Polydatin Prevents Calcium Pyrophosphate Crystal-Induced Arthritis in Mice.

BACKGROUND: Polydatin is a stilbenoid with important antioxidant, anti-inflammatory, and immunomodulating properties. The aim of this study was to assess the anti-inflammatory preventive effect of polydatin in the mouse model of acute arthritis induced by calcium pyrophosphate (CPP) crystals. METHODS: Acute arthritis was induced by the injection of a suspension of sterile CPP crystals into the ankle joint of Balb/c mice. Animals were randomized to receive polydatin or colchicine (the control drug) according to a prophylactic and a therapeutic protocol. The primary outcome was the variation of ankle swelling obtained after crystal injection and treatment, while histological parameters such as leukocyte infiltration, IL-1ß and CXCL1 levels and tissue expression were considered as secondary outcomes. RESULTS: Prophylactic treatment with PD significantly diminished ankle swelling after 48 h from crystal injection. Secondary outcomes such as leukocyte infiltration, necrosis, edema, and synovitis were also decreased. PD caused a reduction in circulating levels of IL-1ß and CXCL1, as well as their tissue expression. By contrast, the therapeutic administration of PD did not have any beneficial effect. CONCLUSIONS: PD can effectively prevent acute inflammatory response to crystals in the mouse model of CPP crystal-induced arthritis. These results suggest that this bioactive compound might be used in the prevention of crystal-induced acute attacks in humans.

FM0807 decelerates experimental arthritis progression by inhibiting inflammatory responses and joint destruction via modulating NF-κB and MAPK pathways.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic articular synovial inflammatory disease. The precise etiology underlying the pathogenesis of RA remains unknown. We aimed to investigate the inhibitory effect of curcumin analog FM0807 (curcumin salicylate monoester, 2-hydroxy-, 4-[(1E,6E)-7-(4-hydroxy-3-methoxyphenyl)-3,5-dioxo-1,6-heptadien-1-yl]-2-methoxyphenyl ester) on experimental RA and investigate its possible mechanisms of action. METHOD: Rats with Freund's complete adjuvant (FCA)-induced arthritis (AIA) were administered aspirin (0.1 mmol.kg-1), curcumin (0.1 mmol.kg-1), FM0807 (0.1, 0.2 mmol.kg-1) and vehicle via gastric gavage, from days 7 to 21, once daily. The hind paw volume and arthritis index (AI) were measured, and radiographic and histological examinations were performed. Twenty-one days later, the animals were killed and left ankle joints were removed to measure protein expression of the elements of the nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathway by Western blot analysis. The enzyme-linked immunosorbent assay (ELISA) was employed to measure synovial fluid levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-1ß and IL-10. RESULTS: Compared with AIA group, FM0807 reduced the AI and swelling of the injected hind paw in a dose-dependent manner, and inhibited increases in inflammatory cell infiltration, pannus formation and cartilage destruction. FM0807 also potently attenuated the increase in the expression of inflammatory factors TNF-α, IL-6 and IL-1ß in synovial fluid, while IL-10 levels were also elevated. FM0807 significantly suppressed phosphorylation of extracellular-signal-regulated kinase (ERK) 1/2 (ERK1/2), c-Jun-N-terminal kinase (JNK) 1/2 (JNK1/2), p38MAPK, inhibitor of NF-κB kinase (IKK), IκB and NF-κB p65 protein, (all P<0.05), which displayed more potential effects compared with those of the aspirin and curcumin groups. CONCLUSION: FM0807 exerts its therapeutic effects on RA by inhibiting cartilage degeneration. FM0807 treatment might be an effective therapeutic approach for RA.

Increased ILC3s associated with higher levels of IL-1ß aggravates inflammatory arthritis in mice lacking phagocytic NADPH oxidase.

The role of redox regulation in immune-mediated arthritis has been previously described. However, the relationship between innate immune cells, including innate lymphoid cells (ILCs) and phagocyte-derived ROS, in this process remains unclear. Here, we characterize ILCs and measure the IL-1 family cytokines along with other cytokines relevant to ILC functions and development in serum-induced arthritic joints in wild type and phagocytic NADPH oxidase (NOX2)-deficient Ncf1-/- mice. We found more severe serum-induced joint inflammation and increased NCR+ ILC3s in inflamed joints of Ncf1-/- mice. Furthermore, in vitro stimulation with IL-1ß on Tbet+ ILC1s from joints facilitated their differentiation into ROR-γt+ ILC3s. Moreover, treatment with IL-1 antagonists effectively lowered the proportions of NCR+ ILC3s and IL-17A producing ILC3s in Ncf1-/- arthritic mice and ameliorated the joint inflammation. These results suggest that NOX2 is an essential regulator of ILC transdifferentiation and may mediate this process in a redox-dependent manner through IL-1ß production in the inflammatory joint. Our findings shed important light on the role of ILCs in the initiation and progression in tissue inflammation and delineate a novel innate immune cell-mediated pathogenic mechanism through which redox regulation may determine the direction of immune responses in joints.

Berberis aristata Ameliorates Adjuvant-Induced Arthritis by Inhibition of NF-κB and Activating Nuclear Factor-E2-related Factor 2/hem Oxygenase (HO)-1 Signaling Pathway.

The present study was carried out to investigate the anti-arthritic activity of Berberis aristata hydroalcoholic extract (BAHE) in formaldehyde-induced arthritis and adjuvant-induced arthritis (AIA) model. Arthritis was induced by administration of either formaldehyde (2% v/v) or CFA into the subplantar surface of the hind paw of the animal. In formaldehyde-induced arthritis and AIA, treatment of BAHE at doses 50, 100 and 200 mg/kg orally significantly decreased joint inflammation as evidenced by decrease in joint diameter and reduced inflammatory cell infiltration in histopathological examination. BAHE treatment demonstrated dose-dependent improvement in the redox status of synovium (decrease in GSH, MDA, and NO levels and increase in SOD and CAT activities). The beneficial effect of BAHE was substantiated with decreased expression of inflammatory markers such as IL-1ß, IL-6, TNF-R1, and VEGF by immunohistochemistry analysis in AIA model. BAHE increased HO-1/Nrf-2 and suppressed NF-κB mRNA and protein expression in adjuvant immunized joint. Additionally, BAHE abrogated degrading enzymes, as there was decreased protein expression of MMP-3 and -9 in AIA. In conclusion, we demonstrated the anti-arthritic activity of Berberis aristata hydroalcoholic extract via the mechanism of inhibition of NF-κB and activation of Nrf-2/HO-1.

Idiopathic haemarthrosis in eight horses.

OBJECTIVES: To review eight horses diagnosed with idiopathic haemarthrosis and to describe the intra-articular use of yttrium-90 ((90) Y) and methylprednisolone acetate (MPA) in recurrent haemarthrosis cases. DESIGN: Retrospective case series. METHOD: The medical records, diagnostic images, histopathology and outcome of all horses diagnosed with idiopathic haemarthrosis between 1998 and 2010 were reviewed. RESULTS: Four Thoroughbred racehorses with haemarthrosis of the antebrachiocarpal joint had severe acute lameness (median, grade 4) and marked joint effusion after high-speed exercise. Another four horses (2 Thoroughbred racehorses, 1 Standardbred racehorse, 1 Warmblood) had haemarthrosis of the tarsocrural joint and presented with mild, intermittent lameness (median, grade 1) and marked, persistent joint effusion. Six of the eight horses had recurrent haemarthrosis prior to treatment. Radiographic and nuclear scintigraphic examinations did not identify bone pathology. Diagnostic arthroscopy (7 cases) identified grossly hypertrophied yellow/brown discoloured synovium. Synovial histopathology of these cases revealed chronic synovial hyperplasia with severe haemosiderosis and granulomatous inflammatory reaction of varying severity. All horses underwent rest, bandaging and phenylbutazone administration. Two horses had subtotal mechanical synovectomy, four horses had intra-articular administration of (90) Y and MPA, and one horse underwent both treatments. Seven cases returned to their previous use (median time, 7 months). Haemarthrosis recurred in three horses, two of which had received the (90) Y and MPA treatment. CONCLUSION: Idiopathic haemarthrosis should be considered a differential for acute and recurrent joint related lameness and effusion. Recurrence appears not uncommon and the use of intra-articular (90) Y and MPA in conjunction with a conservative management treatment protocol warrants further evaluation.

Prostaglandin E(2) binding peptide screened by phage displaying: a new therapeutic strategy in rheumatoid arthritis.

OBJECTIVE: To investigate the therapeutic potential and mechanism of action of the mimotope of PGE(2) receptor EP4 (PBP, named by our team) screened by phage displaying technique in the treatment of adjuvant-induced arthritis (AA). METHODS: Freund's complete adjuvant-induced arthritis was induced in Wistar rats. At the first clinical sign of disease, mice were given with daily injections of PBP or saline for 21 days. Disease progression was monitored by measurement of paw swelling. Inflammation and joint destruction were assessed histologically. The IL-1ß and TNF-α were studied by ELISA in the ankle steeps of arthritis model. The degree of proliferation and apoptosis of synoviocytes of RA patients were assessed by CCK-8 kit and AnnexinV-FITC/PI respectively. RESULTS: PBP-treated animals displayed significantly less cartilage and bone destruction than model controls. Tumor necrosis factor α and IL-1ß expression were reduced after PBP treatment. The proliferation and apoptosis of synoviocytes of RA patients were influenced by PBP. CONCLUSIONS: The data support the view that PBP is a potential therapy for RA that may help to diminish both joint inflammation and destruction. And the activities of PBP are related with the effect on synoviocytes directly.

Cancellous bone formation response to simulated resistance training during disuse is blunted by concurrent alendronate treatment.

The purpose of this study was to assess the effectiveness of simulated resistance training (SRT) exercise combined with alendronate (ALEN) in mitigating or preventing disuse-associated losses in cancellous bone microarchitecture and formation. Sixty male Sprague-Dawley rats (6 months old) were randomly assigned to either cage control (CC), hind limb unloading (HU), HU plus either ALEN (HU + ALEN), SRT (HU + SRT), or a combination of ALEN and SRT (HU + SRT/ALEN) for 28 days. HU + SRT and HU + SRT/ALEN rats were anesthetized and subjected to muscle contractions once every 3 days during HU (four sets of five repetitions, 1000 ms isometric + 1000 ms eccentric). Additionally, HU + ALEN and HU + SRT/ALEN rats received 10 µg/kg of body weight of ALEN three times per week. HU reduced cancellous bone-formation rate (BFR) by 80%, with no effect of ALEN treatment (-85% versus CC). SRT during HU significantly increased cancellous BFR by 123% versus CC, whereas HU + SRT/ALEN inhibited the anabolic effect of SRT (-70% versus HU + SRT). SRT increased bone volume and trabecular thickness by 19% and 9%, respectively, compared with CC. Additionally, osteoid surface (OS/BS) was significantly greater in HU + SRT rats versus CC (+32%). Adding ALEN to SRT during HU reduced Oc.S/BS (-75%), Ob.S/BS (-72%), OS/BS (-61%), and serum TRACP5b (-36%) versus CC. SRT and ALEN each independently suppressed a nearly twofold increase in adipocyte number evidenced with HU and inhibited increases in osteocyte apoptosis. These results demonstrate the anabolic effect of a low volume of high-intensity muscle contractions during disuse and suggest that both bone resorption and bone formation are suppressed when SRT is combined with bisphosphonate treatment.

Protective effect of eotaxin-2 inhibition in adjuvant-induced arthritis.

Eotaxin-2 is a potent chemoattractant for eosinophils, basophils and T helper type 2 (Th2) lymphocytes. The eotaxin-2/CCL24 receptor CCR3 is expressed in human brain, skin, endothelium and macrophages. The aim of the current study was to evaluate the protective effect of a monoclonal anti-eotaxin-2 antibody on the development of adjuvant-induced arthritis in rats (AIA). Adjuvant arthritis was induced in Lewis rats by intradermal injection of incomplete Freund's adjuvant +Mycobacterium tuberculosis. Rats were treated by intraperitoneal (i.p.) injection with three monoclonal antibodies against eotaxin-2 (G7, G8, D8) three times per week. Controls were treated with total mouse immunoglobulin G (IgG), methotrexate (MTX) or phosphate-buffered saline (PBS). Arthritis severity was evaluated by measuring ankle swelling, arthritic score, whole animal mobility and body weight. Sample joints were obtained for pathological evaluation and postmortem X-ray of ankle joints was performed to document erosions. Significant inhibition of arthritis was observed in rats treated with anti-eotaxin-2 antibodies compared to those treated with immunoglobulin or PBS. Inhibition was manifest in ankle diameter, arthritic score and mobility score. The antibody marked D8 showed the greatest efficacy. The effect was observed both in animals treated before the appearance of arthritis and in those where treatment was begun after development of joint inflammation. Combined treatment with D8 and MTX caused additional protection. Significant reduction of inflammation in D8-treated animals was also demonstrated in pathological and X-ray examinations. Inhibition of eotaxin-2 by monoclonal antibodies has a significant protective effect in adjuvant arthritis. These results may introduce a novel therapeutic target in rheumatoid arthritis and additional inflammatory joint disorders.

Double blind investigation of the effects of oral supplementation of combined glucosamine hydrochloride (GHCL) and chondroitin sulphate (CS) on stride characteristics of veteran horses.

REASONS FOR PERFORMING STUDY: Oral chondroprotective supplements are commercially popular for veteran (and other athletic or arthritic) horses prone to joint degeneration, yet lack conclusive scientific support. OBJECTIVES: To quantify the effects of an oral joint supplement (combination glucosamine hydrochloride (GHCL), chondroitin sulphate (CS) and N-acetyl-D-glucosamine) in vivo on stride parameters of veteran horses. METHODS: Twenty veteran horses were randomly assigned to a treatment (n = 15) or placebo group (n = 5). Pre-treatment gait characteristics were recorded at trot using digital video footage (50 Hz). The range of joint motion, stride length, and swing and stance duration were assessed using 2-dimensional motion analysis. Treatment (or placebo) was administered daily for 12 weeks at the manufacturer's recommended dosage. Gait was reassessed every 4 weeks using the pre-treatment protocol. Double blind procedure was implemented throughout. Relationships between variables were analysed using General Linear Model. RESULTS: Differences occurred in the treated horses by week 8. Range of joint motion increased significantly in the elbow (P<0.05), stifle and hind fetlock (P<0.01). Stride length increased significantly (P<0.05) with treatment. Swing duration was significantly increased at week 12 (P<0.05), whilst stance duration remained constant. CONCLUSION: The oral chondroprotective offered symptomatic relief to veteran horses, evidenced by improved stride characteristics. POTENTIAL RELEVANCE: Oral GHCL and CS supplementation may improve welfare by alleviating symptoms of degenerative joint disease.

Chronic intrathecal baclofen treatment and withdrawal: I. Changes in ankle torque and hind limb posture in normal rats.

This study evaluated reflex excitability and locomotor changes during chronic intrathecal infusion of the GABAb agonist baclofen (ITB) and its withdrawal, in the rat. We observed sustained velocity dependent decreases in ankle torque during four weeks of ITB treatment. These changes were correlated with a significant reduction of the EMG burst magnitude time locked to the dynamic phase of ankle dorsiflexion during the first ITB treatment week. However, a considerable recovery of EMG magnitude was observed during the third and fourth weeks of treatment. During baclofen withdrawal, significantly increased velocity dependent ankle torque was observed for 4 weeks. These increases in ankle torque were correlated with increased magnitudes of EMG time locked to the dynamic phase of ankle rotation. Measures of hind limb axis and base of support were obtained using analysis of footprints on a treadmill during ITB treatment and withdrawal periods. During ITB treatment and for up to 7 weeks of withdrawal, hindlimb axis and base of support were significantly altered compared with vehicle controls. These studies were performed to provide a foundation for evaluation of treatment and withdrawal in the setting of experimental chronic contusion spinal cord injury.

Effects of continuous intra-articular infusion of gentamicin on synovial membrane and articular cartilage in the tarsocrural joint of horses.

OBJECTIVE: To determine the effects of a continuous intra-articular infusion of gentamicin on the synovial membrane and articular cartilage in the tarsocrural joint of horses. ANIMALS: 6 healthy adult horses. PROCEDURE: A balloon infusion system attached to a catheter placed in the plantarolateral pouch of both tarsocrural joints in each horse was used for continuous gentamicin solution (GM) or balanced electrolyte solution (BES) delivery for 5 days. Cartilage and synovial membrane specimens were collected on day 5 from 3 horses and on day 14 from the remaining 3 horses. Both infused joints from each horse were assessed, using gross evaluation and histologic scoring systems. RESULTS: Significant differences in the histologic scores of synovial membrane specimens between the GM- and BES-treated joints at either 5 or 14 days were not observed. Safranin-O-fast green staining scores were similar between cartilage specimens from GM- and BES-treated joints. Although the synovial membrane histologic scores and safranin-O-fast green staining scores improved from day 5 to 14, the changes in scores were not significant. Loss of synovial intimal cells from villi was found more commonly in sections of synovial membrane from GM-treated joints, compared with BES-treated joints. CONCLUSIONS AND CLINICAL RELEVANCE: Continuous infusion of GM into the tarsocrural joint of horses does not have significant effects on histologic scores of articular cartilage or synovial membrane, compared with those infused with BES. Continuous infusion of GM into the tarsocrural joint of horses for 5 days is an acceptable method for the treatment of septic arthritis.

The effect of implanting gentamicin-impregnated polymethylmethacrylate beads in the tarsocrural joint of the horse.

OBJECTIVE: To determine the effect of intra-articular gentamicin-impregnated polymethylmethacrylate (PMMA) beads inserted in the equine tarsocrural joint on the synovial fluid, synovial lining, and cartilage, and to determine the peak and sustainable gentamicin concentrations in synovial fluid and plasma. STUDY DESIGN: Pharmacokinetic, cytologic, and histologic study of the effect of gentamicin-impregnated PMMA on normal equine tarsocrural joints. ANIMALS: Five healthy adult horses. METHODS: Gentamicin-impregnated PMMA bead strands (3 strands each of 40 beads, with each strand containing 100 mg gentamicin) were surgically inserted into one radiographically normal tarsocrural joint in 5 horses. Each horse had both joints flushed with 1 L of lactated Ringer's solution before bead administration. Synovial fluid total protein concentration, white blood cell (WBC) count, gentamicin concentration, synovial histology, cartilage integrity, and cartilage glycosaminoglycan (GAG) concentrations were determined. RESULTS: Gentamicin concentration (mean +/- SEM peak concentration, 27.9 +/- 2.27 microg/mL) occurred in the first 24 hours and remained above 2 microg/mL for 9 days. Gentamicin concentrations in control joints and the plasma remained below detectable levels. The synovial fluid WBC count for treated joints was increased compared with control joints for 72 hours, but was similar at day 6. The synovial protein concentration in gentamicin-treated joints remained increased for 21 days. Synovium in treated joints had diffuse synovitis, whereas control joints had less fibrovascular proliferation. Superficial cartilage erosion was present in all treated joints. There was no difference in the GAG content of treated and control joint cartilage. CONCLUSIONS: Short-term implantation of gentamicin (300 mg)-impregnated PMMA beads can provide therapeutic levels of gentamicin (>2 microg/mL) in the normal tarsocrural joint for 9 days; however, gentamicin-impregnated PMMA beads induce synovitis and superficial cartilage erosion. CLINICAL RELEVANCE: Temporary intra-articular administration of antibiotic-impregnated PMMA may be an effective way to treat septic joints that require constant high concentrations of antibiotics.

The tyrosine kinase inhibitor tyrphostin AG126 reduces the development of acute and chronic inflammation.

Protein tyrosine kinases help to regulate the expression of many genes that play important roles in inflammation. Here we investigate the effects of the tyrosine kinase inhibitor tyrphostin AG126 in two animal models of acute and chronic inflammation, carrageenan-induced pleurisy and collagen-induced arthritis. We report here that tyrphostin AG126 (given at 1, 3, or 10 mg/kg i.p. in the pleurisy model or 5 mg/kg i.p. every 48 hours in the arthritis model) exerts potent anti-inflammatory effects in animal models of acute and chronic inflammation in vivo. These include the inhibition of pleural exudate formation and mononuclear cell infiltration (pleurisy model) and the development of clinical signs and tissue injury (arthritis model). Furthermore, tyrphostin AG126 reduced the staining for nitrotyrosine and poly (ADP-ribose) polymerase (by immunohistochemistry) and the expression of inducible nitric oxide synthase and cyclooxygenase-2 in the lungs of carrageenan-treated rats and in the joints from collagen-treated rats. Thus, we provide the first evidence that prevention of the activation of protein tyrosine kinases reduces the development of acute and chronic inflammation, and that inhibition of the activity of certain tyrosine kinases may represent a novel approach for the therapy of inflammation.

Taxol (paclitaxel) involution of articular cartilage destruction in collagen induced arthritis: an ultrastructural demonstration of an increased superficial chondroprotective layer.

OBJECTIVE: To determine the ultastructural changes of Taxol (paclitaxel) involution of articular cartilage destruction in collagen induced arthritis (CIA) and to compare with articular cartilage from normal rats. METHODS: Forty-five Louvain rats were randomized to one of 3 protocols for structural analysis: (1) control group, (2) CIA group, and (3) Taxol treated CIA group. The latter group received 10 mg/kg body weight of Taxol at Days 10, 12, and 14 and 7.5 mg/kg body weight of Taxol on Days 16, 18, and 20 postimmunization with collagen type II. Eight days later, each group was examined by light microscopy and scanning and transmission electron microscopy. RESULTS: In Taxol treated rats, the morphology of the articular cartilage reverted to that observed in naive rats except for a striking increase in the thickness of the superficial amorphous layer covering the articular surface. CONCLUSION: The involution of CIA by Taxol suggests that this agent may be useful in the clinical treatment of RA.

Amelioration of type II collagen induced arthritis in rats by treatment with sodium diethyldithiocarbamate.

OBJECTIVE: Sodium diethyldithiocarbamate (Ditiocarb, DDTC), which is used in the treatment of heavy metal poisoning, effectively inhibits NF-kappaB activation and cytokine secretion in vitro. To investigate the antiinflammatory and immunosuppressive potency of DDTC, we examined its influence on the course of collagen induced arthritis in rats. METHODS: Arthritis was induced in female DA rats by injection of rat collagen type II emulsified in incomplete Freund's adjuvant into the tail base. After onset of arthritis, the animals received DDTC or vehicle by intraperitoneal injections or subcutaneous infusion using osmotic pumps. Disulfiram, which is cleaved into DDTC within the gastrointestinal tract, was administered orally via gastric gavage. The course of arthritis was followed by clinical scoring and measurement of joint swelling. RESULTS: Collagen induced arthritis was significantly ameliorated by intraperitoneal injection (2 x 300 mg/kg/day) and subcutaneous infusion (120 mg/kg/day) of DDTC and by enteral administration of disulfiram (200 and 300 mg/kg/day). CONCLUSION: Dithiocarbamates may provide an effective new approach for the treatment of arthritis and other inflammatory diseases.

Antiarthritic activity of soluble tumor necrosis factor receptor type I forms in adjuvant arthritis: correlation of plasma levels with efficacy.

OBJECTIVE: To determine the importance of tumor necrosis factor (TNF) in the pathogenesis of adjuvant disease in rats and to determine plasma levels of recombinant soluble TNF receptor type I (sTNF-RI) necessary for efficacy, to project dosing for human clinical trials. METHODS: Rats with adjuvant arthritis were treated by continuous infusion with sTNF-RI forms to maintain blood levels of this TNF-alpha inhibitory protein. In addition, rats were given bolus injections of polyethylene glycol linked sTNF-RI and efficacy and plasma levels were determined. Effects of treatment in the rats were monitored by sequential volume or diameter measurement of ankle joints, final paw weights, and histologic evaluation of ankle joints, with particular emphasis on bone erosive lesions. RESULTS: In all studies and regardless of dosing methodology (bolus vs continuous infusion), minimal plasma levels for efficacy were in the 0.3-0.5 microg/ml range. Higher plasma levels resulted in greater efficacy, with maximal effects achieved when plasma levels were in the 5 microg/ml range. Beneficial effects of treatment were seen on body weight, paw swelling, splenomegaly, hepatomegaly, and bone resorption. CONCLUSION: TNF-alpha is an important mediator of all aspects of rat adjuvant disease including both the destructive processes in the joints as well as the systemic manifestations of adjuvant disease. Studies using various forms of sTNF-RI consistently show that plasma levels of 0.3-0.5 microg/ml are required for minimal efficacy and that higher plasma levels show dose-responsive enhanced efficacy.

Sulfate incorporation into organic bone matrix of the tibiotarsus of broiler chicks is reduced by excess dietary methionine.

Two experiments were conducted in broiler chicks to determine whether dietary imbalances of sulfur amino acids (SAA), vitamin A, or interactions between the two nutrients could influence organic bone matrix metabolism measured with L-[35S]-methionine. In the first experiment, in vivo incorporation of 35S into the tibiotarsal bone matrix of 2-wk-old birds was unaffected by vitamin A treatment of 10 and 100 times the requirement when compared with that of birds receiving recommended amounts of vitamin A. However, 35S incorporation was significantly reduced by increasing the SAA concentration of the diet to 1.5 times the requirement relative to lysine. In the second experiment, in vitro incorporation of 35S, derived from L-[35S]-methionine, into bone matrix was reduced in birds consuming a diet containing 1.5 times the methionine requirement relative to lysine (Diet HS) when compared with those receiving .75 (Diet LS), 1.0 (Diet NS), or 1.25 (Diet MS) times the requirement. Birds consuming Diet LS incorporated significantly more 35S into organic bone matrix than birds consuming the other three diets. Although the ratio of SAA to lysine was that recommended (.76:1), on a weight basis the concentration of SAA in diet NS was relatively high (11.48 g/kg diet) compared with the NRC (1984) recommendation of 9.3 g/kg diet. The results show that excess SAA can affect organic bone matrix metabolism and suggest that SAA may play a role in the etiology of tibial dyschondroplasia. They also indicate the importance of distinguishing between nutrient content of the diet expressed as a ratio and that expressed on a weight basis.

Capsaicin effects on substance P and CGRP in rat adjuvant arthritis.

The effects of capsaicin on the sensory neuropeptides substance P and calcitonin gene-related peptide were analyzed in the ankle joints and dorsal root ganglia (L2-L6) of adult female Lewis rats. The study included 23 normal rats and 23 arthritic rats, all injected subcutaneously with capsaicin (total dose 200 mg/kg bw). Another two groups of animals from a previous study, i.e., 23 normal rats and 23 arthritic rats not given capsaicin served as controls. Adjuvant arthritis was induced by inoculation with heat-killed mycobacteria. The morphological distribution of sensory neuropeptides was assessed by immunohistochemistry and the tissue concentrations were determined by radioimmunoassay. In normal rats, capsaicin significantly reduced the concentrations of substance P and calcitonin gene-related peptide in ankle joints (54 and 36%, respectively) as well as dorsal root ganglia (40 and 54%, respectively). In arthritic rats those pretreated with capsaicin had significantly lower concentrations of substance P and calcitonin gene-related peptide in dorsal root ganglia (19 and 42%, respectively) compared to the arthritic controls. In the ankle joints, however, only the SP concentration was reduced (42%). Notably, this was accompanied by a 40% reduction in inflammatory response as assessed by comparing the ankle joint weights of the experimental groups. In general, there was a good correlation between the neuropeptide concentrations in ipsilateral ankle joints and the corresponding dorsal root ganglia as assessed in individual rats. The present study of adjuvant induced arthritis shows that capsaicin administration reduces the otherwise up-regulated levels of sensory neuropeptides in dorsal root ganglia and ankle joints. However, capsaicin at the dose given can only mitigate, not completely prevent the development of joint inflammation. Nonetheless, the findings suggest that antineuronal therapy targeted against specific neurotransmitters may prove useful in inflammatory joint disease.

The effects of capsaicin, bradykinin, PGE2 and cicaprost on the discharge of articular sensory receptors in vitro.

The responses of articular sensory receptors to capsaicin, bradykinin, PGE2, and the selective IP-receptor agonist cicaprost were studied in a rat isolated hindlimb in vitro preparation. Long-term maintenance of normal sensory receptor function was achieved in vitro under conditions of combined superfusion and slow perfusion. Response characteristics to mechanical or chemical stimuli on articular sensory receptors identified in this study did not differ to those reported in vivo. This preparation lacks complex effects mediated via spinal or central reflex mechanisms and allows greater control over the physiological environment of the receptors being studied. These results support the conclusion that the effects of capsaicin, bradykinin and the prostanoids are mediated by distinct pharmacological receptors associated with articular sensory nerve endings. The potent potentiating effects of cicaprost on bradykinin-induced excitation suggests that these actions are mediated via IP-receptors.

PGI2-induced activation and sensitization of articular mechanonociceptors.

The effects of PGE2 PGI2 and the stable PGI2 analogue cicaprost on the afferent discharge of ankle joint mechanonociceptors were studied in the anaesthetized rat. Close-arterial injection of PGI2 (0.01-0.1 micrograms) or cicaprost (0.05-5 micrograms) caused both sensitization to mechanical stimulation and excitation of the majority of mechanonociceptors, whereas PGE2 (0.03-3 micrograms) had only weak effects on a small number of nociceptive units. These results suggest the existence of specific PGI2 sensitive receptors (IP receptors) on rat sensory afferent nerves, and support the hypothesis that in the rat endogenous PGI2 plays an important role in the lowering of nociceptive thresholds in inflamed joints.