马钱子碱抗肝细胞癌作用的实验研究

目的 探讨马钱子碱抗肝细胞癌的作用及其机制.方法 体外培养人肝癌SMMC-7721细胞,加入不同浓度的马钱子碱(2.5～400μg/ml),细胞培养72 h,MTT法测定细胞生长抑制率.Western blotting和荧光定量RT-PCR技术分别测定培养24、48、72 h肝癌细胞PCNA、Cyclin D1、FAS基因mRNA和蛋白表达.结果 随着马钱子碱用量逐渐增加,对人肝癌细胞SMMC-7721生长抑制作用增强,马钱子碱用量为320 μg/ml时对肝癌细胞生长抑制率接近100%.马钱子碱作用肝癌细胞24 h与作用48 h相比,Fas蛋白和mRNA表达差异无统计学意义(分别F=2.547,1.582,均P＞0.05),作用72 h时差异有统计学意义(分别F=1.036,1.137,均P＜0.05);PCNA和Cyclin D1的mRNA和蛋白表达各时间点差异无统计学意义(PCNA分别F=3.612,2.174,3.029;Cyclin D1分别F=2.361,2.915,1.976,均P＞0.05).结论 马钱子碱抑制肝癌细胞生长,可能通过肝癌细胞FAS基因和蛋白表达增加,诱导肝癌细胞凋亡发挥抑制作用,而与PCNA和Cyclin D1作用无关.

Abstract：

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.     

树突状细胞及其亚群在结肠直肠癌术后肝转移中的作用及其临床意义

结肠直肠癌是常见的恶性肿瘤之一。在欧美国家其发病率居恶性肿瘤的第3位,死亡率居恶性肿瘤的第2位;而在我国其发病率居恶性肿瘤的第3～5位,占肿瘤死亡人数的4.9%,且近年来随着人们生活水平的提高和饮食结构改变有逐渐上升趋势。     

大鼠肝硬变和肝癌发生中肝组织病理和甲胎蛋白的变化

目的 探讨大鼠肝硬变和肝癌发生中肝组织病理和甲胎蛋白变化的意义.方法 选取雄性Wistar大鼠,分别采用DENA、四氧化碳和橄榄油诱导建立肝癌和肝硬变模型,于诱导前和诱导后2周、4周、8周、14周、18周和21周分别获取肝、脾组织和外周静脉血,HE染色进行肝脾组织病理学检查,ELISA法测定外周血清AFP水平.结果 大鼠肝硬变诱发过程中18周后出现肝假小叶,淤血性脾肿大;大鼠肝癌诱发过程中14周病理性核分裂,核仁变大、数量增多,肝癌结节形成,脾脏充血性改变.大鼠肝硬变诱发过程中外周血AFP在4周开始升高,14周与对照组比较差异显著(P<0.05);大鼠肝癌诱发过程中外周血AFP在2周开始升高,8周时与对照组比较差异显著(P<0.05);肝癌大鼠外周血AFP表达水平在8周时显著高于肝硬变大鼠(P<0.05).结论 不同诱导因素下大鼠肝脏病理变化出现时间和损害程度不尽相同,DENA对肝脏损害程度较四氯化碳对肝脏损害重;肝细胞受到病毒、细茵、毒素、化学毒物等损害时,动态监测外周血AFP水平,对评估肝细胞损伤程度和癌变具有双重生物学意义.     

纳米技术在抗肿瘤药物中的应用及意义

抗肿瘤药物是治疗恶性肿瘤的重要手段之一,然而许多抗肿瘤药物存在着水溶性差,稳定性差,对正常细胞的杀伤以及由此引发的严重毒副反应等问题,影响了抗肿瘤药物的临床应用.因此,如何提高抗肿瘤药物对肿瘤细胞的选择性,降低其毒副反应,提高难溶性抗肿瘤药物的溶解性显得非常重要.     

IL-10和IL-12在结直肠癌肝转移中作用的临床研究

目的探讨IL-10、IL-12在结直肠癌肝转移中作用。方法分别收集自2008年1月到2010年6月健康成年人、结直肠癌术前、术后和术后肝转移患者外周血,采用ELISA法测定外周血清IL-10、IL-12水平,采用FACS Calibur3.0流式细胞仪测定T淋巴细胞及其亚群比例,进行各组结果统计学分析。结果结直肠癌术后肝转移患者外周血清IL-10水平显著高于健康成年人、结直肠癌术后患者(P<0.05),结直肠癌术后患者外周血清IL-10水平显著低于术前患者(P<0.05);结直肠癌术前、术后和术后肝转移患者外周血清IL-12水平显著低于健康成年人,结直肠癌术前、术后患者之间以及术后和术后肝转移患者之间外周血清IL-12水平存在统计学差异(P<0.05);结直肠癌术后肝转移患者外周血T淋巴细胞总数及Th细胞比例数显著低于健康成年人(P<0.05),Ts细胞比例数显著高于健康成年人(P<0.05)。NK细胞在结直肠癌术后肝转移患者显著低于健康成年人(P<0.05)。结论不同临床时期结直肠癌患者存在细胞免疫或体液免疫严重障碍,纠正机体细胞因子和免疫状态紊乱,不仅能够改善机体免疫功能,可能有助于控制结直肠癌的发生、发展。     

立体化心理干预应用于肺结节患者术前负性情绪和睡眠质量的效果

目的 探讨立体化心理干预应用于肺结节患者对改善术前负性情绪及睡眠质量的效果。方法 2021 年 8 月至 2022 年 8 月 我院收治的肺结节患者 80 例,随机分为对照组和研究组,每组各 40 例。两组均行胸腔镜肺结节切除术治疗。对照组实施常规 围术期干预,研究组实施术前立体化心理干预。对比两组干预前后负性情绪、睡眠质量变化及治疗依从性。结果 研究组术前 1d 的 HAMA、HAMD 评分均较对照组低（P<0.05）。研究组术前 1d 的睡眠质量各维度评分及总分均较对照组低（P<0.05）。研 究组治疗依从性较对照组高（P<0.05）。结论 肺结节患者实施立体化心理干预,能够减轻术前负性情绪,改善睡眠质量,增强 治疗依从性。     

树突状细胞功能异常对结直肠癌肝转移的影响

目的 探讨树突状细胞(DC)功能异常对结直肠癌肝转移的影响及其意义.方法 收集2008年1月至2010年6月健康成年人30例、结直肠癌术前、术后无肝转移者30例和术后肝转移30例患者外周血,分离单核细胞后分别加入GM-CSF(1000 U/ml)、IL-4(1000 U/ml)和'TNF-α(1000 U/ml)诱导分化成熟DC,通过加入HT29大肠癌细胞抗原处理(100 μg/ml)或不处理,DC∶T细胞为1∶10,培养7 d,用ELISA方法测定培养液中IFN-γ和IL-10水平,酶标仪测定CCK8和LDH光密度(OD)值,间接测定T细胞增殖和杀伤能力.结果 结直肠癌术后肝转移患者外周血DC在无肿瘤抗原刺激时与健康人T细胞混合培养液IL-10水平显著低于结直肠癌术前患者和健康人[(11.9±1.3)pg/ml比(29.6±9.7)pg/ml、(23.4±8.0)pg/ml,F=4.475,P＜0.05].结直肠癌术后肝转移患者外周血DC经肿瘤抗原刺激与健康人T细胞混合培养液IFN-γ水平均显著高于结直肠癌术后和健康人[(34 ± 9)pg/ml比(26±12)pg/ml、(24 ±6) pg/ml,F=5.206,P＜0.05].结直肠癌术后肝转移患者DC经HT29肠癌细胞抗原刺激后诱导健康人T淋巴细胞杀伤能力显著高于结直肠癌术前患者和健康成年人[(30.6 ±8.6)pg/ml比(12.1 ± 2.4)pg/ml、(14.9 ±1.7)pg/ml,F=4.147,P＜0.05].结论 结直肠癌患者外周血DC在肿瘤抗原刺激下诱导T淋巴细胞产生IL-10,导致肿瘤免疫逃逸,发生肝转移;外源肿瘤抗原刺激能够提高T淋巴细胞杀伤能力.

Abstract：

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.     

马钱子碱纳米微粒抗肝细胞癌作用的实验研究

目的 观察马钱子碱纳米微粒的抗肝细胞癌作用.方法 采用超声乳化法研制羧基化聚乙二醇-聚乳酸嵌段共聚物马钱子碱纳米微粒,体外观察其对肝癌细胞SMMC-7721生长的影响;采用Western blotting和FQ-PCR检测药物作用72 h对肝癌细胞凋亡相关蛋白Fas蛋白和mRNA表达水平的影响.结果 马钱子碱纳米微粒平均粒径(146±96) nm,载药率:4.2％,包封率:67％.随药物浓度的增加,肝癌细胞生长抑制作用明显,呈现剂量依赖性.5-FU、马钱子碱和马钱子碱纳米微粒对肝癌细胞生长的IC50分别为16.7 μg/ml、90.3 μg/ml和164.9 μg/ml,三者之间差异显著(P＜0.05).与空白纳米微粒相比,马钱子碱纳米微粒显著诱导肝癌细胞Fas蛋白和mRNA表达,且320 μg/ml剂量诱导表达显著高于160 μg/ml (P＜0.05).结论 马钱子碱纳米微粒显著抑制肝癌细胞生长,诱导肝癌细胞凋亡,可能与上调肝癌细胞Fas蛋白和mRNA表达有关.马钱子碱纳米微粒是一种具有潜在发展前景的肝癌治疗药物.