Postheparin Plasma Triglyceride Lipases
Relationships with Very Low Density Lipoprotein
Triglyceride and High Density Lipoprotein2 Cholesterol
Deborah Applebaum-Bowden, Steven M. Haffner, Patricia W. Wahl,
J. Joanne Hoover, G. Russell Warnick, John J. Albers, and William R. Hazzard
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
Hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) probably have major
roles in the removal of triglyceride from triglyceride-rich lipoproteins and in the formation of high density lipoprotein (HDL). However, no population-based study of their
activity and relationship to lipoprotein lipid levels has been reported. To determine
these relationships, we recalled 33 men and 17 women of a randomly selected sample
of the Lipid Research Clinics Pacific Northwest Bell Telephone Company Health Survey. The subjects were 53 ± 7 years old (mean ± SD) with total triglyceride levels of
120 ± 57 mg/dl and total cholesterol levels of 224 ± 35 mg/dl. Postheparin plasma LPL
activity (127 ± 61 nmol/min/ml) was not significantly correlated with either age, sex, or
adiposity. In contrast, HTGL activity was significantly higher in men (235 ± 84 nmol/
min/ml) than women (170 ± 91 nmol/min/ml, p < 0.02), and was correlated with age in
men and with adiposity in women. In both men and women, HTGL activity was related
positively with VLDL triglyceride and inversely with HDL2 cholesterol. When the association between HTGL activity and VLDL triglyceride was examined with values from
men and women pooled, the relationship was not weakened after adjustment for the
linear effect of sex, adiposity, LPL, or HDL2 cholesterol.
(Arteriosclerosis 5:273-282, May/June 1985)
posal, patients lacking HTGL activity have an abnormal VLDL, R-VLDL, pre-B-VLDL in their plasma and
elevated TG levels in both low density lipoprotein
(LDL) and high density lipoprotein (HDL).8 LPL originates from extrahepatic tissues and is thought to
function mainly in the removal of TG from chylomicrons.9 Patients lacking LPL have chylomicronemia,
but do not have (3-VLDL or elevated levels of TG in
either LDL or HDL.1011 Both HTGL and LPL have
reportedly correlated with HDL cholesterol levels12'13
and specifically with the HDL2 subtraction.14'15 They
differ, however, in that HTGL correlates negatively,
while LPL correlates positively. An inverse relationship exists between HDL and the risk of ischemic
heart disease in many studies.1617 Because low
HDL, cholesterol is associated with atherosclerosis
(determined by coronary angiography)18 and HDL2
cholesterol is decreased in survivors of myocardial
infarction,19 it is important to understand how HTGL
and LPL relate to HDL metabolism.
We examined a random sample of men and women from the Pacific Northwest Bell Telephone Company Health Survey20'21 for the behavioral correlates
of the HDL subtractions.22 We also obtained postheparin plasma from these subjects, and in the current report we describe the associations between
postheparin plasma HTGL and LPL and lipoprotein
lipids.
eparin releases into plasma two enzymes, heH
patic triglyceride lipase (HTGL) and lipoprotein
lipase (LPL). The two lipases can be distinguished in
postheparin plasma by the requirement of LPL for
apolipoprotein (apo) C-ll,1 by the inhibition of LPL by
NaCI and protamine sulfate,2 by the different elution
characteristics of these lipases from heparin-Sepharose,3 and by selective inhibition by specific antibodies.45 HTGL originates from the liver and is
thought to function in removing triglyceride (TG) from
very low density lipoprotein (VLDL) and other particles of decreasing size. 67 Consistent with this pro-
From the Northwest Lipid Research Clinic and the Departments
of Medicine, Biostatistics, and Epidemiology, University of Washington, Seattle, Washington.
This work was supported by NIH Grants HL 22381, HL 23474,
HL 30086, HL 30193, NIH Contract HV-12157-L, and a Grant-inAid from the American Heart Association. Dr. Haffner was supported by Training Grant HL-07028. Dr. Albers is an Established
Investigator of the American Heart Association.
Steven M. Haffner is currently at the University of Texas at San
Antonio, Texas. Deborah Applebaum-Bowden and William R.
Hazzard are at Johns Hopkins University School of Medicine,
Baltimore, Maryland.
Address for reprints: Dr. Deborah Applebaum-Bowden, The
Johns Hopkins University School of Medicine, 720 Rutland Avenue, 709 Traylor Building, Baltimore, Maryland 21205.
Received May 16, 1984; revision accepted January 29, 1985.
273
ARTERIOSCLEROSIS
274
Methods
Population Sample
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All subjects were from a 2% random sample of
5000 participants in the Pacific Northwest Bell Telephone Company Health Survey,2021 which was conducted by the Northwest Lipid Research Clinic under
the auspices of the Population Studies component of
the Lipid Research Clinics Program.23 Only individuals who 1) were over age 30 at the time of the initial
screening, 2) were still living in the Greater Seattle
area, and 3) had agreed to participate in a yearly
follow-up were invited to participate in the current
study. Of the original sample, 20 were under the age
of 30 at the original screening (between 1972 and
1974) four had moved out of the area, seven refused
further contact, and two had died; this left 67 possible
participants. There were 50 subjects (aged 38 to 65)
who gave informed consent for the current study; this
was a response rate of 75%. This visit (the fourth for
these subjects) took place in May and June, 1981.
The project was approved by the University of Washington Human Subjects Review Committee.
Study Protocol
The clinic visit included administration of a questionnaire covering demographic information, menopausal status, medication use, smoking (number of
cigarettes presently smoked) and drinking (frequency of alcohol use). Subjects were instructed to fast
for 12 to 14 hours before coming to the clinic, where
they were weighed without shoes, jackets, or coats.
Their height was measured without shoes. Body
mass index (BMI) was determined by dividing weight
(kg) by height squared (m2).
VOL
5, No 3,
MAY/JUNE
1985
was performed with dextran sulfate (15,000 daltons,
Sochibo SA, Boulogne, France) and yielded HDL3.
The cholesterol in these supernatant fractions was
measured by standard Lipid Research Clinics methodology.24 After correction for dilution, the HDL, cholesterol was calculated as the difference between
HDL cholesterol and HDL3 cholesterol. Lipoproteins
and subfractions were analyzed within 72 hours.
HTGL and LPL were measured within 1 month
after the clinic visit by using a modification of the
method of Huttunen et al.26 Both substrates contained, per milliliter, glycerol trioleate (2.9 /i.mol),
glyceryl tri [1 - l4 C]oleate(0.13/xCi), gumarabic(10
mg), and bovine serum albumin (20 mg). The substrate for total postheparin plasma TG lipase activity
contained serum (0.1 ml) and a low salt concentration (0.2 M NaCI, 0.5 ml), whereas that for HTGL
contained 0.9% NaCI (0.1 ml) in place of the serum
and a high salt concentration (2 M NaCI, 0.5 ml). The
assays were run at 28° C for 90 minutes. The released 14C fatty acids were extracted as described
by Belfrage and Vaughan.27 LPL activity was calculated by subtracting the HTGL activity from the total
activity.
HTGL measured by this procedure was found to
correlate closely with the level measured by heparinSepharose chromatography (r3 = 0.980, p < 0.001)
by the method of Applebaum et al.,28 which used
Triton X-100 instead of gum arabic.12 The level of
LPL measured by this procedure did not correlate
with the level obtained with heparin-Sepharose chromatography. Since this lack of agreement may have
been due to a low recovery of LPL from the chromatography step, the modified assay of Huttunen et al.26
was chosen for the current study.
Statistical Methods
Laboratory Methods
Antecubital venous blood was drawn through a 21 gauge butterfly infusion kit and mixed immediately
with dry disodium EDTA (1.5 mg/ml blood) in a vacutainer tube (Becton Dickinson and Company, Rutherford, New Jersey). Heparin (10 units/kg, Upjohn
Company, Kalamazoo, Michigan) was injected, followed by 10 ml of 0.9% sterile saline to flush the line.
Blood was drawn 10 minutes after the heparin injection, was mixed with EDTA in a similar vacutainer
tube, and was placed in ice. Blood was promptly
centrifuged at 1500 g for 30 minutes at 4° C, and the
resulting plasma was collected. The postheparin
plasma was frozen and stored at - 20° C for assay.
The lipoproteins were isolated from the preheparin
plasma by ultracentrifugation and heparin-manganese precipitation by the standard laboratory protocol of the Lipid Research Clinics.24 The preheparin
plasma was also used to measure the cholesterol in
the HDL subclasses with the two-step precipitation
method of Gidez et al.25 The first precipitation was
performed with heparin (40,000 U/ml, Riker Laboratories, Northridge, California) and MnCI2, and the
procedure yielded HDL. The second precipitation
Statistical analyses are based on the entire sample of men and women after ensuring that the inclusion of either the one nonwhite woman or the nine
subjects on antihypertensive drugs did not alter the
results. Descriptive statistics including the mean,
standard deviation (SD), and median (50th percentile) are given separately for men and women. The
Wilcoxon rank sum test statistic29 was used to determine significant differences in HTGL and LPL between men and women. Relationships between two
variables were estimated by using Spearman's rank
correlation coefficient (rj with a corresponding test
of significance.30 First-order Pearson correlation coefficients were used to measure the association between two variables after the linear effects of a third
variable had been removed. To obtain normal distributions, we performed logarithmic transformations
on VLDL-TG and HDL, cholesterol. In addition, we
performed stepwise multiple linear regression to adjust for effects which proved to be significant in univariate analysis. Caution should be used in the interpretation of probability values (p) due to the small
sample and because multiple testing increases the
risk of obtaining a significant result purely by chance.
HEPATIC TRIGLYCERIDE LIPASE AND LIPIDS
Applebaum-Bowden et al.
275
Table 1. Lipid and Lipoprotein Levels in the Study Population
Women (n = 17)
Men (n = 33)
Mean
Measurement
SD
Median
Mean
SD
Median
52.5
7.4
52.2
54.0
7.4
54.9
26.9
2.9
27.5
24.8
3.9
24.1*
Triglyceride (mg/dl)
Total
VLDL
LDL
HDL
129.8
92.2
27.7
9.9
62.9
60.5
7.9
3.4
116.0
78.0
25.8
9.6
100.9
61.0
27.7
12.2
37.2
37.9
7.3
3.3
93.0
52.3*
27.8
11.3*
Cholesterol (mg/dl)
Total
VLDL
LDL
HDL
HDL2
HDL3
220.3
21.5
151.2
47.6
17.6
29.8
35.4
12.6
32.7
13.9
10.8
5.8
216.0
18.3
149.8
45.0
14.8
28.0
231.9
11.9
163.4
56.6
23.8
32.4
34.0
8.0
33.2
11.2
8.4
7.6
Age (yrs)
2
Body mass index (kg/m )
243.0
10.0t
171.0
57.0f
22.0t
31.5
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
*p < 0.05 for men vs women as determined by the Wilcoxon rank sum statistical test,
t p < 0.01 for men vs women as determined by the Wilcoxon rank sum statistical test.
Results
Characteristics of the Study Subjects
The 50 subjects who participated in this study
ranged in age from 38 to 65 years, with the men (n =
33) having a median age of 52 years and the women
(n = 17) having a median age of 55 years (Table 1).
The men had a higher body mass index (BMI) (median = 27.5) than the women (median = 24.1, p =
0.02). Of the men, 33% were cigarette smokers and
76% consumed more than one alcoholic drink per
week. Of the women, 18% smoked cigarettes and
35% consumed more than one alcoholic beverage
per week. The mean and median plasma lipid and
lipoprotein levels for the population are presented in
Table 1. The men had higher levels of VLDL cholesterol (p < 0.01), and VLDL-TG (p < 0.05) but lower
levels of HDL cholesterol (p < 0.01), HDL-TG (p <
0.05), and HDL, cholesterol (p < 0.01) than the
women. The levels of LDL cholesterol and LDL-TG
were similar in both men and women. We have recently reported a detailed analysis of the effects of
smoking, alcohol, and adiposity on the HDL subtractions, apolipoproteins, and lecithin cholesterol acyltransferase.22
Table 2.
HTGL activity was normally distributed in men and
ranged from 89 to 447 nmol/min/ml, with a mean of
235 ± 84 nmol/min/ml (median, 237) (Figure 1).
HTGL activity levels were not normally distributed in
women, and ranged from 56 to 254 nmol/min/ml plus
one value at 424 nmol/min/ml with a median value of
166, which was significantly lower than in men (p =
0.02). Of the 12 postmenopausal women, the four
oldest were taking replacement hormones. The
woman taking depotestadiol, an androgenic sex
steroid, had the highest HTGL activity found in a
woman (Table 2). The three women taking postmenopausal estrogens (mainly natural conjugated
estrogens, Prematin) had a mean HTGL activity of
169 ± 94 nmol/min/ml (median, 185). HTGL activity
for the 13 women not taking sex steroids was 150 ±
59 nmol/min/ml (range, 56 to 252 nmol/min/ml; median, 154), and when these women were categorized
as to pre- and postmenopausal status, the five premenopausal females were found to have HTGL activity levels similar to those of the eight postmenopausal women.
Effect of Gender and Hormone Use on Triglyceride LIpases
Hepatic triglyceride lipase
(nmol/min/ml)
Gender and medication
Lipoprotein lipase
(nmol/min/ml)
Mean
so
Median
Mean
SD
Median
Men (n = 33)
235
84
237
117
61
117
Women (n = 17)
No sex steroids (n = 13)
On estrogens (n = 3)
On depotestadiol (n =: D
170
150
169
424
91
59
94
166*
154
185
149
160
132
52
57
58
2
145
157
130
*p < 0.02, men vs women.
276
ARTERIOSCLEROSIS
VOL
5, No 3,
MAY/JUNE
1985
HTGL
20
LPL
Men
235 ± 84 nmol/min/ml
10
|
20
Men
117 ± 6 1 nmol/min/ml
10
Women
170 ± 91 nmol/min/ml
p < 0.02
20
DC
Woman
149 ± 57 nmol/min/ml
0.05<p<0.10
20
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10
10
"0
200
400
0
Activjty, nmol/min/ml
200
400
Figure 1. Distribution of hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activity in 33 men and 17 women.
The p values are for the Wilcoxon test.
Table 3.
Spearman Rank Correlation Coefficients for Men (n = 33) and Women (n = 17)
Gender
VLDL-TG
LDL-TG
HDL-TG
VLDL-C
LDL-C
VLDL-TG
M
W
—
0.22
0.08
0.15
-0.11
0.76*
0.73*
0.00
0.43$
LDL-TG
M
W
0.08
-0.32
0.34$
0.16
0.34$
0.42$
HDL-TG
M
W
VLDL-C
M
W
LDL-C
M
W
HDL-C
M
W
HDL2-C
M
W
HDL3-C
M
W
HTGL
M
W
LPL
M
W
Age
M
W
BMI
M
W
0.30$
0.06
—
-0.22
0.01
0.09
0.27
—
VLDL-TG = very low density lipoprotein triglyceride; LDL-TG = low density lipoprotein triglyceride; HDL-TG = high
density lipoprotein triglyceride; HTGL = hepatic triglyceride lipase; LPL = lipoprotein lipase; BMI = body mass index; C =
cholesterol. * p < < 0.001.
t p ^ < 0.01. $p £ < 0.05.
HEPATIC TRIGLYCERIDE LIPASE AND LIPIDS
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LPL activity was normally distributed in both men
and women (Figure 1). LPL activity appeared to be
lower in men (117 + 61 nmol/min/ml) than in women
(149 ± 57 nmol/min/ml), although the difference was
not statistically significant. The woman taking depotestadiol had the lowest level of LPL activity (52
nmol/min/ml) (Table 2). The LPL activity in women
taking estrogens was 132 ± 2 nmol/min/ml. The LPL
activity in women not taking sex steroids was 160 ±
58 nmol/min/ml. Premenopausal women had LPL
levels similar to those of postmenopausal women.
Spearman rank correlations between age, BMI,
lipoprotein lipid levels, and the activities of the TG
lipases are shown in Table 3. HTGL activity was
negatively correlated with age in men (rs = -0.36, p
= 0.02) and positively, but not quite significantly,
correlated with age in women (rs = 0.32, p = 0.106).
Thus, the difference between men and women decreased with age. Although HTGL activity was not
significantly correlated with BMI in men, it was highly
correlated with BMI in women (rs = 0.67, p = 0.001)
(Figure 2), and this relationship still existed when the
women taking sex steroids were removed from the
analysis (rs = 0.60, n = 13, p = 0.014). In contrast
to the relationships seen with HTGL activity, LPL
activity was not significantly correlated with age or
Table 3.
Applebaum-Bowden et al.
277
BMI in men or women. HTGL appeared to be negatively correlated with LPL in men and women, but the
relationship was not statistically significant (p >
0.05). Neither HTGL nor LPL was related to cigarette
smoking or alcohol consumption in either men or
women.
HTGL activity was significantly positively correlated with VLDL-TG in men (rs = 0.38, p = 0.014) and
women (rs = 0.74, p = 0.001) (Figure 3). HTGL was
negatively correlated with HDLj cholesterol in men
(r3 = -0.35, p = 0.022) and in women (r3 = -0.50,
p = 0.02) (Figure 4). In men, HTGL activity also was
correlated negatively with LDL-TG (rs = - 0.31, p =
0.04) and HDL-TG (rs = -0.39, p = 0.01). In women, the correlation coefficient for the relationship between HTGL and HDL-TG was similar to that in men,
but was not statistically significant, perhaps because
of the smaller number of women in our sample.
When the women taking sex steroids were removed
from the analysis, HTGL was still significantly positively correlated with VLDL-TG (rs = 0.66, n = 13,
p = 0.007), but the relationship with HDL, cholesterol was not significant (rs = -0.28, n = 13, p =
0.18), possibly due to truncating the range of HDL,
cholesterol. LPL activity was not highly related to
lipoprotein lipid levels, but was negatively related
(Continued)
HDL-C
HDL2-C
HDL3-C
HTGL
LPL
Age
BMI
0.58f
-0.25
-0.59t
-0.37*
-0.78*
-0.14
-0.06
0.38*
0.74*
-0.29
-0.35
-0.05
0.33
0.01
0.01
-0.13
-0.06
0.19
-0.01
-0.31*
0.11
-0.22
0.37
0.26
0.01
-0.16
0.03
-0.11
0.10
-0.15
0.04
0.06
-0.01
-0.39*
-0.33
-0.08
-0.07
-0.09
-0.33
0.23
0.13
-0.42f
-0.45f
-0.44t
-0.52:):
-0.21
-0.12
0.19
0.37
-0.33*
-0.03
0.00
0.13
0.20
0.43*
-0.00
-0.20
-0.31*
-0.37
0.43f
-0.04
0.29*
0.41*
-0.16
0.15
0.05
0.33
-0.07
0.45*
0.76*
0.76*
0.58*
0.69*
-0.21
-0.21
0.00
0.09
0.02
-0.04
-0.37*
-0.17
0.07*
0.12
-0.35*
-0.50*
0.09
0.33
0.14
-0.31
-0.33*
-0.37
—
-0.10
0.22
0.03
-0.28
-0.10
0.25
-0.29
0.05
-0.28
-0.38
-0.36*
0.32
0.21
0.67*
0.17
-0.32
0.04
0.03
—
—
—
—
0.38*
-0.22
-0.04
278
ARTERIOSCLEROSIS
VOL
5, No 3,
1985
MAY/JUNE
HTGL
LPL
A
A
A
30
A A
A A
A*
A
A
^
A
A
A
30
L
A
^
A
A
A
A
A
20
Men
r
f » 0.209
p = 0.121
10
^A
*
. A
&
^
^AA A
'
.
A A
A
A
A
A
A
A
A
20
*
a
A
Men
r, •= 0.041
NS
10
n
m
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
30
O
30
•
O
0
O
O
?>
o
8
o
0
O
O
O
o
20
20
o
10
10
Women
r, » 0.674
p = 0.001
200
400
Women
r, - -0.028
NS
400
200
Activity, nmol/min/ml
Figure 2. Relationships between hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activity and body mass
index (BMI) in the men (A) and women (o). Significance was determined with the Spearman rank correlation coefficient (rs).
with VLDL cholesterol in men (rs = - 0.33, p = 0.03)
and VLDL-TG in men (r, = -0.29, p = 0.052) and
women (rs = - 0 . 3 6 , p = 0.08) (Figure 3).
Partial correlation coefficients between the HTGL
and VLDL-TG and HDL, cholesterol were obtained.
In this analysis, the gender-specific data were
pooled (n = 50) and the logarithms of VLDL-TG and
HDL, cholesterol were used. HTGL activity was normally distributed when the values from men and
women were pooled. The strong relation between
HTGL activity and log VLDL-TG (simple r = 0.508)
was not considerably weakened (r = 0.385 to 0.465)
after adjustment for the linear effect of sex, BMI, LPL,
or log HDL, cholesterol. In contrast, the partial correlation coefficient between HTGL and log HDL, cholesterol (simple r = -0.326) was considerably
weakened by either BMI (r = -0.149) or log VLDLTG (r = -0.188).
To examine further the difference in the interrelationships between the TG lipases and lipoprotein
lipids, the data were analyzed by stepwise multiple
regression (Table 4). With HTGL activity as the dependent variable, the correlation was strongest with
log VLDL-TG and was entered into the regression
equation before HDL-TG, LDL-TG, BMI, LPL activity,
and sex. When stepwise multiple regression was
performed with HTGL as the dependent variable and
with log HDLj cholesterol, but not HDL-TG or LDLTG, less variability in HTGL activity was explained
(42% vs 64%).
Table 4. Regression Statistics with the Dependent
Variable = HTGL Activity (Men and Women Combined, n = 50)
B
SE(B)
F
r2
a. log VLDL-TG
HDL-TG
LDL-TG
BMI
LPL
Sex
(constant)
86.604
-11.631
-2.885
9.284
-0.262
8.016
63.492
35.249
2.522
1.122
3.018
0.155
0.146
6.057
21.277
6.611
9.467
6.264
0.016
0.258
0.438
0.530
0.587
0.638
0.640
b. log VLDL-TG
BMI
LPL
log HDL2-C
Sex
(constant)
65.829
8.279
-0.428
-37.130
-15.874
-17.863
44.024
3.775
0.199
45.467
25.956
2.236
4.810
4.623
0.667
0.405
0.258
0.329
0.399
0.415
0.420
Variable
HEPATIC TRIGLYCERIDE LIPASE AND LIPIDS
Applebaum-Bowden et al.
HTGL
300
LPL
•
300
A
Men
r, - 0.382
p-0.014
200
A
279
Men
r, = -0.288
p - 0.052
200
A
A
A
A
100
A
A
A,
100
A
A
V
*
A
-A
A
A
A
A
A
A
AA
A
AA
0
0
300
300
Women
r, - 0.742
p = 0.001
200
Women
r, - -0.355
p - 0.081
•
200
-
100
O
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
o
100
o
0
oo
n
° °o
o
o
°
o
°
o
o
o
200
400
o
oo°
o° o
o
200
0
Activity, nmol/min/ml
400
Figure 3. Relationships between hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activity and very low
density lipoprotein triglyceride (VLDL-TG) levels in the men (A) and women (o). Significance was determined with the
Spearman rank correlation coefficient (rs).
LPL
HTGL
A
60
Men
r$ = -0.354
p » 0.022
60
Men
r, = 0.092
NS
A
A
40
40
A
A
A
A
A
A
20
A
A
A A
A
A
A
A
&
&
•a
"a
20,
A
^ A
A
A
A
a«&
A
A
A*
AA
A
AA*
*%
A ^
0
0
cf
IN
_l
60
Women
r, - -0.503
p - 0.020
Q
X
o
o
O
0O
o
o
o
o
o
O
CD
o °
0
Women
r, - 0.328
p = 0.099
40
40
20
60
0
°
o
o
o
200
o
400
20
O
O
O
•
o
0
0
Activity, nmol/min/ml
°
gOO
°O
200
400
Figure 4. Relationships between hepatic triglyceride lipase (HTGL) and lipoprotein lipase (LPL) activity and high density
lipoprotein2 cholesterol (HDL2C) levels in the men (A) and women (o). Significance was determined with the Spearman rank
correlation coefficient (rs).
280
Table 5.
ARTERIOSCLEROSIS
VOL
5, No 3,
MAY/JUNE
1985
Comparison of Trlglycerlde Llpase Levels (Mean ± SD)
Reference
Current report
(M, n = 33; W,
Krauss et al.6
(M, n = 40; W,
Greten et al.31
(M, n = 24; W,
Huttunen, et al.26
(M, n = 47; W,
Heparin
dose
(U/kg)
Time
after
injection
(min)
10
10
235 ±84
170±91t
117±61
149±57
10
10
250 ±90
175 + 55*
70 ±25
82 ±38
10
10
90 + 41
53 + 25*
35 ±20
29±16
100
15
447±165
310 + 125*
310±103
430 ±155*
Hepatic triglyceride lipase
Men
Lipoprotein lipase
Men
Women
(nmol/min/ml)
Women
n = 17)
n = 39)
n = 24)
n = 35)
* p < 0.001.
t p < 0.02.
Discussion
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In contrast to earlier reports which measured activity, 62631 the current study was designed to determine the relationships between postheparin plasma
HTGL activity and lipoprotein lipids in a well-defined
population. Due to the relatively small sample size,
caution should be used in interpreting the probability
values. Nevertheless, the mean HTGL values reported here are almost identical to those found by Krauss
et al.6 in a larger sample of normal volunteers aged
19 to 62 years (Table 5). Greten et al.31 reported
HTGL levels about one-third of the activity in the
current study. When Huttunen et al.28 used a higher
heparin dose, they obtained HTGL levels about twice
that reported here. However, regardless of the absolute values, all four studies showed lower HTGL levels in women. Postheparin plasma LPL levels were
not found to be statistically different in men versus
women in the current study, nor in two of the other
studies, but were higher in women in the report of
Huttunen et al. Like HTGL, the levels of LPL activity
differed among the studies and were highest at the
higher heparin dose.
We have previously examined the effect of sex
steroids on HTGL levels and found that ethinyl estradiol administration was associated with lower HTGL
levels,28 while androgen (stanozoiol) administration
was associated with higher HTGL levels.32 Another
androgenic steroid, oxandrolone, also has been reported to increase HTGL levels.33 If endogenous sex
steroids were to act similarly to the exogenous ones,
they might be responsible for the observed lower
HTGL activity in women versus men. Furthermore, a
sex steroid-related difference would be expected to
decrease with age such as that shown for HTGL.
Although the effects of Premarin on HTGL activity
have not been studied, we would have predicted
that, like ethinyl estradiol, it would decrease HTGL
activity. In contrast to this prediction, HTGL activity in
the women taking Premarin was similar to that in
women not taking postmenopausal estrogens. This
similarity in HTGL activity may be due to Premarin
having a much lower potency than ethinyl estradiol.
Alternatively, it may be that Premarin did lower the
activity and that without Premarin the activity would
have been even higher (due to decreasing endogenous estrogens with age). If the latter is true, the
failure to find a significant relationship between
HTGL activity and age in women in our current study
and in that of Huttunen et al. might be due to the
tendency to treat older postmenopausal women with
estrogens.
HTGL levels have been reportedly higher in male
smokers34 and alcoholics.35 In the current study, neither HTGL nor LPL levels were significantly related
to smoking or alcohol consumption. It is tempting to
suggest that one reason for the apparent difference
in the data of smokers is related to the populations
studied. Kuusi et al.34 examined a group of young,
very physically fit men (military academy students)
living under strictly comparable conditions, whereas
our population consisted of older men and women in
a much more diverse environment. We did, however,
find that another enzyme related to HDL cholesterol
metabolism, lecithin-cholesterol acyltransferase,
was lower in the smokers in our population.2136
We have previously shown a negative correlation
between HTGL activity and HDL cholesterol levels in
nromal men.12 In the current study, HTGL levels
were significantly negatively related with the levels of
HDL, cholesterol, but not with total HDL cholesterol
or HDL3 cholesterol. Similarly, Kuusi et al.14 found a
negative correlation between HTGL levels and HDLj
cholesterol but not HDL, cholesterol. VLDL-TG levels have reportedly been inversely related to HDL
cholesterol levels,37-38 but neither of these earlier
studies controlled for VLDL-TG. In the current study,
HTGL activity was significantly positively correlated
with VLDL-TG levels. Since VLDL-TG also was correlated with BMI and HDLj cholesterol, we examined
the effect of controlling for these variables. Although
the positive correlation between HTGL activity and
VLDL-TG levels was not diminished significantly by
controlling for either BMI or HDL-, cholesterol, the
relationship between HTGL activity and HDL, cholesterol was considerably weakened after controlling
for either BMI or VLDL-TG. Thus, the relationship
between HTGL and VLDL-TG appeared to be inde-
HEPATIC TRIGLYCERIDE LIPASE AND LIPIDS
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pendent of BMI or HDL, cholesterol, but the relationship with HDL, cholesterol was not independent of
BMI or VLDL-TG.
If the assay overestimated HTGL activity, it would
underestimate lipoprotein lipase and there would be
an inverse relationship. Thus, the inverse relationship between HTGL and LPL levels may be due to
the fact that LPL was estimated by difference. Nevertheless, LPL levels tended to be inversely related to
VLDL-TG levels in the current study as we described
earlier.2839 Further evidence of this relationship was
evident in the negative correlation with VLDL cholesterol. This inverse relationship is consistent with LPL
playing a rate-limiting role in VLDL catabolism.
The positive correlation between HTGL activity
and VLDL-TG levels in this report appears to present
a paradox. If HTGL removes TG from VLDL, it should
be negatively related to VLDL-TG. The positive relationship suggests that in contrast to LPL, HTGL does
not have a strong role in VLDL-TG catabolism. Consistent with these findings, Huttunen et al.26 and
Reardon et al.40 have examined HTGL levels and
VLDL metabolism and found no evidence that HTGL
played a rate-limiting role in the catabolism of TGrich lipoproteins when LPL activity was normal. However, patients lacking LPL (Type I hyperlipoproteinemia) frequently have normal or only slightly elevated
VLDL levels.41 When Nicoll and Lewis42 studied
VLDL apo B conversion in these patients, they found
that it was normal, as was HTGL activity against
VLDL. Furthermore, at equal lipoprotein TG concentrations, HTGL had increasing activity with lipoproteins of decreasing particle size. Thus, Nicoll and
Lewis calculated that the mean contribution of HTGL
to VLDL-TG hydrolysis by postheparin plasma was
35% in normal controls, but to intermediate density
lipoprotein-TG, hydrolysis was up to 58%. This conclusion is supported by the fact that two brothers who
had no HTGL did have abnormal VLDL ((3-VLDL),
and increased levels of TG in LDL and HDL.8
Krauss et al.6 and Musliner et al.7 have examined
the ability of HTGL to hydrolyze lipoprotein lipids in
vitro. They both found that HTGL hydrolyzes TG
from VLDL 2 to 7 times better than from chylomicrons, although LPL still hydrolyzed VLDL-TG more
avidly than did HTGL. Musliner et al. also showed
that HTGL was better able to hydrolyze LDL-TG and
HDL-TG than was LPL. HTGL, in contrast to LPL,
could release a small amount of fatty acid from TGrich HDL, but LPL could not. These in vitro studies
are consistent with the results of the stepwise multiple regression analysis performed in the current
study, where HDL-TG and LDL-TG were quite
strongly inversely related to HTGL activity, but not to
LPL activity. In the studies by Musliner et al. fatty acid
released by HTGL was from TG, not from phospholipids, and they suggested that in severe hypertriglyceridemia, where TG may replace cholesterol ester as the predominant neutral lipid in LDL and HDL,
the role of HTGL may be even more important. Thus,
the in vitro studies complement the in vivo ones and
Applebaum-Bowden et al.
281
support the suggestion that human HTGL is probably
important in catabolism of VLDL-remnants, IDL,
LDL, and HDL, and may only be important in VLDL
catabolism when LPL is abnormal.
Acknowledgments
The authors appreciate the enthusiastic help of Pam McLean
and Betty King in their work with the subjects at the clinic visits. We
are also grateful to Helen K. Luk and Karen Lum for the lipase
measurements, Joan Benderson for separating the HDL subtractions, Paul Comeaux for helping with the computer programming,
and Donna Sims for typing the manuscript.
References
1. Fielding CJ. Human lipoprotein lipase. I. Punfication and
substrate specificity. Biochim Biophys Acta 1970;206:
109-117
2. LaRosa JC, Levy Rl, Wlndmueller HG, Fredrickson DS.
Comparison of the tnglyceride lipase of liver, adipose tissue,
and post-heparin plasma. J Lipid Res 1972;13:356-363
3. Boberg J, Augustln J, Baglnsky M, Tejada P, Brown WV.
Quantitative determination of hepatic and lipoprotein lipase
activities from human post-heparin plasma. Circulation 1974;
49-50 (suppl lll):21
4. Hernell O, Egelrud T, Ollvecrona T. Serum-stimulated
lipases (lipoprotein lipases). Immunological crossreaction between the bovine and the human enzymes. Biochim Biophys
Acta 1975;381:233-241
5. Huttunen JK, Ehnholm C, Klnnunen PKJ, Nlkklla EA. An
immunochemical method for the selective measurement of
two triglyceride lipases in human postheparin plasm. Clin
Chim Acta 1975:63:335-347
6. Krauss RM, Levy Rl, Fredrickson DS. Selective measurement of two lipase activities in post-heparin plasma from normal subjects and patients with hyperlipoproteinemia. J Clin
Invest 1974:54:1107-1124
7. Musliner TA, Herbert PN, Kingston MJ. Lipoprotein substrates of lipoprotein lipase and hepatic triacylglycerol lipase
from human post-heparin plasma. Biochim Biophys Acta
1979:575:277-288
8. Breckenrldge WC, Little JA, Alaupovlc P, et al. Lipoprotein
abnormalities associated with a familial deficiency of hepatic
lipase. Atherosclerosis 1982:45:161-179
9. Robinson DS. The function of the plasma triglycerides in
fatty acid transport. In: Florkin M, Stotz EM, eds. Comprehensive biochemistry, 18. Upid biochemistry. Amsterdam: Elsevier 1970:51-116
10. Havel RJ, Gordon R. Idiopathic hyperlipidemia. Metabolic
studies in an affected family. J Clin Invest 1960;39:
1777-1790
11. Fredrickson DS, Ono K, Davis LL. Lipolytic activity of postheparin plasma in hypertriglyceridemia. J Lipid Res 1963;
4:24-33
12. Applebaum-Bowden D, Goldberg AP, Hazzard WR, et al.
Postheparin plasma triglyceride lipases in chronic hemodiarysis: Evidence for a role for hepatic triglyceride lipase in lipoprotein metabolism. Metabolism 1979;28:917-924
13. Nlkklla EA, Taskinen M-R, Kekkl M. Relation of plasmahigh-density lipoprotein cholesterol to lipoprotein-lipase activity in adipose tissue and skeletal muscle of man. Atherosclerosis 1978:29:497-501
14. Kuusl T, Saarlnen P, Nikklla EA. Evidence for the role of
hepatic endothelial lipase in the metabolism of plasma high
density lipoprotein2 in man. Atherosclerosis 1980;36:
589-593
282
ARTERIOSCLEROSIS VOL 5, No 3, MAY/JUNE 1985
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
15. Nikkila EA, Kuusi T, Taskinen M-R. Role of lipoprotein
lipase and hepatic endothelial lipase in the metabolism of
high density lipoproteins: a novel concept on cholesterol
transport in HDL cycle. In: Carlson LA, Pernow B, eds. Metabolic risk factors in ischemic cardiovascular disease. New
York: Raven Press, 1982:205-215
16. Barr DP, Russ EM, Eder HA. Protein lipid relationship in
human plasma. II In atherosclerosis and related conditions.
Am J Med 1951 ;11:480-^93
17. Miller GJ, Miller NE. Plasma high density lipoprotein concentration and development of ischemic heart disease. Lancet
1975:1:16-19
18. Miller NE, Hammet F, Saltissi S, et al. Relation of angiographically defined coronary artery disease to plasma lipoprotein subfractions and apolipoproteins. Br Med J 1981;
282:1741-1744
19. Ballantyne FC, Clark FS, Simpson HS, Ballantyne D. High
density and low density lipoprotein subfractions in survivors
of myocardial infarction and in control subjects. Metabolism
1982;31:433-437
20. Hoover JJ, Walden CE, Bergelin RO, et al. Cholesterol and
triglyceride distribution in an adult employee population: The
Pacific Northwest Bell Telephone Company Health Survey.
Lipids 1980:15:895-903
21. Wahl PW, Warnick GR, Albers JJ, et al. Distribution of
lipoprotein triglyceride and lipoprotein cholesterol in an adult
population by age, sex, and hormone use. Atherosclerosis
1981; 39:111-124
22. Haffner SM, Applebaum-Bowden D, Wahl PW, et al. Epidemiological correlates of high density lipoprotein subfractions, apolipoproteins A-l, A-ll and D, and lecithin cholesterol
acyltransferase. Effects of smoking, alcohol and adiposity.
Arteriosclerosis 1985;5:169-177
23. Lipid Research Clinics Program. Reference Manual for
Lipid Research Clinics Program Prevalence Study. University
of North Carolina Chapel Hill: Central Patient Registry and
Coordinating Center for Lipid Research Clinics, Department
of Biostatistics, 1974
24. Lipid Research Clinics Program. Manual of laboratory operations. Vol 1: Lipid and lipoprotein analysis. DHEW publications no (NIH) 75-628. Washington DC: U.S. Government
Printing Office, 1974
25. Gidez LI, Miller GJ, Burstein M, Slagles S, Eder HA. Separation and quantitation of subclasses of human high density
lipoprotein subclasses by a simple precipitation procedure. J
Lipid Res 1982;23:1206-1223
26. Huttunen JK, Ehnholm C, Kekki M, Nikkila EA. Postheparin plasma lipoprotein lipase and hepatic lipase in normal subjects and in patients with hypertriglyceridaemia: Correlations to sex, age and various parameters of triglyceride
metabolism. Clin Sci Mol Med 1976;50:249-260
27. Belfrage P, Vaughan M. Simple liquid-liquid partition system
for isolation of labeled oleic acid from mixtures with glycerides. J Lipid Res 1969:10:341-344
Index Terms:
hepatic triglyceride lipase
28. Applebaum DM, Goldberg AP, Pykalisto OJ, Brunzell JD,
Hazzard WR. Effect of estrogen on postheparin lipolytic activity: Selective decline in hepatic triglyceride lipase. J Clin Invest 1977:59:601-608
29. Armitage P. Statistical methods in medical research. New
York: John Wiley 1971:396-398
30. Snedecor GW, Cochran WG. Statistical methods, 7th ed.
Ames, Iowa: Iowa State University Press 1980:191-192
31. Greten H, DeGrella R, Klose G, Rascher W, DeGennes JL,
Gjone E. Measurement of two plasma triglyceride lipases by
an immunochemical method: Studies in patients with hypertriglyceridemia. J Lipid Res 1976:17:203-210
32. Taggart H, Applebaum-Bowden D, Haffner S, et al. Reduction in high density lipoproteins by anabolic steroid (stanozolol) therapy for postmenopausal osteoporosis. Metabolism
1982:31:1147-1152
33. Ehnholm C, Huttunen JK, Kinnunen PJ, Miettinen TA,
Nikkila EA. Effect of oxandrolone treatment on the activity of
lipoprotein lipase, hepatic lipase and phospholipase A, of
human post-heparin plasma. N Engl J Med 1975;292:
1314-1317
34. Kuusi T, Nikkila EA, Saarinen P, Varjo P, Laitinen LA.
Plasma high density lipoproteins HDL2, HDL3 and postheparin plasma lipases in relation to parameters of physical
fitness. Atherosclerosis 1982:41:209-219
35. Taskinen MR, Valimaki M, Nikkila EA, Kuusi T, Ehnholm
C, Ylikahri R. High density lipoprotein subfractions and
postheparin plasma lipases in alcoholic men before and after
ethanol withdrawal. Metabolism 1982;31:1168-1174
36. Albers JJ, Bergelin RO, Adolphson JL, Wahl PW. Population-based reference values for lecithin-cholesterol acyltransferase (LCAT). Atherosclerosis 1982;43:369-379
37. Olsson AG, Walldius G, Rossner S, Callmer E, Kaijser L.
Studies on serum lipoproteins and lipid metabolism. Analysis
of a random sample of 40 year old men. Acta Med Scand
(suppl637) 1980:1^17
38. Walden CE, Knopp RH, Wahl PW, et al. Hyperlipidemia in
the Pacific Northwest Bell Telephone Company Health Survey. Part 2. Lipoprotein lipid interrelationships. Arteriosclerosis 1983;3:125-131
39. Goldberg AP, Applebaum-Bowden DM, Bierman EL, et al.
Increase in lipoprotein lipase during clofibrate treatment of
hypertriglyceridemia in patients on hemodialysis. N Engl J
Med 1979:301:1073-1076
40. Reardon MF, Sakai H, Steiner G. Roles of lipoprotein lipase
and hepatic lipase in the catabolism in vivo of triglyceride-rich
lipoproteins. Arteriosclerosis 1982:2:396-402
41. Nikkila EA. Familial lipoprotein lipase deficiency and related
disorders of chylomicron metabolism. In: Stanbury JB, Wyngaarden JB, Frederickson DS, Goldstein JL, Brown MS, eds.
New York: McGraw Hill Book Company, 1983:622-642
42. Nicoll A, Lewis B. Evaluation of the roles of lipoprotein lipase
and hepatic lipase in lipoprotein metabolism: in vivo and in
vitro studies in man. Eur J Clin Invest 1980:10:487-495
• lipoprotein lipase
• lipoprotein lipids
Downloaded from http://atvb.ahajournals.org/ by guest on June 15, 2017
Postheparin plasma triglyceride lipases. Relationships with very low density lipoprotein
triglyceride and high density lipoprotein2 cholesterol.
D Applebaum-Bowden, S M Haffner, P W Wahl, J J Hoover, G R Warnick, J J Albers and W R
Hazzard
Arterioscler Thromb Vasc Biol. 1985;5:273-282
doi: 10.1161/01.ATV.5.3.273
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