An association and haplotype analysis of porcine maternal infanticide: a model for human puerperal psychosis?

An association analysis using the Illumina porcine SNP60 beadchip was performed to identify SNPs significantly associated with porcine maternal infanticide. We previously hypothesised that this was a good animal model for human puerperal psychosis, an extreme form of postnatal mood disorder. Animals were selected from carefully phenotyped unrelated infanticide and control groups (representing extremes of the phenotypic spectrum), from four different lines. Permutation and sliding window analyses and an analysis to see which haplotypes were in linkage disequilibrium (LD) were compared to identify concordant regions. Across all analyses, intervals on SSCs 1, 3, 4, 10, and 13 were constant, contained genes associated with psychiatric or neurological disorders and were significant in multiple lines. The strongest (near GWS) consistent candidate region across all analyses and all breeds was the one located on SSC3 with one peak at 23.4 Mb, syntenic to a candidate region for bipolar disorder and another at 31.9 Mb, syntenic to a candidate region for human puerperal psychosis (16p13). From the haplotype/LD analysis, two regions reached genome wide significance (GWS): the first on SSC4 (KHDRBS3 to FAM135B), which was significant (-logP 5.57) in one Duroc based breed and is syntenic to a region in humans associated with cognition and neurotism; the second on SSC15, which was significant (-log10P 5.68) in two breeds and contained PAX3, which is expressed in the brain.

Analysis of X chromosome genomic DNA sequence copy number variation associated with premature ovarian failure (POF).

BACKGROUND: Premature ovarian failure (POF) is a heterogeneous disease defined as amenorrhoea for >6 months before age 40, with an FSH serum level >40 mIU/ml (menopausal levels). While there is a strong genetic association with POF, familial studies have also indicated that idiopathic POF may also be genetically linked. Conventional cytogenetic analyses have identified regions of the X chromosome that are strongly associated with ovarian function, as well as several POF candidate genes. Cryptic chromosome abnormalities that have been missed might be detected by array comparative genomic hybridization. METHODS: In this study, samples from 42 idiopathic POF patients were subjected to a complete end-to-end X/Y chromosome tiling path array to achieve a detailed copy number variation (CNV) analysis of X chromosome involvement in POF. The arrays also contained a 1 Mb autosomal tiling path as a reference control. Quantitative PCR for selected genes contained within the CNVs was used to confirm the majority of the changes detected. The expression pattern of some of these genes in human tissue RNA was examined by reverse transcription (RT)-PCR. RESULTS: A number of CNVs were identified on both Xp and Xq, with several being shared among the POF cases. Some CNVs fall within known polymorphic CNV regions, and others span previously identified POF candidate regions and genes. CONCLUSIONS: The new data reported in this study reveal further discrete X chromosome intervals not previously associated with the disease and therefore implicate new clusters of candidate genes. Further studies will be required to elucidate their involvement in POF.

Stromelysin, gelatinase A and TIMP-1 in prosthetic interface tissue: a role for macrophages in tissue remodelling.

Aseptic loosening of prosthetic components is the most important long-term complication of total joint replacement. To investigate the underlying destructive mechanisms, periprosthetic tissues from both well-fixed and loosened sites from six patients, undergoing surgery for aseptic loosening of knee or hip prostheses, were analysed in detail by immunohistochemical methods for the presence of matrix metalloproteinases and tissue inhibitor of metalloproteinases-1 (TIMP-1). The tissues contained small numbers of cells positive for either collagenase, stromelysin, gelatinase A or TIMP-1; these were randomly distributed, neither specifically next to the bone interface nor to wear particles, and the number of positive cells did not correlate with macroscopic observations at operation. Gelatinase A was co-localized in cells with prolyl-4-hydroxylase, an enzyme involved in collagen synthesis. The predominant cell type in these tissues was shown to be the macrophage by the use of cell marker antibodies. Dual localization was not technically possible but the results strongly suggest that monocyte/macrophages were the primary source of gelatinase A and TIMP-1. Stromelysin was immunolocalized on connective tissue matrix in four patients, and gelatinase A in one patient, and were also observed in tissues in which there was no evidence of cellular synthesis of these enzymes. This suggests that secretion had taken place previously, resulting in enzyme bound to matrix for some time. Taken together, these data indicate that localized focal connective tissue remodelling occurs in periprosthetic tissues from both well fixed and loosened sites.

Immunolocalisation studies on six matrix metalloproteinases and their inhibitors, TIMP-1 and TIMP-2, in synovia from patients with osteo- and rheumatoid arthritis.

OBJECTIVE: To assess the likely importance of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) in the arthritic process. METHODS: Synovial samples from seven joints with rheumatoid arthritis and three osteoarthritic joints were analysed by indirect immunofluorescence microscopy. Using specific human antisera, we documented the frequencies and distributions of collagenase, stromelysins 1 and 2, matrilysin, gelatinases A and B, TIMP-1, and TIMP-2. RESULTS: Stromelysin 1 was found in all synovia, bound to extracellular matrix, within cells, or both, indicating stromelysin synthesis. Matrilysin was present in only one active inflammatory synovium, and focal synthesis of collagenase and gelatinase A was seen in four synovia. Stromelysin 2 and TIMP-2 were not observed, but TIMP-1 synthesis was seen in five synovia, and in two active synovia the distribution of TIMP-1 positive cells was more widespread than that of MMPs. CONCLUSIONS: The presence of stromelysin 1 in all synovia clearly implicates this enzyme in joint damage. Collagenase, gelatinase A and matrilysin may also have a role in rheumatoid arthritis, but are not significant in osteoarthritis. However, marked regional variations were found in the synthesis of these MMPs, indicating not only that these diseases are episodic but that control of enzyme synthesis is focal. Only TIMP-1 may be considered an inhibitory factor.

Metalloproteinase production by rabbit articular cartilage: comparison of the effects of interleukin-1 alpha in vitro and in vivo.

To assess the effects of interleukin-1 on intact To assess the effects of interleukin-1 on intact articular cartilage in vitro, explants from young and adult rabbits were cultured with interleukin-1 and the distributions of the matrix metalloproteinases and tissue inhibitor of metalloproteinases (TIMP-1) were investigated by indirect immunofluorescence microscopy. One to 2-week-old cartilage chondrocytes synthesized collagenase in response to pure or crude interleukin-1 (monocyte conditioned medium), with subarticular cells most responsive. Collagenase synthesis was not stimulated in adult articular chondrocytes when explants were treated with either pure or crude interleukin-1. Stromelysin, gelatinase and TIMP-1 could not be demonstrated within any zone of the cartilage, indicating that their synthesis was not stimulated by either pure or crude interleukin-1. The addition of fibroblast growth factors, either alone or in combination with interleukin-1, did not modify these responses. These results contrast markedly with observations on cultured chondrocyte monolayers, where interleukin-1 treatment induces near co-ordinate expression of metalloproteinases. To assess the effects of interleukin-1 in vivo, it was injected into adult rabbit knee joint spaces and the articular cartilage subsequently analysed for evidence of altered metalloproteinase production by immunocytochemistry. No significant increase in metalloproteinase or TIMP-1 synthesis by chondrocytes was detected, although the cartilage matrix showed a marked loss of toluidine blue metachromasia. We conclude that metalloproteinases are not involved in the rapid loss of proteoglycan from cartilage matrix in these situations.

Rabbit models of arthritis: immunolocalization of matrix metalloproteinases and tissue inhibitor of metalloproteinase in synovium and cartilage.

The distribution of the matrix metalloproteinases, collagenase, stromelysin, gelatinases A and B, and the tissue inhibitor of metalloproteinases in cartilage and synovium removed from rabbits up to 27 days after induction of two models of arthritis was investigated by immunolocalization. Following intra-articular injection of poly-D-lysine/hyaluronic acid coacervate, collagenase and stromelysin were found bound to cartilage matrix, but there was little increase in chondrocyte synthesis of these enzymes. The synovium underwent a complex wound healing response involving invagination and encapsulation of the coacervate and inflammatory cell debris, during which all four metalloproteinases and tissue inhibitor of metalloproteinase could be immunolocalized. The second model, intra-articular injection of ovalbumin into sensitized rabbits, caused considerable chondrocyte necrosis; collagenase was found bound to cartilage matrix on day 13, although again there was little evidence of synthesis by chondrocytes. Inflammatory cell infiltration of meniscoid synovia took place initially, followed by fibrosis involving macrophagelike cells secreting gelatinase A. In both models there was rapid loss of glycosaminoglycan metachromasia from the cartilage matrix. These results are discussed in relation to current knowledge of metalloproteinase involvement in the chronic rheumatoid synovial pannus erosion of cartilage in humans. The data suggest that there are considerable differences between rheumatoid arthritis and these models, and their use must therefore be carefully defined.

The degradation of collagen in pig synovium in vitro and the effect of colchicine.

Colchicine induced a rapid destruction of the collagenous matrix of pig synovial explants in culture in the presence of serum. The most efficacious doses were 0.01-0.1 micrograms/ml (2.5 x 10(-8) M - 2.5 x 10(-7) M). The histological progression of the tissue breakdown induced by colchicine was very similar, although faster, to that described for other agents (Fell et al., 1986), with cells having basophilic nuclei accumulating in areas of fibril degradation. The loss of collagen correlated with an increase in collagenase production and at the peak of resorption (6 to 8 days) active collagenase was present in the culture media. Immunocytochemical methods demonstrated active collagenase bound to collagen fibrils after only 4 days in culture, before significant collagen degradation could be observed histologically. Collagen breakdown was completely inhibited by cortisol, and partially inhibited by indomethacin: only the inhibition by indomethacin could be reversed by exogenous prostaglandin E2. Vinblastine at a higher dose was as effective as colchicine but the lumicolchicines, which do not disrupt microtubules, were ineffective. Although the precise mechanism of action of colchicine is unknown, this culture system provides a useful in vitro model for increasing our understanding of the cellular mechanisms of tissue breakdown and for elucidating the roles of active collagenase and related metalloproteinases.

The promotion and inhibition of collagen-breakdown in organ cultures of pig synovium: the requirement for serum components and the involvement of cyclic adenosine 3':5'-monophosphate (cAMP).

In many pathological situations connective tissue cells acquire the ability to degrade the macromolecular components of their extracellular matrix. To study the destruction of collagen we used organ cultures of porcine synovial tissue. In the presence of 15% rabbit serum explants shrink considerably during 10-14 days, owing to early loss of interfibrillar material followed by retraction and local destruction of collagen fibres, partly by phagocytosis. These changes, and the release of latent collagenase into the medium, are largely inhibited by cortisol and partially by indomethacin. Collagen destruction can be greatly accelerated by the addition to the culture medium of one of the following: sodium fluoride, 3-isobutyl-1-methylxanthine, dibutyryl cyclic adenosine 3':5'-monophosphate or forskolin; these agents are known to affect cyclic adenosine monophosphate metabolism and our results suggest strongly that a change in the intracellular levels of cyclic adenosine monophosphate is a key-step in the process leading to the increased catabolism of collagen. With these compounds the destruction of collagen is largely extracellular; the histological changes and the increased levels of collagenase associated with the destruction can be prevented by cortisol and, except in the case of dibutyryl cyclic adenosine monophosphate, at least partially by indomethacin. Without serum only 3-isobutyl-1-methylxanthine sometimes causes drastic breakdown of collagen. This model system should be of great benefit in exploring the mechanisms involved in collagen destruction.