Tissue Damage In Arthritis Research Articles

GM-CSF drives dysregulated hematopoietic stem cell activity and pathogenic extramedullary myelopoiesis in experimental spondyloarthritis

Dysregulated hematopoiesis occurs in several chronic inflammatory diseases, but it remains unclear how hematopoietic stem cells (HSCs) in the bone marrow (BM) sense peripheral inflammation and contribute to tissue damage in arthritis. Here, we show the HSC gene expression program is biased toward myelopoiesis and differentiation skewed toward granulocyte-monocyte progenitors (GMP) during joint and intestinal inflammation in experimental spondyloarthritis (SpA). GM-CSF-receptor is increased on HSCs and multipotent progenitors, favoring a striking increase in myelopoiesis at the earliest hematopoietic stages. GMP accumulate in the BM in SpA and, unexpectedly, at extramedullary sites: in the inflamed joints and spleen. Furthermore, we show that GM-CSF promotes extramedullary myelopoiesis, tissue-toxic neutrophil accumulation in target organs, and GM-CSF prophylactic or therapeutic blockade substantially decreases SpA severity. Surprisingly, besides CD4+ T cells and innate lymphoid cells, mast cells are a source of GM-CSF in this model, and its pathogenic production is promoted by the alarmin IL-33.

Distinct fibroblast subsets drive inflammation and damage in arthritis.

SummaryThe identification of lymphocyte subsets with non-overlapping effector functions has been pivotal to the development of targeted therapies in immune mediated inflammatory diseases (IMIDs)1,2. However it remains unclear whether fibroblast subclasses with non-overlapping functions also exist and are responsible for the wide variety of tissue driven processes observed in IMIDs such as inflammation and damage3–5. Here we identify and describe the biology of distinct subsets of fibroblasts responsible for mediating either inflammation or tissue damage in arthritis. We show that deletion of FAPα+ fibroblasts suppressed both inflammation and bone erosions in murine models of resolving and persistent arthritis. Single cell transcriptional analysis identified two distinct fibroblast subsets within the FAPα+ population: FAPα+ THY1+ immune effector fibroblasts located in the synovial sub-lining, and FAPα+ THY1- destructive fibroblasts restricted to the synovial lining layer. When adoptively transferred into the joint, FAPα+ THY1- fibroblasts selectively mediate bone and cartilage damage with little effect on inflammation, whereas transfer of FAPα+ THY1+ fibroblasts resulted in a more severe and persistent inflammatory arthritis, with minimal effect on bone and cartilage. Our findings describing anatomically discrete, functionally distinct fibroblast subsets with non-overlapping functions have important implications for cell based therapies aimed at modulating inflammation and tissue damage.

Mechanical strain determines the site-specific localization of inflammation and tissue damage in arthritis

Many pro-inflammatory pathways leading to arthritis have global effects on the immune system rather than only acting locally in joints. The reason behind the regional and patchy distribution of arthritis represents a longstanding paradox. Here we show that biomechanical loading acts as a decisive factor in the transition from systemic autoimmunity to joint inflammation. Distribution of inflammation and erosive disease is confined to mechano-sensitive regions with a unique microanatomy. Curiously, this pathway relies on stromal cells but not adaptive immunity. Mechano-stimulation of mesenchymal cells induces CXCL1 and CCL2 for the recruitment of classical monocytes, which can differentiate into bone-resorbing osteoclasts. Genetic ablation of CCL2 or pharmacologic targeting of its receptor CCR2 abates mechanically-induced exacerbation of arthritis, indicating that stress-induced chemokine release by mesenchymal cells and chemo-attraction of monocytes determines preferential homing of arthritis to certain hot spots. Thus, mechanical strain controls the site-specific localisation of inflammation and tissue damage in arthritis.

Non-classical monocytes as mediators of tissue destruction in arthritis

ObjectivesBone destruction in rheumatoid arthritis is mediated by osteoclasts (OC), which are derived from precursor cells of the myeloid lineage. The role of the two monocyte subsets, classical monocytes (expressing...

<i>In vitro</i> antioxidant and membrane stabilization activities of the fruit extract and fractions of <i>Tetrapleura tetraptera</i> (Schumach & Thonn.) Taub.

Arthritis is an inflammation of one or more joints. There is considerable experimental evidence linking lysosomal enzymes with tissue damage in arthritis. This study investigated anti-arthritic properties of Tetrapleura tetraptera (TT) using membrane stabilization assay (MSA). Powdered TT fruit sample was extracted by maceration in distilled methanol. Phytochemical screening was performed on powdered sample. The crude, partitioned extracts (n-hexane, chloroform and ethyl acetate) and seven pooled column chromatographic fractions were used for MSA (in vitro), using indomethacin and hypotonic solution as positive and negative controls. The total phenolic content (TPC) and antioxidant activity were determined. Phytochemical screening of TT showed the presence of important secondary metabolites. The ethyl acetate fraction showed the best protection of 98.77± 0.02% on red blood cells at 2.0 mg/mL. Fraction M at 0.5 mg/mL from EtOAC extract demonstrated significant (p<0.005) membrane stabilization activity (29.06±0.02%) compared to other pooled fractions (A, L, M, N, O, P, Q). The positive control, indomethacin, at 0.5 mg/mL, exhibited 56.02±0.02% membrane stabilization activity. The TPC ranged from 35.0-128.3 and 85.5 - 172.2 mg GAE/g for TT methanolic extract and pooled fractions, respectively. The IC50 values of the pooled fractions ranged from 41.16 ±0.02 to 91.56±0.04. Five compounds with different Rf values (0.58, 0.50, 0.62, 0.60, and 0.64) were isolated from the EtOAc fraction. The membrane stabilization activities demonstrated by the crude and ethyl acetate extracts and sub-fractions of Tetrapleura tetraptera may be attributed to antioxidant activities on the free radicals generated due to inflammatory reactions observed in most arthritic conditions.Keywords: Tetrapleura tetraptera; Arthritis; Membrane stabilization activity; Anti-oxidant; Lysosomes

Latest approaches on the management of OA in humans

The Sofat laboratory currently investigates the mechanisms responsible for pain and tissue damage in arthritis. Clinical studies include ‘Pain management in osteoarthritis using centrally acting analgesics’ (http://public.ukcrn.org.uk/search/StudyDetail.aspx?StudyID=12175) funded by the Rosetrees Trust. This clinical trial is testing whether drugs that inhibit pain processing pathways in the brain can help with pain in the hand caused by osteoarthritis. Other clinical studies include the ‘Pain Perception in Osteoarthritis’, or PAPO study, investigating tissue damage and pain in people undergoing knee replacement surgery for osteoarthritis. Both studies are on the UK NIHR portfolio of network-adopted studies. Our laboratory has a translational approach, with techniques including magnetic resonance imaging (MRI) to decipher regions of damage in affected joints and evaluating which brain regions are activated by arthritis pain. We also use non-invasive quantitative sensory testing (QST) methods to identify pain pathways. QST identifies sensation and pain thresholds by stimulating the skin. Our laboratory research uses a basic science mechanistic approach to investigate the influence of endogenous damage-associated molecular patterns (DAMPs) in driving chronic inflammation in joints. Molecules of interest include interaction of the DAMPs tenascin-C and fibronectin with oral pathogens.

Mechanisms of tissue damage in arthritis

The destruction of articular structures in the course of inflammatory arthritides such as rheumatoid arthritis (RA) or seronegative spondyloarthropathies is the most serious direct consequence of these diseases. Indeed, joint damage constitutes the "organ damage" of RA and-just like in all other diseases with organ involvement-such damage will usually be irreversible, cause permanent loss of function and subsequent disability. Research has identified a number of mechanisms and mediators of damage to articular structures such as bone and cartilage, ranging from proinflammatory cytokines, signal transduction pathways and cells types, which will be discussed in this review.

Myeloid skewing in murine autoimmune arthritis occurs in hematopoietic stem and primitive progenitor cells

AbstractSkewing toward myeloid cell production is often observed in chronic inflammation and autoimmune diseases. Herein, we determined whether persistent myeloid activation and proinflammatory output occurring in pathologic conditions is at the level of hematopoietic stem and primitive progenitor cells (HSPPCs). By using a mouse arthritis model, we found that even though HSPPCs in arthritis still retained the capacity to differentiate into different lineages, they acquired enhanced in vitro and in vivo propensity in a disease-dependent manner to generate myeloid cells, the key perpetrators of tissue damage in arthritis. This myeloid skewing was cell intrinsic, as arthritic HSPPCs up-regulate myeloid-specific transcripts including S100a8. Exogenous S100a8 promoted myeloid cell output from wild-type HSPPCs, suggesting mechanistic involvement of this gene in the myeloid priming that occurs in arthritic HSPPCs. Therefore, our results indicate that in arthritic mice, HSPPCs adopt a pathologic state that favors disease persistence.

Study On Mechanism of Bone Marrow Mesenchymal Stem Cell in Treating Patients with Rheumatoid Arthritis.

Abstract 4486 ObjectiveTo explore mechanism of Mesenchymal stem cells in treating Rheumatoid arthritis. Methods(1) MSCs were isolated from Bone marrow samples of Rheumatoid arthritis(RA)patients and purified by density gradient centrifagation and cultured in vitro. Morphology, immunophenotype, and proliferative property of bMSC and colony forming unit-fibroblast (CFU-F) were measured and analyzed. (2) In an in vitro co-culture system, MSCs were observed to modulate proliferation,activation, and maturation of T and B lymphocytes of Rheumatoid arthritis(RA)patients. The expression of IL-1?ATNF-a?ATGF-β were obviously changed.(3) Bone marrow- derived BMSCs of wistar rats were isolated and cultured in vitro routinely and the fourth passage as taken for identification of specific surface antigens by flowcytometry, then were labled with 5- BrdU in vitro. The model of collagen-induced arthritis (CIA) rats were established. 5- BrdU labled BMSCs were implanted through tail vein to model rats. At 4 weeks after BMSCs transplantation, immunohistochemical examinations were used to investigate BMSCs aggregate around the knee joints.and identify the contribution of bone marrow– derived cells to joints damage repair. Results(1) The culture expanded cells from RA patients presented a typical fibroblast-like morphology. Cells were positive for SH2(CD105),CD71,and CD44, but negative for CD45.Their proliferative capacity and CFU-F number were similar to those of bMSCs from healthy donors. (2) MSCs significantly inhibited T,B cell proliferation. MSCs also could down-regulating the levels of IL-1, TNF and up-regulating TGF-β. (3) Flow cytometry showed that BMSCs expressed CD44, CD71and CD105, but no CD45,CD34. At 4 weeks after the cells transplantation, the implanted cells were detected in the damaged joints of the model rats, which is not founded in normal knee joints of the rats'.and at same time there are more OPG(osteoprotegerin) positive. Conclusion(1) In the aspect of morphology, immuno -phenotypen, proliferative property and colony forming unit-fibroblast (CFU-F),MSCs from bone marrow of RA patients are not different from those of MSCs isolated from bone marrow of normal donors,MSCs from the bone marrow of RA patients have the potentiality in clinical application.(2) Human bone marrow MSCs inhibited Tcell and Bcell activation and proliferation in patients with RA in vitro. And these immuno –modulatory effects were not MHC-restricted. (3) bone marrow mesenchymal stem cells prevents tissue damage in arthritis. Allogeneic MSCs can engraft at sites of tissue damage,and prepair damage. That provided positive results for developing effective therapy for Rheumatoid arthritis. Disclosures:No relevant conflicts of interest to declare.

12/15-Lipoxygenase Counteracts Inflammation and Tissue Damage in Arthritis

Eicosanoids are essential mediators of the inflammatory response and contribute both to the initiation and the resolution of inflammation. Leukocyte-type 12/15-lipoxygenase (12/15-LO) represents a major enzyme involved in the generation of a subclass of eicosanoids, including the anti-inflammatory lipoxin A(4) (LXA(4)). Nevertheless, the impact of 12/15-LO on chronic inflammatory diseases such as arthritis has remained elusive. By using two experimental models of arthritis, the K/BxN serum-transfer and a TNF transgenic mouse model, we show that deletion of 12/15-LO leads to uncontrolled inflammation and tissue damage. Consistent with these findings, 12/15-LO-deficient mice showed enhanced inflammatory gene expression and decreased levels of LXA(4) within their inflamed synovia. In isolated macrophages, the addition of 12/15-LO-derived eicosanoids blocked both phosphorylation of p38MAPK and expression of a subset of proinflammatory genes. Conversely, 12/15-LO-deficient macrophages displayed significantly reduced levels of LXA(4), which correlated with increased activation of p38MAPK and an enhanced inflammatory gene expression after stimulation with TNF-alpha. Taken together, these results support an anti-inflammatory and tissue-protective role of 12/15-LO and its products during chronic inflammatory disorders such as arthritis.

Granulocyte-macrophage colony stimulating factor is anabolic and interleukin-1β is catabolic for rat articular chondrocytes

Objective: To study the effects of GM-CSF and IL-1β, both implicated in tissue damage in arthritis, on articular chondrocyte proliferation and metabolism, and to explore their agonist/antagonist effects. Methods: Chondrocytes were obtained from 1-month-old rats. First-passage monolayers were incubated for 24 h with or without GM-CSF and/or IL-1β, and labeled with 3H-thymidine, 35S–SO 4 and 14C-proline. Proteoglycan and collagen synthesis were analyzed by liquid chromatography and SDS–PAGE. Gene expression was measured by RT-PCR. Results: IL-1β exerts potent, and GM-CSF weak, inhibitory effects on DNA synthesis. GM-CSF strongly stimulates, and IL-1β inhibits, proteoglycan and collagen synthesis. IL-1β suppresses the effect of GM-CSF, and increases the release of radioactive molecules from pre-labeled cartilage fragments; GM-CSF decreases the IL-1β-induced effect. Interestingly, both cytokines induce the expression of each other’s gene. Conclusions: IL-1β appears to be a catabolic and anti-anabolic agent for chondrocytes, whereas GM-CSF is mainly anabolic, and blocks the IL-1β-induced catabolic effect. It is postulated that both agents are implicated in inflammation: IL-1β promotes tissue catabolism and destruction, whereas GM-CSF enhances tissue reconstruction.

Association of TIMP-4 gene polymorphism with the risk of osteoarthritis in the Korean population

Due to an imbalance in the MMP:TIMP ratio determined a tissue damage in arthritis, it is hypothesized that polymorphic variations of the TIMP genes are associated with regulation of the MMP:TIMP balance. To test this hypothesis, the presence of single nucleotide polymorphisms (SNPs) located in the human TIMP-2 and TIMP-4 genes was confirmed in the Korean RA and OA patients. We performed a case-control study comprising 109 unrelated Korean OA patients, 177 unrelated Korean RA patients and 175 healthy subjects. There were statistically significant differences in the genotype distribution and allele frequencies of the C/T polymorphism of TIMP-4 gene between OA and control groups (P = 0.0002 and P = 0.001, respectively). However, no significant association between TIMP-2 polymorphisms and OA was observed. Also, no difference was observed when allele or genotype frequencies of both TIMP-2 and TIMP-4 gene polymorphisms were compared between RA and controls. We demonstrated that the C/T polymorphism which is located on the 3'-untranslational regions of the TIMP-4 gene might be associated with susceptibility to OA patients.

Perforin deficiency attenuates collagen-induced arthritis

Collagen-induced arthritis (CIA), an approved animal model for rheumatoid arthritis, is thought to be a T cell-dependent disease. There is evidence that CD8+ T cells are a major subset controlling the pathogenesis of CIA. They probably contribute to certain features of disease, namely tissue destruction and synovial hyperplasia. In this study we examined the role of perforin (pfp), a key molecule of the cytotoxic death pathway that is expressed mainly in CD8+ T cells, for the pathogenesis of CIA. We generated DBA/1J mice suffering from mutations of the pfp molecule, DBA/1J-pfp-/-, and studied their susceptibility to arthritis. As a result, pfp-deficient mice showed a reduced incidence (DBA/1J-pfp+/+, 64%; DBA/1J-pfp-/-, 54%), a slightly delayed onset (onset of disease: DBA/1J-pfp+/+, 53 ± 3.6; DBA/1J-pfp-/-, 59 ± 4.9 (mean ± SEM), and milder form of the disease (maximum disease score: DBA/1J-pfp+/+, 7.3 ± 1.1; DBA/1J-pfp-/-, 3.4 ± 1.4 (mean ± SEM); P < 0.05). Concomitantly, peripheral T cell proliferation in response to the specific antigen bovine collagen II was increased in pfp-/- mice compared with pfp+/+ mice, arguing for an impaired killing of autoreactive T cells caused by pfp deficiency. Thus, pfp-mediated cytotoxicity is involved in the initiation of tissue damage in arthritis, but pfp-independent cytotoxic death pathways might also contribute to CIA.

Inhibition of adjuvant-induced arthritis by systemic tissue inhibitor of metalloproteinases 4 gene delivery.

An imbalance in the matrix metalloproteinase:tissue inhibitor of metalloproteinases (MMP:TIMP) ratio in favor of MMP appears to be an important determinant of tissue damage in arthritis. We undertook this study to explore whether reversal of this imbalance in favor of TIMP would alter this process and to examine the mechanism of this alteration. We administered human TIMP-4 by electroporation-mediated intramuscular injection of naked DNA using the rat adjuvant-induced arthritis (AIA) model. Intramuscular naked TIMP-4 gene administration resulted in high circulating TIMP-4 levels and completely abolished arthritis development in the rat AIA model. This inhibition was associated with significantly decreased MMP activity in the joint tissue as well as with significantly decreased serum and tissue tumor necrosis factor alpha levels and serum interleukin-1alpha levels compared with animals with arthritis. The mutation of cysteine at position 1 of TIMP-4 failed to block the development of AIA. Our data indicate that TIMP-4 is a potent antiinflammatory agent, and that its antiarthritis function may be mediated by MMPs. Arthritis-inhibiting effects of TIMP-4 may suggest a unique application of this gene therapy method for arthritis.

Leukaemia Inhibitory Factor (LIF) Suppresses Proteoglycan Synthesis in Porcine and Caprine Cartilage Explants

Leukaemia Inhibitory Factor (LIF) has been implicated in connective tissue damage in arthritis. We have previously shown that LIF stimulates proteoglycan release in pig cartilage explants. The aim of this study was to determine whether LIF modulates proteoglycan synthesis in vitro. The methods used were as follows: slices of pig and goat articular cartilage were incubated overnight in Dulbecco's modification of Eagles medium (DMEM), supplemented with 5% foetal calf serum (FCS) and then cultured for 48 h without FCS and either no cytokines (negative control) or LIF. During the final 6 h the tissue was cultured in sulphate free DMEM containing 35SO4. The radioactivity in the medium and tissue was determined in cetylpyridinium chloride precipitates. Biosynthetic activity was expressed as DPM per mg wet weight of cartilage. Dose-dependent suppression of proteoglycan synthesis was observed with murine and human recombinant LIF in pig and goat cartilage. The degree of inhibition was similar to the maximal suppression observed with IL-lα, but was not IL-1 dependent. In conclusion, LlF is a potent inhibitor of proteoglycan synthesis in cultured pig and goat articular cartilage.