Graham Jennings

Measurement of rat brown-adipose-tissue mitochondrial uncoupling protein by radioimmunoassay: increased concentration after cold acclimation.

A specific antiserum has been raised against the 32 000-mol.wt. uncoupling protein from the mitochondria of rat brown adipose tissue and a sensitive radioimmunoassay for the protein has been developed. The uncoupling protein is present in large amounts in brown adipose tissue; its concentration is increased substantially by cold acclimation. The protein has not been detected in the liver, heart, or parametrial white adipose tissue of rats.

Total energy expenditure in patients with small-cell lung cancer: results of a validated study using the bicarbonate-urea method.

The bicarbonate-urea method for measuring CO2 production was applied to eight free-living patients (mean age, 68 +/- 10 years; mean weight, 69 +/- 10 kg; mean height, 1.65 +/- 0.10 m) with unresectable small-cell lung cancer for a period of 1 day (n = 5) or 2 days (n = 3). The basal metabolic rate (BMR) was measured in all subjects. The technique was first validated against whole-body indirect calorimetry over an additional 24-hour period in five of these subjects. The bicarbonate-urea method predicted net CO2 production to be 102.1% +/- 3.4% of that measured by whole-body indirect calorimetry, and energy expenditure, 101.5% +/- 3.8% of the measured calorimeter value (8.1 +/- 1.6 MJ/d). The 24-hour recovery of label in CO2 excreted by the body was 95.6% +/- 0.5%. In free-living conditions, the bicarbonate-urea method predicted energy expenditure to be 9.0 +/- 2.6 MJ/d. BMR was elevated by a mean of 6% (P < .05) compared with the Schofield standards. The physical activity level ([PAL] the ratio of total energy expenditure [TEE] to BMR) was variable (1.15 to 1.87), but the mean value was only 1.36 +/- 0.22, considerably less than that of moderately active healthy subjects with estimated PAL values of 1.55 (P < .05) to 1.65 (P < .01) and the mean results obtained by doubly labeled water (previous studies) in healthy age- and sex-matched subjects. This is the first time a tracer method for measuring CO2 production and energy expenditure has been validated against whole-body 24-hour indirect calorimetry in patients with lung cancer or a systemic inflammatory reaction. The agreement between the two methods is similar to that observed in normal subjects. This is also the first time a tracer method has been used to measure energy expenditure in free-living patients with lung cancer. The results suggest that TEE and the energy requirements necessary to maintain energy balance were not increased despite basal hypermetabolism, because of the associated decrease in physical activity.

Effect of acclimation temperature on the concentration of the mitochondrial ‘uncoupling’ protein measured by radioimmunoassay in mouse brown adipose tissue

The effect of acclimation temperature on the concentration of the mitochondrial ‘uncoupling’ protein (M r 32 000) from brown adipose tissue of mice has been investigated. The uncoupling protein was measured by a specific radioimmunoassay. Between 33°C (thermoneutrality) and −2°C there was a progressive increase with decreasing environmental temperature in the amount of uncoupling protein. For mice at −2°C the mitochondrial concentration of the protein was 9‐times higher than at 33°C, while the total amount of the protein in interscapulaar brown adipose tissue was estimated to be nearly 80‐times greater at −2°C compared to 33°C.

Bioelectrical impedance analysis in human immunodeficiency virus-infected patients: Comparison of single frequency with multifrequency, spectroscopy, and other novel approaches

Bioelectrical impedance (BIA), a prediction method for estimating body water compartments and body cell mass (BCM), is being increasingly used in studies of human immunodeficiency virus (HIV)-related wasting, but there are few validation studies of the method in this group. The aim of this study is to examine the relationship between impedance measurements and body water compartments in patients with advanced HIV disease, and to investigate whether the newer approaches of multifrequency BIA, BIA spectroscopy, logarithmic transformation using a parallel circuit model, and direct calculation from electrical theory offer any advantage over traditional single-frequency BIA. We measured total body water (TBW) by deuterium dilution and extracellular water by bromide dilution in 33 patients with advanced HIV disease. Intracellular water and BCM were calculated from these results. Impedance was measured over a range of frequencies using a multifrequency analyzer. The relationship between impedance index at various frequencies and body water compartments was assessed by correlation and linear regression. We found that impedance index at higher frequencies had a closer relationship to TBW (r = 0.86, standard error of the estimate [SEE] = 2.96 at 1000 kHz) and at lower frequencies a closer relationship to extracellular water (ECW) (r = 0.47, SEE = 3.13 at 0 kHz) than the traditional 50 kHz measurement (r = 0.84, SE = 3.11 for TBW; r = 0.44 SEE = 3.19 for ECW), but the differences were marginal and not statistically significant. None of the other novel approaches tested were significantly better than traditional single frequency measurement. The 50 kHz equation for BCM developed in this study [BCM (kg) = (0.360331 x Ht2/Z50) + (0.151123 x Wt)-2.95] may be useful to investigators using BIA for hIV-wasting studies.

Impact of tuberculosis on the body composition of HIV-infected men in Brazil.

OBJECTIVE Tuberculosis (TB) is the commonest HIV-related opportunistic infection in many developing countries and is thought to be a frequent underlying cause of HIV-associated wasting. We have used reference water dilution methods to examine the body composition changes associated with TB and to assess the severity and pattern of wasting. METHODS The study was conducted at a charitable support house for poor and homeless HIV-infected people in Rio de Janeiro, Brazil. Male patients who were HIV-positive and receiving treatment for active TB (HIVTB+) and HIV-infected controls without TB (HIVTB-) were studied. Total body water (TBW) and extracellular water (ECW) were measured by giving oral doses of deuterium oxide and sodium bromide, respectively, and determining enrichment in plasma after 4 hours. Intracellular water (ICW), body cell mass (BCM), lean body mass (LBM) and fat mass were calculated from these parameters using standard equations. RESULTS HIVTB+ (n = 11) and HIVTB- (n = 12) groups were similar in age, height, CD4 count and HIV risk factors. HIVTB+ men had significantly lower mean ICW (13.2 versus 16.6 kg; p = .02) and BCM (18.4 versus 23.0 kg; p = .02), a relative expansion of ECW (35.0 versus 30.0 L/kg body weight; p = .04), and small and nonsignificant reductions in total body weight (58.0 versus 62.1 kg; p = .26), LBM (45.5 versus 47.7 kg; p = .33) and fat mass (12.5 versus 14.4 kg; p = .51) compared with HIVTB- controls. BCM in the HIVTB+ group was similar to reference values for severe malnutrition. The relative depletion of BCM appeared excessive in comparison with reference values for uncomplicated starvation. CONCLUSION The nutritional status of HIVTB+ patients was significantly worse than HIVTB- patients. Body weight and LBM underestimated the nutritional deficit, and measurement of BCM is therefore necessary to appreciate the extent of malnutrition in such patients. Malnutrition in HIVTB+ patients is severe and may therefore contribute to decreased survival. Hypermetabolism appears to play a role in the wasting process in patients coinfected with HIV and TB.

Measurement by radioimmunoassay of the mitochondrial uncoupling protein from brown adipose tissue of obese (ob/ob) mice and Zucker (fa/fa) rats at different ages

The concentration of the ‘uncoupling protein’ in brown adipose tissue mitochondria has been measured in lean and obese (ob/ob) mice and Zucker (fa/fa) rats at different ages using a specific radioimmunoassay. During the suckling period the concentration of the protein was similar in normal and mutant animals of both types, despite the decrease in mitochondrial GDP binding observed in the obese. The concentration of uncoupling protein was, however, decreased in adult ob/ob mice and adult Zucker rats compared with their respective lean siblings, in parallel with the decrease in GDP binding. It is concluded that there is a ‘masked’, or inactive, form of uncoupling protein in young ob/ob mice and fa/fa rats.

Brown Adipose Tissue Thermogenesis during Pregnancy in Mice

The thermogenic activity of interscapular brown adipose tissue has been assessed at different stages of pregnancy in mice. In late pregnancy there was a hypertrophy of the tissue which reversed at parturition. Neither the total protein content nor the total cytochrome oxidase activity of the tissue changed significantly throughout pregnancy or into early lactation (2-3 days, post-partum). However, mitochondrial GDP binding, an index of the activity of the proton conductance pathway, was significantly decreased at the end of pregnancy with a further decrease in early lactation. Moderate food restriction had no effect on either cytochrome oxidase activity or mitochondrial GDP binding at the end of pregnancy, as compared with pregnant animals fed ad libitum. Food restriction did, however, prevent the hypertrophy of brown adipose tissue in late pregnancy. It is concluded that brown adipose tissue thermogenesis is not significantly decreased in the pregnant mouse until shortly before parturition, even in animals subject to food restriction. It is also concluded that the normal dietary stimulation of thermogenesis in response to hyperphagia is suppressed in the pregnant animal.

Adaptive changes in the concentration of the mitochondrial ‘uncoupling’ protein in brown adipose tissue of hamsters acclimated at different temperatures

The effect of acclimation at different temperatures on the activity of interscapular brown adipose tissue has been investigated in the hamster, a hibernator. Between 31° and 4°C the cytochrome oxidase activity of the tissue increased 4- to 5-fold, mitochondrial GDP binding per mg of mitochondrial protein doubled, and the amount of uncoupling protein rose from 1.7% to 5.4% of total mitochondrial protein. It is concluded that there are clear adaptive changes induced by temperature in brown adipose tissue of the hamster, but the changes are limited in comparison with those in the mouse.

Dual-energy X-ray absorptiometry for the measurement of gross body composition in rats.

Dual-energy X-ray absorptiometry (DXA) is a novel, non-invasive technique for the measurement of gross body composition in small animals. In the present study the absolute accuracy of the Hologic QDR-1000W scanner was assessed by comparison with direct analysis in twelve rats with a range of body fat and bone mineral content (BMC) values. Fat masses measured by DXA and petroleum-ether extraction were significantly different (P < 0.0023). The DXA technique consistently overestimated fat mass by approximately one third of the measured fat content. BMC derived from the measurement of Ca in ash gave a mean of 8.26 (range 1.57-15.71)g. BMC measured by DXA was not significantly different for the group as a whole. However, there was a trend for DXA to overestimate BMC in animals with low BMC and underestimate in those with higher BMC, compared with direct analysis, such that the 95% limits of agreement for the two techniques were +2.73 to -2.58 g. These results suggest that the present small-animal software developed for use with currently available Hologic machines does not give an accurate measure of gross body composition compared with the results from classical direct analysis.

Effect of caging singly or in groups of different sizes on the thermogenic activity of interscapular brown adipose tissue in mice

The effects on the thermogenic activity of brown adipose tissue of caging mice singly or in groups of different sizes has been investigated. At 23 degrees C the total cytochrome oxidase activity and the level of mitochondrial GDP binding were higher in mice caged singly than in mice caged in groups of three or six. At 4 degrees C GDP binding and cytochrome oxidase activity were lower in mice caged in groups of two, three or six than in mice caged singly. The mitochondrial concentration of uncoupling protein was not clearly affected by the number of mice in each cage.