxisomal dysfunction with congenital rubella has not been described previously. The interaction between rubella virus infection and peroxisomal function may need further investigation.

Rubella virus: Intracellular polypeptide synthesis

Abstract Rubella virus (RV) grown in RK13 cells induces a range of polypeptides ranging in molecular weights from 109K to 28K. As RV does not inhibit host metabolism, detection of these requires the use of elevated concentrations of NaCL in the labelling medium to selectively inhibit cellular translation. Several of the viral polypeptides, including p109, p92, and p72 are transient high molecular weight species which may represent precursors of functional viral polypeptides. Only five or six of the polypeptides are synthesized in large amounts at late stages of infection and these probably include the structural components.

Mapping of Genetic Determinants of Rubella Virus Associated with Growth in Joint Tissue

ABSTRACT Rubella virus (RV) strains vary in their abilities to replicate and persist in cell cultures derived from human joint tissue (synovial cells [SC]), and this arthrotropism appears to be linked to their association with joint symptoms in vivo. In order to map the genetic determinants of arthrotropism, an infectious clone of the Cendehill vaccine strain of RV was constructed, as well as two chimeric clones containing cDNAs from both Cendehill and Therien (wild-type) strains. Replacement of the entire structural gene region of Therien in the infectious clone pROBO302 with the corresponding region of Cendehill did not affect growth in SC. A further observation that Cendehill bound equally well to SC and the permissive Vero cell line indicated that restriction was not at the level of receptor binding, a function of the envelope proteins. Mutations that affected growth in joint cells were mapped to two locations in the nonstructural gene region. The first of these (nucleotides 2803 and 6416) resulted in a 10-fold decrease in yield of progeny virus from SC. This region contained five mutations, at nucleotides 2829, 3060, 3164, and 3528 (near the carboxy terminus of P150 where the protease domain is located) and at nucleotide 4350 in p90. Further substitution of the sequence representing nucleotides 1 to 2803 to give a complete Cendehill infectious clone restricted growth in SC by a further 100-fold to less than 10 PFU/ml. This region contains three mutations, at nucleotides 34, 37, and 55, within the 5′ stem-loop structure. In conclusion, the Cendehill-specific mutations believed to be determinants of joint cell growth are located in two regions, the 5′ nontranslated region and in a sequence that encodes the carboxy-terminal region of p150 extending into the helicase domain of p90.

Murine cytomegalovirus gene expression during nonproductive infection in go-phase 3T3 cells

Abstract Murine 3T3 cells, maintained in the Go-phase by deprivation of serum, were infected with murine cytomegalovirus (MCMV) and examined for the synthesis of viral DNA, RNA, and proteins. No viral DNA replication could be detected, although the viral genome remained viable since infectious centers persisted. Viral RNA was analyzed by reassociation kinetics with 125 I-MCMV DNA. At early and late times after infection, the percentage of viral DNA transcribed amounted to 18% and 21%, respectively (comprising one abundance class), in contrast to 26% and 38%, respectively (two abundance classes), at corresponding times during productive infection in growing 3T3 cells. Five viral polypeptides were detected in the Go-phase cells, but these did not include structural polypeptides. When infected Go-phase cells were exposed to serum and fresh medium, viral DNA synthesis occurred, and progeny virions appeared subsequently. Thus this system may prove useful for analyzing a latent herpes infection in vitro .

The use of hypertonicity to selectively inhibit host translation in murine cytomegalovirus-infected cells

Abstract Elevation of the salt concentration in the medium of MCMV-infected cells causes selective inhibition of cellular relative to viral mRNA translation. This has been used to identify a total of eight early proteins in MCMV-infected cells.