THE EFFECT OF HYDROCORTISONE ON HeLa CELL GROWTH

HeLa Chessen cells have a doubling time of 18 hr when grown in MEM containing 10% calf serum and antibiotics. When hydrocortisone (1.7 μg/ml) is added to exponentially distributed cells in log growth in this medium, a new pattern of growth begins to emerge after 10–12 hr. This pattern is characterized by a transitional state lasting for about 6 hr, and then a new doubling time of about 35 hr is maintained thereafter. Hydrocortisone removes about 5% of the cells from the proliferative pool and extends the generation time of proliferating cells to about 30 hr. The extension of the generation cycle appears to occur almost entirely in late G₁. Cells grown as clones (average 6 cells/clone) prior to the addition of hydrocortisone, undergo these changes with doses as low as 0.00017 μg/ml of medium. When the average clone size is 1.5 cells per clone, the drug concentration must be 0.017 μg/ml or higher to initiate this response. The HeLa S₃ strain continues to grow with an 18-hr doubling time in the presence of hydrocortisone after a temporary delay in growth occurring between the 12th and 16th hour.     

THE EFFECT OF HEMOLYTIC SUBSTANCES ON WHITE CELL RESPIRATION

Substances such as saponin, the bile salts, etc., which produce lysis of red cells also produce cytolysis of white cells from rabbit peritoneal exudates, the arbitrary criterion of their cytolytic effect being their ability to depress the O₂ consumption of the leucocytes. The amount of cytolysis increases regularly as the amount of the added lysin is increased, and sufficiently large quantities of saponin, sodium taurocholate, sodium glycocholate, or sodium oleate are capable of virtually abolishing the O₂ consumption altogether. At the same time, it can be shown that a lysin such as saponin is used up in combining with the white cells in much the same way as it is used up in combining with red cells, and the reduction in oxygen consumption appears to be roughly proportional to the amount so combined. The action of these lytic substances on white cells, in fact, is very similar to their action on red cells, due allowance being made for the fact that the cytolysis of the white cell is probably not an all-or-none process like hemolysis. White cell respiration is also depressed in hypotonic solutions, the respiration being virtually linear with the tonicity.     

AN ANALYSIS OF THE EFFECT OF "CONDITIONED MEDIUM" UPON THE CELL CULTURE AT LOW DENSITY

A favorable effect of “conditioned medium” upon outgrowth of the cell culture with low density in vitro was analysed with the cells of chicken embryos. For preparing “conditioned medium”, cultures with a large number of cells were made with muscle, kidney, lung, liver and skin, while the biological activity of the medium was assayed by using the culture of a small number of the lung secondary cells. A use of “conditioned medium” was found to be necessary for encouraging the outgrowth of the cultured cells below a critical inoculum size. Of the various types of the media tested, the medium conditioned with muscle was most effective. “Conditioned medium” contained at least two different active factors, the first to enhance the plating efficiency of the inoculated cells to the surface of the culture dish, and the second to promote further outgrowth of the plated cells. “Conditioned medium” taken out of the mass culture at its exponentially growing phase had only the second factor, while that taken out of that at its stationary phase contained both factors. An activity of the first factor was not detected, when the mass culture was kept in such condition that the collagen synthesis was inhibited. The factor for enhancing the plating efficiency was eliminated from “conditioned medium” by preincubating the cells, before assaying the effect of the medium.     

THE ALLEGED EFFECT OF ELECTRICAL STIMULATION ON THE METABOLISM OF RED CELL SUSPENSIONS

The apparent increase in the rate of O₂ consumption described when an alternating current, or induction coil current, is passed through a red cell suspension (in a buffered NaCl solution) is not a metabolic effect in any sense of the word. The passage of the current results in a permanent volume decrease in the system, and it is this which has been erroneously interpreted as an "increase in the rate of O₂ consumption." Its magnitude is about 1 part in 1000. The utilization of O₂ is not involved at all, and the same effect is obtained, on a somewhat smaller scale, when the current is passed through a solution of NaCl or of the other halogen salts. The effects occur only with shiny platinum electrodes, and disappear entirely when the electrodes are platinized. Passage of the current through serum, on the other hand, results in a permanent increase in the volume of the system, this effect also disappearing on platinization of the electrodes. The effects are apparently related to obscure electrode phenomena.     

THE EFFECTS OF LOW CONCENTRATIONS OF ACTINOMYCIN D ON THE PROGRESS OF CELLS THROUGH THE CELL CYCLE

The progress of L-cells through the cell cycle in asynchronous and in synchronous culture has been studied at a concentration of actinomycin D which mediates an apparently‘nucleolar-specific’inhibition of RNA synthesis. Under such conditions, cells may be blocked or seriously delayed in the G₁ and G₂ phases of the cycle whilst the processes of DNA synthesis and mitosis once initiated, can still occur at control rates. The results show that the sensitivity of a cell to these blocks depends critically upon the position of that cell within the cycle at the time of drug addition. The possible mechanisms of the drug's action are discussed.     

EFFECT OF UNDERNUTRITION ON CELL FORMATION IN THE RAT BRAIN

Abstract— Rats were undernourished by approximately halving the normal food given from the 6th day of gestation throughout lactation. Growth of the foetuses was nearly normal, in marked contrast to the severe retardation caused by undernutrition during the suckling period. In comparison with controls the size and the DNA content of the brain were permanently reduced by undernutrition during the suckling period: this effect was relatively small, approx. 15 per cent decrease at 21 and 35 days. The rate of ¹⁴C incorporation into brain DNA at 30 min after administration of [2-¹⁴C] thymidine was taken as an index of mitotic activity; compared with controls there was severe reduction in mitotic activity (maximal decrease by about 80 per cent at 6 days in the cerebrum and by 70 per cent at 10 days in the cerebellum). The rate of acquisition of cells was calculated from the slopes of the logistic curves fitted to the estimated DNA contents. In normal animals the maximal slope was attained at 2·7 days and at 12·8 days after birth in cerebrum and cerebellum respectively; the daily acquisition of cells at these times was 4·8 × 10⁶ and 18 × 10⁶ cells respectively. The fractional increase in cell number at the maximum was 5·4 percent per day in the cerebrum and 15·2 per cent per day in the cerebellum. The rate of acquisition of cells relative to the rate of mitotic activity was higher in the brains of undernourished animals than in controls. One of the compensatory mechanisms for the severe depression of mitotic activity in the brain of undernourished animals Seems to involve a reduction in the normal rate of cell loss.     

姜黄素诱导人脑胶质瘤细胞U251分化的实验研究

目的探讨姜黄素对体外培养的人胶质瘤细胞U251的诱导分化作用及其机制。方法将人脑胶质瘤U251细胞分为对照组和药物组,对照组以含10%小牛血清的DMEM培养基常规培养,药物组用16μmol/L姜黄素（本实验先前的研究结果显示,姜黄素作用于U251细胞48h的半数抑制浓度为16μmol/L）处理。倒置显微镜观察细胞形态学变化;流式细胞仪检测细胞周期变化;免疫细胞化学法检测波形蛋白（vimentin）表达变化;免疫印迹（Westernblot）法检测细胞外调节蛋白激酶（ERK）、磷酸化细胞外调节蛋白激酶（pERK）和胶质纤维酸性蛋白（GFAP）表达变化。结果姜黄素作用96h后,与对照组相比,U251细胞突起明显增多变细长。姜黄素培养48h,S期细胞由28.53%明显降低至20.35%（P=0.015）;vimentin强阳性细胞百分率由90%明显降低至50%（P=0.009）;GFAP蛋白与内参β-actin表达量的比值由0.22明显升高至0.50（P=0.01）。Westernblot法显示,姜黄素分别作用于U251细胞48和96h,ERK蛋白水平无明显改变,pERK/ERK分别为0.35和0.22,较对照组明显减少（P=0.03和0.007）。结论姜黄素对体外培养的胶质瘤U251细胞有显著的诱导分化作用,其机制可能与抑制异常激活的ERK信号通路有关。     

大肠杆菌对体外培养的THP-1细胞凋亡的影响

目的探讨大肠杆菌(E.coli)感染与THP-1细胞凋亡的关系.方法以THP-1细胞作为E.coli感染的体外细胞模型,当细胞与细菌浓度比分别为1:5,1:10,1:20,1:50及1:100时,用荧光染色技术、Annexin V/PI双染流式细胞仪检测分析等方法检测细胞凋亡率.结果细胞与细菌浓度比较低时(1:5)即可引起部分THP-1细胞发生凋亡,THP-1细胞凋亡百分率随E.coli感染剂量的增加而增加.Hoechst33258荧光染色及流式细胞仪二种方法所得结果一致.结论 E.coli以剂量依赖方式诱导THP-1细胞凋亡.     

EFFECT OF ACTINOMYCIN D ON THE DIFFERENTIATION OF HATCHING GLAND CELL AND CILIA CELL IN THE FROG EMBRYO

The forehead epidermis of the stage 18–20 R. japonica embryo includes the hatching gland cell (HGC) which contains cell-specific secretory granules. The cilia cell (CC) and common epidermal cell (CEC) constitute the epidermis of the entire body surface, in addition to the forehead region.
Culture of superficial epidermal explants from various embryonic portions at various developmental stages revealed that HGCs are derived from cells localized on the neural crest in the stage 13a (early neural plate) embryo. When explants from the presumptive HGC area were treated with 1 ug/ml actinomycin D (AMD), the formation of secretory granules in HGCs was inhibited either by continuous treatment from stage 13 or by an 8-hr treatment at stage 13b. Similarly, the ciliogenesis in CCs was inhibited. The differentiation of CECs was entirely unaffected by any of the AMD treatment. After release from AMD, mucous vesicles, characteristic of the CEC, were formed in cells whose differentiation into HGC and CC had been suppressed by the antibiotic. Thread complexes and clumps of coiled strings were found in the nuclei of AMD-affected cells.
It is concluded that the DNA-dependent RNA syntheses which direct secretory granule formation in the HGC and ciliogenesis in the CC occur during a limited period at stage 13b, viz. , 20 hr before their cytodifferentiation becomes appreciable.     

EFFECT OF INHIBITORS ON KINETICS OF INDOLEACETIC ACID OXIDATION

Sacher , Joseph A. (Los Angeles State Coll., Los Angeles, Calif.) Effect of inhibitors on kinetics of indoleacetic acid oxidation. Amer. Jour. Bot. 50(2): 116–122. Illus. 1963.—Kinetic studies of the relative effectiveness of a series of phenolic antioxidants in causing a lag in the oxidation of indoleacetic acid by an enzyme from pods of ‘Kentucky Wonder’ pole beans (Phaseolus vulgaris) support the conclusion that they react with free-radical intermediates within the enzyme-substrate complex, and thus greatly inhibit the rate of enzyme turnover. Investigations with a partially purified enzyme show a dependency upon added manganese for any significant oxidation. The enzyme is inhibited by cyanide, but not by carbon monoxide. Cyanide causes complete inhibition (a lag), followed by partial inhibition. The inactivation of phenolic inhibitor requires oxygen, but may occur anaerobically if traces of peroxide are present, attended by irreversible inactivation of most of the enzyme. In the absence of inhibitor, enzymic inactivation occurs under anaerobic conditions in the presence of indoleacetic acid and 5 × 10^–6 MH₂O₂. These results are construed to indicate that a form of the enzyme active in the presence of H₂O₂ and indoleacetic acid is involved in both inactivation of enzyme and the anaerobic destruction of phenolic inhibitors. The enzymic oxidation of indoleacetic acid and inhibitor inactivation are discussed.     

THE EFFECT OF X-RAY INDUCED SYNCHRONY ON TWO-DOSE CELL SURVIVAL EXPERIMENTS

Calculations were made based on experimental data of the variation in radiosensitivity and in cell cycle delay with age in Chinese hamster cells (Sinclair, 1967, 1968). The computed syntheses included: (a) single-dose survival curves; (b) splitdose recovery curves, the pattern of survival; and (c) two-dose survival curves, the shape of the second survival curve. It is clear that, after an initial dose, changes occur in the subsequent dose-response curve which depend upon size of dose and time after irradiation. Following rapid and apparently complete repair of sublethal injury, these changes can be attributed to progression of a partially synchronized population through phases of varying radiosensitivity. These changes may be small in some systems, but important in others. The concept of equal dose-increments after each of several dose fractions is an over-simplification and must be used with care.     

THE EFFECT OF PARTIAL BODY IRRADIATION ON HAEMOPOIETIC STEM CELL MIGRATION

Data obtained after various types of partial body irradiation support the concept of a small rapidly exchangeable pool of CFUs, which seems to be exhausted rapidly after irradiation. The depletion of this pool is the most plausible explanation for the decrease in stem cell migration observed 3 hr after exposure in C₃H mice. After partial body irradiation the size of the rapidly mobilizable pool is reduced in proportion to the areas of bone marrow irradiated. When only one marrow area is shielded, the recovery of this pool does not occur during the first 24 hr after exposure.     

亚硫酸氢钠对大蒜幼苗生长及细胞分裂的作用

研究SO2体内衍生物-亚硫酸钠与亚硫酸氢钠（分子比为3：1）混合液对大蒜幼苗生长和细胞分裂的效应。结果表明：低浓度（0.2mmol/L）的混合液促进幼苗生长,短时间（24h）作用时根尖细胞有丝分裂指数（MI）提高;高浓度（1.5与2.0mmol/L）处理抑制大蒜幼苗生长,细胞周期涎滞,大蒜根尖MI下降,并诱发细胞分裂异常,且呈明显的时间效应与剂量效应关系。     

反义核酸对人大肠癌CCL229细胞侵袭力的抑制作用

为观察反义寡核苷酸(asODN)对培养的高侵袭性人大肠癌CCL229细胞侵袭性的抑制作用,针对尿激酶型纤溶酶原激活物受体(re-ceptorofuPAR)mRNA的翻译起始区,合成了一段反义寡核苷酸(asODN),用脂质体将其导入培养的CCL229细胞中,通过逆转录-聚合酶链反应(RT-PCR)、流式细胞术、羊膜侵袭试验测定uPARmRNA水平、细胞表面uPAR抗原表达及细胞侵袭力的变化,并在扫描电镜下观察细胞的形态改变.结果为(1)RT-PCR检测asODNs组uPAR/β-actin比值为0.44±0.02,与对照组(0.81±0.01)相比,显著降低(P＜0.05);(2)流式细胞术检测asODNs组癌细胞表面与uPA结合的受体和总受体的平均荧光指数分别为0.20±0.07、0.59±0.09,与对照组(分别为0.72±0.12、2.21±0.36)比较,明显下降(P＜0.01,P＜0.05);(3)体外侵袭试验结果,asODN组在48h和72h后穿过羊膜的细胞数分别为12±2、20±3,与对照组(分别为25±4、44±5)相比,明显减少;(4)扫描电镜观察表明细胞经asODNs作用后,表面伪足和针状突起明显减少.以上各指标在随机序列寡核苷酸(rODN)组和对照组相比,无明显变化.CCL229细胞表面uPAR的表达与癌细胞的侵袭力密切相关,asODNs能有效抑制CCL229细胞uPAR基因的表达,从而抑制相关的蛋白质合成,降低其侵袭性,实验结果在癌症的基因治疗上将具有一定意义.