人主动脉硫酸乙酰肝素蛋白聚糖对培养的人血管壁细胞生长的作用

通过培养的人主动脉平滑肌细胞（ｈＡＳＭＣ）及脐静脉内皮细胞（ｈＵＶＥＣ）,应用３Ｈ－ＴｄＲ参入、Ｎｏｒｔｈｅｒｎｂｌｏｔ分析、逆转录多聚酶链反应（ＲＴ－ＰＣＲ）、放射免疫分析（ＲＩＡ）、和紫外比色法等技术观察了人主动脉中硫酸乙酰肝素蛋白聚糖（ＨＳＰＧ）对ｈＡＳＭＣ和ｈＵＶＥＣＤＮＡ合成的作用及对血小板源生长因子（ＰＤＧＦ）、ＰＤＧＦ受体、转化生长因子β（ＴＧＦ－β）、内皮素－１（ＥＴ－１）或碱性成纤维细胞生长因子（ｂＦＧＦ）基因表达和肾素－血管紧张系统（ＲＡＳ）的影响,结果显示,ＨＳＰＧ明显抑制培养的ｈＡＳＭＣ基础的ＤＮＡ合成（ｃｐｍ值为：１０３８５±３２６３ｖｓ,２５５４１±６４２１,Ｐ＜０．０１）及外源性ＰＤＧＦ诱导的ＤＮＡ合成（ｃｐｍ值为：９８７８±１９４７ｖｓ．１３４８１±４４ｌ０,Ｐ＜０．０５）;抑制ＰＤＧＦＡ链、ＴＧＦ－Ｂｐ和ＥＴ－１ｍＲＮＡ表达,提高ＰＤＧＦａ和β受体ｍＲＮＡ的表达;显著降低ｈＡＳＭＣ培养液中血管紧张素Ⅱ（ＡｎｇⅡ）的浓度和血管紧张素转换酶（ＡＣＥ）的活性,推测ＨＳＰＧ抑制ＰＤＧＦＡ链、ＴＧＦ－β及ＥＴ－１ｍＲＮＡ表达,降低ＡＣＥ活性及ＡｎｇⅡ浓度是其抑制ｈＡＳＭＣ增殖的重要机     

人主动脉硫酸乙酰肝素蛋白聚糖对培养的人血管壁细胞生长的作用

通过培养的人主动脉平滑肌细胞（ｈＡＳＭＣ）及脐静脉内皮细胞（ｈＵＶＥＣ）,应用＾３Ｈ－ＴｄＲ参入、Ｎｏｒｔｈｅｒｎ ｂｌｏｔ分析、逆转录多聚酶链反应（ＲＴ－ＰＣＲ）、放射免疫分析（ＲＩＡ）、和紫外比色法等技术观察了人主动脉中硫酸乙酰肝素蛋白聚糖（ＨＳＰＧ）对ｈＡＳＭＣ和ｈＵＶＥＣ ＤＮＡ合成的作用及对血小板源生长因子（ＰＧＤＦ）、ＰＧＤＦ受体、转化生长因子β（ＴＧＦ－β）、内皮素－１（ＥＴ－１）或     

低氧对肺动脉内皮细胞PDGF—B链释放及平滑肌细胞生长的影响

应用蛋白ｄｏｔｂｌｏｔ技术检测了低氧内皮细胞条件培养液（ＨＥＣＣＭ）和常氧内皮细胞条件培养液（ＮＥＣＣＭ）内ＰＤＧＦ相对含量,并利用［３Ｈ］－ＴｄＲ掺入法和流式细胞术观察了ＨＥＣＣＭ和ＮＥＣＣＭ及加入特异ＰＤＧＦ抗体对肺动脉平滑肌细胞（ＰＡＳＭＣ）生长的影响。结果表明,ＨＥＣＣＭ中的ＰＤＧＦ含量明显高于ＮＥＣＣＭ;ＨＥＣＣＭ能明显增强ＰＡＳＭＣ内ＤＮＡ合成,促进ＰＡＳＭＣ从Ｇｏ／Ｇ１期进入Ｓ期;当预先加入ＰＤＧＦ－Ｂ链抗体时,则会明显地抑制ＨＥＣＣＭ对ＰＡＳＭＣ的ＤＮＡ合成,阻止ＰＡＳＭＣ从Ｇｏ／Ｇ１期进入Ｓ期。结果提示,低氧时ＰＡＳＭＣ增殖与肺动脉内皮细胞分泌释放ＰＤＧＦ增加有关     

二甲亚砜与维甲酸对人卵巢癌细胞COC1的生长抑制及转化生长因子β_1表达的影响

本实验以人卵巢癌细胞株（ＣＯＣ１）为模型,观察诱导分化剂二甲亚砜（ＤＭＳＯ）与维甲酸（ＲＡ）对该细胞生长增殖、ＤＮＡ合成和转化生长因子β１（ＴＧＦβ１）在细胞内表达的影响。结果显示：ＤＭＳＯ与ＲＡ对人卵巢细胞ＣＯＣ１生长有明显的抑制作用,生长曲线表明作用５天后其生长抑制率分别为６２．７％和４２．１％;３Ｈ－胸腺嘧啶核苷（３Ｈ－ＴｄＲ）掺入实验说明ＤＭＳＯ组与ＲＡ组的单位时间计数率（ＣＰＭ）明显低于对照组（Ｐ＜０．０１）。用药３天后百分掺入抑制率分别为６０．４％与３７．９％,表明ＤＭＳＯ与ＲＡ抑制ＣＯＣ１细胞的ＤＮＡ合成;免疫细胞化学反应表明,ＤＭＳＯ或ＲＡ处理５天后,对照组细胞ＴＧＦβ１表达为阳性,定位于胞浆,而处理组细胞ＴＧＦβ１呈阴性或弱阳性反应。以上结果提示ＤＭＳＯ和ＲＡ对人卵巢癌细胞有一定的诱导分化作用。     

前列腺素F2α对大鼠肾小球系膜细胞DNA合成的影响

本实验应用（＾３Ｈ）胸腺嘧啶核苷（＾３Ｈ－ＴｄＲ）掺入法测定前列腺素Ｆ２α（ＰＧＦ２α）对大鼠肾小球系膜细胞ＤＮＡ合成的作用,测定系膜细胞合成的二脂酰甘油（ＤＡＧ）及磷酸肌醇（ＩＰ）。结果表明,ＰＧＦ２α促进系膜细胞的ＤＮＡ合成、ＤＡＧ及ＩＰ的生成。提示ＰＧＦ２α使系膜细胞的磷脂酶Ｃ活化,产生ＩＰ及ＤＡＧ,激活蛋白激酶Ｃ,从而促进ＤＮＡ合成及细胞增殖。     

血小板生长因子(PDGF-BB)刺激体外培养兔肺动脉平滑肌细胞DNA和蛋白质合成

应用细胞培养、３Ｈ－ＴｄＲ和３Ｈ－Ｌｅｕｃｉｎｅ掺入方法,观察血小板生长因子ＢＢ（Ｐｌａｔｅｌｅｔ－ｄｅｒｉｖｅｄＧｒｏｗｔｈＦａｃｔｏｒＢＢ）对体外培养兔肺动脉平滑肌细胞ＤＮＡ和蛋白质合成的影响。结果表明：（１）当ＰＤＧＦ－ＢＢ浓度为１０ｎｇ／ｍｌ时,３Ｈ－ＴｄＲ掺入值已较对照组显著增高（６２６２．５±４１２．９ｖｓ８３３．５±１２４．０,Ｐ＜０．０５）;当ＰＤＧＦ－ＢＢ浓度为２０ｎｇ／ｍｌ时,３Ｈ－Ｌｅｕｃｉｎｅ掺入值亦较对照线显著增高（１０２１２．８±６３８．３ｖｓ７３４０．３±１１９７．９,Ｐ＜０．０５）。（２）ＰＤＧＦ－ＢＢ浓度在５－２５ｎｇ／ｍｌ范围内,３Ｈ－ＴｄＲ,３Ｈ－Ｌｅｕｃｉｎｅ掺入值与剂量直线相关（ｒＤＮＡ＝０．９７,ｒｐｒｏｔ＝０．９０Ｐ＜０．０５）。说明ＰＤＧＦ－ＢＢ刺激体外培养兔肺动脉平滑肌细胞ＤＮＡ和蛋白质合成。     

大熊猫基因指纹探针F2ZGP96060801的研制及比较实验分析

利用ＡＢＩ－３９４型ＤＮＡ合成仪合成的寡核苷酸５－「Ａ（Ｘ）ｎ－ｘＴＣＣＡＣ」ｎ－３,经高效液相色谱仪纯化后,制备成了命名为为Ｆ２ＺＧＰ９６０６０８０１的大熊猫基因指纹探针,用同位素标记法标记Ｆ２ＺＧＰ９６０６０８０１,ＬＺＦ－Ｉ、朋５、３３．６和（ＣＡＣ）６／ＧＴＧ）５等５种基因指纹探针,比较检测了大熊猫随机个体的被毛、大熊猫３雄配Ｉ雌所产１个双胞胎计６只个体的福尔马林固定的肝组织和粪便中的胃肠     

细胞松驰素B促微丝解聚对DNA合成的作用

利用微丝（ｍｉｃｒｏｆｉｌａｍｅｎｔ,ＭＦ）解聚药物细胞松驰素Ｂ（ｃｙｔｏｃｈａｌａｓｉｎＢ,ＣＢ）处理Ｇ_０期小鼠Ｃ_３Ｈ_（１０）Ｔ_（１／２）成纤维细胞,对Ｇ_０至Ｓ期ＤＮＡ合成,胸腺嘧啶核苷激酶（ｔｈｙｍｉｄｉｎｅｋｉｎａｓｅ,ＴＫ）活性、ＴＫ基因表达、钙调素（ｃａｌｍｏｄｕｌｉｎ,ＣａＭ）水平和一些细胞周期早期基因的表达进行了观察,Ｇ_０期细胞经３ｍｇ／ＬＣＢ处理２ｈ,促ＭＦ解聚增强了血清对Ｓ期细胞ＴＫ活性、ＴＫ基因表达和ＤＮＡ合成的刺激作用,并促进细胞提前进入Ｓ期．血清刺激Ｇ_０期细胞进入晚Ｇ_１期和Ｓ期时,ＣａＭ水平明显升高,而ＣＢ预处理则使ＣａＭ含量进一步增加,特别是ＣＢ处理促使Ｓ期ＣａＭ增加向核内转移．ＣＢ处理明显增强血清对ｃ－ｊｕｎ、ｃ－ｆｏｓ和ｃ－ｍｙｃ基因表达的刺激作用,而ＰＫＣ抑制剂Ｈ_７则抑制ＣＢ处理对这些基因转录的刺激作用,说明ＣＢ使Ｇ_０期细胞ＭＦ解聚刺激ｃ－ｊｕｎ、ｃ－ｆｏｓ和ｃ－ｍｙｃ的转录活性与ＰＫＣ的作用有关．结果表明Ｇ_０至Ｓ期早期ＭＦ的重组可促进细胞进入Ｓ期,增强ＤＮＡ合成．     

乙型肝炎病人重叠感染丙型肝炎的评价

应用ＥＬＩＳＡ和ＰＣＲ法检测５０２例乙肝病人血清,４０１例ＨＢｓＡｇ阳性血清中,有１１４例（２８．４％）抗－ＨＣＶ和ＨＣＶＲＮＡ双项阳性,２５例（６．２％）ＨＣＶＲＮＡ单项阳性;２１例（５．２％）抗－ＨＣＶ单项阳性。将ＨＢｓＡｇ乙肝病人分成ＨＢＶＤＮＡ,ＨＢｅＡｇ阳性组和ＨＢＶＤＮＡ,ＨＢｅＡｇ阴性组。前者抗－ＨＣＶ阳性率为１１．６％～２０．５％,ＨＣＶＲＮＡ阳性率为１６．２％～２０．５％。后者抗－ＨＣＶ阳性率为２０．２％～５５．６％,ＨＣＶＲＮＡ阳性率为２３％～６０．３％。结果说明长期携带ＨＢＶ者和慢性乙肝病人均可重叠ＨＣＶ感染。ＨＢＶＤＮＡ阳性组抗－ＨＣＶ和ＨＣＶＲＮＡ阳性率明显高于ＨＢＶＤＮＡ阳性组     

转译优化对EPO基因在COS7细胞中表达的影响

利用ＣＯＳ７细胞暂时表达系统,研究转译起始序列对ＥＰＯ－ｃＤＮＡ表达的影响。通过ＤＮＡ重组技术,构建了原ＥＰＯ－ｃＤＮＡ表达载体ｐＣＳＶ－ＥＰＯ（１）,其转译起始序列为５＇ＡＡＴＴＣＡＴＧＧ３＇。同时通过定点突变技术,将起始序列改变成５＇ＣＣＡＣＣＡＴＧＧ３＇,而构建了另一表达载体ＰＣＳＶ－ＥＰＯ（２）。后经序列分析证明无误后和前均通过ＤＥＡＥ－ｄｅｘｔｒａｎ法转染ＣＯＳ７细胞上清,测定结果为     

Partial purification and characterization of a growth factor present in goat's colostrum. Similarities with platelet-derived growth factor.

A factor in goat's colostrum which stimulates DNA synthesis and cell proliferation in Swiss 3T3 fibroblasts has been purified approx. 350-fold by a sequence of acid precipitation, cation-exchange chromatography and gel filtration. The growth factor is a highly basic, heat stable (100 degrees C for 5 min) polypeptide with Mr approx. 35000. The polypeptide resists denaturation by guanidinium chloride or urea but is totally inactivated by treatment with reducing agents. The factor, which we have termed colostric basic growth factor ( CBGF ), inhibits the binding of 125I-labelled epidermal growth factor (125I-EGF) to Swiss 3T3 fibroblasts but does not inhibit 125I-EGF binding to epidermoid A431 cells. CBGF interacts synergistically with plasma in stimulating DNA synthesis in quiescent Swiss 3T3 cells. The chemical and biological properties of CBGF are thus very similar to the properties reported for the human platelet-derived growth factor. Although high concentrations of CBGF are present in the colostrum of goats, cows, and sheep, the milk of these species contains little or no factor. The origin and possible functions of CBGF are unknown.     

Characterization of growth factors in human cartilage

Growth factor activity has been identified in the chondrocytes and extracellular matrix (ECM) fractions of human costal cartilage. There was about five times more growth factor activity in the ECM than was found to be associated with the chondrocytes. The growth factor activity in chondrocytes was found to be associated with chromatin. Both the chromatin-associated growth factor (CAGF) activity and extracellular matrix growth factor (EMGF) activity were characterized for molecular weight, charge, and the effect of reduction by sulfhydryl reducing reagents. Biorex cation exchange chromatography showed that both CAGF and EMGF were cationic. CAGF and EMGF have molecular weights between 15,000 and 18,000 as determined by size exclusion chromatography on HPLC TSK 3000 columns equilibrated with guanidine-HCl and dithiothreitol.     

A uterine cell mitogen distinct from epidermal growth factor in porcine uterine luminal fluids: characterization and partial purification

Uterine luminal fluids (ULF) from early (Days 10 and 12)-pregnant sows contain factors that stimulate DNA synthesis in a variety of cell lines. The major growth factor component in these fluids has been partially purified 200-fold by heat treatment, anion-exchange chromatography, and gel filtration using mouse embryo-derived AKR-2B fibroblasts as an indicator cell line. The ULF mitogen (ULFM) is a polypeptide with an apparent molecular weight of 4800; it is extremely heat stable and resistant to treatment with urea. This mitogen is also present in ULF from cycling sows but is not detectable in uterine cytosolic extracts or in serum isolated from pigs at Day 12 of pregnancy. The addition of this factor to medium containing 0.5% calf serum results in a 50% increase in final cell density of AKR-2B cells. ULFM appears biologically distinct from mouse and human epidermal growth factor (EGF), since its activity is not inhibited by antibody to mouse EGF and it does not compete for binding to human (A431) EGF receptors. In addition, the ULF factor stimulates DNA synthesis in human A431 epidermoid carcinoma cells, whereas EGF is inhibitory. Partially purified ULFM also stimulates DNA synthesis in primary cultures of pig uterine stromal cells. This mitogen activity is dose-dependent and is not inhibited by antibody to mouse EGF. Thus ULFM may act in concert with other peptide growth factors in regulating uterine growth and/or differentiation.     

Partial purification of a low-molecular-weight growth factor from chicken brain

The regenerating amphibian limb serves as a useful model for studying factors influencing cell proliferation and differentiation. In particular, peripheral nerves are thought to provide a stimulus for growth of the blastema, presumably via the elaboration of an as yet unidentified neurotrophic factor. In the present study, pressure ultrafiltration coupled with chromatofocusing have proven to be effective methods of partially purifying a neurotrophic factor from adult chicken brains. This chick brain growth factor (CBGF) appears to be a heat-stable, basic peptide of low molecular weight (less than 6,000). It is a potent mitogen in vitro, at nanomolar concentrations, for both blastema cells and Swiss mouse 3T3 fibroblasts. CBGF is apparently distinct from other peptide mitogens and/or neuromodulators that have been reported to stimulate blastema growth in vivo and in vitro. These include substance P, FGF from bovine brain and pituitary, EGF, transferrin (sciatin), and spinal cord growth factor (SCGF). The possible relationship of CBGF to other neural regulatory molecules is discussed.     

Purification and characterization of a bovine colostrum-derived growth factor

A growth factor in bovine colostrum was purified to homogeneity by a combination of acid extraction, boiling, cation exchange chromatography, isoelectric focusing, and reverse phase HPLC. The bovine colostrum growth factor (BCGF) had an isoelectric point of about 10, a native mol wt of about 30,000, was resistant to inactivation by boiling and exposure to pH 1, but was inactivated by dithiothreitol. BCGF appeared to be structurally related to human platelet-derived growth factor (PDGF) and competed with human PDGF in a radioreceptor assay. However, while human PDGF appeared to be a heterodimer of 17,000 and 14,000 mol wt subunits, BCGF appeared to be a homodimer of 20,000 mol wt subunits. Purified BCGF had a specific activity in stimulating 3T3 cell proliferation of about 3-6 U/ng and was active at about 1-2 ng/ml.     

Isolation of colony stimulating factor from human milk

Human milk contains colony stimulating factor (CSF), a polypeptide growth factor, which stimulates in in vitro bone marrow culture proliferation and differentiation of colony forming granulocytic macrophage progenitor cells (CFU-GM) to form colonies. This activity was not found in either bovine milk or colostrum when assayed in human or mouse bone marrow cells. The human milk CSF activity is destroyed by treatment with proteases. However, neither 6M urea, 4M guanidine hydrochloride, 5 mM dithiothreitol, nor exposure to pH 2 will inactivate the milk derived CSF. Gel filtration and isoelectric focusing indicate that human milk CSF differs biochemically from the other CSFs isolated from various sources and has a molecular weight between 250,000 and 240,000 and an isoelectric point between 4.4 and 4.9.     

Enhancement of DNA synthesis of human epithelial carcinoma cells by acidic fibroblast growth factor

Human epithelial cells that had grown out from a maxillary carcinoma were examined for their responsiveness to putative growth-controlling factors in a serum-free medium. Among the factors examined, bovine brain acidic fibroblast growth factor (FGF) at 1 to 10 ng/ml significantly promoted DNA synthesis of the cells in the presence of 5 U/ml heparin, whereas type beta transforming growth factor inhibited it in a dose-dependent manner. Fetal bovine serum at 0.6% inhibited DNA synthesis of the cells by approximately 15%, but no significant influence was observed at higher concentrations up to 10%. Epidermal growth factor, bovine pituitary gland FGF and basic FGF exhibited no significant effect on DNA synthesis of the cells. The present result suggests that acidic FGF, a known mitogen for endothelial cells, is also mitogenic for human epithelial cells derived from maxillary carcinoma.     

IGF-I, IgA, and IgG responses to bovine colostrum supplementation during training.

This study examined the effect of bovine colostrum (Dynamic colostrum) supplementation on blood and saliva variables (study 1) and the absorption of orally administered human recombinant insulin-like growth factor (IGF)-I (rhIGF-I) labeled with 123I (123I-rhIGF-I) (study 2). In study 1, adult male and female athletes were randomly assigned in a double-blind fashion to either an experimental (Dynamic; n = 19) or a control (Placebo; n = 11) group. The former consumed daily 20 g of Dynamic supplement, and the latter 20 g of maltodextrin during a 2-wk training period. After bovine colostrum supplementation, significant increases were noticed in serum IGF-I (P < 0.01) and saliva IgA (P < 0.01) in Dynamic compared with Placebo. In study 2, gel electrophoresis was carried out in 12 adult subjects with serum samples taken 60 min after ingestion of 123I-rhIGF-I and showed peaks at 0.6 and at 40-90 kDa, with the former inducing 96% and the latter 4% of the total radioactivity. It was concluded that a long-term supplementation of bovine colostrum (Dynamic) increases serum IGF-I and saliva IgA concentration in athletes during training. Absorption data show that ingested 123I-rhIGF-I is fragmented in circulation and that no radioactive IGF-I is eluted at the positions of free, or the IGF, binding proteins, giving no support to the absorption of IGF-I from bovine colostrum.     

The serum-free growth of cultured cells in bovine colostrum and in milk obtained later in the lactation period

Seven established cell lines, including both epithelial cells and fibroblasts (MDCK, Vero, CV-1, NRK, 3T3, F2408, and NIL8) and four early passage cell strains (bovine articular chondrocytes, bovine smooth muscle cells, human foreskin fibroblasts, and rat embryo cells) were cultured in serum-free medium supplemented with milk obtained 1 day after birth (colostrum) or 80 days after birth (older milk). MDCK, Vero, CV-1, NRK, and 3T3 grew readily in colostrum and attained saturation densities ranging from 22% to 63% of that in serum. There was no growth of F2408, NIL8, or the early passage strains in bovine colostrum. None of the 11 cell cultures grew in older milk. The temporal dependence of growth in milk was examined in detail using MDCK cells. Growth equivalent to that in serum occurred in 3% colostrum and in 15% milk obtained 2 days after birth. Milk obtained 3 days and 10 days after birth was not effective as a growth supplement for MDCK cells at any concentration. Those cells, unable to grow in colostrum or in older milk, could be induced to grow if culture dishes were precoated with fibronectin. In addition to fibronectin, it was necessary in some cultures to supplement colostrum or older milk with insulin and/or transferrin in order to achieve growth. In the presence of fibronectin and appropriate factors, the final saturation density attained in colostrum or older milk ranged from 25% to 100% of that in serum. The fibronectin contents of bovine colostrum and milk were determined. The fibronectin level of colostrum was found to be approximately 5% of bovine serum. There was no detectable fibronectin in the 80-day-old milk.     

Purification and partial sequence analysis of insulin-like growth factor-1 from bovine colostrum.

Growth-promoting activity in bovine colostrum has been detected as the capacity to stimulate protein synthesis in L6 myoblasts. By using this assay as a measure of bioactivity, a growth factor has been purified to near homogeneity from centrifuged colostrum by a series of steps including acid extraction, chromatography on sulphopropyl-Sephadex, followed by adsorption to, and elution from, C18 columns using acetonitrile and propan-1-ol gradients. The purified growth factor has a low solubility at neutral and alkaline pH and has an Mr of 7800 by gel-permeation chromatography. Sequence analysis of the first 30 amino acids from the N-terminus indicated complete identity in this region with human insulin-like growth factor-1. Accordingly we conclude that the purified growth factor is bovine insulin-like growth factor-1.