Effects of the antlered form of Ganoderma lucidum on tumor growth and metastasis in cyclophosphamide-treated mice

We examined the alleviation of cyclophosphamide-induced immunodepression by the antlered form of Ganoderma lucidum (G. lucidum AF) and also evaluated the anti-tumor and anti-metastatic effects of G. lucidum AF in cyclophosphamide-treated mice. G. lucidum AF alleviated cyclophosphamide-induced decrease in body weight, natural killer (NK) activity, interferon (IFN)-gamma production, and cytotoxic T lymphocyte (CTL) activity, and inhibited the abnormal increase and decrease in interleukine (IL)-4 level due to cyclophosphamide administration. Post-treatment with cyclophosphamide and G. lucidum AF significantly inhibited tumor growth in MM 46-bearing mice. When Lewis lung carcinoma cells were injected into mice after a cyclophosphamide administration, metastasis of these cells to the lung was increased, but G. lucidum AF suppressed it. The anti-tumor and anti-metastatic effects of the combination of G. lucidum AF and cyclophosphamide might influence the modulatory effects of G. lucidum AF on both cellular and humoral immunity. These findings suggest that G. lucidum AF would be beneficial in alleviating the reduction of immune response by chemotherapeutic anti-cancer drugs.     

洁丽香菇胞外粗多糖对小鼠移植性S180肉瘤的抑制作用和免疫功能调节

对洁丽香菇胞外粗多糖中多糖和蛋白含量进行测定,其中多糖含量为21.87%,蛋白含量为7.14%。通过体内实验研究该多糖对S180肉瘤的生长抑制作用。结果表明洁丽香菇胞外粗多糖高、中、低剂量组均能不同程度抑制S180 肉瘤的生长,其中高剂量组（500mg/kg）抑瘤率最高（为39.44%）,同时能显著增加胸腺指数,降低脾指数,有效改善免疫器官受损现象;中、低剂量（250mg/kg,125mg/kg）亦能抑制肿瘤生长,抑瘤率分别为30.43%和23.40%。实验中采用ELISA法对血清中各种细胞因子含量进行测定,结果表明洁丽香菇胞外粗多糖高剂量组可明显诱导血清中TNF-α、IL-12的分泌,低剂量组可诱导IFN-γ、IL-10的分泌。由此认为洁丽香菇胞外粗多糖通过恢复免疫器官功能,增加血清中TNF-α、IL-12、IFN-γ和IL-10的含量,从而达到增强细胞免疫的作用,发挥抗肿瘤活性。     

参芝发酵产物的抗肿瘤活性及其对小鼠免疫功能的影响

为了探索红参水煎液的灵芝发酵产物对H22荷瘤小鼠的抗肿瘤活性及其对小鼠免疫功能的影响,通过体内抗肿瘤实验和增强免疫功能实验从抑瘤率、对免疫器官的影响指数、对非特异性免疫、体液免疫及细胞免疫的影响5个方面对该产物做了功能性评价。结果表明,在抗肿瘤实验中,参芝发酵产物高剂量组的抑瘤率达到51.65%,脾指数和胸腺指数均高于对照组和环磷酰胺(CTX)组;增强免疫功能实验中,3个实验的给药组小鼠和对照组小鼠相比都有显著性差异(P<0.01)。由此可见,将灵芝与人参在发酵层次上配伍具有显著的抑制荷瘤小鼠肿瘤生长及增强小鼠免疫功能的作用。     

灵芝三萜合成调控与抗肿瘤药理活性的研究进展

三萜类化合物是灵芝的重要药效成分之一,是一类活性天然产物,不仅在保肝、护肾、降血糖、抗缺氧等方面有着重要的应用价值,而且还具有很好的抗肿瘤活性。现就灵芝三萜的发酵培养优化和抗肿瘤药理活性的研究进展进行综述,以期为酸性和中性灵芝三萜类化合物的研究及进一步开发相应抗癌药物提供科学基础。     

Anti-hepatitis B activities of ganoderic acid from Ganoderma lucidum

Ganoderic acid, from Ganoderma lucidum, at 8 μg/ml inhibited replication of hepatitis B virus (HBV) in HepG2215 cells over 8 days. Production of HBV surface antigen and HBV e antigen were 20 and 44% of controls without ganoderic acid. Male KM mice were significantly protected from liver injury, induced with carbon tetrachloride, by treatment with ganoderic acid at 10 mg and 30 mg/kg·d (by intravenous injection) 7 days. Ganoderic acid at the same dosage also significantly protected the mice from liver injury induced by M. bovis BCG plus lipopolysaccharide (from Escherichia coli 0127:B8).     

纳米级灵芝子实体粉末和破壁灵芝孢子粉体外抗肿瘤活性研究

对纳米级灵芝子实体粉末及破壁灵芝孢子粉石油醚提取物(PE)、氯仿提取物(CE)、丙酮(AE)、甲醇提取物(ME)、水提取物(WE)与灵芝子实体及灵芝孢子提取量进行对比,利用GC-MS联用仪对石油醚提取物进行了成分分析鉴定,对水提物中总糖进行了含量测定,并利用宫颈癌细胞Hela和晶体上皮细胞SRA01/04进行了体外增殖作用和剂量效应关系研究,为灵芝资源的保护及进一步开发利用提供理论基础。结果表明,纳米化灵芝子实体及破壁灵芝孢子不同溶剂提取量显著增加,纳米级灵芝子实体粉末水提取物具有抑制宫颈癌细胞Hela和晶体上皮细胞SRA01/04增殖的作用。破壁灵芝孢子各溶剂提取物对宫颈癌细胞Hela和晶体上皮细胞SRA01/04没有明显的增殖抑制作用。     

Studies on the immuno-modulating and anti-tumor activities of Ganoderma lucidum (Reishi) polysaccharides

We describe here the isolation of Reishi polysaccharides for the study of their effect on cytokine expression in mouse splenocytes. A fraction (F3) has been shown to activate the expression of IL-1, IL-6, IL-12, IFN-gamma, TNF-alpha, GM-CSF, G-CSF, and M-CSF, and from this three subfractions have been prepared where F3G1 activates IL-1, IL-12, TNF-alpha, and G-CSF, F3G2 activates all the cytokines as F3 does, and F3G3 activates only IL-1 and TNF-alpha. Together with previous studies, the mode of action on macrophages has been proposed where F3 binds to TLR4 receptor and activates extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 to induce IL-1 expression.     

硬枝树花中抑制H_22荷瘤小鼠肿瘤的活性成分研究

研究了从硬枝树花中提取得到的4个单体化合物松萝酸(usnic acid)、去甲环萝酸(evernic acid)、巴尔巴地衣酸(barbatic acid)和水杨嗪酸(salazinic acid)对H22荷瘤小鼠的抑瘤作用,并且对抑瘤率、胸腺指数、脾指数及小鼠白介素-2含量等各个指标的进行检测,以说明此4种化合物对小鼠肿瘤生长的抑制效果。结果表明,松萝酸高、中剂量组,去甲环萝酸高、中剂量组,巴尔巴地衣酸低剂量组,水杨嗪酸高剂量组对小鼠肿瘤有较好的抑制效果,与阴性对照组比较有极显著差异(P0.01),并且这些组的H22荷瘤小鼠血清中白介素-2的含量显著增加,与抑制肿瘤活性具有相关性。     

Purification and characterization of lectin from fruiting body of Ganoderma lucidum: lectin from Ganoderma lucidum

A novel 114 kDa hexameric lectin was purified from the fruiting bodies of the mushroom Ganoderma lucidum. Biochemical characterization revealed it to be a glycoprotein having 9.3% neutral sugar and it showed hemagglutinating activity on pronase treated human erythrocytes. The lectin was stable in the pH range of 5-9 and temperature up to 50 degrees C. The hemagglutinating activity was inhibited by glycoproteins that possessed N-as well as O-linked glycans. Chemical modification of the G. lucidum lectin revealed contribution of tryptophan and lysine to binding activity. The thermodynamics of binding of bi- and triantennary N-glycans to G. lucidum lectin was studied by spectrofluorimetry. The lectin showed very high affinity for asialo N-linked triantennary glycan and a preference for asialo glycans over sialylated glycans. The binding was accompanied with a large negative change in enthalpy as well as entropy, indicating primarily involvement of polar hydrogen, van der Waals and hydrophobic interactions in the binding.     

Carboxymethylation of a polysaccharide extracted from Ganoderma lucidum enhances its antioxidant activities in vitro

The chemical carboxylmethylated polysaccharide (C-GLP), which derived from water-insoluble crude Ganoderma lucidum polysaccharide (GLP), was prepared. Water solubility, chemical characterization, and antioxidant activities in vitro of C-GLP were determined. The solubility of C-GLP in distilled water reached 100 mg/ml, which was much higher than the solubility of GLP. Chemical analysis indicated that C-GLP was composed of Glc:Man:Gal = 33.0:1.0:3.4 with a molecular weight of 1.8 × 10⁶ Da and a carboxymethyl content of 11.07%. The signals of carboxymethyl were found in IR and ¹³C NMR spectra. Moreover, a high antioxidant activity of C-GLP was observed, especially in scavenging of hydroxyl radical (83.7% at 5 mg/ml) and hydrogen peroxide (51.6% at 10 mg/ml). This study indicates the effects of carboxymethylation on water-insoluble polysaccharide and explores a potential antioxidant in food industry and pharmaceuticals.     

Ganoderma lucidum polysaccharides supplementation attenuates exercise-induced oxidative stress in skeletal muscle of mice

The present study was designed to determine the effects of Ganoderma lucidum polysaccharides (GL-PS) on exhaustive exercise-induced oxidative stress in skeletal muscle tissues of mice. The mice were divided into four groups (three GL-PS administered groups and the control group). The control group was administered with distilled water and GL-PS administered groups were administered with GL-PS (50, 100 and 200 mg/kg body weight per day). After 28 days, the mice performed an exhaustive swimming exercise, along with the determination of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT) activities and malondialdehyde (MDA) levels in the skeletal muscle of mice. The results showed that GL-PS could increase antioxidant enzymes activities and decrease the MDA levels in the skeletal muscle of mice. This study provides strong evidence that GL-PS supplementation possessed protective effects against exhaustive exercise-induced oxidative stress.     

菌草灵芝与段木灵芝的功效成分的比较研究

测定并比较菌草灵芝和不同产地的段木灵芝中的粗多糖、三萜类物质和孢子油中三萜类物质的含量和孢子油得率的差异。结果表明,菌草灵芝和不同产地的段木灵芝中的粗多糖、三萜类物质和孢子油中三萜类物质的含量和孢子油得率存在着差异,菌草灵芝中的粗多糖、三萜类物质和孢子油中三萜类物质的含量和孢子油得率都高于段木灵芝。     

柳生金针菇胞外粗多糖对小鼠肝癌H22移植性肿瘤的抑制作用和免疫功能调节的研究

本研究通过对柳生金针菇胞外粗多糖对肿瘤生长的抑制和机体免疫调节的体内实验,来探索柳生金针菇胞外粗多糖的相关抗肿瘤机制。结果表明柳生金针菇胞外粗多糖高、中、低剂量组能不同程度抑制肝癌H22肿瘤的生长。高剂量组抑瘤率最高,为35.25%,中低剂量也能抑制肿瘤生长,缓解脾脏肿大和胸腺萎缩。同时免疫学实验结果表明柳生金针菇胞外粗多糖可明显增加血清中各种细胞因子的表达,通过增强细胞和体液免疫功能来起到抗肿瘤作用。     

Modulatory effect of Ganoderma lucidum polysaccharides on serum antioxidant enzymes activities in ovarian cancer rats

In the present study, we isolated polysaccharides from Ganoderma lucidum and investigated its effect on serum antioxidant enzymes activity in ovarian cancer rats to explore the mechanism underlying the pharmacological anti-cancer activity of the polysaccharides. Rats were grouped into the control, model and polysaccharides-treated groups. After experiment ended, serum antioxidant enzymes activity in rats were measured. Results showed that polysaccharides from G. lucidum significantly reduced MDA production and increased serum antioxidant enzymes activity. These results suggest that the antioxidant activity of polysaccharides from G. lucidum might be benefical towards ovarian cancer therapy.     

Anti-tumor effects of an antiserum raised in syngeneic mice to a tumor-specific T suppressor factor

Summary DBA/2 mice inoculated with either cells from the syngeneic P815 tumor or tumor cell membrane extracts develop T suppressor cells which suppress the in vitro generation of cytotoxic T lymphocytes with specificity for the tumor. A soluble suppressor factor with similar properties can be isolated from suppressor cell-enriched populations. It can be highly purified by appropriate immunoadsorption. Antisera to this suppressor factor raised in either DBA/2 or C57BL/6 mice can specifically absorb out suppressor factor and eliminate suppressor cells in the presence of complement. The in vivo effects of these antisera were tested for their ability to modulate the growth of P815 tumors in DBA/2 mice. It was found that the antiserum raised in syngeneic (DBA/2) but not allogeneic (C57BL/6) mice was able to significantly slow the rate of tumor growth and to prolong survival in treated mice. The antiserum was effective in this way only if it was administered early in the course of tumor growth. It was shown that this effect was not attributable to the presence in the serum of antibodies directed to antigens present on P815 cells, and it therefore appears to be due to interference with the function of T suppressor cells arising early in the immune response to the tumor cells.     

Effector phenotypes and mechanisms of antitumor immune reactivity of tumor-immunized and tumor-bearing mice in two syngeneic tumors.

By using two different syngeneic tumors, Meth A sarcoma and RL male 1 lymphoma of BALB/c origin, the present study was designed to investigate the subset(s) of T cells mediating in vivo antitumor immune responses and some of the effector mechanisms of in vivo protective immunity in BALB/c mice immunized against tumor or bearing tumor. Spleen cells from the mice immunized against Meth A tumor or bearing Meth A tumor inhibited the growth of Meth A tumor in the Winn assay. In the Meth A-immunized mice, L3T4+ (CD4+) cells played a major role in mediating the inhibitory activity against Meth A tumor growth, whereas in the Meth A-bearing mice, the antitumor protective immunity was mediated by both L3T4+ and Lyt-2+ (CD8+) cells. Spleen cells from the Meth A-immunized or Meth A-bearing mice were not able to generate cytotoxic T lymphocytes (CTL) directed against Meth A tumor after the in vitro restimulation of spleen cells with mitomycin C (MMC)-treated Meth A cells, while fresh spleen cells from the Meth A-immunized or Meth A-bearing mice were able to induce the strong delayed-type hypersensitivity (DTH) responses to Meth A tumor. The DTH response to Meth A tumor was mediated by L3T4+ cells in the Meth A-immunized mice and by both L3T4+ and Lyt-2+ cells in the Meth A-bearing mice. In the similar experiments performed in the RL male 1 lymphoma, the antitumor activity in spleen cells from the RL male 1-immunized or RL male 1-bearing mice depended on Lyt-2+ but not L3T4+ cells in the Winn assay. When spleen cells from the RL male 1-immunized or RL male 1-bearing mice were cultured with MMC-treated RL male 1 cells for 5 days, an appreciable CTL response to RL male 1 tumor was induced. These results suggest that the nature of tumor and/or tumor antigens determines which T cell subset is required to exhibit the protective immunity against tumor and thus the different effector mechanisms could be induced in the different tumor models. Furthermore, these data support the conclusion that antitumor T cell responses are affected by the immune state of host to tumor.     

Schizophyllan (SPG)-treated macrophages and anti-tumor activities against syngeneic and allogeneic tumor cells

Summary We tested anti-tumor activities of macrophages treated with a neutral polysaccharide, schizophyllan (SPG), against syngeneic and allogeneic tumor cell lines. SPG was a macrophage stimulant which was not mitogenic to lymphocytes. That made a sharp contrast with the data that Corynebacterium parvum, BCG, and muramyl dipeptide (MDF) were macrophage stimulants which had lymphocyte-activating properties. Treatment of SPG-treated PEC with Thy12 monoclonal antibody and guinea pig complement did not affect the capabilities of tumor-cell-growth suppression by the treated PEC. Thus, the effector cells were peritoneal adherent cells (macrophages morphologically) and effector-to-target contact seemed to be necessary for effective tumor-cell-growth inhibition, although contradictory data exist for this. Murine peritoneal adherent cells harvested 4 days after a single IP injection of SPG at a dose of 100 mg/kg body weight of mouse showed the most prominent cytostatic and cytotoxic activities against syngeneic and allogeneic tumor cells. The distribution of anti-tumor activity in macrophages of various sizes followed the same pattern as macrophages treated with C. Parvum, i.e., larger macrophages showed more remarkable anti-tumor activity. Crude nonadherent peritoneal cells incubated with SPG at a concentration of 10 g/ml, 100 g/ml, or 1 mg/ml did not secrete lymphokine that rendered macrophages cytotoxic, while ConA-treated nonadherent cells did so. Furthermore, spleen cells treated with SPG in vivo did not secrete macrophage-activating lymphokine in the presence of SPG. On the other hand, addition of 1 mg/ml of SPG-treated peritoneal adherent cells and bone-marrow-derived macrophages in vitro rendered them cytotoxic to a moderate degree. This implies that SPG may activate macrophages directly, allowing them to become cytotoxic in the peritoneal cavity. Lastly, SPG could induce production of II-1-like factor to a moderate degree. SPG, whose molecular structure is well elucidated, will provide us with a strong tool to analyze the mechanism of macrophage activation both in vitro and in vivo.Abbreviations PEC peritoneal exudate cells - SPG schizophyllan - LPS lipopolysaccharide - Con A concanavalin A - CGN carrageenan - B. M. bone marrow - FCS fetal calf serum - BCG bacille Calmétte Guérin - Il-1 interleukin 1 - PPD pure protein derivatives - MDP muramyl dipeptide - C. parvum Corynebacerium parvum Dr. Sugawara is a Research Fellow of the Alberta Heritage Foundation for Medical ResearchDr. Lee is a Research Associate of the National Cancer Institute of Canada     

灵芝子实体-益生菌发酵液对环磷酰胺诱发的肠黏膜损伤及菌群紊乱的调节作用

目的 建立灵芝子实体-益生菌发酵体系,并研究该发酵液对于环磷酰胺诱导的肠黏膜损伤及菌群紊乱的防治作用。 方法 通过吸光度(A值)、多糖含量检测等方法评价嗜酸乳杆菌及短双歧杆菌在灵芝子实体提取液中的生长情况。将混合益生菌菌种与灵芝培养基进行共同发酵,发酵液用于对环磷酰胺诱导的肠黏膜损伤模型小鼠的治疗。 结果 嗜酸乳杆菌、短双歧杆菌均能在灵芝子实体的培养基上生长,且A值及总糖含量均高于对照组。该发酵液对环磷酰胺诱导的肠黏膜损伤模型动物有一定的治疗作用,表现为发酵液干预后由环磷酰胺导致的小鼠体质量降低、结肠组织损伤、炎细胞浸润均有所缓解;并恢复肠道菌群结构,显著增加乳杆菌属丰度,降低肠球菌属和鞘脂单胞菌属丰度。 结论 灵芝子实体-益生菌发酵液可以有效改善环磷酰胺诱导的肠黏膜损伤的症状,其机制可能与调节肠道微生态平衡密切相关。     

Schizophyllan (SPG)-treated macrophages and anti-tumor activities against syngeneic and allogeneic tumor cells

We tested anti-tumor activities of macrophages treated with a neutral polysaccharide, schizophyllan (SPG), against syngeneic and allogeneic tumor cell lines. SPG was a macrophage stimulant which was not mitogenic to lymphocytes. That made a sharp contrast with the data that Corynebacterium parvum, BCG, and muramyl dipeptide (MDF) were macrophage stimulants which had lymphocyte-activating properties. Treatment of SPG-treated PEC with Thy12 monoclonal antibody and guinea pig complement did not affect the capabilities of tumor-cell-growth suppression by the treated PEC. Thus, the effector cells were peritoneal adherent cells (macrophages morphologically) and effector-to-target contact seemed to be necessary for effective tumor-cell-growth inhibition, although contradictory data exist for this. Murine peritoneal adherent cells harvested 4 days after a single IP injection of SPG at a dose of 100 mg/kg body weight of mouse showed the most prominent cytostatic and cytotoxic activities against syngeneic and allogeneic tumor cells. The distribution of anti-tumor activity in macrophages of various sizes followed the same pattern as macrophages treated with C. Parvum, i.e., larger macrophages showed more remarkable anti-tumor activity. Crude nonadherent peritoneal cells incubated with SPG at a concentration of 10 micrograms/ml, 100 micrograms/ml, or 1 mg/ml did not secrete lymphokine that rendered macrophages cytotoxic, while ConA-treated nonadherent cells did so. Furthermore, spleen cells treated with SPG in vivo did not secrete macrophage-activating lymphokine in the presence of SPG. On the other hand, addition of 1 mg/ml of SPG-treated peritoneal adherent cells and bone-marrow-derived macrophages in vitro rendered them cytotoxic to a moderate degree. This implies that SPG may activate macrophages directly, allowing them to become cytotoxic in the peritoneal cavity. Lastly, SPG could induce production of II-1-like factor to a moderate degree. SPG, whose molecular structure is well elucidated, will provide us with a strong tool to analyze the mechanism of macrophage activation both in vitro and in vivo.     

Purification and properties of laccase from Ganoderma lucidum

Summary The enzyme laccase has been partially purified from the culture fluid of Ganoderma lucidum by acetone precipitation, ammonium sulphate fractionation and adsorption on alumina C gel. The enzyme has been shown to be specific for ortho and para hydroxyphenolic compounds, having Km values of 5.5×10^-5 M and 2.86×10^-5 M for catechol and hydroquinone respectively. The optimum pH for the oxidation of catechol and hydroquinone are 5.4 and 5.0 respectively. The enzyme is inactivated above 60°C and is inhibited by enzyme inhibitors and metal chelating agents like azide, cyanide etc.