Moderate exercise protects against joint disease in a murine model of osteoarthritis.

Exercise is recommended as a non-pharmacological therapy for osteoarthritis (OA). Various exercise regimes, with differing intensities and duration, have been used in a range of OA rodent models. These studies show gentle or moderate exercise reduces the severity of OA parameters while high intensity load bearing exercise is detrimental. However, these studies were largely conducted in rats or in mouse models induced by severe injury, age or obesity, whilst destabilization of the medial meniscus (DMM) in mice has become a widely accepted model due to its lower variability, moderate progression and timescale. The present study was undertaken to provide insight into the effect of moderate exercise on early joint pathology in the DMM mouse model. Exercise was induced a week after induction by forced wheel walking for three or 7 weeks. Joints were analyzed by microcomputed tomography and histology. Assessment of skeletal parameters revealed that exercise offered protection against cartilage damage after 7 weeks of exercise, and a temporary protection against osteosclerosis was displayed after 3 weeks of exercise. Furthermore, exercise modified the metaphyseal trabecular microarchitecture of the osteoarthritic leg in both time points examined. Collectively, our findings corroborate previous studies showing that exercise has an important effect on bone in OA, which subsequently, at 8 weeks post-induction, translates into less cartilage damage. Thus, providing an exercise protocol in a surgical mouse model of OA, which can be used in the future to further dissect the mechanisms by which moderate exercise ameliorates OA.

PAR(2) expression in peripheral blood monocytes of patients with rheumatoid arthritis.

OBJECTIVES: Proteinase-activated receptor 2 (PAR(2)) is a G protein-coupled receptor activated by serine proteinases with proinflammatory activity. A study was undertaken to investigate the presence and functional significance of PAR(2) expression on rheumatoid arthritis (RA)-derived leucocyte subsets. METHODS: Venous blood was obtained from patients with RA and osteoarthritis (OA) as well as healthy control subjects. Surface expression of PAR(2) on peripheral blood mononuclear cells (PBMCs) was analysed by flow cytometry and interleukin 6 (IL-6) generation by ELISA. RESULTS: Patients with RA had elevated but variable surface expression of PAR(2) on CD14+ monocytes compared with control subjects (median (1st to 3rd quartiles) 1.76% (0.86-4.10%) vs 0.06% (0.03-0.81%), p<0.0001). CD3+ T cells showed a similar pattern with significantly higher PAR(2) expression in patients with RA compared with controls (3.05% (0.36-11.82%) vs 0.08% (0.02-0.28%), p<0.0001). For both subsets, PAR(2) expression was significantly higher (p<0.00001) in patients with high levels of disease activity: PAR(2) expression for both CD14+ and CD3+ cells correlated to C reactive protein and erythrocyte sedimentation rate. Furthermore, in a cohort of patients with newly diagnosed RA, elevated PAR(2) expression in both CD14+ and CD3+ cells was significantly reduced 3 months after methotrexate or sulfasalazine treatment and this reduction correlated significantly with the reduction in the 28-joint Disease Activity Scale score (p<0.05). PAR(2) expression on cells from patients with OA was low, similar to levels seen in control subjects. Generation of IL-6 by monocytes in response to a selective PAR(2) agonist was significantly greater in patients with RA than in patients with OA and control subjects (p<0.05). CONCLUSIONS: These findings are consistent with a pathogenic role for PAR(2) in RA.

Proteinase-activated receptor-2 mediated inhibition of TNFalpha-stimulated JNK activation - A novel paradigm for G(q/11) linked GPCRs.

In this study we examined the potential for PAR(2) and TNFalpha to synergise at the level of MAP kinase signalling in PAR(2) expressing NCTC2544 cells. However, to our surprise we found that activation of PAR(2) by trypsin or the specific activating peptide SLIGKV-OH strongly inhibited both the phosphorylation and activity of JNK. In contrast neither p38 MAP kinase nor ERK activation was affected although TNFalpha stimulated IkappaBalpha loss was partially reversed. The inhibitory effect was not observed in parental cells nor in cells expressing PAR(4), however inhibition was reversed by pre-incubation with the novel PAR(2) antagonist K14585, suggesting that the effect is specific for PAR(2) activation. SLIGKV-OH was found to be more potent in inhibiting TNFalpha-induced JNK activation than in stimulating JNK alone, suggesting agonist-directed signalling. The PKC activator PMA, also mimicked the inhibitory effect of SLIGKV-OH, and the effects of both agents were reversed by pre-treatment with the PKC inhibitor, GF109203X. Furthermore, incubation with the novel G(q/11) inhibitor YM25480 also reversed PAR(2) mediated inhibition. Activation of PAR(2) was found to disrupt TNFR1 binding to RIP and TRADD and this was reversed by both GF109203X and YM25480. A similar mode of inhibition observed in HUVECs through PAR(2) or P2Y2 receptors demonstrates the potential of a novel paradigm for GPCRs linked to G(q/11), in mediating inhibition of TNFalpha-stimulated JNK activation. This has important implications in assessing the role of GPCRs in inflammation and other conditions.

Protease-activated receptor 2 mediates the proinflammatory effects of synovial mast cells.

OBJECTIVE: Mast cells are hypothesized to play a role in the pathogenesis of rheumatoid arthritis (RA) by mechanisms requiring elucidation. Tryptase released from these cells can activate protease-activated receptor 2 (PAR-2), which was recently shown to have proinflammatory actions. The purpose of this study was to examine the relationship between synovial mast cells and PAR-2. Mast cell proximity to PAR-2-expressing cells was investigated in RA synovium. In murine studies, we assessed the capacity of mast cell tryptase to mediate synovial proinflammatory responses via PAR-2 and whether degranulating mast cells induced synovial hyperemia by PAR-2 activation. METHODS: RA synovial tissue was examined by immunohistochemistry. PAR-2(+/+) and PAR-2(-/-) C57BL/6J mice were used to investigate the PAR-2 dependence of compound 48/80-induced synovial hyperemia, as measured by laser Doppler imaging, and joint swelling and hyperemic responses to recombinant human beta-tryptase. RESULTS: Mast cells and synovial lining cells staining for PAR-2 were colocalized in RA articular tissue. Compound 48/80 administration resulted in vasodilatation in PAR-2(+/+) mice but not in PAR-2(-/-) mice, which showed a vasoconstrictor response. Eliminating the 5-hydroxytryptamine-mediated component of this response with methysergide unveiled an enhanced PAR-2-mediated vasodilatation to compound 48/80 in PAR-2(+/+) mice and ablated the vasoconstrictor response in PAR-2(-/-) mice. Treatment with beta-tryptase resulted in dose-dependent knee joint swelling and synovial vasodilatation in PAR-2(+/+) mice but not PAR-2(-/-) mice. CONCLUSION: This in vivo study is the first to explore the relationship between synovial mast cells and PAR-2. Our results support the hypothesis that mast cells contribute to the pathogenesis of inflammatory arthritis through PAR-2 activation via release of mast cell tryptase.

Angiotensin II type 1 receptor as a novel therapeutic target in rheumatoid arthritis: in vivo analyses in rodent models of arthritis and ex vivo analyses in human inflammatory synovitis.

OBJECTIVE: Angiotensin II (Ang II) is known to have proinflammatory actions, and Ang II type 1 (AT(1)) receptors are up-regulated in the rheumatoid synovium, suggesting that this receptor could be a therapeutic target. The purpose of this study was to investigate the antiinflammatory potential of the selective AT(1) receptor antagonist losartan, which is currently used for the treatment of cardiovascular disease. METHODS: Dose-ranging studies of losartan (1-50 mg/kg) were initially conducted in a rat model of acute (carrageenan/kaolin) arthritis, with subsequent evaluation in a rat model of adjuvant-induced arthritis (Freund's complete adjuvant). Losartan (10(-10) to 10(-6)M) was further tested ex vivo in human inflammatory synovitis, using collagenase-digested synovium. RESULTS: Western blot and immunohistochemical analyses both revealed a substantial increase in AT(1) receptor protein content in synovium from acutely and chronically inflamed rat knee joints. Similarly, synovial Ang I/II protein content was elevated during inflammation. Losartan inhibited acute joint inflammation in a dose-dependent manner, with 15 mg/kg being the optimal dose (and used in subsequent studies). Both prophylactic and therapeutic administration of 15 mg/kg of losartan substantially reduced knee joint swelling in rats with adjuvant monarthritis (> or =50%; P < 0.0001). Losartan also suppressed tumor necrosis factor alpha generation from inflamed human synovium in a dose-dependent manner (P < 0.05). CONCLUSION: Targeting the angiotensin pathway, particularly AT(1) receptors, could have significant therapeutic potential. Randomized placebo-controlled trials are now warranted to establish the extent to which angiotensin receptor blockers may provide antiinflammatory benefits.

Strain dependence in murine models of monoarthritis.

OBJECTIVE AND DESIGN: This study explores the inflammatory response in various murine strains. Utilising several approaches, monoarthritis was induced in the knee, providing an inflammatory model relevant to arthritis. METHODS: Acute (carrageenan/kaolin; C/K) or chronic inflammatory models (Freund's complete adjuvant; FCA) or antigen-induced arthritis (AIA), were induced by peri- and/or intra-articular injection. RESULTS: C/K elicited an acute inflammatory response in various strains of mice, with significant (P < 0.005) phenotypic variation. FCA induction provided a chronic inflammatory response. The magnitude of the response in both acute and chronic models was strain dependent, with BalbC exhibiting the most resistance to swelling in all models. AIA produced only an acute response in three strains tested. CONCLUSIONS: The data presented, demonstrating variation in the magnitude of acute and chronic inflammatory responses in different mice strains, allows informed selection of appropriate strains and models for future experimental studies.

Protective effect of sensory denervation in inflammatory arthritis (evidence of regulatory neuroimmune pathways in the arthritic joint).

OBJECTIVE: To investigate the direct effect of joint innervation on immune mediated joint inflammation in a patient with psoriatic arthritis (PsA). CASE REPORT: The patient developed arthritis mutilans in all digits of both hands with the exception of the left 4th finger, which had prior sensory denervation following traumatic nerve dissection. Plain radiography, ultrasonography and nerve conduction studies of the hands confirmed the absence of articular disease and sensory innervation in the left 4th digit. METHODS: This relationship between joint innervation and joint inflammation was investigated experimentally by prior surgical sensory denervation of the medial aspect of the knee in six Wistar rats in which carrageenan induced arthritis was subsequently induced. Prior sensory denervation--with preservation of muscle function--prevented the development of inflammatory arthritis in the denervated knee. DISCUSSION: Observations in human and animal inflammatory arthritis suggest that regulatory neuroimmune pathways in the joint are an important mechanism that modulates the clinical expression of inflammatory arthritis.

Divergent roles of nitrergic and prostanoid pathways in chronic joint inflammation.

BACKGROUND: Nitrergic and prostanoid pathways have both been implicated in inflammatory processes. OBJECTIVE: To investigate their respective contributions in a rat model of chronic arthritis. METHODS: Male Wistar rats (n = 4-6/group) received either an intra-articular injection of 2% carrageenan/4% kaolin (C/K) or intra- and periarticular injections of Freund's complete adjuvant (FCA; 10 mg/ml M tuberculosis). Joint diameter, urinary nitric oxide metabolites (NO(x)), and prostaglandin E(2) (PGE(2)) levels were measured as indices of the inflammatory process. A prophylactic and therapeutic (day 5) dose ranging study of an inducible nitric oxide synthase inhibitor, L-N-(1-iminoethyl)-lysine (L-NIL), and a cyclo-oxygenase-2 (COX-2) inhibitor, SC-236, was performed with the drugs given subcutaneously. Submaximal doses were identified and used for combination studies. Appropriate vehicle controls were included. RESULTS: L-NIL and SC-236 dose dependently inhibited C/K induced acute joint swelling, the magnitude being greatest when they were given in combination. Both prophylactic and therapeutic administration of SC-236 in the FCA induced model of chronic arthritis produced a dose dependent reduction in all the measures assessed. However, although L-NIL demonstrated similar dose dependent inhibition of urinary NO(x) and PGE(2) levels, joint swelling was significantly exacerbated in this model. Co-administration of the inhibitors nullified the benefits of SC-236. CONCLUSION: Whereas COX-2 derived prostaglandins are proinflammatory in both acute and chronic joint inflammation, NO seems to have divergent roles, being anti-inflammatory in chronic and proinflammatory in acute joint inflammation.

Gender differences in regional cutaneous microcirculatory responses to capsaicin.

Vascular responsiveness between healthy male and female subjects to capsaicin, an agent promoting neuropeptide release, was compared. Changes in skin perfusion were measured non-invasively using laser Doppler imaging. Topical application of a 3% solution of capsaicin to the dorsum of the hand resulted in vasodilatation in nine of 10 male subjects, but in less than half of the female subjects. Responses to capsaicin at the shin were smaller but did not show gender differences. Fingertip temperature was significantly lower in females compared with males and this correlated (r = 0.54, P < 0.01) with the maximum response to capsaicin. These effects were specific to capsaicin as endothelium-dependent and -independent vasodilator mechanisms, assessed non-invasively by iontophoresis of acetylcholine and sodium nitroprusside, respectively, showed no gender differences. These findings suggest a specific anomaly in capsaicin-sensitive vasodilator mechanisms in some subjects, perhaps indicative of subclinical expression of Raynaud's phenomenon.

Combined inhibition of nitrergic and prostanoid pathways in J774 macrophages.

OBJECTIVES: Nitric oxide and prostaglandins are both implicated in the pathogenesis of inflammatory conditions such as rheumatoid arthritis (RA). The hypothesis that simultaneous inhibition of nitric oxide synthase (NOS) and cyclooxygenase (COX) was more effective than inhibition of either enzyme alone was tested. METHODS: J774 macrophages were pre-incubated with L-NAME and/or indomethacin, prior to activation with LPS (10 micrograms/ml). RESULTS: LPS significantly increased NO2-; PGE2 and TNF-alpha levels by 24 h. Quantitative real-time PCR demonstrated a dose-dependent reduction in the expression of COX-2 in the presence of increasing doses of L-NAME. NO2- and PGE2 production were inhibited in a dose-dependent manner by either indomethacin or L-NAME. Combined administration of L-NAME and indomethacin produced a significantly greater inhibition of NO2- and PGE2 than either inhibitor alone. CONCLUSION: The data supports the therapeutic potential of combined inhibition of the prostanoid and nitrergic systems as an anti-inflammatory treatment strategy and supports the progression of this work into models of arthritis.

Expression of constitutive but not inducible cyclooxygenase maintains articular perfusion in the rat knee.

Experiments were performed in the normal rat knee joint to investigate the role of different isoforms of cyclooxygenase (COX) in the regulation of basal joint blood flow. Laser Doppler imaging (LDI) was used to measure articular perfusion, and reverse transcriptase polymerase chain reaction (RT-PCR) for the detection of COX-1 and COX-2 mRNA in joint tissue. Intravenous infusion of indomethacin (a non-selective inhibitor of COX; 0.34 nmol min(-1)) over 40 min produced a time dependent increase in articular vascular resistance (maximum 22.5 % at 40 min; P < 0.0001, one-way ANOVA) whereas vehicle over a similar time period had no effect in a control group. An equimolar concentration of a highly selective inhibitor for COX-2, SC-236, was administered in a further group of rats but this did not increase articular vascular resistance. While there was no significant difference between the response to vehicle and SC-236 (two-way ANOVA; P = 0.686, n = 6) the response to indomethacin was significantly greater than vehicle or SC-236 (two-way Anova; P < 0.0001, n = 6). COX-1, but not COX-2, was detectable by RT-PCR in all joint tissue samples examined (n = 4). The results of this study indicate that prostaglandins (PGs) play an important role in the maintenance of basal perfusion in the rat knee joint, with COX-1 being the physiologically relevant isoform. Experimental Physiology (2001) 86.2, 191-197.

Metacarpophalangeal joints in rheumatoid arthritis: laser Doppler imaging--initial experience.

Laser Doppler imaging is a noninvasive method yielding a spatial perfusion map. With use of a near-infrared laser, elevated perfusion associated with the metacarpophalangeal joints was detectable in patients with active rheumatoid arthritis. Findings at laser Doppler imaging correlated with pain scores and synovitis detected at ultrasonography, whereas the power Doppler sign (red pixels inside the active green box) did not. Laser Doppler imaging has the potential to help assess soft-tissue inflammation.

Novel use of laser Doppler imaging for investigating epicondylitis.

OBJECTIVE: This investigation evaluated a novel form of tissue perfusion measurement, laser Doppler imaging (LDI), in a case of lateral epicondylitis to establish if it might have applications in assessing soft tissue lesions. LDI was used in conjunction with ultrasonography to provide information about tissue oedema as well as the power Doppler signal as an alternative method of assessing blood flow. METHODS: A laser Doppler imager with a near-infrared (NIR) laser source was used to improve tissue penetration and yield measurements of perfusion (flux) from structures under the skin. Skin temperature over the lateral epicondylar region was also measured. Ultrasonography was used in both grey-scale and power Doppler modes. LDI, temperature measurements and ultrasonographic data were obtained before treatment and serially after local injection of methylprednisolone. RESULTS: Before treatment there was increased perfusion and skin temperature and the presence of a power Doppler sign associated with the right lateral epicondyle as well as oedema at the extensor origin. None of these was present at the asymptomatic contralateral epicondylar region. Twenty-four hours after methylprednisolone administration, both perfusion and skin temperature had increased, and they declined over the subsequent 48 h. Although skin temperature had declined to normal (referenced to the contralateral epicondyle) by the third day after injection, it took until the eleventh day after injection for perfusion to normalize. CONCLUSIONS: LDI using an NIR laser source appears to be an effective non-invasive method for the examination of inflammatory responses in soft tissue, with greater sensitivity than thermally based methods. In addition, LDI was found to correlate with power Doppler ultrasonography.

Neurogenic origin of articular hyperemia in early degenerative joint disease.

It has been speculated that joint instability resulting from anterior cruciate ligament (ACL) rupture could be exacerbated by changes in vasomotor activity in the remaining supporting structures. In this study, the effect of ACL transection on medial collateral ligament (MCL) basal perfusion and its responsiveness to calcitonin gene-related peptide (CGRP) and sympathetic adrenergic influences was examined. Using urethan-anesthetized rabbits, we tested the effects of CGRP and its antagonist CGRP-(8-37) by topical application of these agents to the exposed knee while sympathetic influences were tested by electrically stimulating the saphenous nerve. It was found that MCL basal perfusion was elevated in ACL-sectioned joints; however, this effect was abrogated by prior resection of the articular nerve supply. At the doses tested, the normal vasodilator response to CGRP was abolished in ACL-sectioned joints, whereas the response to CGRP-(8-37) was attenuated. Even under the influence of increased constrictor tone, MCL and capsule blood vessels still showed substantially reduced responses to exogenous CGRP administration. By contrast, nerve-mediated constrictor responses were mostly unaffected by joint instability. This study suggests that posttraumatic knee joint hyperemia is neurogenically mediated, possibly by increased secretion of CGRP.

Repetitive activity alters perfusion of proximal interphalangeal joints of the human hand.

OBJECTIVE: To examine whether competitive volleyball players show any difference in perfusion of their proximal interphalangeal (PIP) joints compared with a healthy group of subjects. Also to assess the viability of a dual wavelength laser Doppler imager (LDI) in making these measurements. SETTING: Physiology laboratory. PARTICIPANTS: Ten active volleyball players who had experienced repetitive finger joint injury and 12 age- and sex-matched normal control subjects. MAIN OUTCOME MEASURES: Using a modified LDI incorporating a near- infrared (850 nm) laser as well as a standard red (633 nm) laser, scans were performed over the dorsum of the hands of the volleyball players and the control group. RESULTS: Higher perfusion values were obtained with the 850-nm laser than with the red 633-nm laser. When referenced to adjacent skin blood flow, perfusion over PIP joints of volleyball players was found to be significantly higher than that in control subjects (p=0.00012; n=10-12). CONCLUSIONS: The higher perfusion values obtained using the 850-nm laser suggest that the longer wavelength laser is measuring perfusion in a greater volume of tissue, which could include subcutaneous structures. Volleyball players have significantly higher perfusion over the PIP joints, which is unlikely to be due to differences in skin perfusion over the two regions but is more likely to be related to hyperemia of the underlying PIP joints. The reason for increased PIP perfusion is not clear; it may represent ongoing tissue inflammation due to repeated injury, or it could be an adaptive response to the stresses placed on these joints by this type of repetitive activity. CLINICAL RELEVANCE: Near-infrared laser Doppler imaging has the potential to provide a noninvasive clinical assessment of finger joint injuries.

Spatial variation in sympathetic influences on the vasculature of the synovium and medial collateral ligament of the rabbit knee joint.

1. Laser Doppler perfusion imaging was used to assess the role of the sympathetic nervous system in the control of blood flow to the medial collateral ligament and capsule (synovium and overlying fibrous tissues) of the rabbit knee joint. 2. Electrical stimulation of the saphenous nerve (width 1 ms; amplitude 20V; 1-30 Hz) produced a frequency-dependent vasoconstriction of knee joint vasculature. The response was maximal at 30 Hz and gave the greatest fall in perfusion at the femoral insertion of the ligament (by 33.8 +/- 7.4%, mean +/- S.E.M.; n = 5-6) and the smallest decrease at the tibial insertion of the ligament (by 10.6 +/- 2.9%). 3. Topical application of phentolamine (10(-6) mol) had no significant effect on basal knee joint blood flow. However, it abolished the nerve-mediated constrictor responses in all regions of the medial collateral ligament and synovium at all frequencies. 4. Topical administration of adrenaline (10(-14) to 10(-7) mol) caused a dose-dependent decrease in knee joint blood flow with the highest dose producing > 75% reduction in perfusion at all areas. 5. There was no evidence of a reactive hyperaemia in the 5 min following a 5 min period of femoral artery occlusion. Artificial manipulation of arterial blood pressure by intravenous infusion or withdrawal of blood caused a proportional change in ligament and synovial blood flow. These observations may indicate a lack of autoregulation in the joint and its exclusion from baroreflex modulation. 6. These results suggest a potential role for the sympathetic nervous system in the control of knee joint blood flow. Neuromodulation of ligament perfusion appears to predominate at the femoral insertion and this could prove to have functional significance.

Spatial heterogeneity of the effects of calcitonin gene-related peptide (CGRP) on the microvasculature of ligaments in the rabbit knee joint.

1. Experiments were performed in anaesthetized rabbits to examine the effects of calcitonin gene-related peptide (CGRP) and the CGRP antagonist CGRP8-37 on blood flow to the medial collateral ligament of the knee joint. 2. Topical application of CGRP (10(-13) to 10(-9) mol) to the exposed external surface of eight knee joints resulted in dose-dependent dilatation of vessels in both the ligament and the joint capsule. The magnitude of this response varied significantly in different regions of the medial collateral ligament, with the 10(-9) mol dose of CGRP giving the maximum response (101.5 +/- 25.3% increase) at the femoral insertion site of the medial collateral ligament and lowest (23.1 +/- 8.8%) at the tibial insertion site. 3. Topical application of CGRP8-37 (0.1, 1 and 10 nmol) produced dose-dependent constriction of vessels in the ligament and the joint capsule in five knees, with a trend towards the greatest effect occurring at the femoral insertion site (45.8 +/- 8.1% reduction in blood flow). With the 10 nmol dose, the vasoconstrictor response at the femoral insertion site differed significantly (P<0.05) from the responses obtained at the tibial insertion and joint capsule sites. 4. Topical application of CGRP8-37 (0.1, 1 and 10 nmol) to four chronically denervated knees produced substantially smaller vasoconstrictor responses at all sites. At the femoral insertion site, where 10 nmol CGRP8-37 normally produces a 45.8 +/- 8.1% reduction in blood flow (n=8), ten days following denervation this response was reduced to 6.5 +/- 6.1%, this difference being significant (P=0.01). 5. Adrenaline was applied topically to augment blood vessel tone, in order to establish how effectively co-administration of CGRP would offset this increase in tone. Adrenaline (10(-10) mol) produced vasoconstriction at all sites (n=6). In the capsule this vasoconstriction was virtually abolished when CGRP (10(-9) mol) was co-administered with adrenaline but in the ligament vasodilatation occurred at all sites. This vasodilatation was significantly greater at the femoral insertion site compared to the tibial insertion and mid ligament sites (P<0.05 for both) and the capsule (P<0.01). 6. Topical application of substance P (10(-10) or 10(-9) mol) failed to elicit dilatation of ligament blood vessels. 7. These results suggest that endogenous CGRP may play an important role in regulating blood flow to different structures in and around the knee joint.

Modulation of synovial blood flow by the calcitonin gene-related peptide (CGRP) receptor antagonist, CGRP(8-37).

1. The effect of the calcitonin gene-related peptide (CGRP) receptor antagonist, CGRP(8-37) on blood flow in the knee joint of the anaesthetized rat was investigated. 2. Synovial blood flow in both exposed and intact, skin-covered knees was measured by laser Doppler perfusion imaging. 3. Topical application of CGRP(8-37) caused a dose-dependent fall in synovial blood flow in the exposed knee joint of the rat. At low (1.5 nmol) doses of CGRP(8-37) there was no significant effect on synovial blood flow. In rats treated with 7.5 nmol CGRP(8-37) there was a fall in synovial blood flow (maximum effect at 10 min: -28.8 +/- 4.6%; n=7), which returned to resting levels within 30 min. The highest dose (15 nmol) of antagonist used in this study caused a marked (maximum at 10 min: -35.6 +/- 9.3%; n=8), and prolonged (up to 30 min) fall in blood flow. 4. Ten days after surgical denervation, CGRP(9-37) (15 nmol, topical) had no significant effect on blood flow in the rat exposed knee joint (change in flux at 10 min: -5.1+/-3.6%; n=4). This suggests that CGRP(8-37) acts selectively to antagonize the actions of a neurally derived product, probably CGRP, on the rat synovial vasculature. 5. In skin-covered knee joints, intra-articular injection of CGRP(8-37) (15 nmol; bolus) elicited a significant fall in synovial blood flow (maximum effect at 10 min: -15.5 +/- 5.8%; n=6). 6. CGRP (0.01, 0.1 or 1.0 nmol; topical) caused a dose-dependent increase in exposed knee joint blood flow, which was attenuated by co-administration of 1.5 nmol CGRP(8-37). For example, 1 nmol CGRP elicited a peak increase in flux at 10 min of 94.7 +/- 31.8% (n=8) and 28.8 +/- 8.9% (n=7) in the absence and presence of CGRP(8-37), respectively. The vasodilator responses induced by acetylcholine (ACh) (10 nmol, topical; n=4-5) or sodium nitroprusside (SNP) (10 nmol, topical; n=4-5) were unaltered in the presence of CGRP(8-37) (1.5 nmol, topical). 7. Thus, the CGRP receptor antagonist CGRP(8-37) elicits vasoconstriction in the rat synovium. This suggests that the endogenous, basal release of CGRP may play a physiological role in the regulation of blood flow in the rat knee joint.

Laser Doppler perfusion imaging of synovial tissues using red and near infra-red lasers.

A new laser Doppler perfusion imaging (LDI) system was evaluated by comparing it with the well-established radiolabelled microsphere technique for measuring blood flow in the rabbit knee joint capsule. In this study two laser sources (635 and 835 nm) were compared at three scan speeds (50, 10 and 4 ms/pixel). With blood flow to the rabbit hindlimb controlled via a peristaltic pump, the comparison of LDI and microsphere measurement techniques yielded highly significant correlations for both laser sources (r = 0.9; p = 0.0001; 14 measurements in 7 animals). Comparison of the three scan speeds demonstrated acceptable agreement without significant bias between measurements, suggesting that the inevitable narrowing of the bandwidth at the fastest scan speed does not cause significant deterioration of the signal. The flux values obtained with 635 and 835 nm laser sources were linearly related (r = 0.93, p = 0.0001; 66 measurements in 12 animals), although there was a small but significant bias for higher values with the 635-nm laser (mean ratio of flux values 1.06, 95% confidence interval 1.01-1.12). These results validate the use of LDI with either wavelength laser for the assessment of joint capsule perfusion.