Novel in vitro and in vivo inhibitors of prolyl endopeptidase

Inhibition of prolyl endopeptidase by Z-cyclohexyl prolinal and Z-indolinyl prolinal occurs with slow, tight binding inhibition and K_i values of 2 – 3 nM. In vivo enzyme inhibition is also observed with a half time for recovery of enzyme activity of 3 – 4 h.Inhibition of prolyl endopeptidase by Z-cyclohexyl prolinal and Z-indolinyl prolinal occurs with slow, tight binding inhibition and K_i values of 2 – 3 nM. In vivo enzyme inhibition is also observed with a half time for recovery of enzyme activity of 3 – 4 h.     

Expression of streptococcal plasmid-determined resistance to erythromycin and lincomycin in Escherichia coli

Summary By using recombinant DNA techniques, we have constructed a chimeric plasmid, pSM7322 (10.8 kb), between the streptococcal erythromycin resistance vector plasmid pSM7 (6.4 kb) and the E. coli vector pBR322 (4.4 kb). As judged by the minimum inhibitory concentrations of erythromycin and lincomycin, pSM7-determined resistance to these antibiotics is expressed in E. coli.     

Radiosensitization mechanism of riboflavin in vitro

Riboflavin, suggested to be a radiosensitizer, was studied in murine thymocytes and human hepatoma L02 cell line in vitro with MTT method and fluorescence microscopy. When the murine thymocytes treated with 5–400 μmol/L riboflavin were irradiated by 5 Gy ⁶⁰Co γ ionizing radiation, the low concentration groups, i.e. treated with 5–50 μmol/L riboflavin, showed a different surviving fractions-time relating correlation compared with the high concentration groups, i.e. treated with 100–400 μmol/L riboflavin. The former had a high survival level at the end of irradiation, but which, after 4-h incubation, decreased rapidly to a low level. On the contrary, the high concentration groups showed a low survival level at the end of irradiation, and a poor correlation was found between the surviving fraction and the incubation time, after 4 h a little difference was observed. The results of fluorescence microscopy indicated that under low concentration conditions, the riboflavin localized mainly in nucleus (both perinuclear area and inside of nuclear membrane), while under high concentration conditions, intensive riboflavin also localized around cytoplasmic membranes. Thus we can conclude: the riboflavin had radiosensitivity effect on DNA under low concentration conditions, and enhanced the damage to cytoplasmic membrane under high concentration conditions. Also the most effective concentration of riboflavin can be evaluated to be approximate 100 μmol/L.     

Phagocytosis by carp granulocytes; in vivo and in vitro observations

The phagocytic ability of carp (Cyprinus carpio L.) granulocytes was evaluated in vivo and in vitro. In suspensions of head kidney cells, neutrophil granulocytes incorporated both latex beads and coccidian merozoites. In intestinal tissues from carp with a Goussia carpelli infection, all granulocyte cell types (neutrophils and cells of the basophilic-eosinophilic complex) phagocytosed cell detritus and coccidian developmental stages, mainly merozoites.     

Dietary phytoestrogens and cancer: in vitro and in vivo studies

Thirty postmenopausal women (11 omnivores, 10 vegetarians and 9 apparently healthy women with surgically removed breast cancer) were investigated with regard to the association of their urinary excretion of estrogens, lignans and isoflavonoids (all diphenols) with plasma sex hormone binding globulin (SHBG). A statistically significant positive correlation between urinary total diphenol excretion and plasma SHBG was found which remained statistically significant after elimination of the confounding effect of body mass determined by body mass index (BMI). Furthermore we found a statistically significant negative correlation between plasma SHBG and urinary excretion of 16α-hydroxyestrone and estriol which also remained significant after eliminating the effect of BMI. Furthermore we observed that enterolactone (Enl) stimulates the synthesis of SHBG by HepG2 liver cancer cells in culture acting synergistically with estradiol and at physiological concentrations. Enl was rapidly conjugated by the liver cells, mainly to its monosulfate. Several lignans and the isoflavonoids daidzein and equol were found to compete with estradiol for binding to the rat uterine type II estrogen binding site (the s.c. bioflavonoid receptor). It is suggested that lignans and isoflavonoids may affect uptake and metabolism of sex hormones by participating in the regulation of plasma SHBG levels and in this way influence their biological activity and that they may inhibit cancer cell growth like some flavonoids by competing with estradiol for the type II estrogen binding sites.     

药用植物红根草种质资源的离体保存

以红根草试管苗为材料,研究了不同培养基(MS、1/2MS、1/4MS)、蔗糖浓度和植物生长抑制剂(CCC、PP333、ABA、MH)在红根草试管苗保存中的作用。结果表明:培养基1/2MS对红根草保存最好,保存270d后存活率最高。培养基中不添加蔗糖较添加一定浓度蔗糖时植株的形态和色泽差,但保存时间更长,转继代后能正常恢复生长。添加生长抑制剂能减缓生长速度,延长保存时间,最佳浓度分别为:CCC1.2～1.6mg/L;PP3331.6mg/L;ABA0.5～4.0mg/L;MH0.5mg/L。其中,ABA0.5～4.0mg/L对植株的生长最好,CCC浓度为1.2mg/L和1.6mg/L时,保存时间长,360d时,存活率达90%。     

匙羹藤组织培养条件优化研究

以匙羹藤无菌种子苗为外植体,通过预备实验确定基本培养基后,采用完全实验和正交设计相结合的方法,研究植物生长调节剂两因素配合和多因素组合(NAA、TDZ、2-ip、IAA、6-BA和KT)对匙羹藤芽体诱导及继代培养的影响。结果显示,匙羹藤组织培养最适的基本培养基为N68;N68+NAA0.05mg/L+2-ip0.4mg/L+TDZ0.001mg/L利于芽体增殖,可用于继代培养;N68+NAA0.05mg/L+TDZ0.005mg/L促进植株长根,可用于生根培养。     

Influence of in vitro growth conditions on in vitro and ex vitro photosynthetic rates of easy- and difficult-to-acclimatize sea oats (Uniola paniculata L.) genotypes

Net photosynthetic rates (P _n) of easy (EK 16-3) and difficult-to-acclimatize (EK 11-1) sea oats genotypes were examined under the following culture conditions: (1) photoautotrophic [sugar-free medium, high photosynthetic photon flux (PPF), high vessel ventilation rates and CO₂ enrichment, (PA)]; (2) modified photomixotrophic [sugar-containing medium diluted with sugar-free medium over time, high PPF, and high vessel ventilation rates (PM)]; (3) modified photomixotrophic enriched [same as PM with CO₂ enrichment, (PME)]; or (4) conventional photomixotrophic [sugar-containing medium, low PPF, and low vessel ventilation rates (control)]. Regardless of genotype, plantlets cultured under PA conditions died within 2 wk, whereas under PM and PME conditions, plantlets increased their P _n. After 6 wk, P _n per gram dry weight was 1.7 times greater in EK 16-3 than EK 11-1 plantlets cultured under PME conditions. In vitro-produced leaves of EK 16-3 plantlets were elongated with expanded blades, whereas EK 11-1 produced short leaves without expanded blades, especially under control conditions. After in vitro culture, EK 16-3 PME plantlets exhibited the highest dry weights among treatments. EK 16-3 PME and EK 16-3 PM had similarly high survivability, shoot and root dry weights and leaf lengths ex vitro compared to EK 16-3 control and EK 11-1 PM and PME plantlets. Ex vitro growth, survivability and P _n per leaf area of either genotype were not affected by CO₂ enrichment under modified photomixotrophic conditions. These results suggest that growth and survivability of sea oats genotypes with different acclimatization capacities can be enhanced by optimizing culture conditions.     

单增李斯特菌胎盘感染的体内外模型研究进展

单核细胞增生李斯特氏菌（Listeria monocytogenes）是重要的食源性致病菌,能引发人类的李斯特菌病,是全球公共卫生问题之一。该菌易感染孕妇,引起胎儿和新生儿的侵袭性李斯特菌病,严重威胁母婴健康。因此,建立有效的单增李斯特菌感染胎盘体内外模型,解析和探究单增李斯特菌经胎盘感染机制,是预防和控制单增李斯特菌感染母婴的关键所在。本文综述了可用于研究单增李斯特菌母婴感染的体内外胎盘模型,总结和讨论了各类模型的优势和局限性;并着重分析了体外三维胎盘屏障模型在单增李斯特菌感染方面的研究进展和未来研究方向。以期为深入解析该菌经胎盘感染的途径、发病机制提供支持,并为预防和控制母婴李斯特菌病提供科学参考。     

Cartilage induction in vitro : Ultrastructural studies

When 19-day fetal rat triceps muscle was cultured for 7 to 14 days upon decalcified, sequentially extracted adult rat bone, cartilage formed within clefts and vascular spaces of the decalcified bone. The bone substrata were prepared by extracting tibias and femurs of Sprague-Dawley rats with 1:1 chloroform:methanol, 0.6 N HCl, 2 M CaCl₂, 0.6 M EDTA, 8 M LiCl, and H₂O at 56°C. The culture medium used was CMRL 1066 with 15% newborn calf serum. During cultivation, fibroblastic mesenchymal cells migrated out of muscle and into bone crevices where they secreted a cartilaginous matrix composed of thin, randomly dispersed collagen fibrils and proteoglycan granules. The latter are characteristic for cartilage matrix. Extracted bone matrix contained mature collagen fibrils, some of which retained their typical 640-Å banding. Other collagen fibrils were partially disaggregated and expanded to reveal component 50-Å-thick, beaded micro fibrils. Such an expansion of collagen fibrils is known to result from exposure to proteoglycan solvents such as 2 M CaCl₂. The decalcified bone matrix contained many residual devitalized cells and cell fragments which often were seen in close proximity to chondrifying mesenchymal cells. This finding indicates the possibility that residual cellular material could play a role in stimulating cartilage development.     

睡菜离体快繁技术的研究

以睡菜的幼嫩茎段为外植体,接种到附加不同浓度激素配比(6-BA/NAA)的MS培养基,诱导睡菜愈伤组织、芽及根的生长。研究发现,外植体在1.0mg/L 6-BA+0.1mg/L NAA+MS的培养基上培养10d,可观察到浅绿色的愈伤组织。愈伤组织转接到4.0mg/L 6-BA+0.3mg/L NAA+MS培养基上2周左右可生成芽。对带芽的愈伤组织再进行诱导生根进而形成完整再生植株,最适根诱导培养基为0.3mg/L 6-BA+1.0mg/L NAA+MS培养基。该实验采用植物离体快繁技术成功建立了睡菜再生体系,为睡菜种苗规模化奠定了技术基础。     

斑马鱼胸苷酸合成酶与自身mRNA的相互作用

斑马鱼(zebrafish,Danio rerio)是生物学领域中公认的研究脊椎类生物的模式生物.胸苷酸合成酶(thymidylate synthase,TS)是DNA从头合成的限速酶,多年来一直作为肿瘤化疗的重要靶酶.前期的研究表明,人和大肠杆菌中TS能与自身的mRNA结合,在翻译水平上具有反馈抑制自调控现象.斑马鱼作为药物模型的研究已成为热点研究领域,为了探讨斑马鱼的胸苷酸合成酶的调控规律,以及与人TS的相关性,利用原核表达,纯化获得高均一性斑马鱼TS蛋白,采用凝胶迁移研究了TS和其mRNA的体外结合,采用免疫共沉淀:RT-PCR技术研究了它们在体内的相互作用,实验结果表明,斑马鱼的TS在体内外均与自身的mRNA存在特异性的相互作用.研究说明,斑马鱼和人的TS具有高度生物学过程相关性,为构建斑马鱼抗肿瘤药理模型提供了理论基础.     

丁香(Syringa L.)种间杂交幼胚离体培养研究

采用幼胚离体培养方法克服幼胚败育从而直接获得杂种实生苗,对影响丁香幼胚培养成功的各种因子作了较系统的研究.结果表明,丁香幼胚培养的最适培养基为Monnier;最佳糖浓度为50g·L^-1;幼胚在50～60d胚龄时培养最容易成功;加入适量的椰乳、谷氨酰胺或谷氨酸及活性炭可以促进幼胚的萌发和生长.低浓度(0.01mg·L^-1)BA的加入可以提高幼胚的萌发率;NAA浓度以0.01mg·L^-1为最佳.     

cAMP levels in proliferating rat mammary epithelial cells in vitro and in vivo

Agents that elevate intracellular cAMP levels are required for growth of many cell types in culture including normal rat mammary epithelial (RME) cells. To determine if the intracellular levels of cAMP that result from stimulation by agents such as cholera toxin (CT) or prostaglandin E-1 (PGE-1) are within the physiological range, cAMP levels were determined in RME cells growing in primary culture and compared to levels measured in freshly isolated mammary epithelium. The results indicate that the cAMP levels of mammary epithelial organoids obtained from 45-day-old virgin rats are 4 to 6 pmol/10⁶ cells. Growth of RME cells in primary culture in the presence of CT results in cAMP levels of approximately 15 to 20 pmol/10⁶ cells early in culture when cells are proliferating rapidly. As cells approach confluence, cAMP concentrations decrease to levels observed in fresh organoids. CT-stimulated cAMP levels appear to be within the range of those found in pregnant mammary epithelium in vivo. Growth of RME cells in medium supplemented with PGE-1 instead of CT results in cAMP levels equivalent to those found in fresh mammary epithelial organoids and under these conditions the growth rate is approximately half that found in CT-stimulated cells. These results indicate cAMP to be a positive regulator of cell growth in vivo at levels that are within the physiological range.