![Mean experimental values (symbols) of the tracer to tracee ratio for apo AI-HDL in controls (E) and patients with type II diabetes mellitus (OE). Fits (lines) were calculated using monocompartmental analysis during a primed constant infusion of [ 2 H 3 ]leucine. Data are shown as the mean SEM.](https://www.researchgate.net/profile/Jean-Marie-Bard/publication/11993630/figure/fig1/AS:601733361315846@1520475821497/Mean-experimental-values-symbols-of-the-tracer-to-tracee-ratio-for-apo-AI-HDL-in_Q320.jpg)
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In Vivo Evidence for the Role of Lipoprotein Lipase
Activity in the Regulation of Apolipoprotein AI
Metabolism: A Kinetic Study in Control Subjects and
Patients with Type II Diabetes Mellitus*
R. FRE
´NAIS, H. NAZIH, K. OUGUERRAM, C. MAUGEAIS, Y. ZAI
¨R, J. M. BARD,
B. CHARBONNEL, T. MAGOT, AND M. KREMPF
Human Nutrition Research Center, INSERM, U-539 (R.F., H.N., K.O., C.M., J.M.B., T.M., M.K.), and
Endocrinology, Metabolic Diseases, and Nutrition Clinic (Y.Z., B.C., M.K.), Hoˆtel Dieu, 44093 Nantes
Cedex 01, France
ABSTRACT
The aim of this study was to delineate the role of lipoprotein lipase
(LPL) activity in the kinetic alterations of high density lipoprotein
(HDL) metabolism in patients with type II diabetes mellitus com-
pared with controls. The kinetics of HDL were studied by endogenous
labeling of HDL apolipoprotein AI (HDL-apo AI) using a primed
infusion of D
3
-leucine. The HDL-apo AI fractional catabolic rate
(FCR) was significantly increased (0.32 ⫾0.07 vs. 0.23 ⫾0.05 pool/
day; P⬍0.01), and HDL composition was changed [HDL cholesterol,
0.77 ⫾0.16 vs. 1.19 ⫾0.37 mmol/L (P⬍0.05); HDL triglycerides,
0.19 ⫾0.12 vs. 0.10 ⫾0.03 mmol/L (P⬍0.05)] in diabetic patients
compared with healthy subjects. HDL-apo AI FCR was correlated to
plasma and HDL triglyceride concentrations (r ⫽0.82; P⬍0.05 and
r⫽0.80; P⬍0.05, respectively) and to homeostasis model assessment
(r ⫽0.78; P⬍0.05). Postheparin plasma LPL activity was decreased
in type II diabetes (6.8 ⫾2.8 vs. 18.1 ⫾5.2
mol/mL postheparin
plasma䡠h; P⬍0.005) compared with that in healthy subjects and was
correlated to the FCR of HDL-apo AI (r ⫽⫺0.63; P⬍0.05). LPL
activity was also correlated with HDL cholesterol (r ⫽0.78; P⬍0.05),
plasma and HDL triglycerides (r ⫽⫺0.87; P⬍0.005 and r ⫽⫺0.83;
P⬍0.05, respectively), and homeostasis model assessment (r ⫽
⫺0.79; P⬍0.05). In addition, the LPL to hepatic lipase ratio was
correlated with the catabolic rate of HDL (r ⫽⫺0.76; P⬍0.06). These
results suggest that a decrease in the LPL to hepatic lipase ratio in
type II diabetes mellitus, mainly related to lowered LPL activity,
could induce an increase in HDL catabolism. These alterations in
HDL kinetics in type II diabetes proceed to some extent from changes
in their composition, probably linked to an increase in triglyceride
transfer from very low density lipoprotein particles, in close relation-
ship with LPL activity and resistance to insulin. (J Clin Endocrinol
Metab 86: 1962–1967, 2001)
THE INVERSE RELATIONSHIP between high density
lipoprotein (HDL) cholesterol level and the incidence
of coronary heart disease (1) underlines the need to under-
stand the processes regulating synthetic and catabolic rates
of HDL. Low levels of HDL cholesterol as well as increased
HDL-apo AI fractional catabolic rate (FCR) are often asso-
ciated with hypertriglyceridemia (2), and changes in lipopro-
tein lipase (LPL) and hepatic lipase (HL) activities may partly
explain this atherogenic potential (3). HDL2 particles are
actually formed from the LPL-catalyzed hydrolysis of very
low density lipoprotein triglycerides (VLDL-TG) as apoli-
poproteins (apo) and surface phospholipids released from
VLDL merge with preexisting HDL3 particles (4, 5). How-
ever, although a direct linkage between LPL activity and
HDL concentration has been observed in healthy subjects
(6–9), in dyslipidemia (10, 11), and in diabetes mellitus (12–
14), intervention of the lipase in the kinetic perturbations of
HDL metabolism remains poorly understood. Zech et al. (15)
as well as Magill et al. (10) reported higher FCRs of apo AI
in three subjects with familial LPL deficiency compared with
those in control subjects. Similar results were reported in
monkeys when anti-LPL antibodies were injected to reduce
LPL activity (16). Indirect evidence also supports the role of
LPL activity in HDL catabolism. For example, in type II
diabetes, in which a decrease in LPL activity related to insulin
resistance is usually reported (13, 14, 17–19), we observed an
increased catabolic rate of HDL-apo AI (20). Such a corre-
lation was also reported in patients with impaired glucose
tolerance (21). In addition, in patients with low HDL cho-
lesterol levels and hypertriglyceridemia, the LPL to HL ratio
was correlated with the clearance rate of HDL (3). This study
therefore aims to test the hypothesis that decreased LPL
activity is related to increased HDL catabolism.
Subjects and Methods
Subjects
Kinetic studies of apolipoprotein AI metabolism were performed in
seven healthy subjects (normal LPL activity group) and seven type II
diabetic patients (low LPL activity group). Some relevant clinical and
physiological characteristics are shown in Tables 1 and 2. None of the
subjects had taken any medication that could affect lipid for at least 2
months before the study. All women were postmenopausal. Diabetic
patients had no proteinuria or hypothyroidism, and were not regular
cigarette smokers or alcohol consumers. They had never been treated
with probucol and were not receiving insulin. The subjects were in-
Received May 4, 2000. Revision received November 2, 2000. Rerevi-
sion received January 26, 2001. Accepted February 5, 2001.
Address all correspondence and requests for reprints to: Prof. M.
Krempf, Clinique d’Endocrinologie, Maladies Me´taboliques et Nutri-
tion, Hoˆtel Dieu, 1 place A. Ricordeau, 44093 Nantes Cedex 01, France.
E-mail: mkrempf@sante.univ-nantes.fr.
* Supported by grants from the LIPHA Pharmacological Group and
the French Society of Dietetic and Nutrition.
0021-972X/01/$03.00/0 Vol. 86, No. 5
The Journal of Clinical Endocrinology & Metabolism Printed in U.S.A.
Copyright © 2001 by The Endocrine Society
1962
structed by a dietician to eat a weight maintenance diet composed of 50%
of the usual daily caloric intake as carbohydrate, 35% as fat, and 15% as
protein for at least 1 week before the study. The experimental protocol
was approved by the ethical committee of Nantes University Hospital,
and informed consent was obtained before the study was started.
Infusion of stable isotope tracer
The kinetic protocol was described in a previous study (20). Briefly,
the endogenous labeling of apo AI was performed by the administration
of l-[5,5,5-
2
H
3
]leucine (99.8 atom %
2
H
3
; Cambridge Isotope Labora-
tories, Andover, MA), which was dissolved in a 0.9% saline solution and
tested for sterility and the absence of pyrogens before the study. All
subjects fasted overnight for 12 h before the study and remained fasting
during the entire protocol. Each subject received an iv priming dose of
10
mol/kg tracer, immediately followed by a constant tracer infusion
(10
mol/kg䡠h) for 14 h. Venous blood samples were withdrawn in
ethylenediamine tetraacetate tubes (Venoject, Paris, France) at baseline,
every 15 min during the first hour, every 30 min during the next 2 h, and
then hourly until the end of the study. Plasma was immediately sepa-
rated by centrifugation for 30 min at 4 C; sodium azide, an inhibitor of
bacterial growth, and Pefabloc SC (Interchim, Montluc¸on, France), a
protease inhibitor, were added to blood samples at final concentrations
of 1.5 mmol/L and 0.5 mmol/L, respectively.
Analytical procedures
Measurement, isolation, and preparation of apo. VLDL (density, ⬍1.006
g/mL) were isolated from 3 mL plasma by sequential ultracentrifugation
using an angle rotor at 40,000 rpm for 24 h at 10 C (Himac CP70, Hitachi,
Hialeah, FL). HDL2 (1.063 ⬍density ⬍1.125 g/mL) and HDL3 (1.125 ⬍
density ⬍1.210 g/mL) were then isolated by a modified method of
density gradient ultracentrifugation (22), using a swinging bucket rotor
at 40,000 rpm for 24 h at 10 C (Centrikon T 2060, Kontron Instruments
Ltd., Zurich, Switzerland). Cholesterol and TG levels in plasma and the
TABLE 1. Clinical and physiological characteristics of study subjects
Subject no. Age (yr) Gender BMI
(kg/m
2
)
FBG
(
mol/mL) Insulinemia HbA
1c
HOMA
1 45 Male 29.0 10.0 7.0 6.7 3.1
2 58 Male 32.0 6.1 4.1 7.6 1.1
3 50 Male 27.4 10.9 6.0 8.1 2.9
4 64 Male 33.1 9.0 16.1 8.8 6.4
5 65 Female 30.1 7.6 14.5 7.8 4.9
6 57 Female 28.4 12.4 6.8 10.2 3.7
7 50 Female 35.7 9.4 16.8 7.2 7.0
8 47 Male 28.9 12.4 13.5 6.7 7.4
Diabetic patients (mean) 55 3/5 30.6 9.7 10.6 7.9 4.6
SD 8 F/M 2.8 2.2 5.1 1.2 2.2
9 24 Male 21.8 5.0 4.9 1.1
10 24 Male 21.8 4.6 7.7 1.6
11 48 Male 31.6 4.4 6.3 4.9 1.2
12 47 Female 29.3 5.0 7.5 1.7
13 47 Male 29.6 5.9 15.5 4.3 3.9
14 40 Male 29.6 5.9 6.7 5.3 1.8
15 51 Male 33.5 6.2 7.6 5.0 2.1
Control subjects (mean) 40 1/6 28.2 5.3 8.0 4.9 1.9
SD 12 F/M 4.6 0.7 3.4 0.4 0.9
FBG, Fasting blood glucose; Hba
1c
. glycosylated hemoglobin.
TABLE 2. Lipase activities, plasma and HDL levels of lipids (millimoles per L), and apo AI (milligrams per dL) in study subjects
Subjects no. LPL activity HL activity LPL/HL Plasma apo AI Plasma CH Plasma TG HDL-CH HDL-TG
1 3.6 20.0 0.18 128 6.72 3.25 0.82 0.16
2 5.2 31.9 0.16 93 7.03 2.83 0.70 0.15
3 10.7 55.6 0.19 125 5.66 3.17 0.82 0.12
4 11.4 49.0 0.23 129 8.22 4.19 0.80 O.10
5 4.6 10.1 0.45 134 6.72 3.58 0.83 0.22
6 6.1 28.4 0.21 149 7.47 4.66 0.79 0.26
7 6.3 23.8 0.27 151 8.29 1.69 1.01 0.09
8 6.1 20.5 0.30 111 5.43 3.81 0.43 O.46
Diabetic patients (mean) 6.8 29.9 0.25 127.5 6.94 3.40 0.77 0.19
SD 2.8 15.3 O.09 19.O 1.05 0.91 0.16 012
9 25.5 42.8 0.60 106 2.92 0.73 1.29 0.08
10 23.3 41.5 0.56 100 4.44 0.82 1.52 0.08
11 20.1 22.3 0.90 110 3.59 1.20 O.73 0.10
12 10.9 19.2 0.57 129 6.02 2.23 1.17 O.16
13 17.6 49.7 0.35 111 5.12 0.89 0.94 0.05
14 15.2 26.3 0.58 211 5.74 0.73 1.79 0.10
15 13.9 25.1 0.55 116 6.05 1.34 0.93 0.10
Control subjects (mean) 18.1 32.4 0.59 126.1 4.84 1.13 1.19 0.10
SD 5.2 11.9 0.16 38.5 1.23 0.54 0.37 0.03
P
a
⬍0.005 NS ⬍0.005 NS ⬍0.05 ⬍0.005 ⬍0.05 ⬍0.05
LPL and HL activities, in minsonoles per mL post heparin plasma/h; CH, cholesterol.
a
Diabetic patients (no. 1–8) vs. control subjects (no. 9 –15).
LPL ACTIVITY AND APO AI KINETICS 1963
HDL fraction were measured using commercially available enzymatic
kits (Roche Molecular Biochemicals, Mannheim, Germany). The apo AI
concentration was measured in plasma by immunonephelometry (Be-
hring, Rueil Malmaison, France). The apo AI pool size (milligrams per
kg) was calculated by multiplying the mean plasma apo AI concentra-
tion by 0.032–0.048 L/kg, assuming a plasma volume of 3.2–4.8% of
body weight according to the age, gender, and body weight of each
subject (23). The plasma apo AI concentration was taken to be the HDL
apo AI concentration, with the assumption that more than 90% of plasma
apo AI resides in the HDL fraction (24).
HDL-apo AI and VLDL-apo B100 were concentrated (25) and isolated
from other apolipoproteins by SDS-PAGE using a 4–5-10% discontin-
uous gradient. Apolipoproteins were identified by comparing migration
distances with those of known molecular weight standards (cross-linked
phosphorylase b markers, Sigma, St. Louis, MO; electrophoresis cali-
bration kit, Pharmacia LKB, Biotechnology, Inc., Piscataway, NJ). Apo
bands were excised from polyacrylamide gels and dried in a vacuum
(RC 10–10 Jouan, Saint Herblain, France). The desiccated gel slices were
hydrolyzed with 1 mL 4 nHCl (Sigma, St. Quentin Fallavier, France) at
110 C for 24 h. Hydrolysates were then evaporated to dryness, and the
amino acids were purified by cation exchange chromatography using
Temex 50W-X8 resin (Bio-Rad Laboratories, Inc., Richmond, CA).
Plasma amino acids were esterified with propanol/acetyl chloride and
further derivatized using heptafluorobutyric anhydride (Fluka Chemie
AG, Buchs, Switzerland) before analysis.
Determination of tracer to tracee ratios. Chromatographic separations were
carried out on a 30-m ⫻2.52-mm (id) DB-5 capillary column (J&W
Scientific, Rancho Cordova, CA). The column temperature program was
as follows: initial temperature was held at 80 C, then increased at 10
C/min to a final temperature of 180 C. Electron impact gas chroma-
tography-mass spectrometry was performed on a 5891 A gas chromato-
graph connected to a 5971 A quadrupole mass spectrometer (Hewlett-
Packard Co., Palo Alto, CA). The isotopic ratio was determined by
selected ion monitoring at m/z 282 and 285. Calculations of apo AI
kinetic parameters were based on the tracer to tracee mass ratio (26).
Determination of lipase activities. On the day of the kinetic study, pre- and
postheparin blood samples were drawn into ice-cold ethylenediamine
tetraacetate tubes before and 10 min after iv injection of 100 IU hepa-
rin/kg BW. This bolus of heparin was injected at the end of the tracer
infusion to avoid any effect on VLDL metabolism. Plasma was separated
at 4 C and was stored frozen until assayed. Lipase activities were mea-
sured following the method described by Iverius and Brunzell (27). The
assay was performed using glycerol tri-[1-
14
C]oleate (NEN Life Science
Products, Boston, MA) emulsified with Triton X-100 as substrate. LPL
HL activities were, respectively, inhibited by high salt concentration and
SDS, as previously described (28). Lipases activities were expressed as
micromoles of free fatty acids hydrolyzed by 1 mL postheparin plasma
during1hofincubation at 37 C.
Insulin sensitivity estimate. The insulin resistance level was estimated
with the homeostasis model assessment (HOMA) (29) using the follow-
ing formula: HOMA ⫽[insulin]/(22.5 e
⫺ln [glucose]
).
The plasma insulin concentration (microinternational units per mL)
was measured by radioimmunometric assay (Sanofi Pharmaceuticals,
Inc., Marnes-La-Coquette, France). The fasting blood glucose concen-
tration (micromoles per mL) was evaluated using a glucose oxidase
enzymatic assay (BioMe´rieux, Marcy-l’Etoile, France).
Modeling
For HDL modeling, we used a one-compartment model, as previ-
ously described (20). Kinetic analysis of the tracer to tracee ratio was
achieved by computer software for simulation, analysis, and modeling
(SAAM II version 1.0.1, Resource Facility for Kinetic Analysis, SAAM
Institute, Seattle, WA). VLDL-apo B100 and HDL-apo AI data were
kinetically analyzed using a monoexponential function (26): A(t) ⫽
Ap[1 ⫺exp(⫺k(t ⫺d))], where A(t) is the tracer to tracee ratio at time
t, Ap is the tracer to tracee ratio at the plateau of the VLDL apo B100
curve, d is the delay between the beginning of the experiment and the
appearance of tracer in the apolipoprotein, and k is the fractional pro-
duction rate (FPR) of the apolipoprotein. For the estimation of apo AI
synthesis, we used the plateau of VLDL-apo B100 tracer to tracee ratio
as the precursor pool enrichment. It was assumed that this plateau value,
obtained using a monoexponential function, corresponded to the tracer
to tracee ratio of the leucine precursor pool. This estimation is based
upon the assumption that apo B100 and the majority of apo AI are
synthesized by the liver (30). We estimated the FPR, i.e. the proportion
of apo AI entering the pool per unit time (days), and the absolute
production rate (APR), i.e. the amount of apolipoprotein AI entering the
pool per unit time (milligrams per kg/day). APR was the product of FPR
multiplied by the apo AI mass in the HDL fraction. The apo AI pool was
considered to be constant, as no significant variation was observed
between measurements made at three different infusion times (data not
shown). Under these steady state conditions, FPR equals the FCR.
Statistical analysis
Data are reported as the mean ⫾sd unless otherwise specified.
Statistical analysis was performed using Instat Software package
(GraphPad Software, Inc., San Diego, CA). The Mann-Whitney U test
was used to compare clinical and kinetic data between type II diabetes
and controls. Linear regression and correlation analyses were performed
with a linear correlation analysis, using the StatView 4.5 software pack-
age (Abacus Concepts, Berkeley, CA). A two-tailed Plevel of 0.05 was
accepted as statistically significant.
Results
Kinetic data
Enrichment in plasma free leucine reached a plateau after
30 min of infusion and remained stable to the end of the study
(data not shown). The mean tracer to tracee ratio curves in
HDL are shown in Fig. 1. VLDL-apo B100 isotopic enrich-
ments reached steady state conditions within the infusion
period regardless of the subject investigated (data not
shown). Kinetic parameters of apo AI are shown in Table 3.
Patients with type II diabetes mellitus showed increased
HDL-apo AI FCR (0.32 ⫾0.07 vs. 0.23 ⫾0.05 pool/day; P⬍
0.01), whereas HDL-apo AI APR was not altered (15.8 ⫾3.3
vs. 12.3 ⫾5.5 mg/kg䡠day; P⫽NS). The FCR of HDL-apo AI
was correlated to HOMA (r ⫽0.78; P⬍0.05; Table 4).
Apo and lipid concentrations
Individual data for plasma and HDL composition are pre-
sented in Table 2. Patients with type II diabetes mellitus
FIG. 1. Mean experimental values (symbols) of the tracer to tracee
ratio for apo AI-HDL in controls (E) and patients with type II diabetes
mellitus (Œ). Fits (lines) were calculated using monocompartmental
analysis during a primed constant infusion of [
2
H
3
]leucine. Data are
shown as the mean ⫾SEM.
1964 FRE
´NAIS ET AL. JCE&M•2001
Vol. 86 •No. 5
showed characteristically higher plasma lipids levels com-
pared with controls [total cholesterol, 6.94 ⫾1.05 vs. 4.84 ⫾
1.23 mmol/L (P⬍0.05); TG, 3.40 ⫾0.91 vs. 1.13 ⫾0.54
mmol/L (P⬍0.005)]. HDL composition was also changed
[HDL cholesterol, 0.77 ⫾0.16 vs. 1.19 ⫾0.37 mmol/L (P⬍
0.05); HDL-TG, 0.19 ⫾0.12 vs. 0.10 ⫾0.03 mmol/L (P⬍
0.05)]. Plasma apo AI pool size was not significantly lower in
diabetic patients (49.3 ⫾6.0 vs. 53.6 ⫾16.6 mg/kg; P⫽NS).
The plasma and HDL-TG levels were correlated with the
catabolic rate of HDL-apo AI (r ⫽0.82; P⬍0.05 and r ⫽0.80;
P⬍0.05; Table 4).
Post-HL activities
LPL activity (Table 2) was decreased in diabetic patients
(6.8 ⫾2.8 vs. 18.1 ⫾5.2
mol/mL postheparin plasma䡠dL;
P⬍0.005) and was correlated with the FCR of apo AI (r ⫽
⫺0.63; P⬍0.05; Fig. 2). LPL activity was also correlated with
HDL-cholesterol (r ⫽0.78; P⬍0.05) and plasma and
HDL-TG levels (r ⫽⫺0.87; P⬍0.005 and r ⫽⫺0.83; P⬍0.05,
respectively; Table 4). Correlations were observed between
LPL activity and fasting blood glucose (r ⫽⫺0.87; P⬍0.005)
or HOMA (r ⫽⫺0.79; P⬍0.05; Table 4).
HL activity was similar in diabetic patients and controls
(29.9 ⫾15.3 vs. 32.4 ⫾11.9
mol/mL postheparin plasma䡠dL;
P⫽NS).
The LPL to HL ratio was decreased in type II diabetes
mellitus (0.25 ⫾0.09 vs. 0.59 ⫾0.16; P⬍0.005), and correlated
with the FCR of HDL-apo AI (r ⫽⫺0.76; P⬍0.06) and the
plasma TG level (r ⫽⫺0.92; P⬍0.001).
Discussion
This study was designed to further delineate in vivo the
relationship between LPL activity and the kinetic aspects of
HDL-apo AI metabolism in control subjects and type II di-
abetic patients. We found a negative correlation between LPL
activity and the clearance rate of HDL. As expected, post-
heparin LPL activities were lower in type II diabetic patients
compared with controls and were close to those obtained in
diabetic patients with moderate hypertriglyceridemia (14,
17). In this group the FCR of HDL-apo AI was significantly
increased. Both plasma levels and HDL composition were
altered in type II diabetes mellitus, with an increased
HDL-TG level and a decrease in the HDL cholesterol con-
centration. The HDL-TG level was correlated with the FCR
of HDL-apo AI. In addition, LPL activity was inversely cor-
related with plasma and HDL-TG levels and with HOMA,
but was positively related to the HDL cholesterol concen-
tration, as previously reported (6–13).
Study subjects were recruited according to their potential
level of LPL activity; type II diabetes mellitus was theoret-
ically considered a model of low LPL-mediated hydrolysis of
VLDL-TG, compared with that in control subjects (13, 14, 17,
18). None of them had been included in our previous study
(20). The heparin assay we performed showed, as expected,
that diabetic patients actually presented with low LPL ac-
tivity, whereas controls had normal LPL levels. In addition,
there was an overlap in the activities observed in the two
groups. Therefore, the correlations we found were not re-
lated to two different sets of data, but, rather, corresponded
FIG. 2. Relationship between HDL-apo AI FCR (pool per day) and
LPL activity (micromoles per mL postheparin plasma/h) in type II
diabetic patients and controls.
TABLE 3. Apo Al pool size and HDL kinetic parameters in study
subjects
Subjects no. FCR
HDL-apo Al
APR
HDL-apo
Apo Al
pool size
1 0.31 16.4 52.5
2 0.31 11.5 37.7
3 0.29 15.9 55.0
4 0.44 21.3 48.4
5 0.27 13.5 50.3
6 0.30 17.0 56.6
7 0.25 12.2 48.3
8 0.40 18.2 45.5
Diabetic patients (mean) 0.32 15.8 49.3
SD 0.07 3.3 6.0
9 0.16 7.8 50.4
10 0.17 8.4 48.5
11 0.23 10.0 44.6
12 0.28 14.5 51.0
13 0.28 12.9 46.1
14 0.26 23.4 90.7
15 0.21 9.2 44.1
Control subjects (mean) 0.23 12.3 53.6
SD 0.05 5.5 16.6
P
a
⬍0.01 NS NS
FCR, fractional catabolic rate (pool/day); APR, absolute production
rate (milligrams per (no. kg/day); apo AI pool size in (milligrams
per kg).
aDiabetic patients (no. 1–8) vs. control subjects (no. 9 –15).
TABLE 4. Correlation analysis in study subjects
FCR
HDL-apo Al
LPL
activity
Plasma TG 0.82
a
⫺0.87
b
HDL-TG 0.80
a
⫺0.83
a
HOMA 0.78
a
⫺0.79
b
LPL/HL ⫺0.76
c
HDL-CH 0.78
a
Fasting blood glucose ⫺0.87
b
FCR, Fractional catabolic rate (pool/day).
a
P⬍0.05.
b
P⬍0.005.
c
P⬍0.06.
LPL ACTIVITY AND APO AI KINETICS 1965
to a homogenous plot of points. Although gender and age do
not appear to be key parameters in the control of HDL ca-
tabolism (24, 31, 32), the lack of absolute matching of the two
study groups according to these parameters could constitute
a limitation of the study. We performed an endogenous
labeling of apo AI by infusion of leucine labeled with a stable
isotope because this procedure avoids any change in lipopro-
tein kinetics related to potential alterations of the protein’s
characteristics due to the exogenous labeling (33). Our ex-
perimental enrichment data could not be adjusted on a two-
pool model, as was sometimes previously done, because our
study was designed with a constant infusion of tracer and
our period of sampling did not allow characterization of
tracer exchanges with a second pool. Therefore, as in other
apo AI kinetic studies (21, 34), we applied a single HDL
compartment to our modeling design. As an estimate of apo
AI leucine precursor pool enrichment, we considered VLDL-
apo B100 enrichment at the plateau, which was reached at the
end of the infusion period. This assumed that apo AI was
mainly synthesized by the liver (30), which is likely to occur
in the fasting state.
Our data, although partly speculative, contribute to a
global overview of the metabolic processes that link HDL to
TG-rich lipoproteins. In type II diabetes, the reduced LPL
activity previously reported (13, 14, 17–19) induces a defect
in the clearance of TG-rich lipoparticles from the circulation
(10, 35–37). This combined with the typical overproduction
of VLDL consequently lead to an increase in VLDL-TG. This
may enhance cholesterol ester transfer protein (CETP)-me-
diated TG-cholesteryl ester exchanges, leading to alterations
in HDL composition. The negative correlation between LPL
activity and HDL concentration corroborates this hypothesis
(6–13). This is also in keeping with an in vivo study in an
animal model (38). In transgenic mice expressing the CETP
transgene, LPL activity was correlated with the HDL cho-
lesterol level, but not in the absence of CETP. However,
whereas LPL activity appears to play a strong role in HDL
composition, its effect on HDL-apo AI metabolism in hu-
mans has been poorly studied. A 28% increase in the HDL
clearance rate was also observed by Magill et al. in one subject
with LPL deficiency after exogenous labeling of [
125
I]HDL
(10). Furthermore, Goldberg and co-workers, by infusing
specific monoclonal antibodies into female cynomolgus
monkeys to inhibit LPL, observed that the HDL-apo AI cat-
abolic rate in LPL-inhibited animals was more than double
that in control rabbits (16). Thus, they suggested that the
variations in apo AI level and clearance rate might be a
consequence of differences in LPL-mediated lipolysis of TG-
rich lipoproteins. We now report that LPL activity is corre-
lated to HDL composition and catabolism in humans, and
therefore we suggest that impaired lipase activity on VLDL
could induce an increased CETP-mediated efflux of TG on
HDL, leading to alterations in both their composition and
their clearance rate. This hypothesis is in agreement with the
positive correlation between plasma TG levels and HDL-apo
AI FCR that we previously observed in type II diabetes (20).
In addition, as previously reported (17, 36), HL activities
were similar in the two study groups. As HL activity is
increased and HDL2 cholesterol seems to be specifically re-
duced in obesity (39), we would have probably observed
lowered HL activities among a control group composed of lean
subjects. Furthermore, the LPL to HL ratio was decreased in
diabetic patients and correlated to the FCR of HDL-apo AI, as
previously reported in patients with low HDL cholesterol levels
(3). HL and LPL have opposing effects on HDL composition;
LPL activity catalyzes the degradation of TG-rich lipoproteins
and induces transfer of lipid surface components to HDL,
whereas HL catabolizes HDL phospholipids. Thus, a low LPL
to HL ratio should promote a depletion of HDL surface com-
ponents and an enrichment of these particles in TG, which is in
agreement with their enhanced clearance (3).
As previously reported (40– 43), we found similar plasma
apo AI levels in diabetic patients and controls, contrasting
with other studies (20, 44). The clinical characteristics of
healthy subjects, matched for mean age and body mass index
with diabetic patients in the current study, could explain this
discrepancy. Furthermore, in our previous study the plasma
apo AI level was decreased in diabetes mellitus as the result
of an increased clearance rate and unchanged production
rate of HDL (20). In the current study the slight increase in
HDL APR was sufficient to restore a normal plasma apo AI
level. The heterogeneity of apo AI production rates related
to genetic or environmental factors could therefore be a key
factor in the control of the plasma HDL concentration in the
case of enhanced clearance, and this aspect needs to be clar-
ified in further studies.
The insulin resistance of patients with type II diabetes may
additionally contribute to the down-regulation of LPL ac-
tivity (45, 46) and the increase in HDL-apo AI FCR (32). The
correlations between HOMA and LPL activity or HDL clear-
ance rate support this hypothesis. Therefore, LPL activity
may be a target of hypolipidemic treatment to restore a
normal HDL cholesterol level in type II diabetic patients with
low HDL. Studies have actually shown that fibrates en-
hanced the expression of LPL by activating transcription
factors of the peroxisome proliferator-activated receptors
(47). Other treatments, such as weight loss or biguanides,
may directly act upon insulin resistance to recover suitable
LPL activities.
In conclusion, these results support the hypothesis that
reduced LPL activity, related to resistance to insulin, may
play a major role in disorders of HDL metabolism in humans.
In fact, impaired lipase activity on VLDL could induce an
increased efflux of TG on HDL, leading to alterations in both
their composition and their clearance rate. This study, there-
fore, provides further information about the coordinate reg-
ulation of HDL and TG metabolism.
Acknowledgments
We thank Ms. F. Nazih-Sanderson, C. Le Valegant, and P.
Mauge`re for excellent technical assistance, and D. Darmaun
for review and advice.
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LPL ACTIVITY AND APO AI KINETICS 1967
