白血病细胞和正常细胞生成树突状细胞的比较研究

目的体外诱导髓系白血病原代细胞分化生成树突状细胞(dendritic cell，DC)．并与正常的DC比较。方法分离初诊24例急性髓细胞性白血病和10例慢性髓细胞性白血病患的骨髓单个核细胞以及5例正常人外周血的单个核细胞，用rhGM—CSF 1000U／ml、rhIL-4500U／ml和TNF—α 50U／ml联合培养10天；形态学(Wright染色、倒置显微镜、透射电镜)、免疫学(CD80、CD86、CD83、CD1a、HLA—DR)鉴定，RT—PCR检测白血病克隆以及混合淋巴细胞反应(MLR)检测抗原递呈功能。结果细胞因子诱导后，正常细胞和白血病细胞均出现典型形态的树突状细胞，CD80、CD86、CD83、CD1a的表达明显上调(正常和CML—DC的HLA—DR也明显上调)，急性和慢性髓细胞性白血病以及正常细胞来源的树突状细胞具有不同程度的刺激异基因T淋巴细胞增殖的能力。结论无论是急性和慢性白血病细胞均能诱导分化成DC，并且特征完全相似；白血病与正常骨髓的DC在形态和免疫学表达方面相似，但功能较弱。     

口腔鳞状细胞癌的细胞表面抗原

ABH 同种抗原除了存在于红细胞表面和某些器官的分泌液中,也存在于体内各种组织中。皮肤、口腔、支气管和肺等上皮细胞的细胞表面以及血管内皮均可发现这种抗原,由于这种抗原均存在于表皮细胞而间叶结缔组织通常缺如,提示 ABH 同种抗原是鉴     

小细胞与非小细胞混合型肺癌

关于肺癌的临床处理,主要是以小细胞肺癌(SCLC)与非小细胞肺癌(NSCLC)的组织病理学区别为基础。NSCLC 倘若病变局限。可手术治愈;但若胃转移,对化疗的反应令人失望.SCLC 即使病变局限,一般也不能手术治愈,因早期即有远处微小转移存在,但对化疗有显效,少数病人可获治愈,因此,准确的组织学诊断对决定 SCLC 与 NSCLC 的治疗方案极为重要。至于 SCLC 与 NSCLC 混合型肺癌,迄今尚不清楚究竟是肺癌的一种独立的组织类型,还是     

小细胞肺癌的细胞遗传学研究

在我国,肺癌在恶性肿瘤死亡率的构成统计中占第五位,而小细胞肺癌(SCLC)的发病率约占肺癌的10%。在美国,则为25%。早已证明SCLC 有明显的细胞遗传学改变和生化标记,最近的研究更进一步发现有规律性的染色体改变,即3p(14～23)的缺失。本文结合有关特     

胃癌患者外周血单个核细胞、脾细胞和局部淋巴结细胞的LAK细胞活性

研究表明,IL-2可诱导外周血单个核细胞表达 IFN-γ或TNF-α mRNA及分泌 IFN-γ或TFN-α。作者检测29例胃癌患者外周血单个核细胞(PBMC)、脾细胞(SPC)和局部淋巴结细胞(LNC)经IL-2诱导4天后产生的LAK细胞活性,~(51)Cr释放法测定其细胞毒活     

CIK细胞体内外抗肝癌细胞作用

目的：研究肝癌患者CIK（cytokine-induced killer) 细胞的体外杀伤自体肝癌原代细胞的细胞毒活性以及正常人CIK细胞在裸鼠体内的抗肿瘤作用。方法：分别分离获得肝癌患者和下人的外周血单个核细胞（PBMC）,加入细胞因子,体外诱导成CIK细胞,用流式细胞仪对细胞作动态表型分析,并与正常人的CIK细胞作对比。用^51Cr释放法,测定肝癌患者的CIK细胞体外杀伤自体肿瘤细胞的细胞毒性活性。在Balb/c裸鼠皮下接种肝癌细胞BEL－7402,观察CIK细胞对荷瘤鼠的抑瘤作用,并与LAK、PBMC细胞相对比。结果：肝癌患者的CIK细胞体外增殖力强,至培养28天时达到最大增值倍数300多,表型分析结果表明,CD^3 CD56^ 双阳性细胞得到了大量的扩增,其含量由原来的0．23％上升到第21天的17．8％。体外实验表明,肝癌患者的CIK细胞杀伤自体原代肝癌细胞的细胞毒性活性明显高于自体的PBMC细胞。裸鼠体内实验表明,肝癌患者的CIK细胞能够显著抑制肿瘤的生长,其抑瘤率可达84．7％,高于LAK细胞的52．8％及PBMC的37．1％（P＜0．05和P＜0．01）。结论：CIK细胞具有较强的体内外抗肝癌细胞活性,有可能应用于临床上肝癌的过继性免疫治疗。     

LAK细胞的细胞群体来源初探

LAK细胞既不同于目前淋巴细胞分类中的杀伤性T细胞也不同于NK细胞，其细胞群体来源有待进一步研究．本文报告了利用对不同淋巴细胞群体分别诱导LAK细胞活性的方法，研究LAK细胞的细胞群体来源的初步结果。     

LAK细胞

淋巴因子活化杀伤细胞(Lymphokine-activated killer cell,LAK 细胞)是近年来发现的又一类重要的免疫活性细胞,它具有NK 和 CTL 细胞所不可比拟的肿瘤杀伤效应。LAK 细胞在体内广泛存在,具有广谱和高效杀瘤效应,表明它在肿瘤的免疫监视中占有重     

半抗原修饰的黑素瘤细胞激活树突状细胞诱导特异性T细胞反应

目的：探索半抗原[二硝基氟苯（DNP）]修饰的恶性黑素瘤细胞（恶黑）激活树突状细胞（DC）体外诱导特异性T细胞反应的抗肿瘤效应。方法：采用DNP修饰恶黑细胞B16（H-2^b），然后在体外激活C57BL／6小鼠（H-2^b）外周血来源的DC，观察其诱导T细胞的增殖和特异性T细胞杀伤功能；在小鼠皮肤迟发性超敏反应实验和荷瘤实验中观察DNP修饰瘤苗＋DC的特异性。结果：经DNP修饰的B16细胞激活的DC，诱发T细胞增殖能力和对B16细胞的特异性杀伤效应均明显高于未修饰的B16细胞和DC。动物实验显示DNP修饰瘤苗联合DC具有明显的诱发皮肤迟发性超敏反应作用和抑瘤作用。结论：DNP修饰B16所激活的Dc可以诱导更强的恶黑特异性T细胞效应。     

Alpha-interferon and lymphokine-activated killer cells in hairy cell leukemia

Lymphokine-activated killer (LAK) cell activity generated from peripheral blood was tested in 6 patients with typical hairy cell leukemia, 3 not on treatment with alpha-interferon (alpha-IFN) and 3 receiving therapy. In all cases, substantial killing of the LAK-sensitive target Daudi was observed, but hairy cells, whether or not they had been pretreated with alpha-IFN, were uniformly resistant to LAK lysis. The hairy cells were also resistant to LAK cell killing generated from normal peripheral blood mononuclear cells. alpha-IFN added at various times during LAK generation had little or no effect on LAK activity. It is concluded that LAK cells are not important in mediating the beneficial effects of alpha-IFN in hairy cell leukemia.     

Generation and characterization of lymphokine-activated killer cells against fresh human leukemia cells

Lymphokine-activated killer (LAK) cells generated from 15 acute leukemia patients in remission showed significant levels of cytotoxicity against Daudi 1A4, a natural killer-resistant cell line. This indicates that lymphocytes of leukemia patients in remission could respond to interleukin-2 to generate conventional LAK cells. However, LAK cells caused lysis of autologous leukemia cells at considerably lower levels in seven out of the 15 patients, with the exception of one case (48.6% cytolysis). None of the remaining eight patients exhibited LAK activity against autologous leukemia cells. On the other hand, patients' LAK could lyse allogeneic leukemia cells including those resistant to autologous LAK. Thus, patients' LAK seem not to be defective in lysis of leukemia cells. In the cold target competition analysis, the binding of patients' LAK to leukemia cells could be inhibited by autologous and allogeneic leukemia cell competitors, implying that almost all leukemia cells could be recognized by patients' LAK. Most LAK cells from normal donors showed significant lysis of allogeneic leukemia cells, but some leukemia cells were found to be resistant to lysis. LAK cells against both leukemia cells and Daudi 1A4 were phenotypically heterogenous, and were predominantly observed in the T3- fraction in the precursor phase. In the effector phase, whereas LAK activity against leukemia cells was also predominantly shown in the T cell-depleted fraction, similar levels of LAK activity against Daudi 1A4 were found in both the T cell-depleted and -enriched fractions.     

Effect of human interleukin 3 on the susceptibility of fresh leukemia cells to interleukin-2-induced lymphokine activated killing activity.

Pretreatment of acute myeloblastic leukemia cells with the hemopoietic growth factor interleukin 3 (IL3) increased their susceptibility to lymphokine activated killing (LAK) but did not affect their constitutive resistance to native natural killer activity. In addition, IL3 treatment did not alter the LAK cell-mediated killing of CD34+ hemopoietic progenitors present in normal bone marrow. Increased 3H-thymidine uptake was generally observed after IL3 treatment. However, failure to proliferate in response to IL3, observed in some cases, did not prevent changes in LAK susceptibility. Enhanced lysis of IL3-treated leukemic cells was accompanied by a moderate increase of the effector-target binding. Increased LAK susceptibility was already observed at 18 h, while optimal cytolysis and expression of the cell adhesion molecule (CAM) LFA-3 (CD58) by IL3-treated AML cells were concomitantly observed at later culture times. In contrast, the CAM ICAM-1 (CD54) was not modulated by IL3, nor were significant changes in the expression of either CAMs observed in normal hemopoietic cells. Blocking experiments with the anti-CD58 monoclonal antibody demonstrated a variable neutralizing effect on the IL3-induced increase of LAK activity, depending on the leukemia cell studied. The effect described here, together with the known role of IL3 in normal hemopoiesis makes it a factor of potential therapeutic value for the treatment of leukemic patients.     

Generation and Characterization of Lymphokine-activated Killer Cells against Fresh Human Leukemia Cells

Lymphokine-activated killer (LAK) cells generated from 15 acute leukemia patients in remission showed significant levels of cytotoxicity against Daudi 1A4, a natural killer-resistant cell line. This indicates that lymphocytes of leukemia patients in remission could respond to interleukin-2 to generate conventional LAK cells. However, LAK cells caused lysis of autologous leukemia cells at considerably lower levels in seven out of the 15 patients, with the exception of one case (48.6% cytolysis). None of the remaining eight patients exhibited LAK activity against autologous leukemia cells. On the other hand, patients' LAK could lyse allogeneic leukemia cells including those resistant to autologous LAK. Thus, patients' LAK seem not to be defective in lysis of leukemia cells. In the cold target competition analysis, the binding of patients' LAK to leukemia cells could be inhibited by autologous and allogeneic leukemia cell competitors, implying that almost all leukemia cells could be recognized by patients' LAK. Most LAK cells from normal donors showed significant lysis of allogeneic leukemia cells, but some leukemia cells were found to be resistant to lysis. LAK cells against both leukemia cells and Daudi 1A4 were phenotypically heterogenous, and were predominantly observed in the T3⁻ fraction in the precursor phase. In the effector phase, whereas LAK activity against leukemia cells was also predominantly shown in the T cell-depleted fraction, similar levels of LAK activity against Daudi 1A4 were found in both the T cell-depleted and -enriched fractions.     

Cell-cell adhesion-independent killing due to lymphokine-activated killer cells against glioblastoma cell lines.

Lymphokine-activated killer (LAK) cells can kill several tumor cells. Their killing activity is generally due to cell-cell adhesion. Cell-cell adhesion of the LAK cells to the target cells is essential for LAK lysis. In this report, however, we describe that the LAK cells can also kill the target cells by cell-cell adhesion-independent killing. Killing occurred after the target cells were exposed to the LAK cells. When the LAK cells were added to glioblastoma cell lines T98G and U373MG (which proliferate by adhering to the bottom of a culture flask), the LAK cells killed them by cell-cell adhesion killing within 4 h (early killing). On the other hand, when small numbers of the LAK cells were added, some of the target cells escaped from the early killing. At 4 and 6 h after the adding the LAK cells, when the LAK cells were discarded from the flask by washing with PBS, the escaped cells still adhered and were alive. However, they ultimately died over the next 24-96 h (late killing). The late killing was the cell-cell adhesion-independent killing, because it occurred after the LAK cells were removed. In this killing, numerous granules and vacuoles appeared in the cytoplasm of the cells. The vacuoles enlarged and then the cells died. The cell death was different from apoptosis, because the nucleus was intact until the late stage and no DNA fragment laddering in the degenerated cells was recognized. The vacuoles were stained with acid phosphatase and the cell death was inhibited with 3-methyladenine (an inhibitor of lysosome), suggesting that the late killing may be autophagic cell death due to activated lysosome. Induction of late killing in tumor cells using the LAK cells may become one approach for cancer therapy.     

Membrane-associated Lymphotoxin Expression and Functional Analysis of Lymphokine-activated Killer Cells Derived from Tumor-infiltrating Lymphocytes

The expression of a membrane-associated lymphotoxin molecule (mLT) on lymphokine-activated killer (LAK) cells obtained from 18 patients with malignant tumors and its role in the tumor cell killing mechanisms were investigated. LAK cells from tumor-infiltrating lymphocytes (TIL-LAK cells) were mainly composed of CD3-positive cells, whereas LAK cells from peripheral blood lymphocytes (PBL-LAK cells) were mainly composed of CD16- and CD56-positive cells. However, mLT was found to be expressed on TIL-LAK cells as well as PBL-LAK cells. The degree of mLT expression correlated with the killing activity of LAK cells towards L929 cells (r=0.806, P <0.01, n = 15), but not with that towards Daudi or K562 cells. Although the degree of mLT expression correlated with the amount of secreted lymphotoxin (LT) in the supernatant of LAK cell culture, the secreted LT itself could not account for the tumor cell killing activity of LAK cells. Polyclonal rabbit anti-LT antibody partially inhibited the killing activities of LAK cells towards L929 cells and this inhibition was found in the combination of autologous tumor cells and PBL-LAK cells. These findings suggest the possibility that the mLT-related cytotoxicity is involved in the tumor cell killing mechanisms of TIL-LAK cells as well as PBL-LAK cells.     

In vitro and in vivo susceptibility of human leukemic cells to lymphokine activated killer activity

The susceptibility of human myeloid and lymphoid leukemic blasts to the lytic action of recombinant interleukin-2 (rIL-2)-generated lymphokine activated killer (LAK) cells was analyzed. With the exception of the K562 cell line, all 9 leukemic cell lines tested were resistant to the natural killer activity of freshly isolated peripheral blood lymphocytes (PBL) from healthy donors but were susceptible to the lytic action of PBL cultured for 3 days in the presence of rIL-2. Of the 32 primary myeloid and lymphoid acute leukemia samples investigated, the great majority were natural killer cell-resistant but were variably sensitive to LAK effectors. Variations in LAK activity were observed according to the donor of PBL, while little or no difference was documented in the capacity to elicit LAK activity of PBL cultured with 100 or 1,000 U of rIL-2/ml. Pretreatment of the leukemic target cells with neuraminidase did not increase substantially their sensitivity to LAK activity. LAK cells generated from the PBL of patients at the onset of the disease or in complete clinicohematological remission lysed Raji cells as efficiently as normal LAK effectors. Finally, LAK cells were capable of abrogating the tumor growth in nude mice of a human leukemic T cell line. These findings demonstrate the susceptibility in vitro and in vivo of human leukemic blasts to the lytic effect of LAK cells and point to a possible clinical exploitment of this new form of adoptive immunotherapy in the management of acute leukemia.     

Lymphokine-activated killer cells can kill target cells not only by early killing but also by late killing,and the late killing level against autotumor cell line

Generally, lymphokine-activated killer (LAK) cells kill target cells early after the LAK cells adhere to them. In this study, we describe that LAK cells can also kill at a later time, such as 24-96 h. LAK cells were generated from a cancer patient and healthy volunteers. As target cells, the patient's autotumor cell line H41 was used. When LAK cells were added to the target cells in a culture well, the LAK cells killed the target cells by cell-cell adhesion within 1-4 h (early killing), but not all cells were killed. The LAK cells were then removed. However, the remaining cells ultimately died 24-96 h later (late killing). The late killing was different from the early killing because numerous granules and vacuoles appeared in the cytoplasm. The late killing was not induced by adding supernatant of the LAK cell culture, suggesting that LAK-target cell contact may be necessary for the killing. The cell injury was inhibited by 3-methyladenine (lysosome inhibitor). It suggests that the vacuoles may be caused by activated lysosome. The patient's LAK cells induced late killing at high levels. There was a high percentage of CD8(+)CD16(+) cells in the peripheral blood lymphocytes (PBL). This subset induced late killing more effectively than the CD8(-)CD16(+) subset. Killing was more conspicuous against H41 than against allogeneic cell line T98G. This type of killing is noteworthy for understanding of killing mechanism of LAK cells.     

Increased susceptibility to lymphokine activated killer (LAK) lysis of relapsing vs. newly diagnosed acute leukemic cells without changes in drug resistance or in the expression of adhesion molecules.

The NK and LAK activity of peripheral blood lymphocytes of leukemic patients as well as the susceptibility of their acute myeloid (AML) and lymphoblastic (ALL) leukemia cells to autologous and allogeneic LAKs were examined. In addition, neoplastic cells at diagnosis and at relapse were compared in the same patients for several features, including in vitro susceptibility to LAKs and to the drugs used in the induction phase, expression of MDR phenotype and of adhesion molecules, and differentiation markers. The NK activity of patients' LAK cells on K562 was significantly lower than that of a group of healthy donors whereas no differences were found in LAK activity as evaluated on Daudi cells. Three of 5 AML and 3 of 4 ALL were significantly more susceptible to autologous and allogeneic LAK lysis when blasts obtained at relapse were compared with leukemic cells of the same patients at diagnosis. This different lysability was not associated with in vitro modified sensitivity to drugs used in induction treatment. Moreover, no elevation in the expression of the multidrug-resistance (MDR)-related P170 glycoprotein was noted in relapsing leukemic cells. Even the expression of adhesion molecules and differentiation markers did not correlate with lysability of leukemic cells. These data demonstrate that relapsing leukemic blasts can be significantly lysed by LAK cells and suggest a rationale for adoptive immunotherapy with IL-2 and LAK cells in the treatment of acute leukemic patients.