Bioavailability of suppository acetaminophen in healthy and hospitalized ill dogs.

To determine the plasma pharmacokinetics of suppository acetaminophen (APAP) in healthy dogs and clinically ill dogs. This prospective study used six healthy client-owned and 20 clinically ill hospitalized dogs. The healthy dogs were randomized by coin flip to receive APAP orally or as a suppository in crossover study design. Blood samples were collected up to 10 hr after APAP dosing. The hospitalized dogs were administered APAP as a suppository, and blood collected at 2 and 6 hr after dosing. Plasma samples were analyzed by ultra-performance liquid chromatography with triple quadrupole mass spectrometry. In healthy dogs, oral APAP maximal concentration (CMAX =2.69 µg/ml) was reached quickly (TMAX =1.04 hr) and eliminated rapidly (T1/2 = 1.81 hr). Suppository APAP was rapidly, but variably absorbed (CMAX =0.52 µg/ml TMAX =0.67 hr) and eliminated (T1/2  = 3.21 hr). The relative (to oral) fraction of the suppository dose absorbed was 30% (range <1%-67%). In hospitalized ill dogs, the suppository APAP mean plasma concentration at 2 hr and 6 hr was 1.317 µg/ml and 0.283 µg/ml. Nonlinear mixed-effects modeling did not identify significant covariates affecting variability and was similar to noncompartmental results. Results supported that oral and suppository acetaminophen in healthy and clinical dogs did not reach or sustain concentrations associated with efficacy. Further studies performed on different doses are needed.

Early weaning is beneficial to prevent atopic dermatitis occurrence in young children.

BACKGROUND: Epidemiological data on infant feeding practices and allergic diseases are controversial. The purpose of this study was to explore the association of early weaning with the occurrence of atopic dermatitis (AD). METHODS: We conducted a matched case-control study on incident physician-diagnosed AD in early childhood including 451 cases and 451 controls. Data on several factors, including feeding practices, were collected through an interviewer-administered questionnaire. Odds ratios (OR) and the corresponding 95% confidence intervals (CIs) were estimated through logistic regression models, conditioned on study center, age, sex, and period of interview, and adjusted for potential confounders. RESULTS: Early weaning, defined as the introduction of solid foods at 4 or 5 months of age, was inversely related to the risk of AD, with children weaned at 4 months having lower AD risk (OR = 0.41, 95% CI, 0.20-0.87) compared to those exclusively breastfed. Similar results were observed for weaning started at 5 months of age (OR = 0.39, 95% CI, 0.18-0.83). This association persisted when children with and without family history of allergy were considered separately. Prolonged partial breastfeeding (breastmilk plus milk formulas) was not associated with AD. Consistently, the introduction of a high number of different solid foods reduced the risk of AD (P trend = 0.02 at 4 months of age and P trend = 0.04 at 5 months). CONCLUSION: Our data provide evidence against the preventing role of prolonged exclusive (but not partial) breastfeeding in AD occurrence and confirm recent results indicating a beneficial role of early weaning in AD.

Evaluation of Iron Deficiency Using Reticulocyte Indices in Dogs Enrolled in a Blood Donor Program.

BACKGROUND: People donating blood more than twice annually are at risk of developing iron deficiency. Little is known about the iron status of dogs enrolled in blood donor programs. HYPOTHESIS: Dogs donating blood ≥6 times annually will show evidence of iron deficiency based on their reticulocyte indices. ANIMALS: Thirteen dogs enrolled in a blood donor program donating ≥6 times over the preceding 12 months and 20 healthy nondonor control dogs. METHODS: Prospective observational study. Mature red blood cell (RBC) indices, reticulocyte indices, serum iron, serum ferritin, and total iron-binding capacity (TIBC) were compared between groups. RESULTS: Packed cell volume (median 47%, range 40-52%, P < .01), hematocrit (median 46.4%, range 40.3-52.5%, P < .01), and reticulocyte count (median 16,000/µL, range 9,000-38,000/µL, P < .01) were significantly lower in the blood donor dogs. No statistically significant differences were noted in the mature RBC indices between groups. Both reticulocyte mean corpuscular volume (median 88.8 fL, range 83.4-95.5 fL, P = .03) and reticulocyte hemoglobin content (median 24.6 pg, range 23.1-26.6 pg, P < .01) were significantly lower in the blood donor group. Serum iron and ferritin were similar between groups; however, TIBC was significantly higher in the control group (median 403 µg/dL, range 225-493 µg/dL, P = .02). CONCLUSIONS: The findings in dogs donating ≥6 times annually suggest the presence of iron-deficient erythropoiesis in this population.

Casein-specific IL-4- and IL-13-secreting T cells: a tool to implement diagnosis of cow's milk allergy.

BACKGROUND: Cow's milk allergy (CMA) is a frequent food allergy in young children. The oral food challenge is the gold standard for diagnosis, and there is currently no reliable biological test. Our aim was to evaluate the diagnostic potential of a functional assay quantifying allergen-specific Th2 cells in CMA children. METHODS: A total of 29 children aged 2.8-10.5 years underwent a double-blind, placebo-controlled food challenge (DBPCFC) to cow's milk. Blood was collected before performing the DBPCFC, and peripheral mononuclear cells were cultured in an 18-h ELISpot assay with casein, α-lactalbumin, or ß-lactoglobulin. Numbers of antigen-specific IL-4- and IL-13-secreting lymphocytes and serum-specific IgE, IgG4, and total IgE levels were assessed. Receiver operating characteristic (ROC) curves were generated. RESULTS: A total of 17 (59%) children reacted to cow's milk and were therefore considered as allergic to cow's milk (CMA). The mean number of casein-specific IL-4- and IL-13-secreting T cells was higher in CMA than in non-CMA children (P = 0.009, 0.004, respectively). Moreover, it was inversely correlated with the cumulative dose of cow's milk tolerated (P = 0.003, 0.0009, respectively). ROC curve of combined IL-4 and IL-13 analysis showed an area under the curve of 0.98 (95% CI 0.90-1.06). For a cutoff of 10 IL-4- and 12 IL-13-secreting T cells, sensitivity and negative predictive value were 100%. CONCLUSIONS: Enumeration of casein-specific IL-4- and IL-13-secreting T cells appears a promising tool to improve diagnosis and, if confirmed in larger studies, could permit less frequent use of the oral food challenge.

Quantification of circulating house dust mite-specific IL-4- and IL-13-secreting T cells correlates with rhinitis severity in asthmatic children and varies with the seasons.

BACKGROUND: Defining suitable markers to diagnose and monitor allergy and its severity is essential to correctly assign patients for specific immunotherapy. Circulating levels of specific IgE are good markers of sensitization, but not of clinically symptomatic allergy. OBJECTIVE: To quantify circulating interleukin (IL)-4- and IL-13-secreting T cells specific for house dust mite (HDM) in children presenting HDM-allergic asthma associated or not with rhinitis and correlate results with clinical symptoms. METHODS: We analysed 26 children with HDM respiratory disease (allergic rhinitis and asthma) together with six children with non-allergic asthma. Peripheral blood mononuclear cells were stimulated with HDM extract in a 24-h ELISpot assay to quantify the number of HDM-specific IL-4- and IL-13-secreting T cells. Asthma severity and control, and rhinitis severity were scored according to the Global Initiative for Asthma (GINA) and the Allergic Rhinitis and its Impact on Asthma (ARIA) Guidelines. RESULTS: The number of HDM-specific IL-4- and IL-13-secreting T cells was higher in patients with allergic asthma as compared to patients with non-allergic asthma. It varied with the season of blood sampling with two peaks in the fall and early spring. Independently of the season, the number of HDM-specific IL-4-secreting T cells correlated with rhinitis severity (OR = 2; 95% IC:1.1-3.8; P = 0.04). CONCLUSIONS AND CLINICAL RELEVANCE: Allergen-specific IL-4- and IL-13-producing T cells were only detected in HDM-allergic asthmatic children (not in patients with non-allergic asthma). Their numbers correlated with clinical severity of allergic rhinitis.

Induction of allograft tolerance by monoclonal CD3 antibodies: a matter of timing.

Despite remarkable progress in organ transplantation through the development of a wealth of immunosuppressive drugs highly effective at controlling acute rejection, two major problems still remain, the loss of transplants due to chronic rejection and the growing number of sensitized recipients due to previous transplants, transfusions or pregnancies. Induction of immune tolerance appears to be the only way to curb this complex situation. Here we describe that a therapy, already successfully used to restore immune tolerance to self-antigens in overt autoimmunity, is effective at promoting transplant tolerance. We demonstrate that a short low-dose course with CD3 antibodies started after transplantation, at the time of effector T cell priming to alloantigens, induces permanent acceptance of fully mismatched islet allografts. Mechanistic studies revealed that antigen-specific regulatory and effector T cells are differentially affected by the treatment. CD3 antibody treatment preferentially induces apoptosis of activated alloreactive T cells which is mandatory for tolerance induction. In contrast, regulatory T cells are relatively spared from CD3 antibody-induced depletion and can transfer antigen-specific tolerance thus arguing for their prominent role in sustaining long-term graft survival.

The 'hygiene hypothesis' for autoimmune and allergic diseases: an update.

According to the 'hygiene hypothesis', the decreasing incidence of infections in western countries and more recently in developing countries is at the origin of the increasing incidence of both autoimmune and allergic diseases. The hygiene hypothesis is based upon epidemiological data, particularly migration studies, showing that subjects migrating from a low-incidence to a high-incidence country acquire the immune disorders with a high incidence at the first generation. However, these data and others showing a correlation between high disease incidence and high socio-economic level do not prove a causal link between infections and immune disorders. Proof of principle of the hygiene hypothesis is brought by animal models and to a lesser degree by intervention trials in humans. Underlying mechanisms are multiple and complex. They include decreased consumption of homeostatic factors and immunoregulation, involving various regulatory T cell subsets and Toll-like receptor stimulation. These mechanisms could originate, to some extent, from changes in microbiota caused by changes in lifestyle, particularly in inflammatory bowel diseases. Taken together, these data open new therapeutic perspectives in the prevention of autoimmune and allergic diseases.

99th Dahlem conference on infection, inflammation and chronic inflammatory disorders: immune therapies of type 1 diabetes: new opportunities based on the hygiene hypothesis.

Insulin-dependent (type 1) diabetes is a prototypic organ-specific autoimmune disease resulting from the selective destruction of insulin-secreting beta cells within pancreatic islets of Langerhans by an immune-mediated inflammation involving autoreactive CD4(+) and CD8(+) T lymphocytes which infiltrate pancreatic islets. Current treatment is substitutive, i.e. chronic use of exogenous insulin which, in spite of significant advances, is still associated with major constraints (multiple daily injections, risks of hypoglycaemia) and lack of effectiveness over the long term in preventing severe degenerative complications. Finding a cure for autoimmune diabetes by establishing effective immune-based therapies is a real medical health challenge, as the disease incidence increases steadily in industrialized countries. As the disease affects mainly children and young adults, any candidate immune therapy must therefore be safe and avoid a sustained depression of immune responses with the attendant problems of recurrent infection and drug toxicity. Thus, inducing or restoring immune tolerance to target autoantigens, controlling the pathogenic response while preserving the host reactivity to exogenous/unrelated antigens, appears to be the ideal approach. Our objective is to review the major progress accomplished over the last 20 years towards that aim. In addition, we would like to present another interesting possibility to access new preventive strategies based on the 'hygiene hypothesis', which proposes a causal link between the increasing incidence of autoimmune diseases, including diabetes, and the decrease of the infectious burden. The underlying rationale is to identify microbial-derived compounds mediating the protective activity of infections which could be developed therapeutically.

[Increase of CXCL10 serum level in systemic sclerosis interstitial pneumonia]. / Augmentation de CXCL10 dans le sérum au cours de la pneumopathie interstitielle de la sclérodermie systémique.

INTRODUCTION: CXCL10, a gamma-interferon-induced chemokine seems to play a relevant role in lung involvement that occurs in systemic sclerosis (SSc). The objective of this study was to assess the serum level of CXCL10 in interstitial lung disease (ILD) associated with SSc. METHODS: Serum level of CXCL10 was assayed in 23 healthy volunteers (60.0 years; 58.0-67.3) and 29 SSc patients (63.1 years; 60.1-69.4) by ELISA method. Pulmonary function tests (PFTs), lung CT-scan and echocardiogram were also performed in the patients. Serum levels from patients and healthy controls were compared and a comparison among SSc patients between those with and without ILD, as documented by lung CT-scan, was also performed. RESULTS: Median CXCL10 level from patients with SSc was significantly higher than that from healthy volunteers (110.0 pg/ml; 60.8-223.8 versus 52.0; 41.3-65.8; p<0.001). Fifteen out of the 29 patients had ILD on lung CT-scan; the median CXCL10 level from SSc patients with ILD was significantly higher than that from SSc patients without ILD (210.0 pg/ml; 115.0-307.5 versus 76.0; 55.0-110.0; p=0.02). CONCLUSION: Our findings suggest that CXCL10 is specifically increased in the lung involvement of SSc and plays a role in scleroderma lung disease.

[Therapeutic monoclonal antibodics]. / Les anticorps monoclonaux thérapeutiques.

Less than ten years after the discovery of hybridomas by Kohler and Milstein in 1975, the first monoclonal antibody was injected in humans to prevent graft rejection. The antibodies used were targeted against the CD3 molecule involved in the transduction of T cell signals after recognition of the antigen. Paradoxically, because of pharmaceutical hesitations later found to be unwarranted, the therapeutic class developed slowly. It was not until the 1990s that other formulations emerged. In parallel, work was undertaken to modify the antibodies using genetic engineering techniques to humanize the molecules by changing the majority of the species determinants from the murine producing animals to the human recipients. It was even found possible to produce totally human antibodies using several different strategies based on in vitro phage display. This led to rapid development of new antibodies used for a wide range of indications: organ transplantation (anti-CD2, anti-CD4, antiCD25, antiCD40L), as well as anti-respirator syncytial virus (RSV) for respiratory infections, and anti cancer agents (Anti-CD20 antibodies and anti-tumor antibodies), as well as autoimmune diseases (with significant success with anti-TNF antibodies). It was found the monoclonal antibodies offer an exceptionally effective method in domains where conventional compounds have reached their limit. This specific action of monoclonal antibodies, with fine specificity for the targets (which can be varied to infinity), as well as their capacity to provide positive signals to cells whose functions are not often inhibited by small molecules. An illustration of this mode of action is the effect recently demonstrated by our team on restored self tolerance using anti-CD3 antibodies in newly diagnosed diabetics in whom sustained remission can be injected after less than one week's treatment.

Exacerbated Th2-mediated airway inflammation and hyperresponsiveness in autoimmune diabetes-prone NOD mice: a critical role for CD1d-dependent NKT cells.

The NOD mouse has proved to be a relevant model of insulin-dependent diabetes mellitus, closely resembling the human disease. However, it is unknown whether this strain presents a general biastoward Th1-mediated autoimmunity or remains capable of mounting complete Th2-mediated responses. Here, we show that NOD mice have the capacity to develop a typical Th2-mediated disease, namely experimental allergic asthma. In contrast to what might have been expected, they even developed a stronger Th2-mediated pulmonary inflammatory response than BALB/c mice, a strain that shows a typical Th2 bias in this model. Thus, after allergen sensitization and intra-nasal challenge, the typical features of experimental asthma were exacerbated in NOD mice, including enhanced bronchopulmonary responsiveness, mucus production and eosinophilic inflammation in the lungs as well as specific IgE titers in serum. These hallmarks of allergic asthma were associated with increased IL-4, IL-5, IL-13 and eotaxin production in the lungs, as compared with BALB/c mice. Notwithstanding their quantitative and functional defect in NOD mice, CD1d-dependent NKT cells contribute to aggravate the disease, since in OVA-immunized CD1d(-/-) NOD mice, which are deficient in this particular T cell subset, airway eosinophilia was clearly diminished relative to NOD littermates. This is the first evidence that autoimmune diabetes-prone NOD mice can also give rise to enhanced Th2-mediated responses and might thus provide a useful model for the study of common genetic and cellular components, including NKT cells that contribute to both asthma and type 1 diabetes.

[Prevent and cure insulin-dependent diabetes]. / Prévenir et guérir le diabète insulino-dépendant.

Insulin-dependent mellitus diabetes is an autoimmune disease resulting from the destruction of the beta cells of the islets of Langerhans by autoreactive T cells. Various approaches are being investigated in order to slow down the progression of diabetes using immunotherapy. The most promising results are based on the induction of specific tolerance following the administration of soluble autoantigens or monoclonal anti-CD3 antibodies. These strategies, initially developed in the NOD mouse, have been proven to be efficacious in recently diagnosed diabetics.

A thymulin analogue peptide with powerful inhibitory effects on pain of neurogenic origin.

The effects of a synthetic peptide analog of thymulin (PAT) were tested on nociceptive behavior in two animal models for peripheral mononeuropathy and in another two models for capsaicin-induced hyperalgesia. Treatment with PAT (0.25-25 microg/rat, i.p.) produced significant reduction of the mechanical allodynia and heat hyperalgesia in rats subjected to either chronic constriction injury (CCI) or spared nerve injury (SNI) models for mononeuropathy. Cold allodynia was moderately reduced in the CCI model. The inhibition of neuropathic manifestations peaked at 1-2 h post-treatment and disappeared in 3-4 h. Daily treatment with PAT, however, produced progressive attenuation of all neuropathic manifestations in the SNI model. On the other hand, pretreatment with similar doses of PAT produced dose-dependent reduction of the hyperalgesia induced by intraplantar injection of capsaicin (10 microg in 50 microl). The highest dose of PAT (50 microg) produced significant reduction of abdominal aversive behavior induced by i.p injection of capsaicin (20 microg in 100 microl). Compared with the effects of treatment with morphine or meloxicam (injected at single doses known to produce analgesia), PAT exerted equal or stronger inhibitory effects on neuropathic manifestations. The reported results suggest a possible direct action of PAT on afferent nerve fibers but its mechanisms remain to be determined.

[Current concepts of autoimmunity]. / Les concepts actuels de l'auto-immunité

Autoimmunity is physiological: in every normal individual autoreactive T cells and B cells which produce natural autoantibodies, exist. Auto-immunity becomes pathological, giving rise to an autoimmune disease, when the number of autoreactive cells, and particularly the avidity of their receptors for autoantigens increase. Triggering of the disease depends both on the increase in immunogenicity of the target cell, which may be secondary to a viral infection, and the individual's own capability to recognize the antoantigens (HLA genes, T cell repertoire). More rarely, the disease is caused by an infectious agent leading to a crossed reaction with an autoantigen (Guillain-Barré syndrome). Nevertheless, all these elements are not sufficient to provoke a chronic disease such as multiple sclerosis or myasthenia gravis. The passage to chronicity is usually secondary to a defect in immunoregulation. Several categories of regulatory T cells have been found: Th2 cells, CD25+ cells, Trl cells, NKT cells. It is still difficult to asses the responsibility of the defect of one of these populations in a given disease, or to single out the cytokines implicated, although an essential role is often given to interleukin 10 and/or TGFB. Even if the pathogenic autoimmune reaction is triggered by the autoantigens of the target cell, there is apparently not a unique autoantigen target. The specificity of the reaction spreads progressively from one antigen, which may vary among subjects, to the entire target cell. It is based on these notions that new immunotherapeutic approaches for autoimmune diseases are being developed (soluble autoantigens, or one of their modified peptides: (APL), cytokine, anti-CD3 antibody).

Activation of natural killer T cells by alpha-galactosylceramide treatment prevents the onset and recurrence of autoimmune Type 1 diabetes.

Type 1 diabetes (T1D) in non-obese diabetic (NOD) mice may be favored by immune dysregulation leading to the hyporesponsiveness of regulatory T cells and activation of effector T-helper type 1 (Th1) cells. The immunoregulatory activity of natural killer T (NKT) cells is well documented, and both interleukin (IL)-4 and IL-10 secreted by NKT cells have important roles in mediating this activity. NKT cells are less frequent and display deficient IL-4 responses in both NOD mice and individuals at risk for T1D (ref. 8), and this deficiency may lead to T1D (refs. 1,6-9). Thus, given that NKT cells respond to the alpha-galactosylceramide (alpha-GalCer) glycolipid in a CD1d-restricted manner by secretion of Th2 cytokines, we reasoned that activation of NKT cells by alpha-GalCer might prevent the onset and/or recurrence of T1D. Here we show that alpha-GalCer treatment, even when initiated after the onset of insulitis, protects female NOD mice from T1D and prolongs the survival of pancreatic islets transplanted into newly diabetic NOD mice. In addition, when administered after the onset of insulitis, alpha-GalCer and IL-7 displayed synergistic effects, possibly via the ability of IL-7 to render NKT cells fully responsive to alpha-GalCer. Protection from T1D by alpha-GalCer was associated with the suppression of both T- and B-cell autoimmunity to islet beta cells and with a polarized Th2-like response in spleen and pancreas of these mice. These findings raise the possibility that alpha-GalCer treatment might be used therapeutically to prevent the onset and recurrence of human T1D.