Inflammation In Adjuvant-induced Arthritis Research Articles

Effect of Different Nuts Oil Consumption on Morphological Features and Some Biomarkers of Inflammation in Adjuvant-Induced Arthritis (AIA) Rat Model

This study evaluated the protective effect of different dietary ω-6/ω-3 ratios in oils obtained from various nuts (walnut, peanut, cashew, and hazelnut) against morphological features and markers of inflammation on an adjuvant-induced arthritis rat model. Rheumatoid arthritis (RA) was induced via intradermal injection of heat-killed Mycobacterium tuberculosis. Five groups of rats with RA (n = 5) were randomly categorized as follows: control positive, walnut oil group, peanut oil group, cashew nut oil group, and hazelnut oil group. Another five healthy rats served as a normal non-arthritic (control) group. We assessed the therapeutic effects by measuring arthritis scores during the experiment and serum inflammatory markers at the end of the study. The serum levels of the rheumatoid factor, TNF-α, IL-6, IL-1β, and PGE2, were significantly (p ≤ 0.05) reduced in all treatment groups. The daily consumption of nut oils ameliorates clinical and morphological abnormalities by inhibiting the inflammatory cells that produce inflammatory interleukins and eicosanoids.

Formulation of a dual drug-loaded nanoparticulate co-delivery hydrogel system and its validation in rheumatoid arthritis animal model

Rheumatoid arthritis (RA), a systemic autoimmune disease that dramatically affects patients’ quality of life. Given the intricacy of RA’s pathophysiology, no single treatment can completely halt the disease progression. Here, we attempted to treat RA holistically and synergistically by co-delivering methotrexate (MTX), a standard slow-acting anti-rheumatic drug, and phenethyl isothiocyanate (PEITC), a bioactive phytochemical, using a sodium alginate (SA)-pluronic F127 (PF-127) in situ hydrogel formulation. Therefore, in the current study, the co-delivery of MTX and PEITC in the nanoparticulate form could help enhance stability and solubility and facilitate greater penetration in the target arthritic tissues. The fabricated MTX NP and PEITC NE were found to have a minimum particle size, PDI, and good zeta potential. Results from in vitro release studies showed that MTX and PEITC were simultaneously released from the DD NP HG matrix over 6–7 days through diffusion and erosion mechanisms. An intra-articular (IA) injection of DD NP HG dramatically reduced chronic inflammation in adjuvant-induced arthritis (AIA) rats, delayed the onset of bone erosion, significantly reduced synovitis, and down-regulated the inflammatory cytokine expression. Most notably, the co-delivery strategy almost entirely restored the morphological features of the ankle joints of RA rats. The hepatic and renal function tests indicated good biological safety for DD NP HG in RA conditions. Taken together, these findings indicated that DD NP HG could achieve good anti-inflammatory activity and reverse cartilage disruption through a synergistic effect between two nanoparticulate forms of MTX and PEITC, which can effectively improve the drawbacks of their free forms.

Effects of local cryotherapy on systemic endothelial activation, dysfunction, and vascular inflammation in adjuvant-induced arthritis (AIA) rats

AimThis study explored the systemic vascular effects of local cryotherapy with a focus on endothelial changes and arterial inflammation in the model of rat adjuvant-induced arthritis (AIA).MethodsCryotherapy was applied twice a day on hind paws of AIA rats from the onset of arthritis to the acute inflammatory phase. Endothelial activation was studied in the aorta by measuring the mRNA levels of chemokines (CXCL-1, MCP-1 (CCL-2), MIP-1α (CCL-3)) and adhesion molecules (ICAM-1, VCAM-1) by qRT-PCR. Endothelial dysfunction was measured in isolated aortic and mesenteric rings. Aortic inflammation was evaluated via the mRNA expression of pro-inflammatory cytokines (TNF-α, IL-6) by qRT-PCR and leucocyte infiltration analysis (flow cytometry). Plasma levels of TNF-α, IL-6, IL-1β, IL-17A, and osteoprotegerin (OPG) were measured using Multiplex/ELISA.ResultsAIA was associated with an increased aortic expression of CXCL-1 and ICAM-1 as well as an infiltration of leucocytes and increased mRNA expression of IL-6, IL-1β, and TNF-α. Local cryotherapy, which decreased arthritis score and structural damages, reduced aortic mRNA expression of CXCL-1, IL-6, IL-1β, and TNF-α, as well as aortic infiltration of leucocytes (T lymphocytes, monocytes/macrophages, neutrophils) and improved acetylcholine-induced vasorelaxation in the aorta and mesenteric arteries. Plasma levels of IL-17A and OPG were significantly reduced by cryotherapy, while the number of circulating leucocytes was not. IL-17A levels positively correlated with endothelial activation and dysfunction.ConclusionIn the AIA model, local cryotherapy reduced systemic endothelial activation, immune cell infiltration, and endothelial dysfunction. Mechanistically, the reduction of circulating levels of IL-17A appears as the possible link between joint cooling and the remote vascular effects.

The ROS/GRK2/HIF-1α/NLRP3 Pathway Mediates Pyroptosis of Fibroblast-Like Synoviocytes and the Regulation of Monomer Derivatives of Paeoniflorin.

Hypoxia is an important factor in the development of synovitis in rheumatoid arthritis (RA). The previous study of the research group found that monomeric derivatives of paeoniflorin (MDP) can alleviate joint inflammation in adjuvant-induced arthritis (AA) rats by inhibiting macrophage pyroptosis. This study revealed increased levels of hypoxia-inducible factor- (HIF-) 1α and N-terminal p30 fragment of GSDMD (GSDMD-N) in fibroblast-like synoviocytes (FLS) of RA patients and AA rats, while MDP significantly inhibited their expression. Subsequently, FLS were exposed to a hypoxic environment or treated with cobalt ion in vitro. Western blot and immunofluorescence analysis showed increased expression of G protein-coupled receptor kinase 2 (GRK2), HIF-1α, nucleotide-binding oligomerization segment-like receptor family 3 (NLRP3), ASC, caspase-1, cleaved-caspase-1, and GSDMD-N. Electron microscopy revealed FLS pyroptosis after exposure in hypoxia. Next, corresponding shRNAs were transferred into FLS to knock down hypoxia-inducible factor- (HIF-) 1α, and in turn, NLRP3 and western blot results confirmed the same. The enhanced level of GSDMD was reversed under hypoxia by inhibiting NLRP3 expression. Knockdown and overexpression of GRK2 in FLS revealed GRK2 to be a positive regulator of HIF-1α. Levels of GRK2 and HIF-1α were inhibited by eliminating excess reactive oxygen species (ROS). Furthermore, MDP reduced FLS pyroptosis through targeted inhibition of GRK2 phosphorylation. According to these findings, hypoxia induces FLS pyroptosis through the ROS/GRK2/HIF-1α/NLRP3 pathway, while MDP regulates this pathway to reduce FLS pyroptosis.

PSTPIP2 attenuates joint damage and suppresses inflammation in adjuvant-induced arthritis

Proline–serine–threonine–phosphatase-interacting protein 2 (PSTPIP2) is related to inflammation. In this study, we investigated the function of PSTPIP2 in adjuvant-induced arthritis (AIA) by using adeno-associated virus (AAV) to overexpress PSTPIP2 in rat. AIA rats were developed by injecting Lewis rats with complete Freund's adjuvant (CFA) on day 0. AAV-empty or AAV-PSTPIP2, or PBS was administered intraarticularly into each knee joint on day 8 postinduction. All animals were killed at day 18 after adjuvant injection. WB was used to detect the expression of PSTPIP2 in rat synovial tissues. Fluorescence microscopy showed the transduction efficiency in synovial tissue. The morphology of arthritic joints was examined by HE, safranin O/fast green, or Toluidine blue staining. The bone destruction was examined via X-ray and micro-CT analysis. Immunohistochemical analysis or TRAP staining were used to investigate the role of PSTPIP2 in osteoclasts and the expression of PSTPIP2 in synovial tissue. RT-qPCR and ELISA were used to examine the expression of pro-inflammatory cytokines in synovial tissue or serum. AIA rats were found to have decreased PSTPIP2 expression and AIA-associated bone loss and inflammatory infiltration. We showed that administration of AAV-PSTPIP2 before arthritis onset significantly reduces the severity of AIA. PSTPIP2 was highly expressed in synovial cells. In addition, inflammatory responses and the number of osteoclasts were reduced with AAV-PSTPIP2 treatment. These findings demonstrate that PSTPIP2 may improve the severity of AIA by inhibiting the function of fibroblast-like synoviocytes, suppressing inflammation and reducing the number of osteoclasts.

Local cryotherapy improves adjuvant-induced arthritis through down-regulation of IL-6 / IL-17 pathway but independently of TNFα

ObjectivesLocal cryotherapy is widely and empirically used in the adjuvant setting in rheumatoid arthritis treatment, however its own therapeutic and anti-inflammatory effects are poorly characterized. We aimed to evaluate the effects of local cryotherapy on local and systemic inflammation in Adjuvant-induced arthritis, a murine model of rheumatoid arthritis.MethodsThe effects of mild hypothermia (30°C for 2 hours) on cytokine protein levels (Multiplex/ELISA) were evaluated in vitro in cultured rat adjuvant-induced arthritis patellae. In vivo, local cryotherapy was applied twice a day for 14 days in arthritic rats (ice: n = 10, cold gas: n = 9, non-treated: n = 10). At day 24 after the induction of arthritis, cytokine expression levels were measured in grinded hind paws (Q-RT-PCR) and in the plasma (Multiplex/ELISA).ResultsIn vitro, punctual mild hypothermia down-regulated IL-6 protein expression. In vivo, ice showed a better efficacy profile on the arthritis score and joint swelling and was better tolerated, while cold gas induced a biphasic response profile with initial, transient arthritis worsening. Local cryotherapy also exerted local and systemic anti-inflammatory effects, both at the gene and the protein levels: IL-6, IL-17A and IL-1β gene expression levels were significantly down-regulated in hind paws. Both techniques decreased plasma IL-17A while ice decreased plasma IL-6 protein levels. By contrast, we observed no effect on local/systemic TNF-α pathway.ConclusionsWe demonstrated for the first time that sub-chronically applied local cryotherapy (ice and cold gas) is an effective and well-tolerated treatment in adjuvant-induced arthritis. Furthermore, we provided novel insights into the cytokine pathways involved in Local cryotherapy’s local and systemic anti-inflammatory effects, which were mainly IL-6/IL-17A-driven and TNF-α independent in this model.

Ciclamilast ameliorates adjuvant-induced arthritis in a rat model.

We assessed the effect of a novel and selective phosphodiesterase 4 (PDE4) inhibitor, ciclamilast, on chronic inflammation in adjuvant-induced arthritis (AIA), a rat model of rheumatoid arthritis (RA), and acute inflammation in the rat and mouse model of carrageenan-induced paw edema and peritonitis. Our results showed that daily oral administration of ciclamilast at 1, 3, and 10 mg/kg dose-dependently inhibited the increase in hind paw volume of rats with AIA. The inhibition of paw edema was associated with inhibition of both the production of cytokines such as TNF-α, IL-1β, and IL-6 and cell infiltration assessed in subcutaneous paw tissue. Moreover, there was significantly less tissue destruction in the ciclamilast-treated rats compared to the vehicle-treated rats, as assessed by radiographic analysis and histopathological evaluation. In the two acute inflammation models, ciclamilast inhibited carrageenan-induced paw edema in rats and inflammatory cell migration into the peritoneal cavity in mice in a dose-dependent manner. These results not only suggest that ciclamilast, as a disease-modifying antirheumatic drug (DMARD), can attenuate RA but also provide proof of principle that a PDE4 inhibitor may be useful for the treatment of arthritis.

Therapeutic Effects of Gel Ointments Containing Tranilast Nanoparticles on Paw Edema in Adjuvant-Induced Arthritis Rats

Tranilast (TL), an antiallergic agent, has been clinically used in the treatment of bronchial asthma, although its clinical use has been limited by its poor solubility in water, photodegradation and systemic side effects. In this study, we prepared a gel ointment containing TL nanoparticles (TLnano gel ointment), and investigated its usefulness. In addition, we demonstrated the preventive effects of the TLnano gel ointment on inflammation in adjuvant-induced arthritis (AA) rats. The TLnano gel ointment was prepared using Bead Smash 12 (a bead mill) and additives including sodium docusate, 2-hydroxypropyl-β-cyclodextrin, methylcellulose and Carbopol 934; the mean particle diameter of the TL nanoparticles was 71.0±25.4 nm. In in vitro skin penetration experiments, the amount of penetrated TL, the penetration rate (Jc) and the penetration coefficient through the skin (Kp) of the TLnano gel ointment were significantly higher than those of a gel ointment containing TL microparticles (TLmicro gel ointment; particle diameter 50.5±26.3 µm). The TL concentrations in the skin tissue and plasma of rats receiving the TLnano gel ointment were also higher than in rats receiving the TLmicro gel ointment. In addition, the application of the TLnano gel ointment attenuated the increase in paw edema of the hind feet of AA rats in comparison with AA rats treated with the TLmicro gel ointment. These results suggest that TL nanoparticles can be applied to the formulation of a transdermal system, and that a transdermal formulation using TL nanoparticles might be a delivery option for the clinical treatment of RA.

Endomorphins in rheumatoid arthritis, osteoarthritis, and experimental arthritis

The opioid tetrapeptides endomorphins (EM)-1 and EM-2 are widely expressed in central nervous system and immune tissues of rats and humans. Their analgesic properties are well characterized but they also have anti-inflammatory properties. EM-1 significantly attenuated the onset of hindpaw inflammation in adjuvant-induced arthritis in rats. Immunohistochemical staining demonstrated the presence of EMs in T cells, macrophages, and fibroblasts in synovial tissues from patients with osteo- or rheumatoid arthritis (RA). In an ex vivo superfusion system, EM-1 potently inhibited the release of proinflammatory cytokines interleukin (IL)-6 and IL-8 from synovial tissues from patients with osteo- or RA. These results demonstrate that EMs are endogenously synthesized within human immune cells and have the potential to act as potent therapeutic agents in the treatment of chronic inflammatory disease. We discuss the clinical potential for EM analogues chemically modified to resist proteolytic degradation and identify modified protease-resistant analogues with enhanced bioactivity.

(118) CG100649, a novel dual-acting COX-2 and carbonic anhydrase inhibitor: Preclinical pharmacology

CG100649 is a novel dual-acting cyclooxygenase-2 (COX-2) and carbonic anhydrase (CA) inhibitor which is being developed for the treatment of osteoarthritis, rheumatoid arthritis, and acute pain. CG100649 has moderate COX-2 selectivity over COX-1, ranging from 15-fold in human cells (whole blood, platelets and macrophages) to 45-fold in mouse peritoneal macrophages. In the ex vivo COX-1 activity assay, CG100649 showed weaker COX-1 inhibitory activity than indomethacin in rat whole blood. CG100649 was a potent inhibitor of inflammation in adjuvant-induced arthritis and collagen-induced arthritis in Lewis rats (paw swelling ED50s were 0.10 and 0.22 mg/kg/day, respectively).

MF498 [N-{[4-(5,9-Diethoxy-6-oxo-6,8-dihydro-7H-pyrrolo[3,4-g]quinolin-7-yl)-3-methylbenzyl]sulfonyl}-2-(2-methoxyphenyl)acetamide], a Selective E Prostanoid Receptor 4 Antagonist, Relieves Joint Inflammation and Pain in Rodent Models of Rheumatoid and Osteoarthritis

Previous evidence has implicated E prostanoid receptor 4 (EP4) in mechanical hyperalgesia induced by subplantar inflammation. However, its role in chronic arthritis remains to be further defined because previous attempts have generated two conflicting lines of evidence, with one showing a marked reduction of arthritis induced by a collagen antibody in mice lacking EP4, but not EP1-EP3, and the other showing no impact of EP4 antagonism on arthritis induced by collagen. Here, we assessed the effect of a novel and selective EP4 antagonist MF498 [N-{[4-(5,9-diethoxy-6-oxo-6,8-dihydro-7H-pyrrolo[3,4-g]quinolin-7-yl)-3-methylbenzyl]sulfonyl}-2-(2-methoxyphenyl)acetamide] on inflammation in adjuvant-induced arthritis (AIA), a rat model for rheumatoid arthritis (RA), and joint pain in a guinea pig model of iodoacetate-induced osteoarthritis (OA). In the AIA model, MF498, but not the antagonist for EP1, MF266-1 [1-(5-{3-[2-(benzyloxy)-5-chlorophenyl]-2-thienyl}pyridin-3-yl)-2,2,2-trifluoroethane-1,1-diol] or EP3 MF266-3 [(2E)-N-[(5-bromo-2-methoxyphenyl)sulfonyl]-3-[5-chloro-2-(2-naphthylmethyl)phenyl]acrylamide], inhibited inflammation, with a similar efficacy as a selective cyclooxygenase 2 (COX-2) inhibitor MF-tricyclic. In addition, MF498 was as effective as an nonsteroidal anti-inflammatory drug, diclofenac, or a selective microsomal prostaglandin E synthase-1 inhibitor, MF63 [2-(6-chloro-1H-phenanthro[9,10-d]imidazol-2-yl)isophthalonitrile], in relieving OA-like pain in guinea pigs. When tested in rat models of gastrointestinal toxicity, the EP4 antagonist was well tolerated, causing no mucosal leakage or erosions. Lastly, we evaluated the renal effect of MF498 in a furosemide-induced diuresis model and demonstrated that the compound displayed a similar renal effect as MF-tricyclic [3-(3,4-difluorophenyl)-4-(4-(methylsulfonyl)phenyl)-2-(5H)-furanone], reducing furosemide-induced natriuresis by approximately 50%. These results not only suggest that EP4 is the major EP receptor in both RA and OA but also provide a proof of principle to the concept that antagonism of EP4 may be useful for treatment of arthritis.

Chronic inflammation modulates ghrelin levels in humans and rats.

The aim of this work was to investigate whether changes in plasma ghrelin, the recently discovered 28-amino acid gastric hormone that regulates growth hormone (GH) secretion and energy homeostasis, occur during inflammation in adjuvant-induced arthritis (AA) in rats. For completeness, ghrelin plasma levels were measured in rheumatoid arthritis (RA) patients. AA was induced in male Lewis rats using Freund's complete adjuvant. Animals were monitored for weight and food intake, every 2 or 3 days, along all time-course experiments. Plasma ghrelin concentrations in 31 RA patients and 18 healthy controls, as well as in rats, were determined by a specific double-antibody radioimmunoassay. Gastric ghrelin mRNA expression was evaluated by northern blot analysis. Human GH and insulin-like growth factor (IGF)-1 were determined by quantitative chemiluminescence assay. Compared with controls, arthritic rats gained significantly (P < 0.01) less body weight than controls until the end of the study, when a partial recovery occurred. Ghrelin plasma levels were significantly lower at day 7 after arthritis induction than in controls (AA 7 = 91.2 +/- 5.6 pg/ml vs controls = 124.75 +/- 5.9 pg/ml), but they recovered to control levels by day 15. RA patients had ghrelin plasma levels significantly lower than healthy controls (RA = 24.54 +/- 2.57 pg/ml vs 39.01 +/- 4.47 pg/ml of healthy controls; P = 0.0041). In AA, there is a compensatory variation of ghrelin levels that relates to body weight adjustments. Recovery of ghrelin levels in the latter stage suggests an adaptive response and may represent a compensatory mechanism under catabolic conditions. In RA patients, chronic imbalance in ghrelin levels suggests that this gastric hormone may participate, together with other factors, in alterations of metabolic status during inflammatory stress.

Spinal adenosine receptor activation inhibits inflammation and joint destruction in rat adjuvant-induced arthritis.

To examine the effect of spinal cord adenosine (Ado) receptor stimulation on rat adjuvant-induced arthritis (AIA). Long-term intrathecal (IT) catheters were implanted into rats to provide spinal access for drug delivery. Animals were immunized with complete Freund's adjuvant at the tail base. Eight days later and every other day thereafter until day 20, rats were treated IT with the selective Ado A1 receptor agonist cyclohexyladenosine (CHA) or vehicle. In some experiments, animals received an additional daily intraperitoneal injection of the nonselective Ado antagonist theophylline. Paw swelling was measured by water displacement plethysmometry. The effect of IT CHA on the activation of activator protein 1 (AP-1) was determined by electromobility shift assay. Spinal cord c-Fos expression was determined by immunohistochemistry. Spinal CHA significantly inhibited inflammation in AIA, with a mean +/- SEM 20.9 +/- 16.9% increase in paw swelling in the IT CHA group compared with 81.3 +/- 10.6% in the saline group. The antiinflammatory effect of CHA was mediated through Ado receptors since the effect was reversed by coadministration of systemic theophylline. In addition, radiographs showed significantly less bone and cartilage destruction in the CHA-treated animals. Synovial expression of AP-1, which is a key regulator of metalloproteinase expression, was lower in IT CHA-treated animals. C-Fos expression was localized to spinal laminae I-VI, with a modest decrease observed in the superficial laminae in IT CHA-treated rats. These data demonstrate that the spinal cord can regulate peripheral inflammation. Therapeutic strategies that target the central nervous system might be useful in arthritis.

Protective Effect of Prior Acute Immune Challenge, but Not Footshock, on Inflammation in the Rat

Previous studies have revealed that a single exposure to an acute stress or acute immune stimulus can produce long-lasting changes in the activity and responsiveness of the hypothalamo-pituitary–adrenal (HPA) axis. The HPA axis is believed to be an important component in determining the susceptibility and severity of inflammation in autoimmune disease models such as adjuvant-induced arthritis (AA). In the present study we have tested the hypothesis that a single exposure to either footshock or lipopolysaccharide (LPS) 3 weeks prior to adjuvant injection can alter susceptibility to AA. Changes in HPA axis parameters were also determined. The results demonstrated that prior exposure to LPS conferred resistance to inflammation in AA, which was not related to a delay in onset of inflammation but rather an alteration in susceptibility. In contrast, prior exposure to the acute stress of footshock did not alter susceptibility. HPA axis parameters were increased in adjuvant-injected rats whether inflammation was present or not. These data suggest that prior exposure to acute immune stimuli, but not to acute footshock stress, may alter susceptibility to inflammation in the rat AA model. These changes in susceptibility do not appear to be solely mediated by increases in HPA axis activity, which were apparent in all AA groups irrespective of the presence of inflammation.

The Effect of Hypothalamic Lesions on Hypothalamo-Pituitary-Adrenal Axis Activity and Inflammation in Adjuvant-Induced Arthritis

Adjuvant-induced arthritis (AA) was induced in control and in hypothalamic lesioned Piebald-Viral-Glaxo (PVG) rats. Following discrete paraventricular nucleus (PVN) lesions plasma corticosterone was increased 14 days after adjuvant injection as in controls, when hind paw inflammation was apparent. PVN lesion did not affect the severity of inflammation. In contrast, following medial basal hypothalamus (MBH) lesions adjuvant did not increase corticosterone levels and the increase in paw volume at day 14 was potentiated. Basal proopiomelanocortin (POMC) mRNA expression in the anterior lobe was unchanged by PVN lesions and decreased by MBH lesions. AA increased POMC mRNA in controls and in both PVN and MBH lesioned rats. After complete MBH lesion, surviving anterior pituitary tissue maintained morning levels of corticosterone.Thus, AA may activate the hypothalamo-pituitary-adrenal axis without the mediation of PVN neurones projecting to the median eminence. However, the loss of the corticosterone response to AA and the increase in severity of inflammation in the MBH lesioned rats suggests a central (non-PVN) component mediates effects of inflammation. Furthermore, the increase in POMC mRNA in the MBH lesioned AA rats suggests that part of this process is not mediated by releasing factors in the hypothalamo-hypophysial portal system, and that extrahypothalamic (peripheral) mediators act on the pituitary during chronic inflammation.

The Effect of Stress on Inflammation

Stress can either enhance or suppress aspects of the immune response, depending on the nature, duration, timing and intensity of the stressor. This paper focuses on the effects on inflammation of two behavioural stress paradigms: learned helplessness (LH) and the open-field test (OF), and of the immunological stressor lipopolysaccharide (LPS). We have observed that the onset and severity of inflammation in adjuvant-induced arthritis (AA) in the rat can be altered by experience of the LH paradigm, or by priming with LPS, but not by OF. In the LH test, some rats escape (LH(-)) and others do not (LH(+)). Despite the LH(-) group demonstrating a greater corticosterone response to the LH stressor compared to the LH(+) rats, they exhibited earlier onset and greater severity of AA. In contrast, intraperitoneal injection of LPS several weeks prior to induction of AA protected against inflammation. These results provide further evidence that environmental factors influence the etiology of at least one type of inflammation. The modulation of inflammation by a defined stressor suggests that understanding of the underlying mechanisms may provide a potential for novel therapies.

Onset and severity of inflammation in rats exposed to the learned helplessness paradigm.

To test the hypothesis that there is an association between susceptibility to inflammation and a hyporesponsive hypothalamo-pituitary-adrenal (HPA) axis. Animals were separated on the basis of behaviour in the learned helplessness (LH) paradigm into groups of LH(+) (i.e. animals which did not escape footshock) and LH(-) animals. Adjuvant-induced arthritis (AA) was subsequently induced in the LH(+) and LH(-) animals. Plasma corticosterone was significantly increased in response to the LH test in the LH(-) compared with the LH(+) rats. We observed an earlier onset and increased inflammation in the LH(-) rats in spite of the greater corticosterone response to the acute stress. We noted lower levels of plasma testosterone in the LH(-) animals suggesting a possible influence for this protective factor in AA. These data suggest that increased onset and severity of inflammation in AA is not a simple consequence of an attenuated HPA axis response to stress as proposed in the Lewis rat. Indeed we have observed the converse. Together these data suggest that the balance of pro- and anti-inflammatory factors released in response to stress may influence the progress of AA.

PECAM-1 and Leukosialin (CD43) Expression Correlate with Heightened Inflammation in Rat Adjuvant-Induced Arthritis

A hallmark of both adjuvant-induced arthritis (AIA) and rheumatoid arthritis is chronic joint inflammation characterized by ingress of leuko cytes into the inflamed synovial tissue. The timing of expression of adhesion molecules, which govern the ingress of leukocytes, is im portant in the orchestration of an inflammatory response. We examined the expression of vascular cell adhesion molecule-1 (VCAM-1), sialo adhesin, platelet and endothelial cell adhesion molecule-1 (PECAM-1), and leukosialin (CD43) in AIA, starting at adjuvant injection (day 0), through the peak of inflammation (day 18 postadjuvant injection), until day 54. VCAM-1 is constitutively expressed on the lining layer and ECs and its expression levels do not change throughout the progres sion of AIA. Sialoadhesin synovial tissue lining cell expression is decreased after adjuvant injection. In contrast, PECAM-1 expression is increased on synovial tissue lining cells on day 7 and is elevated through day 54 (peaking on day 54 with six-fold more cells expressing PECAM-1). PECAM-1 expression on endothelial cells peaks on day 7 with three-fold more cells expressing it, while on macrophages expression maximizes on day 25 with six-fold more cells expressing PECAM-1. CD43 expression is increased on synovial tissue lining cells, macrophages, neutrophils, and lymphocytes on days 18 and 25, before going back to basal levels. The increased expression of PECAM-1 and CD43 on leukocytes at the height of inflammation in AIA suggests important roles for these adhesion molecules in potentially binding their EC ligands resulting in leukocyte ingress into the synovial tissue.

Effects of stress on susceptibility and severity of inflammation in adjuvant-induced arthritis.

We have utilized the open field and learned helplessness (LH) models of psychological stress to determine whether a differential response to stress can affect the severity of adjuvant-induced arthritis (AA) within a single rat strain. In response to open field stress, the corticosterone response of the low emotivity rats was significantly lower than that of the high emotivity rats. In spite of the differential corticosterone response to stress, no significant difference was found in paw volumes between the AA high and low emotivity groups. In another study, rats were subjected to a learned LH paradigm and separated into two groups based on failed (LH+) or successful (LH-) avoidance. Plasma corticosterone levels in response to avoidable foot shock in the LH- rats were significantly greater than in the LH+ group. Following injection with adjuvant, paw inflammation occurred earlier and was more severe in the LH- rats compared to the LH+ group. These data show that rats with a greater tendency to avoid foot shock have more severe inflammation, despite having a greater corticosterone response to stress. We conclude that an increased corticosterone response to stress does not affect susceptibility to or severity of inflammation in AA. Indeed, in the LH model a more robust response to stress is associated with increased inflammation and earlier onset of the disease.

Alteration of central serotonin modifies onset and severity of adjuvant-induced arthritis in the rat.

Previous studies have determined that depletion of serotonin reduces the severity of hind-paw inflammation in adjuvant-induced arthritis (AA) in the rat. We wished to (i) test the hypothesis that this effect may be mediated, at least in part, through a central mechanism and (ii) to investigate further the pro-inflammatory role of serotonin we determined whether increasing serotonin using a selective serotonin reuptake inhibitor (SSRI), to increase serotonin availability at the active site of release, would increase inflammation. (i) Serotonin was depleted in the brain of rats with the selective neurotoxin 5'7'-dihydroxytryptamine. (ii) Rats were treated with an SSRI on days 10, 11 and 12 following adjuvant injection. Hind-paw inflammation was determined with plethysmometry as an index of severity of inflammation, and brain, pituitaries and blood were collected for assessment of changes in the hypothalamo -pituitary adrenal (HPA) axis. (i) Serotonin depletion significantly reduced hind-paw inflammation. (ii) SSRI-treated animals developed hind-paw inflammation sooner, and the severity was increased compared to vehicle-treated AA rats. The changes in the HPA axis associated with inflammation were partly reversed by this treatment. These data suggest a pro-inflammatory role for central serotonin in this disease model and indicate that treatment with SSRIs may exacerbate the development of inflammation.