ANALYSIS OF THE PROLIFERATION KINETICS OF BURKITT'S LYMPHOMA CELLS

The in vitro proliferation kinetics of a cell line derived from a patient with American Burkitt's lymphoma were investigated at three different growth phases: lag (day 1), exponential (day 3) and plateau (day 5). The growth curve, labeling and mitotic indices, percentage labeled mitosis (PLM) curves and DNA content distributions were determined. The data obtained have been analysed by the previously developed discrete-time kinetic (DTK) model by which a time course of DNA distributions during a 10-day growth period was characterized in terms of other cell kinetic parameters. The mean cell cycle times, initially estimated from PLM curves on days 1, 3 and 5, were further analysed by the DTK model of DNA distributions and subsequently the mean cell cycle times with respect to DNA distributions during the entire growth period were determined. The doubling times were 39·6, 31·2 and 67·2 hr, respectively, at days 1, 3 and 5. The mean cell cycle time increased from 23·0 to 37·7 hr from day 3 to day 5 mainly due to an elongation of the G₁ and G₂ phases. A slight increase in the cell loss rate from 0·0077 to 0·0081 fraction/hr was accompanied by a decrease in the cell production rate from 0·0299 to 0·0184 fraction/hr. This calculated cell loss rate correlated significantly with the number of dead cells determined by trypan blue exclusion. Analysis of the number of dead cells in relation to the cell cycle stage revealed that a majority of cell death occurred in G₁ (r= 0·908; P < 0·0001). There was a good correlation between the in vitro proliferation kinetics at plateau phase of this Burkitt's lymphoma derived cell line and the in vivo proliferation kinetics of African Burkitt's lymphoma (Iversen et al., 1974), suggesting the potential utility of information obtained by in vitro kinetic studies.     

EZH2蛋白在食管上皮永生化细胞系和恶性转化细胞系中的表达

目的研究EZH2蛋白在食管上皮永生化细胞系(SHEE)和恶性转化细胞系(SHEEC)中的表达。方法采用免疫细胞化学染色、免疫印迹分析和流式细胞术检测两种细胞系EZH2蛋白的表达。结果两种细胞EZH2蛋白染色均呈阳性,阳性反应定位于细胞核,部分细胞胞浆也有阳性着色。免疫印迹分析表明SHEEC细胞和SHEE细胞的总蛋白、核蛋白在分子质量约90ku的位置均出现特异性印迹条带。SHEE细胞中EZH2蛋白的表达强于SHEEC细胞(P<0.05)。流式细胞术显示EZH2蛋白在两种细胞中均有表达,两者的平均荧光强度无明显差别;阳性细胞率均较高,其中SHEE细胞阳性率高于SHEEC细胞。结论EZH2蛋白在SHEE细胞和SHEEC细胞中高表达可能参与了它们的恶性改变;而EZH2蛋白在两种细胞系中的差异表达可能与细胞的分化程度及来源于胚胎食管上皮细胞相关。     

温度对人肺癌细胞 A549蛋白质表达的影响

肺癌蛋白质组的研究,有助于阐明其发病机制并对肺癌的防治有益。本文从研究人肺癌细胞热休克蛋白的表达情况出发,通过比较37℃、42℃和45℃培养条件下的人肺癌细胞A549总蛋白质的双向电泳图谱,获得3个温度敏感的差异蛋白点,依次命名为P_1、P_2、P_3。对差异蛋白进行MALDI-TOF-MS分析和采用SWISS-PROT数据库中的Peptident软件检索后,初步鉴定P_1与2种醛酮还原酶家族成员相匹配,P_2可能为一种新蛋白,P_3为锌指蛋白11A。     

CHANGES IN CELL PROLIFERATION KINETICS OCCURRING DURING THE LIFE HISTORY OF MONOLAYER CULTURES OF A MOUSE TUMOUR CELL LINE

Following inoculation of monolayer cultures of EMT6 mouse tumour cells at 10⁵ cells, a short lag is followed by 3 days of exponential growth with a population doubling time of 12 hr. A plateau cell number is reached between days 4 and 5 and is maintained for at least 8 days. During exponential growth, the pulse ³H-TdR labelling index is 55–60%, all cells are in cycle, and the median cycle time is 11–12 hr. For the first 3 days of plateau phase, the labelling index is about 25 % and there is considerable cell loss. The cell cycle is 32–40 hr, and S-phase is very long. Later in plateau phase, the labelling index falls to <2 % and there is little cell loss. The changes in kinetics occurring in EMT6 cultures are discussed with reference to reported changes occurring in other cell lines.     

ANALYSIS OF THE KINETICS OF GRANULOSA CELL POPULATIONS IN THE MOUSE OVARY

The kinetics of granulosa cell populations in two types of follicles in ovaries of 28-day-old Bagg mice are investigated. The analysis includes estimations of mean values and standard deviations of the transit times (T_G1, T_S, T_G2 and T_C), the doubling time T_D, and the proliferative fraction p. First the percentage of labelled mitosis curve (PLM-curve) and the continuous labelling curve (CL-curve) are estimated. Then a hypothesis concerning the cell kinetics of the granulosa cells in the two follicle types is set up. The normal distribution is chosen to simulate the probability density functions of the transit times. On the basis of the hypothesis mathematical expressions for the PLM- and CL-curves are worked out. By fitting the calculated PLM-curve to the experimental one it is possible to estimate mean values and standard deviations of T_G1, T_S>, and T_G2. As a test of the hypothesis the CL-curve is calculated by means of the estimated parameter values and compared to the experimental one. The calculated PLM- and CL-curves are found to be in good agreement with the experimental data as far as both follicle types are concerned. It is concluded that the method is a useful procedure. The choice of a normal distribution does not imply a significant limitation of the method in these investigations. Moreover it is concluded that the hypothesis is plausible. This means, e.g., that the proliferative fraction is unity in the two follicle types and that there is no cell loss from the cell systems.     

人肝癌细胞株H9101的建立及其细胞生物学特性研究

从厦门市肝癌高发的同安地区肝癌病人肝癌组织建立了人原发性肝细胞癌细胞系H9101.它生长迅速,群体倍增时间平均为38小时,对小牛血清的依赖性较低(1%).体外培养的H9101细胞贴壁生长,扫描和透射电镜下见细胞有丰富细长微绒毛.H9101细胞具有较高的软琼脂克隆形成率(1/200细胞)和裸鼠异种移植成瘤能力(100%).瘤细胞在ConA处理后呈较强的凝集性,在ConA 50ug/ml和100μg/ml时细胞凝集率分别为61%和92%.瘤细胞分泌微量的AFP,用1%DMSO和1.5×10-5mol/L地塞米松处理10天后上升到(140ng/ml/1×105细胞/24小时),且HBsAg转呈阳性.H9101细胞γ-谷氨酰转肽酶活性达5.42U±0.11U/mg,酪氨酸α酮戊二酸转肽酶活性达14.24U±1.50U/mg,与正常人肝细胞者相比差异显著(分别为P＜0.001和P＜0.01).H9101细胞染色体数为非整倍体,众数分布在52条-82条,具人类细胞染色体特征,其DNA经PCR扩增显示HBV DNA特异性条带.H9101细胞具端粒酶活性,是细胞连续传代和建系的保证;它恶性程度高、整合HBVDNA和具人肝细胞癌生物学基本特征,可为肝癌分子生物学研究提供有价值的实验材料.     

THE EFFECT OF HUMAN ENDOTHELIAL CELL-DERIVED PROTEOGLYCANS ON HUMAN SMOOTH MUSCLE CELL GROWTH

Extracellular proteoglycans (PGs) purified from cultured human arterial endothelial cells were tested for their effects on the proliferation of human vascular smooth muscle cells (VSMC). Fractions containing perlecan, the basement membrane heparan sulphate (HS) PG, the large chondrotin sulphate (CS) proteoglycan from connective tissue and other immunoreactive CS did not inhibit the proliferation of human VSMC. Native endothelial extracellular matrix, which was shown to contain the same PGs, demonstrated a pronounced stimulatory effect on the proliferation of human VSMCs. This stimulatory effect was not removed by pre-incubation of the matrix with 1M NaCl, heparin, platelet extract or plasmin. These experiments demonstrate that PGs produced by human arterial endothelial cells do not inhibit the proliferation of VSMC. These data do not support the hypothesis that human endothelial cells, in vivo control the activation or proliferation of VSMCs directly by the secretion of a non-proliferative molecule. Instead they support the hypothesis that the endothelial cells counteract intimal hyperplasia of VSMC indirectly by providing a barrier from activating factors in the plasma.     

羟基脲对人红白血病细胞株中珠蛋白基因表达和细胞分化的影响

人红白血病细胞株(HEL细胞)中珠蛋白基因表达具有自己的特点。即只表达胚胎型的γ-珠蛋白基因而不表达成人型的β-珠蛋白基因。羟基脲(Hydroxyurea)是一种抑制DNA合成的小分子有机化合物。它在临床上被用来治疗地中海贫血和镰刀型贫血。我们的实验结果表明:随羟基脲浓度增加HEL细胞的增殖速度减慢。用常规RT-PCR方法和定量PCR分析证明;用羟基脲诱导HEL细胞后,β-珠蛋白基因表达增加,α-珠蛋白基因表达减少,而γ-珠蛋白基因表达变化不明显。对参与珠蛋白表达的转录因子GATA-1和NF-E2的定量PCR分析发现:这两种转录因子的表达均增加3倍以上。因此,羟基脲可能通过某些信号传递途径促进β-珠蛋白基因表达,从而使HEL细胞趋向终末分化。     

KINETICS OF HUMAN LYMPHOCYTE RESPONSES IN VITRO: THE PHASE OF EXPONENTIAL GROWTH

The kinetics of thymidine uptake in human peripheral lymphocytes stimulated by allogenic cells, antigen E (ragweed allergen) and a variety of mitogens can generally be divided into four consecutive phases. First, a lag period with no increase in thymidine uptake, then a short period of rapid change in uptake, followed by a log-linear growth period and finally a decay phase. In this report we examine in detail the characteristics of the third, log-linear growth phase. Since, as discussed in the preceding paper, thymidine uptake is proportional to the number of cells acumulating thymidine, we can calculate from the log-linear growth period an apparent doubling time. We show that for five different stimulating agents the cells reach a log-linear growth phase of varying length and that the doubling times show little variation. This invariance indicates that, despite possible variation in cell death and recruitment rates, the rate of proliferation is in all cases dominated by the generation time of human lymphocytes.     

人体肝癌细胞系(SMMC-7721)糖皮质激素受体的半衰期

用抑制蛋白质合成的方法,测定了人体肝癌细胞系(SMMC-7721)糖皮质激素受体(GCR)的半衰期。结果显示,在蛋白质合成被抑制79.30±3.69%的情况下,GCR的半衰期为3.5±0.3h,降解速度常数为0.200±0.017h~(-1)。如果正常细胞GCR也具有如此短的半衰期,则提示GCR对内外环境的变化可做出迅速的反应,可能在靶细胞对激素反应的调节中起着活跃的作用。     

CLONE SIZE ANALYSIS: A PARAMETER IN THE STUDY OF CELL POPULATION KINETICS

The method of clone size analysis is described. Such an analysis provides a measure of the distribution of generation times in a cell population. Treatment by ionizing irradiation leads to perturbation of the generation times of a cell population and such perturbations are shown by changes in the clone size distribution. Distributions were compared after neutron irradiation and β-irradiation with those after X-rays; and also between X-irradiation under aerobic and anoxic conditions. At the same level of cell survival no differences were found between the patterns of cell survival with respect to generation times. Other uses of clone size analysis are discussed.     

抗人肺腺癌细胞株SPC-A-1的单克隆抗体的制备和鉴定

人肺腺癌细胞株SPC-A-1,经抗人正常肺抗血清复被后免疫小鼠,脾细胞和同系NS-1或SP 2/0融合,获得17株单克隆抗体。本文报道其建株过程及其中7株单克隆抗体的一些特征。LAC-122、LAC-163与LAC-210对人体肺腺癌和肺鳞癌的阳性反应率分别为75%、78.3%和75%,除LAC-122对一例胃癌细胞株有微弱交叉反应外,三株单抗对肺小细胞癌, 其它肿瘤组织及正常、胚胎组织均呈阴性。对利用抗体复被瘤细胞及肺癌相关抗原问题作了较详细的讨论.     

Bcl—2基因加强表达对SK细胞编程死亡的效应

TNF和OA诱发人神经母细胞瘤SK细胞编程死亡(PCD)。将编码Bcl-2完整蛋白质的cDNA植入pXJ 41neo载体中,由HCMV病毒启动子控制其表达。形成的正向(pBcl-2-S)及反向(pBcl-2-AS)表达质粒经转染导入SK细胞中获得稳定转染子。Western印迹表明正向转染子表达较大量的26kd Bcl-2蛋白,而反向转染子则不表达。增强表达的Bcl-2基因产物能抑制由TNF引发的PCD,但不影响由OA引发的PCD,从而证明Bcl-2基因产物抗细胞死亡效应的特异性。     

TSA杀伤培养的人大肠癌细胞DLD1及机理的研究

我们用曲古抑菌素A(TSA)作用于人大肠癌细胞DLD1,研究小眼相关转录因子(MITF)表达及其酰基化与癌细胞凋亡关系。Northern杂交实验结果表明:TSA作用组MITF表达明显增加,而对照组无明显MITF表达。电镜观察可见TSA引起DLD1超微结构的损伤及凋亡改变。提示TSA可能成为有效的杀伤癌细胞的药物。细胞因子作用组也可见MITF表达增加和细胞超微结构的改变。说明是否TSA引起MITF表达等可能与细胞因子作用通路相关。