Biomedical subjects

Michael T Ryan

Publications and source records attributed to Michael T Ryan.

At least 19 recordsLinked to original sources

PubMed2007

Factors affecting dose estimates for long-term performance assessments: case study-Armagosa Valley.

The purpose of this study was to review and evaluate some of the factors that influence the dose estimates to members of the public due to chronic, long-term releases of radioactive materials into the environment. Although the examples discussed are based on data from the Amargosa Valley located near the proposed Yucca Mountain high-level radioactive waste repository, the factors evaluated are common to any such assessment. While it is recognized that such factors include those related to both the environmental transport of radionuclides from the point of release through the environmental media to the receptor, and the influence of his/her location and living habits, the assessments that follow are primarily limited to the latter. The specific goal in all cases was to illustrate how the assumptions and input values relative to certain factors influence the dose estimates and to quantify, to the extent possible, their relative significance. At the same time, it must be recognized that the assessments presented here were limited to doses due to the ingestion of food and water; those due to the inhalation of airborne radionuclides and external exposures were not considered. The factor that proved most important from the standpoint of the overestimation of doses (i.e., conservatism) was the implementation of the regulatory requirements pertaining to the withdrawal of groundwater from the local aquifer. Another significant source of conservatism was the dose estimates provided by the U.S. Department of Energy, assuming that the Reasonably Maximally Exposed Individual (RMEI) resided at the U.S. EPA designated site, 18 km from the repository, vs. the Amargosa Valley, about 35 km away. Also important, for selected radionuclides, was the impact of their effective half-lives on the committed doses, as estimated, in comparison to those that would actually be received. Having lesser impacts were the status of a local aquaculture farm on the intake of C, and the intake of stable iodine on dose estimates for I. On the basis of these evaluations, one can reasonably conclude that the overall conservatism in the dose assessments, based on these sources that were identified, approaches an order of magnitude. The sole factor that led to an underestimation of the doses (i.e., non-conservatism) was the regulatory requirement that the concept of the RMEI, as defined by U.S. EPA, in contrast to that of the Critical Group (CG), as recommended by the International Commission on Radiological Protection, be applied in estimating the doses to potentially affected population groups. Rather than dwell on the differences in the impacts of the application of each of these two concepts, the next step should be to subject each concept to a systematic and rigorous analysis, the goal being to gain an understanding of the range of dose estimates that would be yielded, the underlying reasons for the differences that are observed, and the lessons to be learned in terms of improving the methodologies for estimating doses due to environmental radionuclide releases.

Artifacts↗

PubMed2007

Mitochondrial-nuclear communications.

Mitochondria cannot be made de novo but replicate by a mechanism of recruitment of new proteins, which are added to preexisting subcompartments. Although mitochondria have their own DNA, more than 98% of the total protein complement of the organelle is encoded by the nuclear genome. Mitochondrial biogenesis requires a coordination of expression of two genomes and therefore cross talk between the nucleus and mitochondria. In mammals, regulation of mitochondrial biogenesis and proliferation is influenced by external factors, such as nutrients, hormones, temperature, exercise, hypoxia, and aging. This complexity points to the existence of a coordinated and tightly regulated network connecting different pathways. Communications are also required for eliciting mitochondrial responses to specific stress pathways. This review covers the mechanisms of mitochondrial biogenesis and the way cells respond to external signals to maintain mitochondrial function and cellular homeostasis.

Calcium↗

PubMed2006

Mitochondrial respiratory chain supercomplexes are destabilized in Barth Syndrome patients.

Mutations in the human TAZ gene are associated with Barth Syndrome, an often fatal X-linked disorder that presents with cardiomyopathy and neutropenia. The TAZ gene encodes Tafazzin, a putative phospholipid acyltranferase that is involved in the remodeling of cardiolipin, a phospholipid unique to the inner mitochondrial membrane. It has been shown that the disruption of the Tafazzin gene in yeast (Taz1) affects the assembly and stability of respiratory chain Complex IV and its supercomplex forms. However, the implications of these results for Barth Syndrome are restricted due to the additional presence of Complex I in humans that forms a supercomplex with Complexes III and IV. Here, we investigated the effects of Tafazzin, and hence cardiolipin deficiency in lymphoblasts from patients with Barth Syndrome, using blue-native polyacrylamide gel electrophoresis. Digitonin extraction revealed a more labile Complex I/III(2)/IV supercomplex in mitochondria from Barth Syndrome cells, with Complex IV dissociating more readily from the supercomplex. The interaction between Complexes I and III was also less stable, with decreased levels of the Complex I/III(2) supercomplex. Reduction of Complex I holoenzyme levels was observed also in the Barth Syndrome patients, with a corresponding decrease in steady-state subunit levels. We propose that the loss of mature cardiolipin species in Barth Syndrome results in unstable respiratory chain supercomplexes, thereby affecting Complex I biogenesis, respiratory activities and subsequent pathology.

Acyltransferases↗

PubMed2006

Mdm38 interacts with ribosomes and is a component of the mitochondrial protein export machinery.

Saccharomyces cerevisiae Mdm38 and Ylh47 are homologues of human Letm1, a protein implicated in Wolf-Hirschhorn syndrome. We analyzed the function of Mdm38 and Ylh47 in yeast mitochondria to gain insight into the role of Letm1. We find that mdm38Delta mitochondria have reduced amounts of certain mitochondrially encoded proteins and low levels of complex III and IV and accumulate unassembled Atp6 of complex V of the respiratory chain. Mdm38 is especially required for efficient transport of Atp6 and cytochrome b across the inner membrane, whereas Ylh47 plays a minor role in this process. Both Mdm38 and Ylh47 form stable complexes with mitochondrial ribosomes, similar to what has been reported for Oxa1, a central component of the mitochondrial export machinery. Our results indicate that Mdm38 functions as a component of an Oxa1-independent insertion machinery in the inner membrane and that Mdm38 plays a critical role in the biogenesis of the respiratory chain by coupling ribosome function to protein transport across the inner membrane.

Calcium-Binding Proteins↗

PubMed2006

Crystal structure of the mitochondrial chaperone TIM9.10 reveals a six-bladed alpha-propeller.

Import of proteins into mitochondria occurs by coordinated actions of preprotein translocases in the outer and inner membranes. Tim9 and Tim10 are translocase components of the intermembrane space, related to deafness-dystonia peptide 1 (DDP1). They coassemble into a hexamer, TIM9.10, which captures and chaperones precursors of inner membrane metabolite carriers as they exit the TOM channel in the outer membrane. The crystal structure of TIM9.10 reveals a previously undescribed alpha-propeller topology in which helical "blades" radiate from a narrow central pore lined with polar residues. The propeller blades are reminiscent of "tentacles" in chaperones Skp and prefoldin. In each TIM9.10 subunit, a signature "twin CX3C" motif forms two intramolecular disulfides. There is no obvious binding pocket for precursors, which we suggest employ the chaperone-like tentacles of TIM9.10 as surrogate lipid contacts. The first reported crystal structure of a mitochondrial translocase assembly provides insights into selectivity and regulation of precursor import.

Amino Acid Sequence↗

PubMed2006

Significance of 14C and 228Ra in terms of the proposed Yucca Mountain high-level radioactive waste repository.

C and Ra are two of the radionuclides that have either been identified as being potentially significant in terms of releases from the proposed Yucca Mountain high-level radioactive waste repository, or are specifically cited for consideration and evaluation in the regulations promulgated by the U.S. Nuclear Regulatory Commission. The purpose of this study was to estimate the concentrations and associated doses for these two radionuclides, if released under conditions of a scenario assumed to apply to a repository containing some of the features of the one proposed at Yucca Mountain, NV, and to compare these estimates to the regulatory limits for that facility. For C, the postulated condition was that an annual fractional release of 10 of its total remaining inventory occurs beginning at 10,000 y after repository closure. For Ra, the same fractional release rate was assumed, but in this case it was presumed to occur when the Ra inventory was projected to reach a maximum at more than 10 y after repository closure. The estimated concentrations and doses were, in turn, compared to the concentration limit, specified in the Ground Water Protection Standards (GWPSs) in the case of Ra, or derived, in the case of C, on the basis of the regulatory dose rate limit. Due to the small inventory of C in the waste, and its short half-life relative to the performance period evaluated, its estimated concentration in the ground water would be slightly more than 4% of the derived GWPS. Due to the relatively small initial inventory of Th, the precursor of Ra, and the correspondingly small quantities of higher atomic number actinides that could, through decay, produce additional quantities of Th, its estimated concentration in the ground water would be less than 3% of the GWPS, leaving the remaining portion of the limit for potential contributions from Ra. At the same time, however, it must be recognized that, in this case, the regulations require that any contributions of naturally occurring Ra and Ra already present in the ground water must be included in the determination of compliance. If this is done, the total concentration of Ra, combined with the naturally occurring concentration of Ra, would be about 10.5% of the limit. In a similar manner, the committed doses due to the annual consumption of each of these two radionuclides in ground water and food, produced in the local biosphere, were evaluated in terms of the Individual Protection Standard (IPS). Based on these analyses, the estimated effective dose for C, using the coefficients in Federal Guidance Report (FGR) No. 13, was 4.15 muSv y, less than 3% of the IPS. For Ra, the comparable estimate at the time of maximum inventory, excluding in this case the contributions from naturally occurring Ra and Ra, was 7.39 muSv y, representing about 5% of the IPS. Based on the value assumed for the fractional release rate (10 y), it was concluded that neither C nor Ra will be significant in terms of either the applicable GWPS or the IPS. While it was recognized that, due to the time spans involved, these analyses were primarily an academic exercise, it is believed that the perspectives and accompanying insights are useful.

Carbon Radioisotopes↗

PubMed2006

Improving the regulation and management of low-activity radioactive wastes.

This paper summarizes the first phase of a study in progress by a committee of the National Research Council's Board on Radioactive Waste Management. The Board initiated the study after observing that statutes and regulations administered by the federal and state agencies that control low-activity radioactive wastes have developed as a patchwork over almost 60 y. These controls usually reflect the enterprise or process that produced the waste rather than the waste's radiological hazard. Inconsistencies in the regulatory patchwork or its application may have led to overly restrictive controls for some low-activity wastes while others were neglected in comparison. In the first phase of this study, the committee reviewed current low-activity waste inventories, regulations, and management practices. This led the committee to develop five categories that encompass the spectrum of low-activity wastes and serve to illustrate gaps and inconsistencies in current regulations and management practices. The committee completed its first phase with four findings that will lead into the final phase of the study. This paper is excerpted from the committee's interim report that was issued in October 2003.

Decision Making↗

PubMed2006

Mitochondrial morphology and distribution in mammalian cells.

It is now appreciated that mitochondria form tubular networks that adapt to the requirements of the cell by undergoing changes in their shape through fission and fusion. Proper mitochondrial distribution also appears to be required for ATP delivery and calcium regulation, and, in some cases, for cell development. While we now realise the great importance of mitochondria for the cell, we are only beginning to work out how these organelles undergo the drastic morphological changes that are essential for cellular function. Of the few known components involved in shaping mitochondria, some have been found to be essential to life and their gene mutations are linked to neurological disorders, while others appear to be recruited in the activation of cell death pathways. Here we review our current understanding of the functions of the main players involved in mitochondrial fission, fusion and distribution in mammalian cells.

Animals↗

PubMed2005

Essential role of Isd11 in mitochondrial iron-sulfur cluster synthesis on Isu scaffold proteins.

Mitochondria are indispensable for cell viability; however, major mitochondrial functions including citric acid cycle and oxidative phosphorylation are dispensable. Most known essential mitochondrial proteins are involved in preprotein import and assembly, while the only known essential biosynthetic process performed by mitochondria is the biogenesis of iron-sulfur clusters (ISC). The components of the mitochondrial ISC-assembly machinery are derived from the prokaryotic ISC-assembly machinery. We have identified an essential mitochondrial matrix protein, Isd11 (YER048w-a), that is found in eukaryotes only. Isd11 is required for biogenesis of cellular Fe/S proteins and thus is a novel subunit of the mitochondrial ISC-assembly machinery. It forms a complex with the cysteine desulfurase Nfs1 and is required for formation of an Fe/S cluster on the Isu scaffold proteins. We conclude that Isd11 is an indispensable eukaryotic component of the mitochondrial machinery for biogenesis of Fe/S proteins.

Amino Acid Sequence↗

PubMed2005

Dissection of the mitochondrial import and assembly pathway for human Tom40.

Tom40 is the channel-forming subunit of the translocase of the mitochondrial outer membrane (TOM complex), essential for protein import into mitochondria. Tom40 is synthesized in the cytosol and contains information for its mitochondrial targeting and assembly. A number of stable import intermediates have been identified for Tom40 precursors in fungi, the first being an association with the sorting and assembly machinery (SAM) of the outer membrane. By examining the import pathway of human Tom40, we have been able to elucidate additional features in its import. We identify that Hsp90 is involved in delivery of the Tom40 precursor to mitochondria in an ATP-dependent manner. The precursor then forms its first stable intermediate with the outer face of the TOM complex before its membrane integration and assembly. Deletion of an evolutionary conserved region within Tom40 disrupts the TOM complex intermediate and causes it to stall at a new complex in the intermembrane space that we identify to be the mammalian SAM. Unlike its fungal counterparts, the human Tom40 precursor is not found stably arrested at a SAM intermediate. Nevertheless, we show that Tom40 assembly is reduced in mitochondria depleted of human Sam50. These findings are discussed in context with current models from fungal studies.

Adenosine Triphosphate↗

PubMed2005

Sensitivity analyses of the standards for the proposed Yucca Mountain repository--a review, evaluation, and commentary.

The standards and regulations for the proposed Yucca Mountain high level radioactive waste repository, which were developed and promulgated by the U.S. Environmental Protection Agency and the U.S. Nuclear Regulatory Commission, respectively, are complex and challenging. A major reason is that they are divided into three parts, an Individual Protection Standard, a Human Intrusion Standard, and multiple Ground Water Protection Standards. Because the individual parts are not fully integrated, the one that controls under a specific set of circumstances depends on the radionuclide being evaluated, its mechanisms of transport, its avenues of intake, and differences in the specified limits. Although the coefficients in Federal Guidance Report (FGR) No. 11 are being used to estimate the doses, other sources (for example, Title 10, CFR, Part 20, and/or FGR No. 13) may deserve consideration. Since the regulations specify that the reasonably maximally exposed individual is an adult, this leaves unanswered the estimated doses to other age groups, such as infants and adolescents. Summarized in this paper are comparisons of the dose coefficients for different age groups, as well as evaluations of the sensitivity of effective and organ dose estimates for adults, depending on the source of the coefficients. All the latter analyses were based only on the consumption of ground water. While the dose estimates are different, depending on the sources of the coefficients, this was not unexpected. What these evaluations demonstrate is the caution that must be exercised to ensure that a full range of considerations is taken into account in interpreting the outcome of the dose assessments being made with respect to the proposed repository.

Body Burden↗

PubMed2005

Impacts of stable element intake on 14C and 129I dose estimates.

The purpose of this study was to evaluate and provide insights related to the influence of the intake of stable isotopes of carbon and iodine on the committed doses due to the ingestion of (14)C and (129)I. This was accomplished through the application of two different computational approaches. The first was based on the assumption that ground (drinking) water was the only source of intake of (14)C and (129)I, as well as stable carbon and stable iodine. In the second, the intake of (14)C and (129)I was still assumed to be restricted to that in the ground (drinking) water, but the intake of stable carbon and stable iodine was expanded to include that in other components of the diet. The doses were estimated using either a conversion formula or the applicable dose coefficients in Federal Guidance Reports No. 11 and No. 13. Serving as input for the analyses was the estimated maximum concentrations of (14)C or (129)I that would be present in the ground water due to potential releases from the proposed Yucca Mountain high-level radioactive waste repository during the first 10,000 y after closure. The estimated contributions of stable carbon and iodine through the consumption of ground water were based on analyses of samples collected in the Amargosa Valley, NV. The contributions through dietary intake were based on surveys conducted in the United States. Based on the accompanying analyses, it was noted that stable isotope intake has a significant effect on the estimated doses due to the intake of radioactive isotopes of the same element. While this is a well-known fact, this observation has international implications in terms of dose estimates for key radionuclides, such as (14)C and (129)I, a primary reason being the wide variations in the intakes of stable carbon and iodine in various countries. For this reason, analysts planning to apply the dose coefficients developed by the International Commission on Radiological Protection (ICRP) should either confirm that the average total intake in their country of stable isotope(s) of the radioactive isotope being evaluated is in reasonable agreement with the value assumed by the ICRP or suitably modify the ICRP dose coefficients to account for any differences. If such a procedure is to be implemented, there is a need for periodic updates of the dietary intakes of various stable elements in countries throughout the world. The importance of this is documented by recent surveys in Asia that revealed that their average total daily intake of stable iodine was less than half of the ICRP value for Reference Man. In this case, application of the ICRP dose coefficients, without modification, would underestimate the dose due to ingested (129)I by a factor of more than two. A related situation exists in the United States where the latest surveys indicate that the daily intake of stable iodine is 75% of the ICRP value.

Administration, Oral↗

PubMed2004

Biochemical and molecular diagnosis of mitochondrial respiratory chain disorders.

Biochemical diagnosis of mitochondrial respiratory chain disorders requires caution to avoid misdiagnosis of secondary enzyme defects, and can be improved by the use of conservative diagnostic criteria. Pathogenic mutations causing mitochondrial disorders have now been identified in more than 30 mitochondrial DNA (mtDNA) genes encoding respiratory chain subunits, ribosomal- and t-RNAs. mtDNA mutations appear to be responsible for most adult patients with mitochondrial disease and approximately a quarter of paediatric patients. A family history suggesting maternal inheritance is the exception rather than the norm for children with mtDNA mutations, many of whom have de novo mutations. Prenatal diagnosis and pre-implantation genetic diagnosis can be offered to some women at risk of transmitting a mtDNA mutation, particularly those at lower recurrence risk. Mutations in more than 30 nuclear genes, including those encoding for respiratory chain subunits and assembly factors, have now been shown to cause mitochondrial disorders, creating difficulties in prioritising which genes should be studied by mutation analysis in individual patients. A number of approaches offer promise to guide the choice of candidate genes, including Blue Native-PAGE immunoblotting and microarray expression analysis.

Biochemistry↗

PubMed2004

Chaperones: inserting beta barrels into membranes.

How beta-barrel proteins are inserted into cellular membranes is poorly understood. New work has identified a sorting and assembly machinery that chaperones beta-barrels into the mitochondrial outer membrane and is evolutionarily conserved from bacteria to man.

Membrane Transport Proteins↗

PubMed2004

Levels of human Fis1 at the mitochondrial outer membrane regulate mitochondrial morphology.

Mitochondria undergo balanced fission and fusion events that enable their appropriate networking within the cell. In yeast, three factors have been identified that co-ordinate fission events at the mitochondrial outer membrane. Fis1p acts as the outer membrane receptor for recruitment of the dynamin member, Dnm1p and the WD40-repeat-containing protein Mdv1p. In mammals, the Dnm1p counterpart Drp1 has been characterized, but other components have not. Here, we report the characterization of human Fis1 (hFis1). hFis1 is inserted into the mitochondrial outer membrane via a C-terminal transmembrane domain that, along with a short basic segment, is essential for its targeting. Although expression of hFis1 does not complement the phenotype of yeast cells lacking Fis1p, overexpression of hFis1 in tissue culture cells nevertheless causes mitochondrial fragmentation and aggregation. This aggregation could be suppressed by expressing a dominant-negative Drp1 mutant (Drp1(K38A)). Knockdown of hFis1 in COS-7 cells using RNA interference results in mitochondrial morphology defects with notable extensions in the length of mitochondrial tubules. These results indicate that the levels of hFis1 at the mitochondrial surface influences mitochondrial fission events and hence overall mitochondrial morphology within the cell.

Amino Acid Sequence↗

PubMed2004

De novo mutations in the mitochondrial ND3 gene as a cause of infantile mitochondrial encephalopathy and complex I deficiency.

Both nuclear and mitochondrial DNA mutations can cause energy generation disorders. Respiratory chain complex I deficiency is the most common energy generation disorder and a frequent cause of infantile mitochondrial encephalopathies such as Leigh's disease and lethal infantile mitochondrial disease. Most such cases have been assumed to be caused by nuclear gene defects, but recently an increasing number have been shown to be caused by mutations in the mitochondrially encoded complex I subunit genes ND4, ND5, and ND6. We report the first four cases of infantile mitochondrial encephalopathies caused by mutations in the ND3 subunit gene. Three unrelated children have the same novel heteroplasmic mutation (T10158C), only the second mutation reported in ND3, and one has the previously identified T10191C mutation. Both mutations cause disproportionately greater reductions in enzyme activity than in the amount of fully assembled complex I, suggesting the ND3 subunit plays an unknown but important role in electron transport, proton pumping, or ubiquinone binding. Three cases appear to have a de novo mutation, with no mutation detected in maternal relatives. Mitochondrial DNA disease may be considerably more prevalent in the pediatric population than currently predicted and should be considered in patients with infantile mitochondrial encephalopathies and complex I deficiency.

Blotting, Western↗

PubMed2004

Characterization of presenilin complexes from mouse and human brain using Blue Native gel electrophoresis reveals high expression in embryonic brain and minimal change in complex mobility with pathogenic presenilin mutations.

The presenilin proteins are required for intramembrane cleavage of a subset of type 1 membrane proteins including the Alzheimer's disease amyloid precursor protein. Previous studies indicate presenilin proteins form enzymatically active high molecular mass complexes consisting of heterodimers of N- and C-terminal fragments in association with nicastrin, presenilin enhancer-2 and anterior pharynx defective-1 proteins. Using Blue Native gel electrophoresis (BN/PAGE) we have studied endogenous presenilin 1 complex mass, stability and association with nicastrin, presenilin enhancer-2 and anterior pharynx defective-1. Solubilization of mouse or human brain membranes with dodecyl-d-maltoside produced a 360-kDa species reactive with antibodies to presenilin 1. Presenilin 1 complex levels were high in embryonic brain. Complex integrity was sensitive to Triton X-100 and SDS, but stable to reducing agent. Addition of 5 M urea caused complex dissolution and nicastrin to migrate as a subcomplex. Nicastrin and presenilin enhancer-2 were detected in the presenilin 1 complex following BN/PAGE, electroelution and second-dimension analysis. Anterior pharynx defective-1 was detected as an 18-kDa form and 9-kDa C-terminal fragment by standard SDS/PAGE of mouse tissues, and as a predominant 36-kDa band after presenilin 1 complex second-dimension analysis. Membranes from brain cortex of Alzheimer's disease patients, or from cases with presenilin 1 missense mutations, indicated no change in presenilin 1 complex mobility. Higher molecular mass presenilin 1-reactive species were detected in brain containing presenilin 1 exon 9 deletion mutation. This abnormality was confirmed using cells transfected with the same presenilin deletion mutation.

Alzheimer Disease↗

PubMed2004

Limitations on upper bound dose to adults due to intake of 129I in drinking water and a total diet-implications relative to the proposed Yucca Mountain high level radioactive waste repository.

The purpose of this report is to comment on the potential annual doses due to the intake by adults of I, an important radionuclide in the proposed high-level radioactive waste repository at Yucca Mountain. An often overlooked, but significant, factor is that, in this case, the ground water, which would be the primary transport vehicle for any releases, contains relatively high concentrations of stable iodine (127I); in fact, the median concentration in the ground water in the vicinity of the proposed repository is 5.0 microg L-1. In comparison, the maximum concentration of 129I in the ground water, due to potential releases of 129I during the first 10,000 y following closure of the repository, is estimated to be approximately 3.7 x 10(-7) Bq L-1 (approximately 10(-5) pCi L-1). This would result in a 127I to 129I ratio in the water of almost 90 million to one. Assuming no other sources of these two isotopes were being consumed, this would place an upper bound on the annual committed thyroid dose of 1.2 x 10(-1) mSv (1.2 x 10(-1) mrem), less than one thousandth of the Ground Water Protection Standard of 4 mrem y-1. When the additional intake of stable and radioactive iodine in other components of the diet is considered, the overall ratio of 127I to 129I would be more than 2 billion to one. The would place an upper bound on the annual committed effective dose of approximately 2.5 x 10(-8) mSv (approximately 2.5 x 10(-6) mrem), less than one millionth of the Individual Protection Standard of 0.15 mSv (15 mrem).

Adult↗