Published online before print October 3, 2007
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* Department of Pharmacology, Faculty of Medicine of Ribeirão Preto, University of São Paulo, São Paulo, Brazil;
Department of Oral Surgery and Pathology, School of Dentistry, Universidade Federal de Minas Gerais, Minas Gerais, Brazil;
Departament of Internal Medicine, School of Medicine, Federal University of Ceara, Fortaleza-CE, Brazil; and
Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
2 Correspondence: Department of Pharmacology, Faculty of Medicine of Ribeirao Preto, University of São Paulo, Avenida Bandeirantes, 3900, 14049-900-Ribeirao Preto, Sao Paulo, Brazil. E-mail: fdqcunha{at}fmrp.usp.br
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Key Words: joint pain • arthritis • hyperalgesia • 5-lypoxygenase • hypernociception
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Cytokines and chemokines such as IL-1β, IL-15, IL-18, TNF-
, IL-8, MIP-1, and MIP-2 orchestrate the recruitment of neutrophils and consequently, tissue lesions [8
9
10
11
12
]. Furthermore, chemotatic activity of those cytokines depends, at least in part, on leukotriene B4 (LTB4) production [13
], which is a potent chemotactic agent for neutrophils in vitro and in vivo [14
]. In arthritic patients, high levels of LTB4 are detected in synovial [15
] and serum fluids [16
], and the increased capacity of the neutrophil to release LTB4 has also been reported. In fact, inhibition of LTB4 synthesis or action ameliorates collagen-induced arthritis in mice [12
] and in other Th1 inflammatory models [17
], therefore suggesting that LTB4 plays an important role in the pathogenesis of arthritis.
In addition, the LTB4 role in pain has been demonstrated, which is apparently a secondary phenomena resulting from its activity in neutrophil migration [18 , 19 ]. In our model of joint inflammation in mice, there is an association between the time-course of the zymosan-induced hypernociception and neutrophil migration, suggesting an interdependence of these phenomena [3 ]. Similarly, a recent study suggests that articular incapacitation in zymosan-induced arthritis in rats is mediated by LTB4, and this model of incapacitation also seems to be associated particularly with the cell influx into the joint [20 ]. The importance of LTB4 for hypernociception was also demonstrated in a Th1 immune inflammatory response in rats [21 ].
Taking into account the above-mentioned data, we investigated the participation of LTB4 and its mechanisms in an experimental model of movement-induced inflammatory joint hypernociception in mice.
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Induction of tibio-tarsal joint inflammation
Joint inflammation was induced by administration of zymosan (30 µg) [3
] or LTB4 (8.3, 25, and 75 ng) diluted in 5 µL saline and injected into the right tibio-tarsal joint of lightly anesthetized mice. The substances were injected with a 29-G hypodermic needle inserted into the joint. Control animals received an intra-articular injection of 5 µL sterile saline.
Articular flexion-elicited hypernociception in the inflamed joint: assessment by a modified electronic pressure-meter test for mice
Experiments were performed as described previously in ref. [3
], which is a modification of the test described earlier in refs. [22
, 23
]. In a quiet room, mice were placed in acrylic cages (12x10x17 cm high) with a wire grid floor 15–30 min before testing for environmental adaptation. Stimulations were performed only when animals were quiet, without exploratory movements or defecation and not resting on their paws. In these experiments, an electronic pressure-meter was used. It consists of a hand-held force transducer fitted with a polypropylene tip (IITC Inc., Life Science Instruments, Woodland Hills, CA, USA). An increasing perpendicular force applied to the central area of the plantar surface induced the flexion of the tibio-tarsal joint. A tilted mirror below the grid provided a clear view of the animal’s hind paw. In the electronic pressure-meter test, the intensity of the stimulus was recorded automatically when the paw was withdrawn. The non-nociceptive tip probe area size used to evaluate the movement-elicited hypernociception was 4.15 mm2. The flexion-elicited withdrawal threshold is expressed in grams.
Measurement of myeloperoxidase (MPO) activity
The tibio-tarsal joint region was separated from the tibio-tarsal bone complex at 5 h after LTB4 and 7 h after zymosan administration. The neutrophil migration to the tibio-tarsal joint region of mice was evaluated by the MPO kinetic-colorimetric assay as described previously [24
]. Samples of joint tissue were collected in 50 mM K2HPO4 buffer (pH 6.0) containing 0.5% hexadecyl trimethylammonium bromide and kept at –80°C until use. Samples were homogenized using a Polytron (PT3100), centrifuged at 16,100 g for 4 min, and the resulting supernatant was assayed spectrophotometrically for MPO activity determination at 450 nm (Spectra max) with three readings within 1 min. The MPO activity of samples was compared with a standard curve of neutrophils. In these experimental conditions, we did not detect MPO activity from peritoneal macrophages (data not shown). Briefly, 10 µL sample was mixed with 200 µL 50 mM phosphate buffer, pH 6.0, containing 0.167 mg/ml O-dianisidine dihydrochloride and 0.0005% hydrogen peroxide. The results were presented as the MPO activity (number of neutrophilsxmg joint tissue–1).
Enzymatic immune assay for LTB4
The animals were killed 1, 3, 5, and 7 h after intra-articular zymosan injection, and the joint tissue samples were collected for the determination of LTB4 levels. Total joint tissue samples were obtained and homogenized in 500 µL of the appropriate buffer containing protease inhibitors [25
]. LTB4 levels were determined by enzyme immunoassay using a commercial kit (BiotrakTM, Amersham Pharmacia Biotech, UK), according to the manufacturer’s instructions. Briefly, recombinant murine LTB4 standards at various dilutions and the samples were added to a microtiter plate precoated with mouse mAb. Then, LTB4 acetylcholinesterase tracer and LTB4 antiserum anti-LTB4 were added to each well. The plate was incubated at room temperature overnight. The plate was washed, 200 µl substrate to acetylcholinesterase (Ellman’s Reagent) was added to each well, and the plate was incubated for 60–90 min to color development. The O.D. was determined at 405 nm, and the results were expressed as pg LTB4/joint, based on a standard curve.
Measurement of PGE2 in joint
The joint tissues were collected 2 h after intra-articular injection of LTB4 (25 ng/joint) or saline as described above. The PGE2 was extracted from tissue as described previously and determined by radioimmunoassay [26
, 27
]. Furthermore, the sample buffer contained 20 µg/ml indomethacin. The results are expressed as pg PGE2 per joint.
Histological analysis
The tibio-tarsal bone complex was harvested and fixed in 10% neutral-buffered formalin for 48 h. After decalcification in 10% EDTA, pH 7.2, at room temperature for 3 weeks, specimens were embedded by routine histological technique in paraffin and sectioned at 7 µm for H&E staining. All sectioning was done in a sagittal plane, and sections, which included the entire tibio-tarsal joint, were selected for analysis. Samples were harvested at 7 h and 96 h after zymosan administration. The histopathological analyses were also done in the fucoidin-treated group. A blind observer evaluated a minimum of six sections per animal. Histopathological changes in the tibio-tarsal joint were evaluated using the following parameters: cell infiltration in the articular space and synovial tissue, the presence of bone resorption, and synoviocyte proliferation. Each sample was scored for the degree of inflammatory infiltration of the periarticular tissue on the following scale: 0, no evidence of inflammation; 1, slight inflammation, characterized by inflammatory cells invading between 1% and 10% of the periarticular tissue; 2, moderate inflammation, characterized by inflammatory cells invading between 10% and 25% of the periarticular tissue; 3, intense inflammation in 25–50% of the periarticular tissue; and 4, severe inflammation, characterized by inflammatory cells invading more than 50% of the periarticular tissue [3
].
Drugs and reagents
MK886 [5LO-activating protein (FLAP) inhibitor] was obtained from Calbiochem (San Diego, CA, USA); CP105696 [LTB4 receptor 1 (BLT1) antagonist] was obtained from Pfizer/Groton Laboratories (Groton, CT, USA); indomethacin (nonselective cyclo-oxigenase inhibitor) was obtained from Prodome (Campinas, SP, Brazil); fucoidan, zymosan, and LTB4 were obtained from Sigma Chemical Co. (St. Louis, MO, USA); and metilcellulose was obtained from Merck (Darmstadt, Germany). The LTB4 ELISA kit was purchased from Cayman Chemical (Ann Arbor, MI, USA). The celecoxib (selective cyclo-oxigenase-2 inhibitor) was purchased from Pfizer (Pfizer Pharmaceuticals LLC, Caguas, PR). The antineutrophil antibody (RB6-8C5) was a generous gift from Prof. Montilo Russo (University of Sao Paulo). The specificity of the antibody was published in ref. [28
].
Statistical analysis
Results are presented as means ± SEM and are representative of two separate experiments of five animals per five animal OVA (ANOVA) and used to compare the groups and doses at all times (curves) when the hypernociceptive responses were measured at different times after the stimulus injection. The analyzed factors were treatments, time and time-versus-treatment interaction. When there was a significant time-versus-treatment interaction, one-way ANOVA followed by Bonferroni’s t-test was performed for each time. Alternatively, when the hypernociceptive responses were measured once after the stimulus injection, the differences between responses were evaluated by one-way ANOVA followed by Bonferroni’s t-test. Differences in the inflammation scores between the groups were expressed as median ± SEM and analyzed by the Kruskal-Wallis test and Dunn’s post-test. Statistical differences were considered to be significant at P < 0.05.
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Figure 1. LTB4 mediates zymosan-induced joint hypernociception. (A) MK886 inhibits zymosan-elicited tibio-tarsal joint hypernociceptive flexion. Mice were treated with MK886 or vehicle (saline) 1 h before the tibio-tarsal injection of zymosan (30 µg/5 µL). (B) Mice were post-treated with MK886 (3 mg/kg/p.o.) 3 h or 24 h after zymosan injection (arrow). (C) The 5LO–/– and wild-type (WT) mice were injected with saline (5 µL) or zymosan (30 µg/5 µL) into the tibio-tarsal joint. The results are expressed as the mean ± SEM of five animals per group. (D) Mice were treated with BLT antagonist [CP105696 (CP), 3 mg/kg i.p.] 45 min before and a reinforcing dose at 2 h and 45 min after zymosan injection. The flexion-elicited hypernociception was evaluated 3 h and 7 h after zymosan injection. (E) Levels of LTB4 in tibio-tarsal joint tissue injected with zymosan. The articular region was colleted 1–7 h after zymosan (30 µg/5 µL), saline group (5 µL) injection, or contralateral joint (CL). The processed samples were used for measurement of LTB4 levels by ELISA. (F) LTB4 induced tibio-tarsal joint hypernociceptive flexion in mice. Injection of LTB4 into the tibio-tarsal joint of mice induced a dose (8.3, 25, and 75 ng/5 µL)- and time (1, 3, 5, 7, 24, and 48 h)-dependent tibio-tarsal joint hypernociceptive flexion. The results are expressed as the mean ± SEM of five animals per group. *, Significant difference when compared with the saline group; #, compared with the zymosan-injected group (P<0.05).
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It is a consensus that the use of knockout mice is also an accurate approach to establish the role of mediators in the inflammatory process. Therefore, 5LO–/– mice were used to confirm the pharmacological approaches described above. The hypernociception induced by intra-articular injection of zymosan was reduced significantly in 5LO–/– mice when compared with WT (Sv129) mice in all the evaluations (1, 3, 5, 7, 24, 48, 72, and 96 h; Fig. 1C ). The hypernociceptive response was inhibited by 61% at 7 h after zymosan injection.
The next step of this study was to evaluate whether LTB4 is the 5LO product responsible for the genesis of joint hypernociception in zymosan-induced articular inflammation. The treatment of mice with the BLT-selective antagonist (CP105696, 3 mg/kg i.p.) 45 min before, plus a reinforcing dose at 2 h and 45 min after zymosan injection, decreased the tibio-tarsal joint hypernociception by 51% and 65% when evaluated 3 h and 7 h after zymosan administration, respectively (Fig. 1D) . Supporting this fact, LTB4 levels were already increased significantly 3 h after zymosan injection when compared with the saline group. The LTB4 levels were maximal between 5 h and 7 h (Fig. 1E) . These results are compatible with a continuous production of LTB4 after zymosan stimulus, which is in agreement with the results obtained with the MK886 treatment, as this drug inhibited zymosan hypernociception when used as pretreatment or post-treatment at different times.
Further supporting the hypernociceptive role of LTB4 in articular inflammation, we observed that the administration of LTB4 (8.3, 25, and 75 ng/5 µL) into the tibio-tarsal joint of mice induced a dose (8.3, 25, and 75 ng/5 µL)- and time (1, 3, 5, 7, 24, and 48 h)-dependent dorsal, flexion-elicited hypernociception. The hypernociceptive responses were maximal at 5 h (Fig. 1F) . The submaximal dose of LTB4, 25 ng, was used for the next experiment. Therefore, the results presented until now suggest that LTB4 is, at least in part, responsible for the joint hypernociception induced by zymosan.
LTB4 mediates zymosan-induced joint hypernociception and neutrophil migration to the joint in mice
Therefore, it seems that there is a sustained production of LTB4 during zymosan articular inflammation. In addition to the antinociceptive effect of MK886, a significant reduction of the zymosan-induced neutrophil migration to the joint was detected, as evaluated by the MPO activity assay (Fig. 2A
). Furthermore, the neutrophil migration induced by zymosan was reduced in 5LO–/– mice (Fig. 2B)
. These results point to a possible association between neutrophil migration and joint hypernociception. It is important to mention that the C57BL/6 is more responsive to neutrophil migration induced by different stimuli than Sv129 (our laboratory observation). Corroborating these data, microscopic analysis revealed significant reduction of inflammation score in the tibio-tarsal joint of 5LO–/– mice 7 h and 96 h after zymosan injection (Fig. 2C)
. Figure 2D
shows an impressive infiltration of cells into the joint of zymosan-injected mice, which was reduced significantly in 5LO–/– mice.
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Figure 2. Role of LTB4 in the neutrophil migration during zymosan-induced articular inflammation. (A) MK886 inhibits zymosan-induced neutrophil migration into the tibio-tarsal joint of mice, which were treated with MK886 (3 mg/kg/p.o.) or vehicle 1 h before the tibio-tarsal injection of zymosan (30 µg/5 µL). (B) Reduced neutrophil migration to the tibio-tarsal joint induced by zymosan in 5LO–/– mice compared with WT mice. The articular region was colleted 7 h after zymosan (30 µg/5 µL) injection, and the MPO activity was measured. (C) Histological score of zymosan-induced inflammation in the tibio-tarsal joint of WT and 5LO–/– (Sv129). The articular region was colleted after 7 h and 96 h for histological analysis. (D) H&E-stained representative joint section of samples collected 7 h after zymosan (Sv129 or 5LO–/– mice) or saline injection (original magnification, 50x). Inflammatory cell infiltration is indicated by the arrow. (E) LTB4 induces an influx of neutrophils into the tibio-tarsal joint of mice. The articular region was colleted 5 h after LTB4 (25 ng/5 µL), saline group (5 µL) injection, or CL, and the MPO activity was measured. The results are expressed as the mean ± SEM of five animals per group. *, Significant difference when compared with the saline group; #, compared with the zymosan and WT groups (P<0.05).
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Prostanoids and LTs mediate LTB4-induced joint hypernociception
The mechanism involved in LTB4-induced articular hypernociception was evaluated by the pretreatment of mice with indomethacin (5 mg kg–1 30 min before i.p., nonselective cyclo-oxigenase inhibitor) [22
], MK886 (3 mg kg–1, p.o.), celecoxib (50 mg kg–1, p.o., cyclo-oxigenase-2-selective inhibitor), or the association of indomehacin and MK886. It was observed that indomethacin, MK886, celecoxib, or the association of indomethacin and MK886 treatments inhibited LTB4-induced articular hypernociception (Fig. 3A
). Furthermore, celecoxib treatment also inhibited zymosan-induced articular hypernociception (data not shown), therefore suggesting that prostanoids derived from cyclo-oxigenase-2 and LTs mediate LTB4-induced articular hypernociception. The possible involvement of LTs in the hypernociceptive effect of LTB4 was also addressed in 5LO–/– mice. Indeed, the joint hypernociception induced by LTB4 was also reduced in 5LO–/– mice when compared with the WT control group (Fig. 3B)
. In addition, indomethacin treatment of 5LO–/– mice did not add further reduction of the hypernociception induced by LTB4 (Fig. 3B)
. Furthermore, the neutrophil migration to the joint induced by LTB4 was abolished in 5LO–/– mice when compared with control group Sv129 (Fig. 3C)
, thus suggesting that LTB4 induces an inflammatory response with the release of additional endogenous LTs, which induce PG production. Further support to an inflammatory characteristic of LTB4-induced articular hypernociception is the demonstration that it was inhibited by the pretreatment of the animals with the glucocorticosteroid dexamethasone (45% of inhibition; data not shown in figure).
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Figure 3. LTs and PGs mediate LTB4-induced joint hypernociception (A) Effect of dexamethasone treatment in the LTB4 tibio-tarsal joint-elicited hypernociceptive flexion. Mice were treated with dexamethasone (3 mg/kg/s.c.) or vehicle (100 µL, saline) 30 min before the tibio-tarsal joint injection of LTB4 (25 ng/5 µL). The flexion-elicited hypernociception was evaluated at indicated times. LTB4 tibio-tarsal joint-elicited hypernociceptive flexion depends on prostanoids. Mice were treated with indomethacin (5 mg/kg/i.p./40 min), MK886 (3 mg/kg/p.o./1 h), indomethacin plus MK886 (at the same doses), celecoxib (10 mg/kg/p.o./1 h), or vehicles (200 µL saline or tris/HCL buffer) before the tibio-tarsal joint injection of LTB4 (25 ng/5 µL). (B) LTB4 hypernociceptive effect is impaired in 5LO–/– mice. The 5LO–/– and WT mice were injected with saline (5 µL) or LTB4 (25 ng/5 µL) into the tibio-tarsal joint. In another group, the 5LO–/– mice were treated with indomethacin (same dose above) or vehicle (200 µL, tris/HCL buffer) 40 min before the tibio-tarsal joint injection of LTB4. The flexion-elicited hypernociception was evaluated at indicated times. (C) Impairment of LTB4-induced neutrophil influx into the tibio-tarsal joint of 5LO–/– mice. The articular region was colleted 5 h after LTB4 (25 ng/5 µL), saline (5 µL), or CL, and the MPO activity was measured. The results are expressed as the mean ± SEM of five animals per group. *, Significant difference when compared with the saline group; #, compared with the LTB4 and WT groups (P<0.05).
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Figure 4. Neutrophils mediate zymosan and LTB4 tibio-tarsal joint-elicited hypernociceptive flexion. (A and B) Mice were treated with fucoidan (20 mg/kg/i.v.), antineutrophil antibody (anti-N ; 100 µg/mice), or vehicle (vei-saline) 15 min before the tibio-tarsal joint injection of zymosan (30 µg/5 µL) or LTB4 (25 ng/5 µL). The flexion-elicited hypernociception was evaluated at indicated times. (C) Effect of the treatment with fucoidan (20 mg/kg/i.v) and antineutrophil antibody (100 µg/mice) on the influx of neutrophils into the tibio-tarsal joint of mice induced by LTB4 or zymosan. The articular region was colleted 5 h or 7 h after LTB4 (25 ng/5 µL) or zymosan (30 µg/5 µL), respectively, and the MPO activity was measured. Saline-injected joints (5 µL) or CL were used as controls. (D) Histological analysis of zymosan-induced inflammation in the tibio-tarsal joint of mice treated with fucoidan (20 mg/kg/i.v.). The articular region was collected after 7 h of stimulus injection for histological analysis. (E) Fucoidan and antineutrophil antibody treatments inhibit zymosan-induced LTB4 release into the joints. Mice were treated with fucoidan (20 mg/kg/i.v.), antineutrophil antibody (100 µg/mice), or vehicle (saline) 15 min before the tibio-tarsal joint injection of zymosan (30 µg/5 µL). The articular region was colleted 7 h after zymosan or saline injection. Processed samples were used for measurement of LTB4 levels by ELISA. (F) Fucoidan inhibits PGE2 release into the joints induced by LTB4. Mice were treated with fucoidan (20 mg/kg/i.v.) or vehicle (saline) 15 min before the tibio-tarsal joint injection of LTB4 (25 ng/5 µL). The articular region was colleted 5 h after LTB4 or saline injection. Processed samples were used for measurement of PGE2 levels by radioimmunoassay. The results are expressed as the mean ± SEM of five animals per group. *, Significant difference when compared with the saline group; #, compared with the LTB4 and zymosan groups (P<0.05).
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Among LTs, LTB4 is the most studied in acute Th1-inflammatory models. Numerous studies reported a significant increase in the levels of this eicosanoid in human inflammatory diseases, which are accompanied by hyperalgesia, including RA [16 , 31 , 33 , 34 ]. Recent studies demonstrated the crucial role of LTB4 acting on the BLT1 receptor in the genesis of arthritis. In arthritis models, it seems that LTB4 mediates neutrophil migration to the joints, which triggers further LTB4 production, in turn, inducing more neutrophil accumulation [34 35 36 ]. Besides its well-recognized chemoattractant effect, LTB4 also plays a role in the induction of inflammatory pain in rats and humans. These effects of LTB4 seem closely related with the presence of neutrophils [18 , 19 ]. In the current study, it seems that LTB4 is the 5LO product, which mediates zymosan-induced joint hypernociception as a BLT antagonist (CP105696) [37 ] reduced joint hypernociception. Moreover, a significant increase in LTB4 levels was observed in the arthritic joint of zymosan-injected mice, and LTB4 administration into the joint of naïve mice induced dose- and time-dependent dorsal flexion-elicited hypernociception.
Considering the above-mentioned evidence, the hypothesis that LTB4 induces hypernociception by a neutrophil-dependent mechanism was evaluated, and in turn, these leukocytes produce additional LTB4, amplifying and maintaining the hypernociceptive inflammatory process. In fact, the zymosan- and LTB4-induced hypernociception as well as neutrophil migration were reduced by MK886, fucoidan, or antineutrophil antibody treatment and in 5LO–/– mice, suggesting the participation of endogenous LTs and infiltrating neutrophils in the zymosan and LTB4 hypernociception. Moreover, fucoidin and antineutrophil antibody treatment reduced zymosan-induced LTB4 production, suggesting that neutrophils are the cells responsible for LTB4 production. In agreement, it was reported that neutrophils participate in the zymosan-induced arthritis in rats through the release of LTB4 [20 ].
Although in rat paws, the LTB4 hypernociceptive effect did not depend on prostanoids, we observed that in mice, LTB4-induced articular hypernociception was inhibited by indomethacin (nonselective cyclo-oxigenase inhibitor) and celecoxib (selective cyclo-oxigenase-2 inhibitor), suggesting that LTB4 mediates the prostanoid component of zymosan-induced hypernociception, mainly through the cyclo-oxigenase-2. Furthermore, administration of LTB4 into mice joints induced the production of PGE2. In accordance with these results, leukocytes stimulated with LTB4 release PGE2 [38 ]. Moreover, indomethacin almost blocked the flexion-elicited hypernociception in zymosan-induced joint inflammation [3 ]. Therefore, it is possible that at the same time that LTB4 induces the neutrophils/LTB4 pathway, it could also induce the production of prostanoids.
It is important to point out that in contrast with our and other mentioned studies, there are also evidences suggesting an analgesic role of neutrophils during the inflammatory process. For instance, it was demonstrated that during inflammation induced by CFA, the stimulation of recruited PMN with IL-1β, corticotropin release factor, or even with stress stimulus produces an antinociceptive effect dependent on PMN production of opioids [39 40 41 ]. These contradictions might be explained by the differences in the inflammatory models [42 ].
The hypernociceptive mechanism of LTB4 also seems to be dependent on other components besides the inflammatory component demonstrated in the present study. It seems that LTB4 might act directly on primary afferent neurons reducing the nociceptive threshold. For instance, LTB4 directly reduced the nociceptive threshold of sensory neurons in vitro and in vivo [43 44 45 46 ]. Moreover, sensitive neurons expressing transient receptor potential vanilloid receptor-1 receptors also express the BLT1 receptor [47 ]. This possible LTB4 effect was not investigated in our model, but it could contribute to the component of zymosan hypernociception. In line, the treatment of animals with dexamethasone, a glucocorticoid, at a dose that blocks the inflammatory component, inhibited the LTB4-induced hypernociception only partially.
Besides LTB4, cytokines also seem important for the genesis of hypernociception during zymosan-induced joint inflammation. For instance, drugs, which inhibited TNF- production, such as thalidomide, pentoxifiline, and residronate, ameliorate joint pain in the rats evaluated by an incapacitation test [48
49
50
]. Although the interaction between cytokines and LTB4 was not evaluated in the present experimental model, there is evidence in the literature that LTB4 mediates TNF--induced neutrophil migration and also that LTB4 stimulates the production of TNF- from murine macrophage [17
, 51
]. This interaction in zymosan-induced articular nociception is under evaluation by our group.
Taking together the present results points out the mechanism involved in LTB4 mediation of inflammatory joint hypernociception. It seems that LTB4 mediates zymosan hypernociception by inducing LTB4 production, which induces neutrophil migration, and in turn, these leukocytes produce additional LTB4 and PGE2 and possibly a direct action on the peripheral ending of sensory neurons. Figure 5 summarizes the hypernociceptive action of LTB4 in the present inflammatory model. Therefore, although a recent clinical trial demonstrated that an orally active BLT antagonist produced only a modest improvement of RA [52 ], more studies with other inhibitors of LT synthesis or action, particularly LTB4, could represent a beneficial therapy for joint inflammatory pain control.
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Figure 5. Representative diagram of the mechanism underling the LTB4 mediation of zymosan inflammatory hypernociception of the joint. The figure summarizes the essential conclusion of the present study, which shows the role of LTB4-stimulating neutrophil recruitment, which in turn produces additional LTB4. This pathway culminates with the release of the direct-acting hypernociceptive mediator PGE2. The possible effect of LTB4 as a direct-acting hypernociceptive mediator is also present [17
, 48
49
50
51
].
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Received February 23, 2007; revised August 31, 2007; accepted September 1, 2007.
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mechanonociceptors in the hairy skin of rat hindlimbs Neuroscience 22,651-659[CrossRef][Medline]This article has been cited by other articles:
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