Developmental consequences of embryo and cell manipulation in mice and farm animals

Advances in biotechnology in recent decades have revolutionized our understanding of early mammalian development and promise to provide ever more finely tuned and precisely targeted techniques for genetic enhancement of domestic animal species. In demonstrating what is both technically and biologically possible, not only in mice but also in larger animal species, research has provided hope that previously intractable diseases and genetic defects can be successfully combated. Crucial to this research is the ability to culture oocytes, embryos and somatic cells in vitro and to sustain their development without inducing adverse short- or long-term consequences. There is a need to refine current culture strategies in farm animal species to avoid jeopardizing their dependent technologies. A key to resolving current limitations of culture strategies is to identify, acknowledge and then address those features of in vitro culture that compromise early regulation of mammalian development. The aim of this review is to appraise critically in vitro embryo and somatic cell production strategies in the context of their impact on developmental competence and normality at embryonic, fetal and later stages. In addition, effects of physically manipulating embryos and cells, most notably via nuclear and gene transfer technologies, are considered with a view to identifying how detrimental consequences can be avoided.     

Widespread microsatellite instability in sebaceous tumours of patients with the Muir-Torre syndrome

OBJECTIVE: Thoracic injury remains a major source of morbidity and mortality in urban trauma centers. With the advent and increasing expertise in video assisted thoracic surgery, this modality has become an attractive alternative in the management of patients with thoracic injury. This report will review our experience with video assisted thoracic surgery at a level I trauma center and attempt to further delineate the indications for and timing of thoracoscopy in thoracic trauma. METHODS: We identified 16 patients who had undergone video assisted thoracic surgery following chest trauma between July 1991 and June 1994. There were 15 penetrating and one blunt trauma. All 16 patients were initially treated with tube thoracostomy. From 0-20 days post-injury, video assisted thoracic surgery was attempted with either diagnostic or therapeutic intentions. RESULTS: Twelve of the 16 patients (75%) had successful thoracoscopy. Three patients had diaphragmatic injury excluded and nine patients had successful evacuation of clotted hemothoraces. Evacuation of clotted hemothorax up to 7 days post-injury was safe and easily accomplished. Four patients (25%) had unsuccessful thoracoscopy and were converted to standard thoracotomy; failure was attributed to either suboptimal single lung ventilation or severe pleural inflammatory reaction. The only death in the entire group occurred 10 days after a thoracotomy for retained hemothorax. The median post-operative hospital stay following successful video assisted thoracic surgery was 3.5 days. CONCLUSIONS: Video assisted thoracic surgery can be utilized as an effective and safe method for the initial diagnostic evaluation and surgical management of stable patients with penetrating thoracic trauma.     

The CTFA Evaluation of Alternatives Program: an evaluation of in vitro alternatives to the Draize primary eye irritation test. (Phase III) surfactant-based formulations

The CTFA Evaluation of Alternatives Program is an evaluation of the relationship between data from the Draize primary eye irritation test and comparable data from a selection of promising in vitro eye irritation tests. In Phase III, data from the Draize test and 41 in vitro endpoints on 25 representative surfactant-based personal care formulations were compared. As in Phase I and Phase II, regression modelling of the relationship between maximum average Draize score (MAS) and in vitro endpoint was the primary approach adopted for evaluating in vitro assay performance. The degree of confidence in prediction of MAS for a given in vitro endpoint is quantified in terms of the relative widths of prediction intervals constructed about the fitted regression curve. Prediction intervals reflect not only the error attributed to the model but also the material-specific components of variation in both the Draize and the in vitro assays. Among the in vitro assays selected for regression modeling in Phase III, the relationship between MAS and in vitro score was relatively well defined. The prediction bounds on MAS were most narrow for materials at the lower or upper end of the effective irritation range (MAS = 0-45), where variability in MAS was smallest. This, the confidence with which the MAS of surfactant-based formulations is predicted is greatest when MAS approaches zero or when MAS approaches 45 (no comment is made on prediction of MAS > 45 since extrapolation beyond the range of observed data is not possible). No single in vitro endpoint was found to exhibit relative superiority with regard to prediction of MAS. Variability associated with Draize test outcome (e.g. in MAS values) must be considered in any future comparisons of in vivo and in vitro test results if the purpose is to predict in vivo response using in vitro data.     

Evaluation of the completion of influenza vaccination

OBJECTIVE: To find the reasons which determine failures to comply with anti-flu vaccinations, so that these can be corrected and the coverage of this preventive action be increased. DESIGN: Observational crossover study, done by means of a telephone survey of people over 65. A questionnaire with closed questions, composed after a pilot study and validated by Cronbach's alpha. SETTING: Primary Care Centre (PCC). PATIENTS: We calculated a population sample for qualitative variables (_ = 0.05; p = 0.60; e = 0.05) of 294 people over 65, chosen from the PCC records, by means of random sampling (K = 4) stratified for age and discounting the telephone selection bias. MEASUREMENTS AND RESULTS: The proportion of vaccinated patients (60.9%) obtained in our study did not significantly differ from that in the general population. The percentage of patients included in the programme for the first time was 14%. Level of satisfaction among those vaccinated was 89.4%, with 8.9% of problems detected being light. Main causes of non-vaccination were: thinking that they didn't need it (63.5%), ignorance of the campaign (35.7%), fear of the reaction (24.3%), forgetting (10.4%). The main form of access to the campaign information was from the PCC, both through individuals and posters. Lack of information was statistically significant (p < 0.00001) as a determinant of non-vaccination, without other factors (age, sex, associated pathologies...) explaining these differences. CONCLUSIONS: Individualised and on-going health education by the PCC is fundamental. This would enable the identification of the group not vaccinated due to their express refusal and the recovery of non-vaccinated patients.     

Familial adenomatous polyposis

Familial adenomatous polyposis (FAP) is an autosomal dominant disease characterized by the development of numerous adenomatous polyps in the colon and rectum with diverse extracolonic manifestations. Recent genetic advances have lead to the sequencing of the FAP gene, with important implications for screening, diagnosis and follow-up. Appropriate management of probands and at-risk patients is of the utmost importance, as untreated carriers will develop colorectal cancer. Identification of FAP families and tracing of pedigrees represent the most important steps. To this end registries are essential, allowing a comprehensive multidisciplinary approach. They have justified their place by decreasing related morbidity and mortality. An overview and discussion of clinical features and management are presented.     

Recombinant fusion proteins for the industrial production of disulfide bridge containing peptides: purification, oxidation without concatamer formation, and selective cleavage

The relative contributions of protein tyrosine kinases (PTKs) and protein kinase C isoenzymes (PKCs), a family of serine/threonine kinases, in integrin alpha(IIb)beta3 (glycoprotein IIb/IIIa) exposure are the subject of much controversy. In the present study we measured the effect of the PTK inhibitor herbimycin A and the PKC inhibitor bisindolylmaleimide I on 125I-fibrinogen binding to alpha(IIb)beta3 and on aggregation/secretion induced by different agonists. Dose-response studies showed complete inhibition of alpha(IIb)beta3 exposure by 30 micromol/L (ADP stimulation) and 35 to 40 micromol/L (alpha-thrombin stimulation) herbimycin A. In contrast, inhibition of exposure by bisindolylmaleimide I varied from none (for ADP and epinephrine), to 30% (for platelet-activating factor), and to approximately 80% (for alpha-thrombin). Studies with a submaximal dose of herbimycin A (approximately 50% inhibition of the ADP-response) and a maximal dose of bisindolylmaleimide I showed that optical aggregation had a similar sensitivity to the inhibitors as alpha(IIb)beta3 exposure with minimal interference by secreted ADP. Thus, the relative contributions of tyrosine and serine/threonine kinases in alpha(IIb)beta3 exposure and aggregation differ among the different agonists, with an exclusive role for PTKs in ADP- and epinephrine-induced responses and a role for both PTKs and PKCs in responses induced by platelet-activating factor and alpha-thrombin.     

Genetics of grass dry matter intake, energy balance, and digestibility in grazing irish dairy cows

The objective of this study was to estimate genetic parameters for grass dry matter intake (DMI), energy balance (EB), and cow internal digestibility (IDG) in grazing Holstein-Friesian dairy cows. Grass DMI was estimated up to 4 times per lactation on 1,588 lactations from 755 cows on 2 research farms in southern Ireland. Simultaneously measured milk production and BW records were used to calculate EB. Cow IDG, measured as the ratio of feed and fecal concentrations of the natural odd carbon-chain n-alkane pentatriacontane, was available on 583 lactations from 238 cows. Random regression and multitrait animal models were used to estimate residual, additive genetic and permanent environmental (co)variances across lactations. Results were similar for both models. Heritability for DMI, EB, and IDG across lactation varied from 0.10 [8 days in milk (DIM)] to 0.30 (169 DIM), from 0.06 (29 DIM) to 0.29 (305 DIM), and from 0.08 (50 DIM) to 0.45 (305 DIM), respectively, when estimated using the random regression model. Genetic correlations within each trait tended to decrease as the interval between periods compared increased for DMI and EB, whereas the correlations with IDG in early lactation were weakest when measured midlactation. The lowest correlation between any 2 periods was 0.10, −0.36, and −0.04 for DMI, EB, and IDG, respectively, suggesting the effect of different genes at different stages of lactations. Eigenvalues and associated eigenfunctions of the additive genetic covariance matrix revealed considerable genetic variation among animals in the shape of the lactation profiles for DMI, EB, and IDG. Genetic parameters presented are the first estimates from dairy cows fed predominantly grazed grass and imply that genetic improvement in DMI, EB, and IDG in Holstein-Friesian cows fed predominantly grazed grass is possible.