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Inhibition of IgE-dependent Mouse Triphasic Cutaneous Reaction by a Boiling Water Fraction Separated from Mycelium of Phellinus linteus

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Phellinus linteus, a mushroom, contains constituents that exhibit potent antitumor effects through activating immune cells. Recently, anti-inflammatory and anti-allergic properties of P. linteus extracts have also been implicated. In the present study, therefore, we separated the constituents of mycelium of P. linteus into five fractions-chloroform-soluble (CF), ethyl acetate-soluble (EA), methanol-soluble (AE), water-soluble (WA) and boiling water-soluble (BW) fractions-and examined their suppressive effects on the IgE-dependent mouse triphasic cutaneous reaction. The triphasic reaction was induced in the ear of BALB/c mice passively sensitized with anti-dinitrophenol IgE by painting with 2,4-dinitrofluorobenzene 24 h later. Ear swelling appeared triphasically with peak responses at 1 h, 24 h and 8 days after the challenge. ME, WA and BW given orally at a dose of 100 mg kg significantly inhibited the first and second phase ear swelling, and BW also inhibited the third phase response. CF only inhibited the second phase. The inhibition by BW was the most potent and almost dose-dependent at doses of 30-300 mg kg. BW also inhibited vascular permeability increase caused by passive cutaneous anaphylaxis and histamine, and ear swelling caused by tumor necrosis factor-alpha. In contrast, BW apparently potentiated the production of interleukin-4 and interferon-gamma from anti-CD3-stimulated mouse splenocytes. These results indicate that BW derived from mycelium of P. linteus contains some constituents with anti-allergic as well as immunopotentiating properties.
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... The proteoglycans of P. nigricans showed antitumor and immunomodulatory activities [74] . P. linteus extract inhibited ear swelling in BALB/c mice by enhancing the generation of interleukin-4 and IFN-γ from anti-CD3stimulated mouse splenocytes [75] . Enhancement of the production of Th1-and Th2-type cytokines in mice was seen with oral administration of P. linteus extract [76] . ...
... Hymenophore Aqueous extract Antibacterial [56] Hymenophore Phelligridin A Antioxidant [107] Phelligridin G Antioxidant and cytotoxic [109] Mycelium Methanol extract and hot water extract Antioxidant [27] Hymenophore Phelligridin H and Phelligridin I Antioxidant [25] Phelligridin J Cytotoxic Aqueous, ethanol (50%, 80%, pure) and ethyl acetate extract Antioxidant [86] Ethanol extract Antioxidant [102] P. linteus Mycelium Exopolysaccharides Antidiabetic [117] Hymenophore Exopolymers [118] Ethanol extract Antioxidant and antiangiogenc [119] Methanol extract and its n-butanol fraction Antibacterial [53] n-butanol subfraction Antiinflammatory [65] Mycelium Different solvent fractions Antitumour [120] Antioxidant [24] Ethanol and Ethylacetate combined extract Antiinflammatory [68] Hymenophore Phellinusfuran A and Phellinusfuran B Anticomplement activity [78] Mycelium Chloroform, ethylacetate, methanol, water and boiled water extract Antiallergic [75] Hot water extract Immunomodulatory activity [71] Mycelium Hispidin, 3,14ˊ-bihispidinyl, hypholomine B and 1,1distyrylpyrylethan Antioxidant [110] Hymenophore Phellifuropyranone A Antiproliferative [111] Ethanol extract Antiallergic [73] Nine different protein glycation inhibitors from ethylacetate fraction Antidiabetic [32] 10 different antioxidant compounds Antioxidant [36] Phellinusfurans A and B Anticomplement activity [78] Aqueous extract Antioxidant [122] Crude methanol, chloroform and ethyl acetate extracts Antimalarial, antioxidant and cytotoxic [49] Aqueous extract CNS activity [80] Methanol and ethyl acetate extract Antibacterial [58] Mycelium Hispidin from mycelial culture Antidiabetic and antioxidant [123] Hymenophore Ethanol extract Anticancer [50] Polysaccharides Antitumour [124] Polysaccharides Antioxidant [18] Hymenophore Aqueous, ethanol (50%, 80%, pure) and ethylacetate extract Antioxidant [86] Hymenophore Ethanol extract Antioxidant [102] Aqueous extract Antibacterial [63] Hispidin Antioxidant [30] Mycelium Ethanol extract and Phellinulin A Hepatoprotective [41] P. lloydii Hymenophore Sesquiterpenes Antibacterial and antifungal [60] P. merrillii Hymenophore ...
... Traditionally, P. linteus (Hymenochaetaceae) was commonly used as medicines or healthy foods in oriental countries to treat several healthy problems such as ulcer, diabetes, cancer and infections. In the previous reports, P. linteus exhibited wide spectrum of bioactivities, such as anti-platelet aggregation, anti-diabetic, anti-dementia, antioxidant, antiinflammatory, cytotoxic and anti-viral effects [13][14][15][16][17][18]. In our previous paper, the mycelium ethanol extract of P. linteus and purified compound phellinulin A (1) displayed significant protection against hepatic fibrosis [19]. ...
... Fraction F3 was isolated by silica gel CC with gradient mixture of chloroform and methanol (29:1 to 1:1) to afford three sub fractions (F3-1-F3-3). F3-1 and F3-2 were further purified by silica gel CC and preparative TLC to yield phellinulins A (1) (7.1 mg), J (10) (11.0 mg), M (13) (1.1 mg) and γionylideneacetic acid (18) (1.1 g), respectively. ...
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In the dimethylnitrosamine (DMN)-induced hepatic fibrosis Wistar rat model, the mycelium extract of Phellinus linteus (PLE) (20 mg/Kg) displayed significant protection against hepatic fibrosis. The present investigation characterized eleven new ionone derivatives, phellinulins D–N (4–14), from the P. linteus mycelium extract and the relative stereochemical structures were constructed according to the spectroscopic and spectrometric analytical results. Some purified compounds were examined for their inhibitory effects on activated rat hepatic stellate cells (HSCs) and several isolates did exhibit significant protection. The results indicated that the mycelium of P. linteus could be explored as a hepatoprotective drug or healthy food candidate in the near future.
... According to traditional applications and empirical practice, PL has various pharmacological activities including regulating blood glucose, improving blood circulation, hepatoprotecting and enhancing immunologic function, etc. Also, PL has been experimentally demonstrated to possess antitumor, immuno-modulatory, anti-inflammatory, anti-oxidant, anti-hyperlipidemic and anti-diabetic activities [13][14][15][16][17]. ...
... Totally 35 flavones, pyranones and furans have been seperated from PL. Among them, four flavones (phelligrins A and B as well as meshimakobnol A and B (1-4)), fifteen pyranones (phelligridimer A, phelligridins A-J, baumin, phellinin C as well as phellinin B1 and B2 (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)) and five furans (phellinusfrans A and B, phellifuropyranone A, phellinstatin as well as phelliusin A (20-24)) were isolated as the characteristic components of PL (Fig. 3). Mo et al. [91] firstly isolated 7 flavones including naringenin, sakuranetin, aromadendrin, folerogenin, eriodictyol coumarin and scopoletin from PL. Afterwards, two new and special ingredients of benzylated dihydroflavones, phelligrins A and phelligrins B, were isolated and identified as 5, 7, 4′-trihydroxyl-6-O-hydroxylbenzyldihydroflavone and 5, 7, 4′-trihydroxyl-8-Ohydroxylbenzyldihydroflavone [65]. ...
... Growing evidence indicates that P. linteus possesses a wide range of therapeutic effects, including antioxidative, antimicrobial, antiviral, antitumor, anti-inflammatory, antidiabetic, neuroprotective, and immunomodulatory [12,13]. In particular, it has been reported that mycelia of P. linteus exhibits a wide range of pharmacological activities, such as hepatoprotective [14,15], antiinflammatory [16], anti-atopic [17], anti-allergic [18], anti-osteoarthritic [19], and immunomodulatory [16] effects. Nevertheless, pharmacological and therapeutic studies on the whole parts or specific extracts of P. linteus mycelium have been explored relatively less than those with fruiting bodies. ...
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Medicinal mushrooms are an important natural resource promoting health benefits. Herein, Phellinus linteus mycelia were prepared under submerged cultivation, the mycelium-containing culture broth was extracted as a whole to obtain the postbiotic materials (PLME), and its effect on the immune system was evaluated in normal C3H/HeN mice. Oral administration of PLME for 4 weeks was well tolerated and safe. In the PLME-administered groups, in addition to the production of immunostimulatory cytokines, such as interferon gamma (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukin 6 (IL-6), the mitogenic activity was significantly increased. PLME administration also significantly increased the levels of serum immunoglobulin G (IgG) and IgA in the small intestinal fluid and Peyer’s patches and enhanced Peyer’s patch-mediated bone marrow cell proliferation activity and cytokine production (IL-2, IL-6, and IFN-γ). Histomorphometric analyses showed an increase in immune cells in the spleen and small intestinal tissues of mice administered PLME, supporting the rationale for its immune system activation. PLME mainly contained neutral sugar (969.1 mg/g), comprising primarily of glucose as a monosaccharide unit. The β-glucan content was 88.5 mg/g. Data suggest that PLME effectively promote immune function by stimulating the systemic immune system through the spleen and intestinal immune tissues. PLME can thus be developed as a functional ingredient to enhance immune functions.
... These are rich in various bioactive constituents like polysaccahrides, alkaloids, tannins, flavonoids, phenols, terpenoids and anthocyanins [14][15]. These bioactive constituents attribute to their significant biological activities such as anti-allergic [16,17], anti-cancer [18][19][20][21][22], anti-diabetic [23], anti-inflammatory [24], anti-microbial [25][26][27], anti-oxidant [28][29], hepatoprotective [30] and immuno-stimulation [31][32]. However physicochemical and biochemical characterization have not been evaluated for all the species belonging to Phellinus. ...
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The genus Phellinus Quél. (Family-Hymenochaetaceae) is a cosmopolitan polypore mushroom including a large number of wood rotting species. However some of these species have pharmacological significance and are in use as folk medicine since ages. The present investigation aims at taxonomic study, evaluation of physicochemical properties and screening of mycochemical composition of the specimen collected from district Dehradun, Uttarakhand (India). Based on morphology and microscopy, the specimen was identified as Phellinus pachyphloeus (Pat.) Pat. This mushroom showed negligible foreign matter, low moisture content (13.67%), high dry weight (86.33%), good flow characteristics, high dispersibility (85.67%), high alcohol soluble extractives, high water absorption capacity, good emulsion properties and high content of water soluble ash (3%). The preliminary mycochemical screening was done as per established standards and showed the presence of carbohydrates, reducing sugars, proteins, amino acids, steroids, terpenoids, phenols, flavonoids, tannins, anthraquinone glycosides and cardiac glycosides. These results for Phellinus pachyphloeus (Pat.) Pat. are an indication of its potential use in pharmaceutical and nutraceutical preparations.
... The most potent fraction was the boiling water-soluble fraction. It inhibited dose dependently (30-300 mg/kg) all phases, inhibited vascular permeability increase caused by passive cutaneous anaphylaxis and histamine, and ear swelling caused by TNFα (Inagaki et al. 2005). ...
Chapter
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Background:  Previous studies have suggested that mice passively sensitized with anti-dinitrophenol (DNP) monoclonal IgE antibody exhibit a triphasic cutaneous reaction with an immediate-phase response (IPR) at 1 h, a late-phase response (LPR) at 24 h and a very late-phase reaction (vLPR) at 8 days after challenge with 2,4-dinitrofluorobenzene. The present study was conducted to elucidate the role of T cells in this tri­phasic cutaneous reaction.Methods:  Mice were passively senseitized by an intra­venous injection of anti-DNP monoclonal IgE antibody. Allergic cutaneous reaction was caused by painting an antigen on the ears of mice and measured an enlargement of the ears.Results:  Whereas the magnitudes of IPR and LPR in BALB/c nu/nu T cell-deficient mice were similar to those in BALB/c +/+ mice, vLPR was not observed in nu/nu mice. In addition, FK 506 (3 and 5 mg/kg) and cyclosporin A (30 and 50 mg/kg) clearly suppressed the onset of vLPR without affecting IPR and LPR. Because these findings suggest the participation of T cells in the onset of vLPR, the character of the T cells was investigated by using anti-CD4 or anti-CD8 monoclonal antibody (mAb) and interleukin (IL)-4 and IL-5 receptor α chain gene-deficient mice. When mice were treated with anti-CD4 or anti-CD8 mAb, the magnitude of the vLPR was augmented by anti-CD4 mAb and suppressed by anti-CD8 mAb, without affecting IPR and LPR. Disruption of the IL-4 gene slightly suppressed IPR, LPR and vLPR, but the lack of the IL-5Rα chain gene did not affect these triphasic responses.Conclusions:  The present findings suggest that vLPR is mainly caused by CD8-positive T cells, probably Tc1 cells, and regulated by CD4-positive T cells.
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Previous studies have reported that the mice passively sensitized with anti-dinitrophenol (DNP) IgE antibody exhibited IgE-mediated cutaneous reaction with an immediate phase response (IPR) at 1 h and a late phase response (LPR) at 24 h after the challenge of dinitrofluorobenzene (DNFB). We found that the third- phase inflammatory reaction with intense and persisting infiltration of eosinophils, named ‘very late phase reaction (vLPR)’, was induced following IPR and LPR in response to DNFB in actively and passively sensitized mice, and that the peak response of vLPR was at 8 days after the challenge. This reaction was slightly observed in non-sensitized mice. Since the accumulation of eosinophils in vLPR was markedly observed when compared with that of LPR at 24 h, the vLPR may be an important reaction in allergic diseases. The development of vLPR was partly decreased in mast cell-deficient WBB6F1-W/Wv mice and was absent in T cell-deficient BALB/c-nu/nu mice in passive sensitization. These results indicate that the vLPR in the triphasic cutaneous reaction may be mainly mediated by T cells and partially by mast cells and/or IgE antibody, and consequently lead to an intense ear swelling accompanying massive infiltration of eosinophils.
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Several attempts were made to elucidate the possible role of histamine, serotonin, leukotrienes C4 (LTC4) and D4 (LTD4), and prostaglandin E1 (PGE1) as vascular permeability increasing factors involved in 48-hour homologous passive cutaneous anaphylaxis (PCA) in the mouse ear. Increased vascular permeability in the mouse ear caused by the mediator injection or PCA was assessed quantitatively by measuring the amount of extravasated dye. In skin reactions, all of the mediators used in the present study significantly increased vascular permeability. The most potent mediator was serotonin, which increased the vascular permeability from a concentration of 10(-8) g/ml, and the activity was about 100 times higher than that of histamine on a weight basis. Vascular permeability increasing activity of LTC4 was about 10 times higher than that of histamine, and LTD4 and PGE1 were also more potent than histamine. Increases of vascular permeability caused by histamine, serotonin, LTC4 and LTD4 were significantly potentiated by injecting 10(-6) g/ml of PGE1 simultaneously. Histamine-, serotonin- and LTC4-induced skin reactions in the mouse ear were suppressed significantly by the administrations of chlorpheniramine, methysergide and FPL 55712, respectively. In contrast, though chlorpheniramine and methysergide suppressed also mouse ear PCA (about 50 and 40%, respectively), neither FPL 55712, indomethacin nor BW 755C suppressed it. These results strongly suggest that the most important mediator involved in mouse ear PCA is histamine and that serotonin also plays an important role in the increase of vascular permeability caused by PCA. Despite their potent vascular permeability increasing activity LTC4, LTD4 and PGE1 do not seem to play an important role in mouse ear PCA.
Passive cutaneous anaphylaxis (PCA) was elicited both in the ear and in the dorsal skin of 13 strains of mice at the same time and assessed quantitatively by measuring the amount of extravasated dye. Body pigments of colored mice such as DBA/2 (chocolate), C3H/He (brown) and C57BL/6 (black) did not interfere with the measurement of dye. In the ear response, ICR was a higher responder, C57BL/6 and BALB/c-nu/nu were lower responder strains. In the dorsal skin response, however, ICR was a lower responder, BALB/c-nu/nu, Hairless and WBB6F1-+/+ were higher responders. WBB6 F1-W/Wv was a nonresponder in both responses. The ear response was highly reproducible and the dorsal skin response of each strain was 1/2-1/10 of its ear response except for BALB/c-nu/nu. The PCA bluing regions on the dorsal skin of BALB/c-nu/nu were clearly delineated and the response was almost comparable to its ear response.
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Several studies have suggested a possible role for IgE antibodies in the pathogenesis of cutaneous hypersensitivity reactions that reach maximum intensity 24 to 48 hr after antigen challenge. The recent availability of murine monoclonal IgE anti-hapten antibodies has made possible the direct examination of the range of cutaneous inflammatory reactions that can be mediated by such antibodies. We have examined the effects of passively sensitizing BALB/c mice with monoclonal IgE anti-dinitrophenyl (DNP) antibody 48 hr before antigen challenge. Inflammatory responses were assessed by measuring ear swelling in mice challenged on the ears with the reactive hapten 2,4-dinitrofluorobenzene (DNFB). Compared with unsensitized controls, the ears of mice passively sensitized with IgE anti-DNP displayed a biphasic pattern of ear swelling after DNFB challenge. An early, transient response (present within 15 to 30 min of challenge and returning to control levels within 4 to 9 hr) was followed by a second, more persistent increase in ear swelling that peaked 24 to 48 hr after challenge. This biphasic pattern of ear swelling seen in IgE-sensitized mice was temporally indistinguishable from that observed in mice conventionally sensitized for allergic contact dermatitis reactions by epicutaneous application of DNFB 5 days before DNFB ear challenge. Antigen specificity of the IgE-mediated contact hypersensitivity reactions was demonstrated by the failure of mice passively sensitized with IgE anti-DNP to display early or delayed ear swelling greater than unsensitized controls when challenged with either of two noncross-reacting haptens, fluorescein isothiocyanate or oxazolone. Mice passively sensitized with a monoclonal IgA anti-DNP antibody (MOPC 315) 48 hr before DNFB challenge failed to display early or delayed ear swelling greater than unsensitized controls. Heat inactivation of the IgE anti-DNP ascitic fluid at 56 degrees C for 30 min completely abolished its capacity to passively sensitize mice for contact hypersensitivity reactions after DNFB challenge. These results document the existence of an antigen-specific, IgE-mediated, delayed-in-time cutaneous hypersensitivity response that can be elicited by epicutaneous challenge (contract) with a reactive hapten.
Passive cutaneous anaphylaxis (PCA) reaction elicited in ears of male ddY mice was studied by means of assessing dye leakage. In the ear, PCA reaction was more sensitive and reproducible than that in the dorsal skin. The reaction was significantly suppressed by ketotifen, one of antianaphylactic agents.