Reduction of the number of mice used for potency testing of human and animal rabies vaccines

Eight different rabies vaccines were tested for their potency in the standard mouse potency test using 3-, 5- and 7-week-old mice. 5-week-old mice seem to be best suited for this purpose, variability from test to test could be reduced considerably. An ELISA was used in parallel for the evaluation of the rabies glycoprotein content of rabies vaccines. Results of the mouse potency test correlated well with those of the ELISA if highly purified human vaccines were tested. Unspecific reactions in the ELISA caused by adjuvanted veterinary vaccines could not be blocked. Further experiments will be needed in order to evaluate the potency of inactivated veterinary rabies vaccines by a in vitro test.     

Studies on the thyroid in transgenic mice expressing the genes for human and bovine growth hormone

Summary The thyroid glands of transgenic mice (TM) expressing the genes for human (h) and bovine (b) growth hormone (GH) were studied. The percentages of larger follicles inhGH TM of either sex were significantly greater than in the corresponding normal littermates, and follicles ranging up to 350 m in diameter were present in malehGH TM. In contrast, thyroid follicles were only slightly enlarged in malebGH TM, and were unchanged in femalebGH TM. The serum concentrations of T4 were significantly decreased in malebGH TM and not altered in the other groups. Serum concentrations of T3 were slightly, but significantly increased in femalehGH TM and femalebGH TM, but were unaffected in male TM of either type. Since the principal difference between these foreign GHs in rodents is the additional lactogenic activity of human GH, these results may indicate that the effects of prolactin can influence the development of the thyroid.     

三维扫描树叶子节点和TSVs数量的优化方法

扫描树结构能够有效地减少集成电路的测试数据量和测试时间,降低电路的测试成本.为减少三维电路中扫描树的叶子节点和硅通孔数量,首先得出了扫描树中叶子节点的最小数量为最大相容组中所含扫描单元数量的结论,然后进一步得到了叶子节点取得最小值的充分必要条件.并在此基础上,提出了一种启发式算法来确定扫描树中相容组的连接顺序,使得叶子节点数量取得最小值的同时能够优化硅通孔的数量.实验结果表明了所提方法的有效性.     

Postimplantation embryo culture for the assessment of the teratogenic potential and potency of compounds

Summary Whole rat embryos cultured during the early stages of organogenesis were subjected to a panel of selected chemicals. Of seventeen known in vivo teratogens, seventeen also induced specific malformations in embryos grown in culture. Of ten chemicals which were reported to be negative in in vivo rat teratogenicity studies, eight also did not provoke dysmorphogenic effects in vitro. Of five additionally tested retinoids, all induced multiple malformations. However, concentrations used to induce these effects varied considerably, isotretinoin inducing malformations at 10^–5M and arotinoid at 10^–11M. The results indicate qualitatively as well as quantitatively a high predictability of this in vitro system and suggest that the postimplantation embryo culture system may also be useful in the prospective testing of new drugs and environmental chemicals.     

The use of microtiter plates for the simple and sensitive determination of insulin by an ELISA method

Summary A method of insulin determination using a commercially available ELISA kit was modified for use in microtiter plates. The adapted assay, based on the binding of procine anti-guinea pig insulin antibodies to microtiter plates and insulin-peroxidase conjugate as displacer, is sensitive between 0.5 and 30 ng/ml. Since it uses only 10–40 l of sample material it enables the determination of 5–100 pg of insulin. The rapid (5–6 h), automatable, reproducible and reliable assay makes it possible to determine many samples in a short time.     

Major contribution of codominant CD8 and CD4 T cell epitopes to the human cytomegalovirus-specific T cell repertoire

Human cytomegalovirus (HCMV) infection or reactivation is a cause of morbidity and mortality in immunocompromised individuals. In immunocompetent individuals, in contrast, HCMV is successfully controlled by specific CD8 and CD4 T cells. Knowledge of CD8 and CD4 T cell epitopes from HCMV and their immunodominant features is crucial for the generation of epitope-specific T cells for adoptive immunotherapy and for the development of a peptide-based HCMV vaccine. Therefore, we investigated the natural frequencies of a large number of CD8 and CD4 T cell epitopes, including 10 novel ones. We determined several epitopes as immunodominant. Surprisingly, no clear hierarchies were found for CD8 T cell epitopes, indicating codominance. These results will be valuable for adoptive transfer strategies and support initiatives towards development of a peptide-based HCMV vaccine.Received 12 August 2004; received after revision 24 September 2004; accepted 29 October 2004 These authors contributed equally to this work.     

Molecular parasitology: Progress towards the development of vaccines for malaria,filariasis, and schistosomiasis

Summary Advances in molecular biology have allowed for the identification of potential vaccine candidates against several parasitic diseases. Antigens from various life stages ofPlasmodium andSchistosoma species and filarial worms have been cloned, sequenced and tested as vaccines. Results to date in animal models have been promising. Modest levels of protection against experimental human malaria have been obtained using both sporozoite and blood-stage antigens. However, a greater understanding of the mechanisms which lead to immunity against parasites is required before effective vaccines can be developed.     

Transgenic and knock-out mice for deciphering the roles of EGFR ligands

Generation of genetically engineered mice with either gain-of-function or loss-of-function mutations is the most popular technique for determining gene functions and the interrelationship between molecules in vivo. These models have provided a wealth of information about the developmental and physiological roles of oncogenes and growth factors. To date, transgenic techniques have been used extensively to study the functions of the epidermal growth factor (EGF) family. This review highlights some of the major recent findings pertinent to the EGF receptor (EGFR) and its ligands with special reference to elucidating how EGF and its related growth factors work together to regulate reproduction, growth and development. Finally, future investigations on ligand-ligand communications, EGFR and its ligands in neural stem cell research, and the mechanisms of EGFR signaling and trafficking in cells are also suggested. Received 24 May 2002; received after revision 15 July 2002; accepted 16 July 2002     

The possible role of glutamine in some cells of the immune system and the possible consequence for the whole animal

Glutamine is important for the function of lymphocytes and macrophages. A role for the high rate of glutamine utilisation by these cells is presented. Since muscle syntheses, stores and releases glutamine, this tissue may play a role in the immune response. Since the number of immune cells utilising glutamine may be large, the demand for glutamine from muscle, especially during trauma sepsis or burns, may be very high. A speculative suggestion is put forward that this requirement for glutamine from muscle may play a role in cachexia under some of these conditions.     

Cellular models for the detection and evaluation of drugs that modulate human phagocyte activity

Summary Simple testing models have been developed for the evaluation of chemical or biological compounds that modulate the activity of human phagocytes. Human neutrophils from buffy coats of donor blood are used. They are stimulated with receptor agonists, and the effects of test compounds on exocytosis of different enzymes, the generation of superoxide (respiratory burst), and cytotoxicity are quantified. All assays are performed in microtiter plates and the responses are evaluated by multi-well photometry or fluorimetry. The models are apt to detect compounds acting on phagocytes as agonists or antagonists, signal transduction activators or inhibitors and primers of agonist responses, and to assess cytotoxic effects.     

Kinetic arguments for the existence of a single form of intestinal ornithine decarboxylase during the postnatal development of normal and sparse-fur mutant mice and after EGF treatment

Summary The K_m for ornithine is remarkably constant during the course of postnatal development in both normal and spf mutant mice even if a large but transient increase in ornithine decarboxylase (ODC) activity is noted. Four hours after EGF injection (4 g/g b.wt) to 17-day-old normal and spf mice, a marked stimulation of ODC activity is observed but K_m remains unaffected. These data argue against the existence of multiple forms of ODC in the intestinal mucosa of mice.Supported by grant MA-8923 from the Medical Research Council of Canada. The author is Chercheur-boursier du Fonds de la Recherche en Santé du Québec.     

Retroviral-mediated gene therapy for the treatment of citrullinemia. Transfer and expression of argininosuccinate synthetase in human hematopoietic cells

Citrullinemia is a recessive genetic disease caused by a deficiency in argininosuccinate synthetase (AS). Retroviruses were used to transduce the human AS gene into cultured human cells. Using amphotropic viruses with high titer (>10⁶ cfu/ml), we were able to correct the defect in cultured fibroblasts from citrullinemic patients. Retroviral transduction of the human AS gene into human bone marrow cells was also studied. Co-cultivation was used to infect the cells and up to 80% of progenitor cells were found to be carrying and expressing the AS retrovirus after infection. When the infected cells were kept in culture, integration and expression of the retrovirus was observed. Retroviral sequences were present and expressed in the cultured bone marrow-derived cells for up to 10 weeks.