Effect of bolus fluid intake on energy expenditure values as determined by the doubly labeled water method.

The doubly labeled water (DLW, 2H(2)18O) method is a highly accurate method for measuring energy expenditure (EE). A possible source of error is bolus fluid intake before body water sampling. If there is bolus fluid intake immediately before body water sampling, the saliva may reflect the ingested water disproportionately, because the ingested water may not have had time to mix fully with the body water pool. To ascertain the magnitude of this problem, EE was measured over a 5-day period by the DLW method. Six subjects were dosed with 2H2(18)O. After the reference salivas for the two-point determination were obtained, subjects drank water (700-1,000 ml), and serial saliva samples were collected for the next 3 h. Expressing the postbolus saliva enrichments as a percentage of the prebolus value, we found 1) a minimum in the saliva isotopic enrichments were reached at approximately 30 min with the minimum for 2H (95.48 +/- 0.43%) being significantly lower than the minimum for 18O (97.55 +/- 0.44, P less than 0.05) and 2) EE values calculated using the postbolus isotopic enrichments are appreciably higher (19.9 +/- 7.5%) than the prebolus reference values. In conclusion, it is not advisable to collect saliva samples for DLW measurements within approximately 1 h of bolus fluid intake.     

Determinants of Free-Living Energy Expenditure in Normal Weight and Obese Women Measured by Doubly Labeled Water

Total free-living energy expenditure (TEE) was measured in 9 normal weight controls and 5 obese women using the doubly labeled water (DLW) method. Resting energy expenditure (REE) and the thermic effect of food (TEF) were measured by indirect calorimetry and the energy cost of physical activity (PA) calculated by deduction, in order to quantify the components and identify determinants of free-living TEE. Although REE was quantitatively the major component of TEE in both groups, PA best explained the variability, contributing 76% to the variance in free-living TEE. The obese women had elevated values for TEE (12397+/-2565 vs. 8339+/-1787 kJ/d, mean+/-SD; p<0.00S), compared with the control women. PA (5071+/-2385 vs. 2552+/-1452; p<0.0S) and REE (6393+/-678 vs. 5084+/-259; p<0.000S) were also raised in the obese, whereas TEF was not significantly different between the groups, accounting for 7.6% of energy expenditure for the obese and 8% for the control subjects. Body weight was the single best determinant of mean daily free-living TEE across both groups. We conclude that PA and body weight are the main determinants of free-living TEE .     

Validation of the doubly labeled water method in rats during isolation and simulated weightlessness

Total energy expenditure (TEE) of rats during simulated microgravity is unknown. The doubly labeled water method (DLW) reliably measures TEE, but the results depend on the methods of calculation. These methods were validated and appraised by indirect calorimetry in eight rats during isolation (7 days) and simulated microgravity (10 days). There were no effects on CO(2) production in the method used to derive constant flux rates as in the regression models. r(CO(2)) estimates were dependent on the assumed fractionation processes, the derivation of constant flux rate methods, and the selected pool models. Use of respiratory or food quotients did not influence TEE estimations, which were similar during isolation and simulation. During either isolation with growth or simulation with a stabilized mass, the one-pool model of Speakman (Speakman JR. Doubly Labelled Water. Theory and Practice. London: Chapman and Hall, 1997) resulted in the more reliable validation (0.8 +/- 2.2 and 2.2 +/- 3.4% vs. calorimetry, respectively). However, during simulation, agreement was also observed with the single pool model of Lifson (Lifson N, Gordon GB, and McClintock R. J Appl Physiol 7: 704-710, 1955) (-2.5 +/- 2.5%), and two two-pool models [Schoeller (Schoeller DA. J Nutr 118: 1278-1289, 1988) (0.5 +/- 3.1%) and Speakman (Speakman, JR. Doubly Labelled Water. Theory and Practice. London: Chapman and Hall, 1997) (-1.9 +/- 2.7%)]. This latter finding seems linked to the stable body mass and to fractionation consideration close to the single-pool model of Speakman. During isolation or simulated microgravity, the other equations underestimated TEE by 10-20%.     

Blood-sucking bugs as a gentle method for blood-collection in water budget studies using doubly labelled water

During doubly-labelled water (DLW) experiments, blood collection by venous puncture may traumatize animals and consequently affect the animals' behaviour and energy budget. Recent studies have shown that blood-sucking bugs (Triatominae; Heteroptera) can be used instead of conventional needles to obtain blood from animals. In this paper, we validate the bug method in captive nectar-feeding bats, Glossophaga soricina, for water budget analysis by comparing the daily water flux estimated with the DLW method with values measured by an energy balance method. As the mean daily water flux of the DLW method was not significantly deviating from the expected value, blood-sucking bugs may substitute more invasive methods of blood collection in DLW experiments. Based on the DLW estimates, daily energy and water intake rates were calculated and compared to values measured with the energy balance method. The DLW method and the energy balance method yielded on average similar results regarding the daily energy intake (DLW method: 48.8+/-14.2 kJ d(-1) versus energy balance method: 48.1+/-9.9 kJ d(-1)) and daily water intake (DLW method: 13.7+/-2.4 mL d(-1) versus energy balance method: 14.7+/-3.0 mL d(-1)). Based on the calculated water and sugar intake per day, we estimated the sugar concentration of ingested nectar to equal on average 16.2+/-2.4% (mass/mass), which fell close to the measured sugar concentration of 17% (mass/mass) bats fed on during the experiment. We conclude that it is possible to extrapolate mean daily energy and water intake for animal groups, populations and species based on DLW estimates, but due to the large variance of results (low accuracy), it seems inadequate to calculate values for single individuals.     

Influence of body composition on physical activity validation studies using doubly labeled water.

This study investigated the influence of two approaches (mathematical transformation and statistical procedures), used to account for body composition [body mass or fat-free mass (FFM)], on associations between two measures of physical activity and energy expenditure determined by doubly labeled water (DLW). Complete data for these analyses were available for 136 African American (44.1%) and Hispanic (55.9%) women (mean age 50 +/- 7.3 yr). Total energy expenditure (TEE) by DLW was measured over 14 days. Physical activity energy expenditure (PAEE) was computed as 0.90 x TEE - resting metabolic rate. During week 2, participants wore an accelerometer for 7 consecutive days and completed a 7-day diary. Pearson's product-moment correlations and three statistical procedures (multiple regressions, partial correlations, and allometric scaling) were used to assess the effect of body composition on associations. The methods-comparison analysis was used to study the effect of body composition on agreement. The statistical procedures demonstrated that associations improved when body composition was included in the model. The accelerometer explained a small but meaningful portion of the variance in TEE and PAEE after body mass was accounted for. The methods-comparison analysis confirmed that agreement with DLW was affected by the transformation. Agreement between the diary (transformed with body mass) and TEE reflected the association that exists between body mass and TEE. These results suggest that the accelerometer and diary accounted for a small portion of TEE and PAEE. Most of the variance in DLW-measured energy expenditure was explained by body mass or FFM.     

Doubly labeled water measurement of human energy expenditure during strenuous exercise

The energy expenditures (EE) of 23 adult male Marines were measured during a strenuous 11-day cold-weather field exercise at 2,200- to 2,550-m elevation by both doubly labeled water (2H2 18O, DLW) and intake balance methods. The DLW EE calculations were corrected for changes in baseline isotopic abundances in a control group that did not receive 2H2 18O. Intake balance EE was estimated from the change in body energy stores and food intake. Body energy-store changes were calculated from anthropometric [-1,574 +/- 144 (SE) kcal/day] and isotope dilution (-1,872 +/- 293 kcal/day) measurements made before and after the field exercise. The subjects kept daily logbook records of ration consumption (3,132 +/- 165 kcal/day). Mean DLW EE (4,919 +/- 190 kcal/day) did not differ significantly from intake balance EE estimated from food intake and either anthropometric (4,705 +/- 181 kcal/day) or isotope dilution (5,004 +/- 240 kcal/day) estimates of the change in body energy stores. The DLW method can be used with at least the same degree of confidence as the intake balance method to measure the EE of active free-living humans.     

Assessment of physical activity in older individuals: a doubly labeled water study.

We compared the accuracy of two physical activity recall questionnaires and a motion detector in 45- to 84-yr-old women (n = 35) and men (n = 32), using doubly labeled water (DLW) in conjunction with indirect calorimetry as the criterion measure. Subjects were administered the Yale Physical Activity Survey (YPAS) and Minnesota Leisure Time Physical Activity Questionnaire (LTA). Physical activity energy expenditure was determined over a 10-day period by using a Caltrac uniaxial accelerometer and DLW in conjunction with indirect calorimetry. In older women, Minnesota LTA (386 +/- 228 kcal/day) and Caltrac (379 +/- 162 kcal/day) underestimated physical activity by approximately 55% compared with DLW (873 +/- 244 kcal/day). No difference was observed between daily physical activity measured by the YPAS (863 +/- 447 kcal/day) and DLW in older women. In older men, Minnesota LTA (459 +/- 288 kcal/day) and Caltrac (554 +/- 242 kcal/day) underestimated daily physical activity by approximately 50-60% compared with DLW (1,211 +/- 429 kcal/day). No difference was found between physical activity measured by the YPAS (1,107 +/- 612 kcal/day) and DLW in older men. Despite no difference in mean physical activity levels between YPAS and DLW in women and men, Bland and Altman (Lancet 1: 307-310, 1986) analyses demonstrated poor concordance between DLW and YPAS (i.e., limits of agreement = -1,310-1,518 kcal/day). Our data suggest that the Minnesota LTA recall and Caltrac uniaxial accelerometer may significantly underestimate free-living daily physical activity energy expenditure in older women and men. Although the YPAS compares favorably with DLW on a group basis, its use as a proxy measure of individual daily physical activity energy expenditure may be limited in older women and men.     

Comparison of methods for evaluating energy expenditure of incubating wandering albatrosses.

Measurements of incubation energetics can vary depending on the method used to measure metabolism of an incubating bird. Therefore, we evaluated the energy expenditure of six male and four female wandering albatrosses (Diomedea exulans Linnaeus) using doubly labeled water (DLW), the rate of mass loss, and estimates of metabolic water production derived from water influx rate (WIR). Incubation metabolic rates (IMR) determined with DLW (169+/-21 kJ x kg(-1) x d(-1) SD) were significantly lower than estimates derived from mass loss (277+/-46 kJ x kg(-1) x d(-1) SD) and WIR (males=289+/-60 kJ x kg(-1) x d(-1) vs. females=400+/-69 kJ x kg(-1) x d(-1) SD). Estimates of IMR from mass loss and WIR were similar to IMR (305+/-39 kJ x kg(-1) x d(-1) SD) determined by respirometry in a previous study, and IMR from DLW was similar to estimates based on heart rate (HR; 147+/-26 kJ x kg(-1) x d(-1) SD) determined in another study. Applying the different measurements of IMR to construct an energy budget, we estimate that a breeding pair of wandering albatrosses spends 124-234 MJ to incubate the egg for 78 d. Finally, IMRs determined with DLW and HR were similar to estimated basal metabolic rates derived from six different allometric equations, suggesting that heat production from adult maintenance metabolism is sufficient to incubate the egg.     

Canadian recommendations underestimate energy needs of women over fifty years as determined by doubly-labelled water

Accurate estimations of energy requirements at the population level are crucial because of disease processes associated with energy imbalance. The present objective was to compare energy expenditure with existing Recommended Nutrient Intakes for Canadians (RNIC) and determine whether the RNIC provides a true index of energy requirement in middle-aged and elderly Canadian women. A second objective was to compare energy expenditure and the RNIC to Food and Agriculture Organization, World Health Organization, United Nations University (FAO/WHO/UNU) predictions. Seventy-six women were recruited for the study (67.3 +/- 11.5 y, 63 +/- 11.7 kg, BMI 24.8 +/- 4.4 kg x m(-2)). The two-point doubly-labelled water (DLW) method was used over 13 days to assess energy expenditure while subjects carried out their routine activities. Subjects were stratified to enable age specific requirements for middle-aged and elderly women. At weight maintenance, energy needs were underestimated using the RNIC (7.1 +/- 1.6 MJ x d(-1), 1698 +/- 391 kcal x d(-1)) compared to total energy expenditure (10.0 +/- 3.2 MJ x d(-1), 2395 +/- 746 kcal x d(-1)) as determined by DLW as a whole and for each age group. The RNIC recommendations were lower than the FAO/WHO/UNU estimations even for light activity. Results indicate that mean energy expenditure was 29% greater than the RNIC recommendations created using formulas based on age and weight, whereas the FAO/WHO/ UNU estimations closely approximated energy expenditure based on heavy activity in women 49-79 y and light activity in women over 80 y old. These data suggest a systematic underestimation of Canadian energy recommendations for women.     

Energy Expenditure Compared to Physical Activity Measured by Accelerometry and Self-Report in Adolescents: A Validation Study

Background

Physical inactivity is responsible for 5.3 million deaths annually worldwide. To measure physical activity energy expenditure, the doubly labeled water (DLW) method is the gold standard. However, questionnaires and accelerometry are more widely used. We compared physical activity measured by accelerometer and questionnaire against total (TEE) and physical activity energy expenditure (PAEE) estimated by DLW.

Methods

TEE, PAEE (TEE minus resting energy expenditure) and body composition were measured using the DLW technique in 25 adolescents (16 girls) aged 13 years living in Pelotas, Brazil. Physical activity was assessed using the Actigraph accelerometer and by self-report. Physical activity data from accelerometry and self-report were tested against energy expenditure data derived from the DLW method. Further, tests were done to assess the ability of moderate-to-vigorous intensity physical activity (MVPA) to predict variability in TEE and to what extent adjustment for fat and fat-free mass predicted the variability in TEE.

Results

TEE varied from 1,265 to 4,143 kcal/day. It was positively correlated with physical activity (counts) estimated by accelerometry (rho  = 0.57; p = 0.003) and with minutes per week of physical activity by questionnaire (rho  = 0.41; p = 0.04). An increase of 10 minutes per day in moderate-to-vigorous intensity physical activity (MVPA) relates to an increase in TEE of 141 kcal/day. PAEE was positively correlated with accelerometry (rho  = 0.64; p = 0.007), but not with minutes per week of physical activity estimated by questionnaire (rho  = 0.30; p = 0.15). Physical activity by accelerometry explained 31% of the vssariability in TEE. By incorporating fat and fat-free mass in the model, we were able to explain 58% of the variability in TEE.

Conclusion

Objectively measured physical activity significantly contributes to the explained variance in both TEE and PAEE in Brazilian youth. Independently, body composition also explains variance in TEE, and should ideally be taken into account when using accelerometry to predict energy expenditure values.     

Validation of water flux and body composition in glaucous gulls (Larus hyperboreus)

Water influx rates (WIR) measured with tritiated water dilution were compared with direct measures of water and energy intake in glaucous gulls (Larus hyperboreus). Total body water (TBW) measured isotopically was also compared with TBW determined by body composition analysis (BCA) of the same birds. Seventeen wild gulls were captured and studied in outdoor enclosures at Ny-Alesund, Svalbard, in July 2002. Gulls were hand-fed known quantities of Arctic cod (Boreogadus saida) or given water on the basis of one of four experimental treatments: (A) fasting, (B) fish only, (C) water only, or (D) fish and water. Water and energy content of Arctic cod was also determined. WIR of gulls (after subtracting metabolic water production) in treatments A, B, C, and D were 0, 101 +/- 5, 62 +/- 19, and 122 +/- 21 SD g d(-1), respectively. Measured water intake in each group was 0, 111 +/- 2, 64 +/- 3, and 134 +/- 15 SD g d(-1), respectively. On average, WIR underestimated measured water intake in each group. Errors were lowest but most variable for gulls fed water only (-2.2% +/- 32.8%) compared with gulls fed fish only (-9.0% +/- 5.4%) or fish and water (-9.0% +/- 7.0%). Compared with measured water intake, errors in WIR were relatively low overall (-6.9% +/- 17.4%) and comparable to previous validation studies. The difference in TBW determined by BCA versus isotopic dilution ranged between -1.02% and +8.59% of mass. On average, TBW measured isotopically (632 +/- 24 g kg(-1)) overestimated true body water by a factor of 1.033.     

Validation of the doubly‐labeled water (DLW) method for estimating CO2 production and water flux in growing poultry chicks

This study is the first validation of the doubly‐labeled water (DLW) method on birds (1) to evaluate the accuracy of 2 points versus multiple points for computing fractional isotopic washout rates (k) and CO₂ production (rCO₂), (2) to measure CO₂ production and water flux each day over a 4‐day period, (3) to compare measured fractional evaporative water loss (r_G) with assumed values that provide DLW estimates of rCO₂ with zero error, and (4) to measure the effect of assumed r_G on the error of estimating water influx and efflux. Percent error of CO₂ production of six growing poultry chicks estimated by the DLW method was not correlated with mean daily relative growth rates of up to 5% nor with daily rates of energy retained in growth of up to 320 kJ/day/kg, nor was it significantly reduced by using multiple points (5 points) rather than 2 points to compute fractional isotopic washout rates (k) and isotope pool sizes. Its seems clear from our study and the previous 5 validations on growing birds that average relative daily growth rates of up to about 20% do not increase the error of estimating rCO₂ by the DLW method. Arithmetic error was significantly less when using one isotopic pool, rather than two pools, to compute rCO₂ and was less when using an assumed fractional evaporative water loss (r_G) of 0.45 rather than an assumed r_G of 0.25 or 0.5 (the two values used predominantly in previous DLW studies). Our study supports Speakman's (1997) suggestion that the one‐pool model is more appropriate than the two‐pool model for birds weighing<1 kg. We recommend using an assumed r_G of 0.45 to compute rCO₂ of poultry, which is a compromise between the two schools of r_G useage, i.e., r_G=0.25 or 0.5, however we hesitate to recommend 0.45 for all birds in all settings. Close agreement between measured r_G and an assumed r_G that produced zero rCO₂ error supports the validity of using the pooled fractionation correction factors (f_pool) of 0.0339 for tritiated water and 0.0249 for deuterated water. Absolute error decreased with the percent washout of during measurement periods of 1 to 4 days. Accuracy of estimating rCO₂ was not significantly different for durations of 2, 3, and 4 days using either tritiated or deuterated water. The arithmetic error of estimating rCO₂ using a one isotopic pool model, 2 points, an r_G of 0.5, and tritiated water was ?1.9% (SD=13.5) for the first day of a 4‐day period and ?4.0% (SD=8.9) for the entire period. Percent arithmetic error of water influx (rH₂O_inf) and efflux (rH₂O_eff) estimated for day 1 from tritiated water washout and an assumed r_G of 0.5 was ?0.5 (SD=6.4) and 0.1% (SD=11.1), respectively. An r_G of 0.5 produced significantly less arithmetic error than an r_G of 0.25 or an r_G of zero (i.e., no fractionation correction), and less absolute error in rH₂O_inf. Errors were slightly more negative (underestimates) with an r_G of 0.25, i.e., ?2.2 and ?2.0%, respectively and even more negative with no correction for isotopic fractionation (i.e., an assumed r_G of zero). Tritiated water estimates of water influx and efflux during the first day had no error when using an r_G of 0.57 and 0.48, respectively. With assumed r_Gs of 0.25 and 0.5, the errors of water influx were ?7.8 and ?5.9%, and the errors of water efflux were 3.4 and 5.6%, respectively, over 4 days. We recommend using an assumed r_G of 0.45 to compute rH₂O_eff for poultry. The error of rCO₂ was about 3 to 4 times more sensitive to values of assumed r_G than the error of water flux.     

Validation of the doubly labeled water technique for measuring energy metabolism in starlings and sparrows

I have tested the idea that doubly labeled water (DLW) can accurately predict CO2 production in savannah sparrows, song sparrows, white-throated sparrows, starlings, and a single house sparrow by comparing DLW estimates with those obtained simultaneously by capturing expired CO2 in Ascarite. In addition I used the energy balance method to see if metabolic rates generated from DLW measurements accurately reflected the actual metabolic rates of these birds. I found close agreement in DLW and the gravimetric and energy balance methods, with DLW underestimating CO2 production on average by -3.5% in sparrows, and -7.1% in starlings. Similarly, the energy balance method indicated a -3.1% underestimate by DLW for sparrows and a -5.1% for starlings. The DLW method can yield reasonable estimates of CO2 production in a variety of passerine birds.     

Combination of high-performance liquid chromatography and radioimmunoassay for the measurement of urodilatin and alpha-hANP in the urine of healthy males

Urodilatin (ANP-(95-126)), a natriuretic peptide in urine, and alpha-hANP (ANP-(99-126)) are crossreactive in the radioimmunoassay of alpha-hANP (ANP-RIA). We therefore developed a method to separate physiological amounts of urodilatin and alpha-hANP in urine by high-performance liquid chromatography (HPLC) followed by ANP-RIA of the separated fractions. We studied urine samples of 10 healthy adult males with a plasma alpha-hANP level of 41 +/- 21 pg/ml (mean +/- SD) and a total urinary ANP-RIA reactivity of 40 +/- 21 pg/ml. In all urine samples we found three peaks of ANP-RIA reactivity, the first one coeluting with synthetic urodilatin, the second one with the retention time of alpha-hANP and a late eluting ANP-RIA-reactive peak, possibly containing degradation products. The ratio of urodilatin/alpha-hANP was 0.77 +/- 0.17.     

Body water handling in response to hypertonic-saline induced diuresis in fasting northern elephant seal pups (Mirounga angustirostris)

During natural fasting conditions in postweaned northern elephant seal (NES) (Mirounga angustirostris) pups, urinary water loss is minimized and percent total body water (TBW) is maintained constant. However, following infusion of hypertonic saline, glomerular filtration rate (GFR) and urine output increased in fasting pups. Therefore, we quantified the magnitude of the hypernatremia-induced diuresis relative to the animal's total body water (TBW) pool and the percentage of filtered water reabsorbed. Following a 24 h control period, naturally fasting NES pups (n=7) were infused (4 ml min(-1)) with hypertonic saline (16.7%) at a dose of 3 mmol NaCl kg(-1) body mass. Total body water was estimated prior to infusion by tritium dilution, GFR was estimated by standard creatinine clearance, and urine output (V) was measured for 24 h during the control and post infusion periods. Percentage of filtered water reabsorbed was calculated as (1-(V/GFR))x100. Twenty-four hours following the infusion, GFR (control: 69+/-12 ml min(-1) and post-infusion: 118+/-19 ml min(-1); mean+/-S.E.) increased 77+/-28% above control and the percentage of filtered water reabsorbed was decreased 0.4+/-0.1%. The increase in urine output (control: 218+/-47 ml d(-1) and post-infusion: 883+/-92 ml d(-1)) accounted for 1.7+/-0.2% of the pups' TBW. The hypernatremia-induced diuresis was accompanied by the loss of body water indicating the lack of water retention. Although the 77% increase in GFR was only associated with a 0.4% decrease in the percentage of filtered water reabsorbed, this decrease was significant enough to result in a 4-fold increase in urine output. Despite the observed diuresis, fasting NES pups appear to possess an efficient water recycling mechanism requiring only a small percentage of body water to excrete an excess salt load. This water recycling mechanism may allow pups to avoid negative perturbations in body water as they initiate feeding in a marine environment following the fast.     

Comparison of organ uptake and disappearance half-time of human epidermal growth factor and insulin

Epidermal growth factor (EGF), which was originally identified in salivary glands and saliva, has been also found in the kidney and urine, suggesting that the kidney may be an alternate source of this peptide. Liver was considered as the major site of the degradation of EGF but the involvement of other organs has been little studied. Therefore, we carried out comparative studies on the organ uptake and the disappearance half-time of EGF and insulin (having similar molecular size) in the same model of anesthetized dog with arterial (from aorta) and venous (from mesenteric, portal, hepatic, renal, femoral and jugular veins) blood sampling from various organs. Basal plasma level of EGF (1.32 +/- 0.33 pmol/l) and insulin (62.1 +/- 13.8 pmol/l) in the aorta was not significantly different from that recorded at various sampling sites. During i.v. infusion of EGF at 41.6 and 166.6 pmol/kg/h, the respective arterial EGF concentrations averaged 103 +/- 21 and 240 +/- 49 pmol/kg/h and the percent reduction in plasma EGF after passage through the head, leg, intestines and liver was about 30-50% and that after passage through the kidney was about 95%. During insulin (6.9 pmol/kg/h) infusion, the arterial hormone level averaged 227 +/- 21 pmol/l and this level was significantly reduced (by 23-42%) after passage through the head, leg, intestine, liver and kidney but no significant difference was found between various venous sampling sites. EGF and insulin appearing in the urine during EGF or insulin infusion accounted for about 40 and 7% of the difference between the entering and leaving renal masses of the peptide. Mean disappearance half time on stopping of EGF and insulin infusion was, respectively, 2.32 +/- 0.58 and 6.88 +/- 1.25 min. We conclude that unlike insulin, which is removed to similar extent by various organs including the kidney and the liver, EGF is taken up mainly by kidney and EGF present in urine originates mainly from renal clearance of peptide.     

Pyruvate shuttling during rest and exercise before and after endurance training in men.

We describe the isotopic exchange of lactate and pyruvate after arm vein infusion of [3-(13)C]lactate in men during rest and exercise. We tested the hypothesis that working muscle (limb net lactate and pyruvate exchange) is the source of the elevated systemic lactate-to-pyruvate concentration ratio (L/P) during exercise. We also hypothesized that the isotopic equilibration between lactate and pyruvate would decrease in arterial blood as glycolytic flux, as determined by relative exercise intensity, increased. Nine men were studied at rest and during exercise before and after 9 wk of endurance training. Although during exercise arterial pyruvate concentration decreased to below rest values (P < 0.05), pyruvate net release from working muscle was as large as lactate net release under all exercise conditions. Exogenous (arterial) lactate was the predominant origin of pyruvate released from working muscle. With no significant effect of exercise intensity or training, arterial isotopic equilibration [(IE(pyruvate)/IE(lactate)).100%, where IE is isotopic enrichment] decreased significantly (P < 0.05) from 60 +/- 3.1% at rest to an average value of 12 +/- 2.7% during exercise, and there were no changes in femoral venous isotopic equilibration. These data show that 1). the isotopic equilibration between lactate and pyruvate in arterial blood decreases significantly during exercise; 2). working muscle is not solely responsible for the decreased arterial isotopic equilibration or elevated arterial L/P occurring during exercise; 3). working muscle releases similar amounts of lactate and pyruvate, the predominant source of the latter being arterial lactate; 4). pyruvate clearance from blood occurs extensively outside of working muscle; and 5). working muscle also releases alanine, but alanine release is an order of magnitude smaller than lactate or pyruvate release. These results portray the complexity of metabolic integration among diverse tissue beds in vivo.     

Validation of the doubly labeled water method in growing precocial birds: the importance of assumptions concerning evaporative water loss

The doubly labeled water (DLW) method was validated against respiration gas analysis in growing precocial chicks of the black-tailed godwit (Limosa limosa) and the northern lapwing (Vanellus vanellus). To calculate the rate of CO2 production from DLW measurements, Lifson and McClintock's equations (6) and (35) were employed, as well as Speakman's equation (7.17) (all single-pool models). The average errors obtained with the first two equations (+7.2% and -11.6%, respectively) differed significantly from zero but not the error obtained with Speakman's equation (average: -2.9%). The latter error could be reduced by taking a fractional evaporative water loss of 0.13, instead of the value of 0. 25 recommended by Speakman. Application of different two-pool models resulted in relative errors of the DLW method of -15.9% or more. After employing the single-pool model with a fractional evaporative water loss value of 0.13, it was found that there was no relationship between the relative growth rate of the chick and the relative error of the DLW method. Recalculation of previously published results on Arctic tern (Sterna paradisaea) chicks revealed that the fit of the validation experiment could be considerably improved by employing a single-pool model and assuming a fractional evaporative water loss of 0.20 instead of the value of 0.50 taken originally. After employing the value of 0.20, it was found that there was no relationship between the relative growth rate of the chick and the relative error of the DLW method. This suggests that isotope incorporation into new body substances does not cause a detectable error. Thus, the DLW method seems to be applicable in young birds growing as fast as 20% d-1, after making adjustments for the fractional evaporative water loss. We recommend Speakman's equation (7.17) for general use in growing birds when evaporation is unknown.     

A comparison of chloride, bromide, and sucrose dilution volumes in neonatal pigs

Use of 36Cl, 82Br, and [3H]sucrose to estimate extracellular water volume was evaluated in 14 piglets (7-14 days old). 36Cl and 82Br were distributed in approximately the same volume, but a period of 5-6 hr after injection was required to reach equilibrium in the neonatal pig. Dilution volumes calculated before equilibration (2-5 hr) for 36Cl (326 +/- 11 ml/kg) and 82Br (328 +/- 13 ml/kg) were different from equilibration (6-8 hr) phase volumes (356 +/- 13 ml/kg and 355 +/- 13 ml/kg, respectively; P less than 0.001). A 3-hr sample estimated the same volume distribution calculated by extrapolation of the 6- to 8-hr period because of the relationship between the two slopes of the plasma clearance curves. After the 82Br and 36Cl had achieved equilibration, each was distributed in a volume equivalent to total body chloride space (362 +/- 29 ml/kg) measured by neutron activation; no statistical differences were found (P = 0.6). The early equilibration phase measured a 10% smaller, faster exchangeable fraction of total body Cl. Sucrose dilution volume (332 +/- 19 ml/kg) required multiple plasma samples for extrapolation and measured a dilution volume 7% smaller (P less than 0.05) than total body chloride space.     

Cardiopulmonary effects of recombinant interleukin-2 infusion in sheep

The systemic administration of recombinant interleukin-2 (rIL-2) with or without lymphokine-activated killer (LAK) cells, a new treatment for patients with advanced cancer, is associated with a presumed "third-space" syndrome. To further define the extent and time course of this toxicity, we established a chronic sheep model and monitored changes in systemic and central vascular pressures, cardiac function, and gas exchange during a 72-h continuous intravenous infusion of rIL-2 at a total dose of 5 (group 3) or 9 x 10(5) U/kg (group 4). At 72 h, caudal mediastinal lymph flow, histology, and extravascular lung water-to-dry lung weight ratio (EVLW/DLW) were obtained. During the rIL-2 infusion there was a dose-dependent significant decrease in systemic blood pressure and arterial Po2 and an increase in core temperature. In group 4, pulmonary arterial pressure increased from a base line of 13 +/- 5 to 21 +/- 6 mmHg (P less than 0.05). Lung lymph flow was significantly increased in groups 3 and 4 compared with animals receiving 0.9% NaCl or excipient infusions (groups 1 and 2). EVLW/DLW values were elevated in groups 3 and 4 (P less than 0.01). In animals receiving rIL-2, histological evaluation revealed a dose-dependent infiltration of lung tissue by lymphoblastoid cells that stained esterase negative. We conclude that rIL-2 infusion in doses comparable to those given to humans results in alterations in systemic and central hemodynamics, gas exchange, high-protein lung lymph flow, and infiltration of lymphoblastoid cells into the lung parenchyma.