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The pro-inflammatory cytokine IL-17A has been implicated in the immunopathology of inflammatory arthritis. IL-17F bears 50% homology to IL-17A and has recently been suggested to play a role in inflammation. We investigated the induction and cytokine profile of IL-17F+ CD4+ T cells, and how IL-17F may contribute to inflammation. Upon culture of healthy donor CD4+ T cells with IL-1β, IL-23, anti-CD3, and anti-CD28 mAb, both IL-17A and IL-17F-expressing cells were detected. In comparison to IL-17A+ IL-17F- CD4+ T cells, IL-17F+ IL-17A- and IL-17A+ IL-17F+ CD4+ T cells contained lower proportions of IL-10-expressing and GM-CSF-expressing cells and higher proportions of IFN-γ-expressing cells. Titration of anti-CD28 mAb revealed that strong co-stimulation increased IL-17F+ IL-17A- and IL-17A+ IL-17F+ CD4+ T cell frequencies, whereas IL-17A+ IL-17F- CD4+ T cell frequencies decreased. This was partly mediated via an IL-2-dependent mechanism. Addition of IL-17A, IL-17F, and TNF-α to synovial fibroblasts from patients with inflammatory arthritis resulted in significant production of IL-6 and IL-8, which was reduced to a larger extent by combined blockade of IL-17A and IL-17F than blockade of IL-17A alone. Our data indicate that IL-17A and IL-17F are differentially regulated upon T cell co-stimulation, and that dual blockade of IL-17A and IL-17F reduces inflammation more effectively than IL-17A blockade alone.

See Zekeridou and Pittock (doi:10.1093/brain/awaa153) for a scientific commentary on this article. Ramberger et al. generate patient-derived monoclonal antibodies against LGI1 and describe their epitopes and sequences, plus their pathogenic potential in vitro and in vivo. They find that binding characteristics of these antibodies inform the molecular mechanisms and relative pathogenicity within polyclonal specificities., Autoantibodies against leucine-rich glioma inactivated 1 (LGI1) are found in patients with limbic encephalitis and focal seizures. Here, we generate patient-derived monoclonal antibodies (mAbs) against LGI1. We explore their sequences and binding characteristics, plus their pathogenic potential using transfected HEK293T cells, rodent neuronal preparations, and behavioural and electrophysiological assessments in vivo after mAb injections into the rodent hippocampus. In live cell-based assays, LGI1 epitope recognition was examined with patient sera (n = 31), CSFs (n = 11), longitudinal serum samples (n = 15), and using mAbs (n = 14) generated from peripheral B cells of two patients. All sera and 9/11 CSFs bound both the leucine-rich repeat (LRR) and the epitempin repeat (EPTP) domains of LGI1, with stable ratios of LRR:EPTP antibody levels over time. By contrast, the mAbs derived from both patients recognized either the LRR or EPTP domain. mAbs against both domain specificities showed varied binding strengths, and marked genetic heterogeneity, with high mutation frequencies. LRR-specific mAbs recognized LGI1 docked to its interaction partners, ADAM22 and ADAM23, bound to rodent brain sections, and induced internalization of the LGI1-ADAM22/23 complex in both HEK293T cells and live hippocampal neurons. By contrast, few EPTP-specific mAbs bound to rodent brain sections or ADAM22/23-docked LGI1, but all inhibited the docking of LGI1 to ADAM22/23. After intrahippocampal injection, and by contrast to the LRR-directed mAbs, the EPTP-directed mAbs showed far less avid binding to brain tissue and were consistently detected in the serum. Post-injection, both domain-specific mAbs abrogated long-term potentiation induction, and LRR-directed antibodies with higher binding strengths induced memory impairment. Taken together, two largely dichotomous populations of LGI1 mAbs with distinct domain binding characteristics exist in the affinity matured peripheral autoantigen-specific memory pools of individuals, both of which have pathogenic potential. In human autoantibody-mediated diseases, the detailed characterization of patient mAbs provides a valuable method to dissect the molecular mechanisms within polyclonal populations.

The pro‐inflammatory cytokine IL‐17A has been implicated in the immunopathology of inflammatory arthritis. IL‐17F bears 50% homology to IL‐17A and has recently been suggested to play a role in inflammation. We investigated the induction and cytokine profile of IL‐17F+ CD4+ T cells, and how IL‐17F may contribute to inflammation. Upon culture of healthy donor CD4+ T cells with IL‐1β, IL‐23, anti‐CD3, and anti‐CD28 mAb, both IL‐17A and IL‐17F‐expressing cells were detected. In comparison to IL‐17A+IL‐17F− CD4+ T cells, IL‐17F+IL‐17A− and IL‐17A+IL‐17F+ CD4+ T cells contained lower proportions of IL‐10‐expressing and GM‐CSF‐expressing cells and higher proportions of IFN‐γ‐expressing cells. Titration of anti‐CD28 mAb revealed that strong co‐stimulation increased IL‐17F+IL‐17A− and IL‐17A+IL‐17F+ CD4+ T cell frequencies, whereas IL‐17A+IL‐17F− CD4+ T cell frequencies decreased. This was partly mediated via an IL‐2‐dependent mechanism. Addition of IL‐17A, IL‐17F, and TNF‐α to synovial fibroblasts from patients with inflammatory arthritis resulted in significant production of IL‐6 and IL‐8, which was reduced to a larger extent by combined blockade of IL‐17A and IL‐17F than blockade of IL‐17A alone. Our data indicate that IL‐17A and IL‐17F are differentially regulated upon T cell co‐stimulation, and that dual blockade of IL‐17A and IL‐17F reduces inflammation more effectively than IL‐17A blockade alone., High‐strength T cell stimulation enhances the frequency of IL‐17F+ CD4+ T cells in part via an IL‐2 dependent manner. Results demonstrate IL‐17A+ and IL‐17F+ CD4+ T cell populations display different cytokine profiles. Combined blockade of IL‐17A and IL‐17F is more effective at reducing inflammation than blockade of IL‐17A alone.

The costimulatory receptor OX40 is expressed on activated T cells and regulates T-cell responses. Here, we show the efficacy and mechanism of action of an OX40 blocking antibody using the chronic house dust mite (HDM) mouse model of lung inflammation and in vitro HDM stimulation of cells from HDM allergic human donors. We have demonstrated that OX40 blockade leads to a reduction in the number of eosinophils and neutrophils in the lavage fluid and lung tissue of HDM sensitized mice. This was accompanied by a decrease in activated and memory CD4(+) T cells in the lungs and further analysis revealed that both the Th2 and Th17 populations were inhibited. Improved lung function and decreased HDM-specific antibody responses were also noted. Significantly, efficacy was observed even when anti-OX40 treatment was delayed until after inflammation was established. OX40 blockade also inhibited the release of the Th2 cytokines IL-5 and IL-13 from cells isolated from HDM allergic human donors. Altogether, our data provide evidence of a role of the OX40/OX40L pathway in ongoing allergic lung inflammation and support clinical studies of a blocking OX40 antibody in Th2 high severe asthma patients.

Interleukin-6 (IL-6) is a critical regulator of the immune system and has been widely implicated in autoimmune disease. Here, we describe the discovery and characterization of olokizumab, a humanized antibody to IL-6. Data from structural biology, cell biology and primate pharmacology demonstrate the therapeutic potential of targeting IL-6 at “Site 3”, blocking the interaction with the signaling co-receptor gp130.

Objective Exposure to supraphysiologic levels of glucocorticoid drugs is known to have detrimental effects on bone formation and linear growth. Patients with sclerosteosis lack the bone regulatory protein sclerostin, have excessive bone formation, and are typically above average in height. This study was undertaken to characterize the effects of a monoclonal antibody to sclerostin (Scl-AbI) in mice exposed to dexamethasone (DEX). Methods Young mice were concomitantly treated with DEX (or vehicle control) and Scl-AbI antibody (or isotype-matched control antibody [Ctrl-Ab]) in 2 independent studies. Linear growth, the volume and strength of the bones, and the levels of bone turnover markers were analyzed. Results In DEX-treated mice, Scl-AbI had no significant effect on linear growth when compared to control treatment (Ctrl-Ab). However, in mice treated with DEX and Scl-ABI, a significant increase in trabecular bone at the femoral metaphysis (bone volume/total volume +117% versus Ctrl-Ab–treated mice) and in the width and volume of the cortical bone at the femoral diaphysis (+24% and +20%, respectively, versus Ctrl-Ab–treated mice) was noted. Scl-AbI treatment also improved mechanical strength (as assessed by 4-point bending studies) at the femoral diaphysis in DEX-treated mice (maximum load +60% and ultimate strength +47% in Scl-AbI–treated mice versus Ctrl-Ab–treated mice). Elevated osteocalcin levels were not detected in DEX-treated mice that received Scl-AbI, although levels of type 5b tartrate-resistant acid phosphatase were significantly lower than those observed in mice receiving DEX and Ctrl-Ab. Conclusion Scl-AbI treatment does not prevent the detrimental effects of DEX on linear growth, but the antibody does increase both cortical and trabecular bone and improves bone mechanical properties in DEX-treated mice.

Background: Intestinal inflammation associated with inflammatory bowel disease (IBD) is typically characterized by an inflammatory cell infiltrate and pro-inflammatory cytokine production. Of particular interest, the frequency of colony stimulating factor-1 (CSF-1)-expressing cells is increased in active lesions. In this study, we have investigated the role of CSF-1 in mucosal inflammation, using a murine model of colitis induced by dextran sulfate sodium (DSS). Methods: A neutralizing anti-CSF-1 antibody was administered to Balb/c mice that received DSS in their drinking water. Signs of colitis, such as clinical disease score, cellular infiltrate, and cytokine production, were assessed. Results: Administration of a neutralizing anti-CSF-1 antibody significantly inhibited DSS-induced colitis. Clinical symptoms, such as weight loss and the appearance of diarrhea or fecal blood, were reduced by CSF-1 blockade; histologic scores were also improved. The cellular infiltrate of macrophages and T cells was inhibited and a trend toward reduced production of pro-inflammatory cytokines was noted. Conclusions: This is the first study to demonstrate that CSF-1 plays an important role in mediating intestinal mucosal inflammation and therefore may prove to be an attractive therapeutic target for intestinal diseases such as inflammatory bowel disease. (Inflamm Bowel Dis 2006)

To examine the extent to which weight gain and weight status in the first 2 years of life relate to the risk of neurodevelopmental impairment in extremely preterm infants.In a cohort of 1070 infants born between 23 and 27 weeks' gestation, we examined weight gain from 7-28 days of life (in quartiles) and weight z-score at 12 and 24 months corrected age (in 4 categories:-2; ≥-2,-1; ≥1,1; and ≥1) in relation to these adverse neurodevelopmental outcomes: Bayley-II mental development index55, Bayley-II psychomotor development index55, cerebral palsy, Gross Motor Function Classification System ≥1 (cannot walk without assistance), microcephaly. We adjusted for confounders in logistic regression, stratified by sex, and performed separate analyses including the entire sample, and excluding children unable to walk without assistance (motor impairment).Weight gain in the lowest quartile from 7-28 days was not associated with higher risk of adverse outcomes. Children with a 12-month weight z-score-2 were at increased risk for all adverse outcomes in girls, and for microcephaly and Gross Motor Function Classification System ≥1 in boys. However, excluding children with motor impairment attenuated all associations except that of weight z-score-2 with microcephaly in girls. Similarly, most associations of low weight z-score at 24 months with adverse outcomes were attenuated with exclusion of children with motor impairment.Excluding children who have gross motor impairment appears to eliminate the association of low weight status with neurodevelopmental impairments at 2 years in extremely preterm infants.

The study investigated the role of P-selectin in the responses of eosinophil transmigration and eotaxin generation in vivo elicited by interleukin (IL)-13, as compared with IL-4. Two murine models of leukocyte transmigration were used, migration into cytokine-stimulated peritoneal cavities and through stimulated cremasteric venules, as observed by intravital microscopy. In mice lacking P-selectin, eosinophil infiltration elicited by the cytokines in the peritonitis model was totally inhibited. In the cremaster muscle, however, although spontaneous leukocyte-rolling flux and stimulated leukocyte firm adhesion were inhibited by ∼97% and ∼48%, respectively, stimulated transmigration was unaffected. However, IL-13-induced leukocyte transmigration was totally blocked in P-selectin-deficient mice treated with an anti-α4 integrin monoclonal antibody (mAb; PS/2). In comparison, treatment of wild-type mice with the anti-α4 integrin mAb resulted in only partial suppression of IL-13-induced leukocyte transmigration. Significant levels of eotaxin were detected in response to IL-13/IL-4 in both tissues in P-selectin-deficient animals. In conclusion, the regulatory role of P-selectin in leukocyte transmigration elicited by IL-13 appears to be tissue-specific, a phenomenon that is independent of the ability of the cytokine to stimulate eotaxin generation.

This article discusses the potential for clinical translation of the large amount of information on the molecular basis of leukocyte-endothelial cell interactions that has been collected over the last twenty years. Areas of current interest include the identification of adhesion molecule expression in inflammation by diagnostic imaging, understanding variability in inflammatory responsiveness and disease susceptibility through identification of adhesion molecule and chemokine polymorphisms and the application to the treatment of inflammatory diseases of monoclonal antibodies and conventional drugs with specific actions on leukocyte adhesion and migration.

Objective. We have hypothesized that the process of monocyte to macrophage differentiation may alter the inflammatory response of mononuclear phagocytes to the uptake of monosodium urate monohydrate (MSU) crystals. Methods. Eight mouse monocyte/macrophage cell lines were arranged in increasing order of differentiation, as judged by expression of the macrophage markers F4/80 and BM 8 and by phagocytic capacity. Secretion of tumor necrosis factor a (TNFa) in response to MSU was measured by enzyme-linked immunosorbent assay. Results. The panel of monocyte/macrophage cell lines revealed a close linkage between the state of differentiation and the capacity of the cells to ingest MSU crystals. TNFa production, however, was not linked to phagocytic ability. Peak TNFa levels were synthesized by cells at an intermediate state of differentiation (3.2‐14.1 ng/ml), whereas mature macrophages, which efficiently phagocytosed crystals, did not secrete TNFa. Mature cell lines produced TNFa when stimulated with zymosan (5.9‐6.2 ng/ml), but this was abolished by coincubation with MSU crystals. Suppression of the zymosan response was not due to apoptosis or steric hindrance by MSU crystals. Culture supernatants from mature macrophages did not stimulate endothelial cell activation, in contrast to MSU-treated cells at an earlier stage of differentiation, which stimulated intercellular adhesion molecule 1 expression on sEND endothelioma cells through the release of TNFa (inhibited 80.6% by anti-TNFa). Conclusion. We demonstrated that phagocytosis and TNFa production are distinct events in the response of mononuclear phagocytes to urate crystals, and these events can be distinguished at the level of macrophage differentiation. The noninflammatory removal of urate crystals by mature macrophages defines a new pathway that may be important in controlling the development of acute gout in patients with hyperuricemia.

The selectins are adhesion molecules that mediate the tethering and rolling of leukocytes on vascular endothelium. Although E-selectin and P-selectin are known to be expressed by endothelial cells (EC) in response to proinflammatory stimuli, their pattern and mechanisms of expression in immune-mediated inflammation remain poorly understood. By quantifying luminal endothelial selectin expression via i.v. administration of radiolabeled mAb, we detected constitutive expression of P-selectin, but not E-selectin, in mouse skin. Both selectins were transiently up-regulated after intradermal TNF-α, IL-1α, or IL-1β. In contrast, during a contact sensitivity response to oxazolone, expression of both selectins was prolonged, with distinct peaks at 6 and 48 h. Experiments with P-selectin gene-targeted mice showed that the P-selectin measured was exclusively expressed by EC rather than platelets. The early and late phases of selectin expression in contact sensitivity were differentiated in terms of their requirement for prior sensitization, and the action of IL-1. Whereas the early phase was a nonspecific ’irritant’ response to oxazolone, the late phase was Ag specific and was partially IL-1 dependent. Therefore, persistence of both E- and P-selectin expression in vivo can occur as a result of sequential and distinct EC activation processes that appear to be at least partially different from those previously reported as stimulating ICAM-1 and VCAM-1 expression. The further elucidation of mechanisms of EC activation in this model may help determine the relative roles of selectins and ligands for leukocyte integrins in the sequential recruitment of T cells and other leukocyte subsets during ongoing immune-mediated inflammatory responses.

Dysfunctional leukocyte-endothelial interactions are thought to play a key role in systemic lupus erythematosus pathogenesis. We questioned the importance of TNF-α and IL-1 for endothelial activation in MRL/lpr lupus-prone mice. Endothelial ICAM-1 and VCAM-1 expression increased significantly with disease evolution in kidney, heart, and brain, as shown by i.v. injected radiolabeled Ab uptake. Lung endothelial VCAM-1 also increased, while lung endothelial ICAM-1 did not rise above a high basal level. Immunoassays showed a significantly raised circulating level of TNF-α by 14 wk, with levels of circulating IL-1α and IL-1β being additionally raised by 20 wk. With 14-wk-old MRL/lpr, anti-TNF-α antiserum inhibited expression of ICAM-1 and VCAM-1 by endothelial cells cultured with sera in vitro, and uptake of anti-ICAM-1 and anti-VCAM-1 mAb in lung, kidney, brain, and heart in vivo. In contrast, both anti-TNF-α and anti-IL-1 antisera were required for maximal inhibition in vitro and in vivo at 20 wk. These data indicate that TNF-α is largely responsible for the early up-regulation of endothelial ICAM-1 and VCAM-1, but that IL-1 enhances expression in late disease. Our observations provide novel insights of possible relevance to understanding endothelial activation in systemic lupus erythematosus, and highlight an approach that can be extended to dissecting other chronic inflammatory diseases.

We have studied vascular endothelial activation and increased expression of ICAM-1 and VCAM-1 at the onset of the elicitation phase of oxazolone contact hypersensitivity in mice. By measuring the local uptake of i.v. administered radiolabeled anti-ICAM-1 and anti-VCAM-1 mAb, we found that endothelial ICAM-1 and VCAM-1 was increased by 4 h after challenge, 2 h later than the first peak of ear swelling and 125I-labeled human serum albumen uptake. Increased expression of endothelial ICAM-1 and VCAM-1 was significantly greater in sensitized animals than in naive animals. Anti-TNF-α antiserum significantly inhibited both the increase in ear thickness (p < 0.01), and the up-regulation of ICAM-1 and VCAM-1 expression (p < 0.01 for both) at 4 h. In contrast, the combination of anti-IL-1α and IL-1β had only a small inhibitory effect on ICAM-1 expression (p < 0.05) and no significant effect on increased ear thickness or on VCAM-1 expression. A mixture of anti-TNF-α, anti-IL-1α, and IL-1β was no more inhibitory for endothelial ICAM-1 and VCAM-1 expression than anti-TNF-α alone. ICAM-1 and VCAM-1 expression at 4 h was unaffected by a combination of mAb against α4 and β2 integrins, whereas expression at 24 h was significantly inhibited (p < 0.05), suggesting that the release of TNF-α and other cytokines involved in the initiation of the response may not require leukocyte traffic or other leukocyte functions involving these integrins. We conclude that the early up-regulation of endothelial ICAM-1 and VCAM-1 during the elicitation of contact hypersensitivity is primarily due to the immune-dependent local release of TNF-α.

In lieu of traditional training of examiners to identify cerebral palsy on a neurologic examination at age 1 year, we proposed an alternative approach using a multimedia training video and CD-ROM we developed after a two-step validation process. We hypothesized that use of CD-ROM interactive training will lead to reliable and valid performance of the neurologic examination by both pediatric neurologists and nonpediatric neurologists. All examiners were asked to take one of six interobserver variability tests found on the CD-ROM on two occasions. In the first interobserver variability evaluation, 89% (531 of 594) of the responses agreed with the gold standard responses. Following annotated feedback to the examiners about the two items that had a 60% correct rate, the correct response rate rose to 93% (114 of 123). In the second interobserver variability evaluation, 88% (493 of 560) of the responses agreed with the gold standard responses. Following annotated feedback to the examiners about the four items that had a 70% correct rate, the correct response rate rose to 96% (104 of 108). Interactive CD-ROM examination training is an efficient and cost-effective means of training both neurologists and non-neurologists to perform structured neurologic examinations in 1-year-old children. It provides an effective means to evaluate interobserver variability, offers a route for feedback, and creates an opportunity to reevaluate variability, both immediately and at periodic intervals. ( J Child Neurol 2005;20:829—831).

Article-at-a-Glance Background In 1993, Intermountain Health Care's three Salt Lake Valley Hospitals formed service lines in four clinical areas, one of which was heart services. After experimenting with various organizational structures, the Salt Lake Valley Hospitals formed a cardiac executive council and three specialty work teams—the clinical process and outcome, satisfaction, and resource teams—to allow for unified planning and greater teamwork. Case study—Open Heart Team The team mapped out the current process and identified areas for potential improvement in the care of patients undergoing coronary artery bypass graft (CABG) surgery. One of the key processes selected for study was extubation. Patients were extubated for an average of 20.41 hours (range, 6 to 120 hours). Analysis of practice patterns demonstrated that extubation was related to staffing patterns, not the patient's readiness. The team created a weaning path, which reduced extubation time to an average of 8.89 hours. Lessons learned A common vision and an organized structure to support integrated services is essential. Cross-training of staff helps ensure that the same standards of care apply across the three campuses. Even when the medical staff and hospital departments each have their own structures for dealing with quality issues, cohesiveness among physicians treating a certain group of patients, such as cardiac patients, can be promoted. In conclusion, a "cardiac culture" that is evident throughout the three hospitals promotes performance improvement.

Chronic inflammation leads to bone loss, and increased fracture rates have been reported in a number of human chronic inflammatory conditions. The study reported here investigates the skeletal effects of dosing a neutralizing antibody to the bone regulatory protein sclerostin in a mouse model of chronic colitis. When dosed prophylactically, an antibody to sclerostin (Scl-AbI) did not reduce the weight loss or histological changes associated with colitis but did prevent inflammation-induced bone loss. At the end of the experiment, Scl-AbI-treated animals had a significantly higher femoral BMD (+27%, p < 0.05) than control antibody (Cntrl-Ab)-treated animals. In a second experiment, treatment with Scl-AbI was delayed until colitis had developed, by which time the mechanical properties of femurs in colitic animals were significantly worse than those of healthy age-matched control mice (maximum load, -26%, p < 0.05; energy, -37%, p < 0.05; ultimate strength, -33%, p < 0.05; elastic modulus, -17%, p < 0.05). A short treatment with Scl-AbI halted bone loss and reversed the decline of both intrinsic and extrinsic mechanical properties of the femur such that, after 19 days of treatment, the bone mechanical properties in the Scl-AbI-treated animals were not significantly different from those of noncolitic age-matched controls. Serum markers of bone formation and resorption suggested that the antibody to sclerostin stimulated osteoblast activity and inhibited osteoclast-mediated bone resorption.

Risk of morbidity and mortality after coronary artery bypass graft surgery (CABG) is higher in patients with clinical diabetes mellitus (DM). We evaluated whether outcomes are affected by postoperative hyperglycemia in CABG patients independent of preoperative DM diagnosis.A total of 2297 consecutive CABG patients were studied. The first glucose value after surgery completion (mean 15 min) was tested as a predictor of outcome. Primary outcome variables were prolonged ventilation (24 h), deep sternal wound infection, renal failure, permanent stroke, any reoperation, length of stay (14 days) and mortality. All outcomes except for prolonged ventilation and length of stay were tracked out to 30 days postoperatively. Patients were stratified by glycemic control: Low (glucose80), normal (referent, glucose 80-110), high (glucose 111-200) and very high (glucose200 mg/dl). Multivariable logistic regression was used to determine the independent predictive value of glycemic groups, adjusted for outcome specific risk scores from the Society of Thoracic Surgeons model.Patient distribution among groups low through very high were 44 (1.9%), 476 (20.7%), 1425 (62.0%) and 352 (15.3%). Greater complication rates were noted in the very high group when compared with the referent group: prolonged ventilation (adjusted odds ratio (OR)=2.66, P.001), length of stay14 days (adjusted OR=2.06, P=.004) and mortality (adjusted OR=7.71, P.001).Patients with blood glucose values200 mg/dl immediately after CABG had an increased risk of complications, including mortality, independent of a clinical diagnosis of DM. This study documents the high risk associated with early postoperative hyperglycemia in this group, suggesting the need for prospective trials of glycemic control.

Intestinal inflammation associated with inflammatory bowel disease (IBD) is typically characterized by an inflammatory cell infiltrate and pro-inflammatory cytokine production. Of particular interest, the frequency of colony stimulating factor-1 (CSF-l)-expressing cells is increased in active lesions. In this study, we have investigated the role of CSF-1 in mucosal inflammation, using a murine model of colitis induced by dextran sulfate sodium (DSS).A neutralizing anti-CSF-1 antibody was administered to Balb/c mice that received DSS in their drinking water. Signs of colitis, such as clinical disease score, cellular infiltrate, and cytokine production, were assessed.Administration of a neutralizing anti-CSF-1 antibody significantly inhibited DSS-induced colitis. Clinical symptoms, such as weight loss and the appearance of diarrhea or fecal blood, were reduced by CSF-1 blockade; histologic scores were also improved. The cellular infiltrate of macrophages and T cells was inhibited and a trend toward reduced production of pro-inflammatory cytokines was noted.This is the first study to demonstrate that CSF-1 plays an important role in mediating intestinal mucosal inflammation and therefore may prove to be an attractive therapeutic target for intestinal diseases such as inflammatory bowel disease.

Colony-stimulating factor-1 (CSF-1) regulates the survival, proliferation and differentiation of macrophages. CSF-1-deficient mice are osteopetrotic due to a lack of osteoclasts, while their tissue macrophage deficiencies and an absence of CSF-1 regulation of CSF-1 receptor-expressing cells in the female reproductive tract contribute to their pleiotropic phenotype. To further understand CSF-1 regulation of macrophages in vivo, we developed a neutralizing anti-mouse CSF-1 antibody which was expressed as a recombinant Fab' fragment and coupled to 40 kDa polyethylene glycol. As developmental regulation by CSF-1 is highest during the early post-natal period, the ability of this anti-CSF-1 reagent to inhibit development was tested by regular subcutaneous injection of mice from post-natal days 0.5-57.5. Antibody treatment decreased growth rate, decreased osteoclast number, induced osteopetrosis, decreased macrophage density in bone marrow, liver, dermis, synovium and kidney and decreased adipocyte size in adipose tissue, thereby inducing phenotypes shared by CSF-1- and CSF-1 receptor-deficient mice. While the antibody blocked macrophage development in some tissues, macrophage densities in other tissues were initially high and were reduced by treatment, proving that the antibody also blocked macrophage maintenance. Since cell surface CSF-1 is sufficient for the maintenance of normal synovial macrophage densities, these studies suggest that anti-CSF-1 Fab'-PEG efficiently neutralizes all three CSF-1 isoforms in vivo, namely the secreted proteoglycan, secreted glycoprotein and cell surface glycoprotein. Since CSF-1 has been shown to enhance chronic disease development in a number of mouse model systems, these studies demonstrate the feasibility of neutralizing CSF-1 effects in these models with an anti-CSF-1 antibody.

Objective. Endothelial activation and dysfunctional leucocyte‐endothelial interactions are thought to play key roles in the pathogenesis of systemic lupus erythematosus (SLE). The object of this study was to investigate directly the effect of increased endothelial adhesion molecule expression on leucocyte‐endothelial cell interactions, using the MRLulpr mouse model. Methods. Leucocyte rolling, arrest and transendothelial migration were quantified in the cremaster muscle microcirculation of 20-week-old MRLulpr mice, using intravital microscopy. Endothelial adhesion molecule expression was quantified using intravenously injected radiolabelled monoclonal antibodies. Results. Basal expression of intercellular adhesion molecule 1 (ICAM-1) by cremaster endothelium was 2-fold greater in MRLulpr than in MRLu++ mice (P

Background The multi-step assembly of the interleukin-6 (IL-6) signaling complex offers the potential for several therapeutic points of intervention in the treatment of rheumatoid arthritis; for example, targeting the cytokine IL-6, the IL-6 receptor (IL-6R; gp80), or gp130. Objectives The objective of this study was to compare affinity-matched anti-murine monoclonal antibodies (mAbs) that target either the IL-6 cytokine or the IL-6 receptor in a range of in vivo assays. Methods Affinity-matched anti-murine reagents 54E07 mAb (anti-IL-6) and 440-1 mAb (anti-IL-6R; gp80) were evaluated for their ability to inhibit either murine IL-6 or CFA-induced serum amyloid A (SAA). IL-6 is also known to have a significant role in B-cell function; therefore, these antibodies (Abs) were evaluated for their ability to inhibit dinitrophenyl (DNP)-specific Ab production. Furthermore, these Abs were dosed to steady state and their capacity to inhibit collagen-induced arthritis (CIA) in DBA-1 mice was evaluated. Results 54E07 mAb produced a ≥90% reduction in the SAA response to both IL-6 and CFA at a dose of 0.1 mg/kg (p Conclusions These murine in vivo studies strongly suggest that targeting IL-6 cytokine rather than the IL-6 receptor is the more efficient therapeutic approach for the treatment of autoimmune diseases such as rheumatoid arthritis. Disclosure of Interest None Declared