人脊索瘤细胞系CM-3l9的建立及其生物学特性的观察

目的 建立人脊索瘤细胞系，为脊索瘤研究提供实验型。方法 取经病理证实的新鲜脊索瘤手术标本，进行体外原代组织块培养。对存活细胞进行形态学观察、组织化学染色、细胞周期检查、染色体分析、电镜观察、异种移植和体外侵袭实验等。结果 建成细胞系CM-319，经近两年的体外培养，已连续传代百余次，其形态学表现、组织化学染色、电镜观察和异种移值等均符合脊索瘤细胞特征。细胞倍增时间为33h。细胞周期测定显示：G1期为55．6％，G2期为21．9％，S期为22．5％，G2／G1为1．90。染色体具有亚三倍体核型。异种移植成瘤率100％，具有侵袭性。结论 CM-319是一株人脊索瘤细胞系，可用于对脊索瘤的研究。     

小鼠子宫颈癌U14细胞系的建立及生物学特性研究

目的:建立小鼠宫颈癌肿瘤细胞系,观察其生物学特性。方法:采用肿瘤原代培养法对小鼠U14肿瘤进行体外传代培养,定期对培养细胞进行形态学观察,进行免疫组织化学鉴定、细胞周期检查、染色体检测、细胞生长测定,进行615、C57BL/c小鼠移植观察体内成瘤情况,并用pEGFP-N1质粒转染U14细胞。结果:细胞呈贴壁和悬浮混合生长,CK阳性特征。体外连续培养10个月,传代50代,细胞倍增时间为21.83小时,细胞周期测定G1期为34%,G2期为26.4%,S期为39.6%。培养U14细胞3-4天处于对数生长期,染色体为亚四倍体核型,众数为64～84条。615小鼠、C57BL/c小鼠移植成瘤率均为100%,无支原体污染。建立了GFP( )的U14-GFP单克隆细胞株。结论:成功建立了小鼠U14子宫颈癌细胞系及GFP标记的U14-GFP细胞株,体内移植可建立常规及带荧光标记的肿瘤模型,可用于体内体外相结合的肿瘤研究。     

人骨肉瘤细胞体外培养与桂皮酸诱导分化的实验研究

目的:体外培养手术切取人骨肉瘤标本并由植物成分桂皮酸(cinnamic acid,CINN)处理,研究其对肿瘤细胞增殖的抑制作用及诱导分化作用.方法:应用原代法培养手术切除标本,传代至第10代获得相当的细胞量后,设立对照组使用CINN处理,对各组骨肉瘤细胞进行形态学观察,组织化学染色,免疫组织化学染色,流式细胞仪(FCM)检测细胞周期和裸鼠移植瘤试验.结果:以浓度为2×10-3 mol/L的CINN处理人骨肉瘤细胞,其存活细胞形态学观察和染色结果均显示分化改变,增殖抑制率第2天为23.3%,第4天为41.7%,第6天为50.6%.FCM检测显示,对照组细胞G1期72.5%,G2期24.9%,S期2.6%;实验组细胞G1期60.9%,G2期9.6%,S期29.5%.处理后的细胞裸鼠移植瘤生长较对照组缓慢,切片呈现大片坏死.结论:原代法体外培养人骨肉瘤细胞成功,体外实验CINN有抑制骨肉瘤细胞增殖和诱导其分化的作用.     

小鼠子宫颈癌U14细胞系的建立及生物学特性研究

目的：建立小鼠宫颈癌肿瘤细胞系。观察其生物学特性。方法：采用肿瘤原代培养法对小鼠U14肿瘤进行体外传代培养，定期对培养细胞进行形态学观察．进行免疫组织化学鉴定、细胞周期检查、染色体检测、细胞生长测定，进行615、C57BL／6j小鼠移植观察体内成瘤情况，并用pEGFP-N1质粒转染U14细胞。结果：细胞呈贴壁和悬浮混合生长，CK阳性特征。体外连续培养10个月，传代50代，细胞倍增时间为21．83h，细胞周期测定G1期为34％，G2期为26．4％，S期为39．6％。培养U14细胞3-4天处于对数生长期，染色体为亚四倍体核型，众数为64～84条。615小鼠、C57BL／6j小鼠移植成瘤率均为100％．无支原体污染。建立了GFP（＋）的U14-GFP单克隆细胞株。结论：成功建立了小鼠U14子宫颈癌细胞系及GFP标记的U14-GFP细胞株，体内移植可建立常规及带荧光标记的肿瘤模型，可用于体内体外相结合的肿瘤研究。     

小鼠乳腺癌细胞系Ca761-03的建立及其生物学特性研究

目的：建立体外培养的小鼠乳腺癌细胞系，明确其生物学特性。方法：利用乳腺癌移植瘤组织，进行体外原代培养，并对其纯化和反复传代，进行生长曲线，倍增时间，软琼脂集落形成，细胞周期，染色体众数，免疫组织化学染色及体内成瘤率，转移率等鉴定。结果：建立了一个小鼠乳腺癌细胞系，命名为Ca761-03。其生长迅速，呈贴壁生长。倍增时间为25．98h；平均软琼脂克隆形成率为5．39％；G1期31.8％，S期57.3％，G2＋M期10．9％；CK染色为阳性，ER，PR均为阴性；体内移植成瘤率100％，肺转移率100％，未见淋巴结转移。结论：成功建立了小鼠乳腺癌细胞系．并对其生物学特性进行了较系统的观察与研究。     

裸小鼠高分化食管鳞状细胞癌株(RE25/N)的建立及其生物学特性的研究

大鼠食管癌裸小鼠细胞株在国内外均很少见,本实验室于1986年11月开始,将体外恶性转化的大鼠食管癌细胞系(RE_(25-3))接种于Swiss及NC两种裸小鼠皮下,并传代获得成功,至今已传至14代,该移植瘤具有生长稳定、接种成功率高(100％),潜伏期短及无自发消退等特点。形态学观察:光镜下肿瘤为高分化鳞癌;电镜观察,超微结构符合高分化鳞癌的改变。流式细胞仪(FCM)的DNA分析,其DNA含量的DI值与其在体外培养的母细胞系相似,但细胞周期各期细胞的比例有明显差异。在裸小鼠体内生长的移植瘤,其S期及G_(2+)M期细胞比例明显降低,说明体内生长的移植瘤其生长增殖能力较体外培养的细胞系为低,生长较慢,可能与宿主对移植瘤的作用有关。     

人肺腺癌细胞系Ch-Huang-1的建立和生物学特性研究

背景与目的肺癌是严重威胁人类生存和发展的恶性疾病之一,本研究旨在建立人肺腺癌细胞系,探讨其生物学特性,为进一步明确肺癌的发生机理,提供一个技术平台。方法取人肺腺癌新鲜手术肿瘤标本,经组织块体外培养并克隆建系,命名为Ch-Huang-1。采用光镜、细胞生长曲线、染色体核型分析、克隆形成率及免疫缺陷小鼠成瘤实验对细胞系生物学特性进行分析。结果细胞系及移植瘤标本证实具有腺癌恶性细胞特征。生长曲线呈S型,细胞群体倍增时间为36 h。分裂指数曲线为倒V字型,培养第30小时处于分裂期的细胞最多。克隆形成率为0.803%±0.078%。观察到肿瘤细胞染色体数目为亚三倍体,有明显的染色体异常。裸鼠皮下异种移植形成移植瘤。结论根据细胞系特征显示该细胞系是一新建的肺腺癌细胞系。     

二烯丙基二硫对裸鼠体内人结肠癌细胞增殖的影响

背景与目的:二烯丙基二硫(diallyl disulfide,DADS)在体外可抑制多种肿瘤细胞增殖,但在体内抗肿瘤作用的研究报道较少.本实验旨在研究DADS对裸鼠体内人结肠癌SW480细胞增殖的影响.方法:建立人结肠癌裸鼠移植瘤模型,实验动物分为DADS组及对照组.观察DADS作用后移植瘤体积和重量的变化,通过光镜和电镜观察移植瘤细胞形态学变化,通过免疫组织化学和形态定量图像分析系统检测瘤组织内增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)蛋白表达变化,用流式细胞术检测移植瘤细胞周期分布.结果:DADS在体内能明显抑制移植瘤的生长,相对肿瘤增殖率为49.85%;组织学分析显示DADS组瘤细胞异型性降低,核质比下降,胞浆内细胞器丰富,部分胞核呈退行性变,可见凋亡小体;免疫组化示两组移植瘤组织PCNA蛋白均有阳性表达,DADS组(149.02±4.26)与对照组(178.86±7.69)比较,裸鼠移植瘤组织内PCNA蛋白表达明显减弱(P＜0.05);流式细胞术分析显示,对照组移植瘤细胞中G2/M期细胞为18.8%,与对照组比较,DADS组G2/M期细胞(38.6%)显著增多.结论:DADS可体内抑制人结肠癌SW480细胞增殖,使癌细胞阻滞于G2/M期.     

胃癌腹膜高转移潜能细胞系的建立及其生物学特性

目的利用人胃癌细胞系(GC9811)在裸小鼠体内反复接种建立一株具有腹膜高转移潜能的胃癌细胞系(GC9811P),并对其生物学特性进行观察,为实验研究提供模型。方法采用胃癌细胞系(GC9811)在裸小鼠腹腔内反复接种,行体外培养腹膜转移灶筛选高转移亚系,绘制细胞生长曲线,光镜、电镜下观察细胞形态,利用流式细胞仪、染色体分析等方法,研究该高转移亚系的细胞周期、增殖、染色体核型等生物学特性。结果母本细胞系GC9811腹膜转移形成率为33.3%(3/10),而GC9811P腹膜转移形成率为100%。两种细胞系腹膜结节组织学形态大体相似。增殖速度较母系加快。细胞周期分析G1期53.5%、G2期12.5%、S期37.1%。且遗传学特性包括染色体形态仍为人类核型,众数维持在104～126之间,占70%。结论具有腹膜高转移的胃癌细胞系GC9811P的建立及裸小鼠体内实验模型,为研究胃癌腹膜转移机制及探索新的治疗途径提供了极为有用的工具。     

C2-神经酰胺对间皮瘤细胞裸鼠移植瘤生长的抑制作用

目的：采用人工合成的外源性C2-神经酰胺研究其对体外培养的间皮瘤细胞和体内移植瘤的移植作用,为是否能将 C2-神经酰胺作为治疗人恶性间皮瘤的新化疗药提供初步理论依据。方法：在体外实验中,C2-神经酰胺作用于人恶性间皮瘤细胞后,采用CCK-8法测定细胞增殖中活细胞数目并制作细胞剂量和时间的生长曲线。荧光激活流式细胞分离术（FACS）观察间皮瘤细胞周期的变化。DNA ladder对凋亡细胞进行分析。Fluorescent assay法检测间皮瘤细胞的Caspase-3活性。在体内实验中,建立荷瘤裸鼠动物模型。随机分为四组,C2-神经酰胺腹腔注射组、C2-神经酰胺皮下注射组、DMSO 组和生理盐水组。观察治疗后裸鼠自然状态、重量,计算肿瘤体积,21天时处死裸鼠,肉眼及镜下观察。免疫组织化学染色观察C2-ceram-ide对人间皮瘤细胞的增殖、凋亡和侵袭的影响。结果：体外实验中,在一定的时间与剂量范围内,C2-神经酰胺对间皮瘤细胞的抑制呈现剂量依从和时间依从性。FACS及DNA电泳细胞凋亡分析均表明,C2-神经酰胺在体外可诱导间皮瘤细胞的凋亡。FACS检测到G0/G1期细胞明显减少,S期细胞增多,细胞周期阻滞于G2/M期。Caspase-3活性测定发现C2-神经酰胺能增加间皮瘤细胞Caspase-3的活性。体内实验中,C2-神经酰胺组裸鼠肿瘤的重量、体积均小于未治疗组。C2-神经酰胺组肿瘤组织 Caspase-3蛋白表达明显强于未治疗组。结论：本实验证实C2-ceramide在体内、外均可抑制恶性间皮瘤细胞增殖、侵袭,诱导间皮瘤细胞凋亡,进一步了解间皮瘤细胞的生物学特性,为临床试验奠定了一定的基础。     

hTERT反义寡核苷酸对脊索瘤细胞周期及增殖的影响

目的：探讨hTERT反义寡核苷酸（ASODN）抑制CM-319细胞端粒酶活性对脊索瘤细胞的细胞周期及增殖的影响。方法：用脂质体介导hTERT正义寡核苷酸（SODN）和反义寡核苷酸（ASODN）转染CM-319脊索瘤细胞株72小时后,MTT法检测细胞活性,流式细胞仪检测细胞周期及细胞凋亡率变化。结果：hTERT的ASODN转染细胞的端粒酶活性及细胞生长都明显受到抑制。流式细胞仪检测显示：ASODN组细胞出现早期凋亡峰,细胞阻滞于S期,而hTERT的SODN则无此作用。ASODN组细胞凋亡率为16.01%,而hTERT的SODN组为2.3%。结论：端粒酶hTERT为靶点的反义寡核苷酸可明显抑制脊索瘤细胞的增殖,且具有促凋亡作用。     

hTERT反义寡核苷酸对脊索瘤细胞周期及增殖的影响

目的:探讨hTERT反义寡核苷酸(ASODN)抑制CM-319细胞端粒酶活性对脊索瘤细胞的细胞周期及增殖的影响。方法:用脂质体介导hTERT正义寡核苷酸(SODN)和反义寡核苷酸(ASODN)转染CM-319脊索瘤细胞株72小时后,MTT法检测细胞活性,流式细胞仪检测细胞周期及细胞凋亡率变化。结果:hTERT的ASODN转染细胞的端粒酶活性及细胞生长都明显受到抑制。流式细胞仪检测显示:ASODN组细胞出现早期凋亡峰,细胞阻滞于S期,而hTERT的SODN则无此作用。ASODN组细胞凋亡率为16.01%,而hTERT的SODN组为2.3%。结论:端粒酶hTERT为靶点的反义寡核苷酸可明显抑制脊索瘤细胞的增殖,且具有促凋亡作用。     

人卵巢肉瘤样癌细胞系BUPH:OVSC-1的建立及其生物学特性

目的:介绍人卵巢肉瘤样癌细胞系的建立,研究其生物学特性.方法:以手术切除的卵巢肉瘤样癌腹壁转移组织为材料,进行体外培养,建立人卵巢肉瘤样癌细胞系.通过光学显微镜、电子显微镜、生长曲线测定、染色体分析、双层软琼脂培养、裸鼠接种、免疫组化等,对其生物学特性进行研究.结果:首次建立一株人卵巢肉瘤样癌细胞系,命名为BUPH:OVSC-1,现已传至100余代.其生物学特性为,形态学观察细胞呈肉瘤样细胞形态,超微结构证实为上皮起源;细胞生长旺盛;染色体为假二倍体,染色体丢失,且有三条标记染色体;具有集落形成能力;裸鼠皮下种植瘤生长快,与原组织形态相似;组织化学奥辛蓝过碘酸雪夫氏反应、免疫组化角蛋白7及波形蛋白染色均阳性.结论:BUPH:OVSC-1为一株高度恶性人卵巢癌细胞系,符合人卵巢肉瘤样癌细胞系的特征,可作为卵巢肉瘤样癌研究的实验模型.     

Establishment of a human pulmonary adenocarcinoma cell line Lu-YePa and its biological characteristics

This paper presents the establishment of a human pulmonary adenocarcinoma cell line Lu-YePa and its biological characteristics. The primary cells of Lu-YePa cell line were derived from a twenty six year old patient with poorly differentiated adenocarcinoma of the lung. After 36 days' culture, transfer was made and the cells proliferated steadily and rapidly afterwards. The doubling time was 34.6 hours. Mitotic index reached 49.5% on day 5. Chromosome number was subtriploid with the mode of 62. The large submetacentric marker chromosome was presented in 68% of cells. PAS positive granules were found in the cytoplasm. The study of concanavalin A condensation, electron microscopic observation, heterotransplantation and plating efficiency indicates that Lu-YePa cells are concordant with the human pulmonary adenocarcinoma cells in morphology and biological behavior.     

Establishment of a human plasma cell line in vitro

A human plasma cell line designated ARH-77 has been established and propagated in culture for the past 2 years. The cells exhibited morphological characteristics of plasma cells under light and electron microscopic examination. An average of 40% cells are positive for immunoglobulin G by direct immunofluorescence, while an immunoglobulin G-specific radioimmunoassay reveals the production of 1.21 X 10(4) ng/10(6) plasma cells. The karyotype is aneuploid with a a modal chromosome number of 45 to 46 and no marker chromosome. Growth kinetics characteristics are: doubling time, 110.4 hr; generation time, 56.4 hr; G1 + G2-phase transit time, 45.5 hr; S-phase transit time, 10.9 hr; growth fraction, 74%; mitotic index, 1.5%; labeling index, 14.3%; and cell loss, 31.0%. Some of the growth kinetics characteristics were markedly similar to the properties displayed in vivo by plasma cells of patients with multiple myeloma and suggest that the cell line might be a useful in vitro model for the study of human myeloma.     

Multidirectional differentiation in a newly established human epithelioid sarcoma cell line (GRU-1) with co-expression of vimentin, cytokeratins and neurofilament proteins.

A new permanent cell line (GRU-1) derived from the lymph-node metastasis of a human epithelioid sarcoma was established in tissue culture. Immunohistochemically, the original tumor had exhibited an intriguing potential for multidirectional differentiation with features of mesenchymal, epithelial and neural differentiation, evidenced by the co-expression of vimentin, cytokeratins and neurofilament proteins, respectively. This capability for multidirectional differentiation was fully preserved in the cultured cells. GRU-1 tumor cells proved to be uniformly positive for vimentin and a considerable proportion of the tumor cells exhibited a positive reaction for cytokeratins and neurofilament proteins. The neural markers neuron-specific enolase (NSE) and synaptophysin were observed in a small proportion of GRU-1 cells. Ultrastructurally, GRU-1 cells showed desmoplastic activity in vitro, being enmeshed by collagen fibrils. DNA distribution, as studied by flow cytophotometry, revealed DNA-diploidy (DNA index = 1) and a G0/G1-proportion of 70.5%. After heterotransplantation in nude mice, GRU-1 tumor cells expressed vimentin and cytokeratin only, whereas the neural markers could not be further demonstrated.     

抗人脊索瘤杂交瘤细胞系的建立及其单抗在病理诊断中的应用

目的:建立一株高特异性抗人脊索瘤单克隆抗体的杂交瘤细胞系,检测其在人脊索瘤组织中及其它骨肿瘤中的特异结合程度.方法:以人脊索瘤细胞株CM-319为抗原,免疫BALB/C小鼠,利用杂交瘤技术,研制出一株MAb-6F2杂交瘤细胞系.通过免疫组织化学方法检测了MAb-6F2在4种不同骨肿瘤石蜡切片和人正常髓核组织上的结合反应.结果:MAb-6F2在骨肉瘤组织上的结合率仅为10%;在软骨肉瘤上的结合率为10%;在骨样骨瘤组织上的结合率为10%;在脊索瘤组织上的结合率为100%;在正常髓核组织上为阴性; MAb-6F2在脊索瘤组织上的结合反应显著高于其它骨肿瘤(P<0.01).结论:MAb-6F2在人脊索瘤组织中的特异性高结合,可以为脊索瘤的诊断提供新的手段.     

Cell culture of small round cell tumor originating in the thoracopulmonary region. Evidence for derivation from a primitive pluripotent cell

The authors describe a 14-year-old girl with small round cell tumor originating in the chest wall analyzed by the extensive studies including light and electron microscopic examination, histochemical study, immunochemical study, cytogenetics, and gene analysis. A cell line producing carcinoembryonic antigen (CEA) and neuron-specific enolase (NSE) has been established from pleural effusion of the pulmonary metastatic tumor. Cytogenetic analysis disclosed a reciprocal translocation (11;22)(q24;q12). Additionally, immunocytochemical studies demonstrated that CEA, NSE, vimentin, cytokeratin, and epithelial membrane antigens are positive, but desmin and S-100 protein are negative. Although neurofilament was negative in the pulmonary metastatic tumor cells, it became positive in cell line in vitro. These results suggest that this tumor may be derived from the primitive and pluripotential cells, differentiating into mesenchymal, epithelial, and neural features in variable proportions.     

人卵巢癌永生化细胞系BUPH:OVSC-2的建立及其生物学特性研究

背景与目的：利用永生化技术建立预期表型的细胞系,已成为建设系的重要手段之一,而人卵巢癌永生化细胞系的建立国内外少有报道。本文介绍人卵巢肉瘤样癌永生化细胞系的建立,以及对其生物学特性的研究。方法：以手术切除的卵巢肉瘤样癌腹壁转移组织为材料,进行体外培养。将永生化基因SV40T抗原基因转染第10代细胞,经筛选、抗性克隆扩大培养,得到永生化细胞系。通过光学显微镜、电子显微镜、生化曲线测定、染色体分析、双层软琼脂培养、裸鼠接种、免疫组化等,研究其生物学特性,并与其来源细胞的生物学特性进行比较。结果：建立一株人卵巢肉瘤样癌永生化细胞系,命名为BUPH：OVSC－2,现已传至90年代。其生物学特性为：形态学观察细胞呈肉瘤样细胞形态,超微结构证实为上皮起源;细胞生长旺盛;具有恶性细胞的特征,恶性度高。通过比较,与其来源细胞的生物学特性无明显差别。结论：BUPH：OVSC－2为一株恶性度高的人卵巢肉瘤样癌永生化细胞系,保留了其来源细胞的生物学特性,可作为卵巢肉瘤样癌研究的实验模型。     

Establishment of a highly metastatic human ovarian cancer cell line (HO-8910PM) and its characterization.

To establish a highly metastatic human ovarian cancer cell line and to study its characteristics tissue from the tumor mass of a nude mouse of 7th subtransplantation of the highly metastasizing human ovarian cancer model (NSMO) was cultured in vitro and the cell line HO-8910PM was established. The cell line grew well through 87 passages and the mitotic index, chromosome analysis, morphology under electron microscope and some oncoprotein expression were studied. The cell doubling time was 34.5 h and the mitotic index was 44.3%. The cells were all of epithelial type and most of them were of polygonal shape. Electron microscopic examination showed malignant nuclei with enlarged nucleoli and abundant microvilli. The plating efficiency in soft agar was 31.2%. The cell agglutination appeared in 4 ug/ml PHA. Chromosomal analysis revealed a mode of 54 per cell. The DNA index was 1.57 measured by FCM. Both of them showed hyperdiploid. Positive ER and PR granules were found in the cells. After hetero-transplantation of the cells into three nude mice all of the latter showed tumor growth with metastasis in lungs or lymph nodes. Eight of the nine kinds of oncoprotein detected by immnohistochemical method were found in the cells. The detection for mycoplasma showed negative. After storage in liquid nitrogen cell growth was stable. The cell line HO-8910PM can meet the criteria for the establishment cell lines. This cell line and NSMO model would be very useful in study of the mechanism of cancer metastasis in identifying various cellular factors regulating local and distant metastasis and also in establishing a rational approach for searching after anti-metastatic agents.