| Summary: | 博士 === 國立臺灣大學 === 微生物與生化學研究所 === 95 === The purpose of this study was to establish both in vitro and in vivo systems to study and explore the possible functional foods for the allergic diseases. It has been well documented that fungi might exert immunomodulatory effect, and thus the Agaricus blazei, Poria cocos, Ganoderma tsugae and Antrodia camphorate were tested for allergic immunomodulatory effect, by using RBL-2H3 mast cell line, primary splenocyte form BALB/c; and OVA-specific CD4+ T cells from T cell receptor (TCR) transgenic DO11.10 mice (nanaïve OVA-specific T
cells). The results showed that ionomycin-stimulated histamine production form RBL-2H3
cells did not affect by Agaricus blazei, Poria cocos and Ganoderma tsugae, but increased by
Antrodia camphorate. OVA-stimulated IL-2 secretion form Th1 cells was increased, but IL-4
and IL-5 secretion from Th2 cells was decreased by all the fungi samples. Furthermore, fungi
treatment increased the IL-2 level and decreased Th2 cytokines production during naïve
OVA-specific T cell differentiation in vitro. To further investigate the effect of fungi on airway
inflammation, the allergen-sensitized mice challenged with aerosol allergen supplemented
with Agaricus blazei, Poria cocos or Ganoderma tsugae. The data showed that the percentage
of eosinophils, IL-5 and total protein content in bronchoalvelor lavage fluid (BALF), and IL-4,
IL-5 levels produced by splenocytes were significantly decreased in mice fed with Agaricus
blazei, Poria cocos and Ganoderma tsugae.
Furthermore, two major fractions of Ganoderma tsugae, triterpenoids (Gt-TRE) and
polysaccharides (Gt-PS) were tested for their contribution for regulation of histamine and
cytokines productions using several different cell models described above. The results showed
that ionomycin-stimulated histamine secretion from RBL-2H3 cell was significantly
suppressed by Gt-TRE, but not Gt-PS fraction. PMA/ionomycin-stimulated IL-4 secreted from
a murine T cell line EL4 was also significantly decreased by Gt-TRE only. To further
investigate whether Gt-PS and Gt-TRE affect CD4+ T cell polarization, naïve OVA-specific T
cells were cultured with Gt-PS or Gt-TRE. The results showed that Gt-PS enhanced Th1
cytokine IL-2 but suppressed IFN-γ significantly. Gt-TRE had no effect on Th1 cytokines but
dose-dependently suppressed Th2 cytokines more significantly than that of Gt-PS. To evaluate
the effects of Ganoderma tsugae on IL-4 production, EL4 cell was transfected a plasmid
containing reporter gene luciferase with IL-4 promoter. The results shown that luciferase activity did not affect by Gt-PS and Gt-TRE treatment, but IL-4 and IL-5 production were
decreased by Gt-TRE. In addition, both EL4 cell and RAW264.7 macrophage cell line
transfected a plasmid containing reporter gene luciferase with NF-κB binding sites promoter
were cultured with Gt-PS or Gt-TRE. The results showed that the luciferase activity was
significantly increased by Gt-PS but significantly decreased by Gt-TRE, suggesting that these
two fractions may exert different effect on NF-κB related cytokine expression.
An in vivo study was meant to investigate if triterpenoid extracts have anti-inflammatory
effects on airway responses and regulatory effects on Th2 responses. BALB/c mice sensitized
intra-peritoneally and challenged with OVA were treated with either TRE or prednisolone for 2
weeks. The effects of TRE on bronchial AHR, airway inflammation, serum antigen-specific
antibody levels, and cytokine secretions from splenocytes were evaluated. TRE treatment
significantly decreased AHR and reduced the total infiltrating leukocytes and eosinophils in
BALF when compared to the control group. Furthermore, TRE decreased the production of
inflammatory mediators, such as IL-4, IL-5, and eotaxin in BALF, as well as secretions from
splenocytes of OVA-sensitized mice. TRE treatment also significantly decreased OVA-specific
IgG1 production, but not IgG2a production. These data suggest that triterpenoids is the fraction
of Ganoderma tsugae that alleviate bronchoalveolar inflammation in allergen-induced
asthmatic animal model through the biological activity for suppression of Th2 responses.
Overall, these data suggest that triterpenoid-rich extracts of Ganoderma tsugae attenuated
histamine release, airway inflammation, airway hyperresponsiveness, Th2 cytokines secretion,
and serum OVA-specific IgG1 production, but not affect Th1 responses. The regulatory
mechanism may exert the NF-κB related Th2 cytokine expression through the PPARγ
activation. Triterpenoid-rich extracts of Ganoderma tsugae exert anti-inflammatory effects on
airway responses and attenuates Th2 responses without the overall immuno-suppression
effects in allergic murine models of asthma.
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