Showing papers in "Clinical Science in 1998"

In utero programming of chronic disease

Abstract: 1 Many human fetuses have to adapt to a limited supply of nutrients In doing so they permanently change their structure and metabolism 2 These 'programmed' changes may be the origins of a number of diseases in later life, including coronary heart disease and the related disorders stroke, diabetes and hypertension 3 This review examines the evidence linking these diseases to fetal undernutrition and provides an overview of previous studies in this area

Anti-inflammatory actions of glucocorticoids : molecular mechanisms

Abstract: 1. Glucocorticoids are widely used for the suppression of inflammation in chronic inflammatory diseases such as asthma, rheumatoid arthritis, inflammatory bowel disease and autoimmune diseases, all of which are associated with increased expression of inflammatory genes. The molecular mechanisms involved in this anti-inflammatory action of glucocorticoids is discussed, particularly in asthma, which accounts for the highest clinical use of these agents. 2. Glucocorticoids bind to glucocorticoid receptors in the cytoplasm which then dimerize and translocate to the nucleus, where they bind to glucocorticoid response elements (GRE) on glucocorticoid-responsive genes, resulting in increased transcription. Glucocorticoids may increase the transcription of genes coding for anti-inflammatory proteins, including lipocortin-1, interleukin-10, interleukin-1 receptor antagonist and neutral endopeptidase, but this is unlikely to account for all of the widespread anti-inflammatory actions of glucocorticoids. 3. The most striking effect of glucocorticoids is to inhibit the expression of multiple inflammatory genes (cytokines, enzymes, receptors and adhesion molecules). This cannot be due to a direct interaction between glucocorticoid receptors and GRE, as these binding sites are absent from the promoter regions of most inflammatory genes. It is more likely to be due to a direct inhibitory interaction between activated glucocorticoid receptors and activated transcription factors, such as nuclear factor-kappa B and activator protein-1, which regulate the inflammatory gene expression. 4. It is increasingly recognized that glucocorticoids change the chromatin structure. Glucocorticoid receptors also interact with CREB-binding protein (CBP), which acts as a co-activator of transcription, binding several other transcription factors that compete for binding sites on this molecule. Increased transcription is associated with uncoiling of DNA wound around histone and this is secondary to acetylation of the histone residues by the enzymic action of CBP. Glucocorticoids may lead to deacetylation of histone, resulting in tighter coiling of DNA and reduced access of transcription factors to their binding sites, thereby suppressing gene expression. 5. Rarely patients with chronic inflammatory diseases fail to respond to glucocorticoids, although endocrine function of steroids is preserved. This may be due to excessive formation of activator protein-1 at the inflammatory site, which consumes activated glucocorticoid receptors so that they are not available for suppressing inflammatory genes. 6. This new understanding of glucocorticoid mechanisms may lead to the development of novel steroids with less risk of side effects (which are due to the endocrine and metabolic actions of steroids). 'Dissociated' steroids which are more active in transrepression (interaction with transcription factors) than transactivation (GRE binding) have now been developed. Some of the transcription factors that are inhibited by glucocorticoid, such as nuclear factor-kappa B, are also targets for novel anti-inflammatory therapies.

Neutrophil Priming: Pathophysiological Consequences and Underlying Mechanisms

Abstract: 1 Neutrophil priming by agents such as tumour necrosis factor-alpha, granulocyte/macrophage colony-stimulating factor and lipopolysaccharide causes a dramatic increase in the response of these cells to an activating agent; this process has been shown to be critical for neutrophil-mediated tissue injury both in vitro and in vivo 2 The principle consequence of priming, aside from a direct effect on cell polarization, deformability and integrin/selectin expression, is to permit secretagogue-induced superoxide anion generation, degranulation and lipid mediator (eg leukotriene B4 and arachidonic acid) release It is now recognized that most priming agents also serve an additional function of delaying apoptosis and hence increasing the functional longevity of these cells at the inflamed site 3 The potential mechanisms underlying priming are discussed; current data suggest a dissociation between priming and changes in receptor number and/or affinity, G-protein expression, phospholipase C and phospholipase A2 activation and changes in intracellular Ca2+ concentration However, more recent studies support a key role for protein tyrosine phosphorylation and enhanced phospholipase D and phosphoinositide 3-kinase activity in neutrophil priming 4 Recent work has also revealed the potential for neutrophils to spontaneously and fully 'de-prime' after an initial challenge with platelet-activating factor This ability of neutrophils to undergo a complete cycle of priming-de-priming (and re-priming) reveals a previously unrecognized flexibility in the control of neutrophil behaviour at an inflamed site

An early prenatal exposure to excess glucocorticoid leads to hypertensive offspring in sheep

Abstract: 1. Recent studies in animals have linked fetal exposure to excess maternal glucocorticoids with the later occurrence of cardiovascular disorders, particularly hypertension. 2. To test the hypothesis that prenatal treatment could impact on adult blood pressure two groups of pregnant ewes were transported from the farm to the Institute at either 22-29 days of pregnancy (pretreatment group 1) or 59-66 days of pregnancy (pretreatment group 2), subjected to 48 h treatment with dexamethasone (0.28 mg day-1 kg-1 for 2 days) and then returned to the farm. The control group remained at the farm for the entire pregnancy. Lambs were then studied at approximately 4, 10 and 19 months after birth. 3. The basal mean arterial pressure in pretreatment group 1 (80 +/- 1 mmHg at 124 days; 83 +/- 1 mmHg at 309 days and 89 +/- 1 mmHg at 558 days; n = 6) was significantly different (P < 0.05 in all groups) from that in the control group of lambs (74 +/- 2 mmHg at 110 days; 76 +/- 1 mmHg at 323 days and 81 +/- 1 mmHg at 568 days; n = 7). However, prenatal glucocorticoid exposure did not alter vascular sensitivity to noradrenaline, angiotensin II and adrenocorticotropic hormone in these sheep at any of the ages studied, nor did it affect basal or adrenocorticotropic hormone-induced concentrations of cortisol or basal plasma renin concentrations in the lambs at any age. 4. These data support the hypothesis that excess glucocorticoid exposure in early pregnancy, during a critical developmental stage or 'window', programmes higher blood pressure that persists in later life.

Non-invasive measurements of arterial structure and function: repeatability, interrelationships and trial sample size.

Abstract: 1. Repeatability of measurements of arterial compliance and flow-mediated dilation of the brachial artery has been infrequently reported, despite increasing use in interventional and risk-factor modification studies. Furthermore, little is known about the interrelationships of the various indices. The purposes of this study were to determine the repeatability and interrelationships of a range of arterial indices.2. Fifty healthy volunteers, 20 men and 30 women, aged 20-70 (mean 46.5) years, were studied on two occasions, using an identical protocol, at a mean interval of 2.5 weeks. Tonometry, ultrasound and Doppler technique were used to measure the following: carotid wall intima-media thickness (IMT), total systemic artery compliance (SAC), arterial pulse wave velocity [PWV aorto-femoral (A-F), and femoral-dorsalis pedis (F-D)], carotid distensibility coefficient (DC) and carotid augmentation index (AI). Brachial flow-mediated dilation was measured in 30 subjects with analysis of diameter change for 4 min post ischaemia.3. There were no systematic differences over the observed range of measurements for any of the reported parameters. Coefficients of variation were as follows: IMT 2.8%, SAC 9.2%, PWV(A-F) 3.2%, PWV(F-D) 5.0%, DC 10.0%, AI 1.3%. Brachial flow-mediated dilation curves were not different between visits; changes were maximum 60-s post ischaemia. All indices of arterial compliance were significantly correlated with age. The three different indices of central arterial compliance [SAC, PWV(A-F) and AI] were significantly correlated with carotid intima-media thickness.4. Under controlled experimental conditions there was good repeatability of measurements of indices between sessions of both intrinsic and functional arterial mechanical properties (central and carotid arterial compliance, intima-media thickness and brachial flow-mediated dilation). Sample size tables for clinical trials using these indices are presented.

Increased oxidative stress in rat liver and pancreas during progression of streptozotocin-induced diabetes.

Abstract: 1. Oxygen free radicals have been suggested to be a contributory factor in complications of diabetes mellitus. There are many reports indicating the changes in parameters of oxidative stress in diabetes mellitus. In this study we aimed to identify whether oxidative stress occurs in the liver and pancreas in the initial stages of development of diabetes. 2. We therefore investigated the lipid peroxide level (thiobarbituric acid-reactive substances, TBARS) and activities of antioxidant enzymes [superoxide dismutase (SOD), catalase and glutathione peroxidase] in liver and pancreas of control and streptozotocin-induced diabetic rats at various stages of development of diabetes. 3. Male Sprague-Dawley rats were divided into two groups: group I, control (n = 42) and group II, diabetic (n = 42). Each group was further subdivided into seven groups consisting of six rats each. Rats in these subgroups were studied at weekly intervals (0 to 6 weeks). Plasma glucose levels, TBARS levels and activities of antioxidant enzymes were measured in liver and pancreas at various time intervals. 4. There was a significant (P < 0.05) and progressive increase in TBARS levels of liver and pancreas in the diabetic group. Total SOD and Cu-Zn-SOD activity increased (P < 0.05) with progression of diabetes while Mn-SOD activity showed no significant change in either tissue. Catalase and glutathione peroxidase activities increased significantly (P < 0.05) in liver and pancreas. 5. Immunohistochemical study of pancreatic islet revealed a decrease in the expression of insulin with progression of diabetes. However, glucagon and somatostatin showed an increase in immunoreactivity and a difference in their distribution pattern. 6. The findings of the present study suggest that oxidative stress starts at early onset of diabetes mellitus and increases progressively. In conclusion, the structural damage to these tissues or complications of diabetes mellitus may be due to oxidative stress.

Vascular endothelial growth factor (VEGF) is released from platelets during blood clotting: implications for measurement of circulating VEGF levels in clinical disease

Abstract: 1. Dysregulated vascular endothelial growth factor (VEGF) expression has been reported in several pathological states based upon evidence of elevated serum VEGF levels. Using two immunoassays for VEGF, this study determines normal plasma and serum VEGF ranges, determines which are more likely to reflect circulating VEGF levels and investigates a potential contribution of VEGF from platelets to VEGF levels detected in serum. 2. The presence of soluble VEGF receptor, sflt-1, at a molar excess of 7:1 significantly reduced measured VEGF levels in both assays. Serum VEGF levels were higher than plasma levels in children [(mean +/- S.E.M.) 306.1 +/- 39.4 versus 107.4 +/- 24.9 pg/ml, P < 0.0001] and adults (249.4 +/- 46.4 versus 76.1 +/- 10.7 pg/ml, P < 0.0001). Serum VEGF increased with clotting time (P = 0.0005 t0 compared with 2 h samples); plasma VEGF levels were not affected by time between sampling and centrifugation. 3. Calcium-induced clotting of platelet-rich but not platelet-poor plasma induced VEGF release with a proportional response between platelet count and VEGF level and isolated platelets released significant quantities of VEGF upon incubation with thrombin. Reverse transcriptase-PCR studies confirmed that platelets express VEGF121 and VEGF165 mRNA. 4. These data suggest that plasma is the preferred medium to measure VEGF levels; a significant and highly variable platelet-mediated secretion of VEGF during the clotting process invalidates the use of serum as an indicator of circulating VEGF levels in disease states.

Measurement and significance of circulating natriuretic peptides in cardiovascular disease.

Abstract: 1. The major cardiovascular and renal actions of alpha-atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) and the fact that the heart is strategically located to sense changes in intravascular volume indicate the importance of these peptides in the overall control of the extracellular fluid volume under normal and pathophysiological conditions.2. This review examines the clinical and diagnostic significance of the measurement of plasma natriuretic peptides in diseases of the cardiovascular system with particular emphasis on the assessment of patients with heart failure. 3. Raised plasma levels of ANP and BNP have repeatedly been found in patients with heart disease originating from diverse causes including tachycardias, valvular stenosis or ventricular dysfunction. The raised circulating levels of natriuretic peptide (ANP, N-terminal proANP and BNP in particular) are associated with (i) raised atrial and pulmonary wedge pressures; (ii) reduced ventricular systolic and diastolic function; (iii) presence (and possibly geometric form) of left ventricular hypertrophy; and (iv) severe myocardial infarction. Although both plasma ANP and BNP are raised in the presence of left ventricular hypertrophy, BNP appears to be a better index of left ventricular hypertrophy.4. Several situations where the measurement of natriuretic peptides may be of benefit in the overall assessment of heart disease are discussed. However, it is emphasized that the measurement of plasma natriuretic peptides alone appears to be of limited value as a specific diagnostic tool, given that raised levels are a consequence of haemodynamic and structural abnormalities arising from diverse pathological processes. Despite these limitations, the major value of plasma natriuretic peptides in the examination of patients with suspected heart disease rests on the premise that: (i) a normal value would not be consistent with cardiac disease; (ii) the presence of markedly raised levels may help to target those for subsequent detailed assessment of underlying cardiac dysfunction; and (iii) markedly raised levels of plasma natriuretic peptides after myocardial infarction can identify those at high risk of death.

Bone mineral changes during pregnancy and lactation: a longitudinal cohort study.

Abstract: 1. The influence of pregnancy, lactation and weaning on bone mineral density in healthy women was investigated during a 2 year prospective study of 59 pregnant and lactating women from the 18th week of gestation. 2. Bone mineral density was measured by dual energy X-ray absorptiometry at the non-dominant radius ultra distally and more proximally in the 18th and 37th weeks of gestation, and 0, 3, 6, 12 and 18 months after delivery. Measurements of bone mineral density of the lumbar spine, the proximal femur and the whole body were performed at all dates after delivery. 3. Reappearance of menstruation after delivery averaged 6.1 months; mean lactating period was 8.7 months. During pregnancy and lactation bone mineral density tended to decrease, but different measuring sites showed different patterns of bone mineral density changes. The reduction in the ultra distal radius during pregnancy amounted to 2%, and no further changes were observed here during lactation. After delivery, reduction in mean bone mineral density was most pronounced in the spine (5.2% in 3 months), but the fall in bone mass tended to revert after resumption of menstruation. Bone mineral density was still reduced by 3.3% after 12 months in women with menstruation resumption later than 8 months after delivery. No significant reduction was observed 18 months after delivery. No association with calcium intake, weight changes or initial bone mineral density was observed. High calcium intake did not protect against bone mineral loss in the spine and the femur. 4. Thus it can be concluded that bone loss during pregnancy and lactation took place mainly from the trabecular skeleton. Resumption of menstruation tended to result in a regain of bone mass towards baseline.

Skeletal progenitor cells and ageing human populations.

Abstract: 1. Stem and progenitor cells present within bone marrow give rise to colony forming units-fibroblastic (CFU-F) which can differentiate into fibroblastic, osteogenic, myogenic, adipogenic and reticular cells. The decrease in skeletal bone formation and rate of fracture repair observed with ageing and in osteoporosis has been suggested to be due to a decrease in numbers of these progenitors, but human studies are limited. 2. We have tested the potential to form CFU-F in a total of 99 patients undergoing corrective surgery (16 controls, 14-48 years of age) or hip arthroplasty for osteoarthritis (57 patients, 28-87 years of age) or osteoporosis (26 patients, 69-97 years of age). Total colony number, alkaline phosphatase-positive colony number and colony size were determined. 3. No decrease in colony forming efficiency under the culture conditions used was observed in all populations examined irrespective of age, disease or gender, as determined by the lack of correlation between colony formation and age. 3. Examination of colony sizes showed a significant reduction in colony size with age in osteoarthritis and in control populations indicating a change in cellular proliferative potential with age. 4. Examination of number and percentage of alkaline phosphatase-positive CFU-F showed a significant decrease in osteoporotic patients compared with controls and osteoarthritis patients, indicating altered differentiation potential. 5. These results suggest that the reduction in bone mass with ageing may be due to reduction of the proliferative capacity of progenitor cells or their responsiveness to biological factors leading to alteration in subsequent differentiation. The maintenance of CFU-F number and alkaline phosphatase activity in these osteoarthritis patients may, in part, explain the inverse relationship observed for the preservation of bone mass between generalized osteoarthritis and primary osteoporosis

Early Administration of Angiotensin-Converting Enzyme Inhibitor Captopril, Prevents the Development of Hypertension Programmed by Intrauterine Exposure to a Maternal Low-Protein Diet in the Rat

Abstract: 1. Associations of intrauterine exposure to maternal undernutrition with later hypertension and coronary heart disease in the human population have been duplicated in the rat. Fetal exposure to low protein diets produces offspring that develop raised systolic blood pressure by the age of weaning. This animal model of 'programmed' hypertension was used to investigate the role of the renin-angiotensin system in the initiation and maintenance of high blood pressure. 2. Pregnant rats were fed diets containing 18 or 9% casein from conception until littering. The offspring from these pregnancies were administered captopril either between 2 and 4 weeks of age, or from 10 to 12 weeks of age. 3. The feeding of low protein diets in pregnancy had no effect upon the reproductive ability of female rats and the offspring generated were of normal birthweight. By 4 weeks of age the male and female offspring of low-protein-fed dams had systolic blood pressures that were 24-25 mmHg higher than those of rats exposed to a control diet in utero. 4. Treatment of 10-week-old female offspring with captopril for 2 weeks indicated that angiotensin II formation may play a role in the maintenance of high blood pressure in low-protein-exposed rats. While captopril had no significant effect upon systolic pressures of rats exposed to the control diet in intrauterine life, the systolic blood pressures of low-protein animals rapidly declined by 31 mmHg. 5. Administration of captopril to male and female offspring between 2 and 4 weeks of age exerted long-term effects upon systolic blood pressure. Eight weeks after cessation of treatment, at an age where maximal blood pressures are achieved, captopril-treated, low-protein-exposed rats had similar blood pressures to normotensive rats exposed to the protein-replete diet in utero. 6. In conclusion, we have demonstrated that the elevation of adult blood pressure associated with fetal exposure to a maternal low-protein diet, is prevented by early administration of an angiotensin-converting enzyme inhibitor. The actions of angiotensin II in the late suckling period may be a critical determinant of long-term cardiovascular functions in these animals.

Administration of albumin to patients with sepsis syndrome: a possible beneficial role in plasma thiol repletion.

Abstract: 1. Albumin is often administered intravenously to critically ill patients as a volume expander, to combat hypoalbuminaemia, and to decrease hyperbilirubinaemia. There is, however, an ongoing debate concerning the therapeutic benefit of the former which is an expensive form of treatment. 2. Albumin has several biological functions, in particular as a ligand binder. It also acts as an extracellular transition metal ion-binding and radical-scavenging antioxidant. These functions are influenced by the presence of an exposed thiol group (cys 34) on the surface of the albumin molecule. 3. The ability of infused albumin to influence the plasma thiol pool, and hence antioxidant potential, was investigated in patients with sepsis syndrome. 4. Plasma thiol levels rose rapidly after albumin infusion and remained elevated even after plasma albumin levels had declined significantly, due to interstitial leakage. Data are suggestive of some form of thiol exchange in the plasma of these patients between albumin and molecules containing oxidized thiol groups. 5. Administration of albumin to patients with sepsis syndrome leads to a sustained increase in plasma thiols. Thiols have several important antioxidant functions, and thiol repletion in these patients, who are known to suffer from oxidative stress, may have beneficial antioxidant effects. Antioxidant repletion may represent an important facet of clinically administered albumin.

Transforming growth factor-β1 gene polymorphisms and coronary artery disease

Abstract: 1. Transforming growth factor-beta1 is a cytokine with a very wide spectrum of biological activities. Previous studies have shown that it is involved in a number of physiological and pathological processes including heart disease. In our study we aimed to scan the transforming growth factor-beta1 locus for polymorphisms and to identify haplotypes significantly associated with a predisposition to coronary atherosclerosis.2. Two patient groups comprising 244 angiographically normal individuals and 655 patients with coronary artery disease were recruited from London and Sheffield. DNA samples from these subjects were screened for mutations in the transforming growth factor-beta1 locus and all subjects were genotyped by a coupled polymerase chain reaction-restriction enzyme digestion method.3. Five polymorphisms have been identified in the transforming growth factor-beta1 gene at positions G-800A, C-509T in the promoter region, Leu10-->Pro, Arg25-->Pro in exon 1 and Thr263-->Ile in exon 5. No significant difference in frequencies for any of the five polymorphisms was found between controls and patients with coronary artery disease. Similarly, there was no correlation between these polymorphisms and hypertension.4. The genotypes of all the individuals participating in the study were assigned to seven main haplotypes of the transforming growth factor-beta1 locus. Based on species comparison data we propose that GCCGC is the ancestral haplotype in humans.5. Our data suggest that these transforming growth factor-beta1 polymorphisms are not associated with coronary artery disease and therefore their presence alone would not be a genetic risk factor for predisposition to coronary artery disease.

Importance of glutamine metabolism in murine macrophages and human monocytes to L-arginine biosynthesis and rates of nitrite or urea production.

Abstract: 1. The intermediates of biochemical cycles are commonly utilized for biosynthetic processes; thus at least one intermediate must be replenished de novo to provide constant flux through the cycle. The utilization of L-arginine for NO synthesis in macrophages may thus reduce the concentration of intermediates of the urea cycle. It is possible that a glutamine-utilizing pathway exists in mononuclear phagocytes that may connect with the urea cycle.2. In this paper we report that mouse peritoneal resident and Bacillus Calmette-Guerin (BCG)-activated macrophages and human monocytes are capable of utilizing glutamine at high rates, contain sufficient activity of the enzymes required to convert glutamine to citrulline (and subsequently citrulline to arginine) to account for observed rates of nitrite synthesis in the absence of extracellular L-arginine, and will release nitrite when exposed to intermediates of the proposed glutamine-->arginine pathway.3. The rate of nitrite production (in the absence of extracellular arginine) was reduced by culturing macrophages or monocytes in the presence of the glutaminase inhibitor 6-diazo 5-oxo norleucine.4. The rate and extent of arginase secretion, glutamine utilization, nitrite production (basal and lipopolysaccharide-stimulated) and phosphate-dependent glutaminase activity from BCG-activated macrophages was increased compared with resident cells.5. We suggest that the elevated arginase secretion rates in activated macrophages would effectively increase the intracellular concentration of arginine available for conversion to NO via inducible nitric oxide synthase, the expression of which is known to increase on activation of macrophages or monocytes. Additionally, the rate of L-arginine biosynthesis from glutamine may be increased on immunostimulation of the macrophage.

Elevated heart rate variability in physically active young and older adult women

Abstract: 1. Low heart rate variability is associated with an increased risk of cardiac sudden death, coronary heart disease and all-cause mortality. We have previously shown that physically active postmenopausal women demonstrate higher levels of heart rate variability and cardiac baroreflex sensitivity compared to their sedentary peers. The purpose of the present prospective study was to test the hypothesis that heart rate variability and cardiac baroreflex sensitivity would be reduced with age in sedentary but not physically active women. To accomplish this, we measured heart rate variability (both time and frequency domain) and spontaneous cardiac baroreflex sensitivity (SBRS, sequence method) in the sitting posture in 23 sedentary women [11 premenopausal and 12 postmenopausal (age, 28 ± 1 and 61 ± 2 years; V o 2 max, 35.3 ± 1.4 and 21.7 ± 1.5 ml · min −1 · kg −1 respectively] and in 22 physically active women [12 premenopausal and 10 postmenopausal (age, 31 ± 1 and 59 ± 2 years; V o 2 max, 52.5 ± 1.4 and 39.7 ± 1.8 ml · min −1 · kg −1 )]. 2. The S.D. of the R—R interval (time domain) was reduced ( P 2 /Hz), total power of heart rate variability and SBRS (11 ± 2 versus 7 ± 2 and 19 ± 3 versus 13 ± 2 ms/mmHg) also demonstrated similar age-related reductions in sedentary and physically active women, respectively (all P P 3. The results of the present study suggest that heart rate variability and cardiac baroreflex sensitivity decline similarly with age in healthy sedentary and physically active women. However, physically active women demonstrate higher levels of heart rate variability and cardiac baroreflex sensitivity compared with their sedentary peers, regardless of age.

Cardiovascular autonomic modulation and activity of carotid baroreceptors at altitude.

Abstract: 1. To assess the effects of acute exposure to high altitude on baroreceptor function in man we evaluated the effects of baroreceptor activation on R-R interval and blood pressure control at high altitude. We measured the low-frequency (LF) and high-frequency (HF) components in R-R, non-invasive blood pressure and skin blood flow, and the effect of baroreceptor modulation by 0. 1-Hz sinusoidal neck suction. Ten healthy sea-level natives and three high-altitude native, long-term sea-level residents were evaluated at sea level, upon arrival at 4970 m and 1 week later.2. Compared with sea level, acute high altitude decreased R-R and increased blood pressure in all subjects [sea-level natives: R-R from 1002+/-45 to 775+/-57 ms, systolic blood pressure from 130+/-3 to 150+/-8 mmHg; high-altitude natives: R-R from 809+/-116 to 749+/-47 ms, systolic blood pressure from 110+/-12 to 125+/-11 mmHg (P<0.05 for all)]. One week later systolic blood pressure was similar to values at sea level in all subjects, whereas R-R remained elevated in sea-level natives. The low-frequency power in R-R and systolic blood pressure increased in sea-level natives [R-R-LF from 47+/-8 to 65+/-10% (P<0.05), systolic blood pressure-LF from 1.7+/-0. 3 to 2.6+/-0.4 ln-mmHg2 (P<0.05)], but not in high-altitude natives (R-R-LF from 32+/-13 to 38+/-19%, systolic blood pressure-LF from 1. 9+/-0.5 to 1.7+/-0.8 ln-mmHg2). The R-R-HF decreased in sea-level natives but not in high-altitude natives, and no changes occurred in systolic blood pressure-HF. These changes remained evident 1 week later. Skin blood flow variability and its spectral components decreased markedly at high altitude in sea-level natives but showed no changes in high-altitude natives. Neck suction significantly increased the R-R- and systolic blood pressure-LF in all subjects at both sea level and high altitude.3. High altitude induces sympathetic activation in sea-level natives which is partially counteracted by active baroreflex. Despite long-term acclimatization at sea level, high-altitude natives also maintain active baroreflex at high altitude but with lower sympathetic activation, indicating a persisting high-altitude adaptation which may be genetic or due to baroreflex activity not completely lost by at least 1 year's sea-level residence.

Clinical and therapeutic significance of the Na+,K+ pump*

Abstract: 1. The Na+,K+-ATPase or Na+,K+-pump, mediating the active transport of Na+ and K+, which was first identified 40 years ago, is a central target for acute and long-term regulation, as well as for therapeutic intervention. Acute stimulation of the Na+,K+-pump in skeletal muscle by insulin, catecholamines, beta2-agonists or theophylline increases the intracellular uptake of K+ and accounts for the hypokalaemia elicited by these agents. Conversely, digitalis intoxication elicits hyperkalaemia via acute inhibition of the Na+, K+-pump. 2. Simple and accurate methods have been developed for the quantification of the total concentration of Na+,K+-pumps in small (0.5-5 mg) fresh or frozen biopsies of human skeletal muscle, myocardium or other tissues. This has allowed the identification of several long-term regulatory changes in the concentration of this transport system in human tissues. In skeletal muscle, upregulation is induced by training, thyroid hormones or glucocorticoids. Downregulation is seen in hypothyroidism, cardiac insufficiency, myotonic dystrophy, McArdle disease, K+ deficiency and after muscle inactivity. 3. Since the skeletal muscles contain one of the major pools of Na+,K+-pumps, these changes are important for the ability to counterregulate the hyperkalaemia elicited by exercise or the ingestion of K+. Moreover, downregulation or inhibition of the Na+, K+-pumps in skeletal muscle interferes with contractile performance. Since digitalis glycosides bind to the Na+,K+-pump, the muscles constitute a large distribution volume for these agents and are therefore an important determinant for their plasma level. 4. In cardiac insufficiency, the decrease in the concentration of Na+, K+-pumps in the myocardium is over a wide range correlated to the concomitant reduction in ejection fraction. The regulatory and pathophysiological changes in the activity and concentration of Na+, K+-pumps are important for the contractile function of skeletal muscle and heart as well as for K+ homoeostasis and the response to digitalization.

Whole Body Protein Metabolism in Human Pulmonary Tuberculosis and Undernutrition: Evidence for Anabolic Block in Tuberculosis

Abstract: 1. Differing patterns of protein metabolism are seen in wasting due to undernutrition and wasting due to chronic infection. 2. We investigated whole body energy and protein metabolism in nine subjects with pulmonary tuberculosis, six undernourished subjects (body mass index < 18.5 kg/m2) and seven control subjects from an Indian population. Fasting subjects were infused with L-[1-13C]leucine (2.3 mumol.h-1.kg-1) for 8 h, 4 h fasted then 4 h fed. Leucine kinetics were derived from 13C-enrichment of leucine and alpha-ketoisocaproic acid in plasma and CO2 in breath. 3. Undernourished subjects, but not tuberculosis subjects, had higher rates of whole body protein turnover per unit lean body mass than controls [163.1 +/- 9.4 and 148.6 +/- 14.6 mumol compared with 142.8 +/- 14.7 mumol leucine/h per kg, based on alpha-ketoisocaproic acid enrichment (P = 0.039)]. 4. In response to feeding, protein oxidation increased in all groups. Tuberculosis subjects had the highest fed rates of oxidation (47.0 +/- 10.5 compared with 37.1 +/- 5.4 mumol.h-1.kg-1 in controls), resulting in a less positive net protein balance in the fed phase (controls, 39.7 +/- 6.2; undernourished subjects, 29.2 +/- 10.6; tuberculosis subjects, 24.5 +/- 9.3; P = 0.010). Thus fed-phase tuberculosis subjects oxidized a greater proportion of leucine flux (33.2%) than either of the other groups (controls, 24.0%; undernourished subjects, 24.0%; P = 0.017). 5. Tuberculosis did not increase fasting whole body protein turnover but impaired the anabolic response to feeding compared with control and undernourished subjects. Such 'anabolic block' may contribute to wasting in tuberculosis and may represent the mechanism by which some inflammatory states remain refractory to nutrition support.

Subcutaneous glucagon-like peptide-1 improves postprandial glycaemic control over a 3-week period in patients with early Type 2 diabetes

Abstract: 1. Glucagon-like peptide-1 (7-36) amide (GLP-1) is released into the circulation after meals and is the most potent physiological insulinotropic hormone in man. GLP-1 has the advantages over other therapeutic agents for Type 2 diabetes of also suppressing glucagon secretion and delaying gastric emptying. One of the initial abnormalities of Type 2 diabetes is the loss of the first-phase insulin response, leading to postprandial hyperglycaemia. 2. To investigate the therapeutic potential of GLP-1 in Type 2 diabetes, six patients were entered into a 6-week, double-blind crossover trial during which each received 3 weeks treatment with subcutaneous GLP-1 or saline, self-administered three times a day immediately before meals. A standard test meal was given at the beginning and end of each treatment period. 3. GLP-1 reduced plasma glucose area under the curve (AUC) after the standard test meal by 58% (AUC, 0–240 ;min: GLP-1 start of treatment, 196±141 ;mmol·min-1·l-1; saline start of treatment, 469±124 ;mmol·min-1·l-1; F = 16.4, P < 0.05). The plasma insulin excursions were significantly higher with GLP-1 compared with saline over the initial postprandial 30 ;min, the time period during which the GLP-1 concentration was considerably elevated. The plasma glucagon levels were significantly lower over the 240-min postprandial period with GLP-1 treatment. The beneficial effects of GLP-1 on plasma glucose, insulin and glucagon concentrations were fully maintained for the 3-week treatment period. 4. We have demonstrated a significant improvement in postprandial glycaemic control with subcutaneous GLP-1 treatment. GLP-1 improves glycaemic control partially by restoring the first-phase insulin response and suppressing glucagon and is a potential treatment for Type 2 diabetes.

A novel method for the visualization of the in situ mucus layer in rat and man.

Abstract: 1. The observed thickness of the gastric mucus barrier varies widely, even appearing discontinuous, depending on the methods used. Here we describe the development and application of a modified periodic acid Schiff/Alcian Blue staining technique for use on cryostat sections of gastric mucosa. This technique for the first time enables the preservation and visualization of the full thickness of the adherent gastric mucus layer and the underlying mucosa. 2. In designing this novel method we have selected those procedures which would result in the least alteration to the mucus layer. The methods used were snap freezing, cryostat sectioning of the whole stomach followed by brief ethanol pretreatment (10 min in 100% ethanol), a prolonged staining with periodic acid Schiff/Alcian Blue (15 min and 2.5 h respectively), a gentle post-fixation (45 min paraformaldehyde vapour at 37 degreesC) and the use of a water-soluble mountant. 3. A continuous, adherent mucus layer was observed over the surface of the rat gastric mucosa (periodic acid Schiff/Alcian Blue stained) and human gastric antral biopsies (periodic acid Schiff stained). In the rat the mean (S.D.) mucus thickness measurements along the antrum to oesophageal axis (which was divided histologically into four regions, A to D) were: A, 166 (47) micrometer; B, 179 (48) micrometer; C, 184 (50) micrometer; D (the non-glandular stratified epithelium at the top of the stomach), Absent. In human gastric antral biopsies the mean (S.D.) mucus thickness was 144 (52) micrometer. 4. This new technique has enabled the visualization and precise measurement of thickness of the gastric mucus layer in rat and man. The adherent gastric mucus layer was observed to be continuous in the rat glandular stomach and human antrum. In validation experiments in rat the mean mucus thickness measurements were found to be twice those measured by conventional histological techniques, in which the mucus layer appeared discontinuous and patchy. However, they were within the range of thickness values seen in unfixed tissues and in the rat in vivo preparation.

Aldosterone blunts the baroreflex response in man

Abstract: I. Recent animal evidence suggests that aldosterone, like angiotensin II, may possess detrimental autonomic modulating properties. Aldosterone has been shown to impair the baroreflex response in animal models. This study is designed to test the hypothesis that aldosterone directly attenuates the baroreflex in vivo in man. 2. Fourteen healthy male volunteers [mean age (S.D.) 25 (9) years] received intravenous d-aldosterone (1 2 pmol min kg 1 ) and 5% dextrose (vehicle) in a double-blind crossover fashion, co-infused with incremental doses of intravenous phenylephrine and sodium nitroprusside. Aldosterone had no significant effect on resting blood pressure, heart rate or baroreflex response to sodium nitroprusside. However, reflex responses to phenylephrine were impaired with aldosterone (P < 0.01) while blood pressure responses were unaltered. Baroreflex sensitivity was significantly blunted in the aldosterone group [8.36 ± 2.19 versus 10.12 ± 2.27 ms/mmHg; P < 0.04]. 3. This study confirms previous observations from animal models that aldosterone impairs the baroreflex response. High aldosterone levels may contribute to the baroreflex dysfunction in cardiovascular diseases such as hypertension and heart failure.

Rate of gastric emptying is a determinant of postprandial hypotension in non-insulin-dependent diabetes mellitus

Abstract: 1. Postprandial hypotension is now recognized as an important clinical problem, particularly in the elderly and in patients with autonomic dysfunction. The mechanisms responsible are poorly understood; however, impaired regulation of splanchnic blood flow and the release of gastrointestinal hormones appear to be important. The effect of gastric emptying on the magnitude of the postprandial fall in blood pressure has not been evaluated. 2. The aim of this study was to determine whether there is a relationship between changes in blood pressure and the rate of gastric emptying after ingestion of 75 g of glucose in patients with non-insulin-dependent diabetes mellitus (NIDDM) and both young and older normal subjects. Sixteen patients with recently diagnosed NIDDM, median age 57 (39-79) years, 10 'young' subjects with a median age of 23 (19-26) years and nine 'older' subjects, median age 48 (40-68) years, were measured simultaneously for gastric emptying of 75 g of glucose in 350 ml of water blood pressure and blood glucose concentrations, commencing at approximately 10.00 hours after an overnight fast. Measurements of blood pressure were made in the sitting position immediately before glucose ingestion and at 15 min intervals up to 180 min. 3. Gastric emptying of glucose was not significantly different between the three groups [50% emptying time (T50): 95 +/- 7.3 min in patients with NIDDM compared with 120 +/- 13.2 min in the 'young' group and 97 +/- 8.1 min in the 'older' group]. There was a significant fall in mean blood pressure after the glucose load in the patients with NIDDM (P or = 20 mmHg) was evident in seven (44%) patients with NIDDM and three (33%) 'older' normal subjects. The area under the change in mean blood pressure curve was related significantly to the gastric emptying T50 (r = 0.67, P < 0.005) in the patients with NIDDM, but not in either control group. 4. In conclusion, in patients with recently diagnosed NIDDM the fall in blood pressure after an oral glucose load is (i) greater than in both young and older normal subjects and (ii) related to the rate of gastric emptying.

Sustained modifications of protein metabolism in various tissues in a rat model of long-lasting sepsis.

Abstract: 1. Sepsis was induced in rats by an intravenous injection of live bacteria. Infected and pair-fed animals were studied before the infection, in an acute septic phase (day 2 post-infection), in a chronic septic phase (day 6) and in a late septic phase (day 10). Protein synthesis rates were measured in vivo after administration of a flooding dose of l[1- 13 C]valine. 2. During the acute phase, muscle protein loss associated with infection resulted from both a decrease in protein synthesis and an increase in proteolysis. During the chronic phase and the late phase, the increase of proteolysis in infected rats as compared with pair-fed animals persisted, worsening muscle atrophy. Skin protein synthesis rates were not significantly modified by infection. However, skin protein content decreased 6 and 10 days after infection, suggesting an increased proteolysis in response to sepsis. 3. Protein synthesis in liver of infected rats was twice that of pair-fed animals. Liver protein synthesis remained elevated in infected rats compared with pair-fed animals until day 10. Hypoalbuminaemia and high plasma concentrations of fibrinogen were evident at all periods studied. α 2 -Macroglobulin and α 1 -acid glycoprotein reached peak concentrations during the acute phase (concentrations increased 50 times in infected rats). On day 10, the levels of these proteins were still about 12-fold higher. 4. Protein synthesis rates were significantly increased in the digestive tract and lung of infected rats compared with pair-fed groups on days 2 and 6, but were similar in the two groups on day 10 postinfection. The fractional protein synthesis rate was increased 3-fold over the entire experimental period in the spleen. 5. The results show that sepsis stimulates protein synthesis in various tissues over a long time, and that skin, like muscle, can provide amino acids to the rest of the body.

Subcutaneous glucagon-like peptide-1 (7-36) amide is insulinotropic and can cause hypoglycaemia in fasted healthy subjects

Abstract: 1.Glucagon-like peptide-1 (7-36) amide (GLP-1) is a gut hormone released postprandially that stimulates insulin secretion, suppresses glucagon secretion and delays gastric emptying. The insulinotropic action of GLP-1 is more potent under hyperglycaemic conditions. Several published studies have indicated the therapeutic potential of subcutaneous GLP-1 in non-insulin-dependent (Type 2) diabetes mellitus.2. We investigated whether subcutaneous GLP-1, at a dose shown to improve glycaemic control in early Type 2 diabetes, is insulinotropic at normal fasting glucose concentrations. A double-blind, randomized, crossover study of 10 healthy subjects injected with GLP-1 or saline subcutaneously after a 16 h fast was performed. The effect on cardiovascular parameters was also examined. 3.GLP-1 caused a near 5-fold rise in plasma insulin concentration. After treatment with GLP-1, circulating plasma glucose concentrations fell below the normal range in all subjects. One subject had symptoms of hypoglycaemia after GLP-1. A rise in pulse rate was found which correlated with the fall in plasma glucose concentration. An increase in blood pressure occurred with GLP-1 injection which was seen at the same time as the rise in plasma GLP-1 concentrations.4. This study indicates that subcutaneous GLP-1 can override the normal homoeostatic mechanism maintaining fasting plasma glucose in man, and is also associated with an increase in blood pressure.

Randomized clinical trial comparing the effectiveness of two dietary interventions for patients with hyperlipidaemia

Abstract: 1. Intervention trials in free-living populations have shown relatively small reductions in risk factors for cardiovascular disease, including lipid levels, and have led some to question whether diet is an effective treatment for hyperlipidaemia. However, behaviour change is a complex process and it is possible that standard intervention methods fail to motivate people sufficiently to comply with dietary advice.2. This study applied motivational interviewing, a style of behaviour change counselling, to dietary education for people with hyperlipidaemia. One-hundred and twenty-one patients with hyperlipidaemia who had been referred to a hospital dietetic department for dietary advice were randomized to receive either standard or motivational dietary interventions for a period of 3 months. Outcomes assessed included dietary knowledge, stage of change, dietary intakes, lipid levels and body mass indices. 3. From baseline, both methods of dietary intervention resulted in self-reported changes in dietary habits and knowledge, statistically significant reductions in intake of total fat (from 32.8% to 28.4%), saturated fat (from 11.4% to 9.2%) and energy intakes [-239 kcal (-999.98 kJ)/day], and in body mass indices (-0.45 kg/m2). Serum cholesterol did not change significantly in either intervention group.4. Motivational and standard dietary interventions achieved statistically significant changes in reported dietary knowledge and behaviour, and led to a reduction in body weight, but not serum cholesterol. Whether this lack of effect is real or due to subjects overestimating true dietary change cannot be determined. Change in body weight was associated with a reported change in energy intake; this provides some support for there having been a real change in intake.

Just how big is the pulmonary granulocyte pool

Abstract: 1. It is widely believed that a large fraction of the blood granulocyte population is located in the pulmonary circulation. 2. Evidence in favour of this belief is based on several independent and complementary techniques including studies of granulocyte deformability in relation to pulmonary capillary diameter, isolated perfused lungs, direct videomicroscopic observations, cellular and capillary morphometry, physiological studies and labelled blood cell kinetics. 3. Inconsistencies in this body of evidence can be identified and traced in many cases to doubts concerning the physiological integrity of labelled granulocytes. 4. In addition to the lungs, other regions of the body undoubtedly pool granulocytes, and there is good quantitative evidence, based mainly on clinical studies, pointing to the liver, bone marrow and especially the spleen. 5. Clinical studies, furthermore, have generally not supported major granulocyte pooling in the lung, except in association with systemic inflammatory diseases, such as inflammatory bowel disease and systemic vasculitis. 6. Compromises based on technical considerations permit reconciliation of the literature, and an overall whole-body model of granulocyte distribution is proposed with which all the existing data are broadly compatible.

Plasma leptin in chronic inflammatory bowel disease and HIV : implications for the pathogenesis of anorexia and weight loss

Abstract: 479 1. Leptin inhibits food intake and is an important regulator of long-term energy balance. In rodents, plasma concentrations of leptin are increased by administration of interleukin-1 and tumour necrosis factor. Hyperleptinaemia may mediate the anorexia and weight loss which is observed in chronic infections and inflammatory conditions. 2. Plasma leptin and soluble tumour necrosis factor receptor (sTNF-r55) concentrations were measured in patients with inflammatory bowel disease and acquired immunodeficiency syndrome (AIDS), and healthy controls. 3. The patients with AIDS were severely wasted lo/. body fat 12 (S16); median (interquartile range)] compared with those with inflammatory bowel disease (25.1 (1%31.5)1 and control subjects (29.4 (23.6-37.8)]. Leptin concentrations were highly correlated with percentage body fat in controls (r = 0.74, P < 0.001) and patients with IBD (r = 0.73, P < 0.001) but not in the patients with AIDS (r = -0.024). Leptin concentrations were similar in the inflammatory bowel disease (4.8 (2.6-10.1) ng/ml] and control groups 18.0 (3.1-14.1) ng/ml] but were significantly lower (P < 0.05) in patients with AIDS 11.8 (1.5-2.3) ng/mll after 23 patients were matched for sex and percentage body fat in patients with inflammatory bowel disease 12.4 (1.8-4.1) ng/ml]. Plasma concentrations of sTNF-r55 were higher in both the patients with inflammatory bowel disease 10.19 (0.16-0.23) ng/ml] and those with AIDS 14.8 (2.8-7.3) ng/ml] compared with controls 10.14 (0.09-0.16) ng/mll but were not correlated with either percentage body fat or plasma leptin concentrations. 4. Hyperleptinaemia does not appear to mediate the anorexia and weight loss associated with inflammatory bowel disease and AIDS. In patients with AIDS with extreme wasting there was no relationship between body fat and leptin and this may be related to the rapid weight loss which occurs in these patients.

Dietary Fish Oil Suppresses Human Colon Tumour Growth in Athymic Mice

Abstract: 1. Human colon tumour growth, initiated by subcutaneous inoculation of HT29 cells, was measured in athymic mice fed ad libitum on high-fat (210 g/kg) diets rich in coconut oil (CO), olive oil (OO), safflower oil (SO) or fish oil (FO); a low fat (LF; 25 g/kg) diet was used as the control. In one experiment the mice were fed the experimental diets for 3 weeks before HT29 cell inoculation and were killed 2 weeks post-inoculation. In a second experiment the mice were maintained on the LF diet until 4 days post-HT29 cell inoculation; they were then fed the experimental diets for 17 days. 2. Compared with mice fed the LF diet, tumour size was increased in mice fed the CO, OO or SO diets for 3 weeks before HT29 cell inoculation; FO feeding did not significantly increase tumour size. 3. Feeding mice the CO or OO diets from 4 days post-inoculation increased tumour growth rate and tumour size compared with feeding the LF, SO or FO diets; tumour growth rate and size did not differ among mice fed the latter diets. 4. The fatty acid composition of the tumours was markedly influenced by the fatty acid composition of the diet. 5. We conclude that human colon tumour growth is influenced by the type of fat consumed in the diet. Human colon tumour growth in this model is promoted by feeding high fat diets rich in medium chain saturated fatty acids (CO) or monounsaturated fatty acids (OO). A high fat diet, rich in long chain n — 3 polyunsaturated fatty acids (FO), does not promote colon tumour growth. The effect of a high fat diet rich in n — 6 polyunsaturated fatty acids (SO) depends upon the time at which it is fed: if fed before tumour cell inoculation such a diet promotes tumour growth, whereas if fed once tumour growth is initiated it does not. This suggests that n — 6 polyunsaturated fatty acids promote the initiation of colon tumour growth, but do not exert growth-promoting effects on colon tumours once they are established.

Fractal dimension and approximate entropy of heart period and heart rate: awake versus sleep differences and methodological issues

Abstract: 1. Investigations that assess cardiac autonomic function include non-linear techniques such as fractal dimension and approximate entropy in addition to the common time and frequency domain measures of both heart period and heart rate. This article evaluates the differences in using heart rate versus heart period to estimate fractal dimensions and approximate entropies of these time series.2. Twenty-four-hour ECG was recorded in 23 normal subjects using Holter records. Time series of heart rate and heart period were analysed using fractal dimensions, approximate entropies and spectral analysis for the quantification of absolute and relative heart period variability in bands of ultra low (<0.0033 Hz), very low (0. 0033-0.04 Hz), low (0.04-0.15 Hz) and high (0.15-0.5 Hz) frequency.3. Linear detrending of the time series did not significantly change the fractal dimension or approximate entropy values. We found significant differences in the analyses using heart rate versus heart period between waking up and sleep conditions for fractal dimensions, approximate entropies and absolute spectral powers, especially for the power in the band of 0.0033-0.5 Hz. Log transformation of the data revealed identical fractal dimension values for both heart rate and heart period. Mean heart period correlated significantly better with fractal dimensions and approximate entropies of heart period than did corresponding heart rate measures.4. Studies using heart period measures should take the effect of mean heart period into account even for the analyses of fractal dimension and approximate entropy. As the sleep-awake differences in fractal dimensions and approximate entropies are different between heart rate and heart period, the results should be interpreted accordingly.

Bronchial reactivity and intracellular magnesium: a possible mechanism for the bronchodilating effects of magnesium in asthma.

Abstract: 1. Increased bronchial smooth muscle contractility with consequent bronchial hyperreactivity are characteristic physiopathological events of asthma. Since magnesium intervenes in calcium transport mechanisms and intracellular phosphorylation reactions, it constitutes an important determinant of the contraction/relaxation state of bronchial smooth muscle. In the present study we investigated the relationship between bronchial reactivity, assessed by methacholine-provocation test, and magnesium concentrations both at extracellular and intracellular levels measured by spectrophotometry. Twenty-two patients with mild-to-moderate asthma and 38 non-asthmatic subjects with allergic rhinitis (24 allergic to Parietaria pollen and 14 allergic to Grass pollen) were recruited to the study. Exclusion criteria included renal failure, hepatic diseases, heart failure and arterial hypertension. 2. The salient finding of our study is that there is a strong positive correlation between bronchial reactivity and the level of intracellular magnesium (r=0.72, P<0.0001), both when the groups are analysed separately or together. Intracellular magnesium concentrations in the group of patients with asthma were significantly lower (1.8+/-0. 01 mmol/l; n=22) when compared with levels in rhinitis subjects allergic to Parietaria (1.9+/-0.01 mmol/l; n=24, P<0.05), and with levels in rhinitis subjects allergic to Grass pollen (2.0+/-0.03 mmol/l; n=14, P<0.05). Serum levels of the ion were similar in all groups. 3. We conclude that the level of intracellular magnesium may be an important determinant of bronchial hyperreactivity, as supported by the significant positive correlation between these two parameters in allergic patients with known bronchial hyperresponsiveness. This finding, in addition to reports of the bronchodilating effects of magnesium administration in patients with asthma, confirms the proposed role of this ion in the pathogenesis and treatment of asthma.