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Biomedical subjects

J Mitchell

Publications and source records attributed to J Mitchell.

At least 127 records · Page 7Linked to original sources
PubMed1997

CDC45, a novel yeast gene that functions with the origin recognition complex and Mcm proteins in initiation of DNA replication.

The CDC45 gene of Saccharomyces cerevisiae was isolated by complementation of the cold-sensitive cdc45-1 mutant and shown to be essential for cell viability. Although CDC45 genetically interacts with a group of MCM genes (CDC46, CDC47, and CDC54), the predicted sequence of its protein product reveals no significant sequence similarity to any known Mcm family member. Further genetic characterization of the cdc45-1 mutant demonstrated that it is synthetically lethal with orc2-1, mcm2-1, and mcm3-1. These results not only reveal a functional connection between the origin recognition complex (ORC) and Cdc45p but also extend the CDC45-MCM genetic interaction to all known MCM family members that were shown to be involved in replication initiation. Initiation of DNA replication in cdc45-1 cells was defective, causing a delayed entry into S phase at the nonpermissive temperature, as well as a high plasmid loss rate which could be suppressed by tandem copies of replication origins. Furthermore, two-dimensional gels directly showed that chromosomal origins fired less frequently in cdc45-1 cells at the nonpermissive temperature. These findings suggest that Cdc45p, ORC, and Mcm proteins act in concert for replication initiation throughout the genome.

Amino Acid Sequence↗
PubMed1997

A targeted mutation at the T-cell receptor alpha/delta locus impairs T-cell development and reveals the presence of the nearby antiapoptosis gene Dad1.

Locus control regions are cis gene regulatory elements comprised of DNase I-hypersensitive sites. These regions usually do not stimulate transcription outside of a chromosomal context, and therefore their ability to regulate the expression of genes is thought to occur through the modification of chromatin accessibility. A locus control region is located downstream of the T-cell receptor (TCR) alpha/delta locus on mouse chromosome 14. This locus control region is known to drive T-cell-specific TCR alpha transcription in transgenic mice. In this report, we describe a targeted deletion of this locus control region and show that this mutation acts at a critical checkpoint in alphabeta T-cell development, between the TCR-intermediate and TCR-high stages. Our analysis further reveals that the antiapoptosis gene Dad1 is at the 3' end of the TCR alpha/delta locus and that Dad1 is required for embryogenesis. We show that mouse Dad1 has a broader expression pattern than the TCR genes, in terms of both tissue and temporal specificity. Finally, we report that the chromatin between TCR alpha and Dad1 is DNase I hypersensitive in a variety of cell types, thus correlating with Dad1 expression and raising the possibility that Dad1 regulatory sequences reside in this region.

Alleles↗
PubMed1997

Dexamethasone increases G alpha q-11 expression and hormone-stimulated phospholipase C activity in UMR-106-01 cells.

Glucocorticoids regulate responsiveness of many cells to hormones that bind to G protein-coupled receptors. We examined the effect of glucocorticoids on parathyroid hormone (PTH) activation of two G protein-activated signal transduction pathways, phospholipase C (PLC) and adenylyl cyclase, in osteosarcoma UMR-106-01 cells. Dexamethasone (100 nM) increased PTH-stimulated and NaF-stimulated PLC activity by > 100% over 4 days (223 +/- 8 and 293 +/- 8.2% of control after 4 days for PTH and NaF-stimulated activity, respectively). The increase in PTH-stimulated adenylyl cyclase response in the same cells was more modest (162 +/- 5.4 and 171 +/- 6.8% of control after 4 days for PTH and NaF-stimulated activity, respectively). PTH activation of PLC was blocked by antiserums to G alpha q-11 and activation of adenylyl cyclase by G alpha s antiserums. Quantification of these G protein subunits in control and dexamethasone-treated cells showed a 78% increase in G alpha q-11 (from 18.1 +/- 1.2 to 32.2 +/- 1.5 pmol/mg), whereas G alpha s was increased only 34% (from 6.2 +/- 0.5 to 8.2 +/- 0.3 pmol/mg) and G beta-subunits were increased 40% (from 54 +/- 2.3 to 75.2 +/- 3.8 pmol/mg). These results suggest that glucocorticoids are more potent regulators of PLC activity than adenylyl cyclase activity in UMR cells, and this is mediated, at least in part, by differential increases in G alpha q-11 proteins.

Adenylate Cyclase Toxin↗
PubMed1997

Effectiveness and economic impact of antidepressant medications: a review.

This article reviews the existing literature on the pharmacoeconomics and effectiveness of antidepressant medications. Although selective serotonin reuptake inhibitors (SSRIs) have not proved to be more efficacious than the older tricyclics, and their prescription costs are significantly higher, they provide superior effectiveness; ie, patients are less likely to discontinue taking them or switch antidepressants. Pharmacoeconomic studies consistently demonstrate a relationship between this superior effectiveness and reductions in overall treatment costs, often through decreased utilization of medical and hospital services. The most conservative study found a cost offset that more than negated the extra cost of drugs, although the cost savings were not statistically significant. Other studies found statistically significant lowering of utilization costs by using SSRIs rather than tricyclics. Studies comparing SSRIs with each other present conflicting findings, although fluoxetine appears to have an edge over sertraline and paroxetine with regards to effectiveness and pharmacoeconomics. More studies employing a prospective outcome design and naturalistic study setting need to be conducted with SSRIs and other new antidepressants.

Antidepressive Agents, Second-Generation↗
PubMed1997

Effect of seizures on hippocampal peptidergic neurons.

Results from animal studies and from human tissue removed from epileptics show that certain subgroups of hippocampal neurons are more vulnerable to seizure activity than others. It is possible that neurons which contain calcium-binding proteins, such as parvalbumin, may be protected from the high calcium overload that results from seizure activity. In the present study, seizures were induced by an injection of tetanus toxin into the rat hippocampus. A morphological and quantitative analysis was made of the parvalbumin-containing neurons and of those which co-localized somatostatin and neuropeptide Y. At 2 weeks there was a generalized increase in immunoreactivity in both groups of neurons. From 1 month through to 3 months after injection, the up-regulation in immunoreactivity was sustained in the surviving hilar neurons which co-localized somatostatin and neuropeptide Y but there was a marked reduction in immunoreactivity of the parvalbumin neurons. Although there was no evidence for a loss of parvalbumin neurons there was a small and significant reduction in the number of somatostatin + neuropeptide Y double-labelled neurons in the contralateral hilus at 3 and 4 months after a tetanus injection. The vulnerability of the somatostatin + neuropeptide Y double-labelled hilar neurons but not of the parvalbumin-containing, presumed, basket cells are considered in terms of their connectivity.

Animals↗
PubMed1996

Comparison of phenotype in uniparental disomy and deletion Prader-Willi syndrome: sex specific differences.

Prader-Willi syndrome (PWS) results primarily from either a paternal deletion of 15q11-q13 or maternal uniparental disomy (UPD) 15. Birth parameters and clinical presentation of 79 confirmed UPD cases and 43 deletion patients were compared in order to test whether any manifestations differ between the two groups. There were no major clinical differences between the two classes analyzed as a whole, other than the presence of hypopigmentation predominantly in the deletion group. However, there was a significant bias in sex-ratio (P < .001) limited to the UPD group with a predominance (68%) of males. An equal number of males and females was observed in the deletion group. When analyzed by sex, several significant differences between the UPD and deletion groups were observed. Female UPD patients were found to be less severely affected than female deletion patients in terms of length of gavage feeding and a later onset of hyperphagia. Although these traits are likely to be influenced by external factors, they may reflect a milder presentation of female UPD patients which could explain the observed sex bias by causing under-ascertainment of female UPD. Alternatively, there may be an effect of sex on either early trisomy 15 survival or the probability of somatic loss of a chromosome from a trisomic conceptus.

Age Factors↗
PubMed1996

Effect of antioxidants on lipopolysaccharide-stimulated induction of mangano superoxide dismutase mRNA in bovine pulmonary artery endothelial cells.

Generation of reactive oxygen species (ROS) is a common event in the pathogenesis of acute lung injury. Endothelial cells may be both a target and a source of the ROS. Exposure of bovine pulmonary endothelial cells (BPAEC) to lipopolysaccharide (LPS) has been shown to result in intracellular generation of both ROS and the antioxidant enzyme, mangano superoxide dismutase (MnSOD). The present study investigates whether alterations in intracellular oxidant state affect LPS-stimulated cytotoxicity and induction of MnSOD mRNA. BPAEC were pretreated with either the free radical scavenger, dimethylsulfoxide (DMSO), the xanthine oxidase inhibitor, allopurinol, or N-acetylcysteine (a cysteine derivate capable of increasing glutathione stores) prior to exposure to LPS (0.1 microgram/ml) for either 4, 8 or 18 hours. We found that pretreatment of BPAEC with DMSO blocked both LPS-induced cytotoxicity and induction of the MnSOD gene. Nuclear run-off experiments demonstrated that LPS-stimulated induction of the MnSOD mRNA occurred at the transcriptional level and that DMSO blocked this event. Pretreatment with allopurinol also prevented the cytotoxicity associated with LPS but, in contrast to DMSO, did not alter induction of MnSOD mRNA. N-acetylcysteine did not affect the LPS-stimulated cytotoxicity but resulted in an early and transient reduction in induction of the MnSOD gene. We conclude that LPS stimulates generation of intracellular ROS that regulate induction of the MnSOD gene at the transcriptional level further, we conclude that LPS-stimulated cytotoxicity involves both the xanthine oxidase pathway and perhaps intracellular generation of hydroxyl radicals. The difference in the protective effect between DMSO, NAC and allopurinol suggest that upregulation of the MnSOD gene does not contribute to LPS-induced cytotoxicity.

Acetylcysteine↗
PubMed1996

Hippocampal NPY neurons project to the fascia dentata in organotypic cultures.

The distribution of neuropeptide Y immunoreactive (NPY-ir) neurons in organotypic cultures of hippocampi from neonates was compared to that seen in adult rats. In addition to the known NPY-ir neurons in the hippocampus proper and in the hilus of the fascia dentata, isolated, large, multipolar, NPY-ir neurons were observed in the subiculum and in areas CA1 and CA3. Their axons projected into stratum radiatum of the hippocampus proper and into the molecular layers and hilus of the fascia dentata where they branched profusely. These NPY-ir neurons were regularly distributed throughout the septo temporal extent of the hippocampus and were present in both neonates and adult hippocampi. The hilar NPY-ir neurons have always been considered the source of the NPY-ir plexus in the outer molecular layer of the dentate gyrus. However, our results show that there is also a contribution from the NPY-ir neurons in the hippocampus proper.

Animals↗
PubMed1996

T-lymphocyte subsets in patients with abnormal body weight: longitudinal studies. in anorexia nervosa and obesity.

OBJECTIVE: In contrast to other types of starvation which are characterized by low CD4+ counts and increased susceptibility to infection, anorexia nervosa is not associated with an increase in infectious complications. To determine why infection risk of anorectics differs from that of other starving populations, we studied T-lymphocytes, including CD4+ and CD8+ phenotypes, in patients with anorexia nervosa, and for comparison, in dieting obese subjects. METHODS: T-lymphocyte phenotypes were determined by flow cytometric analysis of monoclonal antibody-labeled cells obtained from patients with anorexia nervosa before and after successful therapy and weight gain, and in obese subjects before and after weight loss on a very-low-calorie diet. RESULTS: Weight loss in anorectics and obese dieters was associated with normal CD4+ counts. Unexpectedly, CD8+ counts were low in anorectics, both before and after weight gain, and in obese subjects after (but not before) dieting. DISCUSSION: Normal CD4+ counts in anorectics and obese dieters, despite marked weight loss, may explain the lack of increased infection risk in these eating-disordered patients, in contrast to other starving populations. The observation that CD8+ counts are low in anorectics with low and restored body weight and in obese patients after dieting has not been previously reported. The persistence of low CD8+ counts in anorectics even after weight gain suggests that some factors other than weight loss per se may be involved, possibly including effects due to stress, comorbid psychiatric conditions, or unidentified aspects of dysregulated pathophysiology secondary to disordered eating.

Adolescent↗
PubMed1996

OIL FLAIR: optimized interleaved fluid-attenuated inversion recovery in 2D fast spin echo.

Inversion recovery may be used to suppress signal from cerebrospinal fluid, a technique which has been named "fluid attenuated with inversion recovery" (FLAIR). This report describes interleaving a slice selective inversion pulse within a rapid spin-echo sequence to obtain the desirable contrast characteristics of FLAIR in imaging times comparable to standard rapid spin echo. Additionally, the pulse repetition time is allowed to float above a defined minimum, which can further shorten scan times and dramatically ease the optimization process. The optimized interleaved sequence is referred to as OIL FLAIR.

Central Nervous System↗
PubMed1996

Low incidence of BRCA2 mutations in breast carcinoma and other cancers.

Inherited mutant alleles of familial tumour suppressor genes predispose individuals to particular types of cancer. In addition to an involvement in inherited susceptibility to cancer, these tumour suppressor genes are targets for somatic mutations in sporadic cancers of the same type found in the familial forms. An exception is BRCA1, which contributes to a significant fraction of familial breast and ovarian cancer, but undergoes mutation at very low rates in sporadic breast and ovarian cancers. This finding suggests that other genes may be the principal targets for somatic mutation in breast carcinoma. A second, recently identified familial breast cancer gene, BRCA2 (refs 5-8), accounts for a proportion of breast cancer roughly equal to BRCA1. Like BRCA1, BRCA2 behaves as a dominantly inherited tumour suppressor gene. Individuals who inherit one mutant allele are at increased risk for breast cancer, and the tumours they develop lose the wild-type allele by heterozygous deletion. The BRCA2 coding sequence is huge, composed of 26 exons that span 10,443 bp. Here we investigate the rate of BRCA2 mutation in sporadic breast cancers and in a set of cell lines that represent twelve other tumour types. Surprisingly, mutations in BRCA2 are infrequent in cancers including breast carcinoma. However, a probable germline mutation in a pancreatic tumour cell line suggests a role for BRCA2 in susceptibility to pancreatic cancer.

BRCA2 Protein↗
PubMed1996

Pre-exposure to subtoxic levels prevents kainic acid lesions in organotypic hippocampal slice cultures: effects of kainic acid on parvalbumin-immunoreactive neurons and expression of heat shock protein 72 following the induction of tolerance.

The effects of kainic acid on the survival of principal neurons and parvalbumin-immunoreactive (PARV-IR) neurons, and on the expression of heat shock protein 72 immunoreactivity (HSP72-IR) were investigated in organotypic hippocampal slice cultures. Untreated cultures displayed an organotypic organization and the development and morphology of PARV-IR neurons in the hippocampus paralleled that reported to occur in vivo, with the exception of the hilar region of the dentate gyrus which exhibited a marked lack of PARV-IR neurons. No constitutive expression of HSP72 was found in untreated cultures. The lesion of CA3 neurons and the reduction in numbers of PARV-IR neurons in both CA3 and CA1 after chronic exposure to 5 microM kainic acid were similar to those reported to occur in vivo. Exposure to 1 microM doses of kainic acid resulted in a widespread appearance of HSP72-IR and the induction of tolerance to a previously toxic dose of kainic acid. These results suggest the presence of endogenous neuroprotective mechanisms, activated by a stress response which induces HSP72, and is reminiscent of the induced tolerance reported to occur after a mild ischaemic insult.

Animals↗
PubMed1996

d-Sotalol decreases defibrillation energy requirements in humans: a novel indication for drug therapy.

INTRODUCTION: We assessed the effect of d-sotalol on defibrillation voltage and energy requirements in patients undergoing automatic defibrillator implantation. Drugs that primarily prolong cardiac refractoriness generally decrease the energy requirements for defibrillation in animal models. Despite the widespread use of antiarrhythmic drugs in patients with implanted cardioverter defibrillators, the effect of such drugs on defibrillation energy requirements in humans has not been well studied. Sotalol (in the d,l racemic form) is an antiarrhythmic with beta-blocking and cardiac refractoriness prolonging effects. The d-isomer of sotalol is largely devoid of beta-blocking effects; both forms decrease defibrillation energy requirements in animals. We hypothesized that d-sotalol would decrease defibrillation voltage and energy requirements in humans. METHODS AND RESULTS: Fifteen patients undergoing implanted cardioverter defibrillator implantation were studied before and 20 minutes after d-sotalol infusion (2 mg/kg IV in 15 min, followed by 1 mg/kg per hour). The estimated energy (E50) and voltage (V50) for 50% success in defibrillation (estimated from two successive defibrillation "threshold" measurements), ventricular effective refractory period, monophasic action potential duration, and mean cycle length of ventricular fibrillation were measured, along with heart rate, blood pressure, and plasma concentration of d-sotalol. There was a significant decrease in defibrillation energy (E50 = 12.4 +/- 5.0 J before and 8.4 +/- 4.0 J after d-sotalol, P < 0.003) and voltage (V50 = 440 +/- 77 V before and 354 +/- 93 V after d-sotalol, P < 0.001). Consistent with the Class III effect of d-sotalol, ventricular effective refractory period increased from 284 +/- 21 to 330 +/- 24 msec (P < 0.001), and action potential duration was prolonged from 296 +/- 28 to 340 +/- 22 msec (P < 0.001). Following d-sotalol, there was a tendency for induced tachyarrhythmia to self-terminate (23/102 episodes before vs 74/150 after sotalol, P < 0.001), and ventricular fibrillation cycle length was increased from 216 +/- 20 msec before to 274 +/- 23 msec (P < 0.001) after d-sotalol, despite the persistence of a rapid, disorganized rhythm of the surface ECG. No patient suffered adverse effects. CONCLUSIONS: d-Sotalol lowers defibrillation energy by a mean 32% +/- 27% at concentrations producing a 16% +/- 7% increase in ventricular effective refractory period. Along with its other antiarrhythmic effects, d-sotalol may increase the safety margin for defibrillation or allow lower programmed energies in patients with implanted defibrillators.

Adrenergic beta-Antagonists↗