Colorimetric Measurement of Triglycerides Cannot Provide an Accurate Measure of Stored Fat Content in Drosophila

Drosophila melanogaster has recently emerged as a useful model system in which to study the genetic basis of regulation of fat storage. One of the most frequently used methods for evaluating the levels of stored fat (triglycerides) in flies is a coupled colorimetric assay available as a kit from several manufacturers. This is an aqueous-based enzymatic assay that is normally used for measurement of mammalian serum triglycerides, which are present in soluble lipoprotein complexes. In this short communication, we show that coupled colorimetric assay kits cannot accurately measure stored triglycerides in Drosophila. First, they fail to give accurate readings when tested on insoluble triglyceride mixtures with compositions like that of stored fat, or on fat extracted from flies with organic solvents. This is probably due to an inability of the lipase used in the kits to efficiently cleave off the glycerol head group from fat molecules in insoluble samples. Second, the measured final products of the kits are quinoneimines, which absorb visible light in the same wavelength range as Drosophila eye pigments. Thus, when extracts from crushed flies are assayed, much of the measured signal is actually due to eye pigments. Finally, the lipoprotein lipases used in colorimetric assays also cleave non-fat glycerides. The glycerol backbones liberated from all classes of glycerides are measured through the remaining reactions in the assay. As a consequence, when these assay kits are used to evaluate tissue extracts, the observed signal actually represents the amount of free glycerols together with all types of glycerides. For these reasons, findings obtained through use of coupled colorimetric assays on Drosophila samples must be interpreted with caution. We also show here that using thin-layer chromatography to measure stored triglycerides in flies eliminates all of these problems

Long term hemodialysis aggravates lipolytic activity reduction and very low density, low density lipoproteins composition in chronic renal failure patients

<p>Abstract</p> <p>Background</p> <p>Dyslipidemia, particularly hypertriglyceridemia is common in uremia, and represents an independent risk factor for atherosclerosis.</p> <p>Methods</p> <p>To investigate the effects of hemodialysis (HD) duration on very low density lipoprotein (VLDL) and low density lipoprotein (LDL) compositions and lipopolytic activities, 20 patients on 5 to 7 years hemodialysis were followed-up during 9 years. Blood samples were drawn at T0 (beginning of the study), T1 (3 years after initiating study), T2 (6 years after initiating study) and T3 (9 years after initiating study). T0 was taken as reference.</p> <p>Results</p> <p>Triacylglycerols (TG) values were correlated with HD duration (r = 0.70, P < 0.05). An increase of total cholesterol was noted at T2 and T3. Lowered activity was observed for lipoprotein lipase (LPL) (-44%) at T3 and hepatic lipase (HL) (-29%) at T1, (-64%) at T2 and (-73%) at T3. Inverse relationships were found between HD duration and LPL activity (r = -0.63, P < 0.05), and HL activity (r = -0.71, P < 0.01). At T1, T2 and T3, high VLDL-amounts and VLDL-TG and decreased VLDL-phospholipids values were noted. Increased LDL-cholesteryl esters values were noted at T1 and T2 and in LDL-unesterified cholesterol at T2 and T3.</p> <p>Conclusion</p> <p>Despite hemodialysis duration, VLDL-LDL metabolism alterations are aggravated submitting patients to a greater risk of atherosclerosis.</p

Effects of Male Hypogonadism on Regional Adipose Tissue Fatty Acid Storage and Lipogenic Proteins

Testosterone has long been known to affect body fat distribution, although the underlying mechanisms remain elusive. We investigated the effects of chronic hypogonadism in men on adipose tissue fatty acid (FA) storage and FA storage factors. Twelve men with chronic hypogonadism and 13 control men matched for age and body composition: 1) underwent measures of body composition with dual energy x-ray absorptiometry and an abdominal CT scan; 2) consumed an experimental meal containing [3H]triolein to determine the fate of meal FA (biopsy-measured adipose storage vs. oxidation); 3) received infusions of [U-13C]palmitate and [1-14C]palmitate to measure rates of direct free (F)FA storage (adipose biopsies). Adipose tissue lipoprotein lipase, acyl-CoA synthetase (ACS), and diacylglycerol acetyl-transferase (DGAT) activities, as well as, CD36 content were measured to understand the mechanism by which alterations in fat storage occur in response to testosterone deficiency. Results of the study showed that hypogonadal men stored a greater proportion of both dietary FA and FFA in lower body subcutaneous fat than did eugonadal men (both p<0.05). Femoral adipose tissue ACS activity was significantly greater in hypogonadal than eugonadal men, whereas CD36 and DGAT were not different between the two groups. The relationships between these proteins and FA storage varied somewhat between the two groups. We conclude that chronic effects of testosterone deficiency has effects on leg adipose tissue ACS activity which may relate to greater lower body FA storage. These results provide further insight into the role of androgens in body fat distribution and adipose tissue metabolism in humans

Association of Mild Anemia with Cognitive, Functional, Mood and Quality of Life Outcomes in the Elderly: The “Health and Anemia” Study

BACKGROUND: In the elderly persons, hemoglobin concentrations slightly below the lower limit of normal are common, but scant evidence is available on their relationship with significant health indicators. The objective of the present study was to cross-sectionally investigate the association of mild grade anemia with cognitive, functional, mood, and quality of life (QoL) variables in community-dwelling elderly persons. METHODS: Among the 4,068 eligible individuals aged 65-84 years, all persons with mild anemia (n = 170) and a randomly selected sample of non-anemic controls (n = 547) were included in the study. Anemia was defined according to World Health Organization (WHO) criteria and mild grade anemia was defined as a hemoglobin concentration between 10.0 and 11.9 g/dL in women and between 10.0 and 12.9 g/dL in men. Cognition and functional status were assessed using measures of selective attention, episodic memory, cognitive flexibility and instrumental and basic activities of daily living. Mood and QoL were evaluated by means of the Geriatric Depression Scale-10, the Short-Form health survey (SF-12), and the Functional Assessment of Cancer Therapy-Anemia. RESULTS: In univariate analyses, mild anemic elderly persons had significantly worse results on almost all cognitive, functional, mood, and QoL measures. In multivariable logistic regressions, after adjustment for a large number of demographic and clinical confounders, mild anemia remained significantly associated with measures of selective attention and disease-specific QoL (all fully adjusted p<.046). When the lower limit of normal hemoglobin concentration according to WHO criteria was raised to define anemia (+0.2 g/dL), differences between mild anemic and non anemic elderly persons tended to increase on almost every variable. CONCLUSIONS: Cross-sectionally, mild grade anemia was independently associated with worse selective attention performance and disease-specific QoL ratings

Experimental hyperthyroidism in man: effects on plasma lipoproteins, lipoprotein lipase and hepatic lipase

We have studied the effects of triiodothyronine administration (20-40 micrograms three times daily over one week) in six healthy young men, on the activities of lipoprotein lipase and hepatic lipase and on plasma lipoprotein concentrations. Hepatic lipase activity in post-heparin plasma rose by 46 +/- 25% (p less than 0.025), whereas the activity of lipoprotein lipase did not change significantly. Plasma cholesterol concentrations decreased by about 20% (p less than 0.025), whereas there was no change in plasma triglyceride levels. The fall in plasma cholesterol could be accounted for by a reduction of HDL cholesterol (-11%, p less than 0.025) as well as LDL cholesterol (-27%, p less than 0.025). The data emphasize the role of hepatic lipase in the lipoprotein alterations associated with thyroid dysfunction

Enzymes catalyzing the hydrolysis of long-chain monoacyglycerols in rat adipose tissue

Acetone-ether preparations of epididymal fat pads from fasted or fed rats contained two enzymes catalyzing the hydrolysis of long-chain monoacylglycerols. The enzymes were identified as monoacylglycerol lipase (Tornqvist, H. and Belfrage, P., (1976) J. Biol Chem. 251, 813--819) and lipoprotein lipase by their apparent pI values after electrofocusing in non-ionic detergent, selective inhibition properties, substrate specificity and positional specificity. It was estimated that monoacylglycerol lipase accounted for about 90% of the total monoacylglycerol-hydrolyzing activity in acetone-ether preparations from fasted and 70% from fed rats. Its enzyme activity did not change with the nutritional state in contrast to that of lipoprotein lipase. The latter enzyme hydrolyzed 2-monoacylglycerols at a much lower rate than the 1(3)-isomers. Monoacylglycerol lipase was located almost entirely in the adipocytes, thus most of the enzyme activity towards monoacylglycerols in the adipose tissue was found in this site. Fractionated sucrose homogenates of rat epididymal fat pads also contained a third enzyme with monoacylglycerol-hydrolyzing activity, identified as hormone-sensitive lipase by its pI, selective inhibition properties and substrate specificity. It was estimated that hormone-sensitive lipase accounted for less than 20% of the total activity against monoacylglycerols in these tissue preparations from fasted rats. Over-all quantitative estimations emphasized the dominant role of monoacylglycerol lipase over the other two enzymes in the hydrolysis of monoacylglycerols

High-density lipoprotein concentrations increase after stopping smoking.

Concentrations of plasma lipoproteins in 10 men who were habitual smokers were monitored for six weeks after they stopped smoking and related to changes in diet and body weight. The energy intake increased by 10% (p less than 0.05) owing to a higher consumption of carbohydrates and fat, and body weight increased by 2% (p less than 0.01). Plasma triglyceride, cholesterol, and low-density lipoprotein cholesterol concentrations did not change significantly. The most prominent finding was a rapid and pronounced increased in high-density lipoprotein concentrations. From comparatively low values (mean 0.82 mmol/1) they rose by 29% (p less than 0.01) within two weeks and remained at this value throughout the observation period. In three subjects who resumed smoking after the end of the study they again fell to initial values six weeks later. The initial increase in concentration could be accounted for mainly by an increase in the esterified fraction and only to a lesser extent in the free cholesterol fraction. The changes in concentrations were accompanied by similar but less pronounced rises in high-density lipoprotein phospholipid and in apolipoprotein AI concentrations (p less than 0.01), whereas high-density lipoprotein phospholipid and in apolipoprotein AI concentration (p less than 0.01), whereas high-density lipoprotein triglyceride concentrations did not change significantly. These findings confirm and extend those of earlier cross-sectional studies which showed low concentrations of high-density lipoproteins in cigarette smokers, A significant correlation between the rise in high-density lipoprotein cholesterol concentrations and the increase in fat consumption after stopping smoking indicate that the changes in high-density lipoprotein concentrations may be partly due to nutritional factors