Localization of tissue plasminogen activator mRNA in adult rat brain

The distribution of tissue plasminogen activator (tPA) messenger RNA in rat brain was studied using in situ hybridization with ³⁵S UTP-labeled RNA probes derived from a fulllength tPA cDNA. Sense strand controls produced low, even backgrounds, with small elevations in the hippocampus. Full-length antisense probes produced strong signals over cerebral ventricular ependyma (including ependyma of the subcommissural organ), meninges, blood vessels, and Purkinje cell layer of the cerebellum, as well as strong signals over scattered cells throughout the brain. Some of these scattered labeled cells were large with lightly stained nuclei, while others were small with darkly stained nuclei. The large labeled cells, which were probably neurons, constituted 8% and 8% of cells in the brain stem and neocortex, respectively, and 100% of Purkinje cells. The small cells, which were present in all areas of the brain, constituted 3–11 % of cells in individual brain areas.     

高浓度氨基酸对肿瘤细胞体外增殖的影响

本研究首先采用Hanks溶液作为介质,观察了不同氨基酸浓度(1～5mmol/L)对艾氏腹水癌细胞、瓦克癌肉瘤256细胞体外增殖的影响,然后以含有平衡氨基酸组成的Eagle最低必需培养基作为介质进一步观察了不同氨基酸浓度的增加对艾氏腹水癌细胞及人肝癌细胞(PLC/PRF/5)、肺癌细胞(AGTJ-a)体外增殖的影响,同时采用小鼠骨髓细胞观察了高浓度氨基酸对正常组织细胞可能产生的毒性作用。结果表明,尽管一些氨基酸浓度的增加有显著作用,但是不同肿瘤细胞对不同氨基酸浓度增加的反应不尽一致,而且一些氨基酸的作用受培养介质的明显影响;高浓度色氨酸对小鼠骨髓细胞的体外增殖有显著抑制作用。     

Characterization of glial subpopulations in cultures of the ovine central nervous system

The cellular composition and in vitro development of glial cultures derived from the rat CNS has been well studied. However, less information is available on similar cultures from other species, particularly higher mammals. To study ovine glial development in vitro, cultures from 50-day fetal to adult animals were characterized with various immunocytochemical markers, which are frequently used to define neural cell subsets in rat cultures. As in rats, both A2B5+ and A2B5- astrocytes can be identified in ovine cultures. However, ovine A2B5+ and A2B5- could not be reliably differentiated by their morphology, which was more influenced by whether the cells were in serum-free or serum-containing media than by their A2B5-positive or -negative status. In addition, ovine A2B5+ astrocytes were present in cultures from early fetal brain before the development of identifiable oligodendrocytes, unlike rat type II astrocytes, which develop only after the appearance of oligodendrocytes. An A2B5+ cell, morphologically similar to the rat 02-A cell, can be found in cultures from fetal ovine cerebrum or cerebellum. A2B5+/glial fibrillary acidic protein (GFAP)- cells in cultures from 100- to 115-day ovine cerebellum appeared to differentiate into A2B5+ astrocytes in serum-containing media. However, in serum-free media, although the A2B5+ cells assumed a more "oligodendroglial-like" morphology, they did not express galactocerebroside or myelin basic protein, suggesting that these cells may not be bipotential as is the rat 02-A cell. Oligodendroglial differentiation was not induced by treatment with dibutyryl cyclic AMP or insulin-like growth factor I. Many cells in cultures from a variety of fetal ages did not label with any of the immunocytochemical markers used, suggesting the need for more cell-type-specific markers to identify neural cell subsets in higher mammals.     

Decreased antibody-dependent cellular cytotoxicity in minimal change nephrotic syndrome

Secondary IgG response to a tetanus toxoid booster and in vitro measurement of immunoglobulin synthesis, antibody-dependent cellular cytotoxicity (ADCC) and -interferon (IFN-) production were evaluated in 20 healthy controls and in 17 children with minimal change nephrotic syndrome (MCNS), during the acute nephrotic phase and 6 months after remission. Defective responses were observed in all but IFN- production during the acute nephrotic phase; these improved with disease remission. There was a significant correlation between decreases in vitro IgG production and ADCC reaction. These data indicate that defective antibody production is associated with decreased ADCC during the acute nephrotic phase of MCNS.     

基于网络药理和实验验证探究中药通过健脾作用治疗白细胞减少症的作用机制

摘要 目的 运用数据挖掘、网络药理学和实验验证探讨中药通过健脾作用治疗白细胞减少症的用药规律、药效物质基础及潜在的作用机制。方法 在中国知网、万方数据知识服务平台、维普网和PubMed数据库中检索健脾中药复方治疗白细胞减少症的临床文献,运用古今医案云平台在线分析软件和SPSS Modeler 18 统计分析软件寻求核心药物组;利用中药系统药理学数据库和分析平台（TCMSP）、GeneCards数据库、OMIM数据库检索、Cytoscape3.7.2软件、STRING数据库获取关键基因和主要有效成分,利用Metascape进行基因本体（GO）功能富集分析和京都基因与基因组百科全书（KEGG）通路富集分析;采用MTT法检测细胞增殖率,采用qPCR法检测关键基因信使核糖核酸（mRNA）水平表达。结果 共筛选得到90篇有效文献,共计91首中药复方,党参-当归-黄芪为核心药物组合。肿瘤坏死因子（TNF）、白细胞介素-6（IL-6）、白细胞介素-1β（IL-1β）、白细胞介素-10（IL-10）、JUN为核心靶点,槲皮素、木犀草素、山奈酚、芒柄花素、豆甾醇为主要有效成分,其主要作用机制可能通过癌症的途径、TNF信号通路、p53信号通路等来实现。健脾中药黄芪、党参、当归提取物和有效成分芒柄花素、槲皮素对人单核细胞白血病细胞（THP-1）具有促进增殖作用,芒柄花素对化疗诱导的THP-1细胞生长具有促进增殖的作用,芒柄花素能够抑制炎症细胞因子IL-6、IL-1β、TNF-α和刺激抗炎因子IL-10、原癌基因JUN的分泌。结论 初步揭示黄芪等多种中药通过多成分、多靶点发挥健脾作用,参与抗肿瘤、抗炎、免疫调节等相关信号通路,治疗白细胞减少症。     

Kinetic properties of the in vivo accumulation of 3H-(−)-N-n-propylnorapomorphine in mouse brain

Summary (1) The influence of various dopamine (DA) receptor agonists and antagonists on the kinetic properties of the specific binding of ³H(–)-N-n-propylnorapomorphine (NPA) in the mouse striatum in vivo was studied. The specific binding of ³H-NPA, defined as the difference between the radioactivity in the striatum and cerebellum, was completely antagonized by the selective D-2 receptor antagonist raclopride but not by the selective D-1 antagonist SCH 23390, showing that the binding occurs exclusively to the D2 receptors. (2) The selective D-2 receptor agonists pergolide and quinpirole inhibited the ³H-NPA binding biphasically at low doses, indicating that these DA receptor agonists have high affinities for a subfraction (10 to 30%) of the NPA binding sites. (3) Increasing the synaptic DA concentration by DA release [(+)-amphetamine] or uptake blockade (amfonelic acid and methylphenidate) inhibited the ³HNPA binding in a competitive manner (unchanged B _max, increased K _D). Depletion of the DA in the synapses by -butyrolactone or reserpine decreased the apparent K _D value. (4) The possibility of estimating changes in the synaptic DA concentration from changes in the apparent K _D is discussed. According to the results obtained, the normal concentration of DA in the synaptic cleft in mouse striatum in vivo is about 40 nmol/l and this concentration is increased 2 to 3 times by (+)-amphetamine and amfonelic acid in doses which evoke hyperactivity and stereotypic behaviour.Send offprint requests to S. B. Ross at the above address     

Transdermal Dual-Controlled Delivery of Contraceptive Drugs: Formulation Development,in Vitro and in Vivo Evaluations,and Clinical Performance

Several transdermal contraceptive device (TCD) formulations were developed to provide a dual-controlled transdermal delivery of levonorgestrel (LN), a potent progestin, and 17-estradiol (E₂), a natural estrogen. Using a sensitive HPLC method, the in vitro release and skin permeation profiles of LN and E₂ from various TCD formulations were simultaneously characterized in the hydrodynamically well-calibrated Valia–Chien skin permeation cells and both were found to follow zero-order kinetics. The rates of drug release and skin permeation were observed to vary significantly depending upon some formulation parameters. Six-month stability studies were performed on seven formulations at room and elevated temperatures (37 and 45°C), and two (Formulations 4 and 5) were found to be acceptable, based on drug recovery, release rate, and skin permeation rate data. Judging from the 6-month accelerated stability studies, it is projected these two formulations will have shelf-life of at least 2 years. As a result of development of an efficient manufacturing process, Formulation 4 was selected for further evaluation. One-week primary skin irritation evaluation in 6 rabbits indicated that Formulation 4 is nonirritating, and it was thus selected for Phase I clinical bioavailability/dose proportionality studies in 12 healthy female volunteers of child-bearing age. Results of pharmacokinetic and pharmacodynamic analyses demonstrated that it is capable of achieving and maintaining a steady-state serum level of LN throughout the 3-week treatment period by weekly applications of one or two TCD patches (10 or 20 cm²). A dose proportionality was obtained in the serum drug levels, daily dose delivered, and contraception efficacy. An excellent correlation was obtained for the rates of transdermal delivery determined by the in vitro studies using human cadaver skin, the in vivo studies in rabbits, and the clinical studies in living subjects.     

转FL、GM-CSF基因的人骨髓基质细胞促进脐血CD34+细胞体外扩增

研究转FL、GM-CSF基因的基质细胞对脐血CD34+细胞的扩增效应.将转FL、GM-CSF基因的入骨髓基质细胞系与脐血CD34+细胞共培养,观察细胞总数、CD34+细胞数、CFU-GM的变化情况.培养到第4周时,第(4)组(SCF+IL-3+IL-6+GM-CSF+FL)和第(8)组(HFCL/hGM-CSF+HFCL/hFL+SCF+IL-3+IL-6)的细胞总数增加到最大,分别扩增了717±24.47和709±63.63,第1周,第(5)组(HFCL+SCF+IL-3+IL-6)扩增了10.5±2.08倍,较第(8)组减少(P＜0.05).第1周时,CD34+细胞总数第(4)组和第(8)组分别扩增了8.44倍和11.5倍(P＜0.05),CD34+细胞百分率第(7)组(FCL/hFL+SCF+IL-3+II,-6)为50.2%,第(6)组(HFCL/hGM-CSF+SCF+IL-3+IL-6)为28.95%(P＜0.01).第2周,各组CFU-GM增加显著,以第(4)组和第(8)组增加最为明显,以后随扩增时间延长,造血细胞集落数、集落体积逐渐减少.表明转FL、GM-CSF基因的基质细胞,能有效的协同其他细胞因子对脐血CD34+细胞产生明显的扩增作用,能显著改变基质细胞造血功能.     

Propagation of hepatitis A virus in human embryo fibroblasts

human diploid fibroblasts and human primary liver cell carcinoma cells (PLC/PRF/5) were infected with hepatitis A virus (HAV) adapted to growth in cell culture or derived directly from human stool. Viral antigen was expressed in PLC/PRF/5 cells 28 days after infection with cell culture-adapted HAV, and 50 days after infection with virus from human stool. In human fibroblasts the periods until first expression of viral antigen were 90 and 210 days, respectively. During further passages of HAV in fibroblasts the time of first appearance of antigen decreased to about 28 days. Biophysical properties of HAV extracted from infected fibroblasts were comparable to those of HAV derived directly from human stool. Immunofluorescence studies showed that the antigen was located exclusively within the cytoplasm of the infected fibroblasts. Kinetics of antigen production indicated that an equilibrium between virus multiplication and cell metabolism was reached in persistently infected fibroblasts.