Biomedical subjects
N A Taylor
Publications and source records attributed to N A Taylor.
Postoperative radiotherapy increases locoregional control of patients with stage IIIA non-small-cell lung cancer treated with induction chemotherapy followed by surgery.
PURPOSE: To determine the effectiveness of postoperative radiotherapy (RT) in patients with Stage IIB and Stage IIIA non-small-cell lung cancer (NSCLC) treated with induction chemotherapy followed by surgery. METHODS AND MATERIALS: We retrospectively reviewed the treatment records of 98 patients (58 men and 40 women; median age 61 years, range 31-91) with Stage IIB and Stage IIIA NSCLC who were treated with induction chemotherapy followed by surgery at our institution between January 1990 and December 2000. Patients were grouped by treatment (chemotherapy/surgery alone vs. chemotherapy/surgery/RT), by disease stage and nodal classification. The rates of local control (LC), disease-specific survival, disease-free survival, and overall survival (OS) were calculated using the Kaplan-Meier method. RESULTS: Of the 98 patients, 40 had Stage IIB and 58 had Stage IIIA. The clinical disease stage and N stage were significantly greater in those patients who underwent RT than in those who did not; however, no statistically significant differences were identified in the additional characteristics between those receiving and not receiving RT within each stage or nodal group. The overall 5-year actuarial LC rate was 81% in the RT group and 54% in the chemotherapy/surgery-alone group (p = 0.07). Postoperative RT significantly improved the 5-year LC rate in patients with Stage IIIA disease (from 35% to 82%, p = 0.01). Postoperative RT did not significantly improve the 5-year OS rate (30% with RT vs. 49% without) for all patients or for patients with Stage IIIA disease. The disease-specific survival and disease-free survival rates did not differ between the treatment groups. Patients who responded to induction chemotherapy had a significantly greater 5-year OS rate (49%) than did those with stable or progressive disease (22%, p = 0.003). CONCLUSION: Postoperative RT in patients with Stage IIIA NSCLC treated with induction chemotherapy followed by surgery significantly improved LC without improving OS. Significantly improved survival was observed in all patients who responded to induction chemotherapy compared with those with stable or progressive disease.
Whole-body pre-cooling does not alter human muscle metabolism during sub-maximal exercise in the heat.
Muscle metabolism was investigated in seven men during two 35 min cycling trials at 60% peak oxygen uptake, at 35 degrees C and 50% relative humidity. On one occasion, exercise was preceded by whole-body cooling achieved by immersion in water during a reduction in temperature from 29 to 24 degrees C, and, for the other trial, by immersion in water at a thermoneutral temperature (control, 34.8 degrees C). Pre-cooling did not alter oxygen uptake during exercise (P > 0.05), whilst the change in cardiac frequency and body mass both tended to be lower following pre-cooling (0.05 < P < 0.10). When averaged over the exercise period, muscle and oesophageal temperatures after pre-cooling were reduced by 1.5 and 0.6 degrees C respectively, compared with control (P < 0.05). Pre-cooling had a limited effect on muscle metabolism, with no differences between the two conditions in muscle glycogen, triglyceride, adenosine triphosphate, creatine phosphate, creatine or lactate contents at rest, or following exercise. These data indicate that whole-body pre-cooling does not alter muscle metabolism during submaximal exercise in the heat. It is more likely that thermoregulatory and cardiovascular strain are reduced, through lower muscle and core temperatures.
Metabolic habituation following repeated resting cold-water immersion is not apparent during low-intensity cold-water exercise.
This project examined the effects of repeated, resting cold-water immersion on metabolic heat production and core temperature defence during subsequent rest and exercising immersions. Seven males undertook 15 days of cold-water adaptation, immersed to the fourth intercostal space, with cold-water stress tests (CWST) on days 1, 8 and 15 (18.1 SD 0.1 degree C: 60 min seated, followed by 30 min cycling (1 W.kg-1)), and 90-min resting immersions (18.4 SD 0.4 degree C) on each of the intervening days. Adaptation elicited an habituated thermogenic response during the rest phase of CWST3 beyond 20 min, compared to CWST1 (P < 0.05), with oxygen consumption averaging 11.15 (+/- 0.25) ml.kg-1.min-1 and 8.61 (+/- 0.90) ml.kg-1.min-1 by 50 min, for CWST1 and CWST3, respectively. During exercise, this metabolic blunting was only apparent over the first 10-min period (60-70 min). No significant differences were observed during either the rest or exercise phases of the CWSTs for oesophageal temperature (Tes). While repeated cold-water exposures produced an habituated-thermogenic response, for an equivalent drop in Tes during rest, neither this response, nor an elevated thermogenesis, were apparent during subsequent cold-water exercise.
Processing of beta-secretase by furin and other members of the proprotein convertase family.
The amyloid peptide is the main constituent of the amyloid plaques in brain of Alzheimer's disease patients. This peptide is generated from the amyloid precursor protein by two consecutive cleavages. Cleavage at the N terminus is performed by the recently discovered beta-secretase (Bace). This aspartyl protease contains a propeptide that has to be removed to obtain mature Bace. Furin and other members of the furin family of prohormone convertases are involved in this process. Surprisingly, beta-secretase activity, neither at the classical Asp(1) position nor at the Glu(11) position of amyloid precursor protein, seems to be controlled by this maturation step. Furthermore, we show that Glu(11) cleavage is a function of the expression level of Bace, that it depends on the membrane anchorage of Bace, and that Asp(1) cleavage can be followed by Glu(11) cleavage. Our data suggest that pro-Bace could be active as a beta-secretase in the early biosynthetic compartments of the cell and could be involved in the generation of the intracellular pool of the amyloid peptide. We conclude that modulation of the conversion of pro-Bace to mature Bace is not a relevant drug target to treat Alzheimer's disease.
Can skin temperature manipulation, with minimal core temperature change, influence plasma volume in resting humans?
We investigated body-fluid distribution in resting humans, during short-term, whole-body skin temperature modification, in which core temperature changes (DeltaT(c)) were minimal. Seven males participated in hot (36.2 degrees C (s.d. 0.7), 44% relative humidity (rh; s.d. 3)), temperate (22.0 degrees C (s.d. 1.0), 52% rh (s.d. 6)), and cool trials (14.4 degrees C (s.d. 1.6), 74% rh (s.d. 9)), while seated at rest. Total body water (TBW), extracellular fluid (ECF), erythrocyte (RCV) and plasma volumes (PV) were measured using a simultaneous radionuclide dilution technique. In the cold, PV contracted by 205 ml (+/-60) by the end of exposure (p = 0.04), while in the heat, PV expanded 108 ml (+/-123; p = 0.02). Both RCV and TBW remained stable, regardless of the environment. Despite fluid movement across the vascular wall, ECF, interstitial and intracellular volumes were relatively unaffected by skin temperature. It was concluded that, at rest, and with minimal DeltaT(c), the intravascular fluid volume was dependent on prevailing environmental conditions, and its impact on local skin temperature and venomotor tone.
Cardiorespiratory dynamics: sensitivity of the on-transition to endurance-training status.
This project investigated the sensitivity of oxygen uptake (VO(2)) dynamics to training-induced physiological changes, across a broad spectrum of endurance-training histories. Forty subjects participated: sedentary (n = 10), active healthy (n = 10), regular runners (n = 10), and competitive distance runners (n = 10). Subjects completed a cycle step-function protocol, to elicit a steady state at 60% maximal work rate. Breath-by-breath data were collected for VO(2) and cardiac frequency (f(c)), and modelled mathematically, and used to determine the average response times to attain 20%, 40%, 60%, 80% and 100% of the respective steady states. The between-group comparisons for both VO(2) and f(c) revealed significantly faster response times to 40%, 60%, 80% and 100% of the induced response, for the better trained subjects (P < 0.05). In general, this technique permitted differentiation between the VO(2) and f(c) response dynamics of non-elite subjects from a broad range of endurance-training histories, with differences becoming more pronounced as subjects approached the steady state.
Static respiratory muscle work during immersion with positive and negative respiratory loading.
Upright immersion imposes a pressure imbalance across the thorax. This study examined the effects of air-delivery pressure on inspiratory muscle work during upright immersion. Eight subjects performed respiratory pressure-volume relaxation maneuvers while seated in air (control) and during immersion. Hydrostatic, respiratory elastic (lung and chest wall), and resultant static respiratory muscle work components were computed. During immersion, the effects of four air-delivery pressures were evaluated: mouth pressure (uncompensated); the pressure at the lung centroid (PL,c); and at PL,c +/-0.98 kPa. When breathing at pressures less than the PL,c, subjects generally defended an expiratory reserve volume (ERV) greater than the immersed relaxation volume, minus residual volume, resulting in additional inspiratory muscle work. The resultant static inspiratory muscle work, computed over a 1-liter tidal volume above the ERV, increased from 0.23 J. l(-1), when subjects were breathing at PL,c, to 0.83 J. l(-1) at PL,c -0.98 kPa (P < 0.05), and to 1.79 J. l(-1) at mouth pressure (P < 0.05). Under the control state, and during the above experimental conditions, static expiratory work was minimal. When breathing at PL,c +0.98 kPa, subjects adopted an ERV less than the immersed relaxation volume, minus residual volume, resulting in 0.36 J. l(-1) of expiratory muscle work. Thus static inspiratory muscle work varied with respiratory loading, whereas PL,c air supply minimized this work during upright immersion, restoring lung-tissue, chest-wall, and static muscle work to levels obtained in the control state.
The propeptide of prohormone convertase PC2 acts as a transferable aggregation and membrane-association signal.
Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of alpha1-antitrypsin, called pro2alpha1. PC2, alpha1-antitrypsin and pro2alpha1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2alpha1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. alpha1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2alpha1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the propeptide are important in these events.
Differences in the autocatalytic cleavage of pro-PC2 and pro-PC3 can be attributed to sequences within the propeptide and Asp310 of pro-PC2.
PC2 and PC3 are subtilisin-like proteases involved in the maturation of prohormones and proneuropeptides within neuroendocrine cells. They are synthesized as zymogens that undergo autocatalytic maturation within the secretory pathway. Maturation of pro-PC2 is slow (t12 >8 h), exhibits a pH optimum of 5.5 and is dependent on calcium (K0.5 2 mM), while pro-PC3 maturation is relatively rapid (t12 15 min), exhibits a neutral pH optimum and is not calcium dependent. These differences in the rates and optimal conditions for activation of the proteases may contribute to the diversity of products generated by these proteases in different cell types. Although highly similar, there are two major differences between pro-PC2 and pro-PC3: the presence of an aspartate at position 310 in pro-PC2 compared with asparagine at the equivalent position in pro-PC3 (and all other members of the subtilisin family), and the N-terminal propeptides, which exhibit low sequence identity (30%). With a view to establishing the structural features that might be responsible for these differences in the maturation of pro-PC2 and pro-PC3, Asp310 in pro-PC2 was mutated to Asn, and Asn309 in pro-PC3 was mutated to Asp. Chimaeric proteins were also made consisting of the pro-region of PC2 fused to the mature portion of PC3 and the pro-region of PC3 fused to the mature region of PC2. The wild-type and mutant DNA constructs were then transcribed and translated in an in vitro system capable of supporting maturation of pro-PC2 and pro-PC3. The results demonstrated that Asp310 of pro-PC2 is responsible for the acidic pH optimum for maturation. Thus changing Asp310 to Asn shifted the pH optimum for maturation to pH 7.0. However, changing Asn309 of pro-PC3 to Asp had no effect on the optimum pH for maturation of pro-PC3. A chimaeric construct containing the propeptide of pro-PC2 attached to PC3 shifted the pH optimum for maturation from pH 7.0 to 6.0 and slowed down the rate of maturation (t12 >8 h). When attached to PC2, the pro-region of pro-PC3 had no effect on the optimum pH for maturation (pH 5.5-6.0), but it did accelerate the rate of maturation (t12 2 h). These results demonstrate that Asp310 and the pro-region of pro-PC2 contribute to the acidic pH optimum and low rate of maturation of this zymogen relative to its closely related homologue PC3.
Human sudomotor responses to heating and cooling upper-body skin surfaces: cutaneous thermal sensitivity.
The influence of local skin temperature (Tskl) on the control of local and whole-body sweating was evaluated in eight healthy males. A water-perfusion garment (37 degrees C) and a climatic chamber (36.45 +/- 0.78 degrees C; [+/- SD]; relative humidity 60.3 +/- 1.6%) were used to raise and clamp skin and core temperatures. Warm and cool stimuli were applied to four upper-body skin regions (face, arm, forearm, hand) using perfusion patches (249.0 +/- 0.2 cm2). Heating elevated, while cooling suppressed sweat rate (msw) locally, and at other skin surfaces. However, the tendency for Tskl manipulations to induce localized sweat responses was no more powerful than it was at stimulating sweating in non-treated regions (P > 0.05). Accordingly, neither thermal stimulus produced significantly greater local sudomotor influences than were elicited contralaterally (P > 0.05). No statistical support was found for the notion of inter-regional differences in upper-body cutaneous thermal sensitivity for sudomotor control, and, regardless of the stimulation site, whole-body sudomotor responses to localized thermal treatments were equivalent (P > 0.05).
Human body-fluid distribution during exercise in hot, temperate and cool environments.
Using a simultaneous-dilution technique, we investigated body-fluid volume changes during exercise in seven males, during 50 min of cycling (50% maximal work rate) in hot (36.2 degrees C), temperate (22.0 degrees C) and cool conditions (14.4 degrees C). Total body water (TBW), extracellular fluid (ECFV), plasma (PV) and erythrocyte volumes (RCV) were measured, while blood volume (BV), interstitial fluid volume (IFV), extracellular water (ECW) and intracellular water volumes (ICW) were derived. During the initial 10 min of cycling, BV decreased in all environments (P .01), primarily because of a PV reduction (P = 0.01), while IFV, ECFV and ICW were not significantly changed. By 30 min, BV recovered in the temperate and cool conditions, despite mass losses of 563 and 520 mL (respectively), but remained depleted in the hot condition (P = 0.01). The 50-min volume changes revealed that, throughout exercise, body-fluid losses appeared to be drawn primarily from the extracellular space, regardless of air temperature. In the hot condition, the PV change represented 63% of the TBW loss, with the ICW contributing 23%. It was concluded that, during cycling, progressive dehydration mainly affected the extracellular space, with the intravascular and intracellular spaces being defended in less stressful conditions.
Sorting of PC2 to the regulated secretory pathway in AtT20 cells.
PC2 and PC3 are neuroendocrine specific members of the eukaryotic subtilisin-like proprotein convertase (PC) family. Both are sorted via the regulated secretory pathway into secretory granules. In order to identify sequences in PC2 which are involved in targeting to the regulated secretory pathway we expressed a series of PC2 cDNAs containing mutations in the C terminal or propeptide domains in the mouse corticotrophic AtT20 cell line. Sorting of endogenous PC3 was used as a control. PC2 and PC3 were secreted with similar kinetics and sorted to secretory granules with similar efficiencies. Deletions of up to 50 amino acids from the C-terminus of proPC2 had no effect on secretion or sorting, but larger deletions completely prevented maturation or secretion. Two large deletions within the propeptide also prevented secretion. Smaller deletions between the primary and secondary cleavage sites, or of the primary cleavage site, reduced the amount of protein secreted but did not affect sorting to secretory granules. Replacement of the propeptide of PC2 with that of the endogenous PC3 also had no effect on secretion or sorting. The results indicate that targeting of proPC2 to the regulated secretory pathway is dependent on more than one region within the proPC2 molecule.
Mutations within the propeptide, the primary cleavage site or the catalytic site, or deletion of C-terminal sequences, prevents secretion of proPC2 from transfected COS-7 cells.
PC2 is a neuroendocrine endoprotease involved in the processing of prohormones and proneuropeptides. PC2 is synthesized as a proenzyme which undergoes proteolytic maturation within the cellular secretory apparatus. Cleavage occurs at specific sites to remove the N-terminal propeptide. The aim of the present study was to investigate structural requirements for the transfer of proPC2 through the secretory pathway. A series of mutant proPC2 constructs were transfected into COS-7 cells and the fate of the expressed proteins followed by pulse-chase analysis and immunocytochemistry. Human PC2 was secreted relatively slowly, and appeared in the medium primarily as proPC2 (75 kDa), together with much lower amounts of a processed intermediate (71 kDa) and mature PC2 (68 kDa). Mutations within the primary processing site or the catalytic triad caused the protein to accumulate intracellularly, whereas deletion of part of the propeptide, the P-domain or the C-terminal regions also prevented secretion. Immunocytochemistry showed that wild-type hPC2 was localized mainly in the Golgi, whereas two representative mutants showed a distribution typical of proteins resident in the endoplasmic reticulum. The results suggest that proenzyme processing is not essential for secretion of PC2, but peptides containing mutations that affect the ability of the propeptide (and cleavage sites) to fold within the catalytic pocket are not transferred beyond the early stages of the secretory pathway. C-terminal sequences may be involved in stabilizing such conformations.
Sweat distribution before and after repeated heat exposure.
We investigated the impact of short-term, moderate humidity heat acclimation upon sweat distribution. Eight males completed six daily heat exposures [cycling: ambient temperature 39.5 (0.2) degrees C, relative humidity 59.2 (0.8)%], during which auditory canal temperature (T(ac)) was maintained 1.4 degrees C above pre-exposure levels for 70 min by manipulating the work rate. On days 1 and 6, T(ac) and local sweat rates (m(sw): eight sites) were monitored. The pre-exposure, resting T(ac) and the T(ac) sweat threshold decreased from day 1 to day 6 [36.83 (0.05) degrees C vs 36.62 (0.05) degrees C, and 36.90 (0.05) degrees C vs 36.75 (0.05) degrees C, respectively; both P < 0.05]. However, the sweat-onset time, sweat sensitivity (delta m(sw)/deltaT(ac)) and established m(sw) were unaltered (P > 0.05). There was also no evidence of a post-acclimation redistribution in established m(sw) between the eight skin regions, though both the sweat sensitivity and established m(sw) for the forehead and hand were significantly greater than at the remaining sites (P < 0.05). It is concluded that the 5-day heat acclimation regimen provided only a minimal stimulus for sudomotor adaptation.
Effects of artificially-induced anaemia on sudomotor and cutaneous blood flow responses to heat stress.
The influence of artificially induced anaemia on thermal strain was evaluated in trained males. Heat stress trials (38.6 degrees C, water vapour pressure 2.74 kPa) performed at the same absolute work rates [20 min of seated rest, 20 min of cycling at 30% peak aerobic power (VO2pcak), and 20 min cycling at 45% VO2peak] were completed before (HST1) and 3-5 days after 3 units of whole blood were withdrawn (HST2). Mild anaemia did not elevate thermal strain between trials, with auditory canal temperatures terminating at 38.5 degrees C [(0.16), HST1] and 38.6 degrees C [(0.13), HST2; P > 0.05]. Given that blood withdrawal reduced aerobic power by 16%, this observation deviates from the close association often observed between core temperature and relative exercise intensity. During HST2, the absolute and integrated forearm sweat rate (mSW) exceeded control levels during exercise (P < 0.05), while a suppression of forehead mSW occurred (P < 0.05). These observations are consistent with a possible peripheral redistribution of sweat secretion. It was concluded that this level of artificially induced anaemia did not impact upon heat strain during a 60-min heat stress test.
A non-linear effect of ambient temperature on apparent glucose tolerance.
Increased ambient temperature affects apparent oral glucose tolerance to an extent which may have clinical implications for the diagnosis of impaired glucose tolerance and gestational diabetes. As a first step in order to better define the nature of this effect, we have examined, in a climate chamber, the effects of ambient temperature at four levels (20, 25, 30, and 35 degrees C) on glucose and insulin responses to a standard 75 g oral glucose tolerance test in seven non-diabetic male subjects. Plasma glucose responses to ambient temperature were compared with the responses of core (auditory canal) and skin temperatures. The 2-h plasma glucose was affected in a nonlinear manner by ambient temperature (5.4 +/- 0.2, 5.3 +/- 0.4, 6.5 +/- 0.3, 6.4 +/- 0.4 mmol/l at 20, 25, 30, and 35 degrees C, P = 0.015) with the effect localised between 25 and 30 degrees C (P = 0.012). Core temperature responded in a similar manner (36.6 +/- 0.1, 36.6 +/- 0.1, 36.9 +/- 0.1, 37.0 +/- 0.1, (P = 0.0005) with the effect localised 25 and 30 degrees C (P = 0.011). However skin temperature increased significantly with each 5 degrees C increase in ambient temperature (30.2 +/- 0.5, 33.0 +/- 0.5, 34.2 +/- 0.2, 35.2 +/- 0.2, P < or = 0.0001). We conclude that the acute effect of ambient temperature on apparent glucose tolerance is most likely due to redistribution of blood flow between cutaneous and visceral beds driven by changes in core temperature. The absence of temperature effects between the two lowest, and between the two highest temperatures, provides workable guidelines for the standardisation of conditions during oral glucose tolerance tests in circumstances where temperature may have clinically significant effects.
Mutations in the pro-peptide of PC2 prevent transit through the secretory pathway.
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