Objective To investigate th e anti-tumor effects of GeM10 by the natural killer(NK) cells activities and th e production of Interleukin-2 (IL-2) in peripheral blood mononuclear cells (PB MNCs). Methods Assay of hum...Objective To investigate th e anti-tumor effects of GeM10 by the natural killer(NK) cells activities and th e production of Interleukin-2 (IL-2) in peripheral blood mononuclear cells (PB MNCs). Methods Assay of human NK cells activities by dye reject ion assay in vitro and production of IL-2 in PBMNC by IL-2 bioassay with I L-2 dependent cell line CTLL2 and MTT colorometric method. Results GeM10 could significantly stimulate NK activities (60μg·mL -1 G eM10: 17.077±7.665, 120μg·mL -1 GeM10: 24.9±13.04; control: 7.72±4 .64, P< 0.05). GeM10 could up-regulate the production of IL-2 of PBMNCs in tumor patients(60μg·mL -1 GeM10: 2.965± 1.183; 120μg·mL -1 GeM10: 2.28±0.847; control: 1.792±0.823, P<0.05).Conclu si on The GeM10 not only can stimulate the NK activities but also increase the IL-2 production by PBMNCs in tumor patients. These findings indicate that the GeM10 may have promise as an anti-tumor drug and a biological response modi fier in clinic.展开更多
BACKGROUND: The microenvironment within solid tumors has often been shown to exhibit an acidic extracellular pH. Although the morphologic and functional differences in natural killer (NK) cells of the liver and spleen...BACKGROUND: The microenvironment within solid tumors has often been shown to exhibit an acidic extracellular pH. Although the morphologic and functional differences in natural killer (NK) cells of the liver and spleen have been reported previously under physiological conditions, the difference under acidic conditions is still unclear. This study was to investigate the differences in the morphological and functional characteristics between rat liver and spleen NK cells under normal and acidic conditions in vitro. METHODS: Liver and spleen NK cells were isolated and purified from Sprague-Dawley rats by density gradient centrifugation and the Dynabeads FlowComp TM Flexi system, and stimulated for 4 days with or without IL-2 or treated with low pH or control for different times. Morphology was examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), cell death and proliferation assays were performed by flow cytometry, IFN-γ production was tested by ELISA, and cytotoxic activity was evaluated by lactate dehydrogenase (LDH) release assay. RESULTS: Liver NK cells had significantly higher levels of cytotoxic activity than spleen NK cells under normal and acidic conditions, and the maximum difference was observed at pH 5.6. Further analysis revealed that the cytotoxic activity of NK cells was correlated with morphology, cell death, proliferative activity and IFN-γ production. By TEM, liver NK cells contained a greater number of electron-dense granules per cell at pH 5.6.Moreover, a modest elevation of cell death and reduction of proliferation of liver NK cells occurred within a range of 5.6-7.2. Interestingly, an acidic extracellular pH only marginally, and not significantly, suppressed IFN-γ production by liver NK cells. CONCLUSION: The sharp morphological and functional differences shown by the two types of NK cells in vitro indicate that liver NK cells are unexpectedly resistant to pH shock.展开更多
Objective: To study the enhancement of the immune functions and autologous tumor killing (ATK) activity by kappa selenocarrageenan (KSC) in mice bearing sarcoma 180. Methods: To measure the effects of KSC and/or Cy...Objective: To study the enhancement of the immune functions and autologous tumor killing (ATK) activity by kappa selenocarrageenan (KSC) in mice bearing sarcoma 180. Methods: To measure the effects of KSC and/or Cyclophosphamide (Cy) on natural killer (NK) activity, lymphokine activated killer (LAK) activity, the produc tion of interleukin 2 (IL 2), ATK activity and the growth of sarcoma 180 (S 180 ). Results: KSC promoted NK activity, LAK activity and ATK activity in vivo , increased IL 2 production at 40 mg/kg/d×9d. It also enhanced the antitumor action of Cy (20 mg/kg/d×9d) and offset the inhibition of Cy on immunocopetent cells. The ATK activity in splenocytes of S 180 bearing mice could be induced and increased by recombinant interleukin 2 (rIL 2) in vitro . Conclusion: KSC has an up regulating effect on the immune functions and ATK activity in tumor bearing mice. It can be used as a biological response modifier (BRM) in cancer biotherapy.展开更多
AIM： To evaluate the safety and clinical efficacy of a new immunotherapy using both α-Gal epitope-pulsed dendritic cells （DCs） and cytokine-induced killer cells. METHODS： Freshly collected hepatocellular carcino...AIM： To evaluate the safety and clinical efficacy of a new immunotherapy using both α-Gal epitope-pulsed dendritic cells （DCs） and cytokine-induced killer cells. METHODS： Freshly collected hepatocellular carcinoma （HCC） tumor tissues were incubated with a mixture of neuraminidase and recombinant αl,3-galactosyltrans- ferase （αI,3GT） to synthesize α-Gal epitopes on car- bohydrate chains of the glycoproteins of tumor mem- branes. The subsequent incubation of the processed membranes in the presence of human natural anti-Gal IgG resulted in the effective phagocytosis to the tumor membrane by DCs. Eighteen patients aged 38-78 years with stage 111 primary HCC were randomly chosen for the study; 9 patients served as controls, and 9 patients were enrolled in the study group.RESULTS： The evaluation demonstrated that the pro- cedure was safe; no serious side effects or autoimmune diseases were observed. The therapy significantly pro- longed the survival of treated patients as compared with the controls （17.1 ± 2.01 mo vs 10.1 ±4.5 mo, P = 0.00121）. After treatment, all patients in the study group had positive delayed hypersensitivity and robust systemic cytotoxicity in response to tumor lysate as measured by interferon-y-expression in peripheral blood mononuclear cells using enzyme-linked immunosorbent spot assay. They also displayed increased numbers of CD8-, CD45RO- and CD56-positive cells in the peripheral blood and decreased α-fetoprotein level in the se- rum. CONCLUSION： This new tumor-specific immunotherapy is safe, effective and has a great potential for the treat- ment of tumors.展开更多
背景与目的:结直肠癌的发生、发展涉及多个癌基因的激活和抑癌基因的失活,野生型R-脊椎蛋白3(R-spondin 3,RSPO3)在结直肠癌生长中的作用目前尚不清楚,本研究旨在探讨RSPO3对结直肠癌生长的影响并探索其潜在机制。方法:采用生物信息学分...背景与目的:结直肠癌的发生、发展涉及多个癌基因的激活和抑癌基因的失活,野生型R-脊椎蛋白3(R-spondin 3,RSPO3)在结直肠癌生长中的作用目前尚不清楚,本研究旨在探讨RSPO3对结直肠癌生长的影响并探索其潜在机制。方法:采用生物信息学分析RSPO3在结直肠癌及泛癌组织中的表达,分析结直肠癌中RSPO3表达与自然杀伤(natural killer,NK)细胞浸润、NK细胞激活分子表达的相关性。利用短发夹RNA(short hairpin RNA,shRNA)和慢病毒感染建立RSPO3敲减的SW480-RSPO3-KD细胞株、RSPO3过表达的HCT116-RSPO3-OE细胞株及相应的对照细胞株。采用细胞计数试剂盒-8(cell counting kit-8,CCK-8)检测体外各稳定转染细胞株的细胞增殖。采用流式细胞术分析各稳定转染细胞株的细胞周期、裸小鼠脾脏和移植瘤组织中NK细胞的比例。通过裸小鼠皮下移植瘤模型观察RSPO3敲减或过表达的结肠癌细胞在裸小鼠体内的生长。利用双荧光素酶报告基因系统检测RSPO3敲减或过表达对结肠癌Wnt基因转录活性的影响。结果:生物信息学分析显示,RSPO3在多种实体瘤肿瘤组织包括结直肠癌组织中的表达显著低于相应的癌旁组织。RSPO3敲减或过表达不影响体外SW480和HCT116结肠癌细胞的增殖(P>0.05)和细胞周期(P>0.05)。但在裸小鼠体内,与对照细胞相比,RSPO3敲减显著促进SW480细胞移植瘤的生长(260.2±162.4 vs 1311.7±570.1,P<0.05),而RSPO3过表达则显著抑制HCT116细胞移植瘤的生长(1549.0±241.2 vs 512.1±250.0,P<0.05)。流式细胞术分析发现,在荷移植瘤裸小鼠体内,RSPO3敲减显著减少了脾脏和移植瘤组织中NK细胞的比例(脾脏:6.42±0.94 vs 5.25±0.59,P=0.04;移植瘤:8.27±0.29 vs 6.48±1.48,P=0.04);而RSPO3过表达显著增加了脾脏和移植瘤组织中NK细胞的比例(脾脏:5.29±0.16 vs 7.02±0.49,P=0.01;移植瘤:6.39±0.39 vs 8.14±0.34,P<0.05)。癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据相关性分析显示,RSPO3表达与NK细胞表面标志物CD56(r=0.58,P<0.05)和CD16(r=0.64,P<0.05)的表达显著正相关,并与NK细胞激活标志物CD69(r=0.51,P<0.05)和KLRB1(r=0.37,P<0.05)的表达显著正相关。双荧光素酶报告基因实验结果显示,RSPO3敲减后Wnt荧光素酶活性下调(1.0±0.0 vs 0.45±0.09,P<0.05),而RSPO3过表达后Wnt荧光素酶活性上调(1.0±0.0 vs 1.75±0.14,P<0.05)。结论:RSPO3能在体内显著抑制结直肠癌移植瘤的生长,并能增加移植瘤组织中NK细胞浸润,RSPO3是一个潜在的结直肠癌的抑制基因。展开更多
文摘Objective To investigate th e anti-tumor effects of GeM10 by the natural killer(NK) cells activities and th e production of Interleukin-2 (IL-2) in peripheral blood mononuclear cells (PB MNCs). Methods Assay of human NK cells activities by dye reject ion assay in vitro and production of IL-2 in PBMNC by IL-2 bioassay with I L-2 dependent cell line CTLL2 and MTT colorometric method. Results GeM10 could significantly stimulate NK activities (60μg·mL -1 G eM10: 17.077±7.665, 120μg·mL -1 GeM10: 24.9±13.04; control: 7.72±4 .64, P< 0.05). GeM10 could up-regulate the production of IL-2 of PBMNCs in tumor patients(60μg·mL -1 GeM10: 2.965± 1.183; 120μg·mL -1 GeM10: 2.28±0.847; control: 1.792±0.823, P<0.05).Conclu si on The GeM10 not only can stimulate the NK activities but also increase the IL-2 production by PBMNCs in tumor patients. These findings indicate that the GeM10 may have promise as an anti-tumor drug and a biological response modi fier in clinic.
基金supported by grants from the National Natural Science Foundation of China (30671987 and 81000065)
文摘BACKGROUND: The microenvironment within solid tumors has often been shown to exhibit an acidic extracellular pH. Although the morphologic and functional differences in natural killer (NK) cells of the liver and spleen have been reported previously under physiological conditions, the difference under acidic conditions is still unclear. This study was to investigate the differences in the morphological and functional characteristics between rat liver and spleen NK cells under normal and acidic conditions in vitro. METHODS: Liver and spleen NK cells were isolated and purified from Sprague-Dawley rats by density gradient centrifugation and the Dynabeads FlowComp TM Flexi system, and stimulated for 4 days with or without IL-2 or treated with low pH or control for different times. Morphology was examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), cell death and proliferation assays were performed by flow cytometry, IFN-γ production was tested by ELISA, and cytotoxic activity was evaluated by lactate dehydrogenase (LDH) release assay. RESULTS: Liver NK cells had significantly higher levels of cytotoxic activity than spleen NK cells under normal and acidic conditions, and the maximum difference was observed at pH 5.6. Further analysis revealed that the cytotoxic activity of NK cells was correlated with morphology, cell death, proliferative activity and IFN-γ production. By TEM, liver NK cells contained a greater number of electron-dense granules per cell at pH 5.6.Moreover, a modest elevation of cell death and reduction of proliferation of liver NK cells occurred within a range of 5.6-7.2. Interestingly, an acidic extracellular pH only marginally, and not significantly, suppressed IFN-γ production by liver NK cells. CONCLUSION: The sharp morphological and functional differences shown by the two types of NK cells in vitro indicate that liver NK cells are unexpectedly resistant to pH shock.
文摘Objective: To study the enhancement of the immune functions and autologous tumor killing (ATK) activity by kappa selenocarrageenan (KSC) in mice bearing sarcoma 180. Methods: To measure the effects of KSC and/or Cyclophosphamide (Cy) on natural killer (NK) activity, lymphokine activated killer (LAK) activity, the produc tion of interleukin 2 (IL 2), ATK activity and the growth of sarcoma 180 (S 180 ). Results: KSC promoted NK activity, LAK activity and ATK activity in vivo , increased IL 2 production at 40 mg/kg/d×9d. It also enhanced the antitumor action of Cy (20 mg/kg/d×9d) and offset the inhibition of Cy on immunocopetent cells. The ATK activity in splenocytes of S 180 bearing mice could be induced and increased by recombinant interleukin 2 (rIL 2) in vitro . Conclusion: KSC has an up regulating effect on the immune functions and ATK activity in tumor bearing mice. It can be used as a biological response modifier (BRM) in cancer biotherapy.
基金Supported by Hong Kong Wang Kuan Cheng GrantInner Mongolia Stem Cell Grant, No. kjk10jhg
文摘AIM： To evaluate the safety and clinical efficacy of a new immunotherapy using both α-Gal epitope-pulsed dendritic cells （DCs） and cytokine-induced killer cells. METHODS： Freshly collected hepatocellular carcinoma （HCC） tumor tissues were incubated with a mixture of neuraminidase and recombinant αl,3-galactosyltrans- ferase （αI,3GT） to synthesize α-Gal epitopes on car- bohydrate chains of the glycoproteins of tumor mem- branes. The subsequent incubation of the processed membranes in the presence of human natural anti-Gal IgG resulted in the effective phagocytosis to the tumor membrane by DCs. Eighteen patients aged 38-78 years with stage 111 primary HCC were randomly chosen for the study; 9 patients served as controls, and 9 patients were enrolled in the study group.RESULTS： The evaluation demonstrated that the pro- cedure was safe; no serious side effects or autoimmune diseases were observed. The therapy significantly pro- longed the survival of treated patients as compared with the controls （17.1 ± 2.01 mo vs 10.1 ±4.5 mo, P = 0.00121）. After treatment, all patients in the study group had positive delayed hypersensitivity and robust systemic cytotoxicity in response to tumor lysate as measured by interferon-y-expression in peripheral blood mononuclear cells using enzyme-linked immunosorbent spot assay. They also displayed increased numbers of CD8-, CD45RO- and CD56-positive cells in the peripheral blood and decreased α-fetoprotein level in the se- rum. CONCLUSION： This new tumor-specific immunotherapy is safe, effective and has a great potential for the treat- ment of tumors.
文摘背景与目的:结直肠癌的发生、发展涉及多个癌基因的激活和抑癌基因的失活,野生型R-脊椎蛋白3(R-spondin 3,RSPO3)在结直肠癌生长中的作用目前尚不清楚,本研究旨在探讨RSPO3对结直肠癌生长的影响并探索其潜在机制。方法:采用生物信息学分析RSPO3在结直肠癌及泛癌组织中的表达,分析结直肠癌中RSPO3表达与自然杀伤(natural killer,NK)细胞浸润、NK细胞激活分子表达的相关性。利用短发夹RNA(short hairpin RNA,shRNA)和慢病毒感染建立RSPO3敲减的SW480-RSPO3-KD细胞株、RSPO3过表达的HCT116-RSPO3-OE细胞株及相应的对照细胞株。采用细胞计数试剂盒-8(cell counting kit-8,CCK-8)检测体外各稳定转染细胞株的细胞增殖。采用流式细胞术分析各稳定转染细胞株的细胞周期、裸小鼠脾脏和移植瘤组织中NK细胞的比例。通过裸小鼠皮下移植瘤模型观察RSPO3敲减或过表达的结肠癌细胞在裸小鼠体内的生长。利用双荧光素酶报告基因系统检测RSPO3敲减或过表达对结肠癌Wnt基因转录活性的影响。结果:生物信息学分析显示,RSPO3在多种实体瘤肿瘤组织包括结直肠癌组织中的表达显著低于相应的癌旁组织。RSPO3敲减或过表达不影响体外SW480和HCT116结肠癌细胞的增殖(P>0.05)和细胞周期(P>0.05)。但在裸小鼠体内,与对照细胞相比,RSPO3敲减显著促进SW480细胞移植瘤的生长(260.2±162.4 vs 1311.7±570.1,P<0.05),而RSPO3过表达则显著抑制HCT116细胞移植瘤的生长(1549.0±241.2 vs 512.1±250.0,P<0.05)。流式细胞术分析发现,在荷移植瘤裸小鼠体内,RSPO3敲减显著减少了脾脏和移植瘤组织中NK细胞的比例(脾脏:6.42±0.94 vs 5.25±0.59,P=0.04;移植瘤:8.27±0.29 vs 6.48±1.48,P=0.04);而RSPO3过表达显著增加了脾脏和移植瘤组织中NK细胞的比例(脾脏:5.29±0.16 vs 7.02±0.49,P=0.01;移植瘤:6.39±0.39 vs 8.14±0.34,P<0.05)。癌症基因组图谱(The Cancer Genome Atlas,TCGA)数据相关性分析显示,RSPO3表达与NK细胞表面标志物CD56(r=0.58,P<0.05)和CD16(r=0.64,P<0.05)的表达显著正相关,并与NK细胞激活标志物CD69(r=0.51,P<0.05)和KLRB1(r=0.37,P<0.05)的表达显著正相关。双荧光素酶报告基因实验结果显示,RSPO3敲减后Wnt荧光素酶活性下调(1.0±0.0 vs 0.45±0.09,P<0.05),而RSPO3过表达后Wnt荧光素酶活性上调(1.0±0.0 vs 1.75±0.14,P<0.05)。结论:RSPO3能在体内显著抑制结直肠癌移植瘤的生长,并能增加移植瘤组织中NK细胞浸润,RSPO3是一个潜在的结直肠癌的抑制基因。
