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Biomedical subjects

H C Smith

Publications and source records attributed to H C Smith.

At least 19 recordsLinked to original sources
PubMed2001

Ethanol stimulates apolipoprotein B mRNA editing in the absence of de novo RNA or protein synthesis.

Apolipoprotein B (apoB) mRNA editing involves a site-specific modification of cytidine to form uridine. The reaction is catalyzed in the nucleus by a multi-protein editosome. Rat hepatic editing is regulated during development, metabolically and in response to ethanol. Ethanol stimulated editing in hepatocytes within minutes of exposure. In the present study, we show that ethanol stimulated apoB mRNA synthesis and apoB mRNA editing. Significantly, the proportion of edited apoB mRNA also increased following ethanol treatment of transcription or translation arrested cells. These data suggested that ethanol could regulate editing activity using pre-existing editosomal proteins. In addition, the presence of a suppressor of apoB mRNA editing activity was suggested by the finding that inhibition of mRNA or protein synthesis alone was sufficient to increase the proportion of edited RNA. It is proposed that the level of editing activity observed in hepatocytes may be the end result of positive and negative regulatory proteins.

Animals↗
PubMed2001

Commitment of apolipoprotein B RNA to the splicing pathway regulates cytidine-to-uridine editing-site utilization.

A tripartite motif located in the centre of the 7.5 kb exon 26 of apolipoprotein B (apoB) mRNA directs editosome assembly and site-specific cytidine-to-uridine editing at nucleotide 6666. apoB mRNA editing is a post-transcriptional event, occurring primarily at the time exon 26 is spliced or at a time after splicing, but before nuclear export. We show, through reporter RNA constructs, that RNA splice sites suppress editing of precursor RNAs when placed proximal or distal to the editing site. Processed RNAs were edited more efficiently than precursor RNAs. Mutation of both the splice donor and acceptor sites was necessary for RNAs to be edited efficiently. The results suggested that commitment of pre-mRNA to the splicing and/or nuclear-export pathways may play a role in regulating editing-site utilization. The HIV-1 Rev-Rev response element ('RRE') interaction was utilized to uncouple the commitment of precursor RNAs to the spliceosome assembly pathway and associated nuclear-export pathway. Under these conditions, unspliced reporter RNAs were edited efficiently. We propose that pre-mRNA passage through the temporal or spatial restriction point where they become committed to spliceosome assembly contributes regulatory information for subsequent editosome activity.

Animals↗
PubMed2001

Intracellular trafficking determinants in APOBEC-1, the catalytic subunit for cytidine to uridine editing of apolipoprotein B mRNA.

The posttranscriptional deamination editing of apolipoprotein B (apoB) mRNA catalyzed by APOBEC-1 (apoB mRNA editing catalytic subunit 1) is a nuclear process. The signals in APOBEC-1 responsible for its dual cytoplasmic/nuclear distribution have been evaluated. Residues 97-172 in the middle of APOBEC-1 together with its N-terminal 56 residues affect nuclear localization. Mutagenesis studies however revealed no discrete nuclear localization signal in APOBEC-1. Fifteen amino acids (Leu 173-Leu 187) within the previously identified C-terminal domain of APOBEC-1 were sufficient as a determinant for cytoplasmic distribution in that context. These residues failed to demonstrate nuclear export function in a reporter assay. Further, the distribution of APOBEC-1 in the cytoplasm did not respond to leptomycin B, suggesting that APOBEC-1 did not have nuclear export activity. The data suggested that there are at least three regions in APOBEC-1 that participate in its distribution in both the nucleus and the cytoplasm of editing competent cells; however, none of these meet the functional criteria of nuclear localization or nuclear export signals. The findings are discussed in terms of their implications in the regulation of nuclear editing activity and the possibility that interactions with chaperones may play a role in the cellular distribution of APOBEC-1.

APOBEC-1 Deaminase↗
PubMed2001

Identification of the yeast cytidine deaminase CDD1 as an orphan C-->U RNA editase.

Yeast co-expressing rat APOBEC-1 and a fragment of human apolipoprotein B (apoB) mRNA assembled functional editosomes and deaminated C6666 to U in a mooring sequence-dependent fashion. The occurrence of APOBEC-1-complementing proteins suggested a naturally occurring mRNA editing mechanism in yeast. Previously, a hidden Markov model identified seven yeast genes encoding proteins possessing putative zinc-dependent deaminase motifs. Here, only CDD1, a cytidine deaminase, is shown to have the capacity to carry out C-->U editing on a reporter mRNA. This is only the second report of a cytidine deaminase that can use mRNA as a substrate. CDD1-dependent editing was growth phase regulated and demonstrated mooring sequence-dependent editing activity. Candidate yeast mRNA substrates were identified based on their homology with the mooring sequence-containing tripartite motif at the editing site of apoB mRNA and their ability to be edited by ectopically expressed APOBEC-1. Naturally occurring yeast mRNAs edited to a significant extent by CDD1 were, however, not detected. We propose that CDD1 be designated an orphan C-->U editase until its native RNA substrate, if any, can be identified and that it be added to the CDAR (cytidine deaminase acting on RNA) family of editing enzymes.

APOBEC-1 Deaminase↗
PubMed2000

Induction of cytidine to uridine editing on cytoplasmic apolipoprotein B mRNA by overexpressing APOBEC-1.

Post-transcriptional editing of apolipoprotein B (apoB) mRNA is regulated in hepatic cells to achieve a steady state proportion of edited and unedited RNA molecules. This activity is catalyzed by APOBEC-1 (apoB mRNA editing catalytic subunit 1) in what has been widely accepted as nuclear event occurring during or after mRNA splicing. Introns impair the efficiency of editing within an adjacent exon in a distance-dependent manner in reporter RNAs. We show here that this inhibition can be overcome by overexpressing APOBEC-1 and that the enhanced editing efficiency on these reporter RNAs occurred after splicing on cytoplasmic transcripts. Given the absolute requirement of auxiliary proteins in apoB mRNA editing, the data suggested that auxiliary proteins were distributed with APOBEC-1 in both the nucleus and cytoplasm of McArdle cells. In fact, immunolocalization of one such auxiliary protein, APOBEC-1 complementation factor (ACF) demonstrated a nuclear and cytoplasmic distribution. We also demonstrate that in the absence of alterations in APOBEC-1 expression, changes in edited apoB RNA induced by ethanol arise through the stimulation of nuclear editing activity. The finding that apoB mRNA editing can occur in the cytoplasm but normally does not suggests that under biological conditions, restricting editing activity to the nucleus must be an important step in regulating the proportion of the edited apoB mRNAs.

APOBEC-1 Deaminase↗
PubMed2000

Quality of life after coronary revascularization in the United States and Canada.

Cardiac procedures are performed less frequently in Canada than in the United States (US), yet rates of cardiac death and myocardial infarction are similar. We therefore sought to compare long-term symptoms and quality of life in Canadian and American patients undergoing initial coronary revascularization. The 161 patients enrolled in the Bypass Angioplasty Revascularization Investigation at the Montreal Heart Institute were compared with 934 patients enrolled at 7 US sites. Patients' outcomes were documented for 5 years after random assignment to percutaneous transluminal coronary angioplasty or coronary artery bypass graft surgery. Functional status was assessed using the Duke Activity Status Index. Canadian patients were significantly younger and had more angina at study entry. Death and nonfatal myocardial infarction were not significantly different between Canadian and US patients after adjustment for baseline risk. Canadian patients had significantly greater improvements in functional status at 1-year follow-up (Duke Activity Status Index score + 13.5 vs. + 6.0, p = 0.002), but this difference progressively narrowed over 5 years. Angina was equally prevalent in Canadian and US patients at 1 year (16% vs. 19%), but significantly more prevalent in Canadian patients at 5 years (36% vs. 16%, p = 0.001). Repeat revascularization procedures were performed less often over 5 years among Canadian patients (26% vs. 34%, p = 0.08), especially coronary artery bypass graft surgery after initial percutaneous transluminal coronary angioplasty (18% vs. 32%, p = 0.03). These results suggest more anginal symptoms are required in Canada before coronary revascularization, but as a result Canadians receive greater improvements in quality of life after the procedure.

Angina Pectoris↗
PubMed2000

APOBEC-1 dependent cytidine to uridine editing of apolipoprotein B RNA in yeast.

Cytidine to uridine editing of apolipoprotein B (apoB) mRNA requires the cytidine deaminase APOBEC-1 as well as a tripartite sequence motif flanking a target cytidine in apoB mRNA and an undefined number of auxiliary proteins that mediate RNA recognition and determine site-specific editing. Yeast engineered to express APOBEC-1 and apoB mRNA supported editing under conditions of late log phase growth and stationary phase. The cis -acting sequence requirements and the intracellular distribution of APOBEC-1 in yeast were similar to those described in mammalian cells. These findings suggest that auxiliary protein functions necessary for the assembly of editing complexes, or 'editosomes', are expressed in yeast and that the distribution of editing activity is to the cell nucleus.

APOBEC-1 Deaminase↗
PubMed2000

Corticospinal function studied over time following incomplete spinal cord injury.

STUDY DESIGN: Longitudinal. OBJECTIVES: (1) To perform standard clinical neurological examinations and establish the pattern of clinical change with time following incomplete spinal cord injury (iSCI). (2) To establish the pattern of change in corticospinal electrophysiological function with time after iSCI. (3) To correlate clinical with electrophysiological findings. SETTING: The National Spinal Injuries Centre, Stoke Mandeville Hospital, Aylesbury, UK and Imperial College School of Medicine, Charing Cross Hospital, London, UK. METHODS: Neurological assessments and classification were performed according to American Spinal Injuries Association and International Medical Society of Paraplegia (ASIA/IMSOP) standards. Twenty-one patients (ages 18 - 72 years) with iSCI (level C2 - C7, ASIA impairment grades C - D) and 10 healthy control subjects (ages 27 - 57 years) were studied. Electrophysiological tests of corticospinal function were carried out using transcranial magnetic stimulation (TMS) of the motor cortex and electromyographic (EMG) recordings from thenar muscles. Both tests were performed on a number of occasions, beginning 19 - 384 days and ending 124 - 1109 days post-injury, and the group data were pooled into time epochs of 50 or 100 days post-injury for analysis. Seven of the patients were studied on seven or more occasions and were also assessed individually. RESULTS: Individual and pooled data indicated that neurological scores improved progressively and tended to stabilise by around 300 days post-injury. When the patients were first assessed, the mean latency for motor evoked potentials (MEPs) and inhibition of voluntary EMG were significantly different from control values. There was no significant change in latency on subsequent sessions for either the grouped or individual patient data. There was no correlation between clinical assessment and electrophysiological data. CONCLUSION: We conclude that the weakened inhibition seen following iSCI is established within a few days of the time of spinal cord trauma. We argue that reduced corticospinal inhibition may be a prerequisite for the recovery of useful motor function. SPONSORSHIP: The work was supported by a project grant from The Wellcome Trust.

Adolescent↗
PubMed2000

First evidence of premature ventricular complex-induced cardiomyopathy: a potentially reversible cause of heart failure.

Tachycardia-induced cardiomyopathy is a well-recognized and reversible condition, but left ventricular dysfunction due to frequent isolated premature ventricular complexes (PVCs) has not been reported. We observed resolution of dilated cardiomyopathy in a patient after a focal source of PVCs was eliminated by radiofrequency ablation. In a subset of patients with heart failure, PVC-induced cardiomyopathy may be a potentially reversible cause of left ventricular dysfunction.

Adult↗
PubMed2000

Modulation of single motor unit discharges using magnetic stimulation of the motor cortex in incomplete spinal cord injury.

OBJECTIVES: Motor evoked potentials (MEPs) and inhibition of voluntary contraction to transcranial magnetic stimulation (TMS) of the motor cortex have longer latencies than normal in patients with incomplete spinal cord injury (iSCI) when assessed using surface EMG. This study now examines the modulation of single motor unit discharges to TMS with the aim of improving resolution of the excitatory and inhibitory responses seen previously in surface EMG recordings. METHODS: A group of five patients with iSCI (motor level C4-C7) was compared with a group of five healthy control subjects. Single motor unit discharges were recorded with concentric needle electrodes from the first dorsal interosseus muscle during weak voluntary contraction (2%-5% maximum). TMS was applied with a 9 cm circular stimulating coil centred over the vertex. Modulation of single motor unit discharges was assessed using peristimulus time histograms (PSTHs). RESULTS: Mean (SEM) threshold (expressed as percentage of maximum stimulator output (%MSO)) for the excitatory peak (excitation) or inhibitory trough (inhibition) in the PSTHs was higher (p<0.05) in the patients (excitation = 47.1 (5.9) %MSO; inhibition = 44.3 (3.2) %MSO) than in controls (excitation=31.6 (1.2) %MSO; inhibition = 27.4 (1.0) %MSO). Mean latencies of excitation and inhibition were longer (p<0.05) in the patients (excitation=35 (1.8) ms; inhibition = 47.1 (1.8) ms) than in the controls (excitation = 21.1 (1.6) ms; inhibition = 27 (0.4) ms). Furthermore, the latency difference (inhibition-excitation) was longer (p<0.05) in the patients (10.4 (2.1) ms) than in the controls (6.2 (0.6) ms). CONCLUSION: Increased thresholds and latencies of excitation and inhibition may reflect degraded corticospinal transmission in the spinal cord. However, the relatively greater increase in the latency of inhibition compared with excitation in the patients with iSCI may reflect a weak or absent early component of cortical inhibition. Such a change in cortical inhibition may relate to the restoration of useful motor function after iSCI.

Adult↗
PubMed1999

Disproportionate relationship between APOBEC-1 expression and apolipoprotein B mRNA editing activity.

Apolipoprotein B (apoB) mRNA editing is a site-specific (nucleotide 6666) cytidine to uridine transition catalyzed by a cytidine deaminase, APOBEC-1, in the context of a multiprotein complex referred to as the C/U editosome. This report quantifies for the first time the effect of altering APOBEC-1 protein abundance on the proportion of edited apoB mRNAs using transfected McArdle rat hepatoma cells which had been sorted by flow cytometry into populations expressing different levels of green fluorescent protein-APOBEC-1 chimera, GFP-APOBEC. A correlation was observed in which increased expression of GFP-APOBEC protein resulted in a higher proportion of edited apoB mRNA. The number of enzyme molecules required to increase the proportion of edited apoB RNAs was disproportionately high relative to that which might have been predicted from a typical catalytic relationship. Moreover, editing of apoB mRNA at inappropriate sites (promiscuous editing) occurred in response to overexpressing GFP-APOBEC. The data suggest that experimental manipulation of APOBEC-1 abundance in the absence of other regulatory considerations will always result in some level of promiscuous editing. Coordinate expression of APOBEC-1 and the auxiliary proteins and/or regulation of their interactions may be required to increase editing activity without losing editing-site fidelity.

APOBEC-1 Deaminase↗
PubMed1999

Tissue-specific differences in the role of RNA 3' of the apolipoprotein B mRNA mooring sequence in editosome assembly.

Site-specific editing of apolipoprotein B (apoB) mRNA by the cytidine deaminase, APOBEC-1 is proposed to require interactions of auxiliary protein(s) with an eleven nucleotide element, the mooring sequence, located 3' of the C --> U editing site. An analysis of the RNA sequence dependence for protein-RNA interactions and editosome assembly in rat liver and the small intestine demonstrated that the mooring sequence was a minimal requirement for these interactions. Sequences 3' of the mooring sequence either interacted with 66 kDa and 44 kDa proteins or enhanced the interactions of these proteins with the mooring sequence. The data also suggested tissue-specific differences in the relative importance of the 3' cis-acting 'enhancer' elements in the efficiency or stability of editosome assembly. We propose that the previously demonstrated differences in apoB mRNA editing efficiency and its regulation in liver and intestine may in part be due to differences in auxiliary protein interactions with apoB mRNA 3' of the mooring sequence.

Animals↗
PubMed1999

Apolipoprotein B mRNA and lipoprotein secretion are increased in McArdle RH-7777 cells by expression of betaine-homocysteine S-methyltransferase.

The cDNA encoding rat betaine-homocysteine S-methyltransferase (BHMT) was isolated through production of monoclonal antibodies against protein fractions enriched with apolipoprotein B (apo B)-mRNA-editing complexes. BHMT mRNA was expressed predominantly in liver, and also in kidney, but not in small intestine. In stable McArdle RH-7777 (McA) cell lines expressing differing levels of BHMT, the editing efficiency of apo B mRNA was unchanged. Evaluation of apo B-mRNA expression revealed that steady-state levels were increased significantly and in parallel with BHMT protein expression. The highest levels of BHMT mRNA and BHMT enzyme activity expressed in stably transfected McA cells were comparable with those found in rat hepatocytes. In contrast to the changes in apo B-mRNA abundance, levels of other apolipoprotein-encoding mRNAs and several liver-specific and ubiquitously expressed mRNAs were unchanged by BHMT expression. In the cell line expressing the highest level of BHMT, apo B-containing lipoprotein secretion was increased, indicating utilization of increased endogenous message. Results suggest that apo B-mRNA abundance in McA cells is related to the expression of BHMT, an enzyme important in homocysteine metabolism.

Amino Acid Sequence↗
PubMed1999

Comparison of input-output patterns in the corticospinal system of normal subjects and incomplete spinal cord injured patients.

We have examined input-output patterns in the corticospinal system after incomplete spinal cord injury. The amplitude of the motor evoked potential (MEP) to transcranial magnetic stimulation (TMS) was used to study the patterns of recruitment, with increasing stimulus intensity, and facilitation, with increasing voluntary contraction, in thenar muscles of 12 patients with incomplete spinal cord injuries and 13 control subjects. The patients had all suffered spinal cord injury at a segmental level rostral to C8 and T1, the segments supplying innervation of thenar muscles. The patients showed a less pronounced increase in MEP amplitude with increasing strength of TMS compared with the controls. Specifically, at a stimulus strength of 120% threshold and above, the patients showed significantly smaller MEPs relative to the maximum ulnar nerve M-wave response than the controls. The patients also showed a less steep pattern of facilitation with voluntary drive. The MEP continued to increase up to 50% maximum voluntary contraction (MVC) whereas the controls reached a plateau around 10% MVC. The results indicate that the patients show modified corticospinal recruitment and facilitation of the motoneurone pool. We speculate that the function of the adapted corticospinal system after spinal cord injury might be to regulate and modulate drive to motoneurones originating from segmental and other descending inputs. We discuss how such a modified corticospinal system might be of functional benefit to the patients.

Adult↗
PubMed1999

Clinical correlates of the initial and long-term cost of coronary bypass surgery and coronary angioplasty.

BACKGROUND: Medical costs vary substantially among patients. Understanding the baseline factors that predict subsequent cost may allow better selection of therapy for individual patients. Understanding the postprocedure events that increase cost should help to improve efficiency and effectiveness of coronary revascularization. METHODS: Data on 4-year costs were collected from patients randomly assigned to coronary angioplasty or bypass surgery as part of the BARI (Bypass Angioplasty Revascularization Investigation) trial. Regression models first examined factors known at the time of randomization that prospectively predicted initial procedure cost and long-term cost. Subsequent models tested the value of postrandomization events as explanatory variables for cost. RESULTS: The independent baseline predictors of higher initial percutaneous transluminal coronary angioplasty cost included 3-vessel disease (+12%) and acute presentations (+22%), whereas the independent predictors of higher initial coronary artery bypass grafting cost included the number of comorbid conditions (+5% per condition) and female sex (+7%). The independent baseline predictors of 4-year cost included heart failure (+26%), diabetes (+22%), comorbidity (+10%), and angioplasty assignment in patients with 2-vessel disease (-15%). Postrandomization models showed higher initial and long-term costs were strongly correlated with the number of repeat revascularization procedures (+30% to +128%) and the occurrence of clinical complications (+8% to +131%). CONCLUSIONS: Two-vessel disease identifies patients likely to have lower costs after angioplasty, whereas heart failure, comorbid conditions, and diabetes identify patients likely to accrue higher costs after either angioplasty or bypass surgery. Long-term costs can be potentially reduced by interventions that decrease procedural complications or reduce the need for repeat revascularization.

Aged↗
PubMed1999

Synthesis and biological activity of enantiomeric pairs of phosphosulfonate herbicides.

The phosphosulfonates are a new class of soil-active herbicides which control a variety of annual grass and broadleaf weeds. Chirality at the phosphorus atom afforded the opportunity to explore stereospecific requirements for herbicidal activity. Chiral (hydroxymethyl)phosphinate intermediates were enzymatically resolved (Pseudomonas fluorescens lipase) from the racemic mixtures and then used to prepare two pairs of enantiomeric phosphosulfonates. Biological testing of the enantiomeric phosphosulfonate herbicides demonstrated that, in each case, the herbicidal activity was attributed to the (+) enantiomer and that the (+) enantiomer is more active than the racemate.

Herbicides↗
PubMed1999

Motor unit discharge characteristics during voluntary contraction in patients with incomplete spinal cord injury.

Synchronisation of motor unit discharges is commonly seen in hand muscles of normal man but is absent following neurologically complete spinal cord injury and reduced after stroke. These findings support the notion that some corticospinal inputs to motoneurones are shared and contribute to the observed synchrony of discharge. In this study we have examined motor unit discharge in hand muscles below the level of an incomplete spinal cord injury in an attempt to relate strength of synchrony to the integrity of the corticospinal tract. Eight patients with incomplete spinal cord injury (neurological level C3-C7) and eight control subjects took part in the study. The patients had sustained injury 14-191 weeks prior to the recordings and had since regained good motor function in their hands. Two concentric needle electrodes were inserted into the first dorsal interosseus muscle which subjects were instructed to contract weakly so that potentials from individual motor units could be reliably identified on both recordings. Synchrony was detected by constructing cross-correlograms between the discharges of pairs of individual motor units. The amount of synchronous firing was determined from the magnitude of any peak in the cross-correlogram, as the probability above chance (XP) of one motor unit firing with respect to the other and vice versa. The degree of synchrony was lower (P < 0.05) in the patient group (mean XP 0.06) than in the control group (mean XP 0.09). The incidence of significant synchrony was lower in the patient group (41.8 %) than in the control group (92.9 %). The mean (+/- S.E.M.) frequency of motor unit discharge was slightly lower (P < 0.05) in patients (9.7 +/- 0.4 impulses s-1) than controls (10.8 +/- 0.5 impulses s-1). The mean width of synchrony peaks was narrower (P < 0.05) in patients (11.4 +/- 1.1 ms) than controls (13.2 +/- 0.6 ms). We conclude that the weaker synchrony of motor unit discharge in incomplete spinal cord injury may reflect permanent damage to some corticospinal axons.

Adult↗