Isolation and propagation of yolk-sac-derived endothelial cells from a hypervascular transgenic mouse expressing a gain-of-functionFPS/FES proto-oncogene

Summary We report on the isolation and propagation of endothelial cells from the mouse embryonic yolk sac, the earliest site of blood vessel development, and on the advantages of a hypervascular transgenic mouse source of these cells. These transgenic mice express multiple copies of an activated allele of the humanfps/fes proto-oncogene and display hypervascularity progressing to multifocal hemangiomas. This phenotype suggested a role of thefps/fes proto-oncogene in vasculogenesis and angiogenesis and led us to investigate the growth characteristics of yolk-sac-derived endothelial cells from transgenicfps/fes embryos. We have established eight independent cell clones from a mixture of transgenic and control yolk sacs from Day 12 embryos. Southern blot hybridization analysis showed all eight clones to be derived from transgenic cells suggesting a growth advantage of cells carrying the activatedfps/fes gene. A cell line, Clone 166 (C166), established from one of these clones, was more fully characterized. C166 exhibits normal endothelial characteristics, such as rearrangement into tubelike structures when placed on Matrigel, expression of angiotensin converting enzyme, retention of cobblestone morphology at confluence, and the presence of cell surface receptors for acetylated low density lipoprotein. The cells constitutively express murine endothelial cell adhesion molecule VCAM-1 and the vascular addressin identified by antibody MECA-99. As expected, the cell line expresses high levels of the cytoplasmic protein-tyrosine kinase encoded by thefps/fes proto-oncogene. The clone we have described as well as other endothelial cell lines that we have established from the mouse embryonic yolk sac should prove useful for the study of endothelial cell differentiation and for the determination of the mechanisms underlying the establishment of organ-specific endothelial cell heterogeneity.     

Abrogation of the Fc gamma receptor IIA-mediated phagocytic signal by stem-loop Syk antisense oligonucleotides.

The role of Syk kinase in Fc gamma receptor (Fc gamma R) IIA-mediated phagocytosis was examined with two forms of antisense oligodeoxynucleotides (ODNs) designed to hybridize to human Syk mRNA. Monocytes were incubated with linear and stem-loop antisense ODNs targeted to Syk mRNA. When complexed with cationic liposomes, stem-loop Syk antisense ODN with phosphorothioate modification exhibited stability in fetal bovine and human serum. The stem-loop Syk antisense ODN at a concentration of 0.2 microM inhibited Fc gamma RIIA-mediated phagocytosis by 90% and completely eliminated Syk mRNA and protein in monocytes, whereas scrambled-control ODNs had no effect. The Syk antisense ODNs did not change beta-actin mRNA levels and Fc gamma RII cell-surface expression. In addition, stem-loop Syk antisense ODN inhibited Fc gamma RI and Fc gamma RIIIA-mediated phagocytosis. These data indicate the efficacy of stem-loop Syk antisense ODN for targeting and degrading Syk mRNA and protein and the importance of Syk kinase in Fc gamma receptor-mediated phagocytosis. Immunoblotting assay demonstrated that Fc gamma RII tyrosine phosphorylation after Fc gamma RII cross-linking did not change in the absence of Syk protein. These results indicate that Syk kinase is required for Fc gamma RIIA-mediated phagocytic signaling and that Fc gamma RII cross-linking leads to tyrosine phosphorylation of Fc gamma RII independent of Syk kinase.     

GM_3对小鼠腹腔巨噬细胞磷脂代谢转换率的影响

神经节苷脂ＧＭ_３对小鼠腹腔常驻巨噬细胞（Ｒ－Ｍ）和Ｇｅ－１３２体内激活的巨噬细胞（Ｇｅ－１３２－Ｍ）的磷脂代谢转换有显著的影响,当这两种Ｍ在体外用ＧＭ_３处理时,表现出［￣（３２）Ｐ］Ｐｉ和［￣３Ｈ］肌醇参入ＰＩ降低,参入ＰＩＰ、ＰＩＰ_２增加;但在［￣（３２）Ｐ］Ｐｉ和［￣３Ｈ］胆碱参入ＰＣ上,Ｒ－Ｍ与Ｇｅ－１３２－Ｍ不同,即ＧＭ_３促进同位素前体参入Ｒ－Ｍ的ＰＣ,抑制它们参入Ｇｅ－１３２－Ｍ的ＰＣ．以上结果表明ＧＭ３可能提高了ＰＩ或ＰＩＰ的磷酸激酶的活性,致使［￣（３２）Ｐ］ＰＩＰ和［￣（３２）Ｐ］ＰＩＰ_２增多,［￣（３２）Ｐ］ＰＩ减少．激活的Ｍ（Ｇｅ－１３２－Ｍ）本身ＰＣ代谢转换率较Ｒ－Ｍ高,当Ｇｅ－１３２－Ｍ再受ＧＭ_３刺激,ＰＣ代谢转换率降低,这提示ＧＭ_３对激活的Ｍ的ＰＣ代谢转换有调节作用．     

用细胞色素C法检测非真空电子束辐射的间接作用

本文提出了一种检测非真空电束辐射的间接作用的一种方法,根据氧化型和还原型的细胞色素Ｃ的吸收光谱不同,从而测量非真空电子束作用对应的辐射分解的自由基总量。     

人胸腺素α原cDNA的克隆及在大肠杆菌中的表达

采用基因工程方法制取人胸腺素α原获得成功。用２０ｕｇ／ｍｌ植物血球凝集素（ＰＨＡ）和５００Ｕ／ｍｌ重组人白细胞介素２（ＩＬ－２）联合刺激人胎儿胸腺细胞,从中提取总ＲＮＡ,经反转录ＰＣＲ获得了人胸腺素α原ｃＤＮＡ;将之克隆入ｐＵＣ１９中,序列测定表明与已报道序列一致,进一步将之亚克隆入原核表达载体ｐＢＶ２２０,转化大肠杆菌ＤＨ５ａ．观察到在不改变氨基酸编码的前提下,增加胸腺素ａ原上游引物中Ａ、Ｔ含量,可以明显提高胸腺素α原的表达量,同时,不同培养基对它的表达也有影响。胸腺素α原在大肠杆菌中以可溶形式表达,不需复性。初步活性测定显示,它可明显刺激人外周血淋巴细胞Ｅ－玫瑰花结形成率。重组人胸腺素α原在大肠杆菌中表达,为其临床应用及基础研究奠定了基础。     

内蒙古草原退化群落恢复演替的研究II. 恢复演替时间进程的分析

 根据内蒙古典型草原地带的羊草+大针茅草原退化变型一冷蒿群落封育12年(1983—1994)的动态监测数据进行分析,对群落恢复演替轨迹取得以下认识： 1．依据群落优势种的更替及主分量分析结果可将恢复演替过程划分为冷蒿优势阶段、冷蒿+冰草阶段、冰草优势阶段、羊草优势阶段。 2;退化草原群落在恢复演替过程中,群落生产力的变化表现出阶梯式跃变和亚稳态阶面相间的特点。第一次跃变发生在1984年,上升到第二个阶面,第二次跃变发生在1990年,进入了第三个阶面,已接近于原生群落的生产力。 3．群落生产力与水资源量的关系因恢复演替阶段不同而异。第一亚稳态时期,群落地上现存生物量大体处于166g·m-2的水平上,生长季降水量达176mm以上时,增加降水对群落生产力的提高不发生显著影响。第二亚稳态时期,群落生物量与降水量之间的相关性显著。可推算出群落于物质生产用水量介于1.1～1.6mm·g-1之间。此值在1.1mm·g-1时,群落对水资源的利用效率最高,而在1.6mm·g-1时群落生物量达到最大值。 4．在恢复演替进程中,群落密度的位点常数约为271.5株·m-2,循此常数上下波动,表现出拥挤与稀疏交替发生的过程,构成了恢复演替的节奏性变化。群落生物量的跃变与亚稳态的形成,以及群落密度的拥挤与稀疏交替作用是群落恢复演替的内在机制。恢复演替的速度,到第10年发生了1.78个半变的生态距离。5．草原退化群落恢复演替过程中,按照其节奏性及生产力跃变与亚稳态的规律,调控放牧利用强度或采取技术措施,调节群落拥挤和稀疏的交替过程可加速恢复演替进程。     

松嫩草原碱化草地角碱蓬群落水分生态的研究

 本文采用自行设汁的蒸散仪和加拿大Campbe 11科学仪器公司生产的自动气象设备测定了松嫩草原碱化草地角碱蓬群落的蒸散、蒸腾量、太阳辐射及空气温度等环境因子。分析结果表明生长季的睛天条件下,角碱蓬群落的蒸散、蒸腾速率的日进程均为单峰曲线,且各月份间差异很大。群落蒸腾速率与太阳辐射强度、空气温度、相对湿度、风速等环境因子紧密相关,其中与太阳辐射强度呈极显著正相关关系。生长季降雨量和土壤含水量在角碱蓬群落水分循环与平衡的过程中起重要调节作用。1992年6～8月的生长季中,角碱蓬群落总的水分亏缺较少(6.3mm),但各月份间差异很大,其中6月份水分亏缺最高(30.1mm)。     

献血员抗－HCV检测报告

对１５８７名献血员进行了抗－ＨＣＶ检测,总阳性率５．６０％,在不同职业中农民较高,占６．７２％,３０岁以上为１２．７６％。我站自改进采浆工艺,严格使用一次性消毒器具后,未发现ＨＣＶ交叉感染。职业献血者抗－ＨＣＶ阳性率高达３５．４８％,易造成输血后ＨＣＶ感染。对ＨＣＶ感染ＯＤ值阳性３０人又进行４－６个月的观察复测,其中１６人ＯＤ值下降,６人上升,认为ＯＤ值的变化与自限性ＨＣＶ感染有关。     

LDL受体基因cDNA的RT－PCR分离、克隆及其在RFLP中的应用

利用ＲＴ－ＰＣＲ技术分离低密度脂蛋白受体基因ｃＤＮＡ,将长为２６０５ｂｐ的ｃＤＮＡ插入质粒ｐＪＮ６,并经全序列测定证实,该序列与已报道序列相比,存在两个变异碱基,即第７５４位Ｃ→Ｔ,和１６５４位的Ｇ→Ａ,这两个变异碱基并不改变编码的氨基酸,利用ＬＤＬ受体基因ｃＤＮＡ中的ＣｌａⅠ片段作为探针,检测到ＬＤＬ受体基因上外显子８的ＳｔｕⅠ位点是个限制性片段长度多态性位点。所克隆的ｃＤＮＡ含有可译框架的全部密码,因此可作为基因表达材料。