Biomedical subjects

Katalin Mikecz

Publications and source records attributed to Katalin Mikecz.

At least 19 recordsLinked to original sources

PubMed2006

Expression of CD44 and L-selectin in the innate immune system is required for severe joint inflammation in the proteoglycan-induced murine model of rheumatoid arthritis.

Proteoglycan (PG)-induced arthritis, a murine model of rheumatoid arthritis, is characterized by autoimmunity against mouse cartilage PG and chronic joint inflammation. L-selectin (CD62L) and CD44 are major adhesion molecules on leukocytes that regulate their homing to lymph nodes and entry into inflamed tissues. In the present study, we studied the requirement for CD44 and CD62L expression for mediating lymphocyte homing, thus permitting the development of autoimmunity vs mediating the entry of leukocytes into the joints, thus allowing inflammation in PG-induced arthritis. We immunized wild-type, CD44 knockout (KO), CD62L KO, and double (CD44/CD62L) KO BALB/c mice with PG and monitored the effects of gene deficiencies on PG-specific immunity, arthritis severity, leukocyte trafficking, and the ability of lymphocytes to adoptively transfer disease to syngeneic SCID mice. Single and double KO mice demonstrated reduced PG-specific spleen cell proliferation, but the production of Th cytokines and autoantibodies was comparable in KO and wild-type mice. KO leukocytes had reduced ability to adhere tightly to the synovial endothelium in arthritic joints. This diminished leukocyte adhesion correlated with the magnitude of granulocyte (neutrophil) influx and the severity of inflammation, which were both reduced in the joints of KO mice. However, transfer of spleen cells from mildly arthritic KO donors to SCID hosts resulted in development of severe arthritis. Our results indicate that CD44 and CD62L expression in the cells of the innate immune system (granulocytes) is important for their efficient influx into the joints and also suggest that granulocytes play a crucial role in arthritis progression.

Animals↗

PubMed2006

Antigen-induced differential gene expression in lymphocytes and gene expression profile in synovium prior to the onset of arthritis.

To explore early signature genes playing critical roles in the initial steps in an autoimmune murine model of rheumatoid arthritis (RA) (proteoglycan (PG)-induced arthritis; PGIA), we performed gene expression profiling of "arthritogenic" spleen cells stimulated with cartilage PG, and compared them to differentially expressed genes, identified in joints prior to the onset of arthritis, and then in the acute and chronic phases of the disease. A total of 280 genes were up-regulated and 226 genes were suppressed in in vitro PG-stimulated lymphocytes at a minimum of 2-fold expression change. Functional gene classification identified several major clusters of biological activity. Expression of immunoglobulin genes (66 transcripts) was downregulated by approximately 3.7-fold, whereas most of the other genes with immune/inflammation-associated functions such as interleukins (IL-1, -2, -4, -6, -10, -12, -16, -17), chemokine receptors and their ligands (Cxcl1, Ccl2, 7, 8, 9, 10, 22, Ccr2, Ccr5), and major components of the complement cascade were upregulated. Using adoptive disease transfer with stimulated lymphocytes into SCID mice, followed by gene expression profiling of SCID paws, indicated that 37 genes were differentially expressed in yet non-inflamed (pre-arthritic) paws; these genes were related mostly to chemokine, IFN-gamma and TNF-alpha signaling. However, the majority of differentially expressed immune response-related genes were silent in pre-arthritic joints, and only 12 genes were found differentially expressed both in antigen (PG)-stimulated lymphocytes and in the synovium prior to the onset of arthritis. Most of these "arthritis-initiation" genes belonged to chemokine mediated cell motility. Transcripts of chemokine receptor 5 (Ccr5), chemokine ligand 7 (Ccl7) and IFN-gamma-inducible proteins (Ifi47) and GTP-ase 1 were expressed at the highest levels in both antigen-stimulated lymphocytes and pre-inflamed synovium, which suggests a key role of these genes in both lymphocyte maturation and arthritis initiation.

Adoptive Transfer↗

PubMed2005

T-cell recognition of differentially tolerated epitopes of cartilage proteoglycan aggrecan in arthritis.

Proteoglycan (PG) aggrecan, a major macromolecular component of cartilage, is highly immunogenic; it induces arthritis in genetically susceptible BALB/c mice. The present study maps the T-cell epitope repertoire of cartilage PG by identifying a total of 27 distinct T-cell epitopes. An epitope hierarchy, accounting for the different effector functions of PG-specific T cells, and determinant spreading, has been found. T-cell responses to four epitopes were associated with arthritis induction. Some of the T-cell epitopes were full T-cell activators, whereas a number of subdominant and cryptic epitopes proved to be partial activators in vitro, inducing either cytokine secretion or T-cell proliferation, but not both. A few T-cell epitopes of the core protein of cartilage PG were clearly recognized by T cells in PG-immunized arthritic animals, but the corresponding peptides did not induce T-cell responses when injected into naive BALB/c mice; thus these T-cell epitopes were designated as "conditionally immunogenic."

Aggrecans↗

PubMed2005

Two major interacting chromosome loci control disease susceptibility in murine model of spondyloarthropathy.

Autoimmune spondylitis was induced in BALB/c mice and their MHC-matched (BALB/c x DBA/2)F1 and F2 hybrids by systemic immunization with cartilage/intervertebral disk proteoglycan (PG). As in human ankylosing spondylitis, the MHC was the major permissive genetic locus in murine PG-induced spondylitis (PGIS). Two major non-MHC chromosome loci with highly significant linkage were found on chromosomes 2 (Pgis2) and 18 (Pgis1) accounting for 40% of the entire F2 trait variance. The dominant spondylitis-susceptibility allele for Pgis2 locus is derived from the BALB/c strain, whereas the Pgis1 recessive allele was present in the disease-resistant DBA/2 strain. The Pgis1 locus significantly affected the disease-controlling Pgis2 locus, inducing as high incidence of spondylitis in F2 hybrids as was found in the spondylitis-susceptible parent BALB/c strain. Additional disease-controlling loci with suggestive linkage were mapped to the chromosomes 12, 15, and 19. Severity of spondylitis in F2 mice positively correlated with serum levels of amyloid A, IL-6, and Pg-specific Abs, and showed negative correlation with Ag-induced T cell proliferation, IFN-gamma, IL-4, and TNF-alpha production. A major locus controlling serum IL-6 was found on chromosome 14 near osteoclast differentiation factor Tnfsf11. Locus on chromosome 11 near the Stat3 and Stat5 genes controlled serum level of the Ig IgG2a isotype. The two major genetic loci Pgis1 and Pgis2 of murine spondylitis were homologous to chromosome regions in human genome, which control ankylosing spondylitis in human patients. Thus, this animal model of experimentally induced spondylitis might facilitate the identification of spondylitis-susceptibility genes in humans.

Animals↗

PubMed2005

Effects of soluble metals on human peri-implant cells.

Despite reports associating tissue necrosis with implant failure, the degree to which processes, such as metal toxicity, negatively impact implant performance is unknown. We evaluated representative human peri-implant cells (i.e., osteoblasts, fibroblasts, and lymphocytes) when challenged by Al+3, Co+2, Cr+3, Fe+3, Mo+5, Ni+2, and V+3 chloride solutions (and Na+2 as a control) over a wide range of concentrations (0.01-10.0 mM). Cell responses were measured using proliferation assays, viability assays, and microscopic cell morphology assessments. Differential effects were found to be less a function of the cell type than of the composition and concentration of metal challenge. No preferential immunosuppression was demonstrated. Below 0.01 mM, no metal was toxic. The most toxic metals (i.e., Co, Ni, and V) reduced proliferation (IC50), and viability (LC50) and cell morphology of osteoblasts, fibroblasts, and lymphocytes by <50% at challenge concentrations <1 mM. All other metals tested required >5 mM to exact the same responses. Below 1 mM, these toxic metals also induced alterations in all cell morphology consisting of loss of filopodia or lamellipodia or changes in cell shape. Metals that were toxic at clinically relevant concentrations (less than previously reported values in peri-implant tissues/fluids) include Co (0.6 mM), Ni (0.8 mM), V (0.5 mM) for lymphocytes and Co (0.8 mM), V (0.3 mM), Al (1-5 mM), Fe (1-5 mM) for fibroblasts, and Co (0.8 mM), Ni (0.7 mM), V (0.1 mM) for osteoblasts. Only Co and V were toxic in vitro at concentrations below that detected in vivo in synovial fluid (V at 0.1 mM and Co at 0.8 mM for fibroblasts, and V at 0.4 mM and Co at 0.8 mM on osteoblasts). Thus, soluble Co and V released from Co- and Ti-based alloys, respectively, could be implicated as the most likely to mediate cell toxicity in the periprosthetic milieu.

Animals↗

PubMed2005

Genetic control of experimental spondylarthropathy.

OBJECTIVE: To characterize experimentally induced spondylarthropathy (SpA) in arthritis-susceptible inbred mice and in their F(1) and F(2) hybrid generations of susceptible and resistant mouse strains. METHODS: SpA was induced in susceptible BALB/c and C3H/HeJCr (C3H) strains of mice, and in their F(1) and F(2) generations derived from intercrosses with arthritis- and/or spondylitis-resistant DBA/2 and DBA/1 parent strains, by systemic immunization with cartilage proteoglycan (PG) aggrecan. The incidence and severity of PG-induced spondylitis (PGIS) were scored histologically, and these scores for spine involvement were correlated with serum antibody and cytokine levels and with in vitro T cell responses to cartilage PG. RESULTS: PGIS was induced by systemic immunization with cartilage PG in adjuvant, and approximately 60-70% of susceptible mouse strains and their F(2) hybrids developed spondylitis either with or without arthritis. Adjuvants, particularly those activating the innate immune system and enforcing the Th1 dominance, had significant effects on the outcome and progression of SpA. The DBA/1 strain appeared to carry genes protecting this strain and its F(1) and F(2) hybrids from spondylitis, whereas the DBA/2 strain, although resistant to PGIS, harbored genes permitting PGIS in its hybrid generations. Arthritis- and/or spondylitis-susceptible BALB/c and C3H parent strains and their F(2) hybrids exhibited the highest incidence and severity of spondylitis. CONCLUSION: PGIS, a murine model of autoimmune spondylitis, shows similarities to ankylosing spondylitis. Segregation of susceptibility to PG-induced arthritis (PGIA) from that to PGIS in different genetic crosses suggests that PGIA and PGIS are separate diseases. Therefore, this model allows for the elucidation of genetic components involved in the etiology of SpA, independent of those controlling the susceptibility to PGIA.

Aggrecans↗

PubMed2005

Visualization and in situ analysis of leukocyte trafficking into the ankle joint in a systemic murine model of rheumatoid arthritis.

OBJECTIVE: To describe the kinetics of leukocyte migration into a distal joint during the development of chronic inflammation in a murine model of rheumatoid arthritis (RA), to identify leukocyte subpopulations recruited in the synovial vessels, and to test in real time the effects of an antiinflammatory compound on leukocyte-endothelial cell interactions in the arthritic joint. METHODS: We used intravital video microscopy (IVM), which was adapted to the microcirculation of the mouse ankle, to monitor the kinetics of leukocyte-endothelium interactions (rolling and firm adhesion) during the onset and progression of proteoglycan-induced arthritis (PGIA), a chronic autoimmune model of RA. Subpopulations of rolling and adherent leukocytes were identified by in vivo immunostaining. Leukocyte extravasation into the ankle joint was verified histologically. RESULTS: Between the onset of arthritis and the beginning of the destructive phase of PGIA, we found a steady increase in the number of leukocytes that exhibited firm adherence to the endothelium of synovial vessels, which clearly underscores the chronic, self-perpetuating character of joint inflammation in this autoimmune model. We showed, however, that granulocytes, and not T cells, constituted the major cell population that was continuously recruited to the inflamed ankle. Using IVM, we could detect instant changes in leukocyte adhesion behavior in the synovial vessels of the arthritic joint upon administration of a compound that antagonizes leukocyte rolling. CONCLUSION: IVM of the microcirculation of the mouse ankle could become an essential tool for investigating the mechanisms that regulate leukocyte migration to the joint in systemic models of RA as well as for preclinical testing of antiinflammatory therapies.

Animals↗

PubMed2005

CD44, but not l-selectin, is critically involved in leucocyte migration into the skin in a murine model of allergic dermatitis.

CD44 and l-selectin (CD62L) are major adhesion receptors that mediate leucocyte recruitment at inflammatory sites and lymph nodes, by supporting cell rolling under blood flow. Both CD44 and CD62L have been implicated in inflammatory skin disorders, but their specific involvement in an immediate-type allergic reaction remains uncertain. We used mice deficient in CD44 or CED62L or both in order to determine whether one or both of these molecules were required for leucocyte extravasation in an atopic dermatitis-like allergic response. Wild-type (WT) mice and mice deficient in CD44, CD62L or both were immunized with ovalbumin (OVA). Inflammatory reaction in the ear was elicited once by means of intradermal injection of OVA. Effective sensitization of CD62L knockout (KO) mice required intraperitoneal antigen injection; however, OVA-specific T helper 2 (Th2)-type immune responses and IgE production in mice lacking CD44, CD62L or both were comparable to those in WT mice following intraperitoneal immunization. We employed intravital videomicroscopy to monitor the recruitment of fluorescence-labelled leucocytes to the ear tissue following challenge with OVA. The number of adherent leucocytes was significantly reduced in CD44 KO and CD44/CD62L double KO mice, indicating that CD44 was involved in firm adhesion, the committed step of leucocyte extravasation. Histology of the OVA-challenged ears showed a diminished leucocyte infiltration in the ears of CD44 KO and double KO mice. The results of our study demonstrate that CD44, but not CD62L, is required for leucocyte extravasation during a Th2-type inflammatory response.

Animals↗

PubMed2004

Cutting edge: Cbl-b: one of the key molecules tuning CD28- and CTLA-4-mediated T cell costimulation.

Cbl-b negatively regulates CD28-dependent T cell activation. In this report, we tested the hypothesis that CD28 and CTLA-4 have opposite roles in tuning T cell activation threshold by controlling the levels of Cbl-b protein expression. We demonstrate that CD28 costimulation potentiates TCR-induced Cbl-b degradation, whereas CTLA-4-B7 interaction is required for Cbl-b re-expression. In support of this finding, Cbl-b expression in CTLA-4 knockout (KO) T cells is significantly reduced, and treating CTLA-4KO mice with human CTLA-4Ig to block CD28-B7 interaction restores Cbl-b expression on T cells. Furthermore, CD28 and CTLA-4 costimulatory effects are compromised in Cbl-bKO T cells. These observations indicate that CD28 and CTLA-4 tightly regulate Cbl-b expression which is critical for establishing the threshold for T cell activation.

Abatacept↗

PubMed2004

Gene expression profiling in murine autoimmune arthritis during the initiation and progression of joint inflammation.

We present here an extensive study of differential gene expression in the initiation, acute and chronic phases of murine autoimmune arthritis with the use of high-density oligonucleotide arrays interrogating the entire mouse genome. Arthritis was induced in severe combined immunodeficient mice by using adoptive transfer of lymphocytes from proteoglycan-immunized arthritic BALB/c mice. In this unique system only proteoglycan-specific lymphocytes are transferred from arthritic mice into syngeneic immunodeficient recipients that lack adaptive immunity but have intact innate immunity on an identical (BALB/c) genetic background.Differential gene expression in response to donor lymphocytes that migrated into the joint can therefore be monitored in a precisely timed manner, even before the onset of inflammation. The initiation phase of adoptively transferred disease (several days before the onset of joint swelling) was characterized by differential expression of 37 genes, mostly related to chemokines, interferon-gamma and tumor necrosis factor-alpha signaling, and T cell functions. These were designated early arthritis 'signature' genes because they could distinguish between the naive and the pre-arthritic state. Acute joint inflammation was characterized by at least twofold overexpression of 256 genes and the downregulation of 21 genes, whereas in chronic arthritis a total of 418 genes with an equal proportion of upregulated and downregulated transcripts were expressed differentially. Hierarchical clustering and functional classification of inflammation-related and arthritis-related genes indicated that the most common biological activities were represented by genes encoding interleukins, chemokine receptors and ligands, and by those involved in antigen recognition and processing.

Acute Disease↗

PubMed2004

Expression of L-selectin, but not CD44, is required for early neutrophil extravasation in antigen-induced arthritis.

L (leukocyte)-selectin (CD62L) and CD44 are major adhesion receptors that support the rolling of leukocytes on endothelium, the first step of leukocyte entry into inflamed tissue. The specific contribution of L-selectin or CD44 to the regulation of cell traffic to joints in arthritis has not been investigated. We used CD44-deficient, L-selectin-deficient, and CD44/L-selectin double knockout mice to determine the requirement for these receptors for inflammatory cell recruitment during Ag-induced arthritis. Intraperitoneal immunization resulted in similar activation status and Ag-specific responses in wild-type and gene-targeted mice. However, extravasation of neutrophil granulocytes, but not the emigration of T cells, into the knee joints after intra-articular Ag injection was significantly delayed in L-selectin-deficient and double knockout mice. Intravital videomicroscopy on the synovial microcirculation revealed enhanced leukocyte rolling and diminished adherence in mice lacking either CD44 or L-selectin, but CD44 deficiency had no significant effect on the recruitment of L-selectin-null cells. Compared with wild-type leukocytes, expression of L-selectin was down-regulated in CD44-deficient cells in the spleen, peripheral blood, and inflamed joints, suggesting that reduced expression of L-selectin, rather than the lack of CD44, could be responsible for the delayed influx of granulocytes into the joints of CD44-deficient mice. In conclusion, there is a greater requirement for L-selectin than for CD44 for neutrophil extravasation during the early phase of Ag-induced arthritis.

Animals↗

PubMed2004

TSG-6 modulates the interaction between hyaluronan and cell surface CD44.

Interactions between CD44 and hyaluronan are implicated in the primary adhesion of lymphocytes to endothelium at inflammatory locations. Here we show that preincubation of hyaluronan with full-length recombinant TSG-6 or its Link module domain (Link_TSG6) enhances or induces the binding of hyaluronan to cell surface CD44 on constitutive and inducible cell backgrounds, respectively. These effects are blocked by CD44-specific antibodies and are absent in CD44-negative cells. Enhancement of CD44-mediated interactions of lymphoid cells with hyaluronan by TSG-6 proteins was seen under conditions of flow at shear forces that occur in post-capillary venules. Increases in the number of rolling cells were observed on substrates comprising TSG-6-hyaluronan complexes as compared with a substrate containing hyaluronan alone. In ligand competition experiments, cell surface-bound TSG-6-hyaluronan complexes were more potent than hyaluronan alone in inhibiting cell adhesion to immobilized hyaluronan. Link_TSG6 mutants with impaired hyaluronan binding function had a reduced ability to modulate ligand binding by cell surface CD44. However, some mutants that exhibited close to wild-type hyaluronan binding were found to have either reduced or increased activity, suggesting that some amino acid residues outside of the hyaluronan binding site might be involved in protein self-association, potentially leading to the formation of cross-linked hyaluronan fibers. In turn, cross-linked hyaluronan could increase the binding avidity of CD44 by inducing receptor clustering. The ability of TSG-6 to modulate the interaction of hyaluronan with CD44 has important implications for CD44-mediated cell activity at sites of inflammation, where TSG-6 is expressed.

Animals↗

PubMed2004

Achievement of a synergistic adjuvant effect on arthritis induction by activation of innate immunity and forcing the immune response toward the Th1 phenotype.

OBJECTIVE: To apply and analyze the mechanisms of action of dimethyldioctadecylammonium bromide (DDA), a powerful adjuvant that does not have the side effects of the conventionally used Freund's adjuvants, in proteoglycan-induced arthritis (PGIA) and collagen-induced arthritis (CIA). METHODS: PGIA and CIA were generated using standard immunization protocols with cartilage proteoglycan aggrecan (PG) or human type II collagen (CII) emulsified with Freund's complete adjuvant (CFA), and compared with PGIA and CIA generated using immunization protocols in which the same antigens were used in combination with the adjuvant DDA. Immune responses to immunizing and self PGs and CII, and the incidence, severity, and onset of arthritis were monitored throughout the experiments. In addition, a new, inexpensive, and powerful method of inducing arthritis using crude cartilage extracts is described. RESULTS: A significantly reduced onset period and a more severe arthritis were achieved in BALB/c mice immunized with cartilage PGs in DDA. PGs from bovine, ovine, and porcine cartilage, which otherwise have no effect or have only a subarthritogenic effect, and crude extracts of human osteoarthritic cartilage induced a 100% incidence with a very high arthritis score in BALB/c mice. The overall immune responses to either CII or PG were similar in antigen/CFA-immunized and antigen/DDA-immunized animals, but the Th1/Th2 balance shifted significantly toward a Th1 bias in DDA-injected animals with either PGIA or CIA. CONCLUSION: DDA, which was first used in autoimmune models, is a potent nonirritant adjuvant, which eliminates all undesired side effects of the Freund's adjuvants. DDA exerts a strong stimulatory effect via the activation of nonspecific (innate) immunity and forces the immune regulation toward Th1 dominance. These lines of evidence also suggest the possibility that seemingly innocuous compounds may exert an adjuvant effect in humans and may create the pathophysiologic basis of autoimmunity in susceptible individuals via the activation/stimulation of innate immunity.

Adjuvants, Immunologic↗

PubMed2004

Induction of arthritis in HLA-DR4-humanized and HLA-DQ8-humanized mice by human cartilage proteoglycan aggrecan but only in the presence of an appropriate (non-MHC) genetic background.

OBJECTIVE: To determine whether the rheumatoid arthritis (RA)-predisposing class II molecules of the major histocompatibility complex (MHC) can present cartilage proteoglycan (PG) aggrecan, and if so, to determine the epitope repertoire of the human cartilage PG in HLA-transgenic mice and determine whether HLA-transgenic mice develop arthritis in response to immunization with human cartilage PG. METHODS: Mice transgenic for HLA-DR2.Ab(0), DR3.Ab(0), DR4.Ab(0), and DQ8.Ab(0), lacking their own (mouse) class II antigens (Ab(0)), on the original (arthritis-resistant) and the arthritis-susceptible BALB/c backgrounds, were immunized with human cartilage PG. The T cell epitope repertoire presented by these class II MHC alleles was determined using a synthetic peptide library (143 peptides of the core protein of human cartilage PG), and arthritis development was monitored and compared in wild-type and HLA-transgenic/congenic BALB/c mice. RESULTS: Mice of the 4 HLA-transgenic lines, either on the original mixed, arthritis-resistant background or DR4.Ab(0)- and DQ8.Ab(0)-transgenic/congenic mice on the arthritis-susceptible BALB/c genetic background, responded well to PG immunization (as assessed by T cell responses and antibody and cytokine production), and a number of T cell epitopes along the core protein of human cartilage PG were identified. DR4.Ab(0)- and DQ8.Ab(0)-transgenic mice immunized with human cartilage PG developed arthritis, but only when these class II MHC molecules were present on the arthritis-susceptible (BALB/c) genetic background. CONCLUSION: A number of human cartilage PG epitopes can be presented by HLA alleles that predispose to the development of RA, but the epitopes of the cartilage PG presented by HLA-DR4 or HLA-DQ8 can induce arthritis only in the presence of an appropriate genetic (non-MHC) background.

Aggrecans↗

PubMed2004

Enhanced neutrophil extravasation and rapid progression of proteoglycan-induced arthritis in TSG-6-knockout mice.

OBJECTIVE: To gain insight into the mechanisms of the antiinflammatory effect of tumor necrosis factor alpha (TNFalpha)-induced protein 6 (Tnfip6) in arthritis, using Tnfip6-deficient animals. METHODS: TNFalpha-stimulated gene 6 (TSG-6) coding for Tnfip6 was disrupted. Tnfip6-deficient mice were backcrossed into proteoglycan-induced arthritis (PGIA)-susceptible BALB/c mice, and arthritis was induced by systemic immunization with cartilage proteoglycan (PG). Thioglycollate-induced sterile peritonitis was also assessed, to monitor the early events of neutrophil extravasation in wild-type and Tnfip6-deficient mice in the presence or absence of treatment with recombinant murine Tnfip6. RESULTS: The onset of PGIA was similar, but progression and severity were significantly greater, in Tnfip6-deficient mice compared with wild-type BALB/c mice. However, this was not associated with enhanced T or B cell responses to cartilage PGs, but rather, an early and more extensive infiltration of the synovium with neutrophil leukocytes was the most prominent histopathologic feature of PGIA in Tnfip6-deficient mice. This was accompanied by elevated serum levels of interleukin-6 and amyloid A, and significantly increased activities of the enzymes plasmin, myeloperoxidase, and neutrophil elastase in the inflamed paw joints of Tnfip6-null mice, when compared with that of the wild-type littermates. Loss of control over several components of inflammation resulted in extensive and rapid cartilage degradation, bone erosion, joint ankylosis, and deformities in Tnfip6-null animals. In support of the antiinflammatory effect of Tnfip6 via the inhibition of polymorphonuclear (PMN) cell efflux, neutrophil invasion during thioglycollate-induced peritonitis was 2-fold higher in Tnfip6-deficient animals than in wild-type animals, but was dramatically suppressed by intravenous injection of recombinant murine Tnfip6. CONCLUSION: Tnfip6 is a multifunctional antiinflammatory protein that is produced at the site of inflammation and can be retained by the hyaluronan-rich extracellular matrix. A major effect of Tnfip6 is the inhibition of the extravasation of PMN cells, predominantly neutrophils, into the site of inflammation, most likely via a CD44/hyaluronan/Tnfip6-mediated blocking mechanism.

Animals↗

PubMed2004

Proteoglycan aggrecan-induced arthritis: a murine autoimmune model of rheumatoid arthritis.

This chapter describes the major principals, methods, and immunization protocols for the induction of a systemic autoimmune arthritis in genetically susceptible murine strains. The model is called proteoglycan-induced arthritis (PGIA) because the antigenic/arthritogenic material is isolated from cartilage. This autoimmune systemic disease is induced by intraperitoneal immunization of either BALB/c or certain C3H colonies with cartilage proteoglycan, an abundant component in articular cartilage. The chapter presents (a) methodological details on how to purify cartilage proteoglycan aggrecan by cesium chloride gradient centrifugation; (b) substitution of this highly purified antigenic/arthritogenic material with a crude cartilage extract obtained from knee joint cartilages removed during joint replacement surgery; and (c) substitution of human cartilage proteoglycan with pig, dog, sheep, or bovine cartilage proteoglycans for arthritis induction. The cartilage proteoglycan aggrecan requires partial deglycosylation, and necessary materials, methods, and protocols are described. In addition, basic methods for measuring antigen-specific T-cell-dependent immune responses, antibody production, serum cytokine levels, and alternative solutions for adoptive transfers are also described.

Adoptive Transfer↗

PubMed2003

Differential recognition of altered peptide ligands distinguishes two functionally discordant (arthritogenic and nonarthritogenic) autoreactive T cell hybridoma clones.

Intravenous injection of a cartilage proteoglycan (aggrecan)-specific Th1 hybridoma clone 5/4E8 induced joint lesions similar to those seen in either primary or adoptively transferred arthritis in BALB/c mice. A sister clone, TA20, recognizing the same peptide epitope of human aggrecan and using the same Vbeta4 and Valpha1 segments, failed to induce joint inflammation. This study examines the fine epitope specificities of these two clones. Both 5/4E8 and TA20 hybridomas were generated using T cells from the same arthritic animal that has been immunized with human aggrecan, and both clones recognized peptides containing a consensus GRVRVNSAY sequence. However, flanking regions outside this nonapeptide sequence region had differential impact on peptide recognition by the two clones. Similarly, when single amino acid substitutions were introduced to the consensus sequence, significant differences were detected in the epitope recognition patterns of the T cell hybridomas. The 5/4E8 hybridoma showed greater flexibility in recognition, including a higher responsiveness to the corresponding self (mouse) aggrecan peptide, and produced more inflammatory cytokines (IFN-gamma and TNF-alpha), whereas hybridoma TA20 produced IL-5 in response to either human or mouse self peptide stimulation. These results demonstrate that, within the pool of immunodominant (foreign) peptide-activated lymphocytes, marked individual differences of degeneracy exist in T cell recognition, with possible implications to autopathogenic T cell functions.

Aggrecans↗

PubMed2003

IL-4 potentiates activated T cell apoptosis via an IL-2-dependent mechanism.

Activation-induced cell death (AICD) of T cells is one of the major mechanisms of peripheral tolerance. The regulation of AICD by IL-4 is poorly understood. In this study, we report that AICD in IL-4-deficient T cells is significantly reduced compared with that in wild-type T cells. This impaired AICD correlates with the failure to induce degradation of cellular FLIP. IL-4-mediated enhancement of AICD and cellular FLIP degradation requires a Janus kinase/STAT-6 signaling pathway. Unexpectedly, these effects of IL-4 could be blocked by a neutralizing anti-IL-2 Ab, and addition of rIL-2 could completely restore the defective AICD in IL-4-deficient T cells. Furthermore, IL-4 regulates the T cell thresholds for IL-2 signaling during AICD. These data suggest that IL-4 promotes AICD via an IL-2-dependent mechanism.

Adjuvants, Immunologic↗