Mice lacking mature T and B lymphocytes develop arthritic lesions after immunization with type II collagen.

Collagen-induced arthritis in DBA/1 mice is a widely used experimental model of rheumatoid arthritis. The induction phase of the disease is thought to be dependent upon MHC-restricted T and B cell-mediated immune responses to type II collagen, but an influence of additional non-MHC-restricted mechanisms has also been proposed. In this study, we report that type II collagen immunization of DBA/1 mice lacking mature T and B lymphocytes resulted in the development of arthritic lesions, which were characterized by synovial hyperplasia with occasional inflammation as well as cartilage and bone destruction. The specificity of disease induction to type II collagen was confirmed, because arthritis could not be induced when control preparations of OVA or adjuvant alone were administered. A delay in clinical disease onset and a reduction in severity between lymphocyte-positive and -negative DBA/1 mice confirmed that lymphocytes play an important role in disease; however, similar pathologic features and normal incidence suggest that lymphocyte-independent mechanisms of disease induction also operate in the standard collagen-induced arthritis model. We conclude that adaptive immune responses are not the only arthritogenic mechanism and hypothesize that the nonantigenic properties of type II collagen can also lead to arthritis.

Identification of mutations contributing to the reduced virulence of a modified strain of respiratory syncytial virus.

The nucleotide sequences of the genome of the RSS-2 wild type strain of respiratory syncytial (RS) virus, which is known to induce upper respiratory tract infection in adults, and that of the attenuated ts1C candidate vaccine derived from it by three cycles of mutagenesis and selection of temperature-sensitive (ts) mutants, have been determined. Comparison of the sequences has located the genetic changes which contribute to the reduced pathogenicity in adults of the candidate vaccine. Thirty-seven nucleotide changes distinguish the wild type and ts1C, 13 of which confer amino acid substitutions; no mutations are present in extragenic regions. Partial nucleotide sequencing of the genomes of the first stage ts mutant (ts1A) and the second stage ts mutant (ts1B), which were intermediates in the derivation of the third stage mutant ts1C, established that five mutations resulting in amino acid substitutions had been induced in the first cycle of mutagenesis, one in the second cycle, and seven in the third cycle. The unique mutation differentiating ts1B from ts1A substitutes an alanine for a threonine at residue 736 in the polymerase (L) protein. The occurrence of a mutation in ts1C inducing substitution of a phenylalanine for a serine residue at an adjacent site (731) suggests that mutations in this region of the polymerase can have significant attenuating effects. The data suggest also that a mutation in the F gene may contribute to the attenuated phenotype.

Dissection of the pathologies induced by transmembrane and wild-type tumor necrosis factor in transgenic mice.

With increasing awareness that seemingly diverse immune-mediated diseases involve similar pathogenetic mechanisms, and the identification of a growing number of key effector molecules, it is becoming possible to design and generate effective transgenic models for such diseases. Tumor necrosis factor (TNF) plays a prominent role in immune and host defense responses and there is strong evidence that abnormal TNF production contributes to disease initiation and progression in rheumatoid arthritis, systemic inflammatory response syndrome, diabetes, multiple sclerosis, and many other immune-mediated disorders. The generation of TNF transgenic mice, in which TNF production is deregulated, has provided us with direct evidence that, in vivo, this cytokine can indeed trigger the development of such complex disease phenotypes. Transgenic mice that have been engineered to overexpress human or murine TNF molecules in peripheral joints, T cells, or neurons of the central nervous system represent important animal models for human rheumatoid arthritis, systemic inflammation, and multiple sclerosis, respectively. In addition to establishing a central role for TNF in such diseases, these animal models have already proved valuable for identifying additional important disease-effector molecules, and for gaining an insight into the complex in vivo mechanisms that are involved in disease pathogenesis. For example, in the case of arthritis, TNF has been found to transmit its pathogenic effects entirely through interleukin-1, which may therefore represent an additional important target for therapeutic intervention in the human disease. In summary, TNF transgenic models of human disease are expected to serve as important in vivo tools for defining details of disease pathogenesis, potential targets for therapeutic intervention and for evaluating the possible involvement of additional genetic and environmental factors on the disease state.

Transmembrane TNF is sufficient to induce localized tissue toxicity and chronic inflammatory arthritis in transgenic mice.

TNF plays a pivotal role in the pathogenesis of a broad spectrum of infectious, inflammatory, and autoimmune diseases. In addition to the secreted, mature 17 kD form, TNF exists as a bioactive precursor 26 kD transmembrane protein. Transmembrane TNF signaling has been directly associated with specific immune mechanisms, including the contact-dependent lymphocyte and monocyte-mediated cell killing and the CD40 ligand-independent, T cell-mediated polyclonal B cell activation. In previous studies, we have reported that mice expressing 3'-UTR modified human TNF transgenes develop chronic inflammatory polyarthritis with a 100% phenotypic penetrance and timed disease onset. In additional experiments, we have also shown that high-level expression of human TNF in lymphoid cells of transgenic mice results in both local (thymic hypoplasia) and systemic (ischaemia, tissue necrosis, and wasting) TNF-mediated pathology. In this study we show that transgenic mice expressing a T cell-targeted membrane-associated mutant human TNF alpha protein are displaying only local TNF-mediated pathologies, ranging from lymphoid tissue derangements to proliferative synovitis and chronic inflammatory arthritis. These results demonstrate that in vivo, at least part of the pathogenic activities of TNF alpha may be assigned to the functioning of its uncleaved, membrane-associated form.

The type I interleukin-1 receptor acts in series with tumor necrosis factor (TNF) to induce arthritis in TNF-transgenic mice.

The pro-inflammatory cytokines tumor necrosis factor (TNF) and interleukin-1 (IL-1) have been shown to be primary mediators in the pathogenesis of chronic inflammatory joint diseases. However, the relative contributions of these molecules to the development and progression of disease is not known. In the present study, we have investigated the involvement of the type I IL-1 receptor in the development and progression of chronic arthritis in a previously described TNF-transgenic mouse model of this disease. A neutralizing monoclonal antibody to the murine type I IL-1 receptor was injected intraperitoneally three times a week from birth to 9 weeks of age. In the positive control group of untreated transgenic mice the incidence of arthritis was 100% after 9 weeks of age. The injection of normal hamster IgG did not influence the incidence or development of arthritis. In contrast, the injection of antibody to the type I IL-1 receptor completely prevented the development of arthritis. Clinical evaluation of disease was confirmed histologically, anti-receptor antibody-treated mice showed no sign of arthritic change. Moreover, the analysis of sera taken at the end of the study period showed that the neutralization of arthritis by IL-1 receptor antibody treatment was accompanied by significantly lower levels of circulating human TNF compared to the control groups and to untreated transgenic mice prior to disease onset. Taken together, these results show that IL-1 signaling blockade exerts a direct negative feedback effect on TNF production. Our results show that in TNF-transgenic mice the IL-1 receptor accepts the whole pathogenic load of TNF, thereby acting as a potent downstream mediator in the pathogenesis of chronic arthritis.

Variation in the fusion glycoprotein gene of human respiratory syncytial virus subgroup A.

Six different genotypes (designated lineages SHL1-6) of human respiratory syncytial (RS) virus have been defined by partial nucleotide sequence analysis of the variable SH and the hypervariable G membrane protein genes, and by restriction fragment analysis of the conserved N protein gene of viruses isolated in south Birmingham. Viruses of very similar genotype appear to be present worldwide at the present time. We have determined the nucleotide sequences of the fusion protein genes of five viruses isolated in south Birmingham in the same year, but belonging to different lineages, and have compared them with the sequences of four subgroup A viruses isolated at earlier times from diverse localities. The sequence diversity of the F genes of these five viruses, as measured by nucleotide (94.5-98.5%) and inferred amino acid (97.0-99.3%) identifies, is comparable with that of the nine subgroup A viruses considered as a whole. No sequence changes occur in any of the sites of known epitopes. Comparison of the nine subgroup A sequences with the published sequences of a subgroup B strain and three bovine RS viruses confirms that the F protein sequences are most divergent in the F2 region.

Transgenic and knockout analyses of the role of TNF in immune regulation and disease pathogenesis.

Transgenic mutagenesis in whole animals has become without doubt the most rewarding approach to analyse gene structure, expression, and function. In the TNF field, much of what we now question about TNF/TNF receptor function is based, to a large extent, on what we have already learned by overexpressing these molecules in transgenic mice or by ablating their expression in knockout systems. In addition, a clearer view of the involvement of these molecules in disease pathogenesis has emerged, and useful models for human disease have been generated. In this overview, we summarise our experience with TNF transgenic and knockout systems, and highlight advances made in our understanding of the role played by TNF and its receptors in immune regulation and in the pathogenesis of infectious, inflammatory, and autoimmune disease.