1 Nemours Research Programs and 2 Division of Endocrinology, Department of Research, Nemours Children's Clinic, Jacksonville, Florida 32207
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Increased concentrations of plasma fibrinogen, an independent risk factor for cardiovascular disease (CVD), in obese children have been reported. The underlying mechanism for this, however, remains to be defined. In the current study, we measured the fractional synthesis rates (FSR) of plasma fibrinogen in six healthy postpubertal obese girls [body mass index (BMI) 36.6 ± 1.8 kg/m2; age 16.6 ± 0.5 yr] and six age-matched lean normal control girls (BMI 20.8 ± 0.7 kg/m2; age 16.4 ± 0.4 yr) during a primed, continuous infusion of L-[1-13C]leucine in the postabsorptive state. The method involved purification of plasma fibrinogen by use of immunoaffinity chromatography followed by measurement of [13C]leucine enrichment using gas chromatography-combustion-isotope ratio mass spectrometry. The FSR of fibrinogen in obese girls (35.06 ± 2.61%/day) was almost double that in lean girls (17.02 ± 1.43%/day), and this increase was associated with a relative increase in plasma concentration of fibrinogen as well as BMI in the subjects studied. Obese subjects had high fasting insulin levels (138 ± 47 pmol/l) compared with lean subjects (54 ± 11 pmol/l), whereas their glucose concentrations were similar (4.5 ± 0.3 mmol/l in obese and 4.4 ± 0.4 mmol/l in lean subjects), suggesting insulin resistance. The doubling of the FSR of fibrinogen provides novel insight into the mechanism of elevated levels of plasma fibrinogen and suggests a primary role for increased synthesis in producing the hyperfibrinogenemia associated with obesity. This finding may have important implications in the design of therapies for modulating plasma fibrinogen levels in obesity and/or CVD in childhood.
cardiovascular disease; fractional synthesis rate; metabolism; obesity
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
OBESITY CAUSES AND/OR AGGRAVATES many health problems, both independently and in association with other diseases. In particular, it is reported to be associated with the development of cardiovascular disease (CVD) (11, 12, 23, 30). Factors that predispose individuals to CVD are thought to develop during childhood (4, 5). Besides being an acute-phase reactant protein, fibrinogen plays an important role in promoting atherogenesis and thrombogenesis. Recent studies have demonstrated that elevated plasma concentration of fibrinogen is an independent risk factor for CVD (8, 22, 24, 40). Plasma fibrinogen concentrations were even more strongly associated with cardiovascular death than plasma cholesterol (29). Several previous studies have clearly shown increased plasma fibrinogen concentrations in uncontrolled diabetes, vascular disease, and obesity (11, 30); however, the mechanisms that regulate plasma fibrinogen levels in vivo in these conditions are poorly understood.
There are several pathways by which acute or chronic increase in fibrinogen levels may lead to atherosclerotic and cardiovascular events. These include infiltration of the vessel wall by fibrinogen, rheological effects due to increased blood viscosity, increased platelet aggregation and thrombus formation, and increased fibrin formation. In a recent meta-analysis (13), a series of factors such as cigarette smoking, positive energy balance, diabetes mellitus, obesity, pregnancy, high dietary fat intake, increasing age, menopause, inflammation, thrombin, and vascular damage have all been identified to affect plasma fibrinogen concentrations. Genetic and environmental determinants are other important factors contributing to the changes in plasma concentrations of fibrinogen (22, 35). The concentration of plasma fibrinogen represents an imbalance between the production (synthesis) and the disposal (breakdown) rates of this protein. Although currently there is no viable method for the measurement of fibrinogen breakdown in humans, the ability to measure changes in synthesis rate of this protein could offer insight into the mechanisms that regulate this risk factor for CVD.
In humans, fibrinogen FSR was increased by acute insulin deficiency in type 1 diabetes (9) and was decreased by short-term insulin infusion in both normal subjects and subjects with type 1 diabetes (9, 10). Additionally, Tessari et al. (42) showed an acute stimulation of fibrinogen synthesis by glucagon. In a very recent study, Hunter et al. (21) reported a primary role for increased synthesis in producing hyperfibrinogenemia associated with smoking. Fu and Nair (16), however, observed no increase in the synthesis rate of fibrinogen in older people compared with young subjects despite increased levels of plasma fibrinogen levels in the former group. They concluded that the elevated levels of plasma fibrinogen found in the older population are likely due to reduced breakdown of this hepatic protein. Many of the risk factors for CVD, including obesity, begin to develop in childhood. Understanding the impact of the fibrinogen synthesis rate on the elevation of plasma fibrinogen levels may better enable the development of therapeutic research toward alleviating this independent risk factor for CVD. The objective of the present study was to better understand the nature of changes in plasma fibrinogen concentration by measuring the FSR of fibrinogen in obese and lean adolescent females.
![]() |
RESEARCH DESIGN AND METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Materials
L-[1-13C]leucine [99% atom percent excess (APE)] was purchased from Cambridge Isotope Laboratories. Purchased lots of stable isotopes were tested for chemical, isotopic, and optical purity by gas chromatography-mass spectrometry (GC-MS). Solutions of the sterile and bacteria-free isotopes were passed through a 0.22-µm filter and stored in sterile, sealed containers <24 h before each infusion and kept at 4°C until used.Study Subjects and Design
Six obese (BMI >30 kg/m2) and six healthy lean control adolescent girls (age >15 yr <18 yr; Tanner stage IV) were studied. These subjects were a part of a larger study protocol that assessed whole body protein metabolism in these subjects, and some of their data have been published previously (28). The study protocol was approved by the Nemours Children's Clinic Research Review Committee and Baptist Medical Center/Wolfson Children's Hospital Institutional Review Committee. The subject characteristics are given in Table 1. All subjects consumed a weight-maintaining diet for
|
Body weight and height were measured using stadiometers and
calibrated scales. BMI was calculated as body weight in kilograms divided by height in meters squared, and the 1994 clinical guidelines (19) were used to define obesity (BMI 30
kg/m2). The percent fat mass (%FM) was measured using the
sum of the thickness of four skinfolds taken with calipers
(
Technology, Cambridge, MA). The concentration of fibrinogen was
determined immunologically by automated laser rate nephelometry. Serum
albumin concentrations were measured colorimetrically by the bromcresol green method with a kit from Sigma Diagnostics (St. Louis, MO). Free
fatty acids (FFA) were also measured by colorimetric methods with a kit
from Sigma Diagnostics. Plasma glucose was measured by a glucose
oxidase method with a Beckman glucose analyzer (Beckman Instruments,
Palo Alto, CA). Plasma insulin concentrations were measured by RIA at
Endocrine Sciences Laboratories (Calabases Hills, CA).
Analytical Procedures
Precursor pool enrichment.
The plasma enrichment of -[13C]ketoisocaproate
([13C]KIC), an index of intracellular leucine enrichment,
was determined by GC-MS with selected ion monitoring and electron
impact ionization of a t-butyldimethylsilyl derivative, as
previously described (27, 38).
Purification of fibrinogen and measurement of
[13C]leucine enrichments.
Incorporation of labeled leucine into fibrinogen, produced by the liver
and subsequently secreted into plasma, was measured after isolation
(15), subsequent hydrolysis, and GC-combustion-isotope ratio mass spectrometry (GC-Com-IRMS) (2, 49). By use of the precursor-product relationship, the fractional synthesis rate (FSR)
was calculated (46, 48). Briefly, the affinity column of
fibrinogen antibody was prepared by coupling monoclonal anti-human fibrinogen antibodies (the anti-human fibrinogen antibody was generously supplied by Dr. S. Nair, Mayo Clinic, Rochester, MN) onto
the Affi-gel 10 (Bio-Rad). The separation of the protein was carried
out using an automated biological system (model ES-1, Bio-Rad), as
described previously. The system allowed automatic control of buffer
change, loading speed, and elution and collection. Approximately 500 µl of plasma were passed through the affinity column containing
anti-human fibrinogen antibody. The bound protein was eluted with
elution buffer (0.1 M sodium acetate with 2 N urea, pH 3.7) and
collected for further treatment. The entire process was performed at
room temperature. The purity of the protein separated was always
ascertained by analytical SDS electrophoresis followed by silver
staining. The separated proteins were precipitated with 10%
trichloroacetic acid. The proteins were recovered by centrifugation and
then hydrolyzed using 1 ml of 6 N HCl at 110°C for ~24 h. The
hydrolysates were further purified by passing them through a cation
exchange column containing 1 ml of AG 50W-X8, 100-200 mesh,
H+ form. The dried amino acids were reconstituted in 0.01 M
HCl and stored at 80°C until MS analysis.
Determination of [13C]leucine enrichment in purified plasma fibrinogen. We used the GC-Com-IRMS technique as previously described (2) for the measurement of [13C]leucine enrichment of fibrinogen (15). The leucine derived from hydrolysis of fibrinogen was derivatized as its N-heptafluorobutyryl methyl ester, separated on a GC column, and combusted in an online (800-900°C) furnace. The CO2 evolved from the combustion process was subsequently analyzed for its 13CO2-to-12CO2 isotope ratio with an online IRMS (2).
Calculations
The FSR (expressed in %/24 h) of plasma fibrinogen was calculated by dividing the regression slope of isotope enrichment from 180 to 240 min of isotope infusion by the plasma plateau enrichments of [13C]KIC according to the precursor-product relationship
![]() |
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Subjects: Physical and Clinical Characteristics
Table 1 shows the physical and clinical characteristics of the study subjects. Each group, obese and lean control, consisted of six age-matched adolescent girls. By design, the obese subjects had significantly higher BMI (BMI >35 kg/m2; range 34.1-44.4 kg/m2) than lean controls (BMI <25 kg/m2; range 19.4-22.9 kg/m2). Moreover, height was similar between the groups. The %FM and fat-free mass (FFM) values were higher in the obese group (P < 0.01) compared with the lean control group (%FM range: 40.1-46.3 kg for obese and 28.0-35.2 kg for the lean; FFM range: 49.2-65.9 kg for obese and 33.1-42.4 kg for lean). The obese and lean control groups had similar levels of fasting glucose concentrations, whereas the fasting insulin levels were significantly higher in the obese group (P < 0.05). The concentration of plasma fibrinogen was significantly higher in the obese compared with the lean group (2.36 ± 0.07 g/l in the obese and 1.85 ± 0.06 g/l in the lean group; P < 0.01), and there was significant correlation between fibrinogen concentration and BMI (r = 0.81; P = 0.003). Similarly, FFA level was higher in the obese group than in the lean control group (0.61 ± 0.05 mmol/l for the obese and 0.43 ± 0.05 mmol/l for the lean controls; P < 0.01). The albumin concentration, however, was lower in the obese subjects than in lean control subjects (3.9 ± 0.3 g/l in obese vs. 4.6 ± 0.3 g/l in lean; P = 0.04).FSR and ASR Fibrinogen
Figure 1B depicts the increase in tracer enrichment in fibrinogen between 180 and 240 min of isotopic infusion in the obese and lean control groups, showing a linear increment of isotope enrichment in fibrinogen in both groups. The regression slope of the line correlating the change of fibrinogen-bound leucine enrichment vs. time was consistently higher in the obese group than in the lean control group. The plateau value of [13C]KIC was used as the precursor pool enrichment (Fig. 1A) for the calculation of FSR of fibrinogen.
|
Figure 2A shows the FSR values
of fibrinogen in the obese and lean groups. The FSR of fibrinogen in
the control group, 17.02 ± 1.43%/day (range between 12 and
21%/day) on the basis of calculation with plasma KIC as the precursor
pool, was less than that reported (~28%/day) in young adults
(16) but close to that reported by Hunter et al.
(21). The mean FSR of fibrinogen in the obese subjects,
35.06 ± 2.61%/day (range 22-44%/day), was almost double that in the lean control group. The FSR of fibrinogen showed
statistically significant correlation between BMI (r = 0.79; P = 0.002) and plasma concentration of fibrinogen
(r = 0.74; P = 0.009) in the subjects
we studied (Fig. 3, A and
B).
|
|
The ASR of fibrinogen was higher in the obese subjects (Fig.
2B) compared with the lean controls (37.6 ± 2.9 mg
· kg1 · day
1 in obese
children and 14.0 ± 1.2 mg · kg
1 · day
1 in lean;
P < 0.01). The intravascular fibrinogen pool size was also higher (P < 0.01) in the obese group than in the
lean group.
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The current study demonstrates an upregulation of the synthesis rate of fibrinogen in obese postpubertal young females compared with lean age-matched controls. The almost doubling of the FSR of fibrinogen along with significant increase in the plasma fibrinogen concentration suggests a primary role for augmented synthesis of fibrinogen in producing the hyperfibrinogenemia in obese female adolescents. This increase in the FSR was directly associated with BMI and plasma fibrinogen concentration. In addition, the obese subjects had highly elevated fasting insulin levels, whereas the glucose levels were comparable in both groups. This suggests that the increased fibrinogen synthesis may likely be mediated through a mechanism involving insulin resistance. The observed increase in the FSR of fibrinogen in obese adolescent females provides new insight into the mechanism of elevated levels of plasma fibrinogen in these subjects and its potentially detrimental role as a CVD risk factor later in life.
Several studies have established the connection between elevated levels of plasma fibrinogen, obesity, and CVD (4, 6, 8, 14, 24, 25). However, it remains unclear whether an elevated plasma level of fibrinogen is a consequence of increased fibrinogen synthesis or decreased fibrinogen degradation. In addition to genetic and environmental factors (22, 35), impaired fibrinolytic capacity in CVD and obesity may contribute to hyperfibrinogenemia by limiting endogenous fibrinolysis (36, 37). It may, however, be noted that the substantially elevated level of plasma fibrinogen in obesity and CVD is not fully accounted for by this mechanism. The results from the current study, demonstrating higher rates of fibrinogen synthesis in obese than in lean control girls along with its increased plasma concentration, support the hypothesis that increased synthesis of fibrinogen plays an important role in the regulation of plasma levels of fibrinogen in childhood obesity, at least in females. Of interest, we found an increase of fibrinogen ASR (ASR = FSR × intravascular fibrinogen pool) in the obese subjects. However, in contrast to the direct measurement of FSR, the ASR of fibrinogen is an estimated value based on several assumptions. The ASR values are likely influenced by differences in body composition in the two groups, and it is possible that plasma volume is overestimated in the obese group. Additionally, because the ASR of fibrinogen is normalized for body weight, it potentially introduces an error, because the obese group obviously has higher amounts of fat. Regardless of these limitations, the increased ASR and FSR values in the obese group suggest that an upregulation of the synthesis of fibrinogen has a major role in increasing the plasma pool of fibrinogen in the obese subjects. The increase in the FSR of plasma fibrinogen was also significantly correlated with plasma fibrinogen concentration and BMI (Fig. 3, A and B).
The current study, however, is limited, only in postpubertal girls, and needs to be established and validated in a larger population, including both genders. The absolute values reported here are different from the previously reported values of FSR of fibrinogen in adult subjects (16, 44). The current study was performed in female adolescents, and there are no reference values available for FSR of fibrinogen in children with which to compare. However, previous studies have suggested that there are no gender-related differences in the fibrinogen synthesis rate in either young or old populations (16, 44). Although Fu and Nair (16) did not notice any gender effect on fibrinogen synthesis rate, they showed that there was an age-dependent increase in the concentration of plasma fibrinogen that was not associated with an increase in the synthesis rate of fibrinogen. This is suggestive of a decrease in the breakdown of fibrinogen with advancing age. In contrast to the current study, the study by Fu and Nair was performed in a healthy aging population and not in obese young people, which may explain the difference. Additionally, it is unlikely that insulin, glucagon, and glucose homeostasis had a role in the regulation of fibrinogen synthesis in the study in the elderly population, as there was no significant difference in blood concentrations of insulin, glucagon, and glucose observed in the study by Fu and Nair. This is contrary to the apparent insulin resistance observed in the current study of obese girls.
Several assumptions were made for the calculation and interpretation of
the data. We assumed that plasma -KIC is in equilibrium with
intrahepatic
-KIC. Previous studies have shown that plasma
-KIC
enrichment is in reality close to the very low density lipoprotein apoB-100-bound leucine at plateau (7, 43, 45) and also to
the leucyl t-RNA (1). Therefore, plasma
-KIC enrichment can be used as a valid surrogate precursor pool for the calculation of
FSR of hepatic proteins. A recent study by Barazzoni et al. (5) has also demonstrated that hepatic venous leucine
enrichment (likely to represent intracellular leucine in liver) is not
different from circulating KIC enrichment. Therefore, on the basis of
various studies cited (1, 5, 7, 43, 45), it is likely that the calculation of FSR of fibrinogen based on plasma
-KIC as the
precursor pool enrichment is appropriate.
From basic mechanistic as well as therapeutic considerations, the results from this study have important implications. This study for the first time provides evidence to show that increased FSR of plasma fibrinogen has a major role in regulating the plasma concentration of this liver protein in obese postpubertal girls. The underlying molecular mechanism for these changes, however, remains unclear. Although it is well known that obesity is associated with resistance to insulin's effects on glucose metabolism, it is not clear whether it is resistant to its effects on protein metabolism also. Conditions such as Cushing's syndrome (41), old age (3, 17), and non-insulin-dependent diabetes (39, 47) are not associated with increased postabsorptive protein turnover. On the other hand, we observed that obesity in young postpubertal females is associated with insulin resistance in both peripheral carbohydrate and protein metabolism (28). In our study, the glucose concentrations during an intravenous glucose tolerance test (IVGT) in the obese vs. the lean controls were superimposable, similar to a glucose clamp, whereas the insulin concentrations during an IVGT were markedly different (28). This clearly demonstrates marked insulin resistance in the obese group that we studied. The increased FSR of fibrinogen in the obese subjects observed in the current study may thus be related to insulin resistance. Additionally, elevated levels of FFA, along with low levels of albumin, have been observed in the obese subjects compared with the lean subjects in the current study. Because albumin binds to FFAs and FFAs are strong stimuli of hepatic synthesis of fibrinogen, as shown in animal studies (20, 32, 34), changes in the FFA-to-albumin ratio may also contribute to the regulation of the synthesis of fibrinogen (20, 33, 34). Obviously, this needs to be studied further. It is also important to study further whether reported genetic variations in fibrinogen gene expression are important in modulating the response to specific environmental factors.
In conclusion, data from the present study show that, in obese adolescent females, the levels of plasma fibrinogen are elevated primarily because of the increased synthesis rate of this protein. This is an important step in elucidating the pathways through which fibrinogen levels are increased in obese adolescent females. This may contribute to the risk of CVD observed in obese patients and may be a consequence of insulin resistance. It is important to consider this aspect in the optimization therapies to control this independent risk factor in obesity and CVD. Future studies that investigate the effects and mechanisms of pharmacological and nonpharmacological interventions in the modulation of fibrinogen levels (along with other risk factors) in obese children and adults are warranted.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank the volunteers for participating in this study. We are grateful to Burnese Rutledge and the nursing staff of the CRC at the Wolfson Children's Hospital for their superb assistance, Brenda Sager and Lynda Everline for their skilled technical assistance, Annie Rini and Susan Welch for support in conducting the studies, Barbara Tyler for superb secretarial support, Dr. Hossein Yarandi for statistical help, Dr. Robert Olney for helpful comments, and Dr. Sreekumaran Nair for graciously providing the human monoclonal antibody for fibrinogen and for helpful comments.
![]() |
FOOTNOTES |
---|
This study was supported in part by grants from the Nemours Research Programs and the American Heart Association Scientist Development Award.
Address for reprint requests and other correspondence: P. Balagopal, Dept. of Research, Nemours Children's Clinic, 807 Children's Way, Jacksonville, Florida 32207 (E-mail : bbalagopal{at}nemours.org).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
10.1152/ajpendo.00412.2001
Received 17 September 2001; accepted in final form 28 November 2001.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Ahlman, B,
Charlton MR,
Fu AZ,
Berg C,
O'Brien P,
and
Nair KS.
Insulin's effect on synthesis rates of liver proteins. A swine model comparing various precursors of protein synthesis.
Diabetes
50:
947-954,
2001
2.
Balagopal, P,
Ford GC,
Ebenstein DB,
Nadeau DA,
and
Nair KS.
Mass spectrometric methods for determination of [13C]leucine enrichment in human muscle protein.
Anal Biochem
239:
77-85,
1996[ISI][Medline].
3.
Balagopal, P,
Rooyackers OE,
Adey DB,
Ades PA,
and
Nair KS.
Effects of aging on in vivo synthesis of skeletal muscle myosin heavy-chain and sarcoplasmic protein in humans.
Am J Physiol Endocrinol Metab
273:
E790-E800,
1997
4.
Bao, W,
Srinivasan SR,
and
Berenson G.
Plasma fibrinogen and its correlates in children from a biracial community: the Bogalusa Heart Study.
Pediatr Res
33:
323-326,
1993[Abstract].
5.
Barazzoni, R,
Meek SE,
Ekberg K,
Wahren J,
and
Nair KS.
Arterial KIC as marker of liver and muscle intracellular leucine pools in healthy and type 1 diabetic humans.
Am J Physiol Endocrinol Metab
277:
E238-E244,
1999
6.
Cacciari, E,
Balsamo A,
Palareti G,
Cassio A,
Argento R,
Poggi M,
Tassoni P,
Cicognani A,
Tacconi M,
Pascucci MG,
and
Coccheri S.
Haemorheologic and fibrinolytic evaluation in obese children and adolescents.
Eur J Pediatr
147:
381-384,
1988[ISI][Medline].
7.
Cummings, MH,
Watts GF,
Umpleby AM,
Hennessy TR,
Naoumova R,
Slavin BM,
Thompson GR,
and
Sonksen PH.
Increased hepatic secretion of very-low-density lipoprotein apolipoprotein B-100 in NIDDM.
Diabetologia
38:
959-967,
1995[ISI][Medline].
8.
Danesh, J,
Cillins R,
Appleby P,
and
Peto R.
Association of fibrinogen, C-reactive protein, albumin, or leukocyte count with coronary heart disease: meta-analyses of prospective studies.
JAMA
279:
1477-1482,
1998
9.
De Feo, P,
Gaisano MG,
and
Haymond MW.
Differential effects of insulin deficiency on albumin and fibrinogen synthesis in humans.
J Clin Invest
88:
833-840,
1991[ISI][Medline].
10.
De Feo, P,
Volpi E,
Lucidi P,
Cruciani G,
Reboldi G,
Siepi D,
Mannarino E,
Santeusanio F,
Brunetti P,
and
Bolli GB.
Physiological increments in plasma insulin concentrations have selective and different effects on synthesis of hepatic proteins in normal humans.
Diabetes
42:
995-1002,
1993[Abstract].
11.
Eckel, RH,
and
Krauss RM.
American Heart Association call to action: obesity as a major risk factor for coronary heart disease. AHA Nutrition Committee.
Circulation
97:
2099-2100,
1998
12.
Ellis, KJ.
Visceral fat mass in childhood: a potential early marker for increased risk of cardiovascular disease.
Am J Clin Nutr
65:
1887-1888,
1997[ISI][Medline].
13.
Ernst, E,
and
Resch KL.
Fibrinogen as a cardiovascular risk factor: a meta-analysis and review of the literature.
Ann Intern Med
118:
956-963,
1993
14.
Fanari, P,
Somazzi R,
Nasrawi F,
Ticozzelli P,
Grugni G,
Agosti R,
and
Longhini E.
Haemorheological changes in obese adolescents after short-term diet.
Int J Obes
17:
487-494,
1993[ISI].
15.
Fu, AZ,
Morris JC,
Ford GC,
and
Nair KS.
Sequential purification of human apolipoprotein B-100, albumin, and fibrinogen by immunoaffinity chromatography for measurement of protein synthesis.
Anal Biochem
247:
228-236,
1997[ISI][Medline].
16.
Fu, A,
and
Nair KS.
Age effect on fibrinogen and albumin synthesis in humans.
Am J Physiol Endocrinol Metab
275:
E1023-E1030,
1998
17.
Fukagawa, NK,
Minaker KL,
Young VR,
Matthews DE,
Bier DM,
and
Rowe JW.
Leucine metabolism in aging humans: effect of insulin and substrate availability.
Am J Physiol Endocrinol Metab
256:
E288-E294,
1989
18.
Gregersen, MI,
and
Rawson RA.
Blood volume.
Physiol Rev
39:
307-342,
1959
19.
Himes, JH,
and
Dietz WH.
Guidelines for overweight in adolescent preventive services: recommendations from an expert committee. The Expert Committee on Clinical Guidelines for Overweight in Adolescent Preventive Services.
Am J Clin Nutr
50:
307-316,
1994[Abstract].
20.
Hostmark, AT.
Serum fatty acid/albumin molar ratio and the risk of diseases.
Med Hypotheses
44:
539-541,
1995[ISI][Medline].
21.
Hunter, KA,
Garlick PJ,
Broom I,
Anderson E,
and
McNurlan MA.
Effects of smoking and abstention from smoking on fibrinogen synthesis in humans.
Clin Sci (Colch)
100:
459-465,
2001[ISI][Medline].
22.
Kannel, WB.
Influence of fibrinogen on cardiovascular disease.
Drugs
53:
32-40,
1997.
23.
Kannel, WB,
D'Agostino RB,
and
Cobb JL.
Effect of weight on cardiovascular disease.
Am J Clin Nutr
63:
419S-422S,
1996[Abstract].
24.
Kannel, WB,
Wolf PA,
Castelli WP,
and
D'Agostino RB.
Fibrinogen and risk of cardiovascular disease. The Framingham Study.
JAMA
258:
1183-1186,
1987[Abstract].
25.
Licata, G,
Scaglione R,
Avellone G,
Ganguzza A,
and
Corrao S.
Hemostatic function in young subjects with central obesity: relationship with left ventricular function.
Metabolism
44:
1417-1421,
1995[ISI][Medline].
26.
Matthews, DE,
Motil KJ,
Rohrbaugh DK,
Burke JF,
Young VR,
and
Bier DM.
Measurement of leucine metabolism in man from a primed, continuous infusion of L-[1-13C]leucine.
Am J Physiol Endocrinol Metab
238:
E473-E479,
1980
27.
Matthews, DE,
Schwarz HP,
Yang RD,
Motil KJ,
Young VR,
and
Bier DM.
Relationship of plasma leucine and alpha-ketoisocaproate during a L-[1-13C]leucine infusion in man: a method for measuring human intracellular leucine tracer enrichment.
Metabolism
31:
1105-1112,
1982[ISI][Medline].
28.
Mauras, N,
Welch S,
Rini A,
and
Haymond MW.
Ovarian hyperandrogenism is associated with insulin resistance to both peripheral carbohydrate and whole-body protein metabolism in postpubertal young females: a metabolic study.
J Clin Endocrinol Metab
83:
1900-1905,
1998
29.
Meade, TW,
Mellows S,
Brozovie M,
Miller GJ,
Chakrabarti RR,
North WR,
Haines AP,
Stirling Y,
Imeson JD,
and
Thompson SG.
Haemostatic function and ischemic heart disease: principal results of the Northwick Park Heart Study.
Lancet
2:
533-537,
1986[ISI][Medline].
30.
Must, A,
Spadano J,
Coakley EH,
Field AE,
Colditz G,
and
Dietz WH.
The disease burden associated with overweight and obesity.
JAMA
282:
1523-1529,
1999
31.
Nikkila, EA,
and
Kekki M.
Plasma triglyceride transport kinetics in diabetes mellitus.
Metabolism
22:
1-22,
1973[ISI][Medline].
32.
Pickart, LR.
Fat metabolism, the fibrinogen/fibrinolytic system and blood flow: new potentials for the pharmacological treatment of coronary heart disease.
Pharmacology
23:
271-280,
1981[ISI][Medline].
33.
Pickart, LR.
Free fatty acidemia as an inducer of systemic hyperfibrinogenemia and fibrinolytic inhibition.
Inflammation
5:
61-70,
1981[ISI][Medline].
34.
Pickart, LR,
and
Thaler MM.
Free fatty acids and albumin as mediators of thrombin-stimulated fibrinogen synthesis.
Am J Physiol
230:
996-1002,
1976[ISI][Medline].
35.
Ravussin, E,
and
Swinburn BA.
Pathophysiology of obesity.
Lancet
340:
404-408,
1992[ISI][Medline].
36.
Ridker, PM,
Hennekens CH,
Cerkus A,
and
Stampfer MJ.
Plasma concentration of cross-linked fibrin degradation product (D-dimer) and the risk of future myocardial infarction among apparently healthy men.
Circulation
90:
2236-2240,
1994[Abstract].
37.
Schneider, DJ,
Nordt TK,
and
Sobel BE.
Attenuated fibrinolysis and accelerated atherogenesis in type II diabetic patients.
Diabetes
42:
1-7,
1993[Abstract].
38.
Schwenk, WF,
Berg PJ,
Beaufrere B,
Miles JM,
and
Haymond MW.
Use of t-butyldimethylsilylation in the gas chromatographic/mass spectrometric analysis of physiologic compounds found in plasma using electron-impact ionization.
Anal Biochem
141:
101-109,
1984[ISI][Medline].
39.
Staten, MA,
Matthews DE,
and
Bier DM.
Leucine metabolism in type II diabetes mellitus.
Diabetes
35:
1249-1253,
1986[Abstract].
40.
Swahn, E,
Von Schenck H,
and
Wallentin L.
Plasma fibrinogen in unstable coronary artery disease.
Scand J Clin Lab Invest
49:
49-54,
1989[ISI][Medline].
41.
Tessari, P,
Inchiostro S,
Biolo G,
Marescotti MC,
Fantin G,
Boscarato MT,
Merola G,
Mantero F,
and
Tiengo A.
Leucine kinetics and the effects of hyperinsulinemia in patients with Cushing's syndrome.
J Clin Endocrinol Metab
68:
256-262,
1989[Abstract].
42.
Tessari, P,
Iori E,
Vettore M,
Zanetti M,
Kiwanuka E,
Davanzo G,
and
Barazzoni R.
Evidence for acute stimulation of fibrinogen production by glucagon in humans.
Diabetes
46:
1368-1371,
1997[Abstract].
43.
Venkatesan, S,
Cullen P,
Halliday D,
and
Scott J.
Stable isotopes show a direct relation between VLDL apoB overproduction and serum triglyceride levels and indicate a metabolically and biochemically coherent basis for familial combined hyperlipidemia.
Arterioscler Thromb
13:
1118,
1993.
44.
Volpi, E,
Lucidi P,
Bolli GB,
Santeusanio F,
and
De Feo P.
Gender differences in basal protein kinetics in young adults.
J Clin Endocrinol Metab
83:
4363-4367,
1998
45.
Volpi, E,
Lucidi P,
Cruciani G,
Monacchia F,
Reboldi G,
Brunetti P,
Bolli GB,
and
De Feo P.
Contribution of amino acids and insulin to protein anabolism during meal absorption.
Diabetes
45:
1245-1252,
1996[Abstract].
46.
Waterlow, JC,
Garlick PJ,
and
Millward DJ.
Protein Turnover in Mammalian Tissues and in the Whole Body. Amsterdam: North-Holland, 1978.
47.
Welle, S,
and
Nair KS.
Failure of glyburide and insulin treatment to decrease leucine flux in obese type II diabetic patients.
Int J Obes
14:
701-710,
1990[ISI][Medline].
48.
Wolfe, RR.
Radioactive and Stable Isotope Tracers in Biomedicine: Principles and Practice of Kinetic Analysis. New York: Wiley-Liss, 1992.
49.
Yarasheski, KE,
Smith K,
Rennie MJ,
and
Bier DM.
Measurement of muscle protein fractional synthetic rate by capillary gas chromatography/combustion isotope ratio mass spectrometry.
Biol Mass Spectrom
21:
486-490,
1992[ISI][Medline].