新型钙调蛋白拮抗剂EBB对体外培养胶质母细胞瘤增殖能力的影响

本文探讨了国内首先开发的新型钙调蛋白拮抗剂－０－４－乙氧基－丁基小檗胺（ＥＢＢ）对人脑恶性多形性胶质母细胞瘤体外增殖的影响，发现ＥＢＢ可显著抑制细胞生长、集落形成及蛋白合成，且呈剂量依赖性关系，故值得进一步研究以期开发应用于临床。     

外源性IFN-β基因对裸鼠SHG44胶质瘤的诱导凋亡研究

目的:探讨β干扰素基因对人脑胶质瘤的诱导凋亡作用,以及人β干扰素裸DNA治疗人脑胶质瘤的可行性和作用机制.方法:建立裸鼠SHG44胶质瘤模型,用脂质体包埋法将IFN-β基因的真核表达载体pSV2IFNβ注入裸鼠皮下SHG44脑胶质瘤,观察肿瘤生长情况并计算肿瘤体积,通过免疫组化、TUNEL染色以及电镜,了解IFN-β基因诱导人脑胶质瘤细胞凋亡的作用.结果:IFN-β在荷瘤裸鼠瘤体内获得表达,裸鼠皮下胶质瘤生长受到抑制,并诱导SHG44胶质瘤细胞凋亡.结论:IFN-β裸DNA能够抑制人脑胶质瘤生长并诱导胶质瘤细胞凋亡,该实验为IFN-β基因治疗人脑胶质瘤的应用奠定了初步基础.     

冷冻切除联合化疗治疗脑胶质瘤的临床研究

恶性胶质瘤呈浸润性生长，手术不易彻底切除，术后辅以放疗等效果欠佳，我们应用冷冻手术联合化疗治疗6例成人脑胶质瘤，结果总结报道如下。     

微RNAlet-7d抑制胶质瘤生长和侵袭

背景与目的：let-7d与胶质瘤的关系尚不清楚,本研究探讨let-7d在人脑胶质瘤组织中表达及其对人脑胶质瘤U87细胞生物学特征的影响。方法：原位杂交法检测let-7d在人脑胶质瘤组织和正常脑组织中的表达,利用脂质体将let-7d寡核苷酸转入U87细胞,观察各组细胞生长、凋亡、侵袭情况。结果：人脑胶质瘤组织中let-7d较正常人脑组织低表达,转入let-7d寡核苷酸的U87细胞,生长减缓,凋亡增加,侵袭力减低。结论：微RNAlet-7d在胶质瘤的发生发展中可能起抑制作用。     

基因工程改造鼠源性抗人脑胶质瘤单克隆抗体的初步报告

为了提高人脑胶质瘤导向诊治中单抗载体的临床实用价值，必须把鼠源性大分子单抗改造成人源化、小分子单抗。采用ＲＴ－ＰＣＲ法，从人脑胶质瘤单抗杂交瘤细胞系ＳＺ３９中克隆出３４８ｂｐ的重链可变区（ＶＨ）和３１８ｂｐ的轻链可变区（ＶＬ）ｃＤＮＡ片断。又采用重组ＤＮＡ技术，将ＶＨ和ＶＬ与人免疫球蛋白ＩｇＧ１重链ＣＨ１和轻链κ恒定区基因拼接，或将ＶＨ和ＶＬ以通用Ｌｉｎｋｅｒ直接连接，分别构建表达载体ｐＨＥＮ１－ＳＺ３９Ｆａｂ／Ｈｕ（嵌合抗体）和ｐＨＥＮ１－ＳＺ３９ＳｃＦｖ（单链抗体）。而后将此二载体转染至非抑制性大肠杆菌ＨＢ２１５１中表达。结果表明，两者的表达产物均为可溶性，以ＥＬＩＳＡ和Ｗｅｓｔ－ｅｒｎｂｌｏｔ法检测，均与人脑胶质瘤体外细胞系ＳＨＧ４４细胞膜表面抗原发生特异结合，与亲本抗体ＳＺ３９特异识别１８００００膜抗原的条带相一致。该结果显示，基因工程改造的人源化、小分子单抗为今后的临床应用展示了美好的前景。     

人脑胶质瘤与SV40的关系及其p53、RB蛋白的表达

目的：探讨人脑胶质瘤与猴病毒４０（ＳＶ４０）的关系及其ｐ５３和ＲＢ蛋白的表达。方法：采用核酸原位杂交和免疫组织化学技术对２５６例人脑胶质瘤和１１例正常脑组织进行了ＳＶ４０ ＤＮＡ和抑癌基因产物ｐ５３、ＲＢ蛋白检测。结果：胶质瘤ＳＶ４０阳性１０４例,阳性率４０．６％,正常脑组织均未检测出ＳＶ４０ ＤＮＡ,未发现ＳＶ４０感染与胶质瘤病理分级存在明显相关（Ｐ〉０．０５）;ＳＶ４０阳性胶质瘤ｐ５３、ＲＢ蛋     

逆转录病毒载体表达的TGFα反义RNA对人脑胶质瘤细胞BT325的生长抑制作用

逆转录病毒载体表达的ＴＧＦα反义ＲＮＡ对人脑胶质瘤细胞ＢＴ３２５的生长抑制作用惠宏襄，王成济，金明，赵小宁，胡敏西安第四军医大学（７１００３２）转化生长因子α（ＴｒａｎｓｆｏｒｍｉｎｇＧｒｏｗｔｈＦａｃｔｏｒ，ＴＧＦα）是一种小肽分子，其成熟体由５０...     

IFN-β基因诱导裸鼠胶质瘤Fas表达上调及细胞调亡的实验研究

目的研究β干扰素基因对人脑胶质瘤的诱导凋亡作用,以及能否诱导胶质瘤细胞Fas表达上调,并探讨了β干扰素基因诱导细胞凋亡及Fas表达上调之间的联系.方法建立裸鼠SHG44胶质瘤模型,用脂质体包埋法将IFN-β基因的真核表达载体pSV2IFNβ注入裸鼠皮下SHG44脑胶质瘤,观察肿瘤生长情况并计算肿瘤体积,通过免疫组化、TUNEL染色,了解IFN-β基因诱导人脑胶质瘤细胞凋亡的作用以及胶质瘤细胞Fas表达情况.结果 IFN-β在荷瘤裸鼠瘤体内获得表达,裸鼠皮下胶质瘤生长受到抑制,并诱导SHG44胶质瘤细胞凋亡,胶质瘤细胞Fas表达明显高于对照组.结论 IFN-β基因能够抑制人脑胶质瘤生长,并诱导胶质瘤细胞凋亡;IFN-β基因诱导胶质瘤细胞凋亡可能是Fas表达上调的结果.     

IFN-β基因诱导裸鼠胶质瘤Fas 表达上调及细胞凋亡的实验研究

 目的 研究β干扰素基因对人脑胶质瘤的诱导凋亡作用,以及能否诱导胶质瘤细胞Fas表达上调,并探讨了β干扰素基因诱导细胞凋亡及Fas表达上调之间的联系. 方法 建立裸鼠SHG44胶质瘤模型,用脂质体包埋法将IFN-β基因的真核表达载体pSV2IFNβ注入裸鼠皮下SHG44脑胶质瘤,观察肿瘤生长情况并计算肿瘤体积,通过免疫组化、TUNEL染色,了解IFN-β基因诱导人脑胶质瘤细胞凋亡的作用以及胶质瘤细胞Fas表达情况. 结果 IFN-β在荷瘤裸鼠瘤体内获得表达,裸鼠皮下胶质瘤生长受到抑制,并诱导SHG44胶质瘤细胞凋亡,胶质瘤细胞Fas表达明显高于对照组. 结论 IFN-β基因能够抑制人脑胶质瘤生长,并诱导胶质瘤细胞凋亡;IFN-β基因诱导胶质瘤细胞凋亡可能是Fas表达上调的结果.     

人脑胶质瘤细胞原代培养实验模型的构建

目的:探索人脑胶质瘤细胞原代培养的方法,提高原代培养的成功率,快速构建人脑胶质瘤细胞体外试验模型。方法:分别采用组织块培养法和酶消化法对12例人脑胶质瘤细胞进行原代培养,比较两种方法的培养效果;通过倒置相差显微镜和激光共聚焦显微镜观察细胞的形态学特点;通过生长曲线的测定研究体外培养细胞的生长特性。结果:原代人脑胶质瘤细胞短期培养成功,并可传代。经组织块培养法成功率较低为33.3%;经酶消化法短期培养成功率为83.3%,明显高于组织块培养法。培养成功的细胞状态稳定,增殖活跃,有明显的对数生长期。结论:用酶消化法可进行人脑胶质瘤短期体外培养,且成功率较高,适宜建立体外细胞培养模型。     

新型钙调素拮抗剂对体外人脑恶性星形细胞瘤细胞增殖的抑制

采用生长曲线、集落形成试验、蛋白含量测定、超微结构检查、流式细胞术及~3H-TdR掺入试验,观察了新型钙调素持抗剂O-4-Z氧基-丁基-小檗胺[O-(4-ethoxyl-butyl),berb-amine,EBB]对TJ861人脑恶性星形细胞瘤体外细胞系增殖的影响,发现EBB抑制这种细胞的增殖。EBB与化疗药BCNU联合应用有协同作用。     

Influence of the Calmodulin Antagonist EBB on Cyclin B1 and Cdc2-p34 in Human Drug-resistant Breast Cancer MCF-7/ADR Cells

OBJECTIVE To investigate the influence of O-(4-ethoxyl-butyl)- berbamine(EBB)on the expression of cyclin B1 and cdc2-p34 in the human drug-resistant breast cancer MCF-7/ADR cell line. METHODS The MTT assay was used to assess the cytotoxicity of EBB.Different levels of EBB were added to different cell lines at series of time points solely or combined with doxorubicin(DOX) to detect the effect on the expression of cyclinB1 and cdc2-p34 by Western blots.cdc2-p34 tyrosine phosphorylation was detected by immunoprecipitation.In addition,apoptosis and cytoplastic Ca 2 concentrations were systematically examined by laser scanning confocal microscopy(LSCM). RESULTS EBB showed little inhibitory activity on human umbilical vein endothelial cells(ECV304),whereas EBB inhibited cell growth(IC50 range,4.55~15.74μmol/L)in a variety of sensitive and drug-resistance cell lines.EBB also down-regulated the expression of cyclin B1 and cdc2-p34 in a concentration and time dependent manner,which was an important reason for the G2/M phase arrest.EBB was shown to induce apoptosis of MCF-7/ADR cells while increasing the level of cytoplastic Ca 2 . CONCLUSION The low cytotoxicity of EBB suggests it may be useful as a rational reversal agent.The effect of EBB on cell cycle arrest and related proteins,apoptosis,and cytoplastic Ca 2 concentration may be involved in reversing multidrug resistance.     

In vitro and in vivo growth inhibition of human malignant astrocytoma cells by the farnesyltransferase inhibitor B1620

p21-Ras, the protein product of the proto-oncogene Ras is overactivated in malignant astrocytomas despite the absence of mutation. It is known that p21-Ras participates in signaling events from membrane tyrosine kinase receptors and a variety of intracellular biochemical pathways to downstream targets. Signal transduction inhibition by targeting against Ras is now thought to be a promising therapeutic strategy for malignant astrocytomas. This study demonstrates that Ras pathway inactivation by a farnesyltransferase inhibitor, B1620, effectively inhibits in vitro and in vivo growth of human astrocytoma cells, although normal human astrocytes (NHA) derived from fetal brain are resistant to B1620. Anti-proliferative effect of B1620 on in vitro growth of astrocytoma cells was examined by MTT assays and soft agar colony formation assay. B1620 inhibited anchorage-dependent growth of six astrocytoma cell lines with a median effective dose (IC₅₀) ranging from 2.0 to 20.7µM. However, growth of NHA was not significantly affected by B1620 even at the concentration of 100µM. All astrocytoma cells showed apoptotic figures after Hoechst 33258 staining, when treated for 5 days at each IC₅₀ concentration against B1620. Anchorage-independent growth of these astrocytoma cell lines was inhibited at a much lower concentration than that of anchorage-dependent growth. Daily treatment of U87 xenograft-bearing athymic mice with B1620 at 100 or 50mgkg^–1 resulted in significant inhibition of tumor growth. A histological study of the B1620-treated tumor tissue showed decreased vascularity with numerous TUNEL-positive apoptotic cells. These results suggest that the mechanism of the growth-inhibitory effect of B1620 is anti-angiogenesis, apoptosis induction and reversion of the transformed phenotype. The potential clinical use of B1620 could be expanded to malignant astrocytomas.     

阿司匹林对胶质瘤细胞生长增殖及NOS表达的影响

 目的　观察阿司匹林对胶质瘤细胞生长增殖、NOS表达及NO、CEA含量的影响并探讨其作用机制。方法　应用MTT法测定不同浓度阿司匹林对胶质瘤细胞的生长抑制率 ;免疫组化法检测NOS的表达 ;Griess法测定培养基中NO含量 ;微粒子捕捉免疫法测定CEA。结果　阿司匹林对胶质瘤细胞具有抑制作用 ,提高诱导型NOS(iNOS)的表达 ,增加NO的生成 ,并呈浓度依赖关系 ;同时具有降低CEA的作用。结论　阿司匹林可抑制胶质瘤细胞的生长增殖。提高iNOS的表达 ,增加NO含量可能是其部分机制 ,CEA可作为监测胶质瘤疗效的有效指标。     

Increased intracellular cyclic AMP levels suppress the mitogenic responses of human astrocytoma cells to growth factors

Summary It has been shown that the intracellular cAMP levels were decreased in human malignant astrocytomas. On the other hand, various growth factors and their receptors were found to be overexpressed in these tumors. It is therefore intriguing as to whether there is interplay between the two phenomena in the modulation of the astrocytoma cell growth. In a basal medium consisting of 75% DMEM, 25% Ham's F-12 supplemented with 2% FBS, we show that the mitogenic effects of platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), and epidermal growth factor (EGF) on human astrocytoma cells were suppressed by dibutyryl-cAMP. Dibutyryl-cAMP alone neither potentiated nor inhibited the tumor cell growth. Further studies show that PDGF-induced receptor autophosphorylation in human astrocytoma cells is suppressed by increased intracellular cAMP levels as measured by immunoprecipitation with anti-PDGF receptor and antiphosphotyrosine antibodies. Our results indicate that there is antagonistic interplay between the receptor tyrosine kinase pathway and cAMP-dependent protein kinase pathway in the control of the malignantly transformed glial cells. A reduced cAMP level seen in many human astrocytoma cells may favor their response to growth factor mitogenesis.     

Neuritin expression and its relation with proliferation, apoptosis, and angiogenesis in human astrocytoma

Neuritin, a new member of the neurotrophic factor family, plays an important role in promoting neuronal survival, differentiation, function, and repair. However, whether neuritin is expressed in human astrocytoma and involved in their proliferation, apoptosis, and angiogenesis remains unclear. The expression of neuritin messenger RNA, protein and the relationship with proliferation, apoptosis, and angiogenesis were examined in human astrocytoma samples and three glioma cell lines by immunohistochemistry, Western blot, and quantitative real-time RT–PCR and so on. And neuritin immunoreactivity score(IRS), proliferative index (PI), apoptotic index (AI), overall daily growth (ODG), and microvessel density (MVD) in brain astrocytoma were measured. The results showed that neuritin was overexpressed in human astrocytoma samples, and the overexpression correlated positively with the malignancy of astrocytomas as reflected by changes in proliferation, apoptosis, and angiogenesis markers. In our study, we found neuritin is overexpressed in astrocytoma, which may be an important factor in tumorigenesis and progression of astrocytoma, and can be used as a target for biological therapy.     

细胞内p53蛋白积聚：人脑星形细胞瘤恶性进展的因素之一

用单克隆抗体Pabl 801行免疫组化研究48例不同恶性程度的人脑星形细胞瘤,发现17例(35.4%)p53蛋白染色阳性,其中核阳性者12例,胞浆阳性者5例。30例高恶度的星形细胞瘤中有14例(47%)阳性,其余18例低恶度的肿瘤有3例(17%)阳性,二者具有显著性差异。高恶度的肿瘤中p53染色阳性细胞百分比亦高于低恶度的肿瘤。该结果提示细胞内p53蛋白积聚是人脑星形细胞瘤由低恶度进展到高恶度的一个重要因素。     

Differential effects of methionine enkephalin on the growth of brain tumor cells

Summary The effect of methionine enkephalin on the growth of human brain tumor cells was investigated. The results show that this endogenous opioid has dual effects on tumor cell growth. This peptide exerted an inhibitory effect in SK-N-MC human neuroblastoma cell line; in contrast, in U-373 MG human astrocytoma cell line, the peptide showed a stimulatory effect. Treatment with naltrexone, an opioid receptor antagonist, also resulted in a similar alteration of tumor cell growth implicating that its action may be unrelated to opioid receptor blockade. These results suggest that in these tumor cell lines endogenous opioid systems may be involved in cell proliferation. Furthermore, these tumor cell lines may be useful model systems for the study of the signal transduction mechanisms of endogenous opioids.