Effect of polysaccharides from Ganoderma lucidum on streptozotocin-induced diabetic nephropathy in mice

The effects of Ganoderma lucidum polysaccharides (GL-PS) on renal complication in streptozotocin-induced diabetic mice have been investigated in the present study. C57BL/6J mice were made diabetic by injection of streptozotocin and GL-PS (125 and 250 mg kg^- 1) was administered for 8 weeks. Body weight was monitored every week. Serum glucose, creatinine (Cr), blood urea nitrogen (BUN), triglyceride (TG) and urinary albumin excretion (UAE) were measured after 8 weeks of treatment. Glomerular size and mesangial matrix index were assayed by morphometric analysis. Renal expression of transforming growth factor-β₁ (TGF-β₁) were determined by immunochemistry. Renal malondialdehyde (MDA) level and superoxide dismutase (SOD) activities were also evaluated. GL-PS was able to reduce the serum Cr and BUN levels and UAE compared with diabetic model mice in a dose-dependent manner. Increasing serum glucose and triglyceride levels in diabetic mice could also be lowered by GL-PS. Moreover, GL-PS had the capacity to improve the renal morphometric changes and oxidative stress state of diabetic mice. In summary, GL-PS can improve the metabolic abnormalities of diabetic mice and prevent or delay the progression of diabetic renal complications.     

Effect of polysaccharides from Ganoderma lucidum on streptozotocin-induced diabetic nephropathy in mice

The effects of Ganoderma lucidum polysaccharides (GL-PS) on renal complication in streptozotocin-induced diabetic mice have been investigated in the present study. C57BL/6J mice were made diabetic by injection of streptozotocin and GL-PS (125 and 250 mg kg^? 1) was administered for 8 weeks. Body weight was monitored every week. Serum glucose, creatinine (Cr), blood urea nitrogen (BUN), triglyceride (TG) and urinary albumin excretion (UAE) were measured after 8 weeks of treatment. Glomerular size and mesangial matrix index were assayed by morphometric analysis. Renal expression of transforming growth factor-β₁ (TGF-β₁) were determined by immunochemistry. Renal malondialdehyde (MDA) level and superoxide dismutase (SOD) activities were also evaluated. GL-PS was able to reduce the serum Cr and BUN levels and UAE compared with diabetic model mice in a dose-dependent manner. Increasing serum glucose and triglyceride levels in diabetic mice could also be lowered by GL-PS. Moreover, GL-PS had the capacity to improve the renal morphometric changes and oxidative stress state of diabetic mice. In summary, GL-PS can improve the metabolic abnormalities of diabetic mice and prevent or delay the progression of diabetic renal complications.     

Hypoglycemic effect of Ganoderma lucidum polysaccharides

AIM: To investigate the hypoglycemic effect of Ganoderma lucidum polysaccharides (Gl-PS) in the normal fasted mice and its possible mechanism. METHODS: Normal fasted mice were given a single dose of Gl-PS 25, 50, and 100mg/kg by ip and the serum glucose was measured at 0, 3, and 6h after administration. Gl-PS 100mg/kg were also given by ip and the serum glucose and insulin levels were measured at 0min, 30min, 1h, 3h, 6h, and 12h Pancreatic islets were isolated and incubated with glucose 5.6mmol/L and different concentration of Gl-PS, the insulin content of islets and insulin release were examined. The islets fluorescent intensity of [Ca^2 ]i was also studied with a confocal microscope. Verapamil and egtazic acid were used to testify whether the insulin-releasing effect of Gl-PS was mediated by its ability to raise the Ca^2 influx. RESULTS: Gl-PS dose-dependently lowered the serum glucose levels at 3h and 6h after administration. Gl-PS 100mg/kg raised the circulating insulin levels at 1h after administration. In vitro, Gl-PS had no effect on islets insulin content, but it stimulated the insulin release after incubation with glucose 5.6mmol/L. Confocal microscope showed that Gl-PS 100mg/L had the capacity toraise the [Ca^2 ]i. The insulin-releasing effect of Gl-PS was inhibited by verapamil/egtazic acid. CONCLUSION:Gl-PS possesses the hypoglycemic effect on normal mice; one mechanism is through its insulin-releasing activity due to a facilitation of Ca^2 inflow to the pancreatic β cells.     

Diagnosis of the multiple effect of selenium nanoparticles decorated by Asteriscus graveolens components in inhibiting HepG2 cell proliferation

Using plant bio-components for Designing green metal nanoparticles was considered as one of the most important methods in nanomedical application field due to their eco-friendly, cheap source, easily obtainable and having a high detection result. In this report, we fabricated eco-friendly engineering and cost-effective technique for green selenium nanoparticles from 0.01 M H₂SeO₃ solution using Asteriscus graveolens leaves extract as reducing and a capping agent at ambient temperature. Spectral techniques have been used to identify the formatted Selenium nanoparticles such as UV–Vis, pH, XPs, FT-IR, XRD, LDS, Z.P, EDS, TEM and AFM spectroscopy, which showed a size of 20 nm with spherical shape. Herein, the multi-effect of decorated Se-NPs surface have been evaluated, firstly on the hemolysis that showed completely hemocompatibility. Cytotoxicity assay showed that Se-NPs have a high selective effect on the HepG2 apoptosis and which proved by phase-contrast microscopy. Furthermore, the effect of nanoparticles on the action of the mechanism internal revealed that Se-NPs significantly and rapidly increased the level of reactive oxygen species and lipid peroxidation, while caused decreased the potential of mitochondrial membrane and glutathione level, which they together responsible on regulating the HepG2 cells fate. Furthermore, Flow cytometry analysis gave high values about S and G2/M phases of cell cycle resulting from Se-NPs effectiveness. In the end, with all the recorded information that has been measured in this study, this report provides a suitable and effective pathway for the green fabrication of Se-NPs decorated by biomolecules having high anticancer inhibited.     

Antitumor and anti-angiogenic activity of Ganoderma lucidum polysaccharides peptide

AIM: To investigate the antitumor and anti-angiogenic activity of Ganoderma lucidum polysaccharides peptide (GLPP). METHODS: Antitumor effect of GLPP was observed in tumor-bearing mice in vivo. At the same time,the effects of GLPP on proliferation of tumor cells and human umbilical cord vascular endothelial cell (HUVEC) were detected by MTr assay in vitro. Subsequently, spleen lymphocytes proliferation of nude mice was stimulated by LPS or ConA. To investigate the anti-angiogenic effect of GLPP, GLPP 80 μg per disc and GLPP-treated serum 10μL per disc were added to the chick chorioallantoic membrane (CAM) respectively in vivo. RESULTS: GLPP 50, 100, and 200 mg/kg inhibited growth of Sarcoma 180 in BALB/c mice markedly by 35.2 %, 45.2 %, and 61.9 %,respectively. GLPP which was directly added to the cultured medium did not inhibit PG cell proliferation in vitro;but GLPP-treated serum 50, 100, 200 mg/kg potently inhibited PG cell proliferation by 22.5 %, 26.8 %, and 30.3 %,respectively; and reduced the xenograft (human lung carcinoma cell PG) in BALB/c nude mice greatly in vivo by 55.5 %, 46.0 %, and 46.8 %, respectively. Lymphocytes proliferation of nude mice could be stimulated by LPS 5 mg/L but not by ConA 2.5 mg/L, indicating that GLPP could not promote the T lymphocyte proliferation and neutral red phagocytosis of peritoneal macrophages of nude mice. The CAM assay showed that GLPP and GLPP-treated serum had anti-angiogenic effect. GLPP (1, 10, and 100 mg/L) inhibited HUVEC proliferation in vitro with the inhibitory rate of 9.4 %, 15.6 %, and 40.4 %, respectively. CONCLUSION: GLPP has antitumor and antiangiogenic activity. The anti-angiogenesis of GLPP may be a new mechanism underlying its anti-tumor effects.     

灵芝孢子多糖两纯组分的单糖组成研究

目的：对灵芝孢子多糖的2个纯多糖组分GLPSl和GLPS3的单糖组成进行分析。方法：将灵芝孢子多糖原料精制，SephacrylS一400SF型凝胶分离纯化得2个主要组分GLPSl和GLPS3，经酸水解后TLC法和HPLC法鉴别单糖组成，改良的1一苯基一3一甲基一5一吡唑啉酮（PMP）柱前衍生化HPLC法分析单糖组成比例，同时采用LC—MS联用技术鉴定单糖PMP衍生化产物。结果：GLPSl的单糖组成为D一甘露糖：D一葡萄糖：D一半乳糖=1：9．228：1．474；GLPS3的单糖组成为D一甘露糖：D．葡萄糖：D．半乳糖=1：15．900：3．686。结论：灵芝孢子多糖二纯组分均含D一甘露糖，D．葡萄糖和D一半乳糖，仅各单糖的比例不同。     

灵芝多糖对肿瘤细胞与内皮细胞相互作用的影响

目的研究灵芝多糖(GlPS)抗肿瘤作用及其机制。方法通过相差显微镜和间接免疫荧光的方法鉴定了人脐静脉内皮细胞(HUVECs)。采用MTT法检测GlPS对人前列腺癌细胞(PC-3M)和HUVECs增殖的影响。检测GlPS对PC-3M细胞与HUVECs粘附的影响。采用transwell双层小室观察GlPS对PC-3M迁移穿过单层HUVECs的影响。结果GlPS对PC-3M无直接细胞毒作用,对HUVECs增殖无显著性作用。GlPS能够减少粘附和迁移穿过单层内皮细胞的肿瘤细胞数目。结论灵芝多糖通过抑制肿瘤细胞粘附并迁移穿过内皮细胞发挥其抗肿瘤作用。     

灵芝多糖肽对小鼠巨噬细胞自由基的清除作用

目的 :研究灵芝多糖肽对小鼠腹腔巨噬细胞自由基的清除作用。方法 :用四氧嘧啶、叔丁基氢过氧化物 (tBOOH)为氧化剂 ,分别在小鼠体内、体外损伤小鼠腹腔巨噬细胞 ,以DCHF DA为荧光指示剂 ,用共聚焦显微镜观察巨噬细胞的荧光变化 ,并用共聚焦显微镜作时间系列扫描 ,观察巨噬细胞荧光的动态变化。结果 :四氧嘧啶 (75mg·kg-1,iv)、叔丁基氢过氧化物 (7.76× 10 -5mol·L-1)可造成巨噬细胞的氧化损伤 ,使荧光密度增加 ,灵芝多糖肽可减轻其损伤 ,使荧光密度减少。时间系列扫描显示 :随时间改变 ,灵芝多糖肽可减少静息状态下小鼠腹腔巨噬细胞荧光密度 ,也可减少由PMA(5 0nmol·L-1)诱导的呼吸爆发状态下小鼠腹腔巨噬细胞荧光密度。结论 :灵芝多糖肽具有抗氧化作用 ,对小鼠腹腔巨噬细胞自由基有清除作用     

灵芝多糖对体外树突状细胞诱导细胞毒性T-淋巴细胞的调节作用(英文)

目的:研究灵芝多糖(Gl-PS)在抗原提呈阶段对体外培养树突状细胞(DC)诱导特异性细胞毒性T-淋巴细胞(CTL)功能的调节及其机制。方法:体外培养小鼠骨髓来源DC经P815肿瘤细胞冻融抗原冲击致敏,并与不同浓度Gl-PS(0.8,3.2,或12.8 mg/L)共培养。成熟DC与脾淋巴细胞共培养诱导P815特异性CTL生成。第5天收集各组悬浮细胞及培养上清,采用乳酸脱氢酶法比较CTL特异性杀伤活性;RT-PCR法测定T扰素γ(IFNγ)及颗粒酶B mRNA在CTL的表达;ELISA或Western blot法检测IFNγ或颗粒酶B蛋白的表达。结果:3种浓度的Gl-PS均可增高培养上清中释放的LDH活性(P<0.01);增加CTL表达IFN7及颗粒酶B mRNA(IFNγ:Gl-PS 12.8 mg/L组与RPMI-1640组,P<0.05;颗粒酶B:P<0.01);促进CTL培养上清中IFNγ蛋白生成(P<0.05)以及CTL表达颗粒酶B蛋白(Gl-PS 12.8 mg/L组与RPMI-1640组,P<0.05)。结论:Gl-PS可在抗原提呈阶段促进P815肿瘤冻融抗原冲击致敏DC所诱导的特异性CTL的杀伤活性,其机制可能是通过IFNγ及颗粒酶B途径的调节。     

灵芝多糖诱导人肝癌细胞HepG2凋亡的研究

目的：研究灵芝多糖（GLPS）与化疗药氟尿嘧啶联合使用诱导人肝癌细胞HepG2凋亡作用。方法：MTT法测定细胞毒作用,流式细胞术检测细胞凋亡率,比色法测定Caspase-3、Caspase-8酶活性,Western—Blotting法检测细胞色素C、Caspase-3、Caspase-8蛋白表达情况。结果：灵芝多糖与氟尿嘧啶联合使用具有显著的细胞毒作用;流式细胞术检测,随着GLPS浓度增加,细胞凋亡率升高,Caspase-3、Caspase-8酶活性亦增强;Western-Blotting检测发现GLPS作用后细胞色素C、Caspase-3、Caspase-8蛋白表达不同程度增加。结论：灵芝多糖与氟尿嘧啶联合使用可通过引起细胞色素C的释放,活化Caspase诱导肝癌细胞凋亡。     

灵芝多糖肽对氧自由基损伤巨噬细胞的保护作用

目的:研究灵芝多糖肽(GLPP)在离体和整体水平对氧自由基(ROS)(tBOOH为氧化剂)损伤巨噬细胞的保护作用.方法:以tBOOH为氧化剂损伤小鼠腹腔巨噬细胞,以MTT法分析小鼠巨噬细胞存活率,在光镜和电子显微镜下观察细胞的形态改变.结果:GLPP50,100,200 mg/kg腹腔注射5天,能抑制巨噬细胞膜样变性和坏死,细胞存活率提高.在培养的巨噬细胞中加入 GLPP 3.125,12.5,50,200 mg/L,产生相似的保护作用.电镜观察发现,GLPP(100mg/kg)腹腔注射5天可保护细胞器如线粒体免受tBOOH的损伤.结论:GLPP有显著的清除氧自由基和抗氧化作用.     

Effects of Ganoderma lucidum polysaccharides on proliferation and cytotoxicity of cytokine-induced killer cells

AIM: To study the effects (and the mechanisms thereof) of Ganoderma lucidum polysaccharides (Gl-PS) on the proliferation and the anti-tumor activity of cytokine-induced killer (CIK) cells, and to make use of CIK cells as a means to investigate the interactions between Gl-PS and cytokines. METHODS: CIK cells were prepared by using the standard protocol as a positive control. Experimental groups also underwent the standard protocol, except that Gl-PS (400 mg/L or 100 mg/L) was added and the dose of anti-CD3 and interleukin-2 they received was reduced by 50% and 75%, respectively. For negative controls, Gl-PS in the experimental protocol was replaced with soluble starch or methylcellulose (400 mg/L or 100 mg/L). CIK cell proliferation, cytotoxicity, and phenotype were determined by using the Trypan blue exclusion method, MTT assay, and flow cytometry. RESULTS: By synergizing cytokines, Gl-PS (400 mg/L or 100 mg/L) could decrease the amount of cytokine in lymphokine activated killer (LAK) cells and CIK cells culture, but had no significant effect on the proliferation, cytotoxicity, or phenotype of LAK cells, or CIK cells induced by cytokines at higher doses alone, in which CIK cells expanded about 80-fold and the main effectors, CD3+NK1.1+ cells, expanded by more than 15%. The cytotoxicity of CIK cells in experimental groups was 79.3%+/-4.7%, 76.9%+/-6.8% versus the positive control 80.7%+/-6.8% against P815 (P>0.05) and 88.9%+/-5.5%, 84.7%+/-7.9% versus the positive control 89.8%+/-4.5% against YAC-1 (P>0.05). The activity of Gl-PS could mostly be blocked by anti-CR3. CONCLUSION: Gl-PS was shown to be a promising biological response modifier and immune potentiator. The effect of Gl-PS on CIK cells is possibly mediated primarily through complement receptor type 3.     

灵芝多糖对小鼠腹腔巨噬细胞活性氧自由基的影响

为进一步探讨灵芝多糖 ( GLP)免疫调节作用机理 ,采用激光扫描共聚焦显微镜技术 ,动态监测灵芝多糖均一体 GLB7对小鼠腹腔巨噬细胞 ( M )活性氧自由基含量的影响 .以 GLB7( 2 0 mg· L-1)刺激 M ,发现探针 DCHF- DA的荧光强度明显下降 ,与静息水平相比 ,平均下降 ( 36± 6) % ( n=6,P<0 .0 5) ;同时还能拮抗 PMA引起的 M 呼吸爆发 ,荧光强度下降幅度为 ( 1 9± 4) % ( n=6,P<0 .0 5) .结果证明 :GLB7能减少 M 内活性氧自由基的生成 ,具有清除活性氧的作用 ,这也是 GLB7产生免疫增强和抗衰老作用的重要途径 .     

灵芝多糖肽对自由基所致的腹腔巨噬细胞早期损伤的影响

目的　以膜电位方法进一步研究灵芝多糖肽(GLPP)对自由基损伤的巨噬细胞线粒体的影响。方法　以叔丁基氢过氧化物(tBOOH)为氧化剂,建立小鼠巨噬细胞体外氧化损伤模型,以罗丹明1 2 3(Rh1 2 3 )为荧光染色剂,以激光共聚焦显微镜检测细胞线粒体膜电位改变。结果　tBOOH氧化剂可损伤线粒体膜,使线粒体膜电位降低,GLPP体内( 1 0 0mg·kg- 1 ig 5d)及体外给药(GLPP 1 0mg·L- 1)均可对抗tBOOH自由基损伤,可使因自由基损伤而降低的巨噬细胞线粒体膜电位恢复。结论　GLPP体内体外给药可减轻自由基损伤,使损伤的线粒体膜电位恢复     

灵芝多糖对大鼠胰岛细胞分泌胰岛素功能的影响

目的 :探讨灵芝多糖 (Gl PS)对胰岛细胞分泌胰岛素的影响。方法 :分离大鼠胰岛细胞 ,分别观察在葡萄糖为 5 .6和 1 6.7mmol·L- 1 时 ,Gl PS对胰岛细胞分泌胰岛素的影响。进一步加入不同的抑制剂观察Gl PS对胰岛细胞分泌胰岛素的影响。采用Westernblotting观察Gl PS对胰岛细胞葡萄糖转运蛋白 2 (GLUT2 )表达的影响。结果 :在 5 .6mmol·L- 1葡萄糖时 ,1 0 0mg·L- 1 的Gl PS可明显促进胰岛细胞胰岛素的分泌 ;当葡萄糖浓度为 1 6.7mmol·L- 1 时 ,5 0和 1 0 0mg·L- 1 的Gl PS均可明显促进胰岛细胞胰岛素的分泌。维拉帕米或维拉帕米 +EGTA均可显著抑制 5 .6和 1 6.7mmol·L- 1 葡萄糖时胰岛素的分泌 ,维拉帕米只能部分抑制Gl PS的促胰岛素分泌作用 ,而维拉帕米 +EGTA则可完全抑制Gl PS的这种作用。在葡萄糖浓度为 5 .6和 1 6.7mmol·L- 1 时 ,Gl PS 5 0和 1 0 0mg·L- 1 均能明显促进胰岛细胞GLUT2的蛋白表达。结论 :Gl PS可能通过促进胰岛细胞GLUT2蛋白的表达从而有助于葡萄糖转运入B细胞 ,促进葡萄糖的代谢 ,引起胰岛细胞外Ca2 + 内流而起到促胰岛素释放的作用     

Antioxidant activities of Ganoderma lucidum polysaccharides and their role on DNA damage in mice induced by cobalt-60 gamma-irradiation

In this study, the radio-protective effects of Ganoderma lucidum polysaccharides (GLP) were investigated in a mouse animal model exposed to (60)Co gamma-irradiation. Each of three batches of mice were divided into five groups (negative control, positive gamma irradiated control, and low, middle and high dosage GLP groups). Different batches of animals were used to evaluate the impact of GLP on peripheral white blood cell count, immune organ index; DNA damage, lipid peroxidation; micronuclei formation, and nucleated cell count in bone marrow induced by (60)Co gamma-irradiation. DNA strand-break and micronuclei frequency were significantly reduced and glutathione peroxidase activity and nucleated cell count in bone marrow were significantly increased by GLP treatment in a dose-dependent manner. GLP intervention also increased the activity of superoxide dismutase and decreased the level of malondialdehyde in middle and high GLP treatment groups. No adverse effects were observed on peripheral white blood cells and immune organ or body weight in either the control groups or GLP treated gamma exposed mice. These findings suggest that GLP possesses marked antioxidant capacity which plays an important role in the prevention of radiation damage in mice induced by (60)Co gamma-irradiation.     

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.     

灵芝多糖拮抗环孢素A,氢化可的松及抗肿瘤药的免疫抑制作用

灵芝多糖在50,100和200μg·ml~1时可完全拮抗环孢素A,丝裂霉素C,氟尿嘧啶和阿糖胞苷对小鼠混合淋巴细胞反应(MLR)的轻度抑制(抑制50％以下)作用,部分拮抗氢化考的松对MLR的严重抑制(抑制50％以上)作用。     

灵芝多糖引起的小鼠脾细胞核DNA，RNA含量及核质比的变化

灵芝多糖可引起体外培养的小鼠脾细胞核ＤＮＡ，ＲＮＡ含量显著增加，细胞内超微结构改变，细胞质和细胞核的平均截面积明显增加，核质比呈下降趋势，提示灵芝多糖诱导脾细胞ＤＮＡ和蛋白质合成的同时，促进细胞增殖。     

Identification of polysaccharides in pharmaceuticals by capillary gas chromatography

A sensitive method for the identification of polysaccharides in pharmaceuticals is described. Polysaccharides are isolated by gel filtration and subsequently hydrolysed. The monomeric carbohydrates obtained are transformed into oxime-trimethylsilyl derivatives and analysed by capillary gas chromatography. Profiles of 13 different natural or semi-synthetic polysaccharides are discussed. The profiles of the hydrolysis products can be used to identify the polysaccharides mentioned above. Possible interferences by other polymers are given. The method can be used to identify most polysaccharides used as pharmaceutical adjuvants.