浅谈灵芝提取物抗肿瘤作用的研究

目的探讨灵芝及其提取物的抗肿瘤作用及其机制研究情况。方法基于灵芝的中医药理论特点,通过文献收集、归纳、总结,综述了灵芝及其提取物的抗肿瘤作用及其机制研究情况。结果关于灵芝及其提取物抗肿瘤作用的研究和应用日益增加。其中,对灵芝多糖和灵芝三萜抗肿瘤作用及其机制的研究较多。结论探明灵芝及其提取物的抗肿瘤作用机制,在临床治疗、预防保健等层面上有着十分重要的意义。     

Ganoderma lucidum polysaccharides antagonize the suppression on lymphocytes induced by culture supernatants of B16F10 melanoma cells

Objectives Tumour cells produce factors such as interleukin 10 (IL‐10), transforming growth factor β1 (TGF‐β1) and vascular endothelial growth factor (VEGF) that suppress the function of immune cells or induce apoptosis of immune cells. One of the most important goals of tumour immunotherapy is to antagonize this suppression on immune cells. Ganoderma lucidum polysaccharides (Gl‐PS) may have this potential. The purpose of this study was to determine the antagonistic effects of Gl‐PS on the suppression induced by B16F10 melanoma cell culture supernatant (B16F10‐CS) on lymphocytes. Methods Gl‐PS was used on lymphocytes incubated with B16F10‐CS. Enzyme‐linked immunosorbent assay was used to determine the levels of IL‐10, TGF‐β1 and VEGF in B16F10‐CS. The MTT assay was used to determine the proliferation of lymphocytes. Immunocytochemistry and Western blot assay were used to determine perforin and granzyme B production in lymphocytes. Key findings There were elevated levels of IL‐10, TGF‐β1 and VEGF in B16F10‐CS. The lymphocyte proliferation, and perforin and granzyme B production in lymphocytes after induction with phytohemagglutinin, as well as lymphocyte proliferation in the mixed lymphocyte reaction, were suppressed by B16F10‐CS. This suppression was fully or partially antagonized by Gl‐PS. Conclusions B16F10‐CS suppressed lymphocyte proliferation and perforin and granzyme B production in lymphocytes after induction with phytohemagglutinin, as well as lymphocyte proliferation in the mixed lymphocyte reaction. This suppression may be associated with elevated levels of immunosuppressive IL‐10, TGF‐β1 and VEGF in B16F10‐CS. Gl‐PS had antagonistic effects on the immunosuppression induced by B16F10‐CS, suggesting the potential for Gl‐PS in cancer immunotherapy.     

灵芝多糖对高脂性脂肪肝大鼠氧化应激的影响

目的:研究灵芝多糖对高脂性脂肪肝大鼠氧化应激的影响。方法:SD大鼠72只,随机分为6组:正常对照组(N组),模型组(M组),灵芝多糖低(GLPs-L)、中(GLPs-M)、高(GLPs-H)剂量组(200,400,800 mg·kg-1)和辛伐他汀组(SV 1.8mg·kg-1)。用高脂饲料喂养建立大鼠高脂性脂肪肝(HFL)模型。经12周给药后,分别测定各实验组血清总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)、丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)和总抗氧化能力(T-AOC)以及肝组织中MDA,SOD,GSH-Px和T-AOC的含量;光镜观察肝组织脂肪病变程度。结果:M组血清TC,TG,LDL-C含量显著升高(P<0.01);与M组比较,GLPs-M,GLPs-H组血清TC,TG,LDL-C含量均显著降低(P<0.05或P<0.01),GLPs-H组HDL-C含量显著增加(P<0.05);GLPs-M,GLPs-H组血清和肝匀浆中MDA含量较M组相比均明显降低(P<0.05或P<0.01),SOD,GSH-Px和T-AOC水平较M组相比均明显上升(P<0.05或P<0.01)。GLPs-M,GLPs-H组可使肝细胞脂肪变性程度明显减轻。结论:灵芝多糖能明显降低高脂性脂肪肝大鼠血脂水平,有效对抗氧化应激,对大鼠高脂性脂肪肝有一定的防治作用。     

女贞子炮制过程中2种三萜类成分及水溶性低分子糖含量变化规律研究

目的:研究不同炮制条件对女贞子中三萜类成分齐墩果酸、熊果酸及低分子糖含量的影响,探讨女贞子酒制过程中齐墩果酸、熊果酸及低分子糖含量的动态变化规律。方法:对女贞子生品、清蒸品、酒蒸品及不同加热时间酒炖品中齐墩果酸、熊果酸和低分子糖含量进行测定比较。结果:女贞子经不同方法炮制后齐墩果酸和熊果酸含量无明显变化,而水溶性低分子糖含量增加,且随着炖制时间的延长,炮制品中低分子糖含量逐渐增加。结论:炮制对女贞子中齐墩果酸和熊果酸含量无明显影响,对低分子糖含量有显著影响。     

灵芝免疫活性多糖的研究

从云头状灵芝[Ganoderma Lucidum(Leyss.ex Fr.)Karst.]中分离得到三种灵芝多糖BN_3A,BN_3B 及 BN_3C,它们均表现明显免疫调节作用。从灵芝多糖 BN_3B 及 BN_3C 中各分离得到四个多糖均一体,对其中主要成分 BN_3C_1,BN_3C_3,BN_3B_1及 BN_3B_3进行了物理及化学研究。它们的平均分子量依次为1.6×10~4,2.5×10~4,3.5×10~4及4.0×10~4。经完全酸水解、红外光谱测定、过碘酸氧化、甲酸生成、Smith 降解等证明,BN_3B_1及 BN_3C_1均为β-(1→6)(1→3)甙键相连的葡聚糖。BN_3B_3为阿拉伯半乳聚糖。BN_3C_3为由葡萄糖和阿拉伯糖组成的肽多糖。它们均为β-(1→6)(1→3)甙键相连。     

2036灵芝孢子(全破壁)胶囊长期毒性试验报告

目的观察连续给予2036灵芝孢子(全破壁)胶囊对大鼠产生的毒性反应及严重程度。方法根据成人用量和本品最大灌胃浓度,将大鼠设定为高、中、低3个剂量给药组(分别为5.40g/kg、2.70g/kg、1.35g/kg)和1个阴性对照组。大鼠连续口服26周,停药后再观察4周,进行一般症状观察,血象、生化、脏器解剖、脏器系数和组织病理学检查。结果所有剂量动物组活动自如,外观、摄食和粪便等均未见异常,体质量增加与对照组比较无明显差异;血液生化指标与对照组比较均无显著差异;心、肝、脾、肺、肾、肾上腺、脑、胸腺、睾丸、卵巢的脏器系数与对照组比较均无显著性差异;尸检动物主要脏器未见明显病理变化。结论 2036灵芝孢子(全破壁)胶囊长期毒性试验未见异常反应,无毒剂量为5.40g/kg(相当于临床拟定人用剂量的60倍),临床上应用本品是安全可靠的,可以长时间较大剂量服用。     

灵芝颗粒质量标准研究

目的:建立灵芝颗粒的质量标准。方法:采用薄层色谱(TLC)法对方中灵芝、柴胡进行定性鉴别;采用高效液相色谱法测定五味子中五味子甲素、五味子乙素含量。结果:TLC斑点清晰,分离度好,阴性对照无干扰;五味子甲素进样量、五味子乙素进样量分别在0.060～0.60、0.082～0.82μg范围内与各自峰面积积分值呈良好线性关系(r分别为0.9996、0.9995);平均回收率分别为99.02%、99.06%,RSD分别为0.45%(n=6)、0.48%(n=6)。结论:定性定量方法快速、准确、专属性强,所建标准可用于灵芝颗粒的质量控制。     

Polysaccharides from sporoderm-removed spores of Ganoderma lucidum induce apoptosis in human gastric cancer cells via disruption of autophagic flux

The sporoderm-broken spores of Ganoderma lucidum (G. lucidum) polysaccharide (BSGLP) have been demonstrated to inhibit carcinogenesis in several types of cancer. However, to the best of our knowledge, the anticancer effects of polysaccharides extracted from the newly developed sporoderm-removed spores of G. lucidum (RSGLP) have not been assessed. The present study first compared the anticancer effects of RSGLP and BSGLP in three gastric cancer cell lines and it was found that RSGLP was more potent than BSGLP in decreasing gastric cancer cell viability. RSGLP significantly induced apoptosis in AGS cells, accompanied by downregulation of Bcl-2 and pro-caspase-3 expression levels, and upregulation of cleaved-PARP. Furthermore, RSGLP increased LC3-II and p62 expression, indicative of induction of autophagy and disruption of autophagic flux in AGS cells. These results were further verified by combined treatment of AGS cells with the late-stage autophagy inhibitor chloroquine, or early-stage autophagy inducer rapamycin. Adenoviral transfection with mRFP-GFP-LC3 further confirmed that autophagic flux was inhibited by RSGLP in AGS cells. Finally, the present study demonstrated that the RSGLP-induced autophagy and disruption of autophagic flux disruption was, at least in part, responsible for RSGLP-induced apoptosis in AGS cells. The results of the present study demonstrated for the first time that RSGLP is more effective than BSGLP in inhibiting gastric cancer cell viability, and RSGLP may serve as a promising autophagy inhibitor in the management of gastric cancer.     

灵芝孢子油对肺腺癌癌性胸水中原代肿瘤细胞的抗肿瘤作用

目的：研究灵芝孢子油对肺腺癌的癌性胸水中原代肿瘤细胞的抗癌作用。方法：体外培养肺腺癌细胞系A549,用CCK-8法检测灵芝孢子油对A549细胞的体外抗肿瘤作用。收集12例经病理确诊的肺腺癌患者的癌性胸水分离原代肿瘤细胞,用CCK-8法检测灵芝孢子油及临床常用化疗药物对原代肿瘤细胞的药物敏感性;流式细胞仪检测灵芝孢子油对原代肿瘤细胞的凋亡诱导作用,RT-PCR检测灵芝孢子油对凋亡相关蛋白基因bax及bc l-2 mRNA表达的影响。结果：灵芝孢子油对5例癌性胸水中原代肿瘤细胞的IC50平均值为0.28±0.04μl/m l,与对A549细胞的IC500.46±0.08μl/m l无显著差异（P〉0.05）。在各测试药物浓度（TDC）下,灵芝孢子油、多西紫杉醇、顺铂和健择对原代肿瘤细胞的抑制率分别为48.7%±3.3%,31.6%±14.1%,30.2%±11.6%,51.4%±5.6%,灵芝孢子油与这些化疗药物的抗肿瘤作用无显著差异（P〉0.05）。灵芝孢子油显著促进原代肿瘤细胞凋亡,对原代肿瘤细胞中凋亡相关蛋白bax mRNA有明显上调作用,对bc l-2 mRNA有明显下调作用。结论：灵芝孢子油对肺腺癌的癌性胸水原代肿瘤细胞有较好的抗癌作用。     

Ganoderma lucidum polysaccharides reduce methotrexate-induced small intestinal damage in mice via induction of epithelial cell proliferation and migration

Aim:

To study the effects of Ganoderma lucidum polysaccharides (Gl-PS) on methotrexate (MTX)-induced small intestinal damage in mice and the underlying mechanisms.

Methods:

BALB/c mice were used for in vivo study. The mice were administered with Gl-PS (50, 100, or 200 mg/kg, ig) for 10 d, and injected with MTX (50 mg/kg, ip) on d 7 and 8 to induce intestinal damage, and then sacrificed on d 11 for morphological study and tissue malondialdehyde (MDA) and superoxide dismutase (SOD) measurements. Before sacrificing, blood samples were collected to analyze immunoglobulin A (IgA). Rat intestinal IEC-6 cells were used for in vitro study. Cell proliferation and migration were assessed using MTT method and an in vitro wounding model, respectively. Transforming growth factor β (TGFβ) protein expression was determined using ELISA assay. Ornithine decarboxylase (ODC) and c-Myc mRNA expression profiles were determined using RT-PCR.

Results:

MTX treatment caused severe mucosal damage, significantly increased small intestine MDA levels, and decreased SOD and serum IgA levels in BALB/c mice. Gl-PS (100 and 200 mg/kg) markedly reversed the MTX effects. In IEC-6 cells, Gl-PS (0.1, 1, and 10 μg/mL) significantly stimulated the cell proliferation. Furthermore, Gl-PS (10 μg/mL) significantly stimulated the cell migration. In addition, Gl-PS (10 and 20 μg/mL) significantly increased the expression of ODC and c-Myc mRNAs. However, Gl-PS (up to 20 μg/mL) had no effect on the expression of TGFβ protein.

Conclusion:

The results suggest that Gl-PS protects small intestine against MTX-induced injury via induction of epithelial cell proliferation and migration.