多脂鳞伞菌丝体多糖抗肿瘤活性研究

多脂鳞伞[Pholiota adiposa(Fr.)Quel.]菌丝体多糖溶液给S-180荷瘤小鼠连续灌胃10d后,测定对荷瘤小鼠抑瘤率、碳粒廓清指数、小鼠外周血肿瘤坏死因子(TNF-α)和白介素-2(IL-2)含量的影响。结果发现,多脂鳞伞菌丝体多糖对荷瘤小鼠肿瘤的抑瘤率为39.54%,并可提高小鼠碳粒廓清指数,血清中IL-2和TNF-α的含量也明显提高。结果提示,多脂鳞伞菌丝体多糖抑制荷瘤小鼠的肿瘤生长可能与增强宿主的免疫功能相关。     

冬虫夏草菌丝体多酚提取及对3种癌细胞抑制作用研究

以冬虫夏草的菌丝体为原料,提取溶剂为乙醇,采用Folin-Ciocalteu法测定冬虫夏草菌丝体中多酚的含量,目的在于研究乙醇浓度、料液比、提取温度和提取时间对多酚提取率的影响。以单因素试验为前提,以正交试验来指导优化提取参数,测定了所得多酚含量及其对Caco-2结肠癌细胞、Hep G-2肝癌细胞、MCF-7乳腺癌细胞的抑制作用。研究结果表明,在冬虫夏草菌丝体的提取工艺中,最佳的提取工艺条件为:温度为70℃,提取时间60 min,乙醇的浓度70%,料液比(g∶mL)1∶35;冬虫夏草菌丝体中提取的粗多酚对试验中3种癌细胞都表现出一定的抑制作用,当菌丝体粗多酚的浓度为100μg·mL~(-1)时,对3种癌细胞抑制率达到最高,分别为72.12%(Caco-2结肠癌细胞)、83.67%(HepG-2肝癌细胞)和58.78%(MCF-7乳腺癌细胞)。     

蝙蝠蛾拟青霉菌丝体石油醚提取物抗肿瘤活性

选用腹水型H22肝癌荷瘤小鼠,对蝙蝠蛾拟青霉(Paecilomyces hepiali)菌丝体石油醚提取物进行体内抗肿瘤作用的研究.结果表明,800 mg/kg蝙蝠蛾拟青霉菌丝体石油醚提取物具有明显的抑瘤作用,抑瘤率达到59.9%,对H22肝癌荷瘤小鼠的免疫器官指数具有一定程度的影响,但对其血清中白细胞介素2(interleukin-2,IL-2)含量的影响不显著.     

金顶侧耳多糖体外抗肿瘤作用的研究

通过深层发酵获得金顶侧耳菌丝体。用水提法分别提取金顶侧耳菌丝体多糖、胞外(过滤液)多糖和全液(菌丝体 发酵液)多糖。用MTT比色法测定金顶侧耳多糖体外对小鼠S180癌细胞及人结肠低分化腺癌细胞的抑制率。结果表明,金顶侧耳胞外多糖对体外培养的S-180癌细胞有抑制作用,全液多糖和菌丝体多糖无抑制作用。这3种多糖体外对人结肠低分化腺癌细胞有抑制作用,其中金顶侧耳胞外多糖抑制率最高,全液多糖次之,菌丝体多糖最低。     

桑黄菌丝体对小鼠肉瘤S_(180)的抑制作用

建立荷瘤(S180)小鼠模型,用桑黄(Phellinus igniarius)菌丝体灌胃(低、中和高剂量组分别为0.25、0.5和1.0g/kg/d)至模型组肿瘤直径达1cm停止给药,测定抑瘤率和各脏器指数。结果表明:阳性对照组(顺铂3mg/kg/d)显著抑制小鼠体重的增加,而桑黄菌丝体3个剂量组对小鼠体重的增加没有显著的抑制作用;低、中和高3个剂量组抑瘤率分别为18.3%、29.5%和40.7%。桑黄菌丝体对脾脏指数与胸腺指数的影响小于阳性对照药物顺铂。     

裂蹄针层孔菌发酵菌丝体对人肝癌HepG-2细胞的抑制作用

研究了裂蹄针层孔菌(Phellinus linteus)发酵菌丝体对人肝癌Hep G-2细胞的抑制作用。采用MTT比色法测定不同浓度发酵菌丝体对肝癌细胞存活率的影响;建立荷瘤Hep G-2裸鼠模型,检测不同剂量裂蹄针层孔菌发酵菌丝体对肿瘤的抑制作用。MTT法观察结果显示,裂蹄针层孔菌发酵菌丝体可抑制人肝癌Hep G-2细胞的增殖,且呈现剂量效应关系,高剂量(960 mg·L~(-1))能显著抑制肿瘤细胞的增殖,最佳抑制时间为24 h;裂蹄针层孔菌发酵菌丝体中剂量(500 mg/kg·d)、高剂量(1 000 mg/kg·d)对荷瘤Hep G-2裸鼠有显著的抑瘤作用,抑瘤率分别为40.42%、35.06%。裂蹄针层孔菌发酵菌丝体对人肝癌Hep G-2肿瘤细胞有较好的抑制作用。     

冬虫夏草及其菌丝体研究进展

本文从冬虫夏草及其菌丝体的药化、药理(含临床应用)、冬虫夏草的人工栽培及菌丝体的深层培养诸方面概述了近年来的研究工作,对今后的研究工作提出建议与设想。     

冬虫夏草甘露醇含量的高效液相色谱法测定

目的:用高效液相色谱法(High Performance Liquid Chromatography,HPLC)测定冬虫夏草菌丝体中甘露醇的含量,并检验冬虫夏草菌丝体中存在的果糖是否会对比色法测定甘露醇含量产生影响。方法:冬虫夏草菌丝体样品中的甘露醇采用超声辅助提取,用HPLC法测定提取液中甘露醇、果糖的含量。结果:高效液相色谱法测得甘露醇含量为4.41%,平均回收率为102.41%,相对标准偏差(Relative Standard Deviation,RSD)为2.521%。实验HPLC条件下,相同浓度的甘露醇和果糖的出峰面积大致相同。而冬虫夏草水提液中的果糖,相对于甘露醇,峰面积极其微小,其含量不到甘露醇含量的3%。结论:HPLC法特异性强,所受干扰少,适合对组成复杂体系中甘露醇含量的测定。HPLC法测定虫草菌丝体中果糖含量结果表明,果糖对甘露醇比色法测定的干扰极其微小。     

茶树菇菌丝体孢内多糖的抗氧化活性

为探讨茶树菇菌丝体孢内多糖的体内抗氧化活性，分别用水提、酸提、碱提菌丝体粗多特对小鼠灌胃给约（2000mg/kg），检测小鼠血清中超氧化物歧化酶（SOD）的活性以及肝组织中丙二醛（MDA）的含量。结果表明，与对照组相比，菌丝体多糖灌胃各组小鼠血清SOD活力显著增高（P〈0．05），同时肝组织中MDA的含量显著降低（P〈0．05）。     

鳞盖红菇菌丝体多糖对H22荷瘤小鼠Bcl-2和P53表达的影响

用鳞盖红菇(Russula lepida)菌丝体多糖连续灌胃H22荷瘤小鼠10 d.与阴性对照组相比,多糖组的瘤重降低,Bcl-2和突变型P53的表达下调,表明鳞盖红菇菌丝体多糖对H22荷瘤鼠有抑瘤作用,可以抑制H22肝癌细胞的增殖.     

灵雷菌质提取物体内抗肿瘤作用的实验研究

采用小鼠肉瘤(S180)移植性肿瘤模型,观察灵雷菌质乙酸乙酯部位、正丁醇部位、石油醚部位和水部位提取物的肿瘤抑制作用,并测定小鼠的胸腺指数及脾脏指数。结果表明所有的提取物都呈现出明显的抑瘤效果,抑瘤率均大于35%,其中部分正丁醇部位的抑瘤率达到了68.88%。抑瘤效果由弱至强的顺序为石油醚部位<水部位<乙酸乙酯部位<正丁醇部位,而胸腺指数与脾脏指数则显示只有乙酸乙酯部位组有较明显的免疫抑制效果。双向性固体发酵可降低雷公藤的毒性,尤其正丁醇部位和水部位的毒性可能较弱。     

Nucleostemin specific RNAi influences cell proliferation in HeLa cells in vitro and in vivo

AIM: To examine Nucleostemin (NS) expression in tumor cells, and observe the effect of NS specific RNA interference on the cell proliferation in Hela cells. METHODS: Total RNA was extracted from 6 kinds of cultured tumor lines, the NS expression level was measured by RT-PCR and Northern blot. An NS-specific siRNA expression vector was constructed to transfect HeLa cell (NS-siRNA-HeLa), and the proliferation of the cell was observed. RESULTS: NS was highly expressed in 6 kinds of tumor cells. NS expression level in the NS-siRNA-HeLa cells was remarkably reduced, and the percentage of G0/G1 cells increased. The neoplasm forming ability in nude mice by the NS-siRNA-HeLa cells was decreased. CONCLUSION: NS is highly expressed among tumor cells. NS-specific siRNA inhibits the entry of the cell cycle into the S phase, and remarkably reduces the proliferation ability of HeLa cells in vitro and in vivo.     

The action of recombinant human nucleoside diphosphate kinase A (rhNDPK-A) on the growth of S180,H22,Lewis and H460 tumors in vivo

AIM:To study the action of nucleoside diphosphate kinase A (NDPK-A) on the growth of S180,H22,Lewis and H460.METHODS:S180 or H22 cell (5×106) were inoculate subcutaneously into the right armpit of 85 Kunming mice,which were randomized into 8 groups.Lewis lung carcinoma cells (2×105) were inoculate subcutaneously into the right armpit of 85 C57BL/6 mice,which were randomized as Kunming mice.From the 2nd day,the treated groups were given different dose of rhNDPK-A once a day for 8 days (for S180 or H22 by iv) or for 10 days (for Lewis by ip),and the control group was given physiological saline only.H460 tissue pieces about 1.5 mm×1.5 mm×1.5 mm each were inoculated subcutaneously into the armpit of 38 Balb/c/neu mice.After the volume of xenograft become 100 mm×100 mm×100 mm,the nude mice were randomized into 5 groups and given different dose of rhNDPK-A once a day for 17 days.2 days after above treatments,the mice were killed and dissected.The knubs were peeled off and weighted.RESULTS:The growth of S180,H22 and H460 were inhibited by rhNDPK and the growth of H22 was inhibited by rhNDPK at dose of 20 mg/kg combined with cisplatin (0.5 mg/kg).But the growth of Lewis lung cancer was not inhibited.CONCLUSION:rhNDPK-A inhibited the growth of S180,H22 and H460.rhNDPK-A (20 mg/kg) potentiated the antitumor action of cisplatin on H22.     

Angiogenesis and its regulation mechanism in S180 transplanted tumor of mice

AIM: To investigate the angiogenesis in the process of sarcoma 180 (S180) tumor transplantation and changes of regulator factors, and explore the possible mechanism. METHODS: The S180 transplanted tumor in the Km mouse was used to detect the tumor angiogenesis by immunohistochemical examination of FⅧ. The levels of VEGF (V) and endostatin (E) in serum and the homogenate of tumor tissue were measured by ELISA and EIA, and the correlation between tumor weight and microvessel count (MVC) and morphology in tumor was also analyzed by multiple ANNOVA method. RESULTS: MVC, the relative count of total vessels and relative total vessel area increased with the development of transplanted S180. VEGF level in tumor tissue were higher at the 10th and 15th day than the 5th day after tumor transplantation. Endostatin in the tumor tissue and serum both reached the highest level at the 15th day, V/E ratio did not changed in this process. Furthermore, MVC, average vessel area and relative total area had a significant correlation with tumor weight. CONCLUSION: MVC increases in the development of S180 transplantation tumor and is related with the tumor weight; the positive regulator of angiogenesis in the tumor tissue is up-regulated during tumor growth, and the regulators in the tumor tissue maintains a relative balance.     

Effect of mutant β-catenin gene on the proliferation of hepatocytes

AIM:The β-catenin is a key molecule in the Wnt signal pathway, which plays a critical role in normal development and tumorigenesis. However, the mechanisms of the β-catenin on the cell growth control are still not completely defined. The aim of this study was to test the hypothesis that the mutant β-catenin may regulate the hepatocyte proliferation. METHODS: The immortalized murine hepatocyte cell line, AML12, was used for this study. A plasmid that contain mutant β-catenin S33Y was transfected into the AML12 cells and a stable cell line AML12S33Y was established. The cell growth property of this cell line and the parental cell were compared by flow cytometry analysis and direct cell count. The cells were also tested for the ability to form soft agar colonies, and the ability to form tumors in the severe immune deficient mice (SCID). RESULTS:1. The mutant β-catenin containing cell line AML12S33Y has higher proliferating index compared with the parental AML12 cells (P<0.01), suggesting that mutant β-catenin promotes cell growth. 2. The mutant β-catenin cells formed small colonies in soft agar after 4 weeks of culture, but did not generate tumor in SCID mice. CONCLUSION:The mutant β-catenin promotes liver cell growth.     

Study on changes of ET、NO level and tumor weight of mice bearing S180 model and their relationships

AIM: To approach the changes of ET and NO levels of mice bearing sarcoma 180 (S₁₈₀ ) during different period(5、10 and 15 days) and the relationship between ET、NO and tumor’s development METHODS: Adopting mice bearing S₁₈₀ as the models, the levels of ET and NO₂^-/NO₃^- in serum were detected, and the weight of isolated tumors was measured On the basis of the regulation of these changes, their relationships were explored RESULTS: The levels of ET and NO₂^-/NO₃^- of mice bearing S₁₈₀ were higher than that of the control group (not bearing tumor) ( P< 0.05) Along with the development of the tumors, the levels of NO₂^-/NO₃^-and tumors weight both increased ( P< 0 05) ET also had an increasing tendency There was a positive correlation between the level of NO₂^-/NO₃^- and tumor weight ( r =0.995, P< 0.05) Ratio value of (NO₂^-/NO₃^-)/ET decreased at first and then increased CONCLUSION: ET and NO have links with the development of S₁₈₀ There may be cooperation between ET and NO during the development of tumor.     

Knockdown of RRM2 gene by siRNA inhibits human breast cancer cell proliferation,migration and tumor growth in nude mice

AIM: To explore the effect of ribonucleotide reductase M2 (RRM2) gene knockdown by siRNA on the proliferation and migration of human breast cancer MCF-7 cells and the tumor growth in BALB/c nude mice. METHODS: The mRNA and protein expression leves of RRM2 in human breast cancer cell line MCF-7 and human normal breast cell line MCF-10A were determined by real-time PCR and Western blotting. siRNA-RRM2 was constructed and transfected into MCF-7 cells at different time points and different concentrations. The silencing efficiency of RRM2 gene was detected by real-time PCR. The cell proliferation was measured by CCK-8 assay. The migration was observed using Transwell cell migration system. The effect of siRNA-RRM2 on the tumor growth was determined in nude mice. RESULTS: The mRNA and protein levels of RRM2 were higher in MCF-7 cells than those in MCF-10A cells. siRNA-RRM2 down-regulated the expression of RRM2 in MCF-7 cells in a time-and concentration-dependent manner. The results of CCK-8 assay showed that siRNA-RRM2 inhibited the proliferation ability of MCF-7 cells, but not that of MCF-10A cells. The results of Transwell assay indicated that siRNA-RRM2 inhibited the migration ability of MCF-7 cells. siRNA-RRM2 also inhibited the tumor growth in nude mice. CONCLUSION: RRM2 overexpression is associated with the breast cancer proliferation and migration. Suppression of RRM2 function is a potential therapeutic strategy for treating breast cancer.     

Regulation of octreotide on SSTR2 expression in hepatocellular carcinoma

AIM: To observe the regulation of octreotide (OCT) on the expression of somatostatin receptor 2 (SSTR2) in Bel7402 hepatocellular carcinoma (HCC) cells, and the inhibition effect of OCT on the growth of HCC. METHODS: The effect of OCT on proliferative ability of Bel7402 cells was observed by MTT assay. The cell form was observed by light invert microscope. The adhesive and invasive ability was detected by cell adhesion and migration experiments. The cell cycle, SSTR2 expression of 7402 cells were determined by immunofluorescence flow cytometry. Nude mice bearing xenografts in situ were treated with OCT or saline control for 7 weeks since tumor implantation. The immunohistochemistry for SSTR2 was performed. SSTR2 mRNA expression in cell line and xenografts was measured by semi-quantitative RT-PCR. RESULTS: After OCT treatment, the proliferative ability and cell form of 7402 cells didn't change significantly. The adhesive and invasive ability decreased significantly. The ratio of cells in resting state (G₀/G₁) increased, but no apoptosis peak was observed. The SSTR2 expression on 7402 cell membranes decreased significantly. SSTR2 expression in cell line of OCT group was higher than control group, but there was no significant difference between them. The mean tumor weight in mice given OCT was significantly lower than that in control group. SSTR2 immunostaining in tumor cells of treatment group showed stronger positivity, compared with control group. SSTR2 mRNA expression in xenografts after OCT treatment was significantly higher than that in control group. CONCLUSIONS: OCT inhibits the growth of HCC through SSTR2. SSTR2 is regulated by its ligand, the long-term OCT treatment increases the SSTR2 expression and enhances the effect of inhibiting HCC, however, short-term treatment may induces its desensitization and the decrease in anti-tumor effect.     

巴西菇麦角甾醇抗肿瘤活性及作用机理初探

采用小鼠S180移植瘤实验研究了巴西菇子实体中麦角甾醇的体内抗肿瘤活性,并通过巨噬细胞吞噬功能测定(中性红法)、脾淋巴细胞增殖功能检测(MTT法)、体外抑杀肿瘤细胞(MTT法)以及鸡胚绒毛尿囊膜(CAM)等试验初步探讨了麦角甾醇抗肿瘤活性的作用机理.结果表明,巴西菇子实体中提取的麦角甾醇在剂量10 mg·kg-1·d-...     

miR-509 regulates growth,invasion and migration of human hepatocellular carcinoma LM3 cells and survival of nude mice

AIM: To investigate the effect of microRNA-509 (miR-509) on the growth, invasion and migration of human hepatocellular carcinoma (HCC) LM3 cells and survival of tumor-bearing nude mice. METHODS: LM3 cells were transferred with miR-509 mimic and pcDNA Ras-related C3 botulinum toxin substrate 1 (pcRac1), and the expression of Rac1 was measured by Western blot. The relationship between miR-509 and Rac1 was determined by luciferase reporter assay. The invasion ability was determined by Transwell assay, and the migration ability was measured by wound healing assay. Xenograft model of HCC was established by subcutaneous injection with LM3 cells into nude mice. The survival rate of the mice were recorded and the protein level of Rac1 was determined by Western blot. RESULTS: miR-509 mimic inhibited the expression of Rac1 in the LM3 cells (P<0.05). pcRac1 attenuated the effect of miR-509 on Rac1. miR-509 also alleviated luciferase activity of wild Rac1 (P<0.05). Meanwhile, miR-509 mimic decreased the number of invasive LM3 cells and inhibited the migration of LM3 cells (P<0.05). In addition, over-expression of miR-509 up-regulated survival rate of model mice and decreased the protein level of Rac1 in the tumor tissue (P<0.01). CONCLUSION: miR-509 inhibits the invasion and migration of HCC cells and promotes the survival of tumor-bearing nude mice through inhibiting the expression of Rac1.