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Biomedical subjects

M E Ward

Publications and source records attributed to M E Ward.

At least 19 recordsLinked to original sources
PubMed1995

Extent and kinetics of genetic change in the omp1 gene of Chlamydia trachomatis in two villages with endemic trachoma.

Variants of Chlamydia trachomatis in two Gambian villages with hyperendemic trachoma were analyzed by omp1-based polymerase chain reaction and sequencing from conjunctival swabs. Samples collected over a 22-month period included a complete cross-sectional study of each village. Overall, 4 genovar A and 4 B variants were characterized by point mutations in the omp1 gene, resulting in changes in the inferred amino acid sequence. Two genovar A and 2 B variants accounted for 87% of the total ocular chlamydial infection in both villages. Although some flux in the prevalence of individual variants was observed overtime, their overall distribution remained remarkably stable. There was no evidence of major antigenic shift arising from recombination events at the omp1 locus as described for genital tract infection. These results indicate that omp1 variation in these two trachoma-hyperendemic communities is limited and unlikely to hamper development of trachoma vaccines based on the major outer membrane protein.

Amino Acid Sequence↗
PubMed1995

A peptide of Chlamydia trachomatis shown to be a primary T-cell epitope in vitro induces cell-mediated immunity in vivo.

Chlamydiae are a major cause of infertility and preventable blindness and there is currently no effective vaccine in humans or rodents against these organisms. We have previously shown that a peptide of 12 amino acids (termed TINKP) from a conserved region of the major outer membrane protein (MOMP) of Chlamydia trachomatis (C. trachomatis) is a primary T-cell epitope in humans. Here we showed that when dendritic cells (DC) from C3H or BALB/c mice were pulsed in vitro with the peptide they stimulated proliferation of syngeneic T cells in vitro indicating that the peptide is also a primary T-cell epitope in mice. Since the skin is a rich source of DC, we immunized mice from each strain with an intradermal injection of the peptide. Humoral and cell-mediated immunity to peptide, MOMP or whole elementary bodies (EB) of C. trachomatis (F/NI1/GU) were assessed. No antibody response to TINKP was observed. However, immunized mice showed recall responses to all three chlamydial antigens. T-cell-mediated immunity in the absence of antibody was induced by a single injection of the peptide intradermally. C. trachomatis isolated from the human genital tract causes salpingitis in mice. Preliminary studies in susceptible C3H mice indicated that intradermal injection of peptide conferred some protection against the development of salpingitis. Thus, a primary T-cell epitope identified by in vitro stimulation using DC can also initiate cell-mediated immunity in vivo and this approach may be useful in the development of vaccines.

Animals↗
PubMed1994

Effect of inhibition of nitric oxide release on the diaphragmatic oxygen delivery-consumption relationship.

PURPOSE: In the vascularly isolated resting and contracting (3 Hz) canine hemidiaphragm, the hypothesis that nitric oxide (NO) is an important regulator of diaphragmatic O2 extraction was tested. METHODS: The effect of an intra-arterial infusion of an NO-synthase inhibitor NG-nitro-L-arginine (L-NA) on the critical O2 delivery (QO2c), below which O2 consumption becomes dependent on O2 supply, was assessed in two groups of animals in which either saline or L-NA (6 x 10(-4) mol/L) was infused into the phrenic artery over 20 minutes. The diaphragm was then perfused either by left femoral arterial blood (autoperfusion) or by pump perfusion with blood from the femoral artery. QO2 was reduced by stepwise hemorrhage in the autoperfusion groups and by reducing the pump rate in the pump perfusion groups. RESULTS: During autoperfusion, QO2c in the saline- and L-NA-treated groups was not different (0.88 +/- 0.15 and 0.98 +/- 0.12 mL/min/100 g, respectively) for the resting diaphragm. Critical O2 extraction ratios were not different (64.5% +/- 9.9% and 67.8% +/- 6.4%, respectively). In the saline group, QO2c during 3-Hz stimulation was 5.03 +/- 0.9 mL/min/100 g. In the L-NA group, diaphragm flow was lower than the saline group, and no QO2c was found. In the pump-perfused contracting diaphragm, QO2c in both groups did not differ (3.1 +/- 0.5 and 4.05 +/- 0.65 mL/min/100 g, respectively). O2 extraction ratios at these O2 deliveries were different (63.3% +/- 5.2% and 77.4% +/- 4.3%, respectively). However, NO-synthase inhibiton had no effect on maximum diaphragmatic O2 extraction ratio. CONCLUSIONS: These results indicate that NO release is an important modulator of the tone of diaphragmatic resistance vessels, but it does not appear to regulate the processes by which O2 extraction is enhanced to compensate for decreased O2 delivery.

Animals↗
PubMed1994

Reactivity of antibodies to heteroclitic peptides based on the Chlamydia trachomatis major outer-membrane protein.

One problem of peptide vaccines is that antibodies generated against them react poorly with the target sequence on the native protein. Using monoclonal antibodies (mAbs) to the serovar L1 type-specific epitope on the major outer-membrane protein of Chlamydia trachomatis as our model in conjunction with the Pin Technology Epitope Scanning technique, we had previously identified the critical binding site at this epitope as DAVP. Amino acid substitution showed that AV were essential residues for binding. A series of structurally related (heteroclitic) peptides retaining AV were synthesized. Some of these were found to be much more reactive with the model mAb than peptides of cognate sequence. It was hypothesized that the DAVP peptide only approximated to the conformation of the homologous sequence in the native protein, whereas some of the flexible heteroclitic peptides produced conformations which more closely resembled the native constrained sequence. The key question was whether the most reactive heteroclitic peptide would also generate antibody capable of more efficient binding to the native protein. We therefore immunized mice with one of six heteroclitic peptides or one of two native sequence control peptides. The reactivity of these antisera with the peptide immunogens and with native chlamydial elementary bodies was then evaluated by enzyme immunoassay. Pooled antisera to two of the heteroclitic peptides reacted with significantly greater absorbance (P < 0.05) and at higher dilution with whole chlamydiae than did pooled antisera to the control peptides. This suggests that heteroclitic peptides may in some circumstances be useful to increase the reactivity of site-specific antibodies with epitopes on the native protein important for vaccine development or for serodiagnosis.

Amino Acid Sequence↗
PubMed1994

Evidence for naturally occurring recombination in the gene encoding the major outer membrane protein of lymphogranuloma venereum isolates of Chlamydia trachomatis.

The nucleotide sequence of the major outer membrane protein gene (omp1) was determined for three geographically distinct lymphogranuloma venereum isolates which were serologically untypeable. The three omp1 sequences were hybrids of serovars L1 and L2, containing a putative DNA recombination site in variable segment 2. Efforts to manipulate the chlamydial genome in vitro by recombination should be intensified.

Amino Acid Sequence↗
PubMed1994

Molecular epidemiology of trachoma in a Gambian village.

The application of a diagnostic and genotyping technique based on the polymerase chain reaction (PCR) to the study of trachoma epidemiology in the Gambian village of Jali is reported. PCR based on the major outer membrane protein (MOMP) gene of Chlamydia trachomatis appears to be more sensitive than either isolation or antigen detection by enzyme immunoassay; it had a specificity of 95% and sensitivity of 51% against clinical signs. PCR genotyping identified genotypes A and B of Chlamydia trachomatis circulating in Jali. Sequencing revealed a Pst1 restriction endonuclease site in the amplified MOMP gene of some B strains but not others; Pst1 digestion of the PCR product proved an easy method of distinguishing these strains. The distribution of serotypes and B strain variants shows a significant degree of household clustering (p < 0.001). PCR based genotyping combined with strain typing provides a new and powerful epidemiological tool for the study of transmission events in trachoma.

Antigens, Bacterial↗
PubMed1994

Pathological O2 supply dependence of diaphragmatic and systemic O2 uptake during endotoxemia.

Our aim was to assess whether endotoxemia impairs the ability of the diaphragm to extract O2 and whether this defect leads to a greater dependence of O2 uptake on O2 delivery. In two groups of anesthetized mechanically ventilated dogs, the left hemidiaphragm was vascularly isolated. Diaphragmatic blood flow and cardiac output (CO) were measured simultaneously in all animals. Saline (S group) or Escherichia coli endotoxin (100 mg; E group) was infused intravenously over 60 min. In both groups, CO was reduced in stages by controlled hemorrhage, and systemic and diaphragmatic O2 deliveries and consumptions were measured at each stage to construct the O2 delivery-O2 consumption relationships. In the S group, the average systemic O2 delivery below which O2 uptake became supply dependent was 7.2 ml.kg-1.min-1. At this O2 delivery, systemic O2 extraction ratio (ER) averaged 67.9%, whereas the maximum O2 ER was 91.3%. Critical diaphragmatic O2 delivery and critical and maximum diaphragmatic O2 ER, by comparison, averaged 9.0 ml.kg-1.min-1, 65%, and 81.9%, respectively. Endotoxin infusion raised critical systemic O2 delivery to 16.7 ml.kg-1.min-1 (P < 0.05) and reduced critical and maximum systemic O2 ER to 55.5 and 77% (P < 0.05), respectively. Similarly, critical diaphragmatic O2 delivery in the E group increased to 14.8 ml.kg-1.min-1 (P < 0.05), whereas critical and maximum O2 ER declined to 51.8 and 72.8%, respectively (P < 0.05). Thus, endotoxemia impairs diaphragmatic O2 extraction. This, in turn, leads to a greater dependence of diaphragmatic O2 uptake on O2 delivery.

Animals↗
PubMed1994

Diaphragmatic pressure-flow relationship during hemorrhagic shock: role of nitric oxide.

In the vascularly isolated resting and contracting (3 Hz) canine hemidiaphragm, we studied the effect of intra-arterial infusion of the nitric oxide (NO) inhibitor NG-nitro-L-arginine (LNA) on the relationship between phrenic arterial perfusion pressure (Pphr) and blood flow (Qphr). In separate groups of animals, either saline or LNA (final concn 6 x 10(-4) M) was infused into the phrenic artery over 20 min. The diaphragm was then autoperfused by diverting flow from the left femoral artery. Arterial blood pressure was reduced in stages by controlled hemorrhage. The Pphr-to-Qphr relationship was plotted for each animal, and the third-order polynomial of best fit was determined by least squares regression. The inflection point of this relationship was determined for each animal. In the contracting and resting diaphragms, the inflection point corresponded to Pphr values of 83.6 +/- 4.7 and 72.5 +/- 6.8 mmHg, respectively, in the saline-treated group compared with 86.2 +/- 2.7 and 76.8 +/- 5.1 mmHg, respectively, in the LNA-treated group. In the contracting diaphragm, LNA reduced Qphr uniformly across the entire range of perfusion pressures. In the resting diaphragm, the effect of LNA was not uniform. At perfusion pressures below the inflection point, the flow was reduced in proportion to the reduction in inflection point flow. At higher perfusion pressures, Qphr was decreased to a greater extent than could be accounted for by the change in inflection point flow.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Multidomain binding of transforming growth factor alpha to the epidermal growth factor receptor.

Solubilized epidermal growth factor receptor (EGF-R) has been used in an extension of the Geysen epitope mapping protocol in order to provide additional insight into the amino acid residues in human transforming growth factor alpha (hTGF alpha) which are critical to recognition and binding. Overlapping heptapeptides which encompassed the 50 amino acid primary sequence of hTGF alpha were synthesized on a polyethylene solid phase, and the amount of detergent-solubilized EGF-R bound to each peptide was measured using ELISA. EGF-R appeared to bind reproducibly to four heptapeptides cognate to sequences in both the N- and C-domains of hTGF alpha (residues 22-28, 28-34, 36-42, and 44-50). Visualization of these four regions on three-dimensional solution phase structures of hTGF alpha, derived from 1H NMR measurements [Kline, T.-P., Brown, F.K., Brown, S.C., Jeffs, P.W., Kopple, K.D., & Mueller, L. (1990) Biochemistry 29, 7805-7813], indicated that the peptide segments are located on a single face of the protein and suggested the presence of a potential receptor binding cavity. If peptide segments within both the N- and C-domains of hTGF alpha are involved in binding to EGF-R, then this has direct consequences for possible molecular mechanisms by which receptor activation might take place. For example, the observed conformational flexibility in the six NMR-derived hTGF alpha structures due to variations in the main-chain torsion angles of Val-33, in combination with the involvement of residues from both domains in the proposed binding cavity, may imply that receptor activation results from interdomain reorientation in the protein ligand.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1992

Survey of 3765 cardiopulmonary resuscitations in British hospitals (the BRESUS Study): methods and overall results.

OBJECTIVE: To determine the circumstances, incidence, and outcome of cardiopulmonary resuscitation in British hospitals. DESIGN: Hospitals registered all cardiopulmonary resuscitation attempts for 12 months or longer and followed survival to one year. SETTING: 12 metropolitan, provincial, teaching, and non-teaching hospitals across Britain. SUBJECTS: 3765 patients in whom a resuscitation attempt was performed, including 927 in whom the onset of arrest was outside the hospital. MAIN OUTCOME MEASURE: Survival after initial resuscitation, at 24 hours, at discharge from hospital, and at one year, calculated by the life table method. RESULTS: There were 417 known survivors at one year, with 214 lost to follow up. By life table analysis for every eight attempted resuscitations there were three immediate survivors, two at 24 hours, 1.5 leaving hospital alive, and one alive at one year. Survival at one year was 12.5% including out of hospital cases and 15.0% not including these cases. Each hospital year averaged 30 survivors at one year: three who had an arrest outside hospital, seven who had one in the accident and emergency department, seven in the cardiac care unit, 10 in the general wards, and three in other, non-ward areas. Within the hospitals survival rates were best in those who had an arrest in the accident and emergency department, the cardiac care unit, or other specialised units. Outcome varied 12-fold in subgroups defined by age, type of arrest, and place of arrest. CONCLUSION: 71% of the mortality at one year in patients undergoing attempted resuscitation occurred during the initial arrest. Hospital resuscitation is life saving and cost effective and warrants appropriate attention, training, coordination, and equipment.

Age Factors↗
PubMed1992

A longitudinal study of trachoma in a Gambian village: implications concerning the pathogenesis of chlamydial infection.

In order to investigate risk factors for the acquisition of trachoma, and to study the effect of continued exposure to ocular chlamydial infection on the severity of inflammatory trachoma and its cicatricial sequelae, a longitudinal study was conducted in a Gambian village. Over a 20-month period, the incidence of active (inflammatory) trachoma was significantly higher among those sharing a bedroom with an active case (64/561, 11.4%) than among those who were not exposed in this way (37/658, 5.6%) (relative risk 1.97, 95% confidence interval 1.33-2.90). There was a positive trend in the odds ratio for severe to moderate inflammatory disease versus mild disease as the number of active cases in the bedroom increased, but this failed to achieve statistical significance (P = 0.0506). Individuals with inflammatory trachoma of moderate or severe intensity at one survey were significantly more likely than others to have moderate or severe inflammatory changes at a previous or subsequent survey (odds ratio 14.9, 95% confidence interval 3.9-68.0), implying that host factors may be more important determinants of severity than the frequency of exposure to reinfection.

Adolescent↗
PubMed1992

Genotyping of Chlamydia trachomatis from a trachoma-endemic village in the Gambia by a nested polymerase chain reaction: identification of strain variants.

Direct amplification of the major outer membrane protein (MOMP) gene by polymerase chain reaction (PCR) was used to identify Chlamydia trachomatis in eye swabs from clinically active cases of endemic trachoma in a Gambian village. Chlamydial DNA was detected in 51% of 96 subjects with clinically active disease and in 5% of 37 clinically negative individuals. The PCR detection was combined with typing, using nested primers to variable sequences (VS) 1, 2, and 4 of the MOMP genes to distinguish between trachoma genotypes A, B, and C, respectively. Genotypes A and B were detected in the village, with some individuals harboring both genotypes within the same eye. DNA sequencing revealed strain variants of both genotypes. Typing of genotype and strain variants is now in progress to study trachoma transmission within the village.

Bacterial Outer Membrane Proteins↗
PubMed1992

Quantitative assessment of bed rise difficulty in young and elderly women.

OBJECTIVE: To describe the motions which occur during rising from bed, specifically the motions that appeared to characterize difficulty in rising from a bed in older adults. DESIGN: Development of a Mobility assessment tool. SETTING: Retirement center and two university laboratories. PARTICIPANTS: Three groups of female volunteers: young controls (n = 17, mean age 24), community-dwelling older adults (n = 12, mean age 71), and retirement center-dwelling older adults who admitted to difficulty in rising from a bed (n = 15, mean age 86). INTERVENTION: Videotaping of motions occurring during controlled rises from a supine to sitting position. MAIN OUTCOME MEASURES: These motions were rated on the specially developed Bed Rise Difficulty (BRD) scale, a scale designed to measure movements that characterize difficulty in rising from a bed in older adults. Subject groups were compared in total BRD score, individual BRD item score, and total time to rise. Item relationships and scale reliability were also assessed. RESULTS: Older adults with no apparent difficulty in rising based on total time to rise or on the BRD score nevertheless showed differences in upper extremity use when compared to young controls. Older adult subjects with difficulty in rising from a bed, when compared to other older adults with no apparent difficulty, differed more often in their upper extremity and leg use to facilitate the rise. Five BRD scale items, including use of extremity pushes, discontinuity of trunk and leg motion, multiple shoulder/pelvic adjustments, multiple leg adjustments, and poor vertical heel clearance may have best indicated true bed rise difficulty. CONCLUSIONS: These data provide a reliable and valid method to characterize difficulty in rising from a bed and provide the basis for biomechanical analyses of the strength and joint ranges of motion required to rise from a bed.

Activities of Daily Living↗
PubMed1992

Analysis of human chest wall motion using a two-compartment rib cage model.

We present a model of chest wall mechanics that extends the model described previously by Macklem et al. (J. Appl. Physiol. 55: 547-557, 1983) and incorporates a two-compartment rib cage. We divide the rib cage into that apposed to the lung (RCpul) and that apposed to the diaphragm (RCab). We apply this model to determine rib cage distortability, the mechanical coupling between RCpul and RCab, the contribution of the rib cage muscles to the pressure change during spontaneous inspiration (Prcm), and the insertional component of transdiaphragmatic pressure in humans. We define distortability as the relationship between distortion and transdiaphragmatic pressure (Pdi) and mechanical coupling as the relationship between rib cage distortion and the pressure acting to restore the rib cage to its relaxed configuration (Plink), as assessed during bilateral transcutaneous phrenic nerve stimulation. Prcm was calculated at end inspiration as the component of the pressure displacing RCpul not accounted for by Plink or pleural pressure. Prcm and Plink were approximately equal during quiet breathing, contributing 3.7 and 3.3 cmH2O on average during breaths associated with a change in Pdi of 3.9 cmH2O. The insertional component of Pdi was measured as the pressure acting on RCab not accounted for by the change in abdominal pressure during an inspiration without rib cage distortion and was 40 +/- 12% (SD) of total Pdi. We conclude that there is substantial resistance of the human rib cage to distortion, that, along with rib cage muscles, contributes importantly to the fall in pleural pressure over the costal surface of the lung.

Adult↗
PubMed1992

Effect of phrenic afferent stimulation on pattern of respiratory muscle activation.

Ventilation and electromyogram (EMG) activities of the right hemidiaphragm, parasternal intercostal, triangularis sterni, transversus abdominis, genioglossus, and alae nasi muscles were measured before and during central stimulation of the left thoracic phrenic nerve in 10 alpha-chloralose anesthetized vagotomized dogs. Pressure in the carotid sinuses was fixed to maintain baroreflex activity constant. The nerve was stimulated for 1 min with a frequency of 40 Hz and stimulus duration of 1 ms at voltages of 5, 10, 20, and 30 times twitch threshold (TT). At five times TT, no change in ventilation or EMG activity occurred. At 10 times TT, neither tidal volume nor breathing frequency increased sufficiently to reach statistical significance, although the change in their product (minute ventilation) was significant (P less than 0.05). At 20 and 30 times TT, increases in both breathing frequency and tidal volume were significant. At these stimulus intensities, the increases in ventilation were accompanied by approximately equal increases in the activity of the diaphragm, parasternal, and alae nasi muscles. The increase in genioglossus activity was much greater than that of the other inspiratory muscles. Phrenic nerve stimulation also elicited inhomogeneous activation of the expiratory muscles. The transversus abdominis activity increased significantly at intensities from 10 to 30 times TT, whereas the activity of the triangularis sterni remained unchanged. The high stimulation intensities required suggest that the activation of afferent fiber groups III and IV is involved in the response. We conclude that thin-fiber phrenic afferent activation exerts a nonuniform effect on the upper airway, rib cage, and abdominal muscles and may play a role in the control of respiratory muscle recruitment.

Animals↗
PubMed1992

Oxygen delivery-independent effect of blood flow on diaphragm fatigue.

To determine the effect of blood flow on diaphragm fatigue independent of oxygen delivery, the left hemidiaphragm was vascularly isolated in 14 pentobarbital-anesthetized, mechanically ventilated dogs. Fatigue (decline in tension generation) of the left diaphragm was induced by phrenic nerve stimulation at 10 Hz, 12/min, duty cycle of 0.5 for 8 min. Two stimulation periods separated by 30 min of rest were performed in each animal. Diaphragmatic O2 delivery during the two periods was the same. In Group 1 (n = 8), the diaphragm was autoperfused from the femoral artery (high O2-low flow) during the first stimulation period. The tension generated by the diaphragm during this period declined progressively to 47.7% of initial values. In the second period in this group, the diaphragm was pump perfused with arterial blood, diluted with an equal volume of 6% dextran at a flow rate twice that of the first period (low O2-high flow). Tension in this period declined to 76% of initial tension (p less than 0.05 compared with high O2-low flow). In Group 2 (n = 6), stimulation performed while perfusing the diaphragm in the first period with diluted arterial blood at a flow rate twice that recorded during autoperfusion (low O2-high flow) produced a decline in tension to 70% of the initial values. In the second period, the diaphragm was perfused with undiluted arterial blood at a flow rate equal to 50% of that of the first period (high O2-low flow). Tension during this period declined to 56% of initial values (p less than 0.05 compared with low O2-high flow).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗