CpG oligodeoxynucleotide synergizes innate defense regulator peptide for enhancing the systemic and mucosal immune responses to pseudorabies attenuated virus vaccine in piglets in vivo

Oligonucleotides containing CpG motifs (CpG ODN) are strong adjuvants for humoral and cellular immune responses in mice, and innate defense-regulator peptides (IDRs) are known to facilitate the uptake of antigens into antigen presenting cells (APCs), but data on synergistic effects of CpG and IDRs in piglets are scarce. In this report, the combination of porcine-specific CpG ODN and HH2 (a kind of IDR which was selected for its better synergy with CpG ODN) was used as immunoadjuvant to enhance the immune responses of the newborn piglets to Pseudorabies attenuated virus (PRV) vaccine. The titers of specific antibodies and serum IgG1/IgG2 subtypes to PRV vaccine, interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), IL-12 and IL-4 were examined to identify the immune responses of the newborn piglets. The results showed that piglets immunized intranasally (IN) and subcutaneously (SC) with PRV vaccine and CpG-HH2 complex both presented high titers of PRV-specific antibodies and IgG2 isotype, a Th1-dominated (IFN-γ and IL-12) cytokine profiles, high levels of IgA in saliva, broncheoalveolar lavage (BAL) and intestinal washings. The results suggested that, CpG-HH2 complex augmented systemic (IgG in serum) and mucosal (IgA in saliva, BAL and intestinal washings) immune responses against antigen. CpG-HH2 complex stimulated both T-helper type1 (Th1) (IgG2) and Th2 (IgA) responses when delivered IN, and IN route could induce stronger mucosal immune responses than SC route. All these data indicate that CpG-HH2 complex is a potential effective adjuvant for the PRV vaccine in newborn piglets.     

Effects of bisphenol A on antigen-specific antibody production,proliferative responses of lymphoid cells,and TH1 and TH2 immune responses in mice

1. We investigated the effect of bisphenol A (BPA), which binds estrogen receptors, on immune responses including production of antigen-specific antibodies, proliferative responses of lymphoid cells, and Th1 and Th2 responses. 2. For this investigation, mice were p.o. given varying doses including 3, 30, 300, and 3000 micro g kg(-1) of BPA immediately after immunization with hen egg lysozyme (HEL) (day 0) and then daily by day 20. On day 21, anti-HEL IgG antibodies in sera and proliferative responses of spleen cells to the antigen were measured. Anti-HEL IgG2a antibodies and IFN-gamma secreted from splenic lymphocytes were also measured as indicators of Th1 immune responses, while anti-HEL IgG1 antibodies and IL-4, as those of Th2 responses. 3. The results showed that treatment with 3000 micro g kg(-1) of BPA was followed by a significant increase in anti-HEL IgG as well as the antigen-specific cell proliferation. Anti-HEL IgG2a production and IFN-gamma secretion were significantly enhanced in mice treated with 300 and 30 micro g kg(-1) of BPA, respectively, while anti-HEL IgG1 production and IL-4 secretion were augmented in animals given 3000 and 300 micro g kg(-1) of the chemical, respectively. 4. Augmentation of these immune responses was also observed in mice exposed to 0.3-30 micro g kg(-1) of estradiol, although Th1 responses appeared to be more sensitive to the sex hormone than Th2 responses. 5. These results suggest that BPA may play a role in augmenting immune responses, especially Th1 responses.     

Intranasal administration of CpG oligonucleotides induces mucosal and systemic Type 1 immune responses and adjuvant activity to porcine reproductive and respiratory syndrome killed virus vaccine in piglets in vivo

Oligonucleotides containing CpG motifs (CpG ODN) are strong adjuvants for immune responses, particularly in mice. Recently, it has been showed that CpG ODN is a promising mucosal adjuvant in mice, but data on mucosal immune responses induced by CpG ODN in piglets are scarce. We have previously demonstrated that CpG ODN is a potent adjuvant to pseudorabies attenuated virus (PRV) vaccine when administered subcutaneously (SC) in newborn piglets. Herein, we evaluated intranasal (IN) delivery of CpG ODN with porcine reproductive and respiratory syndrome (PRRS) killed virus vaccine (PRRSV) to determine its potential as a mucosal adjuvant to a commercial vaccine. CpG ODN augmented systemic (IgG in serum, Peripheral blood mononuclear cells (PBMC) proliferation) and mucosal (IgA in feces, nasal and oral secretions) immune responses against antigen. CpG ODN stimulated both T-helper type1 (Type 1) (IgG2) and Type 2 (IgA) responses when delivered intranasally. Results from this study indicate that stimulatory CpG ODN may be effective as a mucosal adjuvant with commercial vaccine in husbandry animals.     

Immune response after oral administration of the encapsulated malaria synthetic peptide SPf66

The synthetic peptide SPf66 adsorbed on alum is one of the few Plasmodium falciparum vaccines which have been tested in field trials. We previously reported that subcutaneous administration of SPf66 loaded PLGA microparticles (MP) enhances the antibody response to this antigen compared to the conventional alum formulation. We now evaluate the suitability of polymeric formulations to obtain systemic immune responses by gastric intubation of Balb/c mice. Formulations composed of 1:1 mixtures of PLGA 50:50 and 75:25 (lactic:glycolic) microparticles were administered by the oral route, and when animals were boosted 3 weeks later significant systemic IgG antibody responses were elicited, comparable to alum triple shot and superior to the aqueous vaccine given by the oral route. The finding of IgG2a isotype for PLGA-vaccinated mice compared to the absent levels of this isotype for the alum-vaccinated group could be interpreted as a sign of Th1-like immune response and cellular immune response activation. Our results confirm that using the appropriate schedule the oral administration of PLGA particles is suitable to obtain systemic immune responses to the carried antigen.     

Systemic administration of Bordetella pertussis enhances pulmonary sensitization to house dust mite in juvenile rats.

The incidence of allergies and asthma has increased significantly in the past few decades. The objectives of this study were to establish an allergy model in weanling rats to more closely reflect the developing immune system of children, and to determine whether systemic administration of inactivated Bordetella pertussis could enhance pulmonary and systemic immune responses to locally administered house dust mite antigen (HDM). Three-week old female Brown Norway rats were sensitized with 10 micro g HDM intratracheally or intraperitoneally, with or without a simultaneous injection of 10(8) whole killed B. pertussis organisms. Ten days later, all the rats were challenged with 5 micro g HDM via the trachea. Bronchial lymph nodes and bronchoalveolar lavage fluid (BAL) were collected 0, 2, and 4 days post-challenge. Coadministration of pertussis and intratracheal instillation of HDM enhanced HDM-specific lymphoproliferative responses and increased BAL levels of total protein, lactate dehydrogenase, HDM-specific IgE and IgG antibodies, and the number of eosinophils in BAL to the same extent as had occurred in the systemically immunized animals. The data show that intratracheal instillation of HDM induces a mild allergic sensitization in juvenile rats, and that ip injection with B. pertussis enhances this sensitization process to levels seen in animals injected with antigen and B pertussis together. These results suggest that simultaneous exposure to Th2-inducing vaccine components and allergenic proteins may be a risk factor for allergic sensitization and the development of asthma in susceptible individuals.     

Immune response to Saccharomyces cerevisiae mannan conjugate in mice

Mannan, the surface polysaccharide antigen of Saccharomyces cerevisiae was partially oxidized and conjugated to a protein carrier. Prepared conjugate was immunogenic in mice and re-injection elicited significant increase of anti-S. cerevisiae specific IgG and IgM serum antibodies. There was somewhat less increase in IgM serum antibodies. Serum distribution of IgG subclasses, especially IgG(2(a+b)):IgG(1) ratio throughout the immunization demonstrated effective Th1 predominance of immune response. Newly synthesized S. cerevisiae mannan conjugate could be considered as a perspective vaccine candidate for preventive immunomodulation treatment.     

Suppression of Th1 and Th2 immune responses in mice by Sinomenine, an alkaloid extracted from the chinese medicinal plant Sinomenium acutum

The present study was designed to investigate the effect of sinomenine (SIN), an alkaloid extracted from Sinomenium acutum, on Th1 and Th2 immune responses in mice. For this investigation, mice were S. C. immunized with ovalbumin (OVA) emulsified with complete Freund's adjuvant (day 0). Varying doses of SIN were orally administered daily over a period of 21 days, commencing on day 0. On day 21, anti-OVA IgG and proliferative responses of spleen cells to the antigen were measured. Anti-OVA IgG2a and IFN-gamma were measured as indicators of Th1 immune responses and anti-OVA IgG1, IgE, and IL-5 as those of Th2 responses. TGF-beta was measured as an indicator of Th3 immune responses. The results showed that treatment with SIN was followed by decreases in anti-OVA IgG and the antigen-specific splenocyte proliferation. Production of all isotypes of antibodies including anti-OVA IgG2a, IgG1 and IgE as well as secretion of cytokines such as IFN-gamma and IL-5 was suppressed by SIN, although the suppression of anti-OVA IgG2a and IFN-gamma by the alkaloid appeared to be greater than that of anti-OVA IgG1, IgE, and IL-5. In addition, SIN enhanced the secretion of TGF-beta. These results suggest that SIN appears to have suppressive effects on both Th1 and Th2 immune responses. The results also suggest that Th1 responses may be more preferentially suppressed by the Sinomenium acutum-derived alkaloid compared to Th2 responses. TGF-beta may at least in part contribute to the suppression of Th1 as well as Th2 immune responses.     

Effects of diesel exhaust particle extracts on Th1 and Th2 immune responses in mice

The present study was undertaken to investigate the effects of extracts of diesel exhaust particles (DEP) on Th1 and Th2 immune responses. In order to separate compounds from DEP different in hydrophobicity, a single DEP sample was consecutively extracted with hexane (HEX-DEP), benzene (BEN-DEP), dichloromethane (DIC-DEP), methanol (MET-DEP), and 1M ammonia (AMM-DEP). The last unextracted residue (UNE-DEP) was also used to test its effect on immune responses. To immunize mice, hen egg lysozyme (HEL) was injected i.p. (day 0). Varying doses of DEP, each DEP extract, and UNE-DEP were intranasally administered every 2 days from days 0 to 18. Anti-HEL IgG2a antibodies in sera and IFN-γ secreted from spleen cells were measured as an indicator of Th1 immune responses, while anti-HEL IgG1 antibodies and IL-4 as that of Th2 responses. The results showed that treatment with DEP and DIC-DEP increased both Th1 and Th2 responses to HEL. UNE-DEP facilitated Th1 but not Th2 responses, while MET- and AMM-DEP administration was followed by enhancement of Th2 but not Th1 responses. Neither HEX- nor BEN-DEP modulated Th1 as well as Th2 responses. These results suggest that DEP contain various compounds different in hydrophobicity which may affect both Th1 and Th2, Th1 but not Th2, and Th2 but not Th1 immune responses.     

Diesel exhaust, carbon black, and silica particles display distinct Th1/Th2 modulating activity

Certain particulate air pollutants may play an important role in the increasing prevalence of respiratory allergy by stimulating T helper 2 cell (Th2)-mediated immune responses to common antigens. The study described here examined different particles, diesel exhaust particles (DEP), carbon black particles (CBP), and silica particles (SIP) for their immunomodulating capacity in both primary and secondary immune responses in female BALB/C mice. The primary response was studied after subcutaneous injection of 1 mg of particle together with 10 microgram of reporter antigen TNP-OVA (2,4,6-trinitrophenyl coupled to ovalbumin) into the hind paw. Interferon-gamma (IFN-gamma) and interleukin 4 (IL-4) production was assessed in the popliteal lymph node (PLN) at Day 2 and Day 5 after injection by flow cytometry and ELISA. The number of IL-4-containing CD4(+) T cells increased between Day 2 and Day 5 in DEP- and CBP-exposed mice, in contrast to SIP-treated animals. IL-4 production by cultured PLN cells was also significantly increased for DEP- and CBP-treated animals. The secondary response was studied in different organs after an intranasal challenge with TNP-OVA (50 microgram), which was given 4 weeks after the initial subcutaneous injection. Five days after challenge the number of antibody-forming cells (AFCs) was assessed in peribronchial lymph nodes (PBLN), spleen, bone marrow, and PLN, and antibody levels were determined in weekly obtained blood samples. It appeared that all particles acted as adjuvant, but the different particles stimulated distinct types of immune responses to TNP-OVA. DEP-treated animals show high IgG1 and IgE levels in serum and high IgG1 and IgE-forming AFC numbers in PBLN, bone marrow, and spleen. CBP-treated animals show even higher IgG1 and IgE levels and AFC numbers, and in addition display IgG2a production. SIP-injected animals display predominantly IgG2a responses. It is concluded that DEP are able to skew the immune response toward the T helper 2 (Th2) side, whereas SIP stimulate a Th1 response and CBP have a mixed activity, stimulating both Th1 and Th2 responses in this model.     

The effect of endotoxin on the production of IgE,IgG1 and IgG2a antibodies against the cat allergen Fel d 1 in mice

BACKGROUND: Endotoxin/LPS is ubiquitous in our environment. The question whether lipopolysaccharide (LPS) is beneficial or disease-promoting in relation to asthma and allergy has been raised in several recent studies. Some have reported a positive correlation between the level of LPS in house dust and the symptoms of asthmatic children. Others have found that exposure to LPS appears to protect against the development of atopic disease in children. OBJECTIVES: We performed a study in mice to examine the antibody response after subcutaneous immunization with LPS and the cat allergen Fel d 1. We asked whether LPS would increase the response and direct the antibody production towards an allergic (IgE), or non-allergic (IgG2a) antibody profile. In rodents both IgE and IgG1 are antibodies produced under Th2-dependence and IgG2a antibodies under Th1-dependence. Also, when LPS and Fel d 1 are introduced to the immune system, we asked whether the timing of the two agents relative to each other is crucial. METHODS: The mice were injected subcutaneously with LPS and/or Fel d 1 four times in various orders. IgE, IgG1 and IgG2a antibodies specific to Fel d 1 were measured in serum using ELISA. RESULTS: A strong antibody response, both for IgE, IgG1 and IgG2a, was observed only when Fel d 1 and LPS were injected simultaneously, and in particular after repeated injections. CONCLUSION: A strong specific antibody response was observed, both for IgE, IgG1 and IgG2a, only when LPS was introduced to the immune system together with the cat allergen Fel d 1. No such adjuvant effect was observed when LPS was introduced alone prior to or subsequent to the allergen. The resulting antibody response was not polarized in terms of Th1- or Th2-dependence.     

Exploring the immunopotentiation of Chinese yam polysaccharide poly(lactic-co-glycolic acid) nanoparticles in an ovalbumin vaccine formulation in vivo

Biocompatible and biodegradable poly(lactic-co-glycolic acid) (PLGA) has been approved by the US Food and Drug Administration and has frequently been used to develop potential vaccine delivery systems. The immunoregulation and immunopotentiation of Chinese yam polysaccharide (CYP) have been widely demonstrated. In the current study, cell uptake mechanisms in dendritic cells (DCs) were monitored in vitro using confocal laser scanning microscopy, transmission electron microscopy, and flow cytometry. To study a CYP-PLGA nanoparticle-adjuvanted delivery system, CYP and ovalbumin (OVA) were encapsulated in PLGA nanoparticles (CYPPs) to act as a vaccine, and the formulation was tested in immunized mice. The CYPPs more easily underwent uptake by DCs in vitro, and CYPP/OVA could stimulate more effective antigen-specific immune responses than any of the single-component formulations in vivo. Mice immunized using CYPP/OVA exhibited more secretion of OVA-specific IgG antibodies, better proliferation, and higher cytokine secretion by splenocytes and significant activation of CD3⁺CD4⁺ and CD3⁺CD8⁺ T cells. Overall, the CYPP/OVA formulation produced a stronger humoral and cellular immune response and a mixed Th1/Th2 immune response with a greater Th1 bias in comparison with the other formulations. In conclusion, the data demonstrate that the CYPP-adjuvanted delivery system has the potential to strengthen immune responses and lay the foundation for novel adjuvant design.     

CpG oligodeoxynucleotide promotes protective immunity in the enteric mucosa and suppresses enterotoxigenic E. coli in the weaning piglets

CpG oligodeoxynucleotide (CpG ODN) has been described as an effective activator of the innate immune system, with potential to protect against infection caused by a range of pathogens in a non-specific manner. We therefore investigated if intranasal (IN), oral (OR)-mucosal, and intramuscular (IM)-systemic administrations of CpG ODN without antigen codelivery could all enhance innate immunity in the enteric mucosa and control the extent of enterotoxigenic Escherichia coli (ETEC) infection in weaning piglets. Here our data showed that CpG ODN dosed by IN, OR or IM routes protected weaning piglets against a subsequent challenge with ETEC. The level of protection was greater when CpG ODN was administered IN and OR than IM, demonstrating a clear relationship between the route of CpG dosing and protection. IN and OR treatments with CpG ODN reduced bacterial load in the phases at days 3–5 post challenge. The CXC chemokine (CXCL10 and CXCL11) and CC chemokine (CCL4 and CCL5) mRNA expressions were elevated in the intestinal tissues from animals treated IN or OR with CpG ODN compared to untreated controls. Significantly enhanced mRNA expressions for cathelicidins (PR-39 and protegrin-1), but moderately for β-defensin (pBD1 and pBD2), were observed in IN or OR CpG-treatments. Also, significant production of cytokines (IL-12, IFN-γ, and MCP-1) and F4-specific antibodies (IgG/IgA) was detected in intestinal washings following IN and OR CpG-treatments. In contrast, IM delivery induced marked production of sera F4-specific antibodies. It was possible that these chemokines, cytokines, cathelicidins and antibodies played a role in the clearance of ETEC. These findings suggested that IN or OR administration of CpG ODN without antigen codelivery might represent a valuable strategy for induction of innate immunity against ETEC infection.     

Altered immunomodulating and toxicological properties of degraded Quillaja saponaria Molina saponins

Quillaja saponins are readily hydrolyzed under physiological conditions, yielding deacylated forms that are significantly less toxic than their precursors. Yet, deacylated saponins are unable to stimulate a strong primary immune response. Although deacylated saponins elicit a strong total IgG response, their capacity to stimulate a Thl type IgG isotype profile (i.e. high levels of IgG2a and IgG2b) has been significantly diminished. Instead, an IgG profile closer to that of a Th2 immune response is stimulated (i.e. high IgG1 levels). Deacylated saponins have also lost their capacity to elicit an effective T cell immunity, as shown by their stimulation of a marginal lymphoproliferative response and their inability to elicit the production of cytotoxic lymphocytes (CTL). Modification of the immune-modulating properties brought by the degradation of quillaja saponins during vaccine storage may change the intended immune response from a Th1 to a Th2 type. This alteration would have negligible effects on vaccines depending on Th2 immunity mediated by neutralizing antibodies. However, the performance of vaccines directed against intracellular pathogens as well as therapeutic cancer vaccines may be seriously affected by the loss of their capacity to stimulate both a Th1 immune response and the production of CTL.     

Lactobacillus casei Shirota does not decrease the food allergic response to peanut extract in Brown Norway rats

Probiotics are claimed to beneficially affect the immune system and their involvement in allergy prevention is being investigated extensively. However, the efficacy of probiotics in allergy prevention remains controversial. We investigated whether the probiotic Lactobacillus casei Shirota (LcS) could modulate the food allergic response against peanut extract (PE) in Brown Norway (BN) rats. For this purpose BN rats were sensitized to PE (0, 1 and 10 mg/(rat d)) by daily oral gavage and the LcS-groups were additionally orally dosed with 1 x 10(9) colony forming units LcS/(rat d). LcS administration had minor effects in animals that were not sensitized. LcS increased Th1-(PE-specific IgG1), whereas the Th1/Th2 ratio based on PE-specific IgG1/PE-specific IgG2a shifted towards Th2 dominance in rats sensitized to PE in the presence of LcS as compared to rats that were sensitized to PE only. LcS stimulated PE-specific IgG2a; but for PE-specific IgE the effect was less clear; whereas there was no overall effect, two rats did not show detectable specific IgE antibodies, whereas the remainder showed significantly increased levels. LcS also resulted in increased numbers of basophilic granulocytes in blood. Furthermore, LcS increased levels of both Th1-(IFN-gamma) and Th2-(IL-4) related cytokines in PE stimulated spleen and mesenteric lymph node (MLN) cells, but predominantly IL-4 levels in the supernatants of both spleens and MLNs. Our study does not support the hypothesis that LcS down-regulates food allergic responses in a BN rat model for food allergy to peanut.     

4-[1-(4-Fluorobenzyl)-4-hydroxy-1H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid as a prototype to develop dual inhibitors of HIV-1 integration process

In recent years several potent HIV-1 integrase (IN) inhibitors have been identified and after the successful clinical use of raltegravir, they have gained a definitive place in the treatment of HIV-1 infection. Yet, there is a continuous effort to design newer inhibitors that target different steps in the integration process. Furthermore, the increased understanding of IN structural biology has opened novel approaches to inhibit IN, such as targeting its multimerization or interaction with cellular cofactors. On these bases, we have concentrated our research on the identification of small molecules able to inhibit two different stages of the integration process: the IN strand-transfer phase and the IN–LEDGF/p75 interaction. We found that the 4-[1-(4-fluorobenzyl)-4-hydroxy-1H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid (CHI-1043) is an interesting anti-HIV agent exhibiting dual inhibitory effects. This work has suggested the possibility of also constructing an integration dual inhibitor using a design-in strategy.     

福尔马林所致炎性痛后脑内三种COX亚型的变化及不同选择性COX抑制剂的镇痛效应比较

目的:比较炎性痛后三种环氧合酶(cyclooxygenase,COX)亚型的表达变化,以及选择性COX抑制剂不同应用方式对炎性痛的镇痛效应。方法:小鼠足底注射福尔马林诱导炎性痛。用放射免疫分析及RTPCR分别评估脑COX1、COX2及COX3在福尔马林注射前、注射后1、12h、1、3、7、14、30、60d的变化。在镇痛效应的比较中,动物被分成5组:对照组、SC组、NS组、IN组及NS SC组。前4组分别灌胃生理盐水、SC560、NS398和indomethacin。NS SC组在前一个月接受NS398,后一个月接受SC560。测定各组动物在福尔马林注射前、注射后1、12h、1、3、7、14、30、60d的热痛阈。结果:COX2的表达在炎性痛后12h到3d升高显著,而COX1的表达在2周到2月升高显著。在整个观察时限内COX3的表达无明显变化。与其他组相比,NS SC组动物的热痛阈在整个炎性痛过程中均明显提高。结论:炎性痛后早期COX2升高而晚期COX1升高。COX3变化不明显。COX1抑制剂和COX2抑制剂的结合使用比单纯使用其中一种能取得更好的镇痛效果。     

Intranasal administration of dry powder anthrax vaccine provides protection against lethal aerosol spore challenge

The use of an aerosolizable form of anthrax as a biological weapon is considered to be among the most serious bioterror threats. Intranasal (IN) delivery of a dry powder anthrax vaccine could provide an effective and non-invasive administration alternative to traditional intramuscular (IM) or subcutaneous (SC) injection. We evaluated a dry powder vaccine based on the recombinant Protective Antigen (rPA) of Bacillus anthracis for vaccination against anthrax via IN immunization in a rabbit model. rPA powders were formulated and administered IN using a prototype powder delivery device. We compared serum IgG and toxin neutralizing antibody (TNA) titers of rabbits immunized IN with 10 microg rPA of a powder formulation with those immunized with the same dose of liquid rPA vaccine, delivered either IN or by IM injection. In addition, each group was tested for survival after aerosol spore challenge. Our results showed that IN vaccination with rPA powders elicited serum PA-specific IgG and TNA titers that were equivalent to those raised by liquid rPA administered IN. Serum PA-specific IgG and TNA titers after IN delivery were lower than for IM injection, however, after aerosol spore challenge, rabbits immunized IN with powders displayed 100% protection versus 63% for the group immunized IN with the liquid vaccine and 86% for the group immunized by IM injection. The results suggest that an IN powder vaccine based on rPA is at least as protective as a liquid delivered by IM injection.     

In Vivo Fluorescence Imaging of IgG1 Aggregates After Subcutaneous and Intravenous Injection in Mice

Purpose

To monitor the biodistribution of IgG1 aggregates upon subcutaneous (SC) and intravenous (IV) administration in mice and measure their propensity to stimulate an early immune response.

Methods

A human mAb (IgG1) was fluorescently labeled, aggregated by agitation stress and injected in SKH1 mice through SC and IV routes. The biodistribution of monomeric and aggregated formulations was monitored over 47 days by fluorescence imaging and the early immune response was measured by quantifying the level of relevant cytokines in serum using a Bio-plex assay.

Results

The aggregates remained at the SC injection site for a longer time than monomers but after entry into the systemic circulation disappeared faster than monomers. Upon IV administration, both monomers and aggregates spread rapidly throughout the circulation, and a strong accumulation in the liver was observed for both species. Subsequent removal from the circulation was faster for aggregates than monomers. No accumulation in lymph nodes was observed after SC or IV administration. Administration of monomers and aggregates induced similar cytokine levels, but SC injection resulted in higher cytokine levels than IV administration.

Conclusion

These results show differences in biodistribution and residence time between IgG1 aggregates and monomers. The long residence time of aggregates at the SC injection site, in conjunction with elevated cytokine levels, may contribute to an enhanced immunogenicity risk of SC injected aggregates compared to that of monomers.     

Effect of diesel exhaust particle extracts on induction of oral tolerance in mice.

We examined the effect of diesel exhaust particle (DEP) extracts on oral tolerance in mice. For this examination, a single DEP sample was consecutively extracted with hexane (HEX-DEP), benzene (BEN-DEP), dichloromethane (DIC-DEP), methanol (MET-DEP), and 1 M ammonia (AMM-DEP). Residues unextracted (UNE-DEP) with the last extraction solvent 1 M ammonia were also used to test their ability to induce oral tolerance. To immunize mice, hen egg lysozyme (HEL) emulsified with an equal volume of CFA was injected sc (day 0). Oral tolerance was induced by feeding 10 mg HEL on days -5, -4, -3, -2, and -1. DEP, each DEP extract, and UNE-DEP were intranasally administered immediately after each feeding of HEL. The results showed that oral administration of HEL markedly suppressed production of anti-HEL IgG antibodies as well as proliferative responses of spleen cells to HEL. The suppression of anti-HEL IgG antibody production and the cell proliferation by the oral antigen was significantly blocked by DEP, DIC-, AMM-, and UNE-DEP. Neither HEX-, BEN-, nor MET-DEP modulated the orally induced suppression of these immune responses. When the levels of anti-HEL IgG2a antibodies and IFN-gamma (Th1 responses) and anti-HEL IgG1 antibodies and IL-4 (Th2 responses) were determined, DEP and DIC-DEP diminished the suppression of both Th1 and Th2 responses observed following oral administration of HEL. In contrast, UNE- and AMM-DEP prevented the reduction of Th1 but not Th2, and Th2 but not Th1 oral tolerance, respectively. Thus, UNE-DEP appears to contain compounds that block induction of Th1 but not Th2 oral tolerance, whereas AMM-DEP have compounds that abrogate induction of Th2 but not Th1 oral tolerance. DIC-DEP, as well as DEP, appear to contain components that block induction of both Th1 and Th2 oral tolerance. As oral tolerance is thought to play a critical role in preventing Th1 as well as Th2 food allergy, the blockade of oral tolerance by these DEP extracts suggests that DEP may contain compounds different in hydrophobicity associated with the cause of such adverse immunologic responses to food proteins.     

Th1/Th2-Balancing immunomodulating activity of gel-forming (1-->3)-beta-glucans from fungi.

The immunomodulating effects of various gel-forming (1-->3)-beta-glucans, grifolan (GRN), SSG, sonifilan (SPG) and alkaline-treated SPG (SPG-OH), on balancing helper T cell activity were examined in a murine model. Plasma from mice that were injected with GRN or SPG-OH and trinitrophenyl ovalbumin (TNP-OVA) contained TNP-specific antibodies of both IgG1 and IgG2a isotypes. Administration of SSG and TNP-OVA significantly augmented the synthesis of IgG2a antibodies, while the synthesis of IgG1 was reduced. However, SPG did not enhance the antibody response. In the culture supernatants of splenocytes obtained from GRN- or SPG-OH-administered mice, high levels of IgGI and low levels of IgG2a and IFN gamma were detected. In contrast, high levels of IgG2a and IFN gamma and low levels of IgGI were detected in the case of administration of SSG. Furthermore, it was shown by intracellular cytokine staining that the proportion of IFN gamma+CD4+ double-positive cells among the CD4+ cells from mice administered SSG was most strongly increased by addition of PMA and A23187. On the other hand, the expression of IL-12 p40 mRNA was more markedly elevated in splenocytes after combined administration of TNP-OVA plus SSG than after administration of TNP-OVA alone. The highest IFN gamma production was observed when adherent cells of mice administered TNP-OVA and SSG were cultured with TNP-primed lymphocytes. This effect of administration of SSG on IFN-y production was completely inhibited by addition of anti-IL-12 mAb. In conclusion, our study showed that beta-glucans have various effects on the Th1 or Th2-dependent antibody subclasses, in particular, SSG induces the development of Th1 cells via the IL-12 pathway.