Use of serum-free hormonally defined media to evaluate the effects of growth factors and inhibitors on proliferation of estrogen-responsive mammary and pituitary tumor cells in culture

Summary Serum-free defined media have been developed for assay of the mitogenic effects of growth factors on human MCF-7, human T-47D, and mouse COMMA-D mammary cells as well as for identification of mitogens and inhibitors of GH₄C₁ rat pituitary tumor cell growth. These lines were shown to grow in vivo in response to a variety of hormones including estrogens and thyroid hormones. With mammary cells, complete hormonally and nutritionally defined media were established that supported continuous passage at 50 to 90% of the serum stimulated rate. The strategy used to measure mitogens for mammary cells was to identify nutritional conditions where the growth rate was reduced greatly without impairing the response to picomolar to nanomolar concentration of growth factors. The effects of polypeptide growth factors and tissue extracts were estimated by their addition to basal medium and measuring cell number increases or labeled thymidine incorporation into DNA. In a variation of this methodology, the MTW9/PL2 rat mammary cells were used to identify secreted autocrine growth factors; nutritionally defined conditions were sought for growth of these rat cells in the complete absence of exogenous growth factors. The factors secreted into the medium were detected by bioassays with COMMA-D or MCF-7 mammary cell lines. The effects of growth factors-inhibitors on pituitary cells were measured by a related method; the GH₄C₁ cells were grown at less than optimal rates in a defined medium designated TRM-1. Addition of mitogens to TRM-1 stimulated pituitary cell growth whereas addition of inhibitors caused reduced levels of growth. The methods described in this report offer new means of assaying growth factors-inhibitors for a range of mammary and pituitary tumor cells.     

肝细胞培养过程中酸碱度和溶解氧的关联控制

目的在生物型人工肝支持系统(BAL)中,设计一种能够精确控制溶解氧(D0)与酸碱度(pH)的控制方案.方法根据肝细胞培养过程中所需要的物料衡算,采用比例积分(PI)算法结合开关量控制、预测控制等方案,通过工控机构建关联控制系统,使得D0与pH的值相互关联.结果DO控制范围0%～200%,精度达到±5%;pH控制范围6～8,精度达到±0.05.结论经实验证实,本控制方案工作稳定,无静态误差,解决了培养过程中DO与pH相互影响的问题,可用于BAL中对肝细胞培养环境的控制.     

Detection of Brucella melitensis by the BacT/Alert automated system and Brucella broth culture

This study was conducted to evaluate the ability of the BacT/Alert automated blood culture system to detect Brucella spp. in comparison with traditional Brucella broth culture. Overall, 100 (50 bone marrow and 50 blood samples) paired cultures were obtained, and 59 were positive by at least one method. The Brucella broth culture method detected all 59 positive cultures (100%), and the BacT/Alert system detected 30 (50.8%) ( P  < 0.05). The mean detection times for B. melitensis were 4.5 days in the BacT/Alert system and 5 days in Brucella broth culture ( P  > 0.05). There is no significant difference between the two methods with respect to growth time of the microorganism, but Brucella broth culture is more sensitive than the BacT/Alert system.     

Wg3h细胞系转染PSV_2-neo质粒前后生长特性及表面超微结构改变的研究

本文对转染PSV_2-neo质粒后的Wg3h细胞系(Wg3h-neo)在长期传代中的生长特性和表面超微结构与母系Wg3h细胞进行了比较研究。结果发现:转染后Wg3h细胞的DNA合成及生长速度明显高于母系Wg3h细胞,生长饱和密度增大。在软琼脂培养中不形成集落,接种在裸鼠不长肿瘤。在扫描电镜下,细胞表面的微绒毛较母系Wg3h细胞丰富。Southern印迹杂交实验证明PSV_2-neo质粒已整合到宿主细胞的基因组中。转染后Wg3h细胞的生长特性和表面超微结构均发生了某些转化特征。     

人肺癌细胞A549产生的免疫抑制因子对T细胞的作用及其生物学特性

本文系用人肺腺癌细胞株A549为研究对象,观察了TDSF对T淋巴细胞的作用,并对其某些生物学特性作了初步研究。实验表明:TDSF对T淋巴细胞的增殖、IL-2的产生及其反应性均有明显的抑制作用。TDSF作用相当强烈,这点是应用IL-2对肿瘤进行免疫治疗时应该注意的问题。丝裂霉素等抗代谢药物可抑制TDSF的分泌,提示TDSF为瘤细胞合成和分泌的基因产物。TDSF对酸、碱、热、胰蛋白酶敏感,分子量>150KD,表明其化学本质为蛋白质。     

Membrane carbohydrates of lymphoid cells: the receptor for interleukin 2

The contribution of sialic acids and of N-linked sugars to the biological activity of the receptor for IL 2 has been evaluated by treating activated cells with Neuraminidase or by growing them in the presence of inhibitors of N-linked glycosylation or processing. After treatment with Neuraminidase, Con A-activated spleen cells had not lost their ability to bind IL 2. The IL 2-absorbing capability was, however, strongly reduced after trypsinisation. 6 hours after Trypsin treatment, this property was again expressed. Proliferation of the IL 2-dependent CTLL cells was normal in the presence of Swainsonine but strongly impaired in the presence of Tunicamycin. Glycosylation of the IL 2 receptor may thus be required, but integrity of the sugars is not critical.     

Levetiracetam Inhibits both ryanodine and IP3 receptor activated calcium induced calcium release in hippocampal neurons in culture

Epilepsy affects approximately 1% of the population worldwide, and there is a pressing need to develop new anti-epileptic drugs (AEDs) and understand their mechanisms of action. Levetiracetam (LEV) is a novel AED and despite its increasingly widespread clinical use, its mechanism of action is as yet undetermined. Intracellular calcium ([Ca²⁺]_i) regulation by both inositol 1,4,5-triphosphate receptors (IP3R) and ryanodine receptors (RyR) has been implicated in epileptogenesis and the maintenance of epilepsy. To this end, we investigated the effect of LEV on RyR and IP3R activated calcium-induced calcium release (CICR) in hippocampal neuronal cultures. RyR-mediated CICR was stimulated using the well-characterized RyR activator, caffeine. Caffeine (10 mM) caused a significant increase in [Ca²⁺]_i in hippocampal neurons. Treatment with LEV (33 μM) prior to stimulation of RyR-mediated CICR by caffeine led to a 61% decrease in the caffeine induced peak height of [Ca²⁺]_i when compared to the control. Bradykinin stimulates IP3R-activated CICR—to test the effect of LEV on IP3R-mediated CICR, bradykinin (1 μM) was used to stimulate cells pre-treated with LEV (100 μM). The data showed that LEV caused a 74% decrease in IP3R-mediated CICR compared to the control. In previous studies we have shown that altered Ca²⁺ homeostatic mechanisms play a role in seizure activity and the development of spontaneous recurrent epileptiform discharges (SREDs). Elevations in [Ca²⁺]_i mediated by CICR systems have been associated with neurotoxicity, changes in neuronal plasticity, and the development of AE. Thus, the ability of LEV to modulate the two major CICR systems demonstrates an important molecular effect of this agent on a major second messenger system in neurons.     

脐带血清对骨髓粒—巨噬细胞集落形成的影响

在骨髓细胞半固体琼脂培养体系中，研究脐带血清（ＣＢｓ）对骨髓粒—巨噬细胞集落形成单位（ＣＦＵ－ＧＭ）的影响，单纯应用ＣＢｓ培养可使骨髓ＣＦＵ－ＧＭ（ｘ±ｓ为２６．０３７５±１０．２３２７）显著的高于正常成人外周血清（ＰＢｓ）组（ｘ±ｓ为１１．５９１７±３．６０４７；Ｐ＜０．００１）。ＣＢｓ加粒－巨噬细胞刺激因子，ＧＭ－ＣＳＦ组ＣＦＵ－ＧＭ（ｘ±ｓ为１１９．０７９２±３５．０９９１）是单纯ＣＢｓ组的４．５７倍；而ＰＢｓ加ＧＭＣ－ＳＦ组（ｘ±ｓ为９７．１２５±２６．７５５９）是单纯ＰＢｓ组的８．３８倍。有白细胞介素－６（ＩＬ－６）存在的ＣＢｓ组，ＣＦＵ－ＧＭ（ｘ±ｓ为３１．９３３３±１０．９１４４）明显的高于单纯ＣＢｓ组（Ｐ＜０．０２）。提示ＣＢｓ中含有较高水平的内源性ＧＭ－ＣＳＦ。