垂体瘤细胞增殖与凋亡关系的研究

目的：探讨垂体瘤细胞增殖与凋亡的关系。方法：应用免疫线化法和TUNEL法对20例复发、26例未复发的人脑垂体瘤增殖细胞核抗原（PCNA）与凋亡进行检测。结果：复发组垂体瘤增殖细胞指数（PCNA L1）及凋亡细胞数量较未复发组明显增高,两组有显著性差异（P＜0．05）。结论：肿瘤细胞的增殖细胞核抗原表达指数和凋亡检测是判断垂体瘤预后的指标之一,有临床指导意义。     

脑膜瘤血小板源生长因子及受体表达与脑膜瘤增殖活性及凋亡研究

目的 探讨血小板源生长因子（PDG－F）在脑膜瘤发生和发展中的作用。方法 采用免疫组化检测新鲜脑膜瘤组织中PDGF及其受体表达,用TUＮＥＬ法检测细胞凋亡情况,同时进行细胞增殖核抗原（PCNA）检测,结果 脑膜瘤高表达PDGF及其受体,且以表达PDGFB链及PDGF β受体为主,部分脑膜瘤表达水平很低的PDGFA链,几乎不表达PDGFa受体;非典型性脑膜瘤PDGFBB,PDGFR β蛋白表达水平及PCNA指数高于良性脑膜瘤,但同时其凋亡也增加。结论 PDGFBB／R β自分泌环可能在脑膜瘤的发生和发展过程中起着重要作用。     

青霉素致痫鼠脑组织中凋亡调控基因P^53及Bcl—2的表达及意义

目的 探讨癫痫鼠脑组织中P53及Bcl-2的表达及意义。方法 通过免疫组化方法对青霉素致痫鼠癫痫灶脑组织中的P^53、Bcl-2进行检测。结果 随着痫性发作次数的增多,P^53有显著的上升趋势;致痫组有Bcl-2的表达,发作8次组Bcl－2表达最高,以后随着痫性发作次数的增多,有显著的下降趋势,P^53基因与Bcl-2蛋白表达呈负相关性。结论 P^53基因表达水平与神经元损害程度有一定的关系。P^53基因可诱导B cl-2表达的减少,神经细胞的自我保护是有限的。     

帕金森病大鼠黑质细胞凋亡与Bax蛋白表达的相关性

鉴于在体研究帕金森病(Parkinson disease，PD)模型成功前黑质细胞凋亡的研究罕见报道，我们采用神经损毁剂6-羟基多巴胺(6-OHDA)制作大鼠PD模型，采用免疫组织化学、电镜观察的方法，动态观察PD大鼠模型成功前促凋亡基因Bax蛋白表达与6-OHDA诱发的黑质细胞凋亡的关系。     

白细胞介素8表达与星形细胞肿瘤增殖与凋亡的相关性研究

目的 探讨白细胞介素8(IL-8)mRNA、增殖细胞核抗原(PCNA)在星形细胞肿瘤组织中的表达及其与细胞凋亡之间的关系. 方法 应用RT-PCR方法 检测64例星形细胞肿瘤组织及10例正常大脑组织中IL-8 mRNA的表达情况.用免疫组织化学方法 检测PCNA在肿瘤组织石蜡标本中的表达.并用TUNEL法检测肿瘤细胞的凋亡. 结果 星形细胞肿瘤组织中IL-8mRNA表达与星形细胞肿瘤的恶性程度有关.随着肿瘤恶性程度的增高,PCNA表达增高,细胞凋亡指数增高.IL-8 mRNA表达与PCNA表达呈正相关关系(r=0.938,P<0.01),与细胞增殖亦呈正相关关系(r=0.907,P<0.01). 结论 IL-8mRNA表达与星形细胞肿瘤的恶性增殖呈正相关,星形细胞肿瘤通过抑制凋亡并且上调IL-8分泌促进肿瘤细胞增殖,从而导致肿瘤恶性转化.     

脑胶质瘤中端粒酶活性表达及与增殖、凋亡的相关性

目的观察不同级别人脑胶质瘤中端粒酶活性表达及与增殖、凋亡的相关性。方法采集手术中切除的不同级别人脑新鲜胶质瘤标本26例,脑外伤内减压脑组织8例,用TRAP法及TUNEL法、PCNA免疫荧光染色流式细胞仪法检测不同级别脑胶质瘤端粒酶活性以及凋亡、增殖细胞百分率。结果脑胶质瘤中端粒酶活性阳性表达率及代表端粒酶活性的平均△A值,在Ⅰ、Ⅱ级和Ⅲ、Ⅳ级之间差别显著(P<0.05),各个级别与对照组之间均差别显著(P<0.05),对照组脑组织端粒酶活性均为阴性。平均PCNA阳性细胞率在正常组,Ⅰ、Ⅱ级和Ⅲ、Ⅳ级之间存在显著性差异(P<0.01)。TUNEL阳性细胞率在正常组与其余各组之间均存存显著差异(P<0.01),Ⅳ级与其余各组之间亦均存在显著差异(P<0.05)。代表端粒酶活性的平均△A值(r=0.78,P<0.05)以及平均PCNA阳性细胞率(r=0.86,P<0.01)分别与胶质瘤恶性级别呈正相关。结论脑胶质瘤中,端粒酶活性阳性及PCNA阳性表达细胞率可作为预测胶质瘤化疗效果和脑胶质瘤恶性程度的标志之一,端粒酶活性表达水平和增殖活性与胶质瘤细胞恶性级别具有相关性。     

多巴胺抑制PC12细胞增殖和诱导凋亡作用的研究

目的　研究多巴胺(DA)对PC12细胞的增殖抑制和诱导凋亡的作用,探讨帕金森病(PD)神经元的死亡机制。方法　应用免疫组织化学、流式细胞仪、电镜及电泳技术,研究DA对大鼠嗜铬细胞瘤PC12细胞的增殖抑制及诱导凋亡的作用。结果　适当浓度(0.5mmol/L)DA能显著抑制PC12细胞的生长并诱导其凋亡,在作用时间较短时(<12h)表现为对PC12细胞的生长抑制,此时流式细胞仪检测未见凋亡峰,但细胞周期显示S期细胞明显抑制。此时Bcl-2染色呈强阳性,电镜下细胞形态基本正常,可见线粒体、内质网肿胀及核分裂相减少。当作用时间延长时(>24h),流式细胞仪可见典型亚二倍体凋亡峰,此时电泳可见典型DNA“阶梯状”电泳带,电镜可见核浓缩、染色体边聚等凋亡特征性核结构改变,Bcl-2染色阳性率降低。结论　DA具有抑制PC12细胞增殖和诱导凋亡作用,细胞凋亡参与了PD的病变过程。     

脑肿瘤局部浸润单个核细胞亚群及其免疫功能的原位免疫组织…

应用免疫组化方法对５１例脑肿瘤局部浸润的单个核细胞进行观察。发现主要是辅助／诱导（ＣＤ４）Ｔ细胞、抑制／细胞毒（ＣＤ８）Ｔ细胞和单核巨噬细胞参与肿瘤免疫。仅９例及７例分别有少许Ｂ细胞与ＮＫ细胞。ＣＤ４、ＣＤ８Ｔ细胞的密度与单核巨噬细胞密度呈正相关，与肿瘤体积呈负相关。各病理类型脑肿瘤中ＣＤ４亚群均受抑制，胶质瘤还选择性抑制ＣＤ８亚群。脑肿瘤局部免疫状态不受肿瘤发生部位和浸润Ｔ细胞标记增殖期淋巴细胞     

IGF-1表达与人脑膜瘤细胞增殖活性关系

目的探讨IGF-1在人脑膜瘤中的表达与脑膜瘤细胞增殖活性之间的关系。方法应用免疫组化SP法检测42例脑膜瘤组织、7例正常硬脑膜中IGF-1和增殖细胞核抗原(PCNA)的表达情况,结果进行统计学分析。结果 IGF-1在正常硬脑膜组、WHO I、II和III级脑膜瘤组的表达强度不等,分别为0%、(15.7±12.8)%、(47.2±11.7)%和(62.9±12.9)%,其差异均具有统计学意义(P<0.05)。PCNA在同样四个组别中的PCNA-LI值分别为0%、9.6±5.4%、30.6±5.6%和48.7±15.1%,其差异亦均具有统计学意义(P<0.05)。相关分析结果显示:IGF-1与PCNA-LI两者呈正相关(r=0.8594)。结论 IGF-1表达强度与脑膜瘤细胞增殖活性有关,二者在脑膜瘤的发生、增殖和恶化过程中可能起协同作用,其详细机制有待进一步研究。     

SV40与脑肿瘤相关性研究

SV40在体外可使人及多种动物正常细胞发生恶性转化,在动物体内可诱发多种肿瘤。在人脑肿瘤及其它几种肿瘤组织中新发现有SV40的存在。本文就SV40转化细胞及致瘤机理以及与人脑肿瘤相关性研究作一简要介绍。     

神经上皮肿瘤中端粒酶活性和细胞增殖凋亡的研究

目的探讨神经上皮肿瘤的端粒酶活性及与肿瘤细胞增殖凋亡的关系。方法对35例神经上皮肿瘤组织和8例正常者脑组织采用改良TRAP法检测端粒酶活性，TUNEL法检测肿瘤细胞凋亡指数，免疫组化法检测嗜银蛋白（AgNOR）指数。结果神经上皮肿瘤端粒酶阳性率为23／35，Ⅰ、Ⅱ、Ⅲ及Ⅳ级肿瘤的端粒酶活性分别为37．1TPG、75．2TPG、165．1TPG和243．2TPG；端粒酶活性、AgNOR指数和肿瘤恶性度呈正相关，凋亡指数和肿瘤恶性度成负相关；端粒酶活性和AgNOR指数成正相关，和凋亡指数成负相关。结论在神经上皮肿瘤中端粒酶活性不仅和肿瘤恶性度密切相关，还和肿瘤细胞的凋亡增殖密切相关。     

In situ analysis of cell kinetics in human brain tumors

Summary A newly developed in vitro labeling method with bromodeoxyuridine (BrdU) identifies S phase cells in situ in freshly obtained surgical tissue of human brain tumors which is subsequently fixed and embedded in paraffin for BrdU immunovisualization. For the first time, the BrdU labeling index (LI) is successfully compared here with the LI obtained by immunostaining of frozen sections of the same tumors with monoclonal antibody Ki-67 which identifies all proliferating cells, i.e., the growth fraction. LIs were counted in at least five different areas with high density of labeled cells; at least 1,000 cells were counted. In 13 metastatic tumors, Ki-67 LI was 8.3%–62.6%, and BrdU LI was 5.1%–28.0%. In 18 gliomas, Ki-67 LI was 1.4%–19.3%, and BrdU LI was 0.2%–11.6%. In 7 meningiomas, Ki-67 LI was 0.3%–3.0%, and BrdU LI was 0%–2.0%. Statistical comparison of Ki-67 and BrdU LIs by linear regression analysis revealed a highly significant correlation: BrdU LI=0.99+0.34 Ki-67 LI (r=0.92,P<0.001). A significant heterogeneity of proliferation patterns may occur within one sample from area to area, as well as between different samples of the same tumor, especially in gliomas; thus, some subjective influence on LIs by arbitrary sampling and selection could occur in quantitative evaluation of in situ cell kinetics of human brain tumors. This study indicates that our in vitro BrdU-labeling method allows the in situ identification of S phase cells in excellently preserved fixed tumor tissue which is well suited for further histological examination. This method compares favorably with Ki-67 labeling of frozen sections and might emerge as a powerful new tool for the routine study of cell proliferation in surgical specimens of human brain tumors.Supported by funds from the Commission for Cancer Research of the Medical Faculty of the University of Vienna     

P－糖蛋白在颅脑肿瘤中表达的研究

采用Ｐ－糖蛋白单抗和免疫荧光染色法检测４５例颅脑肿瘤手术标本，Ｐ－糖蛋白阳性表达率如下：原发性胶质瘤１３／２８，脑转移癌１／２，颅骨恶性肿瘤２／２，脑膜瘤、听神经瘤和垂体腺瘤等良性肿瘤Ｏ／１３。在胶质瘤中Ｐ－糖蛋白的表达强度与肿瘤的恶性程度呈正相关。在同一肿瘤中，Ｐ－糖蛋白的分布很不均一，呈明显的异质性。     

Immunohistochemical determination of protein kinase C expression and proliferative activity in human brain tumors

Summary Protein kinase C (PKC), the major receptor for phorbol ester tumor promotors, is a phospholipid- and calcium-dependent phosphorylating enzyme which plays an important role in the intracellular signal transduction necessary for a variety of basic cellular functions including the control of cell proliferation. To determine the expression of PKC in human neurogenic tumors we investigated 121 tumors of the human nervous system by means of immunohistochemistry using the monoclonal antibody C5. The results were compared with immunohistochemical staining for intermediate filament proteins, desmoplakins, and the proliferation-associated nuclear antigen Ki-67. Besides strong staining of normal and reactive astrocytes, C5 immunoreactivity was consistently observed in tumor cells of all types of gliomas. However, the fraction of C5 positive tumor cells varied between the different tumor types with astrocytomas and subependymomas demonstrating the strongest immunoreactivity. In the other gliomas, especially those of higher malignancy, a considerable heterogeneity in C5 expression could be observed. There was a tendency for the percentage of C5 immunostained tumor cells being lower in high-grade gliomas compared to low-grade ones and comparison with Ki-67 staining frequently revealed an inverse relationship between proliferative activity and C5 immunoreactivity. Besides the gliomas we found 3 of 7 neurinomas and 6 of 18 meningiomas which were partially C5 positive. All other tumors investigated including medulloblastomas and metastatic carcinomas were C5 negative. Our results thus indicate that immunohistochemistry for PKC using the monoclonal antibody C5 could be an useful aid for histopathological tumor classification in neurooncology.Supported by the Deutsche Forschungsgemeinschaft, SFB 200     

PC12细胞缺氧后细胞凋亡与DNA损伤相关基因表达关系的研究

目的 探讨PC12细胞缺氧后细胞凋亡与DNA损伤的关系。方法 采用TUNEL法,结合应用流式细胞术观察PC12细胞缺氧培养不同时间点细胞凋亡现象,以及DNA损伤相关基因表达的改变。结果 在缺氧0.5h开始出现凋亡细胞,引时P53、P21^waf1/cip1蛋白表达开始增高。至缺氧1h凋亡细胞达高峰,此时P53、P21蛋白表达最高（P＜0.05）。至缺氧6－12h,则以坏死为主,此时以上基因表达均减弱。结论 PC12细胞在缺氧早期（0.5-2h）主要出现以凋亡为主的细胞死亡,伴有DNA损伤相关基因表达的动脉改变,提示缺氧后PC12细胞凋亡部分是由于DNA损伤严重、损伤不能及时修复所致。     

Argyrophilic nucleolar organizer region proteins (Ag-NORs) in human brain tumors: relations with grade of malignany and proliferation indices

Summary Proliferation indices and mean number of silver-stained nucleolar organizer region-associated proteins (Ag-NORs) are compared in 65 brain tumors, including 34 gliomas, 8 meningiomas, 17 metastatic tumors, and 6 other tumors. Immunocytochemical investigations include labeling with the monoclonal antibody Ki-67 which identifies the whole growth fraction, and with a monoclonal antibody against bromodeoxyuridine (BrdUrd) which detects cells in the S phase of the cell cycle after in vitro incubation with BrdUrd. When all types of tumors are collectively considered, mean numbers of Ag-NORs did not correlate with Ki-67 and Brd-Urd labeling indices (LIs) and mitotic index. Among tumor subtypes, only meningiomas showed significant correlations between Ag-NOR counts, LIs, and malignancy. Mean number of Ag-NORs did not correlate with proliferation indices and tumor grade in low-grade and high-grade gliomas. However, recurrent high-grade gliomas showed a tendency to higher Ag-NOR counts than primary tumors. This study indicates that counting of Ag-NORs in paraffin sections is of limited value in tumor neuropathology. Correlations found in meningeal tumors should be substantiated in larger series.     

脑牵拉引起脑神经元凋亡的实验研究

目的探讨脑牵拉压（BRP）的测量方法及脑牵拉压引起神经元细胞凋亡的规律和机制.方法利用电阻应变计制作脑牵拉力的测量装置,对其定标.选用新西兰大白兔30只,分为30、40、50 g组,牵拉完毕后,用流式细胞仪检测各组脑牵拉压区神经细胞凋亡的百分率和细胞线粒体的活性变化.结果30 g的BRP牵拉30 min引起较低的细胞凋亡率,几乎不影响细胞线粒体的膜电位;40 g的BRP牵拉30 min引起较高的细胞凋亡率.细胞线粒体的膜电位明显降低;50 g的BRP牵拉15 min引起更高的细胞凋亡率,细胞线粒体的膜电位显著降低.结论该装置可作为脑牵拉压的测量工具,脑牵拉可引起神经元细胞凋亡以及细胞线粒体膜电位降低.实验性脑牵拉时,最好将BRP控制在40 g以下.     

Proliferating cell nuclear antigen expression in brain tumors,and its prognostic role in ependymomas: an immunohistochemical study

Summary Proliferating cell nuclear antigen (PCNA)/cyclin is currently often investigated immunohistochemically in tumors as a marker of cell proliferation, but many problems remain open concerning its reliability as a prognostic factor. PCNA has been studied in a series of 123 brain tumors using the monoclonal antibody PC10. A clear intra-and inter-tumor variability of PCNA-positive nuclei has been found, but taking into account the tumor areas with the highest number of positive nuclei, a positive correlation between this number and the histological malignancy of tumors has been demonstrated. The staining intensity of nuclei was variable; very-intensely positive nuclei, counted separately, are hypothesized to represent nuclei in S-phase of the cell cycle. In ependymomas the investigation included a quantitative statistical analysis. The number of PCNA-positive nuclei correlated with cell density and mitotic index, but only very intensely positive nuclei showed a significant statistical correlation with survival. In spite of the many possibilities of wrong interpretation of PCNA expression, the most important of which is its deregulation, the method is useful in the practice for prognostic purposes. Its important advantages are the possibility of a retrospective application and a visual analysis of the proliferation potential of tumors.Supported by CNR, Rome (Special Project A.C.R.O.); AIRC, Milan; Italian Ministry of Scientific Research and University (Progetto Finalizzato 40%); and CSI-Piemonte     

Immunostaining for proliferating cell nuclear antigen: its role in determination of proliferation in routinely processed human brain tumor specimens

Alcohol- and formalin-fixed, paraffin-embedded samples of 71 brain tumors (35 gliomas, 22 metastatic carcinomas, 8 meningiomas and 6 other tumors) were investigated by immunocytochemistry with three different monoclonal antibodies against proliferating cell nuclear antigen (PCNA)/cyclin (19A2; 19F4; PC10). PC10 was found to work best; it is applicable to both alcohol- and formalin-fixed tumor samples. PCNA labeling indices (LIs) were compared in the same tumors with LIs obtained by Ki-67 immunostaining of frozen sections and by in vitro incubation with bromodeoxyuridine (BrdUrd); in the latter preparations, BrdUrd LIs could be compared with PCNA LIs in the very same areas of serial sections. In gliomas, PCNA LIs were 0.7–80.2% (mean 31.7%), in metastases 0–76.0% (mean 47.8%), and in meningiomas 0–53.0% (mean 19.3%). In general, PCNA LIs were highly significantly correlated with Ki-67 LIs (P=0.0002) and BrdUrd LIs (P=0.0001). However, when tumor subgroups are considered, only gliomas show a significant correlation with Ki-67 and BrdUrd LIs. Despite this statistical correlation, PCNA expression was out of proportion to proliferation indices as determined by both other methods in almost one third of all brain tumors. Immunocytochemistry for PCNA produces a broad spectrum of staining intensity of labeled nuclei, whose number is dependent upon the sensitivity of the immunocytochemical technique used. Thus, inter-oberserver and inter-laboratory variabilities in PCNA LI determination may occur. Overlapping of PCNA LIs between tumor subgroups of varying malignancy further limits the informational value for the individual case. In some classic meningiomas, high PCNA scores do not reflect the proliferative activity of the tumor, as Ki-67 and BrdUrd LIs are very low in these cases. We conclude that PCNA immunolabeling is of limited value in the individual tumor, mainly due to overexpression in many tumors, and at present cannot be recommended to replace Ki-67 and/or BrdUrd labeling methods for routine determination of proliferative activity in human tumor specimens.Supported in part by a grant from the Oncology Commission, Medical Faculty, University of Vienna     

脑肿瘤中P53蛋白的表达及Ki－67LI的相关性研究

利用突变型Ｐ５３蛋白的单抗及Ｋｉ－６７单抗对５０例冰冻标本进行检测，结果发现，脑肿瘤中突变型Ｐ５３蛋白的表达阳性率为４６％。低恶度胶质瘤、高恶度胶质瘤及转移癌中Ｐ５３蛋白表达的阳性率不同，分别为１８．２％、５３．８％及１００％。Ｐ５３蛋白表达阳性的肿瘤中高Ｋｉ－６７ＬＩ比例为７０％，其中位数及均数为１４．３％、２０．７６±１８．３（％），而无Ｐ５３蛋白表达的脑瘤中，ｋｉ－６７ＬＩ中位数及均数分别为１．３９及５．１９±９．０（％）。结果表明，突变型Ｐ５３蛋白的表达与脑肿瘤的组织类型，分化程度及细胞的增殖有关，而它的高表达又可能是肿瘤恶性表型或转移的标志之一。