IL-1Ra and vIL-10 gene transfer using retroviral vectors ameliorates particle-associated inflammation in the murine air pouch model

OBJECTIVE: This study examined anti- inflammatory gene therapy to ameliorate tissue responses to ultra high molecular weight polyethylene (UHMWPE) particles in the murine air pouch. METHODS: Retroviruses encoding human interleukin- 1 receptor antagonist (IL-1Ra), viral interleukin-10 (vIL-10), or LacZ (reporter) genes were injected into murine air pouches stimulated by UHMWPE particles. Pouch membranes and fluids were harvested at 1, 3 and 7 days post gene-transduction, and assayed for markers of inflammation using histological, molecular, and immunological techniques. RESULTS: Real time RT-PCR and ELISA showed a strong production of IL-1beta in pouch tissue and lavage fluid induced by particle stimulation, accompanied by a lower expression of IL-6, TNF-alpha and IL-4. Transduction of IL-1Ra or vIL-10 genes resulted in a significant reduction of IL-1beta both at the mRNA and at the protein level. The gene therapy also resulted in diminution of IL-6 and TNF-alpha expression. In addition, significant elevation of TGF-beta expression was observed in IL-1Ra transduced pouches. Histological analysis revealed that the membranes of pouches transduced with vIL-10 or IL-1Ra were significantly less inflamed than the membranes of non-viral and LacZ-transduced pouches, with less cellular proliferation and lowered monocyte/macrophage influx. CONCLUSIONS: IL-1Ra or vIL-10 gene transduction was effective in ameliorating local inflammation by reducing the IL-1 production and subsequent cellular events elicited in response to UHMWPE particles in this model. These findings suggest that IL-1 directed gene therapy might be excellent therapeutic candidates to prevent or retard the inflammatory response to wear debris that contributes to the pathology of aseptic loosening.     

Adenoviral delivery of IL-1 receptor antagonist abrogates disease activity during the development of autoimmune arthritis in IL-1 receptor antagonist-deficient mice

Currently available treatments for rheumatoid arthritis (RA) are limited in terms of their long-term effects and their abilities to control disease progression. Interleukin-1 receptor antagonist (IL-1Ra) is a natural inhibitor of the biologic actions of IL-1, which is known to promote inflammation and degeneration of the joint. In this study, we investigated whether human IL-1Ra gene transfer is effective at treating an established experimental arthritis model. A recombinant adenovirus carrying the gene that encode human hIL-1Ra and GFP (Ad.hIL-1Ra/GFP) was administered by intra-articular injection into the ankle joints of the mice with established the IL-1Ra-deficient Balb/cA mice (IL-1Ra(-/-)), which develop spontaneously chronic inflammatory arthropathy. The effects of two injections of Ad.hIL-1Ra/GFP or control virus with no inserted target gene (Ad.GFP) were compared with the effects of PBS injection with respect to the clinical characteristics of arthritis, as determined by articular index scores, histopathological and immunological assays. We further divided the outcomes of Ad.hIL-1Ra/GFP gene therapy in IL-1Ra(-/-) mice according arthritis stage; early stage and chronic stage corresponding to 8 and 15 weeks of age, respectively. Intra-articular injections of Ad.hIL-1Ra/GFP reduced arthritis severity and footpad swelling compared with control groups treated with Ad.GFP or PBS in early stage IL-1Ra(-/-) mice. Moreover, the histopathology of the ankle joints of IL-1Ra(-/-) mice treated with Ad.hIL-1Ra/GFP showed a significant decrease in synovial proliferation and inflammatory cell infiltration, and preserved proteoglycan levels in the joints of early stage IL-1Ra(-/-) mice compared with the control mice. Moreover, Ad.hIL-1Ra/GFP treated mice showed reduced levels of inflammatory T helper type 1 (Th1) driven IgG2a antibodies to collagen type II but increased levels Th2 driven IgG1 antibody. These results suggest that adenovirus-mediated gene transfer of IL-1Ra may be a promising therapeutic option in the early stage of autoimmune arthritis.     

Therapeutic Potential of the Proteasome Inhibitor Bortezomib on Titanium Particle-Induced Inflammation in a Murine Model

Wear particle-induced aseptic loosening has been recognized as a harmful inflammatory process that jeopardizes the longevity of total joint replacement. The proteasome controls the activation of NF-κB and subsequent inflammatory mediators, such as TNF-α and IL-1β; thus, we investigated whether proteasome inhibition can ameliorate wear particle-induced inflammation in a murine model. A total of 48 BALB/C mice were divided into four groups. Titanium (Ti) particles were injected into the established air pouches of all mice (except negative controls) to provoke inflammation, and then 0.1 or 0.5 mg/kg of Bortezomib (Bzb, a proteasome inhibitor) was administered to ameliorate the inflammation response, while air pouches without Bzb administration were used as loading controls. The air pouches were harvested 2 or 7 days after Bzb injection for molecular and histological analyses. Inflammation responses in the air pouch tissues of Bzb treatment groups are lower than those in the Ti-stimulated group, and this occurs in a dose-dependent manner. Bzb can significantly attenuate the severity of Ti-induced inflammation in air pouches.     

Comparative effects of tetrandrine and berbamine on subcutaneous air pouch inflammation induced by interleukin-1, tumour necrosis factor and platelet-activating factor

The effects of the bisbenzylisoquinoline alkaloids tetrandrine and berbamine on the action of IL-1, TNF and PAF were investigated in the rat subcutaneous air pouch model of inflammation. Both compounds were equipotent in the suppression of leukocyte infiltration into air pouches induced by IL-1 and TNF, with ED₅₀ values in the range 20–30 mg/kg/3 days. Both were also equiptent in suppression of PMN infiltration induced by PAF with ED₅₀ values in the same range as that for IL-1 and TNF. However, tetrandrine was more potent than berbamine as a suppressant of PAF-induced MNC infiltration, but much less potent than berbamine in carageenen-induced PMN infiltration. These results suggest that these bisbenzylisoquinolines may have value in the therapy of chronic inflammatory diseases where IL-1, TNF and PAF have a role in pathogenesis.     

Kakkalide and Its Metabolite Irisolidone Ameliorate Carrageenan-Induced Inflammation in Mice by Inhibiting NF-κB Pathway

The anti-inflammatory activities of kakkalide, a major constituent of the flower of Pueraria thunbergiana, and irisolidone, a metabolite of kakkalide produced by intestinal microflora, against carrageenan-induced inflammation in air pouches on the backs of mice and in lipopolysaccharide (LPS)-stimulated peritoneal macrophages were investigated. Kakkalide and irisolidone down-regulated the gene expression of cytokines [tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β)] and cyclooxygenase-2 (COX-2) and the production of pro-inflammatory cytokines, TNF-α and IL-1β, and inflammatory mediators, NO and prostaglandin E₂ (PGE₂), in LPS-stimulated peritoneal macrophages. These agents also inhibited the phosphorylation of IκB-α and the nuclear translocation of nuclear factor-kappa B (NF-κB). Orally administered kakkalide and irisolidone significantly reduced carrageenan-induced inflammatory markers, leukocyte number, and protein amount in the exudates of the air pouch. These constituents also inhibited PGE₂ production and COX-2 inducible nitric oxide synthase, IL-1β, and TNF-α expression. These agents also inhibited NF-κB activation. The anti-inflammatory effects of irisolidone were more potent than those of kakkalide. Based on these findings, kakkalide and irisolidone may inhibit inflammatory reactions via NF-κB pathway, and irisolidone, a metabolite of kakkalide, may more potently inhibit these inflammatory reactions.     

Increased inflammatory reactivity in newly formed lining tissue

The air pouch has been shown to provide a convenient model for studying the behaviour of synovial lining tissue. Air pouches of different ages were used to study the reactivity of newly developing lining tissue towards irritants known to cause inflammation. Pouches of 1 day in age were relatively inert in their reactivity as judged by the number of cells and volume of the exudate accumulating in the pouch. In contrast, 3-day-old pouches responded to a much greater extent, and 6-day-old pouches were highly responsive with a further increase in cell numbers and fluid volume. The different responses of 1-, 3- and 6-day-old pouches could be explained by (a) developing vascularity of the pouch, (b) formation of an organised lining of phagocytic cells, or (c) an increasingly organised mechanical barrier that retains the irritant and products of the inflammatory response. These studies of air pouch lining development permit a dissection of those components necessary for inflammatory reactivity of a lining tissue and may help explain the sensitivity of synovium to chronic inflammation.     

The IL-1 system in HIV infection: peripheral concentrations of IL-1β, IL-1 receptor antagonist and soluble IL-1 receptor type II

The proinflammatory cytokine IL-1β is thought to be involved in ongoing HIV disease. Furthermore, its naturally occurring inhibitors soluble IL-1 receptor type II (sIL-1RII) and IL-1 receptor antagonist (IL-1Ra) may play a pivotal role in regulating its biological action. To investigate the involvement of the IL-1 system we determined serum levels of IL-1β, IL-1Ra and sIL-1RII in 90 HIV⁺ patients. The obtained values were compared with markers of disease progression such as CD⁺ count, 5′-neopterin, β₂-microglobulin and soluble tumour necrosis factor receptors (sTNF-R) p55 and p75 and then compared with C-reactive protein (CRP), granulocyte count, lL-6 and TNF-α. While IL-1Ra concentrations increased significantly with progressive CDC disease stages, sIL-1RII and IL-1β were not altered in our cohort. IL-1Ra showed statistical relation to decreasing CD4⁺ lymphocytes and increasing 5′-neopterin, β₂-microglobulin, sTNF-R p55, sTNF-R p75. Furthermore, IL-1Ra correlated positively with serum IL-6, TNF-α, CRP and granulocytes. In contrast, sIL-1RII and IL-1β tended to show an inverse correlation or showed no significant relationship to all these parameters. Il-1β was measurable only in a limited number of samples. IL-1Ra showed a clear relationship to acute inflammatory events as well as to the different disease stages. Our data suggest a dissociation between IL-1Ra and sIL-1RII serum levels which may indicate that the two IL-1 binding proteins have different pathophysiological roles in HIV infection.     

Distinct gene expression of receptor activator of nuclear factor-kappaB and rank ligand in the inflammatory response to variant morphologies of UHMWPE particles

Recent studies have examined the role of wear debris-induced bone resorption in the aseptic loosening of orthopedic prostheses. Research has shown that inflammation depends not only on the amount of particulate debris, but also the shape and size of the accumulated wear particles. Our previous studies have demonstrated that variant shapes of ultra-high molecular weight polyethylene (UHMWPE) particles induce diverse cellular and apoptotic responses in a murine inflammation model. Since enhanced osteoclastogenesis is recognized as a hallmark of bone loss in prosthetic loosening, we have now investigated the gene expression of receptor activator of nuclear factor-kappaB (RANK) and receptor activator of nuclear factor-kappaB ligand (RANKL) during the inflammatory response to different shapes of UHMWPE particles. Two shapes of UHMWPE particles (globular or elongated) were implanted in established air pouches on BALB/c mice, and pouches harvested 7 days after stimulation with UHMWPE particles. Gene levels of RANK, RANKL, TNFalpha, IL-1beta, and cathepsin K (CK) were quantified by real time RT-PCR, and TRAP staining of pouch membrane was used to evaluate osteoclastogenesis. We found that (i) elongated particles generated significantly higher RANK and RANKL gene expression than globular particles in pouch tissue; (ii) elongated particles provoked significantly higher IL-1beta and TNFalpha gene expression; (iii) a positive association was found between tissue inflammation status and the gene level of RANK/RANKL; and (iv) elongated particles stimulated significantly higher CK gene expression in comparison with globular particles. Histology revealed that clusters of TRAP+ cells were located in regions in contact with elongated particles. Overall, these data suggest that the morphology of wear debris may be a critical factor in the pathogenesis of prosthetic loosening.     

Macrophage depletion diminishes implant-wear-induced inflammatory osteolysis in a mouse model

The purpose of this study was to determine whether macrophage depletion using clodronate liposomes diminishes wear-debris-induced inflammatory osteolysis in a murine osteolysis model. Ultra high molecular weight polyethylene (UHMWPE) particles were introduced into established air pouches on BALB/c mice, followed by implantation of calvaria bone from syngeneic littermates. Macrophages were depleted by the intraperitoneal injection of clodronate liposome (2 mg) 2 days before bone implantation and re-injection every 3 days (1 mg) until the sacrifice of the mice. Mice without clodronate liposome therapy or treated with empty liposome as well as mice injected with saline alone were included in this study as controls. Pouch tissues were collected 14 days after bone implantation for molecular and histology analysis. Our findings indicated that (1) macrophage depletion in clodronate-liposome-treated mice was achieved, as illustrated by F4/80 immunostaining in both pouch and spleen tissues; (2) clodronate-liposome treatment significantly reduced UHMWPE-induced tissue inflammation, with diminished pouch membrane thickness, reduced inflammatory cellular infiltration, and lowered interleukin 1beta (IL-1beta) and tumor necrosis factor alpha (TNFalpha) expression; (3) clodronate-liposome treatment markedly reduced the number of TRAP(+) cells in pouch tissues and protected against bone collagen depletion. In conclusion, this study demonstrates that macrophage depletion using clodronate-liposome reduces UHMWPE particle-induced inflammatory osteolysis. This observation supports the hypothesis that macrophages contribute to the severity of UHMWPE particles induced inflammatory osteolysis, and suggest that macrophage depletion represents a viable therapeutic approach to the prevention and treatment of patients with aseptic loosening.     

An Interaction Between Genetic Factors and Gender Determines the Magnitude of the Inflammatory Response in the Mouse Air Pouch Model of Acute Inflammation

The widely used mouse air pouch model of acute inflammation is inducible in a variety of inbred strains, but the potential influence of genetic background and gender on inflammation severity has never been examined. We directly compared the degree of inflammation induced in the air pouch model across four commonly utilized inbred strains in both male and female mice. We then applied an in silico mapping method to identify loci potentially associated with determining inflammation severity for each gender. Air pouches were induced by subcutaneous injection 3 (3 cc) and 5 (1.5 cc) days prior to the experiment. 4h after carrageenan injection, exudates were retrieved and leukocyte concentration quantified using a hemocytometer. The in silico mapping method was applied as described below. The strain order for mean leukocyte count/mL in inflamed exudates differed between genders. In males, the order was C57BL/6J > BALB/cByJ > DBA/2J > DBA/1J, while in females the order was BALB/cByJ > DBA/2J > C57BL/6J > DBA/1J. The difference in inflammation severity between genders reached significance only in C57BL/6J mice. Independent in silico analysis based on phenotypic data from male versus female mice identified distinct sets of loci as potentially associated with the exudate count reached. We conclude that the degree of inflammation induced in the mouse air pouch model of inflammation is strain-specific and, therefore, genetically based, and the pattern of interstrain differences is altered in male relative to female mice. The loci identified by in silico mapping likely contain genes with differential roles in determining this phenotype between genders.     

Mice deficient in hepatocyte-specific IL-1Ra show delayed resolution of concanavalin A-induced hepatitis

Interleukin-1 receptor antagonist (IL-1Ra) is a specific IL-1 inhibitor that possesses anti-inflammatory activities. Several studies in human and mouse suggested a protective role for IL-1Ra in liver inflammation, and we previously demonstrated that hepatocytes produce high levels of IL-1Ra in response to inflammatory challenge in vitro and in vivo. In the present study, we investigated the production and the biological function of hepatocyte-derived IL-1Ra in concanavalin A (ConA)-induced hepatitis in mice. We show that the injured liver produces large amounts of IL-1Ra and that secreted and intracellular IL-1Ra isoforms are produced with different kinetics during the course of hepatitis. By using hepatocyte-specific IL-1Ra-deficient mice (IL-1Ra(ΔH)), we demonstrate that hepatocytes represent the major cellular source of local IL-1Ra. Most interestingly, hepatic necrosis and inflammation were increased in IL-1Ra(ΔH) as compared with wild-type mice during the late phase of the disease, leading to a delayed resolution of hepatitis in IL-1Ra(ΔH) mice. In conclusion, our results show that the local production of IL-1Ra by hepatocytes contributes to the resolution of hepatitis.     

Modulation of infection‐mediated migration of neutrophils and CXCR2 trafficking by osteopontin

Osteopontin (OPN) is a pro‐inflammatory protein that paradoxically protects against inflammation and bone destruction in a mouse model of endodontic infection. Here we have tested the hypothesis that this effect of OPN is mediated by effects on migration of innate immune cells to the site of infection. Using the air pouch as a model of endodontic infection in mice, we showed that neutrophil accumulation at the site of infection with a mixture of endodontic pathogens is significantly reduced in OPN‐deficient mice. Reduced neutrophil accumulation in the absence of OPN was accompanied by an increase in bacterial load. OPN‐deficiency did not affect neutrophil survival, CXCR2 ligand expression, or the production of inflammatory cytokines in the air pouch. In vitro, OPN enhanced neutrophil migration to CXCL1, whereas in vivo, inhibition of CXCR2 suppressed cellular infiltration in air pouches of infected wild‐type mice by > 50%, but had no effect in OPN‐deficient mice. OPN increased cell surface expression of CXCR2 on bone marrow neutrophils in an integrin‐α_v‐dependent manner, and suppressed the internalization of CXCR2 in the absence of ligand. Together, these results support a model where the protective effect of OPN results from enhanced initial neutrophil accumulation at sites of infection resulting in optimal bacterial killing. We describe a novel mechanism for this effect of OPN: integrin‐α_v‐dependent suppression of CXCR2 internalization in neutrophils, which increases the ability of these cells to migrate to sites of infection in response to CXCR2 ligands.     

Enhancement of neutrophil infiltration in histidine decarboxylase-deficient mice

The roles of histamine in the anaphylactic increase in vascular permeability and leucocyte infiltration were analysed in an air pouch-type allergic inflammation model in histidine decarboxylase-deficient (HDC^−/−) mice and wild-type mice. In the immunized wild-type mice, histamine content in the pouch fluid and vascular permeability in the anaphylaxis phase were increased by injection of the antigen solution into the air pouch. However, in the immunized HDC^−/− mice, the antigen challenge did not increase histamine content in the pouch fluid and vascular permeability in the anaphylaxis phase. Number of leucocytes (more than 83% are neutrophils) in the pouch fluid 4–24 hr after the antigen challenge in the HDC^−/− mice was significantly higher than that in the wild-type mice. Simultaneous injection of histamine with the antigen solution into the air pouch of the immunized HDC^−/− mice reduced the antigen-induced leucocyte infiltration at 4 hr. Simultaneous injection of the H2 antagonist cimetidine but not the H1 antagonist pyrilamine with the antigen solution into the air pouch of the immunized wild-type mice further increased leucocyte infiltration at 4 hr. The levels of macrophage inflammatory protein-2 at 2 hr and of tumour necrosis factor-α at 4 hr in the pouch fluid of the HDC^−/− mice were significantly higher than those of the wild-type mice. These findings indicate that histamine plays significant roles not only in the anaphylactic increase in vascular permeability via H1 receptors but also in the negative regulation of neutrophil infiltration via H2 receptors in allergic inflammation.     

Interleukin-1 receptor antagonist attenuates airway hyperresponsiveness following exposure to ozone

The role of an interleukin (IL)-1 receptor antagonist (IL-1Ra) on the development of airway hyperresponsiveness (AHR) and airway inflammation following acute O(3) exposure in mice was investigated. Exposure of C57/BL6 mice to O(3) at a concentration of 2.0 ppm or filtered air for 3 h resulted in increases in airway responsiveness to inhaled methacholine (MCh) 8 and 16 h after the exposure, and an increase in neutrophils in the bronchoalveolar lavage (BAL) fluid. IL-1beta expression, assessed by gene microarray, was increased 2-fold 4 h after O(3) exposure, and returned to baseline levels by 24 h. Levels of IL-1beta in lung homogenates were also increased 8 h after O(3) exposure. Administration of (human) IL-1Ra before and after O(3) exposure prevented development of AHR and decreased BAL fluid neutrophilia. Increases in chemokine levels in lung homogenates, tumor necrosis factor-alpha, MIP-2, and keratinocyte chemoattractant following O(3) exposure were prevented by IL-1Ra. Inhalation of dexamethasone, an inhibitor of IL-1 production, blocked the development of AHR, BAL fluid neutrophilia, and decreased levels of IL-1 following O(3) exposure. In summary, acute exposure to O(3) induces AHR, neutrophilic inflammation, epithelial damage, and IL-1. An IL-1Ra effectively prevents the development of altered airway function, inflammation, and structural damage.     

Compartmentalized transgene expression of granulocyte-macrophage colony-stimulating factor (GM-CSF) in mouse lung enhances allergic airways inflammation

To investigate the role of GM-CSF in asthmatic airways inflammation, we have targeted GM-CSF transgene to the airway cells in a mouse model of ovalbumin (OVA)-induced allergic airways inflammation, a model in which there is marked induction of endogenous IL-5 and IL-4 but not GM-CSF. Following intranasal delivery of a replication-deficient adenoviral gene transfer vector (Ad), transgene expression was found localized primarily to the respiratory epithelial cells. Intranasal delivery of 0.03 × 10⁹ plaque-forming units (PFU) of AdGM-CSF into naive BALB/c mice resulted in prolonged and compartmentalized release of GM-CSF transgene protein with a peak concentration of ≈ 80 pg/ml detected in bronchoalveolar lavage fluid (BALF) at day 7, but little in serum. These levels of local GM-CSF expression per se resulted in no eosinophilia and only a minimum of tissue inflammatory responses in the lung of naive mice, similar to those induced by the control vector. However, such GM-CSF expression in the airways of OVA-sensitized mice resulted in a much greater and sustained accumulation of various inflammatory cell types, most noticeably eosinophils, both in BALF and airway tissues for 15–21 days post-OVA aerosol challenge, at which times airways inflammation had largely resolved in control mice. While the levels of IL-5 and IL-4 in BALF and the rate of eosinophil apoptosis were found similar between different treatments, there was an increased number of proliferative leucocytes in the lung receiving GM-CSF gene transfer. Our results thus provide direct experimental evidence that GM-CSF can significantly contribute to the development of allergic airways inflammation through potentiating and prolonging inflammatory infiltration induced by cytokines such as IL-5 and IL-4.     

Lymphocyte kinetics in a murine model of chronic inflammation

A modification of the air pouch system [1], has been developed to investigate the role of lymphocytes in an inmmunologically driven model of chronic inflammation. Radiolabelled spleen and lymph node mononuclear cells from mice presensitised withBordatella pertussis vaccine (BPV) were infused intravenously into mice also presensitised with BPV and challenged with BPV into 6 day old air pouches. Cell migration was monitored by gamma counting of tissues sampled 4 hours after infusion. The percentage increase in counts obtained in air pouch tissue compared to control skin, over a time course of 30 days, reached a peak at 10 days after BPV challenge. Vessels with features of high endothelial venules, and clusters of lymphocytes have been demonstrated histologically and immunohistochemically in air pouch tissue at this time.     

Inhibitory effects of erythromycin on wear debris-induced VEGF/Flt-1 gene production and osteolysis

Objectives  A highly vascularized and inflammatory periprosthetic tissue augments the progress of aseptic loosening, a major clinical problem after total joint replacement. The purpose of this study is to investigate the effect of erythromycin (EM) on ultra high molecular weight polyethylene (UHMWPE) particle-induced VEGF/VEGF receptor 1 (Flt-1) gene production and inflammatory osteolysis in a mouse model. Methods  UHMWPE particles were introduced into established air pouches on BALB/c mice, followed by implantation of calvaria bone from syngeneic littermates. EM treatment started 2 weeks after bone implantation (5 mg/kg day, i.p. injection). Mice without drug treatment as well as mice injected with saline alone were included. Pouch tissues were harvested 2 weeks after bone implantation. Expression of VEGF, Flt-1, RANKL, IL-1, TNF and CD68 was measured by immunostain and RT-PCR, and implanted bone resorption was analyzed by micro-CT (μCT). Results  Exposure to UHMWPE induced pouch tissue inflammation, increase of VEGF/Flt-1 proteins, and increased bone resorption. EM treatment significantly improved UHMWPE particle-induced tissue inflammation, reduced VEGF/Flt-1 protein expression, and diminished the number of TRAP⁺ cells, as well as the implanted bone resorption. Conclusion  This study demonstrated that EM inhibited VEGF and Flt-1 gene expression. The molecular mechanism of EM action on VEGF/Flt-1 signaling-mediated osteoclastogenesis warrants further investigation.     

Modulation of proinflammatory cytokine production in tumour necrosis factor-alpha (TNF-α)-transgenic mice by treatment with cells engineered to secrete IL-4, IL-10 or IL-13

TNF-α is one of the major proinflammatory cytokines involved in the pathogenesis of chronic inflammatory joint disease, in human rheumatoid arthritis as well as in murine models of this disease. It was previously described that a highly destructive chronic spontaneous inflammatory arthritis develops in mice expressing a human TNF-α transgene modified with the 3′ untranslated region of β-globin. The present study investigates in this mouse model the effects of the anti-inflammatory cytokines IL-4, IL-10 and IL-13 administered in vivo on proinflammatory cytokine expression. Groups of TNF-α-transgenic mice were engrafted with xenogeneic transfected Chinese hamster ovary (CHO) fibroblasts secreting murine IL-4, IL-10 or IL-13. In vivo treatments consisted of 3 or 4 weekly engraftments, starting when the mice were 4 weeks old. Control groups of transgenic mice were engrafted with β-galactosidase gene-transfected CHO cells or injected with medium. A significant decreased expression of TNF-α transgene, endogenous mouse TNF-α and IL-1 mRNA was observed in splenocytes of mice treated for 3 or 4 weeks with CHO/IL-4 and CHO/IL-13, and, to a lesser extent, with CHO/IL-10, compared with controls. Finally, attenuation of histological scores of arthritides was statistically significant only in the group of CHO/IL-4-treated mice after 3 weeks of treatment (P < 0.05), and was not significant in any other group. These results show that IL-4, IL-10 or IL-13, administered by gene therapy, can decrease the mRNA steady state levels of both endogenous and transgenic cytokines in human TNF-α transgenic mice. In addition, IL-4 can slightly attenuate the development of arthritides in this model.     

The release of interleukin-1-like activity by macrophages in two models of immunological inflammation in the rat

Interleukin-1 (IL-1) is a putative mediator of inflammation released by activated macrophagesin vitro. The IL-1 release by rat macrophages collected either from exudates in pertussis-induced air pouches or from the peritoneum during adjuvant arthritis has been investigated. In air pouch inflammation LPS-stimulated macrophages collected from sensitized animals release more IL-1 than cells from control rats at day 6 after challenge. This enhanced IL-1 release parallels the extent of mononuclear cell migration in the inflammatory lesion. In adjuvant arthritis LPS-stimulated macrophages collected from sensitized animals release more IL-1 than cells from control rats at days 16 and 23 after adjuvant injection. The secondary inflammation in arthritic rats was statistically significant at days 16 to 28. These results indicate that during immunological inflammation macrophages either from the inflamed area or from a non-inflamed region release more IL-1 than control cells. This release parallels the extent of the inflammation and may be important in its pathogenesis.