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T T Glant

Publications and source records attributed to T T Glant.

At least 19 recordsLinked to original sources
PubMed1997

Human monocyte/macrophage response to cobalt-chromium corrosion products and titanium particles in patients with total joint replacements.

The responses of human peripheral blood monocytes of 10 normal volunteers and 14 patients with total hip replacements to particles of commercially pure titanium and chromium orthophosphate (a corrosion product from cobalt-chromium alloy implants) were studied. In addition, these phagocytosable particles were added to cultured mononuclear cells isolated from the interfacial membrane of 14 patients with failed implants. Peripheral blood monocytes from patients who had had a total hip replacement produced significantly higher levels of interleukin-1 (both interleukin-1 alpha and interleukin-1 beta) and prostaglandin E2 following particulate stimulation than those from normal volunteers. Supernatants from both titanium and chromium orthophosphate-stimulated peripheral blood monocytes from the volunteers and patients with total hip replacement induced bone resorption (assayed in organ cultures of newborn mouse calvariae) and the proliferation of human fibroblasts. The levels of bone resorption were significantly higher (p < 0.05) in patients with implants than in normal volunteers. There were no significant differences in the responses of cells between patients with focal osteolysis and those without osteolysis. Interfacial membrane mononuclear cells also produced high levels of interleukin-1 alpha, interleukin-1 beta, and prostaglandin E2 and expressed bone resorptive activities following stimulation with either titanium or chromium orthophosphate. More importantly, interfacial membrane mononuclear cells "spontaneously" produced high levels of prostaglandin E2 that were comparable with the response of peripheral blood monocytes stimulated with particulate wear debris. The clinical relevance of this study is 2-fold. First, mononuclear cells from patients with total hip replacement were some-how "sensitized" to metal particles in comparison with mononuclear cells from individuals without an implant. Second, the chromium orthophosphate corrosion product was a potent macrophage/monocyte activator and may contribute to macrophage-mediated osteolysis and aseptic loosening.

Adult↗
PubMed1997

CD44 expression by leucocytes in rheumatoid arthritis and modulation by specific antibody: implications for lymphocyte adhesion to endothelial cells and synoviocytes in vitro.

Anti-CD44 MoAb IM7 induced the loss of CD44 from mouse leucocytes thereby inhibiting leucocyte migration and joint inflammation in murine arthritis. Thus, targeting CD44 with MoAb may have potential for the treatment of patients with inflammatory joint diseases. Expression of CD44 by peripheral blood (PB) and synovial fluid (SF) leucocytes from rheumatoid arthritis (RA) patients was compared and the ability of IM7 to modulate this expression determined. RASF lymphocytes showed increased CD44 expression compared with those in PB indicative of an activated phenotype. As inflammatory SF did not up-regulate CD44 expression on PB lymphocytes, the increased CD44 expression by SF lymphocytes was a result of the selective homing of CD44(high) cells to the synovium rather than an effect of the synovial environment. RASF granulocytes showed reduced CD44 expression compared with those in PB, again indicative of an activated phenotype. However, this reduction could be induced on PB granulocytes following culture with inflammatory SF and was inhibited by anti-TNF-alpha MoAb, implying that soluble factors in inflammatory SF such as TNF-alpha induced granulocyte activation and CD44 loss. IM7 induced the loss of CD44 from lymphocytes (both from PB and SF) and granulocytes in vitro, but was subsequently re-expressed after 24 h culture in the absence of the MoAb. This loss of CD44 was blocked by serine- and metalloprotease inhibitors implying that IM7 induced the proteolytic cleavage of CD44 by a mechanism similar to that reported for the loss of CD44 from PMA-activated granulocytes. Furthermore, IM7-treated CD44(low) lymphocytes showed reduced adherence to both an endothelial cell line and RA synovial fibroblasts in vitro. The unique ability of IM7 to reduce CD44 expression by lymphocytes suggests that it could prevent lymphocyte extravasation and synovial infiltration in RA as previously reported in murine arthritis.

Antibodies, Monoclonal↗
PubMed1997

Suppression of osteoblast function by titanium particles.

In order to understand the effect of particulate debris on osteoblast function, we studied the effect of different particles, including titanium and polystyrene, on bone collagen mRNA (messenger RNA) with the use of Northern blot hybridization analysis, and we studied the effect of the particles on the biosynthesis of bone collagen with analysis of 3H-proline incorporation and with the Western blot technique. The steady-state levels of mRNA for procollagens alpha1(I) and alpha1(III) were markedly suppressed in human MG-63 osteoblast-like cells exposed to phagocytosable titanium particles that were smaller than three micrometers. Both titanium and polystyrene particles smaller than three micrometers suppressed the expression of the gene that codes for collagen, and the suppression of the expression of the gene was related to the size but not to the composition of the particles. The biosynthesis of both type-I and type-III collagen also was decreased in cells that had been treated with titanium particles. Neither the viability nor the proliferation of cells was affected by particulate debris. These data indicate that phagocytosable titanium particles can significantly suppress the expression of the gene that codes for collagen in osteoblast-like cells (p < 0.05).

Blotting, Northern↗
PubMed1996

Species-specific alternative splicing of the epidermal growth factor-like domain 1 of cartilage aggrecan.

Aggrecan transcripts contain two alternatively spliced exons that code for two epidermal growth factor-like domains (EGF1 and EGF2). Whereas the EGF2 sequence is expressed at a uniform level among different species, the EGF1 sequence has been detected only in human aggrecan transcripts. In this study we have used the nested primer reverse transcription-PCR (RT-PCR) method to compare the expression of the EGF1 exon in human, bovine and dog aggrecan transcripts. Our results indicate that this exon is expressed in a species-specific manner. In addition to its significant expression level in human transcripts, the EGF1 sequence can be detected in a small portion of bovine aggrecan transcripts as shown with nested primer RT-PCR. In contrast, the same module is not detectable in dog aggrecan transcripts, although an EGF1 sequence is present in the dog aggrecan gene. The expression level of the EGF1 exon in the aggrecan transcripts correlates with the strength of the polypyrimidine tract upstream of the exon. The EGF1 sequence also shows much less conservation between the species than the EGF2 sequence. The species-specific expression and high sequence variation of the EGF1 exon imply that this sequence is likely to code for an aggrecan domain having no cartilage-specific function.

Aggrecans↗
PubMed1995

A proteoglycan (aggrecan)-specific T cell hybridoma induces arthritis in BALB/c mice.

Aggrecan, the high buoyant density cartilage proteoglycan (PG), has been shown to induce progressive polyarthritis and ankylosing spondylitis in genetically susceptible BALB/c mice. To further characterize the nature of the autopathogenic effector T cells operating in these mice and to determine the region(s) of the PG molecule recognized by these T cells, we generated PG-specific T cell hybridomas from arthritic mice. One of the PG-specific T cell hybridomas (5/4E8), when injected into naive irradiated BALB/c mice, was capable of inducing clinical and histopathologic signs of arthritis. Massive swelling and redness of the paws dominated the clinical picture. A reactive synovial cell proliferation, the accumulation of hybridoma and inflammatory cells in the enlarged joint space, the loss of PG from the superficial layer of the articular cartilage, and the erosion of articular surface were identical histopathologic signs to those found either in primary or adoptive transfer of PG-induced arthritis. The PG-specific and arthritogenic T cell hybridoma (5/4E8) expressed TCR-alpha beta + (V beta 4), CD4+, and CD8- phenotypes and belonged to the Th1 subset, as the cells secreted IL-2 and IFN-gamma, but not IL-4 upon PG stimulation, and the response was MHC class II (I-Ad)-restricted. These observations provide direct evidence that PG-specific Th cells play crucial roles in autoimmune arthritic processes.

Aggrecans↗
PubMed1995

Large and small proteoglycans of osteoarthritic and rheumatoid articular cartilage.

OBJECTIVE: To identify characteristic changes in large aggregating (aggrecan) and small proteoglycan (PG) populations in articular cartilages during osteoarthritis (OA) and rheumatoid arthritis (RA). METHODS: Aggrecan populations in guanidine extracts of femoral condylar cartilages of 46 OA and 8 RA patients who underwent total knee arthroplasty, as well as of 2 fetuses and 6 normal adults, were separated in agarose-polyacrylamide composite gels. Small PGs (biglycan, decorin, and fibromodulin) in the same extracts were analyzed in 12% polyacrylamide gels. Gels were stained or electrophoretically transferred and probed with antibodies to aggrecan epitopes and to small PGs. Epitope contents of the samples were also compared by inhibition radioimmunoassay. RESULTS: There were significant differences found among normal and diseased samples in their electrophoretic mobilities, band distributions, and antibody staining. OA and especially RA samples were heavily degraded, lacked certain aggrecan populations, and contained fewer keratan sulfate and chondroitin-6-sulfate epitopes compared with normal samples. Levels of chondroitin-4-sulfate and "fetal-type" epitopes were elevated in the OA samples compared with the normal ones. More core proteins of small PGs were found in diseased than in normal cartilages, but they were more heterogeneous in size and glycosaminoglycan substitution. CONCLUSION: There is extensive degradation of both large and small PGs in diseased cartilages, but a repair process does exist, especially in OA cartilages. Chondrocytes of diseased cartilages are able to synthesize fetal-type aggrecans. Small PGs are glycosylated differently in diseased cartilages than in normal ones.

Adult↗
PubMed1995

The potential role of fibroblasts in periprosthetic osteolysis: fibroblast response to titanium particles.

Periprosthetic osteolysis with or without aseptic loosening is a major clinical problem in total hip arthroplasty. While the macrophage response to prosthetic wear debris and its role in periprosthetic osteolysis has been extensively studied, information regarding other cell types (fibroblasts, osteoblasts) is limited. This study explored the response of fibroblasts to particulate wear debris. Fibroblasts isolated from interfacial membranes of patients with failed total hip replacements and normal synovial tissue, when challenged with small-sized ( < 3 microns) titanium (Ti) particles, responded with significantly enhanced expressions of collagenase, stromelysin and, to a much lesser extent, their tissue inhibitor of metalloproteinases (TIMP). These "regulated" expressions at both mRNA and protein levels were correlated with the size and composition of particles. De novo protein synthesis was required for the regulation of these mRNAs. A similar effect could be induced by the treatment of the cells with particle-free conditioned medium from Ti particle-stimulated fibroblasts. Furthermore, this conditioned medium significantly suppressed the mRNA levels of procollagen alpha 1 (I) and alpha 1 (III) in osteoblast-like MG-63 cells. It is concluded that fibroblasts stimulated with certain particle debris may play an important role in periprosthetic osteolysis by releasing bone-resorbing metalloproteinases and mediator(s) which resulted in suppressed collagen synthesis in osteoblasts.

Base Sequence↗
PubMed1995

Human monocyte response to particulate biomaterials generated in vivo and in vitro.

We studied the ability of four clinically relevant particle species to stimulate human peripheral blood monocytes to release bone-resorbing agents, including interleukin-1 (both interleukin-1 alpha and interleukin-1 beta), interleukin-6, and prostaglandin E2. The species studied were titanium-6% aluminum-4% vanadium (TiAlV), commercially pure titanium, fabricated ultrahigh molecular weight polyethylene, and polyethylene retrieved from interfacial membranes of failed uncemented total hip arthroplasties. For all species, the mean size was less than 1 micron. Human peripheral blood monocytes were challenged with these particles in a uniform manner on the basis of surface area. Phorbol 12-myristate acetate, zymosan, and nonphagocytosable titanium particles served as controls. Stimulation of human monocytes is a function of the composition and concentration of particles. In this study, TiAlV particles appeared to be the most competent to elicit the synthesis and release of inflammatory mediators. Particles of commercially pure titanium and of fabricated ultrahigh molecular weight polyethylene also could induce the release of various cellular mediators, albeit at a lower level, whereas the particles of polyethylene retrieved from interfacial membranes were less stimulatory in these short-term in vitro experiments.

Alloys↗
PubMed1995

Interferon-gamma but not granulocyte/macrophage colony-stimulating factor augments proteoglycan presentation by synovial cells and chondrocytes to an autopathogenic T cell hybridoma.

Immunization of BALB/c mice with human cartilage proteoglycan (aggrecan) produces a progressive polyarthritis, similar in many aspects to human rheumatoid arthritis, and autoreactive T cells are necessary for initiation of the disease. To study the immunopathological mechanisms operating in the synovium of arthritic mice, we isolated a proteoglycan (PG)-specific arthritogenic T-cell hybridoma, 5/4E8, and examined the presentation of PG to this T-cell hybridoma by mouse synovial cells and chondrocytes. Both cell types expressed very low levels of major histocompatibility complex (MHC) class II following isolation and culture and were unable to present PG to the hybridoma. However, following stimulation with interferon-gamma (IFN-gamma), both synovial cells and chondrocytes showed a marked increase in MHC class II expression and consequently were able to present PG very effectively. The PG-specific responses of the hybridoma were abrogated by an anti-Ia monoclonal antibody. Granulocyte-macrophage colony-stimulating factor (GM-CSF), one of the most abundant cytokines in the rheumatoid synovium, had no effect on the antigen-presenting capacity of synovial cells and chondrocytes, either on its own or together with IFN gamma.

Aggrecans↗
PubMed1995

Cellular mediators secreted by interfacial membranes obtained at revision total hip arthroplasty.

The interfacial membrane between implant and host-bone in aseptically loose total hip arthroplasties has a potential role in the etiology of local bone resorption and loosening of the prosthetic component. Inflammatory/potential "bone-resorbing" agents (cytokines/mediators) released by the cells of the interfacial membranes of loosened uncemented and cemented total hip arthroplasties were measured. Synovial tissues from patients with acute femoral neck fractures, patients with osteoarthritis, and cadavers without joint disorders were used as control subjects. Control synovial tissue from osteoarthritic patients secreted the highest levels of prostaglandin E2, interleukin-8, and tumor necrosis factor alpha. Interleukin-1 alpha was the only cytokine whose levels were elevated as much as 4-fold around uncemented implants compared with cemented implants, and up to 16-fold compared with control synovial tissue. An apparent inverse relation between interleukin-1 alpha and interleukin-6 interfacial membranes of total hip arthroplasties compared with control synovial tissues suggests a complex cellular mechanism through a cytokine/prostaglandin cascade; this may regulate the observed bone resorption in aseptic loosening.

Adult↗
PubMed1995

Anti-CD44 treatment abrogates tissue oedema and leukocyte infiltration in murine arthritis.

A ubiquitous cell adhesion receptor, CD44, preferentially binds hyaluronan, a polysaccharide macromolecule that is present in most extracellular matrices. Hyaluronan molecules have large hydrodynamic volumes that entrap substantial amounts of water and can therefore control tissue hydration (swelling). CD44 is overexpressed by synovial cells and leukocytes, and hyaluronan is overproduced in the rheumatoid synovium and in other inflammatory sites. Nevertheless, the role of the CD44-hyaluronan interaction during inflammation is unclear. Our evidence shows that the CD44 receptor plays a critical role in governing the migration of inflammatory leukocytes into the extravascular compartment of the synovium in murine arthritis. An anti-CD44 antibody induces a rapid loss of CD44 from both leukocytes and synovial cells and displays an inhibitory effect on cell-extracellular matrix interactions in the synovium. As a result, the administration of such an antibody abrogates tissue swelling and leukocyte infiltration, two major components of inflammation.

Amino Acid Sequence↗
PubMed1995

Antigen-specific B cells present cartilage proteoglycan (aggrecan) to an autoreactive T cell hybridoma derived from a mouse with proteoglycan-induced arthritis.

Cartilage proteoglycan (aggrecan)-induced polyarthritis in BALB/c mice is characterized by chronic inflammation and destruction of joint tissues similar to that observed in human rheumatoid arthritis. The immunization of mice with fetal human proteoglycan (PG) elicits specific antibodies to the immunizing antigen of which a population cross-reacts with native mouse PG. This (auto)antibody production is immediately followed by an explosive proliferation of autoreactive T cells, suggesting that PG-specific B cells may participate in antigen presentation of PG to autoreactive T cells. We therefore isolated B cells from the spleens and lymph nodes of PG-immunized mice and examined their ability to present PG to a PG-specific T cell hybridoma. The antigen-specific T cell responses elicited by B cells from PG-immunized mice (both arthritic and clinically asymptomatic) were markedly higher than those of non-immune mice and keyhole limpet haemocyanin (KLH)-immunized mice, and these B cells could present low PG concentrations. Levels of B cell presentation corresponded with the serum levels of PG-specific antibodies, implying that these B cells were presenting the PG specifically via their surface immunoglobulin. This B cell-T cell interaction was strongly dependent on MHC class II/T cell receptor (TCR), LFA-1/intercellular adhesion molecule-1 (ICAM-1) and CD28/B7 interactions, as antibodies to Ia, ICAM-1 and B7-2 (but not to B7-1) markedly reduced presentation. These data indicate that PG-specific B cells may play an essential role in governing the development of PG-induced arthritis.

Aggrecans↗
PubMed1995

Presentation of cartilage proteoglycan to a T cell hybridoma derived from a mouse with proteoglycan-induced arthritis.

Immunization of BALB/c mice with human fetal cartilage proteoglycan (PG) produces progressive polyarthritis, and T cells play key roles in the development of the disease. To gain an understanding of how PG is presented to autoreactive T cells by synovial antigen-presenting cells (APC), we examined the abilities of various syngeneic APC in presenting PG to a specific T cell hybridoma 5/4E8, derived from a mouse with PG-induced arthritis. A20 B lymphoma cells and spleen cells were strong presenters of PG, but synoviocytes and P388D1 macrophages could only present PG effectively after stimulation with interferon-gamma (IFN-gamma). The IFN-gamma exerted its effect by up-regulating both MHC class II and intercellular adhesion molecule-1 (ICAM-1) expression by these cells as neutralizing antibodies to Ia, LFA-1 and ICAM-1 inhibited presentation. Our studies also showed that synoviocytes and spleen cells took up and processed PG more rapidly than the cell lines. Cysteine and serine protease-dependent antigen presentation of PG was blocked at 4 degrees C, 18 degrees C and by chloroquine treatment, indicating that presentation required active uptake and processing in an acidic compartment, probably in lysosomes. Also, keratan sulphate-depleted and cyanogen bromide (CNBr) and 2-nitro-5-thiocyanobenzoic acid (NTCB)-cleaved PG elicited stronger T cell responses, as they were more easily processed than the native molecule. Furthermore, CNBr-generated peptides were presented by fixed APC, indicating that core protein fragments of cartilage PG can be presented directly by APC in context with MHC class II molecules.

Animals↗
PubMed1995

VDIPEN, a metalloproteinase-generated neoepitope, is induced and immunolocalized in articular cartilage during inflammatory arthritis.

The destruction of articular cartilage in immune inflammatory arthritic disease involves the proteolytic degradation of its extracellular matrix. The role of activated matrix metalloproteinases (MMPs) in the chondrodestructive process was studied by identifying a selective cleavage product of aggrecan in murine arthritis models initiated by immunization with either type II collagen or proteoglycan. We conducted semiquantitative immunocytochemical studies of VDIPEN341 using a monospecific polyclonal antibody requiring the free COOH group of the COOH-terminal Asn for epitope detection. This antibody recognizes the aggrecan G1 domain fragment generated by MMP [i.e., stromelysin (SLN) or gelatinase A] cleavage of aggrecan between Asn341-Phe342 but does not recognize intact aggrecan. VDIPEN was undetectable in normal mouse cartilage but was observed in the articular cartilage (AC) of mice with collagen-induced arthritis 10 d after immunization, without histological damage and clinical symptoms. This aggrecan neoepitope was colocalized with high levels of glycosaminoglycans (GAGs) in pericellular matrices of AC chondrocytes but was not seen at the articular surface at this early time. Digestion of normal (VDIPEN negative) mouse paw cryosections with SLN also produced heavy pericellular VDIPEN labeling. Computer-based image analysis showed that the amount of VDIPEN expression increased dramatically by 20 d (70% of the SLN maximum) and was correlated with GAG depletion. Both infiltration of inflammatory cells into the synovial cavity and early AC erosion were also very prominent at this time. Analysis of adjacent sections showed that both induction of VDIPEN and GAG depletion were strikingly codistributed within sites of articular cartilage damage. Similar results occurred in proteoglycan-induced arthritis, a more progressive and chronic model of inflammatory arthritis. These studies demonstrate for the first time the MMP-dependent catabolism of aggrecan at sites of chondrodestruction during inflammatory arthritis.

Amino Acid Sequence↗
PubMed1995

Mapping of arthritogenic/autoimmune epitopes of cartilage aggrecans in proteoglycan-induced arthritis.

Immunization of BALB/c mice with chondroitin sulfate-depleted proteoglycan (aggrecan) of fetal human cartilage produces progressive polyarthritis and ankylosing spondylitis. The development of the disease in genetically susceptible BALB/c mice is dependent upon the expression of both cell-mediated and humoral immune responses against the host mouse cartilage proteoglycan (PG). Although cartilage PGs from various species have many biochemical and immunological similarities, only a select group of PGs from fetal and newborn human, fetal pig and canine articular cartilages, human osteophytes and human chondrosarcomas are able to induce arthritis in BALB/c mice. Arthritis develops only in mice that also develop autoantibodies to self-cartilage PGs, although autoantibodies occasionally are present in non-arthritic animals as well. The protease-sensitive auto/arthritogenic epitope(s) is located in, or close to, the chondroitin sulfate (CS) attachment region of the PG molecule. The primary structure of the core protein is responsible for the autoimmune/arthritogenic effect of this select group of PGs, whereas the core protein epitopes are masked by glycosaminoglycan (GAG)-side chains. The CS side chains seem to inhibit antigen recognition in all aggrecans with arthritogenic potential, whereas a similar effect with keratan sulfate (KS) appears only in PGs of aging cartilages.

Aggrecans↗
PubMed1994

Mediators and autopathogenic effector cells in proteoglycan-induced arthritic and clinically asymptomatic BALB/c mice.

Proteoglycan (aggrecan)-induced arthritis is an autoimmune inflammatory animal model produced in genetically susceptible BALB/c mice. This animal model shows many similarities to human rheumatoid arthritis as indicated by clinical assessments, histopathological studies, and immunological parameters. The systemic immunization of mice with a select group of cartilage proteoglycans provokes immune responses to the immunizing antigen and then the production of cross-reactive antibodies to self proteoglycans. This is followed by an explosive proliferation of autoreactive T cells, especially in joint draining lymph nodes, accompanied by local (joint) inflammatory events. In the current experiments we found that lymphocytes from arthritic, or potentially arthritic but yet clinically asymptomatic animals, produced more IL-2 than those T cells obtained from animals immunized with nonarthritogenic PGs. In addition, synoviocytes isolated from prearthritic or arthritic animals produced several-fold more interleukin-1 beta (IL-1 beta) than cells from normal animals. Flow cytometric analysis indicated an autoantigen (mouse PG)-specific selective proliferation of surface Ig+/CD45R+ cells in prearthritic stages followed by the proliferation of predominantly T helper (CD4+) cells during and after the development of arthritis.

Animals↗
PubMed1994

Complete coding sequence, deduced primary structure, chromosomal localization, and structural analysis of murine aggrecan.

We have isolated and sequenced overlapping cDNA clones encoding the entire core protein of aggrecan (the large aggregating chondroitin sulfate/keratan sulfate proteoglycan of cartilage) from three chondrocyte cDNA libraries of BALB/c mice and localized the aggrecan gene in mouse chromosome 7. We determined 7386 bp of the cDNA sequence, including 132 and 854 nucleotides of 5' and 3' untranslated regions, respectively. The core protein precursor is 2132 amino acids long (M(r) 222,008), including a 19-residue secretory signal peptide. The overall amino acid sequence of the mouse aggrecan shows 91.6% identity to rat and 72.5% to human aggrecan. Comparison of the amino acid sequences of various domains and subdomain structures of mouse aggrecan to known sequences of other species and related proteins (versican, neurocan, link protein, and lymphocyte homing receptor CD44) revealed high levels of identity of the G1, G2, and G3 globular domains and relatively less conserved structures in the interglobular and glycosaminoglycan-attachment regions. Epidermal growth factor (EGF)-like module was detected in only a minor fraction of aggrecan clones, while the complement regulatory protein (CRP)-like domain was regularly expressed in all samples.

Aggrecans↗