An oxygen-rich atmosphere or systemic fluoxetine extend the time to respiratory arrest in a rat model of obstructive apnea.

Audiogenic seizure-prone mice can be protected from seizure-associated death by exposure to an oxygen atmosphere or treatment with selective serotonergic reuptake inhibitors (SSRIs). We have shown previously in a rat model that epileptic seizure activity can spread through brainstem areas to cause sufficient laryngospasm for obstructive apnea and that the period of seizure-associated obstructive apnea can last long enough for respiratory arrest to occur. We hypothesized that both the oxygen-rich atmosphere and SSRIs function by prolonging the time to respiratory arrest, thus ensuring that seizure activity stops before the point of respiratory arrest to allow recovery of respiratory function. To test this hypothesis, we evaluated each preventative treatment in a rat model of controlled airway occlusion where the times to respiratory arrest can be measured. Adult male Sprague Dawley rats (median age = 66 days) were studied in the absence of any seizure activity. By directly studying responses to controlled airway occlusion, rather than airway occlusion secondary to seizure activity, we could isolate the effects of manipulations that might prolong respiratory arrest from the effects of those manipulations on seizure intensity. All group sizes were ≥ 8 animals per group. We found that both oxygen exposure and fluoxetine significantly increased the time to respiratory arrest by up to 65% (p < .0001 for 5 min oxygen exposure; p = .031 for 25 mg/kg fluoxetine tested 60 min after injection) and, given that neither treatment has been shown to significantly alter seizure duration, these increases can account for the protection of either manipulation against death in sudden death models. Importantly, we found that 30 s of exposure to oxygen produced nearly the same protection as 5 min exposure suggesting that oxygen exposure could start after a seizure starts (p = .0012 for 30 s oxygen exposure). Experiments with 50% oxygen/50% air mixtures indicate that the oxygen concentration needs to be above about 60% to ensure that times to respiratory arrest will always be longer than a period of seizure-induced airway occlusion. Selective serotonin reuptake inhibitors, while instructive with regard to mechanism, require impractical dosing and may carry additional risk in the form of greater challenges for resuscitation. We conclude that oxygen exposure or SSRI treatment prevent seizure associated death by sufficiently prolonging the time to respiratory arrest so that respiratory function can recover after the seizure abates and eliminates the stimulus for seizure-induced apnea.

Serpin protein CrmA suppresses hypoxia-mediated apoptosis of ventricular myocytes.

BACKGROUND: In this study, we ascertain whether caspase 8 activation and mitochondrial defects underlie apoptosis of ventricular myocytes during hypoxia. As an approach to circumvent the potential shortcomings surrounding the limited permeability and short half-life of the synthetic peptide inhibitors designed to block caspase activation, we constructed a replication-defective adenovirus encoding the serpin caspase inhibitor protein CrmA to ensure efficient and continual inhibition of caspase 8 activity during chronic hypoxia. METHODS AND RESULTS: In contrast to normoxic cells, oxygen deprivation of postnatal ventricular myocytes for 24 hours resulted in a 9-fold increase (P<0.05) in apoptosis as determined by Hoechst 33258 staining and nucleosomal DNA laddering. Moreover, hypoxia provoked a 1.5-fold increase (P<0.01) in caspase 8-like activity. Furthermore, hypoxia provoked perturbations to mitochondria consistent with the mitochondrial death pathway, including permeability transition pore (PT) opening, loss of mitochondrial membrane potential ((m)), and cytochrome c release. Importantly, CrmA suppressed caspase 8 activity, PT pore changes, loss of (m), and apoptosis but had no effect on hypoxia-mediated cytochrome c release. Furthermore, Bongkrekic acid, an inhibitor of PT pore, prevented hypoxia-induced PT pore changes, loss of (m), and apoptosis but had no effect on hypoxia-mediated cytochrome c release. CONCLUSIONS: To our knowledge, we provide the first direct evidence for the operation of CrmA as an antiapoptotic factor in ventricular myocytes during prolonged durations of hypoxia. Furthermore, our data suggest that perturbations to mitochondria including PT pore changes and (m) loss are caspase-regulated events that appear to be separable from cytochrome c release.

Behavioral aspects of experimental autoimmune encephalomyelitis.

Acute inflammation is known to induce a depressive-like sickness behavior syndrome in humans and in experimental animals. In the present study, we sought to determine whether a chronic neuroautoimmune inflammation is also associated with a similar behavioral syndrome. Experimental autoimmune encephalomyelitis (EAE) was induced in SJL/J female mice by adoptive transfer of lymph node cells, and sickness behavior symptoms, including anorexia, loss of body weight, reduced social exploration, and decreased preference for sucrose solution were measured. We report that these components of sickness behavior were induced during the acute phase of the disease, and recovered in later phases. Moreover, the onset and recovery of the behavioral symptoms preceded the onset and recovery of the neurological signs, respectively. Since EAE is considered a model for multiple sclerosis (MS), it is suggested that EAF-induced behavioral changes may serve as a model for the depressive symptomatology that characterizes most MS patients.

[Drowning in Israel--1990-1992].

There are no epidemiological studies of drowning in Israel in the scientific literature, despite prominent reports in the media. We analyzed the extent of mortality from drowning in Israel during 1990-92, attempting to identify and characterize groups at high risk, and to determine trends and differences between Israel and other countries. Computerized death certificate files were provided by the Israel government Central Bureau of Statistics; deaths from drowning were included among external causes codes E-830, 832, E-910, E-954, E-964, E-984. In Israel, as in the United States, mortality from drowning is the fourth cause of death among all unintentional causes, and the second cause in age-groups 1-24, ranking after transport accidents. During 1990-92 there were 1.2 drownings a year/100,000 population. The highest rates were found among young non-Jews 15-24 years old (7.8/100,000) and among elderly Jews (3.5/100,000). Unintentional drowning accounted for 89% of all deaths; while about 10% were defined as suicides. Males had a rate almost 3 times greater than females, and the among Arabs was 2.4 times greater than among Jews. The 1990-92 drowning rate was slightly lower than in previous years. Drowning rates in Israel are lower than in the United States, except in the elderly. Apparently the principal reason for this difference is difference in sites of drowning. In Israel most drownings occur in the sea, so site data are unavailable for international comparisons. Careful consideration of the categories (E-codes) included in the rates, and of local registration procedures, is necessary for international comparisons. Foreign workers, tourists and other nonresidents are not included in national vital statistics. But estimations based on Ministry of Interior sources show that foreign workers (most, recent arrivals) during the last few years are an extremely high risk group. Population-based drowning rates are not an accurate estimation of drowning risk, since universal exposure to the "opportunity to drown" is assumed.

[Gastric functional disorders in patients with various hemodynamic types of acute myocardial infarction]. / Funktsional'nye narusheniia zheludka u bol'nykh s razlichnymi gemodinamicheskimi variantami ostrogo infarkta miokarda.

The paper is concerned with the results of investigation of acid- and enzyme-producing, motor and evacuatory function of the stomach on days 2, 10-15, 28-35 after myocardial infarction. Correlation of the type and severity of gastric dysfunction and the type of hemodynamic reactions developing in myocardial infarction, was established. The most serious and prolonged disorders were detected in patients with hyper- and especially hypodynamic disorders of central hemodynamics which were indicative of close relationships between the type of hemodynamic reactions in myocardial infarction and gastric function. Drug correction of gastric dysfunction in myocardial infarction is achieved by administration of sulpiride and protein-oxygen cocktails which are recommended for prophylactic purposes.

Diagnosis of renal tumors on needle biopsy specimens by histological and molecular analysis.

PURPOSE: We diagnosed the subtypes of renal cell carcinoma on needle core biopsies using a combination of histopathology and a molecular diagnostic algorithm. MATERIALS AND METHODS: Core biopsies were taken of renal tumors following nephrectomy. RNA was extracted and quantitative real-time polymerase chain reaction was performed for 4 gene products to differentiate among renal cell carcinoma subtypes. Histopathological diagnosis was achieved on a second core before and after obtaining the molecular diagnostic algorithm results. RESULTS: Based on the nephrectomy diagnosis 6 of 77 renal masses were nonneoplastic and 71 were tumors, including 65 renal cell carcinoma/oncocytomas. The overall diagnostic accuracy using histology and our molecular diagnostic algorithm combined was 90.0% (70 of 77). Side by side comparison of histology vs molecular diagnostic algorithm was feasible for 60 classifiable renal cell carcinoma/oncocytomas (31 clear cell, 14 papillary renal cell carcinoma, 6 chromophobe renal cell carcinoma, 2 mucinous tubular and spindle cell carcinoma, and 7 oncocytoma). In this group histology correctly predicted the final histological subtype in 83.3% (50 of 60) of cores. Addition of the molecular diagnostic algorithm to histology improved the subtyping accuracy to 95% (57 of 60), whereas the molecular diagnostic algorithm alone was accurate in 50 of 60 cases (83.3%). Dividing these 60 specimens into clear cell and nonclear cell neoplasms, the addition of the molecular diagnostic algorithm improved the sensitivity for the diagnosis of clear cell carcinoma from 87.1% (27 of 31) to 100% and the negative predictive value from 87.5% to 100%. CONCLUSIONS: Core biopsies of renal tumors provide adequate material for diagnosing and subtyping renal cell carcinoma. The addition of our molecular diagnostic algorithm to histology improved the diagnostic accuracy of core biopsies of renal masses.

Mutant telomeres inhibit transcriptional silencing at native telomeres of the yeast Kluyveromyces lactis.

We report the identification and characterization of transcriptional silencing at native telomeres in the budding yeast Kluyveromyces lactis. We show that K. lactis telomeres are able to repress the transcription of a gene located at the junction between the telomeric repeat tract and the subtelomeric domain. As in Saccharomyces cerevisiae, switching between the repressed and derepressed transcriptional states occurs. C-terminal truncation of the telomere binding protein Rap1p, which leads to a regulated alteration in telomere length, reduces telomeric silencing. In addition, telomeric silencing is reduced dramatically in telomerase RNA mutants in which telomere length control has been lost. This is consistent with the possibility that the structure of the entire telomere affects the silencing functions exhibited by its internal domain.

Isolation and characterization of a ColE1 plasmid containing the entire bio gene cluster of Escherichia coli K12.

ColE1amp plasmids carrying the entire bio gene cluster were constructed in vitro using ColE1amp as the cloning vehicle and a lambda transducing phage, lambdaatt2, as the source of bio DNA. Restriction endonuclease EcoRI digests of ColE1amp and lambdaatt2 DNA were joined by polynucleotide ligase and plasmids bearing the entire bio gene cluster were selected, after transformation, in bio deletion strains of E. coli. Recombinant DNA molecules contained one ColE1amp fragment (7.4 X 10(6) daltons) and one lambdaatt2 DNA fragment (5.4 X 10(6) daltons). Clones carrying ColE1 amp-bio plasmids produce elevated levels of biotin and biotin synthetase activity.

Caspase activation and mitochondrial cytochrome C release during hypoxia-mediated apoptosis of adult ventricular myocytes.

Oxygen deprivation for prolonged periods leads to cardiac cell death and ventricular dysfunction. The ability to prevent myocardial cell death would be of significant therapeutic value in maintaining cardiac function after injury. While caspases have been suggested to play a critical role in apoptosis, their involvement during hypoxic injury has not been formally determined. In this report, we show that adult ventricular myocytes subjected to hypoxia for 1 h undergo a three-fold increase (P<0.05) in the incidence of apoptosis as determined by TUNEL analysis and Hoechst 33258 nuclear staining. Western blot analysis of hypoxic myocytes revealed a 10-fold increase in the proteolytic processing of caspase 3 to p17 with a concomitant cleavage of the caspase 3 substrate PARP from 116 kd to p85 kd compared to normoxic controls. Defects in mitochondrial membrane integrity were also observed as evidenced by the translocation of cytochrome c from the mitochondrial to cytosolic compartment of hypoxic cells. Pretreatment of ventricular myocytes with the peptide-caspase inhibitor known to block caspases related to caspase 1 (Ac-YVAD-CHO) attenuated cytochrome c release, processing of caspase 3, and apoptosis. While the caspase inhibitor (Ac-DEVD-CHO) which blocks caspases related to caspase 3, suppressed the cleavage of PARP and apoptosis, it had no effect on cytochrome c release by mitochondria. The data provide direct evidence for the proteolytic activation of caspases during hypoxia-mediated apoptosis of adult ventricular myocytes. Furthermore, the data suggest a hierarchical scheme for caspase activation with mitochondrial cytochrome c release occurring proximally to DEVD-CHO-inhibitable caspases.

Genetic transformation of obligately chemolithotrophic thiobacilli.

Genetic transformation of Thiobacaillus thioparus auxotrophs to prototrophy was obtained at frequencies of up to 10(-2) when proliferating cell populations were exposed to chromosomal DNA from a nutritionally independent strain of the same bacterium. The rate at which transformation occurred depended on recipient growth rate and could be drastically reduced by depriving otherwise competent cells of either nitrogen or exogenous energy substrate. Interspecies marker transfer was also shown among several obligately chemolithotrophic members of the genus.