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N Dean

Publications and source records attributed to N Dean.

50 records · Page 3Linked to original sources
PubMed1994

Factors influencing the outcome of in-vitro fertilization with epididymal spermatozoa in irreversible obstructive azoospermia.

Microsurgical epididymal sperm aspiration (MESA) and in-vitro fertilization (IVF) were found to offer limited opportunity for fatherhood to 45 men with obstructive azoospermia, due principally to poor embryo implantation. Adequate sperm preparations were obtained in 46/50 treatment cycles (92%), with the best motility found in the caput epididymis in 89% of cases. The mean fertilization rate was 11.2% and fertilization occurred in 23 cycles (50%), with embryo transfer arising from 12/26 men with vas aplasia (CAV), 4/9 with genital tract obstruction (EV) and 7/11 with irreversible vasectomy (VV). The overall implantation rate was low, 8.7% per embryo transfer (11.7% per 2-3 embryo transfers) and was not improved by Fallopian transfer. There were two pregnancies (4% per cycle), both in the EV group where embryo formation and implantation (2/4, 50% per cycle) were optimum even though sperm preparations were paradoxically inferior to the CAV and VV groups. The spermatozoa retrieved in the two successful EV cycles were appreciably blood contaminated. Analysis of the 21 failed embryo transfers showed delayed fertilization in 10 cycles, cystic fibrosis (CF) mutation or familial disease in 7/12 CAV men and the VV men were older (P < 0.001). A pregnancy which miscarried arose from a case of Young's syndrome, a carrier of CF mutation DF508. Male factors could thus be implicated in the high embryo wastage of MESA cycles and might also be influencing implantation in other IVF procedures. Where feasible, male reconstructive surgery is preferable unless fertilization can be improved, possibly by speedier retrieval techniques or by permitting sperm capacitation in vitro, but probably more effectively by micro-assisted insemination.

Adult↗
PubMed1994

Ethical implications of standardization of ICU care with computerized protocols.

Ethical issues related to the use of computerized protocols to control mechanical ventilation of patients with Acute Respiratory Distress Syndrome (ARDS) are identical to the ethical issues surrounding the use of any therapy or intervention. Four ethical principles must be considered: nonmaleficence, beneficence, autonomy, and distributed justice. The major ethical challenges to computerized protocol use as a specific application of clinical decision support tools are found within the principles of nonmaleficence and of beneficence. The absence of credible outcome data on which ARDS patient survival probabilities with different therapeutic options could be based is a constraint common to most ICU clinical decision making. Clinicians are thus deprived of the knowledge necessary to define benefit and are limited to beneficent intention in clinical decisions. Computerized protocol controlled decision making for the clinical management of mechanical ventilation for ARDS patients is ethically defensible. It is as well supported as most ICU therapy options.

Clinical Protocols↗
PubMed1993

Potential health benefits of nutrition label changes.

OBJECTIVES: The Nutrition Labeling and Education Act of 1990 mandates the Food and Drug Administration to promulgate changes in nutrition labeling regulations. This study investigates the potential health benefits associated with expected changes in food consumption resulting from the act. METHODS: This paper provides four estimates of the potential health benefits from the dietary changes expected to occur as a result of the 1990 act. The upper bound estimates begin with the premise that all consumers will adopt the daily reference values of total fat, saturated fat, and cholesterol. The lower bound estimate is based on consumers' responses to a shelf-labeling program sponsored by the Food and Drug Administration in the 1980s. A computer model developed by Dr. Warren Browner and his associates was used to estimate the health benefits from reduced nutrient intakes. RESULTS: Estimates of the number of discounted life-years gained nationwide for the first 20 years after the implementation of the act range from a high of 1.2 million to a low of 40,000. CONCLUSIONS: The results of the study highlight that relatively small changes in nutrient intakes may generate large public health benefits.

Adult↗
PubMed1990

ERD2, a yeast gene required for the receptor-mediated retrieval of luminal ER proteins from the secretory pathway.

Resident proteins of the ER lumen carry a specific tetrapeptide signal (KDEL or HDEL) that prevents their secretion. We have previously described the isolation of yeast mutants that fail to retain such resident proteins within the cell. Here we describe ERD2, a gene required for retention. It encodes a 26 kd integral membrane protein whose abundance determines the efficiency and capacity of the retention system. Reduced expression of ERD2 leads to secretion of proteins bearing the HDEL signal, whereas overexpression of ERD2 improves retention both in wild-type cells and in other mutants. These results are consistent with other evidence that ERD2 encodes the HDEL receptor (see accompanying paper). The gene is also required, perhaps indirectly, for normal protein transport through the Golgi, and hence for growth. We discuss possible roles for ERD2 in the secretory pathway.

Amino Acid Sequence↗
PubMed1990

ERD1, a yeast gene required for the retention of luminal endoplasmic reticulum proteins, affects glycoprotein processing in the Golgi apparatus.

We have previously shown that the C-terminal sequence HDEL acts as a retention signal for luminal endoplasmic reticulum (ER) proteins in Saccharomyces cerevisiae, and that it is possible to isolate mutants that fail to retain an invertase fusion protein bearing this signal. Analysis of many such mutants defines two genes, ERD1 and ERD2. Cells lacking the ERD1 gene secrete the endogenous ER protein, BiP. Under normal growth conditions, the rate of secretion is equivalent to the rate at which wild-type cells secrete a modified form of BiP that lacks the HDEL signal altogether. Thus, erd1 cells show a profound disruption of the retention system. The mutant cells have no gross abnormality of their intracellular membrane system, but show defects in the Golgi-dependent modification of glycoproteins. We suggest that sorting of luminal ER proteins normally occurs in the Golgi, and that the function of ERD1 is required for the correct interaction of an HDEL receptor with its ligands. The sequence of ERD1 predicts a membrane protein with several transmembrane domains, a conclusion supported by analysis of ERD1-SUC2 fusion proteins.

Amino Acid Sequence↗
PubMed1990

Recycling of proteins from the Golgi compartment to the ER in yeast.

In the yeast Saccharomyces cerevisiae, the carboxyl terminal sequence His-Asp-Glu-Leu (HDEL) has been shown to function as an ER retention sequence (Pelham, H. R. B., K. G. Hardwick, and M. J. Lewis. 1988. EMBO (Eur. Mol. Biol. Organ.) J. 7:1757-1762). To examine the mechanism of retention of soluble ER proteins in yeast, we have analyzed the expression of a preproalpha factor fusion protein, tagged at the carboxyl terminus with the HDEL sequence. We demonstrate that this fusion protein, expressed in vivo, accumulates intracellularly as a precursor containing both ER and Golgi-specific oligosaccharide modifications. The Golgi-specific carbohydrate modification, which occurs in a SEC18-dependent manner, consists of alpha 1-6 mannose linkages, with no detectable alpha 1-3 mannose additions, indicating that the transit of the HDEL-tagged fusion protein is confined to an early Golgi compartment. Results obtained from the fractionation of subcellular organelles from yeast expressing HDEL-tagged fusion proteins suggest that the Golgi-modified species are present in the ER. Overexpression of HDEL-tagged preproalpha factor results in the secretion of an endogenous HDEL-containing protein, demonstrating that the HDEL recognition system can be saturated. These results support the model in which the retention of these proteins in the ER is dependent on their receptor-mediated recycling from the Golgi complex back to the ER.

Amino Acid Sequence↗
PubMed1988

Ordering promoter binding of class III transcription factors TFIIIC1 and TFIIIC2.

The separation of the mammalian class III transcription factor TFIIIC into two functional components, termed TFIIIC1 and TFIIIC2, enabled an analysis of their functions in transcription initiation. Template competition assays were used to define the order with which these factors interact in vitro to form stable preinitiation complexes on the adenovirus VAI and Drosophila melanogaster tRNA(Arg) genes. The interaction between these genes and TFIIIC2, the factor that binds with high affinity to the B block, was both necessary and sufficient for template commitment. When either the VAI or tRNA(Arg) gene was preincubated with TFIIIC2 alone, transcription of a second gene added subsequently was excluded, indicating that TFIIIC2 bound stably to the first template. Furthermore, the interaction between TFIIIC2 and these genes must occur prior to that of TFIIIC1 or TFIIIB. Once TFIIIC2 was bound, TFIIIC1 could bind to the tRNA(Arg) and VAI genes, although its interaction with the VAI gene was less stable than that with the tRNA(Arg) gene. TFIIIB activity bound stably to the complex of both genes with TFIIIC2. These results demonstrate that TFIIIC2 is the first transcription factor to bind to these genes and that TFIIIB and TFIIIC1 can then interact in either order to form a preinitiation complex.

Adenoviridae↗
PubMed1987

Separation of TFIIIC into two functional components by sequence specific DNA affinity chromatography.

Recently, it has been shown that mammalian transcription factor IIIC (TFIIIC) activity can be separated by anion exchange FPLC chromatography into two functional components (1), both of which are required for transcription of tRNA and the adenovirus VA RNA genes. Here we show that these two functional components, designated TFIIIC1 and TFIIIC2, can also be separated by sequence specific DNA affinity chromatography. These results confirm the observation that TFIIIC can be fractionated into two components, which are both required for transcription of VA I and tRNA genes in vitro. Thus in the mammalian reconstituted system, a minimum of three proteins, in addition to RNA polymerase III, are required for the transcription of the VA and tRNA genes in vitro. The DNA binding component, TFIIIC2, binds specifically to the 3' segment of the internal promoter (the B block), demonstrated by its ability to protect this region from digestion by DNase I. TFIIIC2 is the limiting, titratable component in the phosphocellulose C fraction required for the formation of a stable pre-initiation complex on the VAI RNA gene in vitro, as demonstrated with a template competition and rescue assay.

Base Sequence↗
PubMed1986

Adenovirus stimulation of transcription by RNA polymerase III: evidence for an E1A-dependent increase in transcription factor IIIC concentration.

Human cells expressing adenovirus E1A proteins transcribe transfected tRNA and adenovirus VAI genes at greater than 10-fold higher levels than uninfected HeLa cells. Here we show that the increased transcription observed in vivo is reflected in the in vitro transcriptional activity of cell extracts. Depletion of E1A protein from these extracts by immunoprecipitation with a monoclonal antibody did not diminish the activity, suggesting that E1A proteins do not stimulate transcription directly. Fractionation of the extracts by chromatography on phosphocellulose suggests that the higher activity of extracts of adenovirus-infected cells was due to increased activity of the transcription factor (TF) which is the limiting component required for specific initiation of tRNA and VAI transcription in extracts of uninfected HeLa cells, i.e. TFIIIC. Template commitment titrations further suggest that the increased TFIIIC activity was due to an increase in the concentration of active TFIIIC. On the basis of these results and recent genetic analyses of early adenovirus promoters, we suggest that E1A proteins stimulate transcription of adenovirus genes indirectly by increasing the effective in vivo concentration of the limiting cellular transcription factors required for their transcription.

Adenovirus Early Proteins↗
PubMed1986

Oxidation of reduced menaquinone by the fumarate reductase complex in Escherichia coli requires the hydrophobic FrdD peptide.

Plasmids carrying cloned segments of the frd operon of Escherichia coli have been used in genetic complementation studies to identify two independent mutants defective in the frdD gene, which encodes the hydrophobic FrdD polypeptide of the fumarate reductase complex. Mutations in the frdA and frdB genes have also been mapped by this technique. One of the FrdD peptide mutants, DW109 (frdD-109), showed that fumarate reductase was not as tightly bound to the membrane in this mutant. In addition, the mutation in the FrdD peptide caused an almost total loss of the ability of the enzyme to oxidize either menaquinol-6, a physiological donor for fumarate reduction, or reduced benzyl viologen. However, the mutation did not impair the ability of the membrane-bound fumarate reductase complex to function with succinate as substrate, as evidenced by unchanged turnover numbers for phenazine methosulfate and 2,3-dimethoxy-5-methyl-6-pentyl-1,4-benzoquinone (a quinone analogue) reductase activities. These data establish the essential role of the FrdD polypeptide both in the interaction of the enzyme with reduced menaquinone and thus in anaerobic respiration with fumarate as electron acceptor, and in binding the enzyme to the membrane.

Benzoquinones↗