Juvenile sudden death in a family with polymorphic ventricular arrhythmias caused by a novel RyR2 gene mutation: evidence of specific morphological substrates

We report on a family with a history of sudden death and effort-induced polymorphic ventricular arrhythmias. The index case was a 17-year-old boy who died suddenly and at postmortem had evidence of fibrofatty replacement in the right ventricular free wall, consistent with arrhythmogenic right ventricular cardiomyopathy, as well as calcium phosphate deposits within the myocytes. A molecular genetics investigation carried out in the paraffin-embedded myocardium of the subject and in blood samples of family members disclosed a missense mutation in exon 3 (230C-->T; A77V) of the cardiac ryanodine receptor type 2 gene. The carriers showed effort-induced polymorphic ventricular tachycardia in the setting of normal resting electrocardiogram and trivial echocardiographic abnormalities, consistent with catecholaminergic polymorphic ventricular tachycardia. The observation of both arrhythmogenic right ventricular cardiomyopathy type 2 and catecholaminergic polymorphic ventricular tachycardia in the same family suggests that the two entities might correspond to different degrees of phenotypic expression of the same disease. This experience underscores the importance of a precise autopsy diagnosis in the case of sudden cardiac death, including molecular genetics, and the mission of pathologists to guide further clinical investigation of family members.     

Establishment of diagnostic cutoff points for levels of serum antibodies to pertussis toxin, filamentous hemagglutinin, and fimbriae in adolescents and adults in the United States

Numerous reports have documented that serologic methods are much more sensitive than culture for the diagnosis of pertussis in adolescents and adults. However, a standardized serologic test for pertussis is not routinely available to most clinicians, and the serologic test levels or cutoff points correlated with diseases have not been determined. The goal of the present study was to examine the distribution of immunoglobulin G (IgG) levels against three Bordetella pertussis antigens (pertussis toxin [PT], filamentous hemagglutinin [FHA], and fimbria types 2 and 3 [FIM]) and to determine population-based antibody levels for the purpose of establishing such diagnostic cutoff points. Enzyme-linked immunosorbent assays (ELISAs) were performed with sera from >6,000 U.S. residents aged 6 to 49 years who participated in the Third National Health and Nutrition Examination Survey. Mixture models were developed to identify hypothesized exposure groups and establish diagnostic cutoffs. Quantifiable (>20 ELISA units/ml [EU]) anti-FHA and anti-FIM IgG antibodies were common (65 and 62% of individuals, respectively), but quantifiable anti-PT IgG antibodies were less frequent (16%). Given the distributions of antibody levels, an anti-PT IgG level of > or =94 EU was proposed as the diagnostic cutoff point. Application of this cutoff point to culture-confirmed illness in a prior study investigating cough illness yielded a high diagnostic sensitivity (80%) and specificity (93%). A standardized ELISA for anti-PT IgG with a single serum sample appears to be useful for the identification of recent B. pertussis infection in adolescents and adults with cough illness. The PT cutoff point will be further evaluated in prospective studies of confirmed B. pertussis infection.     

The fluorolysis assay,a highly sensitive method for measuring the cytolytic activity of T cells at very low numbers

We have developed a highly sensitive cytolysis test, the fluorolysis assay, as a simple nonradioactive and inexpensive alternative to the standard 51Cr-release assay. P815 cells were stably transfected with a plasmid expressing the enhanced green fluorescent protein (EGFP) gene. These target cells were coated with or without cognate peptide or anti-CD3 Ab and then incubated with CD8(+) T cells to allow antigen-specific or nonspecific lysis. The degree of target cell lysis was measured using flow cytometry to count the percentage of viable propidium iodide(-) EGFP(+) cells, whose numbers were standardized to a reference number of fluorochrome-linked beads. By using small numbers of target cells (200-800 per reaction) and extended incubation times (up to 2 days), the antigen-specific cytolytic activity of one to two activated CD8(+) T cells of a CTL line could be detected. The redirected fluorolysis assay also measured the activity of very few (> or =6) primary CD8(+) T cells following polyclonal activation. Importantly, antigen-specific lysis by small numbers (> or =25) of primary CD8(+) T cells could be directly measured ex vivo. This exquisite sensitivity of the fluorolysis assay, which was at least 8-33-folds higher than an optimized 51Cr-release assay, allows in vitro and ex vivo studies of immune responses that would otherwise not be possible due to low CTL numbers or frequencies.     

Comorbid psychiatric disorders in depressed outpatients: demographic and clinical features

BACKGROUND: This study evaluated the clinical and sociodemographic features associated with various degrees of concurrent comorbidity in adult outpatients with nonpsychotic major depressive disorder (MDD). METHODS: Outpatients enrolled in the STAR*D trial completed the Psychiatric Diagnostic Screening Questionnaire (PDSQ). An a priori 90% specificity threshold was set for PDSQ responses to ascertain the presence of 11 different concurrent DSM-IV Axis I disorders. RESULTS: Of 1376 outpatients, 38.2% had no concurrent comorbidities, while 25.6% suffered one, 16.1% suffered two, and 20.2% suffered three or more comorbid conditions. Altogether, 29.3% met threshold for social anxiety disorder, 20.8% for generalized anxiety disorder, 18.8% for posttraumatic stress disorder, 12.4% for bulimia, 11.9% for alcohol abuse/dependence, 13.4% for obsessive-compulsive disorder, 11.1% for panic disorder, 9.4% for agoraphobia, 7.3% for drug abuse/dependence, 3.7% for hypochondriasis, and 2.2% for somatoform disorder. Those with more concurrent Axis I conditions had earlier ages at first onset of MDD, longer histories of MDD, greater depressive symptom severity, more general medical comorbidity (even though they were younger than those with fewer comorbid conditions), poorer physical and mental function, health perceptions, and life satisfaction; and were more likely to be seen in primary care settings. LIMITATIONS: Participants had to meet entry criteria for STAR*D. Ascertainment of comorbid conditions was not based on a structured interview. CONCLUSIONS: Concurrent Axis I conditions (most often anxiety disorders) are very common with MDD. Greater numbers of concurrent comorbid conditions were associated with increased severity, morbidity, and chronicity of their MDD.     

Acrocentric interconnections and NOR variants in human lymphocytes

Acrocentric interconnections and NOR (nucleolus organizer region) variants are frequently observed in silver-stained metaphase preparations from lymphocytes of phenotypically normal individuals. The types of interconnections and of NOR variants are outlined. It is speculated that the satellite acrocentrics (both normal and variant) are the consequence of breakage and recoiling of these interconnections. Awareness of these two features of the human genome may facilitate understanding of the NOR/nucleolus interaction(s) in such important processes as nucleolus formation and in development and/or diagnosis of disease states (i.e., malignancy).     

Client choice of treatment and client outcomes

Participants in this study suffered from severe mental illness and were homeless at baseline. They were given their choice of five different treatment programs. The current study investigated two major questions: (1) what is the impact of positive expectancies about the efficacy of the chosen program on number of contacts with the chosen program and client outcomes; and (2) what is the impact of positive views about nonchosen programs (alternative choice variables) on contact with the chosen program and client outcomes. Client outcomes assessed were psychotic symptoms, days homeless, and client satisfaction. Positive expectancy variables were the number of reasons for choosing a program and confidence that the program would help. Alternative choice variables were the number of nonchosen programs visited and the attractiveness of a nonchosen program. Only the number of reasons for choosing the program was significantly related to program contact with the chosen program. Both of the positive expectancy variables and program contact were significantly correlated with consumer satisfaction. In general, neither the positive expectancy variables nor the alternative choice variables predicted changes in psychotic symptoms nor days homeless. © 2003 Wiley Periodicals, Inc. J Comm Psychol 31: 339–348, 2003.     

Effect of clarithromycin on cytokines and chemokines in children with an acute exacerbation of recurrent wheezing: a double-blind, randomized, placebo-controlled trial.

BACKGROUND: Clarithromycin is postulated to possess immunomodulatory properties in addition to its antimicrobial activity. OBJECTIVE: To evaluate the effect of clarithromycin on serum and nasopharyngeal cytokine and chemokine concentrations in children with an acute exacerbation of recurrent wheezing. METHODS: Children with a history of recurrent wheezing or asthma and who presented with an acute exacerbation of wheezing were enrolled in a double-blind, randomized trial of clarithromycin vs placebo. Concentrations of tumor necrosis factor alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-1beta (IL-1beta), IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, granulocyte-macrophage colony-stimulating factor, RANTES, eotaxin, macrophage inflammatory protein 1alpha, macrophage inflammatory protein 1beta, and monocyte chemoattractant protein 1 were measured in serum and/or nasopharyngeal aspirates before, during, and after therapy. Mycoplasma pneumoniae and Chlamydophila pneumoniae infection were evaluated for by polymerase chain reaction and serologic testing. RESULTS: Nasopharyngeal concentrations of TNF-alpha, IL-1beta, and IL-10 were significantly and persistently lower in children treated with clarithromycin compared with placebo. There tended to be a greater effect of clarithromycin on nasopharyngeal cytokine concentrations in patients with evidence of M. pneumoniae or C. pneumoniae infection. No significant differences were detected in serum cytokines for children treated with clarithromycin compared with placebo. CONCLUSION: Clarithromycin therapy reduces mucosal TNF-alpha, IL-1beta, and IL-10 concentrations in children with an acute exacerbation of recurrent wheezing.     

Differential localization of IL-2- and -15 receptor chains in membrane rafts of human T cells

We studied whether cytokine receptors (Rs) on T cells associate with lipid microdomains ("rafts"). Low-dose phytohemagglutinin (PHA)-stimulated human T cells were separated into cytoplasmic, membrane, and raft fractions by buoyant density centrifugation. Examination of these fractions for the presence of interleukin (IL)-2- and -15R chains and associated signaling molecules by Western blotting revealed marked, selective enrichment of the IL-2/15R beta-chain in rafts before IL-2 stimulation. After IL-2 stimulation, a substantial amount of the beta-chain was found in the membrane fraction. This partial translocation was also observed for the beta-chain-associated molecules JAK-1, p56(lck), and grb-2. Finally, raft disruption with methyl-beta-cyclodextrin (MBCD) attenuated IL-2-induced tyrosine phosphorylation events and selectively decreased the surface expression of the IL-2/15R beta-chain detected by flow cytometry. These results show that the IL-2/15R beta-chain is enriched in rafts obtained from low-dose, PHA-stimulated T cells, that IL-2 binding alters this enrichment, and that this enrichment may be functionally relevant as a possible mechanism to ensure cytokine selectivity and specificity.     

Accuracy of reporting endocervical component adequacy--a continuous quality improvement project

Inaccurate reporting of the absence of an endocervical (EC) component on Pap smears often results in slide rescreens, amended reports, clinician dissatisfaction, and sometimes unnecessary repeat smears. Therefore, the accuracy of reporting EC component adequacy was selected as a quality indicator for the laboratory continuous quality improvement program (CQI). The process consisted of problem identification, analysis of the situation, collection of data, implementation of solutions, and evaluation of results. The objective of the study was to determine if the accuracy of reporting EC component adequacy on Pap smears improved after application of such a program. During the first phase, 150 Pap smears originally reported with the absence of an adequate EC component and 150 smears reported with the presence of an adequate EC component were rescreened to measure the baseline accuracy of EC component adequacy reporting. The improvement process was then implemented. A cause-and-effect diagram was developed and root cause was determined. A presentation was then made to the cytology staff. Criteria for EC component adequacy were reviewed, examples were shown, and standardized marking of EC component was implemented. Following improvement actions, a second audit of 150 Pap smears reported with the absence of an adequate EC component as well as 150 smears reported with the presence of an adequate EC component was undertaken to measure change in performance in assessing EC component adequacy. For the baseline rescreening, before initiation of the CQI program, 98% accuracy was achieved with smears that were reported as adequate for EC component present. However, the accuracy with smears reported as absence of an adequate EC component was only 71%, i.e., an adequate EC component was identified in almost 1/3 of these cases on rescreen. After the implementation of improvement actions, the accuracy with smears reported with the presence of EC component remained high (98%) and the accuracy of reporting the absence of EC component was 90%. The difference of the latter before and after the implementation was statistically significant (P = 0.015, z-test). The accuracy of reporting EC component adequacy increased following the CQI process. Using reporting EC component adequacy as an example, we demonstrate that by treating clinical problems as quality control issues and applying basic quality improvement tools, a positive outcome can be effected.     

Summary of complementation groups of UV-sensitive CHO cell mutants isolated by large-scale screening

A summary is given for the lineage and complementation groupassignments of 153 UV-sensitive mutants of the CHO AA8 cellline. The distribution of mutants among six complementationgroups was highly non-random, with the great majority of theisolates belonging to groups 1 and 2. This asymmetry is consistentwith the known hemizygosity of these two linked loci in CHOcells. The relative numbers of mutants induced in group 2 wasfound to depend greatly on the type of mutagen used. Mutagenesiswith UV radiation, ethyl methanesulfonate (EMS), N-methyl-N'-nitro-N-nitroso-guanidine and 7-bromomethylbenz[a]anthraceneproduced high frequencies of group 2 mutants. In contrast, ICR170and ICR191, which are thought to produce mostly frameshift mutations,yielded very few mutants in group 2. These results are of particularimportance in light of the recent finding that the human ERCC2gene, which corrects group 2 mutants, has very strong homologywith the yeast gene RAD3. RAD3 is an essential gene for viabilityin yeast, and the low recovery of group 2 mutants using theframeshift agents strongly suggests that frameshift mutationstend to be lethal in the hamster ERCC2 locus. Several mutagen-sensitivedouble mutants were isolated in two-step selections from EMS-,mitomycin C- or UV-sensitive parental cells, including the lineUVU1, the first mammalian line with two mutations that affectUV sensitivity. The first mutation inactivated excision repair,and the second mutation appears to have affected some otherrecovery process. UVU1 should be useful for studying recoveryprocesses that are separate from nucleotide excision repair. ¹To whom correspondence should be addressed