[Evaluation of the Acute Toxicity and Genotoxicity of the Rice Biofortified with ß-Glucan].

OBJECTIVE: To provide a scientific evaluation of the food safety of the rice biofortified with ß-glucan. METHODS: The acute toxicity and genotoxicity of the rice were evaluated by 14-day feeding experiment, Ames experiment, erythrocyte micronucleus test and mouse lymphoma thymidine kinase gene ( TK) mutation assay respectively. RESULTS: In the acute toxicity test, there was no obvious toxicity of rice biofortified with ß-glucan, and no abnormality was found in anatomical observation. The median lethal dose (LD 50) to rats and mice wereall greater than 15 mg/kg, which belonged to the actual non-toxic level. Whether with S 9 activation or not, no genotoxicity was found to the tested strains TA97a, TA98, TA100, TA102 and TA1535. No induction of polychromatic erythrocytes and inhibition of bone marrow were found in erythrocyte micronucleus test. The results of TK gene mutation assay did not show the mutagenicity of ß-glucan bioaugmentation rice. All results of the three genotoxicity tests were negative. CONCLUSION: Under the current experimental conditions, ß-glucan biofortified rice showed no obvious acute toxicity and genotoxicity.

Study on chemopreventive effects of raw and roasted ß-glucan-rich waxy winter barley using an in vitro human colon digestion model.

Due to their unique dietary fibre composition, in particular ß-glucan, the consumption of barley may contribute to a healthy diet and the prevention of colon cancer. The aim of the present study was to analyse chemopreventive effects of barley flakes obtained from a ß-glucan-rich barley cultivar. In order to address the impact of heat treatment on potential chemopreventive effects, barley flakes were roasted (160 °C-180 °C, approx. 20 min). The flakes were subjected to in vitro digestion and fermentation. Fermentation supernatants (FS) were analysed for the concentrations of short-chain fatty acids (SCFA) and ammonia. Chemopreventive endpoints (growth inhibition, apoptosis, DNA integrity, gene expression of detoxifying enzymes) were analysed in LT97 colon adenoma cells. Concentrations of SCFA were increased in barley FS (2.5-fold, on average) with a shift of molar ratios towards butyrate production, while ammonia levels were significantly decreased (0.7-fold, on average) compared to the fermentation control. The growth of LT97 cells was significantly reduced by barley FS in a time- and dose-dependent manner, and caspase-3 activity of treated cells was significantly enhanced (up to 6.3-fold, on average). On average, treatment of cells resulted in increased mRNA levels of CAT (2.1-fold), SOD2 (2.2-fold) and GSTP1 (3.9-fold), while expression of GPx1 (0.3-fold) was significantly decreased in some cases. The roasting process did not cause genotoxic effects of barley FS and had no impact on chemopreventive properties. Our results indicate chemopreventive potential of in vitro fermented waxy winter barley, mediated primarily by growth inhibitory and apoptotic effects, which is largely unaffected by roasting.

Dual delivery of tuberculosis drugs via cyclodextrin conjugated curdlan nanoparticles to infected macrophages.

In tuberculosis, macrophages serve as a host for Mycobacterium tuberculosis and hence targeting them with nanoparticles-based drug delivery could be the best strategy to achieve high therapeutic efficacy. Two tuberculosis drugs, namely rifampicin and levofloxacin, which have different mechanism of action on the bacteria, were complexed with cyclodextrin and conjugated to curdlan nanoparticles, to achieve simultaneous sustained release of both the drugs over a prolonged period of time. They are non-cytotoxic to both RAW 264.7 and L929 cells. They are taken up ˜1.8 times more by the macrophage cells through dectin-1 receptor than the fibroblast cells. They are also able to kill more than 95% of Mycobacterium smegmatis residing within the macrophages in 4 h. These results demonstrate that curdlan-CD nanoparticles can be a promising system for the loading and intracellular release of hydrophobic drugs into macrophages for various therapeutic applications.

Protection against Gut Inflammation and Sepsis in Mice by the Autodigested Product of the Lingzhi Medicinal Mushroom, Ganoderma lingzhi (Agaricomycetes).

Ganoderma lingzhi (reishi) (GL) is a widely used medicinal mushroom in the treatment of several diseases, including metabolic syndrome and cancer. We recently performed autodigestion of GL and found enhanced release of hypotensive peptides and immunomodulating beta-1,3-glucan. In the present study, we examined the protective effects of G. lingzhi and its autodigested product (AD-GL) against gut inflammation and endogenous sepsis induced in mice by the oral administration of indomethacin (IND). Gut inflammation was assessed by measuring the lengths of the intestines and colon, and sepsis was evaluated by the survival period. G. lingzhi and AD-GL were mixed with animal feed (2.5%) that was available ad libitum during the experimental period. The murine model was established by the repeated oral administration of IND (once a day, 5 mg/kg from day 0). On day 3, the lengths of the small intestine and colon were measured, and the average lengths of the intestines were significantly shorter in the control and G. lingzhi-administered groups than in the AD-GL-administered group. This finding suggests that AD-GL protected against gut inflammation due to IND-induced ulceration and subsequent microbial translocation. Furthermore, the median numbers of survival days in the control group, the G. lingzhi group, and the AD-GL group were 5, 6, and 11, respectively. The concentrations of the inflammatory cytokines, tumor necrosis factor (TNF)-α and interleukin-6, in the blood were significantly reduced in the mice administered AD-GL. In the in vitro cell culture, G. lingzhi and AD-GL fractions released a significantly higher concentration of TNF-α from the spleen, and the splenocytes of mice administered AD-GL hot water extract showed a greater potential to produce cytokines in response to pathogen-associated molecular patterns. These results strongly suggest the protection of the gut mucosa from inflammation, and therefore the prevention of sepsis, by the administration of AD-GL. Autodigestion appears to be a promising protocol that enhances the usefulness of G. lingzhi as a functional food.

Chitosan:ß-glucan particles as a new adjuvant for the hepatitis B antigen.

The development of new vaccine adjuvants is urgently needed not only to enable new routes of vaccine administration but mostly to go beyond protective humoral immunity, often insufficient to fight infectious diseases. The association of two or more immunopotentiators or mimicking pathogen physicochemical properties are strategies that can favor powerful and more balanced Th1/Th2 immune responses. Therefore, the present work aimed to combine both chitosan and ß-glucan biopolymers in the same particle, preferably with surface ß-glucan localization to simulate the cell wall of some pathogens and to stimulate the immune cells expressing the Dectin-1 receptor. Chitosan:ß-glucan particles (ChiGluPs) were developed through a chitosan precipitation method. The chitosan was precipitated into a ß-glucan alkaline solution followed by genipin crosslink. The optimized method produced particles with a mean diameter of 837 nm for ChiPs and 1274 nm for ChiGluPs. ß-glucan surface location was confirmed by zeta potential measurements (+24 mV for ChiGluPs and +36 mV for ChiPs) and zeta potential titration. These new particles showed high antigen loading efficacy and low cytotoxicity. Mice vaccination studies revealed that both ChiPs and ChiGluPs had an adjuvant effect for the hepatitis B surface antigen (HBsAg), with ChiGluPs resulting in serum anti-HBsAg total IgG 16-fold higher than ChiPs, when administered with 1.5 µg HBsAg per dose. Specifically, IgG1 subclass was 5-fold higher and IgG3 subclass was 4-fold higher for ChiGluPs comparing to ChiPs. Overall, the preparation method developed allowed the advantageous combination of ß-glucan with chitosan, without chemical functionalization, which represents an additional step toward tailor-made adjuvants production using simple precipitation techniques.

A water soluble ß-glucan of an edible mushroom Termitomyces heimii: Structural and biological investigation.

A water soluble ß-glucan (PS-I) with an average molecular weight ∼ 1.48 × 10(5)Da was isolated from the alkaline extract of an edible mushroom Termitomyces heimii. PS-I contained (1 → 3)-, (1 → 6)-, (1 → 3, 6)-linked and terminal ß-d-glucopyranosyl moieties in a ratio of nearly 2:1:1:1. Based on the total hydrolysis, methylation analysis, periodate oxidation, Smith degradation, partial hydrolysis and 1D/2D NMR experiments the structure of the PS-I was elucidated. On the basis of these experiments, the repeating unit of the polysaccharide was found to consist of a backbone chain of two (1 → 6)-ß-D-glucopyranosyl residues, one of which was branched at O-3 position with the side chain consisting of two (1 → 3)-ß-D-glucopyranosyl and a terminal ß-D-glucopyranosyl residue. Cytotoxic effect of PS-I on human blood lymphocytes at varied concentrations was studied. Moreover, it also exhibited potent antioxidant activities by diminishing the ROS and NO in the nicotine stimulated lymphocytes up to 200 µg/ml.

Cell Type-Specific Delivery of RNAi by Ligand-Functionalized Curdlan Nanoparticles: Balancing the Receptor Mediation and the Charge Motivation.

Tissue-specific delivery of therapeutic RNAi has great potential for clinical applications. Receptor-mediated endocytosis plays a crucial role in targeted delivery of biotherapeutics including short interfering RNA (siRNA). Previously we reported a novel Curdlan-based nanoparticle for intracellular delivery of siRNA. Here we designed a nanoparticle based on ligand-functionalized Curdlan. Disaccharides were site-specifically conjugated to 6-deoxy-6-amino Curdlan, and the cell line specificity, cellular uptake, cytotoxicity, and siRNA delivery efficiency of the corresponding disaccharide-modified 6-deoxy-6-amino-Curdlan were investigated. Observation by fluorescence microscopy as well as flow cytometry showed that galactose-containing Curdlan derivatives delivered fluorescently labeled short nucleic acid to HepG2 cells expressing ASGPR receptor but not in other cells lacking surface ASGPR protein. Moreover, highly galactose-substituted Curdlan derivatives delivered siRNA specifically to ASGPR-expressing cells and induced RNAi activities, silencing endogenous GAPDH gene expression. Our data demonstrated that galactose-functionalized 6-deoxy-6-amino-Curdlan is a promising carrier for short therapeutic nucleic acids for clinical applications.

Evidence for Proinflammatory ß-1,6 Glucans in the Pneumocystis carinii Cell Wall.

Inflammation is a major cause of respiratory impairment during Pneumocystis pneumonia. Studies support a significant role for cell wall ß-glucans in stimulating inflammatory responses. Fungal ß-glucans are comprised of d-glucose homopolymers containing ß-1,3-linked glucose backbones with ß-1,6-linked glucose side chains. Prior studies in Pneumocystis carinii have characterized ß-1,3 glucan components of the organism. However, recent investigations in other organisms support important roles for ß-1,6 glucans, predominantly in mediating host cellular activation. Accordingly, we sought to characterize ß-1,6 glucans in the cell wall of Pneumocystis and to establish their activity in lung cell inflammation. Immune staining revealed specific ß-1,6 localization in P. carinii cyst walls. Homology-based cloning facilitated characterization of a functional P. carinii kre6 (Pckre6) ß-1,6 glucan synthase in Pneumocystis that, when expressed in kre6-deficient Saccharomyces cerevisiae, restored cell wall stability. Recently synthesized ß-1,6 glucan synthase inhibitors decreased the ability of isolated P. carinii preparations to generate ß-1,6 carbohydrate. In addition, isolated ß-1,6 glucan fractions from Pneumocystis elicited vigorous tumor necrosis factor alpha (TNF-α) responses from macrophages. These inflammatory responses were significantly dampened by inhibition of host cell plasma membrane microdomain function. Together, these studies indicate that ß-1,6 glucans are present in the P. carinii cell wall and contribute to lung cell inflammatory activation during infection.

Induction of Dectin-1 and asthma-associated signal transduction pathways in RAW 264.7 cells by a triple-helical (1, 3)-ß-D glucan, curdlan.

People living in damp buildings are typically exposed to spore and mycelial fragments of the fungi that grow on damp building materials. There is experimental evidence that this exposure to triple-helical (1, 3)-ß-D glucan and low molecular weight toxins may be associated with non-atopic asthma observed in damp and moldy buildings. However, the mechanisms underlying this response are only partially resolved. Using the pure (1, 3)-ß-D glucan, curdlan, and the murine macrophage cell line, RAW 264.7, there were two objectives of this study. The first was to determine whether signal transduction pathways activating asthma-associated cell signaling pathways were stimulated using mouse transduction Pathway Finder(®) arrays and quantitative real-time (QRT) PCR. The second objective was to evaluate the dose and temporal responses associated with transcriptional changes in asthma-associated cytokines, the signal transduction receptor gene Dectin-1, and various transcription factor genes related to the induction of asthma using customized RT-PCR-based arrays. Compared to controls, the 10(-7) M curdlan treatment induced significant changes in gene transcription predominately in the NFkB, TGF-ß, p53, JAK/STAT, P13/AKT, phospholipase C, and stress signaling pathways. The 10(-8) M curdlan treatment mainly induced NFkB and TGF-ß pathways. Compared to controls, curdlan exposures also induced significant dose- and time-dependent changes in the gene translations. We found that that curdlan as a non-allergenic potentiator modulates a network of transduction signaling pathways not only associated with TH-1, TH-2, and TH-3 cell responses including asthma potentiation, but a variety of other cell responses in RAW 264.7 cells. These results help provide mechanistic basis for some of the phenotypic changes associated with asthma that have been observed in in vitro, in vivo, and human studies and open up a hypothesis-building process that could explain the rise of non-atopic asthma associated with fungi.

Depletion of CD4+CD25+Foxp3+ regulatory T cells with anti-CD25 antibody may exacerbate the 1,3-ß-glucan-induced lung inflammatory response in mice.

1,3-ß-Glucan was a major cell wall component of fungus. The existing studies showed that 1,3-ß-glucan exposure could induce lung inflammation that involved both Th1 and Th2 cytokines. Regulatory T cells (Treg cells) played a critical role in regulating immune homeostasis by adjusting the Th1/Th2 balance. The role of Treg cells and regulatory mechanism in 1,3-ß-glucan-induced lung inflammation is still unclear. In our study, mice were exposed to 1,3-ß-glucan by intratracheal instillation. To investigate the role of Treg cells in response to 1,3-ß-glucan, we generated Treg-depleted mice by intraperitoneal administration of anti-CD25 mAb. The Treg-depleted mice showed more inflammatory cells and severer pathological inflammatory change in lung tissue. Depletion of Treg cells led to increased Th1 cytokines and decreased Th2 cytokines. Treg-depleted mice showed a decreased expression of anti-inflammation cytokine and lower-level expression of CTLA-4. In all, our study indicated that Treg cells participated in regulating the 1,3-ß-glucan-induced lung inflammation. Depletion of Treg cells aggravated the 1,3-ß-glucan-induced lung inflammation, regulated the Th1/Th2 balance by enhancing Th1 response. Treg cells exerted their modulation function depending on both direct and indirect mechanism during the 1,3-ß-glucan-induced lung inflammation.

Induction of GSNO reductase but not NOS in the lungs of mice exposed to glucan-spiked dust.

Both in vivo and in vitro studies have suggested that airborne organic dusts may induce inflammatory responses in the lungs, characterized by typical patterns of cytokine up-regulation and secretion. Recent work showed that exposure to glucan-spiked dust might influence nasal and pulmonary function, without an accompanying inflammatory response. However, effects of glucan-spiked dust exposure on NOS and GSNO reductase (enzymes important to NO signaling) remain less clear. This study aims to determine the effects of simultaneous exposure to glucan-spiked dust on NO signaling pathway in the airway. Danish Office dust was spiked with 1% (1-3)-ß-glucan (curdlan). Mice were exposed to 20 µL PBS (controls), 20 µL 25 µg/20 µL OVA and 20 µL 100 µg/20 µL glucan-spiked dust, respectively, daily for 12 days. NOS and GSNO reductase activity were measured in lung homogenate. Glutathione concentration and SOD activity in lung tissue were also determined to evaluate changes in oxidative stress. IL-6 concentration was measured in lungs to quantify the inflammatory response. Results showed that 12 day OVA and glucan-spiked dust exposure did not significantly influence NOS activity, GSH concentration, SOD activity, or IL-6 concentration. An insignificant increase in GSNOR activity and expression was observed in 12 day OVA-exposed mice, whereas glucan-spiked dust exposure significantly increased GSNOR activity and expression. Our results suggested that repeated glucan-spiked dust exposure to the airway could activate GSNO reductase but not NOS. Since GSNO reductase plays a pivotal role in NO signaling, these results may have clinical importance.

Deoxycholic acid modified-carboxymethyl curdlan conjugate as a novel carrier of epirubicin: in vitro and in vivo studies.

Deoxycholic acid hydrophobically modified-carboxymethylated-curdlan (DCMC) conjugate was developed as a novel carrier for the anticancer drugs. Epirubicin (EPB), as a model drug, was physically loaded into DCMC self-assembled nanoparticles. EPB-loaded DCMC nanoparticles were almost spherical in shape and their size, in the range of 327.4-511.5 nm, increased with the EPB-loading content increasing. In vitro release of EPB from DCMC self-assembled nanoparticles showed sustained drug release pattern and the release rate was related to pH of release media and drug loading content. The cytotoxic activity of EPB-loaded DCMC nanoparticles was assayed by the MTT colorimetric assay. Compared with free drug, EPB-loaded DCMC nanoparticles showed the higher cytotoxicity, which may be attributed to the enhanced cellular uptake. In vivo toxicity study indicated that DCMC conjugate did not induce unexpected side effects. Tissue biodistribution study was performed in tumor-bearing mice. The result showed that DCMC increased the uptake of EPB in the tumor and decreased the uptake of EPB in kidney and heart, compared to free drug. Moreover, tumor volume reductions induced by DCMC conjugate, free EPB and EDNs were 24.3%, 58.9% and 70%, respectively, which suggested that EDNs could effectively retard the growth of the tumor.

Beta-glucan-indomethacin combination produces no lethal effects.

BACKGROUND: The most important quality of beta-glucans and the reason why so much attention has been devoted to them are their physiological effects. They are typical biological response modifiers with pronounced immunomodulating activity. However, some questions about possible side effects remain. AIM: Several papers reported the lethal side effects of non-steroidal anti-inflammatory drugs in glucan-treated mice, probably due to the peritonitis by enteric bacteria. However, these results were never independently confirmed. The purpose of this study is to evaluate these claims using several different types of glucans. METHODS: Effects of combined treatment with four different types of glucans and indomethacin were measured by evaluation of phagocytosis of HEMA particles by peripheral blood leukocytes and production of IL-6 in mouse serum. In addition, the level of blood glucose, colon length and survival after 30 days of treatment was measured. RESULTS: Our finding showed that simultaneous treatment with glucan and indomethacin caused a small decrease of phagocytic activity and IL-2 production. Two other tested parameters-blood glucose levels and colon length-that had been found to be significantly affected by this treatment, were virtually unchanged. In the final, yet most important part of the study, we found absolutely no mortality, regardless of the type of glucan or the routes of glucan administration. CONCLUSION: No adverse negative effects due to simultaneous treatment with glucan and non-steroidal anti-inflammatory drug dose was found, despite testing two different routes of glucan administration and four different types of glucan.

Asian sand dust aggravates allergic rhinitis in guinea pigs induced by Japanese cedar pollen.

Asian sand dust (ASD) contains microbial materials, sulfate (SO(4)(2-)), and nitrate (NO(3)(-)), and is derived from air pollutants in East China. ASD reportedly causes adverse respiratory health effects; a case in point is aggravated allergen-associated experimental lung eosinophilia. Guinea pigs were administered normal saline (control), ASD (0.3 mg/animal), ASD (0.6 mg/animal), Japanese cedar pollen (JCP) (0.2 mg/kg body weight), JCP + ASD (0.3 mg/animal), or JCP + ASD (0.6 mg/animal), into their nasal cavities at seven weekly intervals. The number of sneezes, amount of nasal secretions, and nasal obstructing response were measured as indices of nasal responses. Total immunoglobulin E (IgE) antibodies in serum and the number of eosinophils, histamine, and arachidonic acid metabolites in nasal cavity lavage fluids (NCLF) were also measured. ASD enhanced the JCP-associated nasal obstructing response, but not the number of sneezes or amount of nasal secretions. ASD enhanced JCP-associated cysteinyl leukotrienes (C(4), D(4), E(4)) and histamine production in NCLF. ASD augmented the number of eosinophils in NCLF and total IgE in serum induced by JCP. ASD enhanced eosinophil recruitment in the nasal mucosa, and goblet cell proliferation in the nasal epithelium induced by JCP. These results suggest that ASD enhances the nasal allergic reaction induced by repeated JCP administration in guinea pigs.

Epigenetic and phenotypic changes result from a continuous pre and post natal dietary exposure to phytoestrogens in an experimental population of mice.

BACKGROUND: Developmental effects of exposure to endocrine disruptors can influence adult characters in mammals, but could also have evolutionary consequences. The aim of this study was to simulate an environmental exposure of an experimental population of mice to high amounts of nutritional phytoestrogens and to evaluate parameters of relevance for evolutionary change in the offspring. The effect of a continuous pre- and post-natal exposure to high levels of dietary isoflavones was evaluated on sexual maturity, morphometric parameters and DNA methylation status in mice. Adult mice male/female couples were fed ad libitum either with control diet (standard laboratory chow) or ISF diet (control diet plus a soy isoflavone extract at 2% (w/w) that contained the phytoestrogens genistein and daidzein). In the offspring we measured: i) the onset of vaginal opening (sexual maturation) in females, ii) weight and size in all pups at 7, 14, 21 and 42 days post-natal (dpn) and iii) DNA methylation patterns in skeletal alpha-actin (Acta1), estrogen receptor-alpha and c-fos in adults (42 dpn). RESULTS: Vaginal opening was advanced in female pups in the ISF group, from 31.6 +/- 0.75 dpn to 25.7 +/- 0.48. No differences in size or weight at ages 7, 14 or 21 dpn were detected between experimental groups. Nevertheless, at age 42 dpn reduced size and weight were observed in ISF pups, in addition to suppression of normal gender differences in weight seen in the control group (males heavier that females). Also, natural differences seen in DNA methylation at Acta1 promoter in the offspring originated in the control group were suppressed in the ISF group. Acta1 is known to be developmentally regulated and related to morphomotric features. CONCLUSION: This study demonstrates in mammals that individuals from a population subjected to a high consumption of isoflavones can show alterations in characters that may be of importance from an evolutionary perspective, such as epigenetic and morphometric characters or sexual maturation, a life history character.

Branched fungal beta-glucan causes hyperinflammation and necrosis in phagocyte NADPH oxidase-deficient mice.

Chronic granulomatous disease (CGD), a genetic disorder characterized by the absence of a functional phagocyte NADPH oxidase, is a severe immune deficiency. However, non-infectious hyperinflammation is a second hallmark of the disease. In CGD mouse models, sterile hyperinflammation can be induced by A. fumigatus cell wall preparations. In this study, we used subcutaneous injection of microbial cell walls and cell wall components to identify causes of CGD hyperinflammation and to characterize its histological features. Sterile cell wall preparations from fungi (A. fumigatus, C. albicans, S. cerevisiae), but not from bacteria (S. aureus, P. aeruginosa, E. coli), caused prolonged and severe skin inflammation in CGD mice. To identify fungal cell wall elements responsible for this process, we investigated microbial cell wall-derived monosubstances. Injection of beta(1-3)(1-6)-glucan induced severe hyperinflammation in CGD mice, while other fungal cell components [mannan, (1-3) beta-glucan] or bacterial cell wall components (lipopolysaccharide, lipoteichoic acid) caused no or only moderate inflammation. beta-glucan-induced hyperinflammation was predominantly due to a defect in termination of inflammation, as in the initial stage (2 days), the severity of inflammation and the extent of cell death were comparable in wild-type and CGD mice. At later stages (7 days), beta(1-3)(1-6)-glucan-induced inflammation had subsided in wild-type mice. In contrast, CGD mice showed persistent severe inflammation with central necrosis, containing abundant apoptotic and necrotic cells. In summary, branched fungal beta-glucan induces a severe inflammatory reaction in the absence of phagocyte NADPH oxidase. As opposed to the commonly perceived notion that reactive oxygen species are the cause of cell death, our results demonstrate that tissue necrosis can be caused by the absence of a superoxide-producing enzyme.

Candida soluble cell wall beta-D-glucan induces lung inflammation in mice.

Bioactivity of cell wall component(s) of fungi has not been fully elucidated, especially in vivo. We isolated Candida soluble beta-D-glucan (CSBG) from Candida albicans (C. albicans). We investigated the effects of airway exposure to CSBG on the immune systems in the airways in mice. CSBG exposure induced neutrophilic and eosinophilic inflammation in the lung, which was concomitant with the increased local expression of proinflammatory cytokines including tumor necrosis factor-alpha, interleukin (IL)-1beta, IL-6, macrophage inflammatory protein -1alpha, macrophage chemoattractant protein -1, RANTES (regulated on activation and normal T cells expressed and secreted), and eotaxin. The lung inflammation with enhanced expression of proinflammatory proteins caused by CSBG was directly related to its structure, since structurally degraded products of CSBG by formic acid induced negligible responses in the lung. CSBG enhanced nuclear localization of phosphorylated signal transducer and activator of transcription (STAT)-6 in the lung. These results suggest that airway exposure to CSBG induces lung inflammation, at least partly, via the enhanced expression of proinflammatory cytokines and the activation of STAT-6 pathway, and can be a proper murine model for fungal lung inflammation.

Cell wall beta-glucan derived from Candida albicans acts as a trigger for autoimmune arthritis in SKG mice.

SKG mice are a recently established experimental model for rheumatoid arthritis (RA). Although they spontaneously develop chronic autoimmune arthritis under conventional conditions, SKG mice failed to develop chronic arthritis in a strictly controlled specific pathogen-free (SPF) environment. Beta-glucan (BG) from Laminaria digitata, laminarin (LAM), induced arthritis under SPF conditions, thus BG would be a pathogenic factor for arthritis in SKG mice. Therefore, we prepared BG from Candida albicans, a pathogenic fungus and investigated whether BG from C. albicans induced arthritis in SKG mice under SPF conditions. SKG mice were injected intraperitoneally with particulate BG (oxidative-Candida albicans (OX-CA)), soluble BG (Candida soluble beta-glucan (CSBG)) from C. albicans and LAM as a positive control. In addition, schizophyllan (SPG) from Schizophyllum commune or Mycobacterium whole cells were injected into SKG mice to induce arthritis. Mice injected with OX-CA, CSBG and SPG had more severe arthritis than with LAM, and whole Mycobacterium cells. IL-6 concentration in sera from SKG mice injected with OX-CA or CSBG was high, whereas not detected in sera from mice treated with LAM. In histological analysis, infiltration of inflammatory cells was observed in SKG mice injected with BG. These results suggest that fungal infection may be a factor to induce and exacerbate autoimmune diseases such as RA.

Effect of nitric oxide on beta-glucan/indomethacin-induced septic shock.

We have previously shown that repeated administration of nonsteroidal anti-inflammatory drugs (NSAIDs) to mice treated with beta-glucan, a biological response modifier, induced severe lethality. The lethality would be strongly related to the translocation of enterobacterial flora to the peritoneal cavity and disruption of the cytokine network. Reports suggest that nitric oxide (NO) can have an effective or detrimental role in septic shock. In the present study, we examined the effect of NO, an inflammatory mediator, on beta-glucan/indomethacin (IND)- induced septic shock by inhibiting its synthesis with N(G)-nitro-L-arginine methyl ester (L-NAME), a nonselective NO synthase (NOS) inhibitor. Nitrite concentration was used as an indicator of NO generation. Mortality in beta-glucan/IND-treated mice was increased by administering L-NAME. Numbers of bacteria in various organs of mice treated with beta-glucan/IND rose significantly within a couple of days of the administration of L-NAME. Additionally, TNF-alpha, IL-1beta, and IL-6 concentrations were enhanced in peritoneal exuded cells in culture. These results suggest a significant loss of the bactericidal activity of macrophages on the administration of a NOS inhibitor which enhanced the rate of enterobacterial invasion to the peritoneal cavity, resulting in systemic inflammatory response syndrome. The production of NO, therefore, provides a protective effect in beta-glucan/IND-induced sepsis.