Differences in LH receptor down-regulation between rat and mouse Leydig cells: effects of 3',5'-cyclic AMP and phorbol esters.

The role of cyclic AMP and phorbol esters in luteinizing hormone (LH) receptor down-regulation in Leydig cells has been studied. Dibutyryl cyclic AMP (db-cAMP) (0.01, 0.1 and 1 mM), forskolin (80 microM) and cholera toxin (1.19 nM) caused a 30-50% loss of [125I]hCG binding sites and an inhibition of receptor-[125I]hCG complex internalization in mouse tumour Leydig (MA10, MLTC-1) cells during 2 h. In contrast, db-cAMP had no effect on the level of binding sites or internalization of the hormone receptor complex in rat testis Leydig cells or a rat tumour (R2C) Leydig cell. Phorbol 12-myristate 13-acetate (PMA) at concentrations from 10(-9) to 10(-5) M had no effect on hormone binding or hormone-receptor complex internalization in any of the Leydig cells. In contrast a 2 h preincubation of MLTC-1 cells with 10(-7) M PMA caused a loss of subsequent LH-stimulated cyclic AMP and pregnenolone production. These results indicate that LH receptor down-regulation is mediated by cyclic AMP dependent kinase, but not protein kinase C, in mouse Leydig cells. No down-regulation of rat Leydig cell LH receptor occurs with either kinase.     

Increasing prevalence of penicillinase‐producing Neisseria gonorrhoeae and the emergence of high‐level, plasmid‐mediated tetracycline resistance among gonococcal isolates in The Gambia

Summary One hundred and three strains of Neisseria gonorrhoeae isolated from a periurban STD clinic in The Gambia were studied for antimicrobial susceptibility, plasmid profile, and serogroup using standard procedures. Seventy-nine (77%) were penicillinase producers (PPNG) and fully resistant to penicillin (MIC ≥8 mg/l). One isolate showed chromosomally induced resistance to penicillin (MIC 2 mg/l). None of the isolates was sensitive to tetracycline; 16 (16%) showed intermediate resistance (MICs 1–8 mg/l) and 87 (84%) showed high-level plasmid-mediated resistance (TRNG) (MICs >10 mg/l). This is the first report of TRNG in The Gambia. Only 6 (6%) strains were fully sensitive to trimethoprim-sulphamethoxazole (MIC <8 mg/l); 78 (76%) showed intermediate level resistance (MICs 8–16 mg/l) and 19 (18%) were fully resistant (MIC >32 mg/l). Indications of an increase in MIC to ciprofloxacin and ceftriaxone were found in 6 (6%) and 1 (1%) strains, respectively, although all remained fully sensitive (MICs 0.004–0.03 mg/1 and 0.001–0.015 mg/l). All PPNG and TRNG strains carried the 3.2 MDa and 25.2 MDa plasmids, respectively. All isolates carried the 2.6 MDa cryptic plasmid and 9 (3 PPNG and 6 non-PPNG) carried the 24.5 MDa conjugative plasmid. Forty-four (43%) strains were typed group W1, 58 (56%) W11/111 and 1 had cross-reacting antigens. Because PPNG are frequently encountered and high-level TRNG is now prevalent, the newer cephalosporins and quinolones must now be considered as first-line drugs for the treatment of gonorrhoea in The Gambia.     

Transport and epithelial secretion of the cardiac glycoside, digoxin, by human intestinal epithelial (Caco-2) cells.

1. Human intestinal epithelial Caco-2 cells have been used to investigate the transepithelial permeation of the cardiac glycoside, digoxin. 2. Transepithelial basal to apical [3H]-digoxin flux exceeds apical to basal flux, a net secretion of [3H]-digoxin being observed. At 200 microM digoxin, net secretory flux (Jnet) was 10.8 +/- 0.6 nmol cm-2 h-1. Maximal secretory flux (Jmax) of vinblastine was 1.3 +/- 0.1 nmol cm-2 h-1. Cellular uptake of digoxin was different across apical and basal cell boundaries. It was greatest across the basal surface at 1 microM, whereas at 200 microM, apical uptake exceeded basal uptake. 3. Net secretion of [3H]-digoxin was subject to inhibition by digitoxin and bufalin but was not inhibited by ouabain, convallatoxin, and strophanthidin (all 100 microM). Inhibition was due to both a decrease in Jb-a and an increase in Ja-b. Uptake of [3H]-digoxin at the apical surface was increased by digitoxin and bufalin. All cardiac glycosides decreased [3H]-digoxin uptake at the basal cell surface (except for 100 microM digitoxin). 4. The competitive P-glycoprotein inhibitors, verapamil (100 microM), nifedipine (50 microM) and vinblastine (50 microM) all abolished net secretion of [3H]-digoxin due to both a decrease in Jb-a and an increase in Ja-b. Cellular accumulation of [3H]-digoxin was also increased across both the apical and basal cell surfaces. I-Chloro-2,4,-dinitrobenzene (10 microM), a substrate for glutathione-S-transferase and subsequent ATP-dependent glutathione-S-conjugate secretion, failed to inhibit net secretion of [3H]-digoxin. The increase in absorptive permeability Pa-b (= Ja-b/Ca) and cellular [3H]-digoxin uptake upon P-glycoprotein inhibition, showed that the intestinal epithelium was rendered effectively impermeable by ATP-dependent extrusion at the apical surface. 5. A model for [3H]-digoxin secretion by the intestinal epithelium is likely to involve both diffusional uptake and Na(+)-K+ pump-mediated endocytosis, followed by active extrusion at the apical membrane.     

Efficacy of SIV/deltaB670 glycoprotein-enriched and glycoprotein-depleted subunit vaccines in protecting against infection and disease in rhesus monkeys

Immunization with an inactivated whole-virus vaccine is highly effective in preventing lentivirus infection. The viral protein(s) essential to the induction of protective responses, however, have not been identified. To define the role of virion components in the induction of protective immunity, we evaluated the efficacy of glycoprotein-enriched and glycoprotein-depleted simian immunodeficiency virus (SIV) subunit vaccines prepared by lentil-lectin affinity chromatography of gradient-purified virions using the immunization and challenge regimen previously found successful with an inactivated whole-virus vaccine. Infection was determined by successful recovery of virus, the induction of SIV-specific antibody responses, and infection of naive recipients by inoculation with lymph-node-derived lymphocytes from the vaccinates. Immunization with the glycoprotein-enriched preparation prevented infection in two out of four monkeys, whereas the glycoprotein-depleted vaccine failed to prevent infection in all four vaccinates tested. However, the glycoprotein-depleted vaccine appeared to moderate the progression of SIV-induced disease compared with non-immunized infected control monkeys inoculated with the same challenge dose. These data suggest that subunit vaccines containing sufficient quantities of viral glycoproteins can protect against SIV infection, whereas subunit vaccines composed predominantly of viral core proteins cannot. The development of effective vaccines against HIV infection should include studies on the optimum presentation of the viral envelope glycoproteins to produce long-term broadly protective immune responses.     

赖型钩体flaB2与VR1012中的CpG基序分析

目的：对问号赖型钩端螺旋体（赖型钩体）DNA疫苗[包括内鞭毛蛋白基因（flaB2)和质粒DNA表达载体（VR1012）]的CpG基序（CpG motifs)进行分析，为DNA疫苗免疫机制的阐明和提高DNA疫苗的效能奠定基础。方法：以flaB2与VR1012构建重组DNA的免疫原，对flaB2及VR1012全核苷酸序列进行计算机分析（分类、计数和定位）。结果：CpG的“C”的侧翼为两个嘌呤，“G”的侧翼为两个嘧啶，在flaB2中共3个，分别为GACGCT，GACGTC和GACGCC；在VR1012中共11个，分别为GACGTC1个，GACGCT2个，GACGCC1个，GACGTT1个，GGCGTT2个，GGCGCT2个，GGCGCC1个，AACGCT1个，其中特别重要的TGACGTCA4个和TAACGCCA有1个，位于5'端456－463；509－516；592－599；778－785和486－493；4个TGACGTCA和1个TAACGCCA均位于5'端且相对集中。结论：赖型钩体flaB2与VR1012构成的DNA疫苗含有TGACGTCA等CpG，这些基序又称免疫刺激序列，构成了DNA疫苗中的佐剂。     

二氯胺基酚诱发的染色体畸变及染毒时间对畸变率的影响研究

２，４－二氯胺基酚（ＤＣＡＰ）是８３—１除草剂在哺乳动物体内的主要代谢产物。本研究以三种染毒计划观察了ＤＣＡＰ诱发Ｖ７９细胞的染色体畸变。结果表明：ＤＣＡＰ是一种染色体损伤剂，诱发的畸变主要为染色单体断裂和交换；３ｈ染毒和染毒后培养１７ｈ诱发的染色体畸变率最高，２０ｈ染毒观察不到染色体畸变，说明以高浓度短期染毒对高细胞毒性化合物的细胞遗传毒性研究可能是较好的染毒方案。     

Ag—NOR计数在区分良恶性甲状腺病变中的意义

随机选择经长期随访证实的滤泡性甲状腺癌和腺瘤各10例,正常甲状腺组织5例作为对照。按Ploton的染色方法和Crocker推荐的计数方法分别计算三组每例各50个细胞的Ag-NOR嗜银颗粒平均数,再算出三组各自的AgNOR均数,经统计学检验三者有极显著性差异。可望成为甲状腺滤泡性肿瘤良恶鉴别的一项辅助指标。     

A cell line from Wilms' tumour with deletion in short arm of chromosome II

A cell line (T3/73) from a Wilms' tumour has been established from a 9 month-old boy with aniridia. The tumour was removed in 1973. On histological examination a diagnosis of Wilms' tumour was made which showed undifferentiated areas, marked tubule formation and abundant striped muscle fibres. The tumour cells, which are fusiform, grew rapidly in culture without the addition of growth factors, and have undergone over 100 passages. Approximately 95% and 5% were positive for desmin and cytokeratin, respectively. The cell doubling time was 28 hr. Cytogenetic studies revealed a karyotype of 46,XY,del(11) (p12::p14). Although the cells stained very intensely with a monoclonal antibody that detects oncogene ras p 21 antigen, Southern blot analysis using c-Ha-ras as a probe failed to reveal an obvious deletion or amplification of either Ha-ras allele.