肝癌患者外周血LAK细胞活性测定及其影响因素的研究

近年来通过白细胞介素2(IL-2)诱导产生的淋巴因子活化性杀伤细胞(Lym-phokine-activated killer cell,LAK)在继承性免疫治疗肿瘤动物模型和人体肿瘤中已显出一定的疗效,但至今尚未见到有关研究肝癌患者LAK细胞活性变化的     

LAK细胞对不同靶细胞的杀伤作用及其意义

我们研究了重组白细胞介素２（ｒＩＬ２）活化的杀伤细胞（ＬＡＫ）对Ｋ５６２细胞、Ｒａｊｉ细胞、ＨｅｐＧ２细胞和２．２．１５细胞的杀伤活性。结果表明：ＬＡＫ细胞对不同靶细胞，其杀伤活性不同。在诱导早期（３小时），对Ｋ５６２细胞有明显的杀伤活性；在诱导晚期（３天和７天），对Ｒａｊｉ、ＨｅｐＧ２和２．２．１５细胞则有杀伤活性。揭示ｒＩＬ２诱生的ＬＡＫ细胞可能由不同的免疫杀伤细胞组成，在不同的诱导时期，活化不同的杀伤细胞。我们还建立ＨｅｐＧ２、２．２．１５细胞的靶细胞系统，可以在体外测定ＬＡＫ细胞对ＨＢＶ感染细胞的杀伤活性，亦可在体外筛选增强免疫杀伤细胞杀伤ＨＢＶ感染细胞活性的药物。     

IL—2／LAK细胞联合应用治疗原发性肝癌

用IL-2/LAK细胞联合治疗81例原发性肝癌,分别采用经静脉输注、肌肉注射及肿瘤局部用药。治疗后作CT、B超检查,观察肿瘤占位、淋巴细胞亚群、肝功能及临床症状、体征改变等。结果表明,局部用药组总有效率显著高于其他各组(P<0.01)。T细胞亚群检测表明,各治疗组治疗后的细胞免疫水平均显著提高。     

胎脾,脐血及成人外周血LAK细胞诱导的研究

用白细胞介素－２（ＩＬ－２）在体外诱导胎儿脾脏、新生儿脐血及成人外周血单个核细胞形成ＬＡＫ细胞，并行免疫标记和杀伤活性测定，发现胎脾、脐血ＬＡＫ细胞对ＨＬ－６０细胞的杀伤活性高于成人外周血ＬＡＫ细胞。     

人肝癌细胞与LAK细胞凋亡的超微结构比较

目的比较观察人肝癌细胞株ＢＥＬ７４０２细胞与ＬＡＫ细胞体外凋亡的超微结构．方法将ＢＥＬ７４０２细胞、人血树突状细胞和人ＬＡＫ细胞（淋巴因子和ＰＨＡ激活的杀伤细胞）共同置于含１００ｍＬ／Ｌ新生牛血清的１６４０培养液内，在３７℃，５０ｍＬ／Ｌ，ＣＯ２，饱湿条件下培养６ｈ．另将ＬＡＫ细胞单独置于含有环磷酰胺的上述培养液内，在同样条件下培养６ｈ．收集沉淀细胞制备电镜样本．细胞经２５ｇ／Ｌ戊二醛和２０ｇ／Ｌ锇酸双固定，ＰＤＡＰ包埋，超薄切片，常规染色，透射电镜观察．结果凋亡的ＢＥＬ７４０２细胞与ＬＡＫ细胞，其细胞核染色质浓缩、边集，核碎裂，离散的碎片外包以双层膜，一些线粒体也变得致密．然而，在有些凋亡的ＢＥＬ７４０２细胞内，可见浓缩的细胞碎片向细胞外出芽隆起，形成膜包的凋亡小体；其线粒体和粗面内质网的变化也较为复杂．结论凋亡的ＢＥＬ７４０２细胞与ＬＡＫ细胞均具有凋亡细胞的基本特征，但是两种凋亡细胞具体的形态表现不尽一致．表明细胞内的主要细胞器主动参与了凋亡过程．     

LAK细胞和单克隆抗体MGb2的协同抗胃癌作用

在LAK细胞对胃癌细胞KATOⅢ的杀伤过程中,加入单克隆抗体MGb2能产生明显协同作用。我们认为此作用的原理是抗体依赖细胞介导的细胞毒性作用(ADCC)。此作用随着制备LAK细胞时白细胞介素2(IL-2)浓度的增加而增高.与LAK活性呈平行关系:制备LAK细胞时,IL-2的诱导时间既影响LAK活性,也影响相应的ADCC作用。     

Endogenous lymphokine activated killer cell activity and cytogenetic response in chronic myelogenous leukaemia treated with α-interferon

The capacity of α-interferon (α-IFN) to induce lymphokine activated killer (LAK) cytotoxicity in the absence of interleukin-2 (IL2) has prompted us to test whether or not its ability to reduce dramatically the number of Ph1 ⁺ clones in chronic myelogenous leukaemia (CML) patients is in part mediated through the generation of natural killer (NK) or LAK activity. The latter were tested using NK-sensitive (K562) and NK-resistant (Raji) cell lines in a target-cell colony-growth inhibition assay. Effector cells (E) were patient blood mononuclear cells (MC) without in vitro activation prior to their coculture with targets (T). Out of 16 patients tested so far, three failed to undergo cytogenetic remission under α-IFN therapy. No NK nor LAK cells could be detected in the MC from two of them while the other displayed NK activity within upper normal limits. 13 patients underwent complete (eight) or partial (five) cytogenetic remission together with significantly high NK and/or LAK activity as compared to normal controls. These observations could favour the hypothesis of an indirect effect of α-IFN on leukaemic cells, mediated by cells involved in immune surveillance.     

Defective immunological functions associated with abnormal lymphokine-activated killer activity in patients with hepatocellular carcinoma

The in vitro lymphokine-activated killer (LAK) activity of peripheral blood mononuclear cells (PBMC) from 36 patients with hepatocellular carcinoma was investigated. The activity was greatly diminished in 13 patients and enhanced in seven patients. A flow cytometric study showed that the percentage of OKM1+, Leu-7+-11b+, and Leu-7-11b+ fractions in PBMC was decreased and the percentage of OKT8+ and Leu7+11- fractions was increased significantly in the 13 patients with lower LAK activity, compared with the values of the seven higher LAK activity patients. Furthermore, the response of PBMC to interleukin-2 (IL-2) was deficient in the lower activity group. However, there was no significant difference in IL-2 production by PBMC, IL-2 receptor (p55) expression of PBMC and mitogen (Con-A, PHA) response of PBMC between the two groups. These findings indicate the possibility that diminished LAK activity in hepatoma patients is due to a decreased number of LAK precursor cells and a defective response of LAK precursor cells to IL-2.     

Natural and activated cytotoxic lymphocytes reactivity to human hepatocellular carcinoma cell lines in hepatocellular carcinoma patients

The status of cellular cytotoxic activity in Hepatocellular Carcinoma (HCC) patients was compared to that in normal individuals by testing the cytotoxicity against K562 and five established HCC cell line targets. Natural killer (NK) activity of fresh peripheral blood mononuclear (PBM) cells in HCC patients to K562 cell line target was lower than that in normal donors. NK activity of unstimulated PBM cells from either source was minute against all five HCC cell line targets. Three different activation systems were employed to examine the cellular cytotoxicity of activated PBM cells: (1) conventional mixed lymphocyte culture (MLC), (2) allogeneic mixed lymphocyte tumor culture (MLTC), and (3) lymphokine-activated killer (LAK) cell culture. The cytotoxic effects of PBM cells in all three activation conditions were significantly lower in HCC patients than in normal donors (P less than 0.05 to P less than 0.01). These results suggest that, in addition to naturally present NK cells, the degree of in vitro activation of PBM cells may also have decreased in HCC patients.     

Pulmonary natural killer cell activity is reduced in patients with bronchogenic carcinoma

Natural killer (NK) cells are a subpopulation of lymphocytes capable of killing a variety of tumor targets. They can limit pulmonary metastases in vivo and thus might be important effectors of tumor defense in human lung. Lymphocytes were purified from whole lung specimens obtained from patients with lung cancer undergoing curative resection, and their NK activity was compared with that of lymphocytes purified from normal lung specimens obtained from cadavers undergoing medicolegal autopsy. The NK activity of pulmonary lymphocytes obtained from the patients with lung cancer was significantly lower (p less than 0.05) than the NK activity of normal lungs. This reduction occurred despite high levels of blood NK activity in the patients with cancer, suggesting that NK cells might be locally suppressed in the lungs of patients with bronchogenic carcinoma. Because human pulmonary macrophages (PM) are known to be potent inhibitors of NK function, we investigated the role that PM might play in the reduction of NK activity in these patients. The PM obtained from the patients with lung cancer released soluble inhibitors of NK activity when stimulated with lipopolysaccharide. Release of these inhibitors was blocked by indomethacin, strongly suggesting a role for arachidonic acid metabolites as an inhibitor of pulmonary NK function. Inhibition of NK function by PM may occur in vivo, as a significant inverse correlation (r = -0.71, p less than 0.001) existed between the NK activity of lymphocytes obtained from a lung and the number of PM present in the lung.(ABSTRACT TRUNCATED AT 250 WORDS)     

自然杀伤细胞抗肝癌的研究新进展

肝癌是最常见的恶性肿瘤之一,预后差,复发率高。免疫治疗已成为继手术、放疗、化疗后的第四种治疗模式,其中自然杀伤细胞可以在细胞、分子及基因水平抑制肿瘤转移及复发,在肝癌的免疫治疗中起关键性作用,在开展新的治疗模式以及提高肝癌患者生活质量方面具有很广阔的应用前景。     

Impaired T cell function and decreased natural killer activity in patients with liver cirrhosis and their significance in the development of hepatocellular carcinoma

It is well known that the incidence of hepatocellular carcinoma (HCC) in patients with liver cirrhosis (LC) is very high. We investigated the immunological state in patients with LC and HCC. T cell population of peripheral blood lymphocytes (PBL) and blast transformation of PBL by phytohaemagglutinin (PHA) were significantly decreased in patients with LC. Natural killer activity against HeLa cell was also significantly decreased in these patients. These results suggest that immunological surveillance is impaired in patients with LC and this may be one of the aetiological factors in genesis of hepatocellular carcinoma in patients with LC.     

Defective function of lymphokine-activated killer cells and natural killer cells in patients with hepatocellular carcinoma

Lymphokine-activated killer activity and natural killer activity in hepatocellular carcinoma patients were assessed. Maximum lymphokine-activated killer activity was induced at 3 to 5 days of incubation, and lymphokine-activated killer activity tended to increase in a manner dose dependent of recombinant interleukin-2. However, the maximum increase of lymphokine-activated killer activity in hepatocellular carcinoma was not as high as that of normal subjects or liver cirrhosis patients. Lymphokine-activated killer activity was impaired in hepatocellular carcinoma as compared to that in normal subjects. Hepatocellular carcinoma seemed to consist of two groups: i.e. a high-lymphokine-activated killer activity group and a low-lymphokine-activated killer activity group. Reduction of natural killer activity was also observed in hepatocellular carcinoma as compared with that in normal subjects and patients with liver cirrhosis. No correlation could be demonstrated between natural killer activity and lymphokine-activated killer activity in normal subjects, liver cirrhosis patients and hepatocellular carcinoma patients. With regard to the presence of HBsAg or alpha-fetoprotein concentration in the sera, there was no significant difference in natural killer and lymphokine-activated killer activity in hepatocellular carcinoma patients. Patients with a small mass lesion showed a low lymphokine-activated killer activity, and depressed lymphokine-activated killer activity was not necessarily related to tumor size. In comparison with the high-lymphokine-activated killer group, the low-lymphokine-activated killer group showed a significant decrease in gamma-interferon production and a preserved function of indocyanine green clearance.     

Depressed lymphokine-activated killer activity and analysis of the precursor cells in peripheral blood of patients with hepatocellular carcinoma

The in vitro lymphokine-activated killer activity and natural killer activity of peripheral blood mononuclear cells from 33 patients with hepatocellular carcinoma were investigated. Lymphokine-activated killer and natural killer activities of patients were significantly decreased compared with those of healthy volunteers. Peripheral blood mononuclear cells showed significantly lower lymphokine-activated killer and natural killer activities in patients with larger tumors (greater than or equal to 5 cm in diameter) than in patients with smaller tumors (less than 5 cm in diameter). Of 20 patients with larger tumors, 8 and 6 generated very little or no lymphokine-activated killer and natural killer activities. respectively. Lymphokine-activated killer precursors and natural killer cells were present mainly in the Leu-11+ fraction and partially in the Leu-7+ fraction in patients and normal volunteers. A flow cytometric study showed that the percentage of Leu-7+ 11+ and Leu-7-11+ fractions in peripheral blood mononuclear cells was lower in patients than in normal volunteers. The percentages of Leu-7-11+ and Leu-7+ 11+ fractions were diminished in the peripheral blood mononuclear cells of the patients with little or no lymphokine-activated killer activity. It is suggested that deficient lymphokine-activated killer and natural killer activities partially results from a reduction in the number of their precursor cells in patients with hepatocellular carcinoma.