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Objective To observe the clinical significance and effect of dysfunction of dendritic cell( DC) in colorectal cancer with liver metastasis.Methods Peripheral blood were respectively collected from healthy adult donors (30 cases), preoperative and postoperative coloreatal cancer patients without liver metastasis (30 cases) , and 30 postoperative coloreatal cancer patients with liver metastasis from Jan 2008 to Jun 2010.Peripheral blood mononuclear cells were separated, GM-CSF( 1000 U/ml) , IL-4( 1000 U/ml) and TNF-α ( 1000 U /ml) were added into cell culture fluid to induce the mononuclear cells for mature dendritic cells.There were two subgroups, and in antigen processing subgroup the lysate of HT29 colonic carcinoma cells (100 μg/ml) were added into the cell culture fluid.The T lymphocytes from healthy adults were added into two subgroups by ratio of 1∶10 ( DCs∶ T cells) , cocultured for 7 days.The level of INF-γ and IL-10 in cell culture fluid was assayed with ELISA method.The optical density (OD) of CCK8 ans LDH was assayed with ELIASA to indirectly measure the reproductive activity and the killing efficacy of T lymphocytes.Results The IL-10 level in cocultured fluid of peripheral blood DCs in postoperative colorectal carcinoma patients with liver metastasis and T lymphocytes of healthy adults was significantly higher than that of preoperative patients of colorectal carcinoma and health adults without tumor antigenic stimulation(11.9±1.3) pg/ml vs.(29.6±9.7) pg/ml, (23.4±8.0) pg/ml, F =4.475, P ＜0.05).The IFN-γ level in cocultured fluid of peripheral blood DCs in postoperative colorectal carcinoma patients with liver metastasis and T lymphocytes of healthy adults was significantly higher than that of postoperative patients of colorectal carcinoma and healthy adults with or without tumor antigenic stimulation ( 34 ± 9) pg/ml vs.(26 ± 12 ) pg/ml, (24 ± 6) pg/ml, F = 5.206, P ＜ 0.05).The killing activity of healthy adults T lymphocytes induced by HT29 colonic carcinoma cells in postoperative colorectal carcinoma patients with liver metastasis was significantly higher than that of preoperative patients of colorectal carcinoma and healthy adults (30.6 ±8.6) pg/ml vs.(12.1 ±2.4) pg/ml, (14.9 ± 1.7) pg/ml, F =4.147, P ＜ 0.05).Conclusions T lymphocytes produce IL-10 when indued by DCs from patients with colorectal carcinoma under stimulation of tumor antigen leading to tumor immune escape and liver metastasis.The killing activity of T lymphocytes can be enhanced when stimulated by exogenous tmuor antigen.
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Objective To study the effect of brucine on the growth of a hepatocellular carcinoma cell line in vitro. Methods Brucine was added into a liver cancer cell line of SMMC-7721 in vitro, at drug concentration of brucine from 2. 5 μg/ml to 400 μg/ml. The inhibition rate of cell growth was measured by MTT technique after the cells were cultured for 72 hours. The protein and mRNA expression of PCNA,cyclin D1 and FAS were respectively assayed with Western blotting and fluorescent quantitation RT-PCR techniques at 24, 48, 72 h. Results The inhibition rate of liver cancer cell was near 100% when the brucine concentration was at 320 μg/ml. The protein and mRNA expression of FAS were of no significant difference at 24 h vs 48 h ( seperately F = 2. 547,1. 582, all P ＞ 0. 05 ), and significant difference existed at 24 h vs 72 h( seperately F = 1. 036, 1. 137, all P ＜ 0. 05 ). The protein and mRNA expression of PCNA,Cyclin D1 were of no significant difference between various time period( seperately PCNA F = 3.612,2. 174,3.029;Cyclin D1 F=2.361,2.915,1.976,all P＞0.05). Conclusions Brucine inhibits the growth of liver cancer cells, by inducing increased apoptosis of the cells probably through FAS overexpression.
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Objective To observe the therapeutic effect of brucine nanoparticles on hepatocellular carcinoma.Methods Brucine nanoparticles with block copolymer of carboxylation polyethylene glycol and polylactic acid were manufactured by ultraphonic emulsification.The effect brucine nanoparticles on growth of SMMC-7721 cell line was observed in vitro.The protein and mRNA levels of Fas were measured with Western blotting and FQ-PCR respectively after the brucine nanoparticles were added into cell culture fluid for 72 hours.Results The mean diameter,the carried drug rate and the entrapment rate of brucine nanopaticles was 146±96 nm,4.2％ and 67％,respectively.The growth inhibition of liver cancer cells was enhanced significantly with the increasing drug dose.The IC50 of growth inhibition of 5-FU,brucine and brucine nanoparticles was 16.7 μg/ml,90.3 μg/ml and 164.9 μtg/ml,respectively.There was significant difference among them (P＜0.05).The protein and mRNA expression of Fas in brucine nanoparticles treated SMMC-7721 cells increased significantly compared with that in blank control group(P＜0.05).Conclusion Brucine nanoparticles may potentially be a novel therapeutic drug for hepatocellular carcinoma.