THE EXPRESSION OF C－MYC AND N－RAS ONCOGENES IN HUMAN HEPATOCELLULAR CARCINOMA－AN IN SITU HYBRIDIZATION STUDY ON PARAFFIN EMBE

ＴＨＥＥＸＰＲＥＳＳＩＯＮＯＦＣ－ＭＹＣＡＮＤＮ－ＲＡＳＯＮＣＯＧＥＮＥＳＩＮＨＵＭＡＮＨＥＰＡＴＯＣＥＬＬＵＬＡＲＣＡＲＣＩＮＯＭＡ－ＡＮＩＮＳＩＴＵＨＹＢＲＩＤＩＺＡＴＩＯＮＳＴＵＤＹＯＮＰＡＲＡＦＦＩＮＥＭＢＥＤＤＥＤＴＩＳＳＵＥＳＥＣＴＩＯＮ...     

TUMOR NECROSIS FACTOR－α ALTERS PROTEINMETABOLISM AND CELL－CYCLE KINETICSIN MALIGNANT TUMOR

ＴＵＭＯＲＮＥＣＲＯＳＩＳＦＡＣＴＯＲ－αＡＬＴＥＲＳＰＲＯＴＥＩＮＭＥＴＡＢＯＬＩＳＭＡＮＤＣＥＬＬ－ＣＹＣＬＥＫＩＮＥＴＩＣＳＩＮＭＡＬＩＧＮＡＮＴＴＵＭＯＲＹｅＳｈｅｎｇｌｏｎｇ叶胜龙；ＴａｎｇＺｈａｏｙｏｕ汤钊猷；ＢｒｕｃｅＲＢｉｓｔｒｉａｎ...     

THE IN VITRO POTENTIATION OF LAK CELL CYTOTOXICITY IN CANCER AND AIDS PATIENTS INDUCED BY F3—A FRACTIONATED EXTRACT OF ASTRAG

ＴＨＥＩＮＶＩＴＲＯＰＯＴＥＮＴＩＡＴＩＯＮＯＦＬＡＫＣＥＬＬＣＹＴＯＴＯＸＩＣＩＴＹＩＮＣＡＮＣＥＲＡＮＤＡＩＤＳＰＡＴＩＥＮＴＳＩＮＤＵＣＥＤＢＹＦ３—ＡＦＲＡＣＴＩＯＮＡＴＥＤＥＸＴＲＡＣＴＯＦＡＳＴＲＡＧＡＬＵＳＭＥＭＢＲＡＮＡＣＥＵＳＣｈｕ...     

T－LYMPHOCYTE MEDIATED TUMOR CELL DESTRUCTIONIN VIVO ASSOCIATING WITH A SPECIFICFEATURE OF APOPTOSIS

Ｔ－ＬＹＭＰＨＯＣＹＴＥＭＥＤＩＡＴＥＤＴＵＭＯＲＣＥＬＬＤＥＳＴＲＵＣＴＩＯＮＩＮＶＩＶＯＡＳＳＯＣＩＡＴＩＮＧＷＩＴＨＡＳＰＥＣＩＦＩＣＦＥＡＴＵＲＥ　ＯＦ　ＡＰＯＰＴＯＳＩＳＹｕＤａ鱼达；ＹａｎｇＨｕａ杨骅；ＺｈｅｎｇＳｈｕ郑树；ＷａｎｇＸｉａ...     

PATHOLOGICAL STUDIES ON THE ANTI－INVASIVE CHARACTER BY RECOMBINANT HUMAN INTERLEUKIN－6 GENE－TRANSFECTED MOUSE LEUKEMIA CELLS

ＰＡＴＨＯＬＯＧＩＣＡＬＳＴＵＤＩＥＳＯＮＴＨＥＡＮＴＩ－ＩＮＶＡＳＩＶＥＣＨＡＲＡＣＴＥＲＢＹＲＥＣＯＭＢＩＮＡＮＴＨＵＭＡＮＩＮＴＥＲＬＥＵＫＩＮ－６ＧＥＮＥ－ＴＲＡＮＳＦＥＣＴＥＤＭＯＵＳＥＬＥＵＫＥＭＩＡＣＥＬＬＳＧｅＬｉｎｆｕ葛林阜，Ｃａｏ...     

ADYNAMIC STUDY OF THE CYTOTOXICEFFECTS OF HYPERTHERMIA COMBINED WITH CIS－DIAMINE DICHLOROLPLATINUM（DDP） ON HUMAN GASTRIC CANC

ＡＤＹＮＡＭＩＣＳＴＵＤＹＯＦＴＨＥＣＹＴＯＴＯＸＩＣＥＦＦＥＣＴＳＯＦＨＹＰＥＲＴＨＥＲＭＩＡＣＯＭＢＩＮＥＤＷＩＴＨＣＩＳ－ＤＩＡＭＩＮＥＤＩＣＨＬＯＲＯＬＰＬＡＴＩＮＵＭ（ＤＤＰ）ＯＮＨＵＭＡＮＧＡＳＴＲＩＣＣＡＮＣＥＲＣＥＬＬＬＩＮＥＳＭＫＮ...     

TWO－STAGE RESECTION FOR ADVANCED HEPATOCELLULAR CARCINOMA─PRELIMINARY RESULTS OF 16 CASES

ＴＷＯ－ＳＴＡＧＥＲＥＳＥＣＴＩＯＮＦＯＲＡＤＶＡＮＣＥＤＨＥＰＡＴＯＣＥＬＬＵＬＡＲＣＡＲＣＩＮＯＭＡ─ＰＲＥＬＩＭＩＮＡＲＹＲＥＳＵＬＴＳ　ＯＦ１６ＣＡＳＥＳＬｉａｎｇＬｉｊｉａｎ；Ｌｕｍｉｎｇｄｅ；ＨｕａｎｇＪｉｅｆｕ；ＰｅｎｇＢａｏｇａｎｇ梁...     

THE EXPRESSION OF CYTOKERATINS IN HUMAN HEPATOCELLULAR AND CHOLANGIOCELLULAR CARCINOMAS

ＴＨＥＥＸＰＲＥＳＳＩＯＮＯＦＣＹＴＯＫＥＲＡＴＩＮＳＩＮＨＵＭＡＮＨＥＰＡＴＯＣＥＬＬＵＬＡＲＡＮＤＣＨＯＬＡＮＧＩＯＣＥＬＬＵＬＡＲＣＡＲＣＩＮＯＭＡＳ￥ＳｕＱｉｎ；苏勤；Ｌｉｕｙａｎｆａｎｇ；刘彦仿（ＤｅｐａｒｔｍｅｎｔｏｆＰａｔｈｏｌｏｇｙ，...     

THE ANTICANCER EFFECT AND ANTI－DNA TOPOISOMERASE II EFFECT OF EXTRACTS OF CAMELLIA PTILOPHYLLA CHANG AND CAMELLIA SINENSIS

ＴＨＥＡＮＴＩＣＡＮＣＥＲＥＦＦＥＣＴＡＮＤＡＮＴＩ－ＤＮＡＴＯＰＯＩＳＯＭＥＲＡＳＥＩＩＥＦＦＥＣＴＯＦＥＸＴＲＡＣＴＳＯＦＣＡＭＥＬＬＩＡＰＴＩＬＯＰＨＹＬＬＡＣＨＡＮＧＡＮＤＣＡＭＥＬＬＩＡＳＩＮＥＮＳＩＳＸｉｅＢｉｎｓｆｅｎ；谢冰芬；ＬｉｕＺ...     

Basic Investigations EXPRESSION OF GAP JUNCTION PROTEIN Cx43 IN CULTURED HUMAN NORMAL AND MALIGNANT LUNG CELLS

ＢａｓｉｃＩｎｖｅｓｔｉｇａｔｉｏｎｓＥＸＰＲＥＳＳＩＯＮＯＦＧＡＰＪＵＮＣＴＩＯＮＰＲＯＴＥＩＮＣｘ４３ＩＮＣＵＬＴＵＲＥＤＨＵＭＡＮＮＯＲＭＡＬＡＮＤＭＡＬＩＧＮＡＮＴＬＵＮＧＣＥＬＬＳＺｈａｎｇＺｈｉｑｉａｎ；张志谦；ＬｉｎＺｈｏｎｇｘｉａｎｇ...     

Isolation and characterization of cytotoxic granules from human lymphokine (interleukin 2) activated killer cells

Cytotoxic granules were isolated from human lymphokine-activated killer (LAK) cells and analyzed for their biochemical properties. Isolated granules of approximately 85-95% purity were obtained by differential centrifugation followed by discontinuous Percoll gradient centrifugation. The murine lymphocyte granule marker N-alpha-carbamazepine-L-lysine thiobenzyl ester-esterase as well as cytotoxic activity toward the human tumor cell lines K562, Raji, Daudi, and CEM were associated with LAK granule fractions. Granule-associated N-alpha-carbamazepine-L-lysine thiobenzyl ester-esterase activity increased in recombinant interleukin 2 expanded human LAK cells in parallel with cytotoxic activity for Raji tumor cell targets. Cytotoxic LAK cell granules mediated calcium-dependent killing of the tumor cell lines K562, Raji, Daudi, and CEM. However, no calcium-dependent hemolytic activity was found. Preincubation of human granules with calcium, a treatment which totally inactivates the hemolytic and cytotoxic activity of murine lymphocyte granules [perforin 1 (P1)] had no effect on human LAK granule cytotoxicity for nucleated cells. Human LAK granules appear to contain P1 detected as cross-reactive antigen detected by mouse anti-P1 and human anti-C9 in Western blot analysis. In addition, Northern blot analysis of polyadenylated RNA isolated from human LAK cells using a murine P1 complementary DNA probe showed a cross-hybridizing 2.8- to 3.0-kilobase mRNA species identical in size to murine P1 mRNA. These results demonstrate that despite similar biochemical composition, functional differences exist between human and murine cytotoxic granules. Human LAK granules were synthesized in response to recombinant interleukin 2 activation and appeared in parallel with cytotoxicity for tumor targets, suggesting an important role for LAK granules in tumor cell cytotoxicity by human LAK cells.     

Mechanism of trifluorothymidine potentiation of oxaliplatin-induced cytotoxicity to colorectal cancer cells

Oxaliplatin (OHP) is an anticancer agent that acts by formation of Platinum-DNA (Pt-DNA) adducts resulting in DNA-strand breaks and is used for the treatment of colorectal cancer. The pyrimidine analog trifluorothymidine (TFT) forms together with a thymidine phosphorylase inhibitor (TPI) the anticancer drug formulation TAS-102, in which TPI enhances the bioavailability of TFT in vivo. In this in vitro study the combined cytotoxic effects of OHP with TFT were investigated in human colorectal cancer cells as a model for TAS-102 combinations. In a panel of five colon cancer cell lines (WiDr, H630, Colo320, SNU-C4 and SW1116) we evaluated the OHP-TFT drug combinations using the multiple drug-effect analysis with CalcuSyn software, in which the combination index (CI) indicates synergism (CI<0.9), additivity (CI=0.9-1.1) or antagonism (CI>1.1). Drug target analysis was used for WiDr, H630 and SW1116 to investigate whether there was an increase in Pt-DNA adduct formation, DNA damage induction, cell cycle delay and apoptosis. Trifluorothymidine combined with OHP resulted in synergism for all cell lines (all CI<0.9). This was irrespective of schedule in which either one of the drugs was kept at a constant concentration (using variable drug ratio) or when the two drugs were added in a 1 : 1 IC(50)-based molar ratio. Synergism could be increased for WiDr using sequential drug treatment schedules. Trifluorothymidine increased Pt-DNA adduct formation significantly in H630 and SW1116 (14.4 and 99.1%, respectively; P<0.05). Platinum-DNA adducts were retained best in SW1116 in the presence of TFT. More DNA-strand breaks were induced in SW1116 and the combination increased DNA damage induction (>20%) compared with OHP alone. Exposure to the drugs induced a clear cell-cycle S-phase arrest, but was dose schedule and cell line dependent. Trifluorothymidine (TFT) and OHP both induced apoptosis, which increased significantly for WiDr and SW1116 after TFT-OHP exposure (18.8 and 20.6% respectively; P<0.05). The basal protein levels of ERCC1 DNA repair enzyme were not related to the DNA damage that was induced in the cell lines. In conclusion, the combination of TFT with the DNA synthesis inhibitor OHP induces synergism in colorectal cancer cells, but is dependent on the dose and treatment schedule used.     

以SMMC—7721为靶细胞MTT法检测LAK细胞活性方法的建立

目的建立以贴壁生长的癌细胞为靶细胞、MTT法检测LAK细胞活性的方法。方法用MTT比色法测定LAK细胞对体外培养的K562、Raji、SMMC－7721细胞的杀伤活性。结果LAK细胞对已知的NK敏感细胞K562及NK耐受细胞Raji均有明显杀伤作用,贴壁生长的SMMC-7721肝癌细胞对NK细胞不敏感,而对LAK细胞敏感,在效靶比为40：1时,LAK细胞对肝癌细胞的杀伤率达90％以上,其杀伤效应随效靶比增高而增高。结论SMMC-7721细胞可作为检测LAK细胞活性的靶细胞。以贴壁细胞作为靶细胞的MTT法检测LAK细胞活性较传统的悬浮细胞如Raji作为靶细胞具有灵敏、准确、简单、可靠的优点,更适用于国内一般实验室开展。     

Antitumor effect of resveratrol oligomers against human cancer cell lines and the molecular mechanism of apoptosis induced by vaticanol C

Twenty resveratrol (3,5,4'-trihydroxystilbene) (Res) derivatives, which were isolated from stem bark of Vatica rassak (Dipterocarpaceae), were evaluated for in vitro cytotoxicity against a panel of human tumor cell lines. Among them, seven compounds displayed marked cytotoxicity. Vaticanol C (Vat C) as a major component induced a considerable cytotoxicity in all cell lines tested and exhibited growth suppression in colon cancer cell lines at low dose. Vat C caused two cell lines (SW480 and HL60) to induce cell death at four to seven times lower concentrations, compared with Res. The growth suppression by Vat C was found to be due to apoptosis, which was assessed by morphological findings (nuclear condensation and fragmentation) and DNA ladder formation in the colon cancer cell lines. The apoptosis in SW480 colon cancer cells was executed by the activation of caspase-3, which was shown by western blot and apoptosis inhibition assay. Furthermore, the mitochondrial membrane potential of apoptotic SW480 cells after 12 h treatment with Vat C was significantly lost, and concurrently the cytochrome c release and activation of caspase-9 were also detected by western blot analysis. Over-expression of Bcl-2 protein in SW480 cells significantly prevented the cell death induced by Vat C. Taken together, the findings presented here indicate that Vat C induced marked apoptosis in malignant cells mainly by affecting mitochondrial membrane potential.     

人LAK细胞的诱导及其生物特性的研究

LAK cell activity of peripheral blood from normal donors was induced by interleukin 2 (IL-2). Optimal induction of LAK cells was obtained by 3-4 day incubation of PBMC in the presence of 30-40 units/ml of IL-2. Several cell lines Raji, CEM, HeLa, 109 (esophageal carcinoma) and OC (ovarian carcinoma) which were NK cell resistant tumor cells could be lysed by LAK cells. PHA was unable to markedly induce the LAK cell activity. Although both LAK and NK cell activity was decreased by mitomycin C treatment, their degree of susceptibility to such treatment was quite different. NK cells were more sensitive to mitomycin C than LAK cells. In contrast to the significant inhibition of the NK cell-mediated lysis by the addition of anti-CD 2 monoclonal antibody during cytotoxicity assay LAK cell-mediated lysis was not affected. These results indicate that LAK cells are capable of lysing a variety of tumor cells that are resistant to NK cells and their biological characteristics are also different.     

S-acetyl-glutathione selectively induces apoptosis in human lymphoma cells through a GSH-independent mechanism.

Reduced apoptosis is associated to cancer development. Agents able to restore the programmed cell death responsiveness of cancer cells are foreseen as potential effective cancer therapies. In this study, we report that a glutathione-S-derivative, S-acetyl-glutathione (Sag), induces significant apoptosis in three human lymphoma cell lines, including Daudi, Raji and Jurkat cells while it had no or little effect on either Hut-78 lymphoma cells or the normal BT lymphocytes. We used Annexin-V FACS analysis and DNA laddering to demonstrate that Sag activated apoptosis in the three sensitive cell lines in a dose- and time-dependent-fashion. Using mercury orange staining and FACS analysis, we showed that Sag generated an intracellular GSH depletion in Daudi, Raji and Jurkat cells but not in Hut-78 or the normal BT cells. These data provide direct evidence that Sag specifically activates programmed cell death in lymphoma cells through, at least in part, a depletion of intracellular GSH rather than an increase, as previously suggested. Because of its selective effect on cancer cells, Sag appears as a promising new lymphoma cell apoptosis inducer with potential clinical value for lymphoma patients.     

Cisplatin induces fas expression in esophageal cancer cell lines and enhanced cytotoxicity in combination with LAK cells

OBJECTIVE: To establish a new therapeutic approach for the treatment of esophageal cancer, we investigated an alternative mechanism of immunotherapy for sensitizing target cells to effector cells. METHODS: Six human esophageal cancer cell lines were used. The expression of Fas antigen on tumor cells was determined by flow cytometry. The cytotoxic effect of cis-dichlorodiammineplatinum (CDDP) and anti-Fas antibody was evaluated using an MTT assay. The cytotoxic activity of LAK cells was measured by a (51)Cr release assay. RESULTS: Five out of six esophageal cancer cell lines expressed Fas antigen at various levels (26.2-61.5%), and Fas expression increased after CDDP treatment. The antitumor effect of anti-Fas antibody on the esophageal cancer cell line and the antitumor effect of LAK cells activated by IL-2 were enhanced by pretreatment with CDDP. After concanamycin A treatment to specifically evaluate Fas-dependent cytotoxicity, LAK cells expressing Fas ligand killed only Fas-positive cells, but not Fas-negative cells. An anti-Fas neutralizing antibody inhibited this cytotoxicity. DNA fragmentation was shown in a cell line that was treated with CDDP and anti-Fas antibody, and also in the targeted esophageal cancer cell line cocultured with LAK cells. CONCLUSION: Our results suggest a potential clinical application of CDDP as a Fas inducer to make esophageal tumors susceptible to Fas antigen and LAK cytotoxicity.     

Augmentation of lymphokine-activated killer cell cytotoxicity by monoclonal antibodies against human small cell lung carcinoma

This study examined the lymphokine-activated killer (LAK) cell cytotoxicity on monoclonal antibody (MoAb)-bound tumor cells from the human small cell lung carcinoma cell lines H69 and H128. LAK cells were generated from normal peripheral blood mononuclear cells by incubation with interleukin 2 for 3 or more days. Cells from the LAK culture were cytotoxic to natural killer-sensitive (K562, 84% cytotoxicity) and natural killer-resistant (Daudi, 85%; H69 and H128, 69% and 97%, respectively) cell lines, and to freshly excised human lung (49%) and breast (57%) tumors. LAK cytotoxicity to H69 or H128 cells was significantly augmented by target cell preincubation with the small cell lung carcinoma-reactive MoAbs 1096 (increases of up to 271%) or 5023 (up to 223%). SCLC 5023 or 1096 did not enhance LAK cytotoxicity to Daudi cells of lymphoblastoid origin. Pretreatment of LAK cells with an anti-Fc receptor antibody blocked MoAb augmentation by 1096 or 5023 (but not LAK cytotoxicity), suggesting that LAK-MoAb interaction may be mediated by Fc binding. LAK activity coincided with emergence of a large cell [interleukin 2-stimulated large mononuclear leukocyte (LML)] subset expressing the CD16 and NKH-1 surface determinants. Serial immunophenotyping of the LAK cell culture harvested at Days 3, 5, and 7 indicated that the level of LAK cytotoxicity, with or without MoAb augmentation, correlated with frequency of NKH-1-reactive LMLs. These observations support the hypothesis that LAK cytotoxicity is mediated by a NKH-1-reactive LML subpopulation. Antitumor cytotoxicity may be augmented by tumor-reactive MoAbs through Fc binding to this LML subset.     

B-NHL患者NK细胞中CD16ζ表达及利妥昔单抗与LAK细胞的联合抗瘤作用

背景与目的:自然杀伤细胞(nature killer cell,NK)是抗体依赖细胞介导的细胞毒作用的主要效应细胞,肿瘤患者普遍存在NK细胞活化功能的缺陷可能会影响单克隆抗体的治疗效果.因此如能逆转NK细胞的CD16ζ链信号转导的功能缺陷,并与单克隆抗体联合免疫治疗,可能会产生协同抗肿瘤作用.本研究的目的是了解B细胞非霍奇金淋巴瘤(B-cell non-Hodgkin's lymphoma,B-NHL)患者是否存在NK细胞的活化障碍,体外白细胞介素-2(interleukin-2,IL-2)是否能完全逆转其活化障碍,并观察利妥昔单抗与LAK细胞联合对肿瘤细胞的杀伤作用.方法:使用密度梯度离心方法分别分离69例B-NHL患者和30例健康志愿者外周血单个核细胞(peripheral blood mononuclear cell,PBMC),将两种PBMC在体外与1 000 U/ml IL-12共同培养制备LAK细胞,流式细胞仪检测PBMC和LAK细胞中CD16ζ链的阳性率和平均荧光强度.流式细胞仪检测Raji细胞表面CD20的表达;Annexin V/PI方法检测利妥昔单抗单药对Raji细胞的促凋亡作用,乳酸脱氢酶(lactate dehydrogenase,LDH)释放实验进行杀伤活性的检测.结果:在B-NHL组和健康对照组,CD56 细胞表达CD16ζ链的阳性率为(63.3±16.4)%、(97.8±3.1)%(P＜0.001),CD16ζ链MFI值分别为1.3±1.3和3.6±1.7(P＜0.001).在体外1 000 U/ml的IL-2共培养的LAK细胞中,两组CD16ζ链的阳性率分别为(99.3±4.1)%和(99.7 3.9)%,其MFI值分别为29.2±12.5和31.4±13.8,均无显著性差异(P=0.15和P=0.44).40 μg/ml利妥昔单抗可以完全结合细胞表面CD20抗原,在24 h时才开始出现对Raji细胞的明显的凋亡作用.利妥昔单抗与LAK细胞联合对Raji细胞的杀伤率在不同的浓度组均明显高于不加利妥昔单抗组(P＜0.05).LAK细胞与Herceptin(40 μg/ml)联合的杀伤率与不加Herceptin组相比,在各效靶比浓度梯度均无明显提高(P＞0.05).LAK细胞与利妥昔单抗联合对Jurket细胞的杀伤率在各效靶比浓度梯度均与不加利妥昔单抗组无显著性差异(P＞0.05).结论:B-NHL患者普遍存在NK细胞CD16ζ链的表达下调,高剂量的IL-2可以显著增强CD16ζ链的表达,利妥昔单抗与LAK细胞联合可增强对Raji细胞的抗肿瘤作用.     

高表达bcl-2卵巢癌细胞系参与药物抵抗和细胞凋亡作用的研究 *

目的：探讨化疗的敏感性与细胞凋亡的关系,观察细胞凋亡抑制分子ｂｃｌ－２在卵巢癌化疗抵抗中的作用。方法：构建了逆转录病毒ｂｃｌ－２表达载体ｐＬＸＳＮ／ｂｃｌ－２,应用ｌｉｐｏｆａｃｔｉｎ法建立转染ｂｃｌ－２基因和转染空载体ｐＬＸＳＮ的卵巢癌细胞系。ＭＴＴ法观察细胞系抵抗药物的细胞毒作用。ＦＡＣＳ和ＤＮＡ琼脂糖凝胶电泳观察ｂｃｌ－２高表达细胞系在阿霉素诱导细胞凋亡中的变化。结果：转染 ｂｃｌ－２的卵巢癌细胞系ＯＣ３／ｂｃｌ－２稳定表达ｂｃｌ－２分子并抵抗药物介导的细胞毒作用,明显提高细胞的存活率,与此同时也抑制细胞凋亡的发生。结论：化疗的敏感性与细胞凋亡的调节密切相关,凋亡抑制基因ｂｃｌ－２在肿瘤耐药中起重要作用。