Doses Of Ovalbumin Research Articles

PSXI-29 Influence of a Direct Fed Microbial on Growth Performance, Carcass Data, and Immune Function in Feedlot Finishing Steers

Abstract One hundred and eighty crossbred beef steers (405.9 ± 2.3 kg) were used to determine the impact of a direct fed microbial (DFM) on growth performance, carcass characteristics, and immune response in finishing beef cattle. Steers were blocked by body weight and randomly assigned, within block, to 1 of 2 treatments (3 replicates/treatment: 30 steers/replicate). Groups within a replicate were randomly assigned to treatments. Treatments included: 1) no DFM (control) and 2) DFM supplementation at 50 mg/animal/d [BOVAMINE DEFEND Plus containing Lactobacillus animalis, Propionibacterium freudenreichii, Bacillus licheniformis, and Bacillus subtilis]. All steers were fed a high concentrate, steam-flaked corn-based finishing diet. Individual feed intake was recorded daily via the GrowSafe system. Body weights were collected every 28 d. Jugular blood samples were collected from all steers on day 0 and 112. On d 55 of the experiment, 10 steers per pen (30 steers per treatment) were randomly selected and injected, subcutaneously with 4.0 mg of ovalbumin (OVA). Jugular blood samples were collected from each steer on d 0, 7, 14, and 21 post injection. On d 112 of the experiment, the same steers were injected with the same dose of OVA and intramuscularly with 5 mL of a 25 % pig red blood cell (PRBC) solution. Jugular blood samples were collected from each steer on day 0, 7, 14, and 21 post injection. After receiving the finishing diet for 185 d, cattle were transported to a commercial abattoir and slaughtered. Hot carcass weights were collected at the time of slaughter. Carcasses were allowed to chill for 24 h before carcass data were collected. Data were analyzed as a completely randomized block design and individual animal within pen was considered the experimental unit. Initial body weight, final body weight, and overall feed efficiency were similar across treatments. Steers receiving DFM had greater (P < 0.01) body weights and average daily gains on d 28 and 55 and greater (P < 0.04) dry matter intakes over the entire experiment compared with controls. Hot carcass weight, longissimus muscle area, quality grade, and marbling score were similar across treatments. However, dressing percentage (P < 0.02) was greater in steers receiving DFM compared with controls. Antibody titers to OVA and PRBC and d 112 interferon gamma concentrations were similar across treatments. However, red blood cell superoxide dismutase activity was greater (P < 0.05) in DFM-supplemented cattle compared with controls on d 112 of the experiment. These data suggest that BOVAMINE DEFEND Plus supplementation improves growth performance during certain periods of the finishing phase, increases overall dry matter intake and carcass yield, and may impact antioxidant status in beef cattle.

A Food Matrix Triggers a Similar Allergic Immune Response in BALB/c Mice Sensitized with Native, Denatured, and Digested Ovalbumin.

The search for an animal model to evaluate the allergenic potential of processed food products is still ongoing. Both the sensitization to ovalbumin (OVA) in different structural states and the allergic response triggered after intragastric or food challenges were assessed. BALB/c mice were sensitized intraperitoneally to OVA (50 µg) in different structural states (native OVA, N-OVA; denatured OVA, D-OVA; formaldehyde- and lysine-treated OVA, FK-OVA; denatured OVA-FK, OVA-DFK; peptides from pepsin digestion, Pep-OVA). Anti-OVA-specific IgE responses were evaluated using ELISA. Anaphylactic signs and mMCP-1 serum levels were evaluated after intragastric (2.0 mg/OVA) and food (0.41 mg/OVA) challenges. IgE reactivities to N-OVA and D-OVA were similar among groups (p > 0.05). After the challenges, all OVA-sensitized mice developed mild to severe anaphylactic signs (p < 0.05 vs. control). Mice sensitized to N-OVA and D-OVA had the highest mMCP-1 serum levels after challenges (p < 0.05 vs. control). Allergic responses were similar despite the different OVA doses used for the challenges. The N-OVA-sensitized murine model of egg allergy proposed in the present study holds the potential for evaluating the impact of food matrix composition and processing on the threshold of egg-allergic responses.

A Preliminary Study in Immune Response of BALB/c and C57BL/6 Mice with a Locally Allergic Rhinitis Model.

BALB/c and C57BL/6 mouse strains are commonly used in allergy research. The current study investigated the immunological differences between these two mouse strains with a locally allergic rhinitis model. Eighteen BALB/c and eighteen C57BL/6 mice received different doses of ovalbumin (OVA) intranasally for eight weeks (each mouse strain has three subgroups, 25 mg/mL group, 0.25 mg/mL group, and the PBS group). The allergic symptoms, OVA-specific serum antibody (IgE, IgG1, IgG2a), cytokines (IL-4, IFN-γ, IL-10) in the splenic culture supernatant, infiltrating eosinophils and goblet cells in local nasal mucosa were measured. RNA-seq technology was applied to detect differential gene expression in the local nasal mucosa. With the same dose of OVA stimulation, the exacerbation of allergic symptoms was more pronounced in C57BL/6 than in BALB/c. BALB/c serum IgE, IgG1, and IgG2a gradually increased, and C57BL/6 produced fewer serum antibodies IgE and IgG1, while IgG2a never increased. BALB/c spleen cell culture supernatant IL-4 and IL-10 increased with increasing dose, and IFN-γ increased significantly in the intermediate dose group, while IL-4, IL-10, and IFN-γ did not increase in C57BL/6. The infiltration of eosinophils and goblet cells in both mice was proportional to the dose, while C57BL/6 was elevated more than BALB/c. RNA-seq suggested that the innate immune response, immune system process function, Jun kinase (JNK) pathway, and MAPKK pathway were upregulated in C57BL/6 compared to BALB/c. The core genes responsible for the differential immune response in both mice with allergic rhinitis were Kng2, Kng1, Gnb3, Lpar3, Lpar1, Pik3r1, Pf4, Apob, Rps9, and Fbxo2. There are significant differences in the immunologic responses between BALB/c mice and C57BL/6 mice. BALB/c mice developed mild local allergic inflammatory reactions and strong systemic immune responses. In contrast, C57BL/6 mice had stronger local allergic inflammatory responses and relatively mild systemic immune responses. Different mice strains can be selected according to the research purpose.

IMPROVEMENT OF IMMUNOLOGICAL RESEARCH METHODOLOGY IN THE SYSTEM OF NOVEL FOOD SAFETY ASSESSMENT

Immunological studies have an important role in the comprehensive safety assessment of novel food. This publication presents the data of the experiment with different doses of ovalbumin, that confirms the informative value of the study of morphological indicators of the immune system’s organs (thymus, spleen, Payer's patch).

An initial assessment of the involvement of transglutaminase2 in eosinophilic bronchitis using a disease model developed in C57BL/6 mice

The detailed pathogenesis of eosinophilic bronchitis (EB) remains unclear. Transglutaminase 2 (TG2) has been implicated in many respiratory diseases including asthma. Herein, we aim to assess preliminarily the relationship of TG2 with EB in the context of the development of an appropriate EB model through ovalbumin (OVA) sensitization and challenge in the C57BL/6 mouse strain. Our data lead us to propose a 50 μg dose of OVA challenge as appropriate to establish an EB model in C57BL/6 mice, whereas a challenge with a 400 μg dose of OVA significantly induced asthma. Compared to controls, TG2 is up-regulated in the airway epithelium of EB mice and EB patients. When TG2 activity was inhibited by cystamine treatment, there were no effects on airway responsiveness; in contrast, the lung pathology score and eosinophil counts in bronchoalveolar lavage fluid were significantly increased whereas the cough frequency was significantly decreased. The expression levels of interleukin (IL)-4, IL-13, IL-6, mast cell protease7 and the transient receptor potential (TRP) ankyrin 1 (TRPA1), TRP vanilloid 1 (TRPV1) were significantly decreased. These data open the possibility of an involvement of TG2 in mediating the increased cough frequency in EB through the regulation of TRPA1 and TRPV1 expression. The establishment of an EB model in C57BL/6 mice opens the way for a genetic investigation of the involvement of TG2 and other molecules in this disease using KO mice, which are often generated in the C57BL/6 genetic background.

Ovalbumin/lipopolysaccharide induced vasculitis in rats: a new predictive model.

Currently, there are several animal models for vasculitis. Ovalbumin and lipopolysaccharide (OVA, LPS) are well established for causing inflammation and used as an adjunct in the vasculitis induction. However, to date, none has established the effect of OVA and LPS in disease induction. Therefore, in the present study, an attempt has been made to develop a new animal model for vasculitis using OVA/LPS in rats. A total of 42 Wistar rats were divided randomly into seven groups (n=6/group), normal control, and three different doses (0.5, 1, and 5mg/kg) of OVA and LPS treated groups. Half of the rats in each group received only intraperitoneal sensitization, while the remaining half rats were additionally subjected to a one-week intranasal challenge. Results showed that both OVA/LPS in their respective groups have significantly increased circulating inflammatory cells, C-reactive protein (CRP), Inflammatory cytokines (IL-1β, IL-6, TNF-α), Kidney damage markers (BUN, Creatinine), and liver function enzymes (AST, ALT) in a dose-dependent manner. OVA/LPS induced vascular inflammation in a dose-dependent manner. However, the higher (5mg/kg) dose of ovalbumin and lipopolysaccharide has contributed to severe vascular inflammation through increasing inflammatory cytokines. These findings suggest that OVA/LPS may contribute as a possible model for vasculitis in rats.

Different doses of ovalbumin exposure on dendritic cells determine their genetic/epigenetic regulation and T cell differentiation.

It has been reported that allergen dosage can impact the differentiation of dendritic cells (DCs)-mediated T cells. However, the mechanisms of such dose-dependent differentiation are poorly understood. In this study, bone marrow-derived immature DCs stimulated with Ovalbumin (OVA) of different concentrations (0, 10, 100, 1000, 10000μg/ml, respectively). DCs were then co-cultured with naïve T cells. RNA-sequencing detection and DNA methylation of DCs were performed. We show that when DCs were stimulated with low-dose (10μg/ml), 77 genes were up-regulated and 87 genes down-regulated. Most activated genes were related to ribosome synthesis and ion channel inhibition. At the medium-dose (100μg/ml), 339 genes were up-regulated and 168 genes down-regulated. Most activated genes involved cytokine synthesis and regulation of immune responses. At high-dose (10000μg/ml), 2497 genes were up-regulated and 1156 genes down-regulated. TNF signaling pathway, NF-kappa B signaling pathway, antigen processing and presentation signaling pathway were mostly up-regulated. The related co-stimulators, co-inhibitory molecules, inhibitory cytokines, negative regulating enzymes were highly expressed. The monocarbate, coenzyme, fatty acid, glucolipid, starch, sucrose and other metabolism-related signaling pathways were down-regulated. The profiles of DNA methylation and RNA synthesis of DCs varied with different doses of OVA, which serves to induce T cells to differentiate in various directions.

Experimental protocol for development of adjuvant-free murine chronic model of allergic asthma

BackgroundMouse models of allergic asthma play a crucial role in exploring of asthma pathogenesis and testing of novel anti-inflammatory drugs. Widely used acute asthma models usually developed with adjuvant (aluminum hydroxide (alum)) do not reproduce one of the main asthma feature - airway remodeling while chronic asthma model mimic the pathophysiology of human disease. Moreover, the use of alum causes distress in experimental animals and impedes the test of adjuvant-containing drugs. In this study, we aimed to develop a chronic adjuvant-free asthma model with pronounced asthmatic phenotype. MethodsFemale BALB/c mice were divided into 3 groups. The first group was sensitized with intraperitoneal injections of ovalbumin (OVA) emulsified in aluminum hydroxide on days 0, 14, 28 followed by two stages of intranasally challenge with OVA on days 41–43 and 62–64. The second group was subcutaneously sensitized with the same dose of OVA without adjuvant and challenged on the same days. The third group (negative control) included mice which did not received any kind of treatment (i.e. sensitization and challenge). Serum levels of OVA-specific IgE, IgG2a and IgG1 antibodies were detected by ELISA. Airway hyper-responsiveness was measured by non-invasive plethysmography on days 44 and 65. Bronchoalveolar lavage fluids (BALF) sampled in all groups on days 45 and 66 were analyzed by light microscopy. The left lung was removed for histological analysis. The IL-4 and IFNγ mRNA expression in BALF cells was evaluated by RT-PCR. ResultsThe OVA-specific IgE antibody response was two-fold increased in mice from adjuvant-free group compared to the adjuvant group that reflects reorientation of immune response towards Th2 phenotype. At the same time, the level of OVA-specific IgG1 and IgG2a antibodies was increased in the adjuvant group. Airway hyperresponsiveness to methacholine in mice of both experimental groups was two-fold higher than in control. Analysis of cell composition in BAL has shown a significant increase in eosinophil count in both experimental groups that indicate the development of allergic inflammation. Lung histology revealed airway remodeling in both experimental groups including goblet cell hyperplasia/metaplasia, thickening of airway walls, collagen deposition in the wall of distal airways. Additionally, the tendency to develop hypertrophy of bronchial smooth muscle layer was observed. Study of gene expression in BAL cells revealed the increase of IL-4 level in both adjuvant and adjuvant-free groups while IFNγ expression in both experimental groups was similar to control group. ConclusionWe have developed a chronic adjuvant-free mouse asthma model which possesses all necessary features of the disease including airway remodeling and is more suitable for pre-clinical evaluation of novel therapeutic approaches including adjuvant-containing drugs.

Chronic allergic pulmonary inflammation is aggravated in angiotensin-(1-7) Mas receptor knockout mice.

The angiotensin-(1-7) [ANG-(1-7)]/Mas receptor pathway is currently recognized as a counterbalancing mechanism of the renin-angiotensin system in different pathophysiological conditions. We have previously described that treatment with ANG-(1-7) attenuates lung inflammation and remodeling in an experimental model of asthma. In the present study, we investigated whether lack of the Mas receptor could alter the inflammatory response in a model of chronic allergic lung inflammation induced by ovalbumin (OVA). Mas receptor wild-type (MasWT) and knockout (MasKO) mice were subjected to four doses of OVA (20 μg/mice ip) with a 14-day interval. At the 21st day, nebulization with OVA (1%) was started, three times per week until the 46th day. Control groups received saline (0.9% ip) and were nebulized with saline (0.9%). MasWT-OVA developed a modest inflammatory response and minor pulmonary remodeling to OVA challenge. Strikingly, MasKO-OVA presented a significant increase in inflammatory cell infiltrate, increase in extracellular matrix deposition, increase in thickening of the alveolar parenchyma, increase in thickening of the smooth muscle layer of the pulmonary arterioles, increase in proinflammatory cytokine and chemokine levels in the lungs, characteristic of chronic asthma. Additionally, MasKO-OVA presented an increase in ERK1/2 phosphorylation compared with MasWT-OVA. Furthermore, MasKO-OVA showed a worse performance in a test of maximum physical exercise compared with MasWT-OVA. Our study shows that effects triggered by the Mas receptor are important to attenuate the inflammatory and remodeling processes in a model of allergic lung inflammation in mice. Our data indicate that impairment of the ANG-(1-7)/Mas receptor pathway may lead to worsening of the pathophysiological changes of asthma.

Antibody Production, Anaphylactic Signs, and T-Cell Responses Induced by Oral Sensitization With Ovalbumin in BALB/c and C3H/HeOuJ Mice

PurposeTwo mouse strains, BALB/c and C3H/HeOuJ, broadly used in the field of food allergy, were compared for the evaluation of the allergenic potential of ovalbumin (OVA).MethodsSensitization was made by administering 2 different OVA doses (1 and 5 mg), with cholera toxin as Th2-polarizing adjuvant. Antibody levels, severity of anaphylaxis, and Th1 and Th2 responses induced by the allergen were assessed. In addition, because the mice selected had functional toll-like receptor 4, the influence of contamination with lipopolysaccharide (LPS) on the immunostimulating capacity of OVA on spleen cells was also evaluated.ResultsBoth strains exhibited similar susceptibility to OVA sensitization. The 2 protein doses generated similar OVA-specific IgE and IgG1 levels in both strains, whereas C3H/HeOuJ mice produced significantly more IgG2a. Oral challenge provoked more severe manifestations in C3H/HeOuJ mice as indicated by the drop in body temperature and the severity of the anaphylactic scores. Stimulation of splenocytes with OVA led to significantly higher levels of Th2 and Th1 cytokines in BALB/c, and these were less affected by protein contamination with LPS.ConclusionsThe antibody and cytokine levels induced by OVA in BALB/c mice and the observation that BALB/c spleen cell cultures were more resistant than those of C3H/HeOuJ mice to the stimulus of LPS make this strain prone to exhibit Th2-mediated food allergic reactions and very adequate for the study of the features of OVA that make it allergenic.

In-vivo antioxidant and anti-inflammatory activity of rosiglitazone, a peroxisome proliferator-activated receptor-gamma (PPAR-γ) agonists in animal model of bronchial asthma.

Peroxisome proliferator activated receptor-gamma (PPAR-γ) has been shown to play an important role in the control of immunological and inflammatory responses. This study aims at investigating the potential role of rosiglitazone, a strong PPAR-γ agonist in a murine model of bronchial asthma. Adult male guinea pigs were administered ovalbumin 100 mg/kg subcutaneous (SC) and 100 mg/kg intraperitoneal (IP). Treatment with rosiglitazone [5 mg/kg/day, per oral (PO)] was assessed for 21 days. On day 21, the animals were challenged with the same dose of ovalbumin. The forced expiratory volume in 1 s (FEV1 ) to forced vital capacity (FVC), FEV1 /FVC, was measured using a spirometer to diagnosis lung obstruction. Serum levels of interleukin-5 (IL-5) and immunoglobulin E (IgE) were assessed. The activity of superoxide dismutase (SOD) and catalase and the level of reduced glutathione (GSH) were determined in lung tissue homogenates. Our results demonstrated that treatment with rosiglitazone resulted in a statistically significant improvement in lung function and histopathological features. Significant decrease in the serum levels of IL-5 and IgE were observed. The activity of SOD and catalase as well as the GSH level were significantly increased in the lung tissues of treated animals compared with untreated asthmatic animals. Serum IgE concentrations and IL-5 levels were directly correlated to each other and inversely correlated to the SOD, GSH and catalase levels in the all studied guinea pigs. Our results provide evidence that the PPAR-γ agonist rosiglitazone may have potential in the development of therapies for bronchial asthma.

Chronic Allergic Pulmonary Inflammation is Aggravated in Angiotensin‐(1‐7) Mas Receptor Knockout Mice

Angiotensin‐(1‐7)/Mas receptor axis of the renin‐angiotensin system is associated with attenuation of different inflammatory processes. In this study, we investigated the involvement of Mas receptor in the inflammation and airway remodelling in a model of chronic allergic pulmonary inflammation induced by ovalbumin (OVA). FVBN mice groups: Mas KO control (Mas‐/‐CTRL), Mas KO sensitized with OVA (Mas‐/‐OVA), WT control (Mas+/+CTRL) and WT sensitized with OVA (Mas+/+OVA). The allergic groups received 4 doses of OVA (20µg/mice, i.p.) 14 days apart. After the 2nd sensitization, mice were nebulized with OVA (1% for 30 min; 3 times/week for 4 weeks) or saline (CTRL). Mice were sacrificed 72h after the last nebulization. Mas+/+OVA did not develop inflammation or pulmonary remodelling, as it is observed with this protocol in BalbC mice. However, Mas‐/‐OVA showed a worse performance in the maximum load exercise test and presented a significant increase in inflammatory cell infiltrate in the lung, collagen deposition in the airways, increased thickening of the alveolar parenchyma, increased thickening of the smooth muscle layer of the pulmonary arterioles and hypertrophy of the cardiomyocyte of the right ventricle. Furthermore, Mas‐/‐OVA showed an increase in ERK1/2 and SAPK/JNK phosphorylation and an increase in pro‐inflammatory cytokine (IL13) and chemokines (MCP‐1/CCL2 and RANTES/CCL5) in the lungs. These data indicate that genetic deletion of Mas receptor aggravates lung inflammation and remodelling in mice subjected to a model of chronic asthma.

IL-13 and STAT6 signaling involve in low dose lipopolysaccharide induced murine model of asthma

BackgroundWe reported that level of lipopolysaccharide (LPS) exposure determined the type of airway inflammation in a murine model of asthma.ObjectiveThe purpose of this study is to evaluated the role of IL-13 in low dose LPS induced murine model of asthma using IL-13 and signal transducer and activator of transcription 6 (STAT6) deficient mice.MethodsMice were sensitized with an intranasal application of LPS-depleted ovalbumin (OA) and different doses of LPS (0.1 and 10 µg), and then challenged intranasally with OA alone. The phenotype changes between wild type (WT) and IL-13-/- mice and between WT and STAT6-/- mice were evaluated.ResultsWe confirmed again that low and high dose LPS resulted in different phenotypes of murine asthma. In the present study, we observed that phenotypes of murine asthma induced by low dose LPS were abolished in the homozygous null mutation of the IL-13 and STAT6 gene. However, those changes were not shown in mice sensitized OA plus high dose LPS.ConclusionIL-13 plays an important role in low dose LPS induced murine model of asthma. Our results provided a new insight in understanding of the potential role of IL-13 in innate immunity in human allergic asthma.

Ultradeformable archaeosomes as new topical adjuvants

Ultradeformable archaeosomes (UDA) are vesicles made of soybean phosphatidylcholine (SPC), sodium cholate (NaChol) and polar lipids from Halorubrum tebenquichense (3:1:3 wt/wt). Although ultradeformable liposomes (UDL, made of SPC and NaChol at 6:1 wt/wt) and UDA were neither captured nor caused cytotoxicity on keratinocytes, UDA was avidly captured by macrophages, their viability being reduced by 0.4-1.6 mg/mL phospholipids by 25 to 60%. Instead, UDL were poorly captured and caused no toxicity. Balb/C mice immunized by the topical route with four doses of ovalbumin (OVA)-loaded UDA, at 75 μg OVA/600 μg phospholipids (125 nm mean size and -42 mV zeta potential), induced IgG titers tenfold to 100-fold higher than those immunized with OVA-loaded UDL at the same dosage. Both matrices penetrate to the same skin depth (nearly 10 μm after 1 hour on excised human skin), being the higher topical adjuvancy and higher phagocytic uptake of UDA related to its glycolipid content. From the Clinical EditorThis work summarizes key findings related to the development of ultradeformable archaeosomes as vehicles utilized in transdermal delivery systems with improved skin penetration.

Two-and Three-Layered Dissolving Microneedles for Transcutaneous Delivery of Model Vaccine Antigen in Rats

Purpose: Comparison of transcutaneous immunization of ovalbumin (OA) between two-and three-layered dissolving microneedles (MN) in rats. Methods: We prepared 500 μm long two-layered and three-layered dissolving microneedle (2-MN and 3-MN, respectively) arrays from chondroitin sulfate as the base, and OA as the model antigen. The 2-MN containing OA at the acral portion and 3-MN with OA at the second portion were administered to rat skin transcutaneously. As a positive control, OA solution was injected subcutaneously (sc). The OA delivery and diffusion in the rat skin were studied using confocal microscopy with fluorescein-conjugated OA (FL-OA). Results: The formulated positions of OA were 0-155 ± 5 μm for 2-MN and 175 ± 4 – 225 ± 5 μm for 3-MN. The administered doses of OA were 2.2 ± 0.1 μg, 12.0 ± 0.2 μg and 22.0 ± 0.2 μg for 2-MN, 1.8 ± 0.2 μg, 12.6 ± 0.7 μg, and 20.4 ± 0.3 μg for 3-MN, 10 μg, 100 μg and 1000 μg for sc injection. At 4 weeks after the first administration, 3-MN showed about 2.5-7.0 fold and 5.4 fold higher total Ig (G + A + M) antibody than 2-MN and sc injection of the OA solution. Conclusions: The 3-MN, which delivered OA to the epidermis, is a useful drug delivery system for transcutaneous antigen delivery.

Toll-like receptor 4 signalling attenuates experimental allergic conjunctivitis

Allergic conjunctivitis from an allergen-driven T helper type 2 (Th2) response is characterized by conjunctival eosinophilic infiltration. Association between signalling through Toll-like receptor 4 (TLR-4) and adaptive immune responses has been observed in allergic airway disease. We examined whether administration of bacterial lipopolysaccharide (LPS), a prototypic bacterial product that activates immune cells via TLR-4, could affect the development of allergic conjunctivitis and modify the immune response to ovalbumin (OVA) allergen in an experimental allergic conjunctivitis (EAC) model. Mice were challenged with two doses of OVA via conjunctival sac after systemic challenge with OVA in alum. Several indicators for allergy were evaluated in wild-type and TLR-4(-/-) mice with or without adding of different doses of LPS into OVA in alum. Mice challenged with OVA via conjunctival sac following systemic challenge with OVA in alum had severe allergic conjunctivitis. Of interest, LPS administration markedly suppressed immunoglobulin (Ig)E-mediated and eosinophil-dependent conjunctival inflammation. In addition, mice sensitized with OVA plus LPS had less interleukin (IL)-4, IL-5 and eotaxin secretion than mice sensitized with OVA only. The suppression of allergic response by LPS administration was due to Th1 shift. In contrast, the presence of LPS during sensitization with OVA had no effect on severity of allergic conjunctivitis and Th2 responses in TLR4-4(-/-) mice. Our findings demonstrate, for the first time, that LPS suppresses Th2 responses via the TLR-4-dependent pathway in the EAC model.

Antigen-induced arthritis in rabbits: a comparative study between high-resolution ultrasound and contrast-enhanced ultrasound and pathologic findings

The objective of this study was to explore the feasibility and value of high-resolution ultrasound and contrast-enhanced ultrasound in evaluation of synovitis in rabbit knee joint in antigen-induced arthritis (AIA). Thirty-six rabbits were divided into three groups, each injected with different doses of ovalbumin (OVA) into the right knee joint. On week 1 and 4, 6 randomly selected in each group were killed. Each knee joint undergone the high-resolution ultrasound to measure capsule thickness and contrast-enhanced ultrasound to measure synovium thickness. The results from the ultrasound examinations were compared with those of pathologic examinations. Different OVA doses resulted in different modeling success rate, different pathological synovitis score, and different capsule and synovium thickness measured by ultrasound. The diagnostic accuracy of synovitis by ultrasound was high. The ultrasound measurement revealed that the capsule thickness on week 4 was lessened than that on week 1, while the synovial thickness on week 4 was greater than that on week 1. Both the joint capsule and synovium thickness measured by ultrasound were significantly and positively correlated to the pathologic synovitis score (P<0.05). The synovial thickness on week 4 is more correlated to the synovitis score than the capsule thickness, rather than that on week 1. Injection of OVA of different doses results in different modeling success rate and synovitis severity. The high-resolution ultrasound and contrast-enhanced ultrasound can be used to determine whether the AIA was made successfully and evaluate the synovitis severity. In chronic inflammatory phase, the contrast-enhanced ultrasound has better effect.

Establishment of airway eosinophilic bronchitis mouse model without hyperresponsiveness by ovalbumin

Eosinophilic bronchitis (EB) is a useful tool for studying airway hyperresponsiveness (AHR), but an exact EB animal model is lacking. Our objective was to establish an EB mouse model using ovalbumin (OVA). Mice were divided into asthma, normal saline (NS) control and three model groups. Asthma mice were challenged intranasally with 200 μg OVA. Model groups were challenged with one of the three OVA doses (10, 20 or 100 μg), and NS control mice received normal saline. Changes in lung resistance (R(L)), serum OVA-specific immunoglobulin-E (IgE) and differential inflammatory cell counts in bronchoalveolar lavage fluid (BALF) were determined after exposure to increasing doses of methacholine (MCh). Lung histological sections were examined for inflammatory infiltration. R(L) in the 10-μg OVA-challenged model group was not significantly different compared with the NS group at any MCh concentration but was significantly different compared with the asthma group (P < 0.01). R(L) in the other two model groups was intermediate between the asthma and NS groups. Serum OVA-specific IgE and eosinophils in BALF were increased significantly in all model and asthma groups compared with the NS group, but no significant differences were observed between model and asthma groups. Inflammatory cells were seen around bronchioles and capillaries in model and asthma groups but not the NS group. A mouse model of EB without AHR can be established by 10 μg OVA challenge and provides a useful tool for studying AHR mechanisms.

Ligation of CD11c during vaccination promotes germinal centre induction and robust humoral responses without adjuvant

In this study, we investigated the mouse dendritic cell (DC) receptor, complement receptor 4 (CR4; CD11c/CD18), as an immunotarget for triggering humoral immunity. Comparison of antibody titres generated against a panel of 13 anti-antigen-presenting cell receptor monoclonal antibodies, with or without conjugated ovalbumin (OVA), revealed uniquely rapid and robust responses following CR4 targeting, with antibody titres approaching 1 : 100 000 7 days after a single dose of antigen. Furthermore, using just 100 ng OVA conjugated to anti-CD11c Fab', we generated anti-OVA titres greater than those produced by a 100-fold higher dose of OVA in complete Freund's adjuvant at day 28. These anti-OVA antibody titres were sustained and could be boosted further with targeted OVA on day 21. Investigations to explain this vaccine potency showed that, in addition to targeting splenic DC, anti-CDl1c antibodies delivered a powerful adjuvant effect and could boost humoral immunity against OVA even when the OVA was targeted to other molecules on DC, such as major histocompatibility complex class II, CD11a and CD11b. However, interestingly, this adjuvant effect was lost if OVA was targeted to other cells such as B cells via CD21 or CD19. The adjuvant effect was mediated through a marked enhancement of both germinal centre and extrafollicular plasma cell formation in responding spleens. These results demonstrate that anti-CD11c monoclonal antibody can both target antigen and act as a powerful adjuvant for rapid and sustained antibody responses. They also point to an interesting role for CR4 on DC in triggering B cells during humoral immunity.

Contributions of Age and Sex to Heterogeneity of Symptoms and Effectiveness of Secondary Prevention Strategies in Asthma as Modeled in the Guinea Pig

Previous studies in a guinea pig model of asthma have suggested that age and sex contribute both to the profile of asthma symptoms, i.e., asthma heterogeneity, as well as to the success of primary prevention strategies. The present study investigated the contributions of age and sex to the severity of central vs. peripheral airway hyperresponsiveness as well as to the effectiveness of secondary preventions strategies for asthma as modeled in the guinea pig. Experimental groups: Young/Young, sensitized and challenged before sexual maturity; Young/Adult, sensitized young and challenged after sexual maturity; Adult/Adult, sensitized and challenged after sexual maturity. Males and females were sensitized IP with 0.5 mg/kg ovalbumin (OVA) and challenged intratracheally with varying doses of OVA. Cellular infiltration into lung and lavage fluid, OVA specific IgG1 as well as airway hyperresponsiveness to intravenous methacholine were determined 24 hr later. Airway hyperresponsiveness in central airways and peripheral lung was assessed by measurement of airway resistance, tissue damping and tissue elastance. Airway hyperresponsiveness with allergen sensitization and challenge was evident in male and female Adult/Adult animals and male Young/Young animals. Airway hyperresponsiveness in female Young/Young animals was not significant, despite marked airway eosinophilia. Changes in tissue elastance were more evident in OVA treated Adult/Adult compared to Young/Young animals. As allergen exposure decreased, a reduction in inflammation was seen in young females before other age sex groups. OVA induced increases in eosinophils were more pronounced in Young/Adult compared to Adult/Adult animals. Our results suggest that in asthmatic children, females may clinically benefit most from secondary prevention strategies to limit allergen exposure. In adult asthmatics, changes in tissue elastance may be significant, and secondary prevention strategies may be more effective in those sensitized as children compared to those sensitized as adults.