人肺癌细胞转录因子--C/EBP和HMG样蛋白的EMSA检测

目的：检测人肺癌细胞中是否存在与Na^ /H^ 交换蛋白-1（NHE-1）基因转录相关的重要转录因子--C/EBP和HMG样蛋白。方法：采用电泳迁移率改变分析法（EMSA）检测人肺癌细胞株核蛋白中C/EBP和HMG样蛋白的表达及其水平。结果：EMSA检测到A549细胞和转染NHE-1反义载体的A549细胞均有转录因子C/EBP和HMG样蛋白的表达，且后者表达水平高于前者；采用50倍未标记的片段能完全抑制相应转录因子与^32P标记的相同片段的结合。结论：A549细胞核蛋白中存在能与C/EBP结合序列和Poly(dA：dT)区结合的转录因子，即C/EBP和HMG样蛋白，后者的表达水平高于前者。特异性竞争抑制试验表明外源性特异DNA片段能抑制相同片段与转录因子的结合。     

精子染色质结构分析的临床意义

目的探讨女方不明原因流产和不明原因不孕与其配偶精子染色质性状之间的关系;方法通过精子染色质结构分析(SCSA)对22例不明原因流产和36例不明原因不孕妇女配偶精子,以及20例正常对照者精子进行DNA完整性检测,并比较它们之间的差异;结果对照组与不明原因流产组以及不孕组之间精子DNA完整性存在差异,且差异具有统计学意义(P<0．05),而不明原因流产组与不孕组之间精子DNA完整性比较无明显差异(P>0．05);结论精子染色质结构分析有可能作为不明原因流产和不孕的辅助诊断方法在临床应用。     

抗人甲状腺球蛋白单克隆抗体的制备及其对抗原决定簇特异性的研究

用杂交瘤技术建立三株分泌抗甲状腺球蛋白(TG)单克隆抗体的杂交瘤细胞株。所获三种单克隆抗体经兔抗小鼠IgG亚类、IgM及IgA血清鉴定表明,两种为IgG_1,一种为IgA;三种含单克隆抗体小鼠腹水的滴度(ELISA法)均为51200以上;以竞争性固相抗体结合试验测定它们对抗原决定簇的特异性差异,结果发现三种单克隆抗体的抗原结合部位是不同的,但对抗原的结合有一定的相互竞争抑制现象。     

Oral lichenoid lesions, mercury hypersensitivity and combined hypersensitivity to mercury and other metals: histologically-proven reproduction of the reaction by patch testing with metal salts

We report 11 patients seen between 1991 and 1994 with oral lichenoid lesion(OLL). In 10 cases, there was contact with dental amalgam fillings, and in patient no. 10 with both amalgam restorations and a gold crown. The last patient had, in addition to her OLL, lichen planus of the skin and genital mucosa. In 5 cases, combined sensitization to mercury and other metal salts, particularly gold sodium thiosulfate (GST) and palladium chloride (PDC), was observed. In 10 patients, the lesions considerably improved or totally cleared within 1 to 9 months of replacement of restoration materials. Histological examination of biopsies from the test sites of amalgam, mercuric chloride, GST and PDC, taken 10 or 17 days, after application of patch tests, showed lichenoid changes in 7 patients with at least 1 of the allergens. At least 2 patients had inflammatory lesions of the oral mucosa related to both amalgam and gold restorations, combined sensitization to inorganic and organic mercury derivatives. GST and, in 1 case, PDC, a "dental restoration metal intolerance syndrome" is proposed.     

Synthesis of the antigen bovine serum albumin‐ergosterol and its immunocharacterization

Fungal contamination of agricultural commodities leads to their spoilage and renders them unfit for human consumption. Ergosterol, a predominant sterol in most fungi and a major constituent of the cell membrane, has been established as a reliable biochemical marker for fungal growth. Various chemical and physico‐chemical methods to quantify ergosterol as an index of fungal contamination are in practice. Yet, immunoassays are the methods of choice in food analysis due to their increased specificity, sensitivity and rapidity. This paper reports the synthesis of an antigen, bovine serum albumin‐ergosterol conjugate, and its immunocharacterization. Ergosterol was converted to ergosterol hemisuccinate (EHS) by succinylation. Subsequently, EHS was conjugated to bovine serum albumin by the mixed anhydride reaction. The molar ratio was found to be 1:28. Antisera raised against the synthesized antigen in rabbits was characterized by the Ouchterlony double‐diffusion technique and an antibody capture assay. Ouchterlony analysis showed a titre of 1:2. Further, characterization by an antibody capture assay, using 50 ng well (10 ng equivalent of ergosterol) of the antigen, gave a titre of 1:4000 dilution of antiserum, with an absorbance of 1.0 at 405 nm. The synthesized antigen may find an application in the development of an immunoanalytical method for ergosterol quantification as a measure of food quality in relation to fungal contamination.     

盐酸三环哌酯在小鼠体内的药代动力学

目的研究抗胆碱新药盐酸三环哌酯（ＴＣＰＮ）在小鼠体内的药代动力学及组织分布。方法药代动力学采用放射受体分析法研究，组织中的分布用放射同位素分析法研究。结果小鼠ｓｃＴＣＰＮ后，血液中药物浓度的经时过程符合一级吸收二室模型（Ｔ１／２β为２．２８ｈ，Ｔｍａｘ为０．１２５ｈ，Ｃｍａｘ为３．４４μｇ·Ｌ－１，Ｃｌ为２３．０Ｌ·ｋｇ－１·ｈ－１）。小鼠ｓｃ［３Ｈ］－ＴＣＰＮ后，分布高峰放射性依次为肾＞肝＞脑＞颌下腺＞肠１９９８－０４－０３收稿，１９９８－０７－２６修回＊国家自然科学基金资助课题，Ｎｏ３９１３００９０－２作者简介：葛召恒，男，３２岁，硕士，助理研究员；阮金秀，男，６２岁，研究员，博士生导师，中国毒理学会副理事长，中国药理学会药物代谢专业委员会主任委员＞心＞肌肉，唯有脑组织的放射性以较高的水平维持较长的时间。预先给小鼠ｓｃ不同剂量的ＱＮＢ，可不同程度地降低脑中放射性的分布。结论ＴＣＰＮ从血中的消除较快，但在脑组织分布时间长，这与药物与脑中Ｍ受体的特异性结合有关。     

Immunology: Diversity of the blocking effects of antisperm antibodies on fertilization in human and mouse

The blocking effects of complement-dependent sperm immobilizingantibodies in the sera of infertile women and monoclonal antispermantibodies against humans and mice on fertilization were investigated.The hemizona assay (HZA) and sperm penetration assay (SPA) wereused to study the inhibitory effects of sera from 22 infertilepatients positive for sperm immobilizing antibodies. Use ofthese tests allowed us to differentiate whether the antibodyblocked sperm—zona pellucida tight binding and/or spermpenetration into the ooplasm. The zona pellucida penetrationassay (ZPA) was also used to study the effects of four monoclonalantibodies (mAbs) on human sperm penetration into the zona pellucida.Seven mAbs against murine spermatozoa were tested for theirinhibitory effects on in-vitro fertilization (IVF) and HZA inmice. Of 22 patient sera with sperm immobilizing antibodies,21 (95.5%) inhibited HZA attachment and penetration, whereasthis did not occur in any of 13 patient sera without these antibodies.However, 19 of 22 (86.4%) patient sera with sperm immobilizingantibodies and eight of 13 (61.5%) patient sera without theseantibodies inhibited the SPA. Two (2C6, 1G12) of four mAbs againsthuman spermatozoa showed strong inhibitory effects in all theassays (HZA, ZPA and SPA). One mAb (3B10) did not inhibit HZAbut blocked ZPA and SPA. Another mAb (H6-3C4) seemed to haveno inhibitory effects on fertilization. Two (Vx 5 and Vx 8)of seven mAbs against murine spermatozoa inhibited IVF in micebut did not block mouse HZA. These findings suggest that antispermantibodies block fertilization at specific stages. Some of themmay inhibit sperm capacitation and thus prevent all processesof fertilization that follow. Some other antibodies may notaffect capacitation and sperm binding to zona pellucida butinhibit the acrosome reaction, followed by the blocking of spermpenetration through zona pellucida and ooplasm.     

免疫层析法用于HCG快速检测的研究

目的：研制用于尿液HCG快速检验试纸条。方法：采用胶体金免疫层析分析方法。结果：检测灵敏度可达50IU／L，与高含量的LH、FSH、TSH均无交叉反应，检测214例临床样品，准确率为100％。结论：该方法简单、快速、特异性强，是检测尿液HCG较好的免疫学方法。     

Development of Activity Assays for High-Volume Evaluation of Human Immunodeficiency Virus (HIV) Protease Inhibitors in Rat Serum: Results with Ditekiren

We showed previously that a commercially available synthetic tetradecapeptide, Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu-Leu-Val-Tyr-Ser, produces authentic angiotensin I (Ang I) upon incubation with the HIV-1 protease (S. K. Sharma et al., Anal. Biochem. 198:363, 1991). Therefore, we developed an Ang-I based activity assay for HIV protease inhibitors based on the technology developed earlier (M. J. Ruwart et al., Pharm. Res. 7:407, 1990; S. K. Sharma et al., Anal. Biochem. 186:24, 1990) for tracking renin inhibitors in rat sera. Ditekiren was either extracted from sera with ethyl acetate or assayed after the interfering substances in sera were precipitated with acetonitrile. Purified recombinant HIV-1 protease was added to extracted rat serum and the enzymatic reaction was initiated in the presence of the tetradecapeptide substrate. The inhibition of Ang I production was measured by a commercially available RIA kit. The cleanup methodology also enabled a commercially available Proteinase Scintillation Proximity Assay (SPA, Amersham) to quantify ditekiren in rat serum through the addition of recombinant HIV-1 protease and cleavage of substrate from SPA beads. Results were confirmed by HPLC or by the renin assay for ditekiren, which inhibits both aspartyl proteases. These technologies should prove useful for assessing serum levels of HIV protease inhibitors in rat.     

Relationship of actin filament assembly to clearance of fibrinogen gold, GPIIb-IIIa complexes on spread platelets

Abstract. The present study has evaluated the influence of high concentrations of cytochalasins B and E on the detergent-resistant actin levels in fully spread platelets by PAGE gel electrophoresis, and the effects of the two inhibitors of new actin filament assembly on translocation of fibrinogen gold (Fgn/Au) labelled GPIIb-IIIa receptors on the surface-activated cells. Concentrations of 10^{- 4}m and 10^-5 m cytochalasin B and E reduced detergent-resistant actin in fully spread platelets to levels present in resting discoid platelets in suspension. Despite reduction of actin filaments to levels in resting cells, cytochalasin B did not prevent translocation of Fgn/Au from platelet margins into channels of the open canalicular system (OCS). Similar concentrations of cytochalasin E completely blocked translocation of receptor-ligand complexes and produced a patching phenomenon not observed in previous studies. Rinsing of the spread cells to remove cytochalasin, followed by incubation of the treated platelets in Hank's buffered salt solution (HBSS) restored levels of detergent-resistant actin to those found in untreated, spread platelets. Resting grids of 10 ⁵ m cytochalasin E-treated platelets on drops of HBSS for 15min restored their ability to clear FGN/Au linked to GPIIb-IIIa from exposed surfaces to the OCS, but 10^-4 m cytochalasin E-treated cells remained anergic after incubation on drops of HBSS. Thus a fully assembled cytoplasmic actin filament cytoskeleton does not appear to be essential for translocating receptor-ligand complexes on the platelet surface to the OCS, nor does its presence guarantee that the ability to clear GPIIb-IIIa receptors will be restored.