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Biomedical subjects

S Gay

Publications and source records attributed to S Gay.

At least 109 records · Page 6Linked to original sources
PubMed1999

Analysis of the p53 tumor suppressor gene in rheumatoid arthritis synovial fibroblasts.

OBJECTIVE: To determine whether mutations in the tumor suppressor gene p53 may contribute to the transformed-appearing phenotype of rheumatoid arthritis (RA) synovial fibroblasts. METHODS: We performed p53 gene mutation analysis using different molecular approaches. Synovial fibroblasts of 10 patients with RA were cultured and RNA and DNA were harvested after 3-5 passages in cell culture. Sequence analysis of all exons of the p53 gene was performed using 3 different techniques: 1) single-strand conformational polymorphism, 2) nonisotopic RNase cleavage assay, and 3) base excision sequence scanning T-scan, followed by sequence analysis of specific gene segments. RESULTS: Although p53 antigen could be detected by immunocytochemistry in numerous cultured fibroblasts, gel electrophoresis analysis of products obtained using all 3 methods and subsequent sequence analysis showed no specific mutation pattern in the genome of the synovial fibroblasts from patients in Germany, including the known "hot spots" within the p53 genome. However, p53 mutations were identified in different clones of 3 additional RA synovial fibroblast populations from the United States. Sequence analysis of the p53 promoter did not reveal mutational base changes. CONCLUSION: The findings of the study support the hypothesis that the majority of the mutations of the p53 gene observed in RA synovium are not derived from the genome of RA synovial fibroblasts, and that the variability of the mutation pattern reflects, in part, the heterogeneity of the disease.

Arthritis, Rheumatoid↗
PubMed1999

Expression of RANTES in biopsies of skin and upper gastrointestinal tract from patients with systemic sclerosis.

Inflammatory infiltrates and upregulated collagen production are hallmarks of systemic sclerosis (SSc). There are indications that chemokines are involved in accumulation of inflammatory and matrix-synthesizing cells in SSc skin lesions. Therefore, we searched for the expression and localization of the chemokine RANTES ("regulated upon activation and normal T cells expressed and secreted") in skin and esophageal biopsies from patients with SSc. Using immunohistochemistry and in situ hybridization, skin biopsies derived from clinically involved and noninvolved skin of 18 patients with early and long-term SSc were examined for RANTES expression and compared with nondiseased skin sections of seven patients without SSc. In addition, esophageal snap biopsies were taken in a subgroup of six SSc patients. Strong expression of RANTES could be detected in the epidermis in keratinocytes of patients with short-term and long-term disease, both on the mRNA and protein level. The percentage of RANTES-expressing cells were significantly higher in clinically noninvolved skin sections than in involved skin areas. In contrast, no RANTES expression was found in esophageal biopsies or in the control group. The results indicate that RANTES is present in human sclerodermatous skin. RANTES may be involved in early pathogenesis of SSc as well as in fibrosis pathways, either by chemoattraction of immunocompetent cells and/or by modulation of collagen production.

Adult↗
PubMed1999

Prolactin and prolactin-like polypeptides in rheumatoid arthritis.

A bidirectional communication network exists between the neuroendocrine and immune systems, and a dysfunctional communication may contribute to the development of autoimmune diseases in various species, including humans. Experimental, epidemiological, and clinical data suggest that breast feeding and hyperprolactinemia constitute a risk factor for the development of diseases with autoimmune components, including rheumatoid arthritis (RA). We hypothesized that the anterior pituitary hormone prolactin (Prl) and locally produced Prl-like polypeptides may act as endocrine, autocrine, and paracrine regulators of synovial cell functions. They may participate not only in enhancing T-lymphocyte immune reactivity, but also in the exacerbation of RA lesions through their influence on synovial fibroblasts. In RA synovial tissue, Prl-like polypeptides could participate in a bidirectional communication between immunocytes and fibroblasts. Both Prl and Prl-like polypeptides might act via proto-oncogenes and transcriptional factors, leading to cell proliferation, i.e., synovial tissue hyperplasia, neo-angiogenesis, and the production of catabolic enzymes such as matrix metalloproteinases and cathepsins. In such cases, they could represent important regulators of the T-cell independent mechanism of joint destruction.

Animals↗
PubMed1999

Assessment of resident knowledge: subjective assessment versus performance on the ACR in-training examination.

RATIONALE AND OBJECTIVES: The authors assessed the ability of faculty and residents to predict the ranked performance of residents on the American College of Radiology (ACR) In-Training Examination. MATERIALS AND METHODS: Radiology faculty at Penn State Geisinger Health System (PSGHS), the Medical College of Virginia (MCV), and the University of Virginia (UVA) and residents at PSGHS and MCV ranked the expected performances of residents taking the 1997 ACR In-Training Examination. Surveyed faculty and residents were blinded to the actual performances on the examination. Forty-nine residents took the examination (21 at PSGHS, 22 at MCV, six at UVA), and 37 faculty members (11 at PSGHS, 11 at MCV, 15 at UVA) participated in the study. Correlation analysis was performed to assess the agreement between the subjective and actual ranking of residents in each residency class. RESULTS: Faculty were moderately accurate in the overall ranking of resident performances (r = 0.34). High levels of concordance for ranking individual residents correlated with accuracy in only certain cases. Differences in agreement and accuracy of the respondents existed between PSGHS and MCV (P = .0001 and .0014, respectively). The concordance of respondents increased significantly from the 1st- to the 2nd-year class at MCV (P = .0002), whereas accuracy increased significantly between these classes for the PSGHS (P = .042). CONCLUSION: Faculty are only moderately successful in ranking resident performances on the ACR In-Training Examination, and a high level of agreement is not necessarily indicative of increased accuracy. The concordance and accuracy of subjective rankings differ among residency programs and classes.

Clinical Competence↗
PubMed1999

Anticytokines.

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Animals↗
PubMed1999

Collagenase, cathepsin B and cathepsin L gene expression in the synovial membrane of patients with early inflammatory arthritis.

OBJECTIVE: To examine the expression of the matrix metalloproteinase, MMP-1, and the cysteine proteases, cathepsin B (CB) and cathepsin L (CL), in the synovial membrane (SM) of patients with early inflammatory arthritis. METHODS: Samples of SM were obtained by blind needle biopsy or needle arthroscopy from inflamed knees of 28 patients with early inflammatory arthritis (mean disease duration 10.2 months, range 2 weeks-18 months). Sixteen patients had rheumatoid arthritis (RA), nine psoriatic arthritis and there was one each with ankylosing spondylitis, gout and an undifferentiated arthritis. Comparison was made with tissue from two patients with established erosive RA and three normal synovial tissue samples. In situ hybridization was performed using digoxigenin-labelled RNA probes. RESULTS: MMP-1, CB and CL were expressed in all patients with early arthritis and in established erosive RA, whereas normal synovium showed only scanty expression. The three proteases were prominent in perivascular infiltrates and endothelial cells of early arthritis tissue. MMP-1 was observed primarily in the lining layer, but was also evident in the sublining area. CB and CL were expressed to a lesser extent in the lining layer, and were present mainly in the subintima. The three proteases were not found in lymphoid aggregrates. No differences were observed between the disease categories. CONCLUSIONS: The detection of MMP-1, CB and CL in the synovium shortly after symptom onset implies that the potential for joint destruction exists at a very early stage in the disease. In addition, the perivascular and endothelial cell expression suggests a role for these proteases in mononuclear cell influx to the inflamed synovium and in angiogenesis.

Adult↗
PubMed1999

MIA (melanoma inhibitory activity): a potential serum marker for rheumatoid arthritis.

OBJECTIVE: MIA (melanoma inhibitory activity) is correlated with metastasis in patients with malignant melanoma. As MIA is not only produced by melanoma cells, but also by differentiated chondrocytes, we examined whether serum levels of MIA are correlated with inflammation and/or joint destruction in rheumatic diseases. METHODS: MIA serum concentrations of patients with different rheumatic diseases were examined and compared with healthy individuals and malignant melanoma patients. In addition, MIA concentrations were correlated to inflammatory parameters and joint destruction. RESULTS: Increased MIA serum concentrations were found only in patients with rheumatic diseases associated with joint destruction, such as rheumatoid arthritis (RA), osteoarthritis, HLA B27-associated oligoarthritis, and psoriatic arthritis. Of these rheumatic diseases, a significant increase in MIA serum concentrations was seen only in patients with RA, associated with rheumatoid factor (RF) positivity and joint destruction. CONCLUSIONS: In addition to RF, MIA might therefore be useful in the differential diagnosis of RA vs non-destructive rheumatic diseases, and the presence of elevated levels of MIA in serum very likely reflects joint destruction in RA.

Arthritis, Rheumatoid↗
PubMed1999

Rheumatoid arthritis.

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Anti-Inflammatory Agents↗
PubMed1999

Signaling and effector pathways.

There is increasing evidence that distinct signaling and effector pathways in the rheumatoid synovium result in a cascade of pathophysiologic events. These interactions, which finally lead to progressive joint destruction, are different from all other joint diseases in numerous aspects. As outlined in this review, molecular biology techniques allow detection of key pathways ranging from external stimuli to subcellular gene regulation mechanisms operative in various cells within the rheumatoid synovium. To alter these pathways, inhibitory factors need to be applied to these "hot zones" for an extended period, which can be achieved either by repeated drug administration or by local synthesis using genetically altered synovial cells. Both adenovirus and retroviral constructs, as well as ex vivo and in vivo strategies, can be used for gene transfer into these cells, and routine delivery of "protective" genes into the affected joints might be achieved within the next decade.

Arthritis, Rheumatoid↗
PubMed1999

Membrane-type-1 matrix metalloproteinase is abundantly expressed in fibroblasts and osteoclasts at the bone-implant interface of aseptically loosened joint arthroplasties in situ.

OBJECTIVE: To investigate the distribution pattern of membrane-type-1 matrix metalloproteinase (MT1-MMP) within the synovial-like interface membranes of failed prosthetic joints. METHODS: Interface tissue around loose arthroplasties containing both fibrous membrane and attached bone was obtained from 6 patients at revision surgery. In situ hybridization with digoxigenin labeled RNA probes was applied to investigate MT1-MMP expression in paraffin sections of the samples. In addition, double labeling using immunohistochemistry was performed to characterize MT1-MMP producing cells. RESULTS: Apart from being present in fibroblasts, MT1-MMP was also found expressed in osteoclasts at sites of bone resorption. Our results revealed no expression of MT1-MMP at parts of the membrane that originally had been located next to the prosthesis. In contrast, abundant staining for MT1-MMP was observed at sites attached to bone. MT1-MMP mRNA expression was more intense at those sites of bone resorption covered by a thicker interface membrane. CONCLUSION: These results indicate a role for MT1-MMP not only in matrix degradation by fibroblasts but also in osteoclast mediated bone resorption. Given the ability of MT1-MMP to activate MMP2 and MMP13, they suggest also that osteoclasts might contribute to matrix degradation by activating these MMP. This could be of potential interest not only for other conditions in which bone resorption by fibroproliferative tissue plays a role, but also to design novel strategies to prevent loosening of prosthetic joints.

Arthroplasty, Replacement↗
PubMed1998

Activation of CD4+ and CD8+ T-lymphocytes in bone marrow associated with reduced erythropoiesis in patients with chronic inflammation and anaemia.

We hypothesised that, in "anaemia of chronic disorders" (ACD), a local immune activation in the bone marrow alters erythropoiesis. We determined the activation status of bone marrow T-lymphocytes and the levels of cytokines. Patients (n = 27) with chronic inflammatory diseases and low hemoglobin values who had undergone bone marrow aspiration were divided into 2 groups according to the aetiology of anaemia: (1) unknown or (2) known causes (i.e. iron, folic acid and/or vitamin B12 deficiencies). Cytokines in peripheral blood and bone marrow serum were measured by ELISA and T-lymphocytes were phenotyped by flow cytometry. Peripheral blood from 27 age-matched healthy donors and bone marrow from 3 transplantation donors served as control. The erythrocyte count in the peripheral blood of patients with unknown aetiology of anaemia (group 1) was lower than in patients with known aetiology (group 2). This indicated that group 1 included most patients with ACD. Both groups had active disease, as shown by elevated levels of C-reactive protein and the erythrocyte sedimentation rate. The levels of IL-6 and IL-7 in the peripheral blood were higher in patients of group 1 than in patients of group 2 or normal subjects. The levels of IL-6, IL-7 and TNF-alpha in the bone marrow also were higher in group 1 compared to group 2. The CD4+ and CD8+ T-lymphocytes of the bone marrow showed a more activated phenotype than peripheral blood T-lymphocyte with the number of T-lymphocytes that expressed the early activation marker, CD69, being higher in group 1 than in group 2. A smaller proportion of "naive" cells (CD45RO-, CD69-) was accompanied by a higher proportion of "false naive" cells (CD45RO-, CD69+). In the bone marrow, elevated levels of IL-7 were correlated with fewer "naive" and more "false naive" cells in the CD4+ T-lymphocyte subpopulation. In group 1 only, a significant inverse relation was observed between the erythrocyte counts in blood and the levels of IL-7 in bone marrow. Thus, in patients with unknown aetiology of anaemia--a group including most patients with ACD-, an interrelationship may exist between the local activation of bone marrow T-lymphocytes, increased production of cytokines, and reduced erythropoiesis.

Adult↗
PubMed1998

Interleukin-16, produced by synovial fibroblasts, mediates chemoattraction for CD4+ T lymphocytes in rheumatoid arthritis.

The massive infiltration of synovium with CD4+ T cells during the course of rheumatoid arthritis (RA) implies the expression of chemoattractant factors by resident synovial cells. Therefore, we analyzed the expression of IL-16, a potent chemoattractant for CD4+ T cells, to account for the accumulation of CD4+ T cells in RA. Indeed, IL-16 was found to be significantly elevated in synovial fluid (SF) from patients with RA as compared to non-RA arthritis (p < 0.001), osteoarthritis (p < 0.001) and controls (p < 0.001). Chemotaxis studies showed IL-16 to contribute to the strong chemotactic activities of RA-SF. In situ hybridization (ISH) revealed IL-16 mRNA-expressing cells located within the lining layer of rheumatoid synovial tissue. In the sublining area, only scattered IL-16 transcript-positive cells could be detected, mainly adjacent to blood vessels. By a double-labeling technique, combining ISH for IL-16 mRNA and immunohistochemistry for CD68, synovial fibroblast-like, CD68-negative cells were identified as a major source of IL-16 mRNA within RA synovium. This study demonstrates that synovial fibroblasts produce IL-16 in RA and thus mediate chemoattraction of CD4+ cells into synovial tissue.

Adult↗
PubMed1998

Differential expression of cathepsins B and L compared with matrix metalloproteinases and their respective inhibitors in rheumatoid arthritis and osteoarthritis: a parallel investigation by semiquantitative reverse transcriptase-polymerase chain reaction and immunohistochemistry.

OBJECTIVE: To study the expression of the cysteine proteinases cathepsin B and L and their most potent inhibitor cystatin C in the synovial membrane of patients with rheumatoid arthritis (RA) and osteoarthritis (OA) on both the messenger RNA (mRNA) level and the protein level. METHODS: The expression of both cysteine proteinases and cystatin C was investigated in synovial tissue from 15 RA and 11 OA patients and compared with the expression of matrix metalloproteinase 1 (MMP-1; collagenase), MMP-3 (stromelysin), and tissue inhibitor of metalloproteinases 1 (TIMP-1). Expression of mRNA was analyzed by semiquantitative reverse transcriptase-polymerase chain reaction. Protein expression was evaluated by immunohistochemistry. The results were correlated with the histologic evidence of inflammatory activity. RESULTS: A significantly more pronounced expression of MMP mRNA was observed in RA synovium compared with OA. In contrast, the mRNA expression of cysteine proteinases, as well as TIMP-1 and cystatin C, did not differ between the patient groups. However, the protein expression of both MMP and cysteine proteinases was significantly more prominent in RA synovial lining compared with OA, whereas cystatin C and TIMP-1 protein were expressed equally. CONCLUSION: The data indicate a more pronounced expression of MMP mRNA compared with cysteine proteinases in RA. The higher levels of cathepsin B and L proteins in RA synovial lining cells compared with OA are consistent with previous studies that assert a post-transcriptional up-regulation of these enzymes in RA.

Adult↗
PubMed1998

Immunohistochemical and molecular studies of serotonin, substance P, galanin, pituitary adenylyl cyclase-activating polypeptide, and secretoneurin in fibromyalgic muscle tissue.

OBJECTIVE: Because former investigations have reported abnormal changes in the expression of serotonin (5-hydroxytryptamine [5-HT]) and substance P (SP) in serum and cerebrospinal fluid, this study sought to determine whether 5-HT and pain-modulating neuropeptides (SP, galanin [GA], pituitary adenylyl cyclase-activating polypeptide, and secretoneurin) were expressed abnormally in the muscle tissue of patients with fibromyalgia (FM). METHODS: Snap-frozen muscle tissue specimens (deltoid muscles) from 10 patients with FM (mean disease duration 15 years) and from 10 healthy control subjects were examined by reverse transcriptase-polymerase chain reaction (RT-PCR) of RNA preparations from muscle cells, and by immunohistochemistry methods (alkaline phosphatase-anti-alkaline phosphatase and immunogold-silver) using specific primers as well as antibodies. When specific messenger RNA (mRNA) was detected by RT-PCR, in situ RT-PCR was performed for mRNA localization. RESULTS: Specific mRNA for the examined substances was absent in 9 of 10 FM patients and in 10 of 10 controls. No differences between the FM patients and controls could be detected in the muscle tissue by immunohistochemistry. In 1 FM patient, mRNA for the GA receptor could be shown. CONCLUSION: This study showed that 5-HT and neuropeptides are not produced in the muscle of patients with FM, and therefore do not appear to be involved in the peripheral induction of pain in this chronic, painful disorder.

Adult↗
PubMed1998

Persistent high serum levels of cartilage oligomeric matrix protein in a subgroup of patients with traumatic knee injury.

The objective was to assess whether changes of cartilage oligomeric matrix protein (COMP) serum levels can predict the development of osteoarthritis following traumatic knee injury. Sera and synovial fluids were acquired at surgery (T0) and postoperatively during the first (T1) and second (T2) year from 30 knee-injured patients. COMP levels and anti-COMP autoantibodies were quantified by ELISA. Radiographs and patient questionnaires were used to assess outcomes. At T0, compared with controls (1.6 +/- 1.6 micrograms/ml), the serum COMP concentration was significantly elevated (6.5 +/- 2.8 micrograms/ml) with a tendency to further increase (T0 vs. T1, P = 0.076) and subsequently decrease (T1 vs. T2, P = 0.074). However, individual variations are observed, e.g. persistently high (8/30) or increasing (T0 to T2, 8/30) serum COMP. Ten of these patients have elevated COMP at T2 that increased from T0. COMP levels in serum and synovial fluid correlated significantly (P = 0.012). Interestingly, some patients who revealed increasing serum levels of COMP from T0 to T2 displayed anti-COMP autoantibodies. These data suggest that local immune response could contribute to further joint damage. The subgroup of 10 patients (33%) with elevated and increasing serum COMP levels and in particular the patients with antibodies against cartilage matrix molecules appear at increased risk for developing posttraumatic osteoarthritis.

Adult↗
PubMed1998

Analysis of cartilage oligomeric matrix protein (COMP) in synovial fibroblasts and synovial fluids.

We investigated the expression of cartilage oligomeric matrix protein (COMP) in normal and rheumatoid arthritis (RA) synovial fibroblasts. In situ hybridization (ISH) was conducted on synovial specimens from five RA patients applying specific probes for COMP or fibroblast collagen type I. ISH was combined with immunohistochemistry, applying antibodies to the macrophage marker CD68. Ribonuclease protection assay (RPA) and rapid amplification of 3'-cDNA ends (3'-RACE) were performed on total RNA from normal and RA synovial fibroblast cultures. Protein extracts from fibroblasts and culture supernatants were compared with synovial fluids and protein extracts from isolated chondrocytes by Western blot utilizing polyclonal and monoclonal antibodies (18-G3 mAb) to COMP. COMP mRNA was detected in fibroblasts of RA synovium by ISH, and in normal and RA synovial fibroblast cultures by RPA. 3'-RACE demonstrated sequence homology of chondrocyte and synovial fibroblast COMP along the coding sequence. COMP protein was detected in synovial fibroblasts and culture supernatants by immunoblot. Using polyclonal antibodies, the major portion of COMP from fibroblasts and culture supernatants was present as low-molecular-weight (LMW) bands, corresponding to those found in synovial fluids. These LMW COMP bands, however, were not detected in any of the cells or tissues tested using 18-G3 mAb. In protein extracts from chondrocytes and in COMP purified from cartilage, these LMW bands could not be detected. In conclusion, the data suggest that certain forms of COMP detected in synovial fluid are secreted from synovial fibroblasts and could be distinguished by specific mAbs from COMP secreted by chondrocytes.

Arthritis, Rheumatoid↗