Protein synthesis, development, growth and life span.

To test the hypothesis that reduced protein synthesis may increase life span by retarding genetic informational transfer during early life and reducing the use of the genetic code and thereby minimizing genetic imperfections as they may occur during late life, two approaches were used. In the first protein synthesis was depressed by the administration of cycloheximide, in the second by reducing the dietary protein level. One-day-old chick embryos were injected with either 0.8 gamma or 1.0 gamma of cycloheximide. On the second and third day of incubation both stage of development and heart rate were lower in the treated embryos. Growth was retarded throughout the 17 days of incubation as measured by size and DNA contents. As estimated by the activities of various enzymes per unit DNA, cells of the treated embryos were the same as normal ones of the same age. Sixteen-month-old female Wistar rats which had been previously maintained on a commercial diet (23.4% protein) were fed diets which contained either 24, 12, 8 or 4% casein throughout their remaining life span. Except for a lowering of the body weights of the animals fed the 4% casein diet, the body weights of the remaining animals were unchanged. Reducing the dietary protein level from 24% to 12% increased the life span (25%) of the animals.     

Airway hyperresponsiveness in allergic rhinitis. A risk factor for asthma

In order to study whether the methacholine inhalation challenge could predict which patients with allergic rhinitis were at risk to develop asthma, we prospectively studied a group of ragweed-sensitive patients over a four to five year period. On the initial study, 16 of 40 patients (40 percent) were found to be hyperresponsive to methacholine. On the follow-up study, three of these 16 patients (19 percent) were found to have developed asthma from one and one-half to five years after the initial testing. Each had greater methacholine responsiveness on repeat study. The degree of methacholine hyperresponsiveness, judged by the PD20, could not predict which of the initial responders would develop asthma. Twenty-four (60 percent) of our patients showed normal responses to methacholine on initial study; none developed asthma and 88 percent remained nonresponders on repeat study. Our study shows that allergic rhinitis patients hyperresponsive to methacholine are at greater risk to develop asthma than those with normal bronchial challenges (p less than 0.05).     

Makaluvamines vary in ability to induce dose-dependent DNA cleavage via topoisomerase II interaction

The makaluvamines are marine natural products that were originally isolated because of their cytotoxicity in a cell-based mechanism screen. They have significant anti-cancer activity in animal models. There is, however, disagreement in the literature as to whether these compounds target topoisomerase II via a clinically relevant mechanism. This work shows that the makaluvamines can induce dose-dependent DNA cleavage via topoisomerase II. For most of the makaluvamines the levels of cleavage are significantly below those achieved by equimolar concentrations of etoposide. To some extent these results might explain the discrepancies present in the literature.     

The sumatriptan/naratriptan aggregated patient (SNAP) database: aggregation, validation and application

Pooled data from multiple clinical trials can provide information for medical decision-making that typically cannot be derived from a single clinical trial. By increasing the sample size beyond that achievable in a single clinical trial, pooling individual-patient data from multiple trials provides additional statistical power to detect possible effects of study medication, confers the ability to detect rare outcomes, and facilitates evaluation of effects among subsets of patients. Data from pharmaceutical company-sponsored clinical trials lend themselves to data-pooling, meta-analysis, and data mining initiatives. Pharmaceutical company-sponsored clinical trials are arguably among the most rigorously designed and conducted of studies involving human subjects as a result of multidisciplinary collaboration involving clinical, academic and/or governmental investigators as well as the input and review of medical institutional bodies and regulatory authorities. This paper describes the aggregation, validation and initial analysis of data from the sumatriptan/naratriptan aggregate patient (SNAP) database, which to date comprises pooled individual-patient data from 128 clinical trials conducted from 1987 to 1998 with the migraine medications sumatriptan and naratriptan. With an extremely large sample size (>28000 migraineurs, >140000 treated migraine attacks), the SNAP database allows exploration of questions about migraine and the efficacy and safety of migraine medications that cannot be answered in single clinical trials enrolling smaller numbers of patients. Besides providing the adequate sample size to address specific questions, the SNAP database allows for subgroup analyses that are not possible in individual trial analyses due to small sample size. The SNAP database exemplifies how the wealth of data from pharmaceutical company-sponsored clinical trials can be re-used to continue to provide benefit.     

Clinical behaviors and skills that faculty from 12 institutions judged were essential for medical students to acquire

This paper describes a 1988-1989 collaborative mail survey of faculty opinion about clinical behaviors and skills that students should be expected to demonstrate prior to graduation from undergraduate medical school (hereafter called "exit objectives"). Selected faculty from 12 American and Canadian medical schools indicated whether each of 77 objectives was essential for every student to know or demonstrate prior to graduation; useful but not essential at the undergraduate level; or not applicable to their undergraduate program. Their responses provide a glimpse into faculty expectations regarding some of the exit behaviours and skills they deemed essential. Forty-two percent (32) of the 77 objectives were regarded as essential by 75% or more of the faculty members who responded. Essential objectives involved conducting organ system examinations, formulating problems and hypotheses, and gathering fundamental interview, physical, and screening examination data, including emergency examinations of the airway and circulatory systems. Other essential objectives involved collaboration and communication, demonstrating concern for legal and ethical values, and keeping abreast of current information within the discipline. Exit objectives related to the diagnosis and management of specific conditions were regarded as useful but not essential at the undergraduate level. Implications for medical education are discussed.     

Nitric oxide inhibits Coxiella burnetii replication and parasitophorous vacuole maturation

Nitric oxide is a recognized cytotoxic effector against facultative and obligate intracellular bacteria. This study examined the effect of nitric oxide produced by inducible nitric oxide synthase (iNOS) up-regulated in response to cytokine stimulation, or by a synthetic nitric oxide donor, on replication of obligately intracellular Coxiella burnetii in murine L-929 cells. Immunoblotting and nitrite assays revealed that C. burnetii infection of L-929 cells augments expression of iNOS up-regulated in response to gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha). Infection in the absence of cytokine stimulation did not result in demonstrable up-regulation of iNOS expression or in increased nitrite production. Nitrite production by cytokine-treated cells was significantly inhibited by the iNOS inhibitor S-methylisothiourea (SMT). Treatment of infected cells with IFN-gamma and TNF-alpha or the synthetic nitric oxide donor 2,2'-(hydroxynitrosohydrazino)bis-ethanamine (DETA/NONOate) had a bacteriostatic effect on C. burnetii replication. Inhibition of replication was reversed upon addition of SMT to the culture medium of cytokine-treated cells. Microscopic analysis of infected cells revealed that nitric oxide (either cytokine induced or donor derived) inhibited formation of the mature (large) parasitophorous vacuole that is characteristic of C. burnetii infection of host cells. Instead, exposure of infected cells to nitric oxide resulted in the formation of multiple small, acidic vacuoles usually containing one C. burnetii cell. Removal of nitrosative stress resulted in the coalescence of small vacuoles to form a large vacuole harboring multiple C. burnetii cells. These experiments demonstrate that nitric oxide reversibly inhibits replication of C. burnetii and formation of the parasitophorous vacuole.     

Aberrant methylation of liver DNA in rats during hepatotoxicity

Liver DNA of young adult male Fischer 344 rats given orally a necrogenic dose of hydrazine (60 mg/kg body wt) rapidly became methylated at the 7-position of guanine, and more slowly and to a lesser extent at the O⁶-position of guanine. The identity of 7-methylguanine from liver DNA of hydrazine-treated rats was confirmed by four high-pressure liquid chromatographic methods, in one paper chromatography system, and by ultraviolet absorption spectrometry. Similar evidence was obtained to support the identification of O⁶-methylguanine. Liver DNA isolated from rats given hydrazine and [methyl-³H]methionine contained 7-methylguanine and O⁶-methylguanine which were labeled to a greater extent than was the parent base, guanine, suggesting that S-adenosylmethionine was the methyl source in the alkylation of DNA. Aberrant methylation of liver DNA was also observed in rats after administration of hepatotoxic doses of carbon tetrachloride and ethanol, indicating that the DNA methylation may be a nonspecific response to chemical injury to the liver.     

RNAi screening reveals requirement for host cell secretory pathway in infection by diverse families of negative-strand RNA viruses

Negative-strand (NS) RNA viruses comprise many pathogens that cause serious diseases in humans and animals. Despite their clinical importance, little is known about the host factors required for their infection. Using vesicular stomatitis virus (VSV), a prototypic NS RNA virus in the family Rhabdoviridae, we conducted a human genome-wide siRNA screen and identified 72 host genes required for viral infection. Many of these identified genes were also required for infection by two other NS RNA viruses, the lymphocytic choriomeningitis virus of the Arenaviridae family and human parainfluenza virus type 3 of the Paramyxoviridae family. Genes affecting different stages of VSV infection, such as entry/uncoating, gene expression, and assembly/release, were identified. Depletion of the proteins of the coatomer complex I or its upstream effectors ARF1 or GBF1 led to detection of reduced levels of VSV RNA. Coatomer complex I was also required for infection of lymphocytic choriomeningitis virus and human parainfluenza virus type 3. These results highlight the evolutionarily conserved requirements for gene expression of diverse families of NS RNA viruses and demonstrate the involvement of host cell secretory pathway in the process.