Energy expenditure, insulin, and VLDL-triglyceride production in humans
2006; Elsevier BV; Volume: 47; Issue: 10 Linguagem: Inglês
10.1194/jlr.m600175-jlr200
ISSN1539-7262
AutoresLars Christian Gormsen, Michael D. Jensen, Ole Schmitz, Niels Møller, Jens Sandahl Christiansen, Søren Nielsen,
Tópico(s)Muscle metabolism and nutrition
ResumoHypertriglyceridemia is considered a cardiovascular risk factor in diabetic and nondiabetic subjects. In this study, we aimed to determine potential regulators of very low density lipoprotein-triglyceride (TG) production. VLDL-TG kinetics were measured in 13 men and 12 women {body mass index [mean (range)]: 24.8 (20.2–35.6) kg/m2}. VLDL-TG production was assessed from the plasma decay of a bolus injection of ex vivo labeled VLDL particles ([1-14C]triolein-VLDL-TG). Similar VLDL-TG production (μmol/min) was found in men and women. VLDL-TG production was not significantly correlated with palmitate flux ([9,10-3H]palmitate) (r = 0.09, P = 0.67) or palmitate concentration (r = −0.29, P = 0.2) but was correlated significantly with fasting insulin concentration (r = 0.46, P < 0.05) and resting energy expenditure (REE) (r = 0.45, P < 0.05). The latter correlation improved when adjusted for sex. The best multivariate model with VLDL-TG production as the dependent variable and REE, body composition, hormones, and substrate levels as independent variables included fasting insulin (P = 0.02) and REE (P = 0.02) (r2 = 0.32, P < 0.001). We conclude that VLDL kinetics are similar in men and women and that REE and plasma insulin are significant independent predictors of VLDL-TG production. FFA availability and body fat distribution are unrelated to VLDL production. We suggest that REE plays a greater role in VLDL-TG production than previously anticipated. REE and insulin should be taken into account when VLDL-TG production comparisons between groups are made. Hypertriglyceridemia is considered a cardiovascular risk factor in diabetic and nondiabetic subjects. In this study, we aimed to determine potential regulators of very low density lipoprotein-triglyceride (TG) production. VLDL-TG kinetics were measured in 13 men and 12 women {body mass index [mean (range)]: 24.8 (20.2–35.6) kg/m2}. VLDL-TG production was assessed from the plasma decay of a bolus injection of ex vivo labeled VLDL particles ([1-14C]triolein-VLDL-TG). Similar VLDL-TG production (μmol/min) was found in men and women. VLDL-TG production was not significantly correlated with palmitate flux ([9,10-3H]palmitate) (r = 0.09, P = 0.67) or palmitate concentration (r = −0.29, P = 0.2) but was correlated significantly with fasting insulin concentration (r = 0.46, P < 0.05) and resting energy expenditure (REE) (r = 0.45, P < 0.05). The latter correlation improved when adjusted for sex. The best multivariate model with VLDL-TG production as the dependent variable and REE, body composition, hormones, and substrate levels as independent variables included fasting insulin (P = 0.02) and REE (P = 0.02) (r2 = 0.32, P < 0.001). We conclude that VLDL kinetics are similar in men and women and that REE and plasma insulin are significant independent predictors of VLDL-TG production. FFA availability and body fat distribution are unrelated to VLDL production. We suggest that REE plays a greater role in VLDL-TG production than previously anticipated. REE and insulin should be taken into account when VLDL-TG production comparisons between groups are made. An increased concentration of plasma triglyceride (TG) is recognized as an independent cardiovascular risk factor (1Hokanson J.E. Austin M.A. Plasma triglyceride level is a risk factor for cardiovascular disease independent of high-density lipoprotein cholesterol level: a meta-analysis of population-based prospective studies..J. Cardiovasc. Risk. 1996; 3: 213-219Crossref PubMed Google Scholar) and is a frequent finding in insulin-resistant conditions such as obesity and type 2 diabetes. Although plasma TG concentration is a well-described parameter in health and disease, our knowledge about plasma TG kinetics and factors affecting TG metabolism is still limited. This is probably attributable to the lack of easily available, robust methods to determine TG production, as indicated by the use of a wide variety of approaches (2Magkos F. Sidossis L.S. Measuring very low density lipoprotein-triglyceride kinetics in man in vivo: how different the various methods really are..Curr. Opin. Clin. Nutr. Metab. Care. 2004; 7: 547-555Crossref PubMed Scopus (25) Google Scholar). In the postabsorptive state, circulating TGs are found predominantly in VLDL particles. The absolute and relative turnover of this TG pool determines the plasma TG concentrations, reflecting the balance between the hepatic secretion and peripheral clearance of VLDL-TG. The latter event is largely determined by endothelial lipoprotein lipase activity and receptor-mediated VLDL uptake. Among factors proposed to regulate VLDL-TG metabolism is FFA availability (3Lam T.K. Carpentier A. Lewis G.F. van de Werve G. Fantus I.G. Giacca A. Mechanisms of the free fatty acid-induced increase in hepatic glucose production..Am. J. Physiol. Endocrinol. Metab. 2003; 284: E863-E873Crossref PubMed Scopus (204) Google Scholar, 4Lewis G.F. Fatty acid regulation of very low density lipoprotein production..Curr. Opin. Lipidol. 1997; 8: 146-153Crossref PubMed Scopus (246) Google Scholar), which has long been considered a key regulator of VLDL production. However, fasting FFA availability is in turn influenced by other factors, such as resting energy expenditure (REE) (5Nielsen S. Guo Z. Albu J.B. Klein S. O'Brien P.C. Jensen M.D. Energy expenditure, sex, and endogenous fuel availability in humans..J. Clin. Invest. 2003; 111: 981-988Crossref PubMed Scopus (112) Google Scholar), body composition (5Nielsen S. Guo Z. Albu J.B. Klein S. O'Brien P.C. Jensen M.D. Energy expenditure, sex, and endogenous fuel availability in humans..J. Clin. Invest. 2003; 111: 981-988Crossref PubMed Scopus (112) Google Scholar, 6Jensen M.D. Haymond M.W. Rizza R.A. Cryer P.E. Miles J.M. Influence of body fat distribution on free fatty acid metabolism in obesity..J. Clin. Invest. 1989; 83: 1168-1173Crossref PubMed Scopus (547) Google Scholar, 7Jensen M.D. Fate of fatty acids at rest and during exercise: regulatory mechanisms..Acta Physiol. Scand. 2003; 178: 385-390Crossref PubMed Scopus (67) Google Scholar), catecholamine availability (5Nielsen S. Guo Z. Albu J.B. Klein S. O'Brien P.C. Jensen M.D. Energy expenditure, sex, and endogenous fuel availability in humans..J. Clin. Invest. 2003; 111: 981-988Crossref PubMed Scopus (112) Google Scholar, 8Aarsland A. Chinkes D. Wolfe R.R. Contributions of de novo synthesis of fatty acids to total VLDL-triglyceride secretion during prolonged hyperglycemia/hyperinsulinemia in normal man..J. Clin. Invest. 1996; 98: 2008-2017Crossref PubMed Scopus (192) Google Scholar, 9Carpentier A. Patterson B.W. Leung N. Lewis G.F. Sensitivity to acute insulin-mediated suppression of plasma free fatty acids is not a determinant of fasting VLDL triglyceride secretion in healthy humans..Diabetes. 2002; 51: 1867-1875Crossref PubMed Scopus (38) Google Scholar, 10Lewis G.F. Uffelman K.D. Szeto L.W. Steiner G. Effects of acute hyperinsulinemia on VLDL triglyceride and VLDL apoB production in normal weight and obese individuals..Diabetes. 1993; 42: 833-842Crossref PubMed Scopus (283) Google Scholar, 11Lewis G.F. Uffelman K.D. Szeto L.W. Weller B. Steiner G. Interaction between free fatty acids and insulin in the acute control of very low density lipoprotein production in humans..J. Clin. Invest. 1995; 95: 158-166Crossref PubMed Google Scholar), and sex (5Nielsen S. Guo Z. Albu J.B. Klein S. O'Brien P.C. Jensen M.D. Energy expenditure, sex, and endogenous fuel availability in humans..J. Clin. Invest. 2003; 111: 981-988Crossref PubMed Scopus (112) Google Scholar). Fasting plasma insulin has also been reported to have an effect on VLDL metabolism (9Carpentier A. Patterson B.W. Leung N. Lewis G.F. Sensitivity to acute insulin-mediated suppression of plasma free fatty acids is not a determinant of fasting VLDL triglyceride secretion in healthy humans..Diabetes. 2002; 51: 1867-1875Crossref PubMed Scopus (38) Google Scholar), and it has been demonstrated that hyperinsulinemia caused by massive carbohydrate ingestion is associated with increased VLDL-TG production independent of the concomitant increase in FFA levels (8Aarsland A. Chinkes D. Wolfe R.R. Contributions of de novo synthesis of fatty acids to total VLDL-triglyceride secretion during prolonged hyperglycemia/hyperinsulinemia in normal man..J. Clin. Invest. 1996; 98: 2008-2017Crossref PubMed Scopus (192) Google Scholar). On the other hand, an acute exposure to high levels of insulin decreases VLDL-TG output by the liver (10Lewis G.F. Uffelman K.D. Szeto L.W. Steiner G. Effects of acute hyperinsulinemia on VLDL triglyceride and VLDL apoB production in normal weight and obese individuals..Diabetes. 1993; 42: 833-842Crossref PubMed Scopus (283) Google Scholar). Recently, we reported significantly greater FFA (palmitate) turnover in women compared with men and a strong relationship between FFA turnover and REE (5Nielsen S. Guo Z. Albu J.B. Klein S. O'Brien P.C. Jensen M.D. Energy expenditure, sex, and endogenous fuel availability in humans..J. Clin. Invest. 2003; 111: 981-988Crossref PubMed Scopus (112) Google Scholar). The greater FFA turnover in women was noted not only at comparable levels of REE and relative oxidation rates of glucose and lipids but also at similar plasma FFA and TG concentrations. These findings can only be explained by an increased nonoxidative clearance of FFAs in women compared with men. One nonoxidative mechanism by which FFAs can be readily cleared from the circulation is by the hepatic production and subsequent release of VLDL-TG particles into the circulation destined for adipose tissue storage. Whether women channel more FFAs toward VLDL-TG than men and whether a relationship with REE also exists for VLDL-TG is unknown. This study was designed to further investigate differences in VLDL turnover in men and women. We specifically wanted to establish whether a relationship exists between REE and VLDL-TG production and, if so, to what extent this relationship was modified by sex, FFA availability, body composition, and hormone levels. If a significant relationship between VLDL-TG production and REE can be demonstrated, perhaps independent of FFA availability, it would change our present perception of VLDL-TG from serving merely as a passive transporter of TG for adipose tissue storage toward an additional direct role of VLDL particles in substrate delivery to energy-consuming tissues. We used a recently validated method involving ex vivo labeling of native VLDL-TG with subsequent calculation of VLDL-TG production based on the monoexponential decay of VLDL-TG specific activity (SA) (12Gormsen L.C. Jensen M.D. Nielsen S. Measuring VLDL-triglyceride turnover in humans using ex vivo-prepared VLDL tracer..J. Lipid Res. 2006; 47: 99-106Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). Twenty-six healthy subjects (13 women and 13 men) were included in the study. All participants were nonsmokers, not taking any medication, and had been weight-stable for the previous 3 months. All were normotriglyceridemic and had normal blood pressure, normal hematological indices, and normal liver and renal function. All women were premenopausal. Subjects were recruited such that a wide range of body fat distribution was included among both sexes. Informed consent was obtained from all participants, and the Ethics Committee gave its approval. One woman had to be excluded during the study because of technical difficulties. Dual-energy X-ray absorptiometry (QDR-2000; Hologic, Inc., Waltham, MA) and computed tomography of the L2–3 interspace (13Jensen M.D. Kanaley J.A. Reed J.E. Sheedy P.F. Measurement of abdominal and visceral fat with computed tomography and dual-energy x-ray absorptiometry..Am. J. Clin. Nutr. 1995; 61: 274-278Crossref PubMed Scopus (285) Google Scholar) was performed in the days preceding the examination to determine the body composition [total fat mass (FM) and fat-free mass (FFM)] of the participants. REE and substrate oxidation rates (14Ferrannini E. The theoretical bases of indirect calorimetry: a review..Metabolism. 1988; 37: 287-301Abstract Full Text PDF PubMed Scopus (1298) Google Scholar) were measured by indirect calorimetry (Deltatrac monitor; Datex Instrumentarium, Helsinki, Finland). The initial 5 min was used for acclimatization to the hood, and REE was then measured from 65 to 90 min. Systemic palmitate flux was measured at 30–60 min using the isotope dilution technique with a constant infusion of [9,10-3H]palmitate (0.3 μCi/min) (Lægemiddelstyrelsen, Copenhagen, Denmark) (15Guo Z. Nielsen S. Burguera B. Jensen M.D. Free fatty acid turnover measured using ultralow doses of [U-13C]palmitate..J. Lipid Res. 1997; 38: 1888-1895Abstract Full Text PDF PubMed Google Scholar) from 0 to 60 min. Blood samples for the measurement of palmitate concentration and SA were drawn at baseline and at 10 min intervals over the last 30 min of the infusion. The steady-state SA was verified (30, 40, 50, and 60 min) for each individual. Plasma palmitate concentration and SA were determined by HPLC using [2H31]palmitate as the internal standard. Systemic palmitate flux (μmol/min) was calculated using the [9,10-3H]palmitate infusion rate (dpm/min) divided by the steady-state palmitate SA (dpm/μmol). VLDL was isolated from plasma by ultracentrifugation. Approximately 3 ml of each plasma sample was transferred into Optiseal tubes (Beckman Instruments, Inc., Palo Alto, CA), covered with a saline solution (d = 1.006 g/ml), and centrifuged (50.3 rotor; Beckman Instruments) for 18 h at 40,000 g and 10°C. The top layer, containing VLDL, was aspirated, and the exact volume was recorded. A small proportion was analyzed for TG content, and the plasma concentration of VLDL-TG was calculated. This procedure results in VLDL-TG concentrations that are highly correlated with total plasma TG concentration (r = 0.86, P < 0.000001). However, because the aspiration of VLDL particles is not complete, VLDL-TG concentrations are slightly underestimated. The remaining VLDL-TG was transferred to a scintillation glass vial, 10 ml of scintillation liquid was added (Optiphase HiSafe 2; Wallac), and the sample was measured for 14C activity using dual-channel counting. One week before the examination, a 40 ml blood sample was obtained aseptically from each participant. The VLDL fraction was separated from plasma as described above and transferred to a sterile glass test tube containing 20 μCi of [1-14C]triolein dissolved in 200 μl of ethanol. The solution was then gently sonicated in a water bath at 37°C for 30 min and subsequently filtered through a sterile 20 μm filter (Filtropur). A 300 μl sample of the solution was tested for bacterial growth to rule out contamination during the process. To ensure that [1-14C]triolein was incorporated into the lipoprotein particles, each sample was assayed by size-exclusion HPLC as described previously (12Gormsen L.C. Jensen M.D. Nielsen S. Measuring VLDL-triglyceride turnover in humans using ex vivo-prepared VLDL tracer..J. Lipid Res. 2006; 47: 99-106Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). An HPLC radiochromatogram demonstrated that the radioactivity coeluted with VLDL-sized particles, indicating successful incorporation of radioactive tracer into the VLDL lipoproteins. Because VLDL particles are essentially confined to plasma, the VLDL-TG pool can be viewed as a single compartment system. Therefore, the decay of SA in radiolabeled VLDL-TG after a bolus injection is monoexponential and can be described using the following general equation: SAt=SApeak×e−FCR×twhere SAt = SA of VLDL-TG (dpm/μmol) at time t, SApeak = SA of VLDL-TG at the peak of enrichment, FCR = fractional catabolic rate (pools/min), and t = time (min). The FCR of the VLDL-TG pool is easily calculated from the slope of the log SA (dpm/μmol VLDL-TG) versus time curve as described previously (16Reaven G.M. Hill D.B. Gross R.C. Farquhar J.W. Kinetics of triglyceride turnover of very low density lipoproteins of human plasma..J. Clin. Invest. 1965; 44: 1826-1833Crossref PubMed Scopus (130) Google Scholar): FCR=−InSAt−InSApeakt Because the VLDL tracer could potentially be "contaminated" by small amounts of free [1-14C]triolein, the slope was calculated from 120 min onward. This time point was chosen because control experiments performed in our laboratory have shown that any free [1-14C]triolein injected at time 0 has been cleared from the circulation and contributes in a nominal way to the VLDL SA decay curve after 2 h (12Gormsen L.C. Jensen M.D. Nielsen S. Measuring VLDL-triglyceride turnover in humans using ex vivo-prepared VLDL tracer..J. Lipid Res. 2006; 47: 99-106Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). VLDL-TG kinetics were computed as follows: VLDL-TG production rate (μmol/min)=FCR/60 × CVLDL-TG×PV VLDL-TG secretion rate (μmol/L plasma/min) = FCR/60 × CVLDL-TG VLDL-TG clearance (ml/min) = VLDL-TG production rateCVLDL-TGwhere CVLDL-TG = average VLDL-TG concentration from time 0 to 300 min and PV = plasma volume. Plasma volume was calculated based on each participant's FFM (17Boer P. Estimated lean body mass as an index for normalization of body fluid volumes in humans..Am. J. Physiol. Endocrinol. Metab. 1984; 247: F632-F636PubMed Google Scholar). PV = 0.055 × FFM Serum growth hormone was analyzed with a double monoclonal immunofluorometric assay (Delfia; Wallac Oy, Turku, Finland). Serum insulin was measured with an immunoassay (DAKO). Serum FFA was determined by a colorimetric method using a commercial kit (Wako Pure Chemical Industries, Ltd., Neuss, Germany). Epinephrine and norepinephrine were measured by HPLC. Plasma TG concentration was analyzed using a COBAS Fara II. Blood samples were collected in chilled tubes and placed in an ice bath, and plasma was then separated as quickly as possible by centrifugation (3,600 rpm at 4°C for 10 min). Aliquots of plasma (3 ml) were refrigerated at 4°C for isolation of VLDL upon completion of the examination. The remaining samples were stored at −20°C for later analysis. One week before the start of the study, the participants were instructed by a clinical dietician to consume a weight-maintaining diet and to refrain from physical exercise for 3 days before examination. A blood sample was drawn to ensure that the participants had normal values for hematological, liver, and renal indices. A second blood sample was drawn to isolate and label VLDL. All female participants had a negative pregnancy test. On the evening preceding the study day, subjects were admitted to the research laboratory at 11:00 PM and fasted overnight. Before the examination, they were allowed to use the toilet and were then allowed to either sit or lie in bed wearing light hospital clothing in a room with ambient temperature of 22–24°C. They remained in bed throughout the study. Two intravenous catheters (Venflon; Viggo AB, Helsingborg, Sweden) were inserted, one in an antecubital vein of the left arm and the other in a dorsal hand vein on the right. The hand was then placed in a heated box at 65°C, allowing for arterialized blood samples to be drawn. At 8:00 AM (time 0), after drawing baseline blood samples (insulin, TG, catecholamines, growth hormone), 14C-labeled VLDL was administered as a bolus injection over 10 min. At the same time, a constant infusion of [3H]palmitate (0.3 μCi/min) was initiated and maintained for 1 h. Blood samples were drawn at 0, 30, 40, 50, and 60 min and analyzed for palmitate concentration and SA, insulin, growth hormone, and catecholamines. At 0, 30, 60, 120, 180, 240, and 300 min, blood samples were drawn and analyzed for VLDL-TG SA and total TG concentration. Indirect calorimetry was performed between 30 and 60 min. After completion of the study, intravenous catheters were removed and the participants were dismissed. All data are expressed as means ± SEM unless stated otherwise. Variables that were not normally distributed were log-transformed before statistical processing. Between-group differences were analyzed using Student's t-test or the Mann-Whitney two-samples test. Correlations were evaluated by Pearson's r. Two-way ANOVA was performed to test the effect of sex and time on VLDL-TG concentration during the investigation. Univariate correlation analysis was used to identify variables that potentially affected VLDL-TG production (see Table 2 below). Accordingly, variables displaying the strongest univariate correlation (P < 0.25) with VLDL-TG production (REE and insulin level) were selected for consideration in a multivariate linear regression analysis to assess significant determinants of VLDL-TG production. In addition to REE and insulin, the following candidate independent variables were also considered: palmitate flux, FM, visceral fat area, and sex. Palmitate concentration was also entered instead of palmitate flux in the analysis. Using the approach of stepwise variable selection (stepping up), we accepted variables with a significance level of 0.1 or less into the final model. Because of the well-known colinearity between REE and FFM and because FFM is used in the equation to calculate VLDL production, models in which only REE and FFM were included were analyzed first. In both nonstepped and stepped multiple linear regression analyses, REE consistently emerged as a significant independent predictor of VLDL-TG production, whereas FFM was consistently eliminated. The latter was also true when FFM was forced into the model before REE. Thus, REE turned out to be a stronger determinant of VLDL-TG production despite the fact that FFM is used in the calculation of VLDL-TG production. Therefore, FFM was not included in the final models. To ensure that FM, palmitate flux, and palmitate concentration were not prematurely excluded from consideration, they were forced separately into nonstepwise models that also included the variables found to be significant in the stepwise model. Differences were considered significant at P < 0.05.TABLE 2.Metabolic parametersParameterMen (n = 13)Women (n = 12)PMean p-VLDL-TG (mmol/l)0.26 ± 0.030.27 ± 0.04NSMean p-total TG (mmol/l)0.77 ± 0.070.87 ± 0.09NSVLDL-TG FCR (pools/h)0.23 ± 0.020.26 ± 0.01NSVLDL-TG production (μmol/min)3.20 ± 0.322.82 ± 0.42NSVLDL-TG secretion (μmol/l plasma/min)0.9 ± 0.11.1 ± 0.2NSVLDL-TG clearance (ml/min)14.2 (7.2–23.6)10.0 (6.8–13.7)NSp-Palmitate (μmol/l)118 ± 11117 ± 9NSPalmitate flux (μmol/min)128.5 (91–226)120.5 (78–250)NSLipid oxidation (kcal/24 h)693 ± 55450 ± 74<0.05Carbohydrate oxidation (kcal/24 h)683 ± 59628 ± 45NSProtein oxidation (kcal/24 h)466 ± 50360 ± 40NSFCR, fractional catabolic rate; p, plasma; TG, triglyceride. Data are means ± SEM or median and (range). Open table in a new tab FCR, fractional catabolic rate; p, plasma; TG, triglyceride. Data are means ± SEM or median and (range). The mean and range of body mass index values of our volunteers are given in Table 1 . The expected differences in FFM, FM, and percentage body fat between men and women were present. There were no statistically significant differences in intra-abdominal fat volumes between men and women, but women had significantly greater abdominal subcutaneous fat volume. REE was significantly greater in men than in women; however, the respiratory exchange ratio was not different, indicating similar relative fuel oxidation rates in men and women.TABLE 1.Subject characteristicsCharacteristicMen (n = 13)Women (n = 12)PAge (years)25.9 ± 1.024.7 ± 1.6NSBody mass index (kg/m2)23.8 (21.1–29.8)23.8 (20.2–35.6)NSPlasma volume (liters)3.55 ± 0.992.42 ± 0.80<0.0001Fat-free mass (kg)64.5 ± 1.844.1 ± 1.5<0.0001Total fat mass (kg)12.3 (5.1–20.4)17.9 (12.1–49.1)0.01Body fat (%)15.2 ± 1.631.4 ± 2.8<0.0001Visceral fat area (cm2)93 ± 1475 ± 17NSSubcutaneous fat area (cm2)65 (31–139)101 (61–395)<0.05REE (kcal/24 h)1836 ± 371438 ± 48<0.0001Respiratory exchange ratio0.85 ± 0.010.87 ± 0.01NSGrowth hormone (ng/ml)0.93 (0.04–15.93)0.83 (0.01–7.81)NSInsulin (pmol/l)24 (13–48)26 (14–87)NSEpinephrine (pg/ml)31.5 (25–67)65 (25–222)<0.05Norepinephrine (pg/ml)128 (110–283)172 (119–339)NSREE, resting energy expenditure. Data are means ± SEM or median and (range). Open table in a new tab REE, resting energy expenditure. Data are means ± SEM or median and (range). Plasma norepinephrine, human growth hormone, and insulin concentrations measured at time 0 were comparable in men and women, whereas plasma epinephrine concentrations were significantly greater in women than in men (Table 1). Because epinephrine is a known stimulator of hormone-sensitive lipase, the principal enzyme responsible for adipocyte FFA release, correlations between epinephrine and FFA concentration (palmitate), FFA turnover (palmitate flux), and VLDL-TG production were analyzed subsequently. However, no significant correlations between epinephrine concentration and palmitate concentration (r = −0.09, P = 0.64), palmitate flux (r = 0.08, P = 0.70), or VLDL-TG production (r = 0.01, P = 0.96) were observed; therefore, epinephrine was not considered for inclusion in the final multivariate regression model. Plasma total TG concentration and VLDL-TG concentration remained constant throughout the study (Fig. 1, bottom panel), with no sign of a sex or time effect. The rate of decline in VLDL-SA is shown in Fig. 1, top panel. VLDL-SA was significantly higher in women compared with men but declined similarly in both sexes. Thus, no significant difference was observed for VLDL-TG FCR, VLDL-TG production, or VLDL-TG clearance (Table 2). Because the initial multiple regression analysis (see below) revealed independent effects of REE and plasma insulin, these factors were plotted against VLDL-TG production. The top panel of Fig. 2 depicts the relationship between VLDL-TG production and REE. For the entire group, there was a significant positive correlation between VLDL-TG production and REE (r = 0.45, P = 0.02). Adjusting for sex, however, displayed two parallel regression slopes steeper than the combined slope (men, r = 0.57, P = 0.043; women, r = 0.58, P = 0.048) with significantly different y intercepts (P < 0.01). The relationship between VLDL-TG production and fasting plasma insulin was also significant. To better appreciate the apparent sex effect shown in the upper panel plot (VLDL-TG production vs. REE), VLDL-TG production was plotted against the residuals derived from the linear regression of VLDL-TG production on REE (Fig. 2, bottom panel). It appears that the effect of insulin is independent of a sex effect. One woman diverged markedly from the remaining subjects (upper left corner of the plot). However, exclusion of this subject could not be justified. Table 3 shows the univariate correlations between VLDL-TG production and the candidate independent predictors. Based on a cutoff value of P < 0.25, we identified the determinants most likely to have a major impact on VLDL-TG production to include them in a multivariate regression analysis. Because sex changed the intercept value between REE and VLDL-TG markedly (Fig. 2), it was also included in the final model. Tables 4 and 5 include both the nonstepped and the stepwise (forward) regression analyses. Using the stepwise selection method, we found that the best multivariate model to predict VLDL-TG production included log-insulin (slope, 1.06; P = 0.02) and REE (slope, 0.0021; P = 0.02). The adjusted r2 for this model was 0.32. Forcing sex or indices of body composition into this model did not improve the overall r2. However, if data from the outlier in Fig. 2 (lower panel) were excluded, sex emerged as a significant independent determinant of VLDL-TG production.TABLE 3.Univariate correlationsVLDL Production (μmol/min)FCR (pools/h)VariablerPrPp-Palmitate (μmol/l)−0.26720.1970.21920.292Palmitate flux (μmol/min)0.09160.670.30590.146REE (kcal/24 h)0.45450.022−0.17010.416Intra-abdominal fat (cm2)0.27440.2290.41050.065Fat mass (g)0.24630.2350.41430.039p-Insulin (pmol/l)0.45930.021−0.17480.403VLDL production (μmol/min)——−0.14320.495VLDL pool (μmol)0.7565 0.5 are not shown. Open table in a new tab TABLE 4.Multivariate, nonstepped, regression analysisVariableParameter EstimateStandard ErrorPIntercept−7.293.390.044Fat mass (g)−0.0000170.00003510.63Log insulin (pmol/l)0.890.540.114REE (kcal/24 h)0.00430.00190.036Sex (woman)1.341.080.227Dependent variable: VLDL-TG production (μmol/min). Open table in a new tab TABLE 5.Multivariate, stepwise forward, regression analysisVariableParameter EstimateStandard ErrorPIntercept−3.951.920.05Log insulin (pmol/l)1.060.430.02REE (kcal/24 h)0.00210.00090.02Dependent variable: VLDL-TG production (μmol/min). Open table in a new tab Univariate correlations with P > 0.5 are not shown. Dependent variable: VLDL-TG production (μmol/min). Dependent variable: VLDL-TG production (μmol/min). Figure 3 shows the relationship between VLDL-FCR and VLDL-TG pool size. A significant inverse correlation was found, indicating a progressively slow relative turnover of the VLDL-TG pool with increasing VLDL-TG pool size. This study was undertaken to determine the factors that affect VLDL-TG production. We measured VLDL-TG production using a recently validated method based on the plasma disappearance rate of a bolus infusion of ex vivo labeled VLDL-TG particles (12Gormsen L.C. Jensen M.D. Nielsen S. Measuring VLDL-triglyceride turnover in humans using ex vivo-prepared VLDL tracer..J. Lipid Res. 2006; 47: 99-106Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar). Our results show that tota
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