人体恶性实体瘤TIL增殖力、表型和杀伤力研究

在建立高活力肿瘤浸润淋巴细胞（ＴＩＬ）分离培养方法的基础上，对８３例恶性实体瘤ＴＩＬ的细胞增殖动力学、部份ＴＩＬ表型与杀伤力等作了较为系统的研究。结果提示，本实验室所建立分离培养的ＴＩＬ增殖能力较强且稳定，有６５％的增殖倍数大于１０００倍；３Ｈ－ＴｄＲ掺入高峰在培养的第４５～７５天；对其自身的原代肿瘤细胞杀伤力可以维持至５６天以上；对５６份经ｒＩＬ２诱导的ＴＩＬ（培养时间１７±５．２天）表型分析：ＣＤ３８０±２１％，ＣＤ４３７±２１％，ＣＤ８４４±１８％，ＨＬＡＤＲ６９±２４％，与ｒＩＬ２诱导前比较，ＣＤ３、ＣＤ４、ＣＤ８和ＨＬＡＤＲ均增多，尤以ＣＤ３与ＣＤ８表型增多明显。     

Cytological detection of epithelial dysplasia in the oral mucosa using Feulgen-DNA-image cytometry

Cytological scrape material of the oral mucosa from 114 patients with epithelial dysplasia and with oral cancer was stained with the Feulgen-reaction and investigated with an image analyzer. The size and the integrated optical density of cell nuclei, and four chromatin texture features were measured. All tumor slides contained cell nuclei with DNA greater than 5c, 16% of the slides had cell nuclei with DNA greater than 8c. A total of 14.5% of the tumor patients showed significantly increased DNA values in nuclei distant from the tumor. Two smears with severe epithelial dysplasia showed nuclei with DNA greater than 5c both in the tumor material and far from the tumor. Texture analysis allowed discrimination between benign, dysplastic and malignant smears. No correlation was found between DNA content and tumor staging. Image cytometry was a reliable method for detecting tumor cells. Epithelial dysplasia in areas distant from the tumor is probably due to "field canceration" of the epithelium.     

Cytokine flow cytometry differentiates the clinical status of multiple sclerosis (MS) patients

In this study we have examined intracellular cytokines in peripheral blood mononuclear cells (PBMC) of MS patients by flow cytometry (cytokine flow cytometry). MS progressive patients showed an increased number of cells producing interferon-gamma (IFN-gamma) after activation with phorbol 12-myristate 13-acetate and ionomycin, compared with patients with clinically inactive forms (P < 0001) and with healthy controls (P = 0001). These cells belonged to the CD4+ and CD8+ subsets in similar proportions. Clinically inactive patients showed a lower level of cells producing IL-2 than controls (P = 0.03) and active MS patients (P = 0.03). Most IL-2-producing cells were CD4+ lymphocytes, although a small part of the IL-2 was also produced by CD8+ cells. The percentage of cells producing simultaneously IL-2 and IFN-gamma was increased in active MS and they were mainly CD4+ lymphocytes. No differences in the production of IL-4 were observed between groups. However, we found an increased IL-10 production in clinically active MS patients (P = 0.03). Treatment with IFN-beta of active MS patients showed lower levels of cytokines when compared with untreated MS patients. This methodological approach could help in the follow up and therapeutic monitoring of MS patients.     

经动脉化疗栓塞兔VX2肝癌后早期肿瘤细胞凋亡

目的　研究介入治疗兔VX2肝癌肿瘤细胞凋亡的早期动态改变 .方法　建立 2 7只新西兰大白兔VX2肝癌模型 ,随机分成肝动脉化疗栓塞组 (Transarterialchemoembolization ,TACE)、灌注化疗组 (Transarterialinfusion ,TAI)、灌注肝素生理盐水组 (对照组 ,Control) ,每组各 9只 .介入治疗后每组再随机分成 3个亚组 ,每亚组 3只 .在治疗后第2 4、72、12 0小时分别处死 3组中一亚组 ,取材肿瘤生长活跃的外带组织用流式细胞仪AnnexinV和PI双标记法定量检测细胞凋亡 ,组织病理切片苏木精 -伊红染色 (HE染色 )和甲基绿 -派洛宁染色 (MG -P染色 )光镜下形态学方法观察不同时间肿瘤细胞的凋亡变化 .结果　TACE、TAI、Control组在治疗后 2 4、72、12 0小时流式细胞仪法定量检测肿瘤细胞凋亡比率分别为 11.4 4± 2 .15、10 .99± 1.74、6 .0 0± 0 .5 8,5 .84± 0 .6 8、4 .6 5± 0 .11、2 .88± 1.2 3和 2 .80± 0 .15、2 .19± 1.6 9、2 .5 1± 2 .13.TACE、TAI、Control组凋亡在各时间点均有显著差异 (p <0 .0 1) ,TACE组 2 4、72小时的凋亡明显高于 12 0小时 (p <0 .0 5 ) .TACE组诱导的肿瘤细胞凋亡比率明显高于TAI、Control组 ;TACE组诱导的肿瘤细胞早期凋亡比率随时间呈下降趋势 ,在 12 0小时凋亡仍明显高于其     

Positive selection is an early event in thymocyte differentiation: high TCR expression by cycling immature thymocytes precedes final maturation by several days

T cell antigen receptor expression by cycling and post-cycling thymocytes has been analysed by flow cytometry. Normal mice were pulsed with 5-bromo-2'-deoxyuridine (BrdUrd), a thymidine analogue detectable with a monoclonal antibody. Thymocytes were surface-stained with antibodies against several V beta gene products and against whole alpha beta receptors and detection of BrdUrd in the nuclei was performed after enzymatic generation of single-stranded DNA. A significant (10%) percentage of thymocytes expressing high levels of alpha beta TCR were found in the cycle: these cells were immature, as shown by the CD4+8+ phenotype and by high HSA expression. After division, most alpha beta high BrdUrd+ cells entered a resting state and their number remained constant for 3 days, decreasing in two steps thereafter. This post-mitotic evolution was not modified by injection of an anti-mitotic drug. After day 4, a majority of the studied subset acquired a single positive phenotype. Location of BrdUrd+ V beta 8.2 high cells studied on frozen sections was found cortical at early times and medullary after day 3. V beta 6 expression by cycling and post-cycling thymocytes was analysed in various mouse strains, and early high expression by cycling thymocytes was found to be restricted to MIs 1b strains. These results suggest that high alpha beta TCR expression by cycling immature thymocytes corresponds to positive selection, which must therefore be considered as an early event in intrathymic differentiation.     

Large B-cell lymphomas: fine-needle aspiration plays an important role in initial diagnosis of cases which are falsely negative by flow cytometry

Large B-cell lymphomas (LBCLs) have significant false-negative results when immunophenotyped by flow cytometry (FC). To clarify the role fine-needle aspiration (FNA) in reducing this false-negative rate, 28 cases ultimately diagnosed as LBCL that had FNA as part of the workup and a negative FC were identified. We examined their clinical and cytologic features, in comparison with cases of LBCL with FNAs that were positive by FC. In 24/28 FC-negative cases (86%) a cytologic diagnosis of suspicious or positive for malignancy was rendered. We conclude that cytologic analysis is more sensitive than FC in the diagnosis of malignancy in FNA of LBCL, particularly in aspirates with low cellularity and/or low viability. Examination of cytospin preparations of the actual material analyzed by FC may provide an indication that an FC result is falsely negative. It is important to recognize the potential of false-negativity by FC of LBCLs when interpreting FNAs with features suggesting lymphoma.     

Fine-needle aspiration of a primary mediastinal large B-cell lymphoma: a case report with cytologic, histologic, and flow cytometric considerations

Fine-needle aspiration (FNA) cytology and immunophenotyping by flow cytometry (FCM) are increasingly being used for diagnosing and subclassifying lymphoma in the REAL/WHO classification. Herein, we report a case of primary mediastinal large B-cell lymphoma (PMBL), a subtype of diffuse large B-cell lymphoma in the WHO classification, diagnosed by FNA cytology in conjunction with FCM. This, to our knowledge, has not previously been reported. A 57-yr-old woman presented with bilateral axillary lymphadenopathy and intermittent shortness of breath. CT scan revealed a 5-cm anterior mediastinal mass and mediastinal lymphadenopathy. Endoscopic ultrasound-guided FNA of a 4.5-cm subcarinal lymph node showed medium to large atypical lymphocytes with scant to moderate finely vacuolated cytoplasm. Nuclei were enlarged, cleaved, noncleaved, lobulated, and hyperchromatic. The background showed lymphoglandular bodies. Malignant large cell lymphoma was cytologically diagnosed. FCM, performed on a portion of the FNA specimen, demonstrated large B cells devoid of surface immunoglobulin expression, the characteristic immunophenotype of PMBL. The histologic diagnosis was PMBL. Touch-imprint cytology of the histologic specimen showed large cells with a narrow rim of clear cytoplasm and prominent outer cell border. Nuclear features were similar to the FNA specimen. In the presence of a mediastinal mass, FNA cytology in conjunction with FCM can effectively diagnose PMBL in the appropriate clinical setting.     

异体角膜刺激引起的人外周血T细胞及其亚群CD25分子表达

目的：观察异体角膜体外对人外周血T细胞及其亚群CD25分子表达的影响。方法： 异体角膜与外周血淋巴细胞体外共同培养后，进行单克隆抗体标记和流式细胞分析。结果： 对照组T细胞CD25表达为25.2％；受角膜或沸波醇酯（PDB）激活后T细胞CD25表达分别为56.8％和80.9％；受角膜和沸波醇酯共同激活后T细胞CD25表达为70.2％；受异体角膜激活后，CD4和CD8 T淋巴细胞CD25表达分别为67.3％和52.3％。结论： 异体角膜组织体外能够刺激人外周T细胞CD25表达和T细胞活化，CD4 T细胞比CD8 T细胞活化更明显。     

流式细胞术检测肿瘤患者外周血T淋巴细胞亚群研究

目的 探讨肿瘤患者T淋巴细胞亚群的变化及临床意义。方法 采用流式细胞术检测27例肿瘤患者和25例正常对照组的T淋巴细胞亚群。结果 肿瘤患者CD3^＋T细胞、CD4^＋T细胞和CD4^＋／CD8^＋比值明显低于对照组并有统计学意义，而CD8^＋T细胞显著高于对照组（P〈0．01）。结论 肿瘤患者的免疫功能下降。流式细胞术对肿瘤患者的免疫功能的检测具有快速、敏感、准确的特点，对于评价疗效、判断预后具有重要价值。     

Effect of Gardenia extract ZG on the adsorption quantity of herpes simplex virus type 1 （HSV-1）

To observe the effect of Gardenia extract ZG on the adsorption quantity of herpes simplex virus type 1 (HSV-1) so as to explore the mechanism of its antiviral activity, fluorescein isothiocyanate (FITC) was used as the fluorescent probe to label viruses and heparin sodium was used as control. Meanwhile , the effect of Gardenia extract ZG on the adsorption quantity on the surface of Hep-2 cells was determined by flow cytometry. It was demonstrated that adsorption of HSV-1 on the surface of Hep-2 cells exhibited the character of saturation and specificity and heparin sodium could prevent attachment of viruses on these cells. These results are in accord with those reported previously. It was also proved that the manner of drug-use prior to adsorption or simultaneous use of drug and adsorption was better than adsorption prior to drug-use, and the inhibition rates of the former and latter manner were 84. 76% and 82.92% respectively. Three manners of drug-use with Gardenia extract ZG were all effective to reduce the adsorption quantity of viruses, especially the manner of simultaneous use of drug and adsorption with an adsorption inhibition rate of 68.46% . From the above observation, it is apparent that the mechanism of anti-viral activity of Gardenia extract ZG may be via several steps involved in the HSV-1 adsorption.