Growth factors, extracellular matrix, and oncogenes in scleroderma.
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Extracellular Matrix↗
Biomedical subjects
S Gay
Publications and source records attributed to S Gay.
Intestinal intraepithelial lymphocyte T cells are resistant to lpr gene-induced T cell abnormalities.
The mucosal immune system of the gastrointestinal (GI) tract consists of Peyer's patches (PP), which are IgA inductive sites, and more diffuse effector regions which include cells in the intraepithelial lymphocyte (IEL) compartment. Since autoimmune MRL lpr/lpr (MRL/lpr) mice develop a proliferating CD3+, CD4-, CD8- (double negative; DN), B220+ T cell subset in systemic lymphoid tissue, we have initiated studies to determine the distribution of CD3+, DN, B220+ T cells (B220+ T cells or lpr/lpr T cells) in the GI immune system. Specifically, we examined T cell subsets separated according to expression of CD4, CD8, Thy-1, B220, alpha/beta T cell receptor (TcR) and gamma/delta TcR in PP and IEL of MRL/lpr mice at 6, 12 and 21 weeks of age. Increased numbers of CD3+ T cells were noted in both PP and spleen of 12- and 21-week-old mice in which the development of autoimmune disorders were also evident. However, normal numbers of CD3+ IEL T cells were seen in MRL/lpr mice in all three age groups tested. When the presence of T cell lymphadenopathy was examined in both IgA inductive and effector tissues, the PP followed the B220+ T cell pattern seen in the spleen, where approximately 30%-50% of CD3+ T cells in the PP of 12- and 21-week-old MRL/lpr mice expressed the phenotype of lpr/lpr T cells and greater than 90% were alpha/beta TcR+. On the other hand, B220+ T cells had not developed in PP or spleen of 6-week-old MRL/lpr mice. Of interest was the finding that IEL from lpr/lpr homozygous mice did not contain B220+ T cells in any age group tested. In this regard, the IEL of MRL/lpr mice comprised an identical pattern and frequency of CD4-/CD8+, CD4+/CD8-, DN and CD4+/CD8+ (double positive, DP) T cell subsets as their normal counterparts (i.e. MRL +/+, BALB/c and C3H/HeN mice) which consisted of approximately 75%, approximately 7.5%, approximately 7.5% and approximately 10%, respectively. Further, Thy-1, gamma/delta TcR and alpha/beta TcR expression in these four subsets of MRL/lpr IEL were very similar to normal mice. These results suggest that the intestinal IEL compartment is minimally affected by the lpr/lpr mutation which induces T cell abnormalities and indicate that B220+ T cells do not preferentially home to IEL. Further, our results support the concept that IEL T cells develop as a separate T cell lineage from thymus-derived cells.
Animals↗
Scanning electron microscopic evaluation of the arthritis in MRL/lpr mice.
The articular surfaces of disarticulated knee joints from MRL/lpr and MRL/n mice, aged 4-33 weeks were examined by light microscopy (LM) and scanning electron microscopy (SEM). Light microscopy did not reliably predict SEM findings. Most of the abnormalities detected by SEM were related to surface disruption of articular cartilage. However, areas of articular cartilage covered by tightly adherent non-confluent monolayers of stellate-shaped cells with intertwining cytoplasmic processes were observed. In these areas the integrity of the underlying cartilage matrix was disrupted, with exposure of collagen fibers. These findings suggested that outgrowth of proliferating synovial cells in the joints of arthritic MRL/lpr mice may lead to cartilage destruction.
Animals↗
Spontaneous expression of immediately-early response genes c-fos and egr-1 in collagenase-producing rheumatoid synovial fibroblasts.
In view of the important role of interstitial collagenase in the pathogenesis of rheumatoid arthritis (RA), we studied the expression of fibroblast-type collagenase in rheumatoid synovium and searched for its potential transcription factors, namely the oncoprotein c-fos and the early-growth-response gene-1 (egr-1), an inducible zinc-finger encoding gene. Elevated levels of RNA sequences complimentary to c-fos and egr-1 cDNA probes could be detected in cytoplasmic extracts of collagenase-expressing synovial fibroblast-like cells when compared to equivalent RNA amounts isolated from control fibroblasts. Utilizing immunocytochemistry, immunoreactivity for c-fos oncoprotein was found in 13 of 19 joint specimens obtained from patients with active RA. These oncoprotein data were positively correlated to the collagenase expression in the same specimens. Moreover, immunohistochemical analysis confirmed the localization of both oncoprotein c-fos and fibroblast-type collagenase within synovial fibroblast-like cells attached to bone erosions.
Arthritis, Rheumatoid↗
Long-term treatment of the MRL/lpr mouse with methotrexate and 10-deazaaminopterin.
Female MRL/lpr mice were treated with I.P. doses of methotrexate (MTX) and 10-deazaaminopterin (DAAM) in the range of 1 to 100 mg/kg body weight/week, in two equally divided doses. Treatment began at 7 weeks of age and continued to 30 weeks of age. Joint histopathology scores were tightly correlated with skin lesion-proteinuria scores at 30 weeks of age. MTX at levels of 5, 25, and 100 mg/kg body weight/week and DAAM at a level of 25 mg/kg body weight/week significantly reduced skin lesion-proteinuria scores below controls in a dose dependent manner. Animals receiving MTX at 25 mg/kg body weight/week had a significantly longer median life span and animals receiving MTX at 100 mg/kg body weight/week had a greater than 15% suppression of growth when compared with controls. Longevity and skin lesion-proteinuria scores appeared to be good indicators of drug efficacy while growth suppression appeared to be a good indicator of drug toxicity.
Aminopterin↗
Expression of collagenase and potential transcriptional factors in the MRL/l mouse arthropathy.
Typical erosions of articular joint structures in rheumatoid arthritis and in the spontaneous destructive hind-limb arthropathy of autoimmune MRL-lpr/lpr (MRL/l) mice occur predominantly in areas contiguous with proliferating synovial lining cells, suggesting release of proteolytic enzymes from these cells. Synovial lining cells were isolated from arthritic MRL/l mice, and the spontaneous expression of the interstitial procollagenase and its potential transcriptional factors, egr-1 and c-fos, was examined in vitro. The data indicate that basal collagenase RNA expression was stronger in MRL/l cells than in virus-transformed cells. Moreover, elevated RNA levels of the c-fos gene could be detected in the collagenase-expressing synovial lining cells in vitro. In a related immunohistochemical study, collagenase was detected in situ in proliferating synovial lining cells as well as in chondrocytes of the first stage of pathological changes in the MRL/l mouse arthropathy.
Animals↗
Binding of serum immunoglobulins to collagens in IgA nephropathy and HIV infection.
The mechanism of the binding of IgA to the mesangium in IgA nephropathy (IgAN) is unknown. Interactions between IgA and components of the mesangial matrix may contribute. We measured by enzyme-linked immunosorbent assay the binding of serum IgA, IgG, and IgM from patients with IgAN, human immunodeficiency virus type I (HIV) infection, and healthy controls to purified native collagen types I to VI, and to an extract of normal kidney tissue. HIV infection is an appropriate disease control because of the lack of mesangial IgA deposits, despite high serum levels of IgA and IgA1-containing immune complexes. Increased levels of IgA-binding to collagen types I and V and the kidney extract were found only in IgAN. Both IgAN and HIV-infected patients had increased IgA-binding to collagen types II, III, and VI. Preabsorption of the sera with gelatin substantially reduced the IgA-binding to collagen types I to IV, but not to types V and VI. This finding suggests that the binding to collagen type V is not fibronectin-mediated, but may reflect autoantibody formation. Thus, fibronectin-mediated IgA-collagen interactions are not specific for IgAN, and their pathogenetic role is questionable. The role of IgA anti-collagen type V antibodies requires further study.
Adolescent↗
Expression of collagenase and potential transcriptional factors c-fos and egr-1 in periodontal gingival fibroblasts.
In view of the important role of fibroblast-type collagenase in the pathogenesis of periodontal disease (PD), we investigated the expression of this metalloproteinase in primary cultures of non-stimulated fibroblasts dissected from gingival tissues of patients with generalized moderate and localized severe chronic adult PD. Enhanced hybridization signals for collagenase RNA were observed in 8/8 PD-cases when compared with equivalent RNA amounts extracted from normal fibroblasts. Since both the proto-oncogene c-fos and the "early growth response" gene egr-1 might be involved in the transcriptional regulation of the collagenase gene expression in vivo, we also compared the relative expression of both potential transcriptional factors with collagenase RNA in the same fibroblast cytoplasmic extracts. Hybridization signals indicated elevated RNA amounts for c-fos in 8/8 PD-cases and for egr-1 in 7/8 PD-cases when compared with the cells from non-inflamed tissue. In periodontitis gingival tissue specimens, immunolocalization of collagenase could be confirmed in fibroblasts, macrophages and epithelial cells in situ. Collagenase label was not widely distributed within the tissues, but concentrated at the interface between epithelium and connective tissue. The data provide the first evidence that gingival fibroblasts producing elevated levels of collagenase RNA amounts express also c-fos and egr-1 indicating a crucial role for both genes in cellular proliferation and collagenase expression in gingival and periodontal tissue destruction in vivo.
Alveolar Bone Loss↗
Oncogene activation in rheumatoid synovium.
Rheumatoid arthritis (RA) is a chronic systemic disorder that is dominated by the debilitating sequelae associated with the progressive destruction of articular joints. The molecular and cellular basis of rheumatoid joint destruction is characterized by an abnormal expression of oncogenes modulating cellular proliferation and the induction of lysosomal and metalloproteinases. Based on the observation that the synovial hyperplasia in RA is associated with the proliferation of transformed-appearing synovial lining cells and an overexpression of such oncogenes, the possibility that a hitherto unknown HTLV related retrovirus is involved in the etiopathogenesis of RA is discussed.
Arthritis, Rheumatoid↗
Characterization of heterotypic adherence between transformed human lymphoblastic cells and marrow stromal cells: VCAM-1 is a ligand for one of the leukemia cell adhesion proteins.
Marrow stromal cells are important in normal myelopoiesis and support growth of leukemia/lymphoma (LL) cells in vitro. We have previously described the heterotypic adherence of a human B-lymphoblastic cell line (UTMB-460) to marrow stromal cells (MSC). We have extended these observations to a human T-lymphoblastic cell line (CEM) and characterized the heterotypic adherence of B- and T-lymphoblastic cell lines to human MSC. Electron microscopy demonstrated UTMB-460 cells were in very close apposition to the MSC, but no specific intercellular junctions were noted. Under the conditions employed, these MSC express extracellular fibronectin, collagen types I and IV, intracellular laminin, and vimentin, but no factor VIII-R antigen. In addition, the MSC had receptors for the lectin Ulex europaeus agglutinin I. UTMB-460 and CEM cells do not adhere to extracellular matrix (ECM) proteins secreted by the MSC, i.e., fibronectin, collagen types I, III, or IV, or laminin. Monoclonal antibodies (MoAbs) against CD11a, CD11b, CD18, and CD54 and a polyclonal anti-human fibronectin antibody do not inhibit attachment of either B- or T-lymphoblastic cells to MSC. Peptides GRGES and GRGDS did not inhibit adherence of UTMB-460 and CEM cells to MSC. In contrast, the anti-vascular cell adhesion molecule (VCAM)-1 MoAb (4b9) caused significant inhibition (p < 0.01) of the adherence of both UTMB-460 and CEM cells to normal human MSC monolayers. These data suggest: (1) that MSC to which lymphoblastic cells adhere are specialized mesenchymal cells; (2) that the membrane interactions between T- and B-lymphoblastic cells and MSC involve close apposition of cell membranes of MSC and the lymphoblastic cells; (3) that the heterotypic adherence between B- and T-lymphoblastic cell lines (UTMB-460 and CEM) and MSC does not involve the RGD recognition sequence of the integrin family, the B2 leukocyte integrins, CD44, LAM-1, or the ECM proteins examined; and (4) that VCAM-1 may at least be partially responsible for heterotypic adherence between human MSC and B- and T-lymphoblastic cells.
Amino Acid Sequence↗
A comparison of magnetic resonance imaging to bone scintigraphy in early traumatic ischemia of the femoral head.
In adult rabbits, a unilateral subcapital osteotomy of the femoral neck was performed to induce avascularity. One half of the osteotomy sites were fixed with a magnetic resonance imaging (MRI) compatible absorbable pin and the other osteotomies had no fixation. The femoral heads were studied at three, five, and 12 days with roentgenographs, bone scintigraphy, and MRI, and MRI only at four weeks and six weeks after osteotomy. Histologic studies were performed after imaging to evaluate the viability of the femoral heads. At three, five, and 12 days after osteotomy, bone scintigraphy showed a decrease in uptake of radioisotope in the region of the femoral head on the operated side relative to the acetabulum and greater trochanter in 17 of 18 rabbits. A comparison of the surgically treated hip to the normal hip in fixed and unfixed osteotomies showed no change in the signal behavior of T1- or T2-weighted images in all rabbits Days 3, 5, and 12 (n = 18) after operation. The rabbit femoral heads with fixation of the osteotomy 28 days after operation showed a decrease in signal intensity in the subcapital region of the femoral head. Six weeks after operation, the fixed femoral head shows a loss of signal in a portion of the femoral head near the osteotomy. The MRI signal intensity appears to increase in the unfixed femoral heads six weeks after operation.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Ultrastructural pathway of human fibroblast collagenase and distribution in tissues of rheumatoid joint destruction.
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Animals↗
Cathepsin B in synovial cells at the site of joint destruction in rheumatoid arthritis.
Based on the concept that proteolytic enzymes, like cathepsins, are associated with tissue destruction, we investigated the expression of the matrix-degrading cysteine proteinase cathepsin B in synovial tissues from the joints of patients with rheumatoid arthritis. The data indicate an enhanced transcription of cathepsin B in synovial cells when compared with normal fibroblasts, cathepsin B-producing epithelial tumor cells (SW1116), or fibroblasts derived from inflamed tonsils. Immunolocalization of cathepsin B appeared to be restricted mainly to the synovial cells attached to cartilage and bone at sites of rheumatoid joint erosion.
Arthritis, Rheumatoid↗
Detection of p24 in HIV-1 infected cells embedded in LR White and Lowicryl K4M.
In this study we present a postembedding on-grid immunogold labelling procedure for the ultrastructural localization of the HIV-1 core protein p24. HIV-1 infected cells were fixed in 0.1% glutaraldehyde, incompletely dehydrated and embedded in LR White or in Lowicryl K4M. Antigenic sites were detected by incubation of ultrathin sections with primary mouse monoclonal antibody anti-HIV-1 p24, followed by the secondary antibody goat anti-mouse IgG coupled to 10nm gold particles. Antigenicity of p24 was found to withstand the applied fixation and was shown to be preserved in LR White as well as in Lowicryl. The described procedure permits the uncomplicated and easy detection of p24 in HIV-1 infected cells and tissues.
Acrylic Resins↗
Collagen autoantibodies in patients with vasculitis and systemic lupus erythematosus.
In order to determine whether collagen autoantibodies are present in sera from patients with vascular diseases, a highly sensitive solid-phase radioimmunoassay was utilized to evaluate the presence of antibodies against collagen types I to VI and laminin in sera of 20 patients with vasculitis and 20 patients with systemic lupus erythematosus (SLE). Autoantibodies to interstitial collagen types I and II were noted in 20 and 35% of the patients, respectively. Most significantly, 70% of the polyarteritis nodosa patients and 55% of the total vasculitis group had autoantibodies to collagen type IV. In SLE, autoantibodies to collagen type V were detected in 70% and to collagen type IV in 85% of the patient sera. These data indicate that basement membrane and basement membrane-associated collagens may become immunogenic in patients with vasculitis and SLE. The role of these autoantibodies is most likely a result of endothelial damage secondary to the initial inflammatory disease process. However, it can be concluded from the present studies that collagen types IV and V are involved in the immune response and thereby may perpetuate further vascular damage.
Adolescent↗
Immunolocalization of types V and XI collagen in cartilage using monoclonal antibodies.
Monoclonal antibodies produced against pepsin-solubilized newborn rat skin type V collagen [alpha 1(V)]2 alpha 2(V), and chondrosarcoma type XI collagen [alpha 1(XI) alpha 2(XI) alpha 3(XI)] are used to localize the collagens in sections of the chondrosarcoma as well as the normal rat knee joint by indirect immunofluorescence. Immunostaining for type V collagen shows strong cellular staining of chondrocytes; while the interstitial matrix as well as the lacunae are not stained. In contrast, antitype XI stains not only chondrocytes, but the extracellular compartments as well. In ELISA, rat anti-type XI collagen reacts with its native antigen, but does not cross-react with native types I, II, III, or V collagen from rat. The distinct locations of type V and XI collagens in cartilaginous tissue suggest varied functional roles for these constituents in the tissue.
Animals↗
Immunoglobulin isotype distribution of locally produced autoantibodies to collagen type I in adult periodontitis. Relationship to periodontal treatment.
Production of antibodies to collagen type I was analyzed by means of an enzyme-linked immunospot (ELISPOT) assay in patients with chronic adult periodontitis (AP) before and after periodontal hygiene treatment. Anti-collagen type I antibody-secreting cells were found among mononuclear cells enzymatically eluted from inflamed gingiva in 9 of 15 patients with untreated AP and in 4 of 14 hygiene-treated patients with a varied isotype distribution. A notably high prevalence of IgG and IgM isotypes was observed for the anti-collagen antibodies in untreated patients. With wide variation, chronic AP was characterized by a high frequency of spontaneous IgG and low numbers of IgA and IgM-producing cells. Periodontal hygiene treatment significantly reduced the number of IgA and IgM-secreting cells. Although AP is not an autoimmune disease in the accepted sense, our results indicate that local autoimmune reactions to collagen type I are common in untreated AP, implying an interplay between periodontal infection and autoimmunity.
Autoantibodies↗