重组人钙调素对白芷细胞增殖及结核杆菌生长的影响

目的 :探讨重组人钙调素 ( rh Ca M)对白芷悬浮细胞及结核杆菌增殖作用的影响。　方法 :用基因重组技术获得人钙调素基因 ( h Ca M c DNA)重组表达质粒 h Ca M / p BV2 2 0 ,将其转化大肠杆菌 DH5α。用 Phenyl-Sepharose CL -4 B疏水亲和层析法纯化重组菌超声上清表达产物。将纯化 rh Ca M加入悬浮培养的白芷细胞及结核杆菌培养基中 ,观察 rh Ca M对其增殖作用的影响。　结果 :含 h Ca M / p BV2 2 0的重组菌经温度诱导可高效表达Ca M蛋白 ,经 15 % SDS-PAGE分析 ,可观察到一相对分子质量为 170 0 0的表达条带 ,Western blot结果证实 ,此表达条带可与标准鼠抗 Ca M Mc Ab起特异反应。每 1L菌液经 Phenyl-Sepharose CL -4 B疏水亲和层析法纯化可获Ca M纯品 3～ 4mg。rh Ca M对白芷细胞增殖作用影响的研究结果表明 ,rh Ca M在低细胞密度培养条件下对白芷细胞有促进增殖作用 ,效果高于标准植物 Ca M。本研究同时还发现一定浓度的纯化 rh Ca M( 1、10μg/ ml)可促进牛型结核杆菌的生长。　结论 :rh Ca M对植物细胞和细菌均有细胞外促增殖作用     

P^27的研究概况

目的：综述Ｐ＾２７的研究进展。方法：光盘检索有关献,归纳综合整理。结果：Ｐ＾２７是细胞周期的相关蛋白,Ｐ＾２７白过度表达可抑制细胞周期的进程,阻断细胞生长于Ｇ１期。结论：Ｐ＾２７是细胞周期中重要的负性调控控因子,是一种新的侯选的肿瘤抑制基因。     

人原发性肝细胞癌核基质蛋白的研究

目的比较正常肝与原发性肝细胞癌（ＨＣＣ）核基质蛋白的异同,观察是否有ＨＣＣ特异性核基质蛋白的存在。方法用双向电泳方法比较了３例正常肝和８例ＨＣＣ核基质蛋白成分。结果正常肝和ＨＣＣ的核基质蛋白组成极为相似,但在ＨＣＣ中发现至少有４个ＨＣＣ的特异性核基质蛋白。其中以分子量为６２０００、等电点为５．３的蛋白最具代表性,它存在于所研究的８例ＨＣＣ中。其余３个ＨＣＣ特异性蛋白亦存在于大多数ＨＣＣ中。３例正常肝组织中未见有这４个ＨＣＣ特异性核基质蛋白。结论ＨＣＣ确实有ＨＣＣ特异性核基质蛋白的存在,它的发现可能会为ＨＣＣ的发生、发展、发病机理的研究提供一个新途径。     

1H-NMR and NOE studies of 2-cephems and 3-cephems

The¹H-NMR signals of 2-cephems and 3-cephems have been assigned and the Nuclear Overhauser Effect (NOE) study of these compounds was undertaken.     

Regulatory volume response of erythrocytes exposed to a gradual and slow decrease in medium osmolality

 A sudden decrease in external medium osmolality (90 mosmol/kg) causes an immediate swelling of trout erythrocytes, followed by a regulatory volume decrease (RVD) due to activation of both a KCl cotransporter and a taurine transport pathway. Here, we determined how trout red cells respond when they are exposed to a gradual and slow decrease in medium osmolality (80 mosmol/kg at a rate of 0.7 mosmol/kg per min). Erythrocytes were unable to regulate their volume efficiently when swollen gradually and it increased continuously throughout the experimental period (120 min). As long as volume was increased slowly by 15–25%, regulatory pathways remained essentially inactivated, erythrocytes losing no significant amount of intracellular osmotically active solutes. Above this swelling threshold, a response was triggered but the quantity of solutes lost via the regulatory pathways was still not sufficient to counterbalance the continuous entry of water due to the slow and gradual decrease in medium tonicity. Received: 18 January 1999 / Received after revision: 10 February 1999 / Accepted: 11 February 1999     

Flow-mediated cell stress induction in adherent leukocytes is accompanied by modulation of morphology and phagocytic function

Leukocytes adherent to the surfaces of both vascular biomaterials and normal blood vessels experience blood flow induced shear stress. The goal of the reported studies was to investigate the effect of fluid flow on the morphology, phagocytic function and stress response induction in adherent immune cells. Shear approximating arterial, venous and intermediate levels were applied onto glass-adherent IC21 macrophages in a temperature-controlled parallel plate flow system. The results indicate that fluid flow induces a shear-dependent physiological stress response in adherent macrophages and that significant morphological changes accompany macrophage responses to shear stress. In addition, arterial flow conditions induce not only significant cell polarisation, but also enhanced phagocytic ingestion in glass-adherent IC21 macrophages. These findings suggest that blood flow induced shear stress may not only be consequent to adherent leukocyte activation, but may also be integral to the regulation of adherent leukocyte behaviour in vivo.     

Cellular dynamics of corneal wound re-epithelialization in the rat. I. Fate of ocular surface epithelial cells synthesizing DNA prior to wounding

The fate of ocular surface epithelial cells in response to injury of the cornea was examined. Corneal epithelial cells were labeled during DNA synthesis with [3H]thymidine 1 h prior to wounding. A 3-mm diameter epithelial defect was made in the center of the rat cornea, with the basement membrane remaining intact. Within 12 h of abrasion, labeled cells were detected in the regenerating surface. At 18 h, there was a 2.7- and 17-fold increase of labeled basal and suprabasal cells, respectively, in the epithelium adjacent to the wound, and at 24 and 30 h there was an excessive number of cell layers (up to 7) at the margin of the abrasion. Re-epithelialization progressed as a gradient of cell layers that became diminished towards the center of the wound. Completion of layers 1, 2, 3, and 4 were recorded at 24, 30, 36, and 72 h, respectively. No changes in the labeling index of the limbus or conjunctiva were noted. These results suggest that processes of centripetal and vertical migration, as well as events related to cell division, in the uninjured corneal surface are not impeded by wounding of the corneal epithelium. However, wound healing appears to require cells with a basal phenotype, presumably because of this cell type's migratory capability.     

Intracellular ATP depletion inhibits swelling-induced -[H]aspartate release from primary astrocyte cultures

Volume expansion-sensing outward rectifier (VSOR) anion channel, also referred to as volume-sensitive organic osmolyte-anion channel (VSOAC), appears to be responsible for cell swelling-induced amino acid release in a variety of cells. One prominent feature of the VSOR/VSOAC is that non-hydrolyzed intracellular ATP binding to the channel or an accessory protein is required for its activation. In this study, the effect of intracellular ATP depletion on the swelling-induced release of -[³H]aspartate from rat primary astrocyte cultures due to exposure to either high K⁺ or hypotonic media was studied. When the cells were pretreated for 10 min with a combination of the metabolic inhibitors 2-deoxyglucose and rotenone, 100 mM K⁺ media- or hypotonic media-induced -[³H]aspartate release was completely suppressed. Added separately, each inhibitor showed only partial or no inhibition of -[³H]aspartate release, which correlated with its relative effectiveness in decreasing intracellular ATP levels. These data are consistent with the view that during high [K⁺]_o or hypotonic media-induced swelling of primary astrocyte cultures an ATP-dependent swelling-activated VSOAC channel is responsible for -[³H]aspartate release and close to normal ATP is required for full channel activation.     

Hypoxia induces differential changes of dopamine metabolism in mature and immature mesencephalic and diencephalic cell cultures

Summary. Perinatal hypoxia is known as a high risk factor for the development of long-lasting abnormalities in dopaminergic system. The early developmental alterations of dopamine (DA) metabolism induced by hypoxia could contribute to these abnormalities. To understand the hypoxia-induced changes of intra- and extracellular dopamine levels and its main metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), in immature dopaminergic neurons, we compared these changes in rat mesencephalic and diencephalic cell cultures on day in vitro (DIV) 2 (immature cells), DIV 8 and DIV 13 (mature cells). Cell cultures were exposed to an oxygen-free gas mixture in a Billups chamber for 2–4 hours. Mature cell cultures responded to hypoxia with an increase of DA levels in the cells and in the medium during the first 45 min (by an average of 57 and 114% respectively). Thereafter, DA levels decreased, and returned to the baseline within the next 30 min. The cellular DA levels continued to decrease up to 15% of the baseline during 255 min hypoxia whereas the extracellular DA content stabilized at the prehypoxic levels. Immature cell cultures (DIV 2) in contrast to mature ones, were unable to maintain normal extracellular DA levels during hypoxia and showed a decrease of the cellular and extracellular levels to 50% of the prehypoxic levels. DOPAC and HVA changes mimick, however, at a lower level, the pattern of DA changes during the exposure to hypoxia. In principle, in the diencephalic cell culture similar effects of hypoxia exposure on the investigated parameters were found (studied during 0–120 min). The present study demonstrates that mature and immature dopaminergic cells differ in the regulation of the extra- and intracellular DA levels during hypoxia. In immature cells the low synthetic capacity of tyrosine hydroxylase and the deficient capacities of the transport and storage processes result in decreased extracellular DA levels. This could be an important factor for the long-term modulation of the expression of tyrosine hydroxylase and subsequent long-term behavioral and/or neurological abnormalities induced by perinatal hypoxia. Received June 8, 1998; accepted July 21, 1998