Antibody catalysis of peptidyl-prolyl cis-trans isomerization in the folding of RNase T1

An antibody generated to an alpha-keto amide containing hapten 1 catalyzes the cis-trans isomerization of peptidyl-prolyl amide bonds in peptides and in the protein RNase T1. The antibody-catalyzed peptide isomerization reaction showed saturation kinetics for the cis-substrate, Suc-Ala-Ala-Pro-Phe-pNA, with a kcat/Km value of 883 s-1.M-1; the reaction was inhibited by the hapten analog 13 (Ki = 3. 0 +/- 0.4 microM). Refolding of denatured RNase T1 to its native conformation also was catalyzed by the antibody, with the antibody-catalyzed folding reaction inhibitable both by the hapten 1 and hapten analog 13. These results demonstrate that antibodies can catalyze conformational changes in protein structure, a transformation involved in many cellular processes.     

Chromosome-specific aneusomy in carcinoma of the breast

Fluorescence in situ hybridization was performed on touch preparations from 55 primary infiltrating ductal carcinomas of the breast to determine numeric chromosome abnormalities. The frequency of aneusomy, measured by both nondisomy and chromosomal gain, was determined for chromosomes X, 4, 6-12, 17, and 18 with the use of chromosome-specific, alpha-satellite DNA probes. The presence of chromosome-specific numeric abnormalities was correlated with established clinicopathological parameters, including tumor size, lymph node involvement, tumor grade, estrogen receptor level, and menopause status. In addition, a case-control study was performed to explore a possible association between chromosome-specific aneusomy and recurrence in lymph-node-negative patients. Although chromosomes 8 and 6 were most frequently aneusomic, numeric abnormalities of chromosomes 4 and 11 were most strongly associated with established prognostic factors. For chromosomes 4 and 11, strong associations were found with tumor involvement of lymph nodes and increased tumor size, along with a weaker association with tumor grade. In addition, numeric abnormalities of the following chromosomes were associated with the corresponding prognostic factors: chromosomes X, 7, and 12 with lymph node status; chromosomes 10, 17, and 6 with tumor size; and chromosomes 7, 12, 17, and X with tumor grade. No correlations were observed with estrogen receptor level or menopause status. In the case-control study performed on isolated nuclei of paraffin-embedded tissue from lymph node-negative breast cancer patients (19 cases and 19 controls), the gain of chromosome 4 was correlated with disease progression. These findings suggest that chromosome-specific aneusomy is associated with certain established prognostic factors and may be associated with disease progression.     

Anatomic correction of transposition of the great vessels

We present a new approach for anatomic correction of transposition of the great arteries. The two coronary arteries, with a piece of the aortic wall attached, are transposed to the posterior artery. The two aortic openings are closed with a patch. The aorta and pulmonary artery are transected, contraposed, ant then anastomosed. The interventricular septal defect is closed with a patch, through a right ventriculotomy approach, because the right ventricle is no longer part of the systemic circulation. Two patients, aged 3 months and 40 days weighing 4,200 and 3,700 grams, respectively, were operated upon with deep hypothermia and total circulatory arrest. There was good recovery from the operation, with normal cardiocirculatory conditions. Renal failure developed in the first patient, and she died on the third postoperative day. During this time the cardiocirculatory conditions were good. The second patient made an uneventful recovery. Hemodynamic studies 20 days after the operation showed complete correction of the malformation. Five and one-half months after the operation, he weighs 7,500 grams, and his development is very good. We believe that this operation will be reproducible by most cardiovascular septal defect and pulmonary hypertension.     

Protection against adoptive transfer of autoimmune diabetes mediated through very late antigen-4 integrin

The very late antigen-4 (VLA-4) integrin expressed on the surface of lymphocytes and macrophages can regulate their migration to inflammatory sites as well as control cellular activation. The role of VLA-4 in the establishment of autoimmune diabetes is not easily predicted given the multiplicity of adhesion pathways and their differential use by various cell types. The contribution of VLA-4 to insulin-dependent diabetes mellitus was investigated by administration of VLA-4-specific monoclonal antibodies (MoAb) in an adoptive transfer model of disease in NOD mice. This study shows that VLA-4-specific MoAbs profoundly inhibit the development of diabetes with protection sustained by repeated MoAb exposure. Insulitis was completely inhibited during treatment and progressed to a severe degree once MoAb treatment was suspended, yet approximately 40% of treated recipients failed to become diabetic during 1-2 months post-treatment. Although we cannot rule out depletion of a relatively minor subpopulation of cells by prolonged anti-VLA-4 MoAb exposure, this inhibition of diabetes onset by treatment with MoAbs to VLA-4 supports a dependence on VLA-4 for cellular functions leading to diabetes and demonstrates that a significant disease modifying effect can be mediated by targeting the VLA-4 integrin.     

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.     

Effects of cold and warm blood cardioplegia assessed by myocardial pH and release of metabolic markers

The optimal temperature of blood cardioplegia remains controversial. Interstitial myocardial pH was monitored online with a probe that was inserted in the anterior wall of the left ventricle. Venous pH, lactate production, and creatine kinase and troponin T release were measured in coronary sinus blood obtained in 14 dogs after ischemic arrest periods of 5, 10, 20, and 40 minutes with warm (n = 7; mean myocardial temperature, 35 degrees +/- 2 degrees C) and cold (n = 7; mean myocardial temperature, 12 degrees +/- 1 degree C) blood cardioplegic protection. Blood cardioplegic solution was delivered at a rate of 100 mL/min during the 10 minutes between each ischemic arrest. The interstitial myocardial pH decreased significantly (p < 0.05) from 7.1 +/- 0.3 to 6.53 +/- 0.3 after ischemia in animals perfused with warm blood cardioplegia and from 7.04 +/- 0.3 to 6.64 +/- 0.1 in those receiving cold blood cardioplegic protection; however, the difference between the groups was not significant (p > 0.05). Lactate production and creatine kinase and troponin T release increased significantly after ischemia, but there was no difference in the changes between the warm and cold blood cardioplegia groups. In conclusion, ischemia caused significant changes in all variables measured, and these changes were directly proportional to the duration of ischemia. However, there was no significant difference (p > 0.05) in the myocardial metabolic changes between the warm and cold blood cardioplegia groups in terms of the duration of ischemic arrest studied.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role strain in couples with and without a child with a chronic illness: associations with marital satisfaction, intimacy, and daily mood

The tryptophan depletion test is a research strategy to investigate the functional consequences of decreasing the brain serotonin metabolism. Because serotonin is involved in sleep regulation and the regulation of affective states, we studied the acute polysomnographic effects of tryptophan depletion and expected to induce similar changes of sleep EEG as observed in depressed patients. A total of 12 healthy subjects (mean age 34 +/- 3 years) had eight polysomnograms, divided in two blocks of 4 consecutive nights. After one adaptation and 1 baseline night, subjects received a low-protein diet on day 3 and 4 until midday. On day 4 at 18.00 h, they drank an amino acid mixture either devoid of tryptophan or containing 2.3 g of tryptophan (placebo control) in randomized and double-blind order, resulting in an 85% decrease (tryptophan depletion) and a 144% increase (placebo control) of serum tryptophan at 22.00 h. After tryptophan depletion but not placebo, significant effects on sleep EEG were observed in terms of decreased non-rapid eye movement (non-REM) stage 2, increase of wake %, and of rapid eye movement (REM) density compared with baseline. REM latency was not altered, however the first and second REM period interval were significantly shorter after tryptophan depletion. This study underlines the impact of the serotonergic system on sleep maintenance and on REM sleep.