1 Department of Physiology, Göteborg University, 405 30 Goteborg; and 2 AstraZeneca Research and Development, 431 83 Molndal, Sweden
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ABSTRACT |
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The effects of long-term chronic
growth hormone (GH) excess on lipid and lipoprotein metabolism were
investigated in 8-mo-old bovine GH (bGH)-transgenic mice. Total body
weight, serum cholesterol, insulin-like growth factor-I, and insulin
levels were higher, whereas serum levels of glucose, free fatty acids,
and triglycerides were lower in transgenic mice. Very low-density
lipoprotein (VLDL) cholesterol levels were lower, and low-density
lipoprotein (LDL) cholesterol levels were higher, in transgenic mice
irrespective of gender, whereas only transgenic male mice had higher
high-density lipoprotein cholesterol levels. Total serum apolipoprotein
B (apoB) levels were not affected, but the amount of apoB in the LDL
fraction was higher in transgenic mice. Hepatic LDL receptor expression was unchanged, whereas apoB mRNA editing and hepatic triglyceride secretion rate were reduced in bGH-transgenic male mice. Both lipoprotein lipase activity in adipose and heart tissue and
-adrenergic-stimulated lipolysis were increased in transgenic male
mice. The relative weight of adipose tissue was lower in transgenic
mice, whereas hepatic triglyceride content was unchanged. Fat feeding
of the mice equalized serum triglycerides and free fatty acids in
bGH-transgenic and control mice. In summary, long-term GH excess is
associated with marked alterations in lipid and lipoprotein metabolism,
indicating decreased production and increased degradation of VLDL and
preferential flux of fatty acids to muscle tissues.
very low-density lipoprotein; low-density lipoprotein; high-density lipoprotein; apolipoprotein B; low-density lipoprotein receptor; apolipoprotein B mRNA editing
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INTRODUCTION |
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TRANSGENIC MICE with chronic overexpression of growth hormone (GH) have been extensively used to investigate long-term effects of GH (19). Besides increased body size, linear growth, and organomegaly, these mice have been shown to have decreased body fat mass (13, 31, 41) and increased lean body mass (61). Metabolic studies of GH-transgenic mice have consistently shown hyperinsulinemia, normal blood glucose levels, and slightly impaired (26) or normal (19) glucose tolerance. Bovine (b)GH-transgenic mice have reduced insulin receptor concentration but increased insulin receptor substrate-1 (IRS-1) phosphorylation and phosphatidylinositol 3-kinase activity in the liver (10). However, these mice were insensitive to further stimulation with insulin with respect to insulin signaling (10). Furthermore, bGH-transgenic mice have been shown to have impaired cardiac function and bioenergetics (4) and increased locomotor activity (52). Thus many aspects of metabolism, including carbohydrate metabolism, insulin signaling, and cardiac bioenergetics, have been studied, but few studies concerning lipid and lipoprotein metabolism in the GH-transgenic mice have been performed. bGH (5, 42) and GRF-transgenic (42) mice have been shown to have increased cholesterol levels, whereas serum triglycerides were unchanged. Many treatment studies of GH-deficient patients and hypophysectomized rats have revealed that GH influences several aspects of lipid and lipoprotein metabolism. These aspects include alterations in serum lipoprotein levels (12, 16, 33-35, 51), hepatic lipoprotein production (14, 24, 49, 50), hepatic low-density lipoprotein (LDL) receptor expression (1, 45), and lipoprotein lipase (LPL) activity (35, 36) after GH treatment. However, many of the underlying mechanisms of GH action on lipoprotein metabolism, including which genes are involved, are to a large extent still unknown.
Transgenic mice overexpressing or lacking a gene have been used extensively over the last years to study the importance of various genes in lipid and lipoprotein metabolism (6, 62). Transgenic mice have also made it possible to investigate hormone regulation of human genes such as apolipoprotein a (56). The potential of using transgenic mice for the understanding of which genes are responsible for the action of GH on lipid and lipoprotein metabolism led us to investigate the effects of chronic overexpression of bGH on lipid and lipoprotein metabolism in mice.
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MATERIALS AND METHODS |
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Generation of Transgenic Mice
bGH-transgenic mice used in this study were generated as previously described (46). The animals were housed with alternating 12-h periods of light (0700-1900) and dark (1900-0700). The animals had free access to mouse standard chow (rat/mouse standard diet, B&K Universal, Sollentuna, Sweden) and tap water. If not otherwise stated, the mice were killed at the age of 8 mo. All mice were killed at a similar time interval of the day (0900-1200) to avoid the effects of diurnal variation. In one experiment, female mice had free access to a high-fat diet for a period of 2 mo; these mice were killed at 6 mo of age. The high-fat diet contained 57.6% energy from fat, 26.6% from carbohydrates, and 15.8% from protein (Bio-Serv, Frenchtown, NJ). The animals were anesthetized with xylazine (Rompun, 9 mg/kg; Bayer, Lever-kusen, Germany) and ketamine hydrochloride (Ketalar, 77 mg/kg; Parke-Davis, Detroit, MI). Blood was collected by heart puncture using nonheparinized syringes. Adipose tissue depots, heart, and liver were excised, blotted, weighed, and instantly frozen in liquid nitrogen. Serum and tissues were kept atSerum Analyses
Triglyceride and cholesterol concentrations were determined by enzymatic colorimetric assays (MPR2: TG/GPO-PAP and Chl/CHOD-PAP, Roche Diagnostics, Mannheim, Germany). The intra-assay coefficient of variation (CV) was 3% for the triglyceride assay and 4% for the cholesterol assay. Serum apolipoprotein B (apoB) concentrations were determined by an electroimmunoassay as previously described (33, 51). The intra- and interassay CVs for the apoB assay were 5.3 and 8.6%, respectively. Standardization of apoB measurements was made by isolation of a narrow-density cut of LDL (density 1.030-1.055 g/ml) containing only apoB (33). The protein concentration of the narrow-density cut of LDL was determined by the Bio-Rad protein assay (Bio-Rad Laboratories, Hercules, CA) using albumin as standard. Serum concentration of glucose was enzymatically determined in 15-µl samples on a YSI 2700 SELECT biochemical analyzer (Yellow Springs Instrument, Yellow Springs, OH). Serum free fatty acid (FFA) concentrations were measured with a NEFA kit (Wako Chemicals, Neuss, Germany). Serum insulin concentration was determined by a rat insulin RIA (Linco Research, St. Charles, MO) with 100% cross-reactivity to mouse insulin. Serum insulin-like growth factor I (IGF-I) concentrations were determined by a hydrochloric acid-ethanol extraction followed by RIA with the use of human IGF-I for labeling (Nichols Institute of Diagnostics, San Juan Capistrano, CA).Hepatic Lipid Content
Frozen liver was homogenized in ice-cold distilled water (6 ml/g tissue). Total lipid content was extracted from the homogenate according to Bligh and Dyer (3). The extracted lipids were dissolved in isopropanol after evaporation of the chloroform phase by use of a stream of N2. Triglyceride and cholesterol concentrations were determined as already described.Size Distribution of Serum Lipoproteins
Determination of size distribution of lipoproteins was performed by gel filtration with the use of fast protein liquid chromatography (FPLC) equipment (Pharmacia Upjohn, Uppsala, Sweden). Briefly, serum from six mice was pooled to a total volume of 1.5 ml, and the density was adjusted to 1.215 g/ml with KBr in 0.9% NaCl. After ultracentrifugation (35,000 g, 4°C, 24 h), the total lipoprotein fraction was recovered by aspiration, and the final volume was adjusted to 2 ml with FPLC buffer (0.15 M NaCl, 0.01% EDTA, 0.02% NaAz, pH 7.3). After filtration through a 0.45-µm low-protein filter, the sample was loaded on a 25-ml Superose 6B column (Pharmacia Upjohn) using a constant flow rate of 0.35 ml/min. Eluted samples were collected in 0.5-ml fractions, and the fractions were stored atIn Vivo Hepatic Triglyceride Secretion Rate
Triglyceride secretion rate in vivo was measured by intravenous administration of Triton WR-1339 (22). After a 5-h period without access to mouse standard chow (0700-1200), anesthetized wild-type and bGH-transgenic male mice were injected intravenously with Triton WR-1339 (Sigma Chemical) diluted in saline (200 mg/ml) (500 mg/kg body wt) via the jugular vein by means of catheters. Blood samples (70 µl) were taken before and 30 and 60 min after Triton WR-1339 injection. The triglyceride accumulation was linear during this time period. The triglyceride concentration was analyzed as described in the previous section.Western Blot Analyses
ApoB. Western blotting was performed using an enhanced chemiluminicence (ECL) protocol (Amersham Pharmacia Biotech, Buckinghamshire, UK). In each lane, adjacent FPLC fractions were pooled in pairs (9 µl each) and separated on 4-20% Tris-glycine polyacrylamide gels (Novex, San Diego, CA). After electrophoresis, the proteins were transferred to Hybond-P polyvinylidene difluoride transfer membrane (Amersham Pharmacia Biotech) in transfer buffer (25 mM Bis-Tris, pH 7.4, with 25 mM bicine, 0.8 mM EDTA, and 10% methanol) for 3 h at 400 mA (Transblot cell, Bio-Rad). The membrane was blocked overnight at 4°C in PBS containing 0.1% Tween-20 (PBS-T) and 5% nonfat milk before 1-h incubation with polyclonal rabbit anti-rat apoB antiserum (33) diluted 1:1,000 in PBS-T. The membrane was then incubated for 1 h with horseradish peroxidase-linked donkey anti-rabbit IgG (Amersham Life Science) diluted 1:4,000. Detection and development was performed by using ECL detection assay and Hyperfilm ECL (Amersham Pharmacia Biotech).
LDL receptor. To quantify the LDL receptor, livers were homogenized in 4 volumes of PE buffer (10 mM potassium phosphate buffer, pH 6.8, and 1 mM EDTA) containing 10 mM 3-([3-cholamidopropyl]dimethylammonio)-1-propanesulfate (18), 4 mM Pefabloc, 20 nM leupeptin, 15 nM pepstatin, and 0.15 nM aprotinin (Boehringer Mannheim, Mannheim, Germany). The homogenates were sonicated (3 × 10 s) and centrifuged (10,000 g, 10 min), and protein concentrations were determined in the supernatant by use of the Bio-Rad DC protein assay with albumin as standard. Proteins (30 µg/lane) were separated and transferred as described, but under nonreducing conditions. Primary antibody, rabbit anti-bovine LDL receptor antibody (a generous gift from Professor Joachim Herz, Dept. of Molecular Genetics, University of Texas, Dallas, TX), and secondary antibody were diluted 1:2,000 in blocking buffer. Detection and quantification were done with Fluor-S Multi Imager and Quantity One software (Bio-Rad). The detection capacity was found to be linear to 60 µg protein (data not shown). As a negative control, liver protein preparations from an LDL receptor-deficient mouse and a corresponding wild-type mouse (kindly provided by Dr. Marcela Pekna, Dept. of Medical Biochemistry, Göteborg University) were used (data not shown).
ApoB mRNA Editing
Total RNA was isolated from mouse livers according to Chomczynski and Sacchi (7). To analyze the extent of apoB mRNA editing, primer extension analysis was performed according to Sjöberg et al. (50). RNA (10-20 ng) was annealed overnight at 45°C to a 32P- and 5' end-labeled rat apoB oligonucleotide (2). The primer was labeled using a 5' end-labeling kit and 50 µCi of [32P]ATP (Amersham). The gel bands corresponding to the edited and nonedited apoB mRNA were cut out, and the radioactivity was counted after digestion of the gel. The intra-assay CV was <3%.LPL Activity
LPL activity in heart and adipose tissue depots was determined according to the method of Peterson et al. (40). In brief, frozen tissue (Lipolysis
Epidydimal adipose tissue was excised from four transgenic mice and four wild-type littermates. Adipocytes were isolated with collagenase A digestion (0.5 mg/ml, Sigma) in Dulbecco's medium (Life Technologies, Paisley, Scotland, UK) at 37°C for 1 h, as described by Rodbell and Krishna (44) and with the modifications made by Gause et al. (17). After digestion, liberated adipocytes were collected by filtration through a mesh filter (250 µm) and washed three times with medium. Thereafter, a 50/50 cell suspension was prepared using isolated fat cells and incubation medium. For each incubation, 100 µl of the 50/50 cell suspension was added to plastic vials containing 2 ml of incubation medium, supplemented with 4% BSA (fraction V, Sigma), in the absence (basal lipolysis) or presence of 10Statistics
Values are given as means ± SE. Comparison between groups was performed with unpaired Student's t-test or one-way ANOVA followed by Student-Newman-Keul's multiple range test. P values <0.05 were considered significant. When appropriate, values were normalized by logarithmic transformation. ![]() |
RESULTS |
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Chronic overexpression of bGH in transgenic mice resulted in a
significant increase in body weight and absolute (not shown) and
relative weights of liver and heart in both sexes (Table
1). The absolute weight of
retroperitoneal adipose tissue depots was significantly lower in both
male and female transgenic mice compared with wild-type mice, but there
was no significant difference in absolute weights of the reproductive
adipose tissue depots (not shown). The relative weights (%body wt) of
retroperitoneal and reproductive adipose tissue were significantly
lower in both male and female transgenic mice (Table 1).
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Serum IGF-I concentrations were higher in male bGH-transgenic mice than
in their controls (Table 2). Serum
insulin levels were severalfold higher, and serum levels of glucose and
FFA were lower, in both male and female bGH-transgenic mice compared
with their gender controls (Table 2). Serum cholesterol levels were higher in both male and female bGH-transgenic mice compared with their
controls (Fig. 1A). However,
serum cholesterol levels were more markedly elevated in transgenic
males than in females (P < 0.05, one-way ANOVA
followed by Student-Newman-Keul's test). Serum triglyceride levels
were reduced to the same extent in male and female transgenic mice
(Fig. 1B). Changes in size distribution of lipoproteins were
determined by FPLC analysis of total lipoproteins (d < 1.215 g/ml). Size distribution of serum lipoproteins in transgenic male and
wild-type male mice are shown in Fig. 2,
A and B. Transgenic male mice showed a marked
reduction in cholesterol and triglyceride content of the VLDL fraction
and increased cholesterol content of the LDL and HDL fractions. The
distribution of apoB48 and apoB100 was
determined by Western blotting. As shown in Fig. 2C, the
distribution of apoB was changed. The amount of B48 and
B100 in VLDL was markedly reduced in transgenic male mice
compared with their controls. The size distribution of serum
lipoproteins in transgenic female mice and their controls was also
determined, since the effect of bGH overexpression on total serum
cholesterol levels was less marked than in transgenic males (Fig.
3, A and B).
Compared with littermates, the cholesterol (Fig. 3A) and
triglyceride content (Fig. 3B) of the VLDL fraction was
lower and LDL cholesterol was higher in transgenic female mice.
However, no obvious difference in HDL cholesterol content was observed
between female bGH mice and their littermates. The distribution of apoB
was changed in a similar way from VLDL to LDL to what was observed in
males (data not shown).
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The lower triglyceride and FFA levels in the bGH-transgenic mice could be due to a decreased availability of fatty acids for hepatic VLDL production and increased uptake of fatty acids in other tissues. If a decreased hepatic fatty acid supply was important for the observed effects, an increased amount of dietary triglycerides would reduce the difference in serum triglycerides and FFA between the transgenic mice and their controls. We therefore gave a diet enriched in triglycerides to bGH-transgenic females (n = 5) and their littermate controls (n = 5) for 2 mo. In fact, there was no difference in serum triglycerides between bGH-transgenic females and gender controls after 2 mo of fat feeding (bGH-transgenic mice: 1.25 ± 0.27 mmol/l; control mice: 0.70 ± 0.11 mmol/l, not significant). Moreover, no difference in serum FFA levels was observed between transgenic and wild-type female mice fed the high-fat diet (data not shown).
Total serum apoB levels were not different between transgenic male and
wild-type control mice (Table 3). Because
the distribution of apoB was changed (Fig. 2C), it could be
concluded that the apoB content of the LDL fraction was higher, and the
apoB content of the VLDL fraction was lower, in transgenic male mice
compared with controls. Because a reduced LDL receptor expression could explain the increased LDL-apoB levels, the LDL receptor expression was
measured in protein extract from whole liver. However, LDL receptor
expression was not different between transgenic male and wild-type male
mice (Table 3).
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Overexpression of bGH resulted in slightly lower editing of apoB mRNA
compared with wild-type littermates (Table 3). Increased editing of
apoB mRNA has been shown to be associated with increased hepatic
triglyceride production in different models (2, 24). The
reduced editing of apoB mRNA in bGH-transgenic mice therefore indicates
decreased triglyceride synthesis and VLDL secretion in bGH-transgenic
mice. In fact, the hepatic triglyceride secretion rate was decreased
~50% in transgenic male mice compared with wild-type littermates
(Fig. 4). The secretion rate was related to the body weight and plasma volume estimated by Li et al.
(22). Long-term GH treatment of GH-deficient humans
results in an increased plasma volume (27). Therefore, the
bGH-transgenic mice may have increased plasma volume per gram of body
weight, leading to an underestimation of the true triglyceride
secretion rate. However, it is unlikely that an increased plasma volume
can explain the difference in the triglyceride secretion rate, because
the plasma volume per gram of body weight would have had to be
increased by ~100% to balance out the difference observed between
control and bGH-transgenic mice. The 5-h period of fasting before the hepatic triglyceride secretion rate was measured did not influence the
previously described difference in serum triglycerides between transgenic and wild-type male mice (0.76 ± 0.08 vs. 1.74 ± 0.09 mmol/l, P < 0.05). The hepatic triglyceride
content was not different between transgenic male mice and their
controls (Table 3), indicating that triglycerides were not accumulating
as a result of impaired VLDL secretion. However, hepatic cholesterol
content was somewhat higher in the transgenic male mice than in
wild-type male mice (Table 3).
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Despite a marked reduction in the hepatic triglyceride secretion rate,
LDL and HDL levels were higher in the transgenic male mice. This
finding could be at least partly explained by an increased turnover of
VLDL via increased LPL activity (39, 62). The LPL activity
was therefore measured in heart and adipose tissue of male mice. The
LPL activity in heart was nearly twofold higher in male transgenic mice
compared with wild-type littermates (Fig. 5A). LPL activity in
epididymal and retroperitoneal adipose tissue was 2- to 2.5-fold higher
in both adipose tissue depots in transgenic male mice compared with
wild-type controls (Fig. 5, B and C). The
increased LPL activity in adipose tissue may result in increased availability of triglyceride-derived fatty acids, leading to increased triglyceride accumulation if not balanced by increased lipolysis and/or
decreased lipogenesis. We therefore investigated the lipolytic response
in epidydimal adipose tissue. Lipolytic response in adipose tissue was
measured as glycerol release from isolated adipocytes during basal and
-adrenergic stimulation. Basal lipolysis was not changed, but
norepinephrine-stimulated lipolysis was significantly higher in
transgenic males compared with their controls (Fig. 6).
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DISCUSSION |
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This study demonstrates that chronic excess of GH leads to
significant alterations in lipid and lipoprotein metabolism. The marked
reduction in VLDL levels could be explained both by decreased hepatic
secretion of triglycerides and by increased LPL activity. Despite an
increased lipolytic response to -adrenergic stimuli in adipose
tissue, serum levels of FFA were reduced in transgenic mice. These
findings, together with reduced adipose tissue mass, reduced hepatic
triglyceride secretion, and unchanged mass of hepatic triglycerides,
indicate decreased triglyceride biosynthesis in both liver and adipose
tissue of bGH-transgenic mice. The similar serum triglyceride levels in
transgenic mice and controls after being fed a triglyceride-enriched
diet indicates that the reduced triglyceride production in the liver
was most likely due to a reduced fatty acid supply. Fatty acids
liberated through the lipolytic effects of GH in adipose tissue and
increased LPL activity in adipose tissue and heart may therefore be
rerouted preferentially to heart and muscle tissue. Moreover, these
findings, in combination with low glucose levels, high insulin levels,
and increased locomotor activity in mice kept on a low-fat diet
(52), indicate a substantial carbohydrate oxidation for
energy production in the bGH-transgenic mice. In line with this
assumption, subjects with acromegaly have enhanced carbohydrate
oxidation after a glucose tolerance test compared with healthy controls
(38). Because these effects were correlated to IGF-I
levels (38) and IGF-I preferentially increases glucose
uptake in skeletal muscle (15), it is suggested that increased IGF-I levels may contribute to increased glucose oxidation in
muscle tissue also in bGH-transgenic mice.
Overexpression of GH in transgenic animals produces supraphysiological levels of GH (19). The line of bGH-transgenic mice used in this study has been shown to have mean serum bGH levels of 1,057 (46) and 1,124 ng/ml (28), which are markedly higher than in normal mice (25). The difference in serum IGF-I levels between the bGH-transgenic mice and the control mice in this study is similar to that reported previously (46), indicating that the serum levels of bGH in these mice are in the same range as that reported previously (28, 46). GH is also overexpressed in several tissues that normally do not express GH, which might lead to higher concentrations of GH locally than what is observed in the circulation.
The present results confirm previous findings of decreased adipose tissue mass (13), high serum insulin (26, 42, 60), and IGF-I levels (19, 42) in GH-transgenic mice. In contrast to our findings, comparable glucose levels in GH-transgenic and wild-type mice have been observed in other studies (19, 26). The reason for the glucose levels in the bGH-transgenic mice being lower than in their controls is unclear. The mice were not fasted, indicating that the lower glucose levels were not a result of impaired glucose production but may have been due to an enhanced glucose oxidation in these mice. Serum FFA levels were reduced in the bGH-transgenic mice, suggesting that these bGH-transgenic mice possess the ability to handle liberated FFA efficiently, thereby preventing a deterioration of insulin sensitivity induced by high FFA levels (43).
The transgenic mice overexpressing GH demonstrated a marked reduction in serum triglycerides in both the fed and fasted states. This finding is in contrast to the unchanged serum triglyceride levels in GH-transgenic mice observed in other studies (5, 42) and the unchanged or slightly reduced triglyceride levels after GH treatment of hypophysectomized rats (33, 34). Treatment of GH-deficient adults results in small or no changes in serum triglycerides, but interestingly, those subjects with the highest serum triglyceride levels before treatment show a marked decrease in serum triglycerides after GH treatment (12). In acromegaly, serum triglyceride concentrations have been shown to be higher than (29) or similar to (8) the levels in a control population, but therapy of acromegaly has been shown to decrease serum triglycerides (37, 59). The reason for the difference in serum triglycerides between acromegaly and bGH-transgenic mice is not clear from the present results. However, the similar serum triglyceride levels in bGH-transgenic mice and controls after 2 mo of fat feeding indicate that a decreased supply of fatty acids to the liver may be important. FFA are regarded as rate limiting for hepatic triglyceride synthesis, and VLDL assembly and secretion (32) and FFA levels were reduced in the bGH-transgenic mice consuming a chow diet. Thus the dietary intake of fat may be a difference between the human situation and our mouse model, which may contribute to the different effects on serum triglycerides.
When consuming ordinary chow, the bGH-transgenic mice showed a markedly lower triglyceride secretion rate than controls. In contrast, acromegalic patients have an increased triglyceride production rate (29). Thus it is possible that that an increased content of fat in the diet would increase the hepatic triglyceride secretion rate also in bGH-transgenic mice above that in control mice. However, low hepatic triglyceride secretion and reduced apoB mRNA editing in bGH-transgenic mice is in contrast to the increased VLDL secretion and increased apoB mRNA editing in GH-treated, hypophysectomized rats consuming ordinary low-fat chow (14, 49, 50). Thus other explanations than the dietary intake of fat may be important for the lower triglyceride secretion. The very high serum insulin levels in the bGH-transgenic mice may have contributed to the lower triglyceride secretion. Interestingly, hyperinsulinemic ob/ob mice also have reduced hepatic triglyceride secretion and slightly lower apoB mRNA editing (23). However, long-term incubation of hepatocytes with insulin increases both triglyceride synthesis and apoB mRNA editing (24, 58). Thus hyperinsulinemia may play a role in the observed effects, but the mechanisms behind the possible importance of insulin for these effects are unclear.
In line with the present results, we have observed that heart and gastrocnemicus muscle LPL activity was increased by GH treatment of hypophysectomized rats (36). Mainly skeletal muscle and heart LPL activity has been shown to affect circulating levels of triglycerides and to be rate limiting for the uptake of triglyceride-derived fatty acids into muscle tissue (21, 62). Thus the increased LPL activity in heart may contribute to the lower VLDL levels in the bGH-transgenic mice.
LPL activity was also increased in adipose tissue in the GH transgenic mice. This finding is in contrast to the lack of effect of GH treatment of hypophysectomized rats on LPL activity in adipose tissue (36) and the decrease in LPL activity observed after GH treatment of GH-deficient adults (35). Thus the reduced adipose tissue mass in acromegaly does not need to be explained by the decreased adipose tissue LPL activity (48). LPL activity in adipose tissue, in contrast to muscle LPL activity, has been shown to increase after insulin treatment both in vivo (36) and in vitro (11). Thus it is possible that the markedly increased serum insulin levels mediate the increased LPL activity in adipose tissue of bGH-transgenic mice.
We observed increased circulating levels of LDL cholesterol and LDL-apoB, although LDL receptor expression was unchanged. Hepatic lipase is of importance for hepatic LDL uptake and degradation (47). Recent data from a DNA microarray study investigating hepatic gene expression in bGH-transgenic mice indicated a marked reduction in hepatic lipase mRNA expression in these mice (unpublished data by B. Olsson, M. Bohlooly-Y, O. Brusched, O. G. P. Isaksson, S.-O. Olofsson, J. Oscarsson, and J. Törnell). Thus reduced hepatic lipase activity may contribute to the increased LDL levels in the bGH-transgenic mice. Increased production of LDL is less likely, because the hepatic triglyceride secretion rate was decreased, indicating decreased VLDL secretion. However, we cannot exclude the possibility that the number of apoB-containing lipoprotein particles secreted from the liver was unchanged, because apoB secretion was not measured. The reduced hepatic mRNA editing in the bGH-transgenic mice may have resulted in a larger proportion of apoB100-containing particles, which may have contributed to higher LDL levels (57). It is also possible that the increased LPL activity contributed to increased production of LDL in the bGH-transgenic mice (62). In acromegaly, the LDL levels are unchanged after therapy (59), but LDL cholesterol has also been shown to be higher in acromegalic patients than in controls (8). In line with the present results, apoB levels are unchanged after therapy of acromegaly (37).
HDL cholesterol (59) and apoA-I levels (37) have been shown to increase after therapy of acromegaly, indicating lower HDL levels in acromegaly. However, there are some studies that did not detect a difference in HDL levels between acromegalic patients and a control population (8). HDL cholesterol levels were higher in male transgenic mice than in their controls, but no difference was observed in female mice. We have previously observed that the higher HDL levels in female rats than in male rats were due to the more continuous GH secretion in female rats compared with male rats (33, 34). The secretion of GH is sexually dimorphic (25) and regulates sex differences in hepatic metabolism also in mice (30). Therefore, the effect of a continuous secretion of GH in the transgenic mice may affect the HDL levels to a larger extent in males than in females, because the secretion is more profoundly shifted from a pulsatile to a continuous secretion in male mice.
In conclusion, the present study shows that there are marked changes in
lipid and lipoprotein metabolism in mice overexpressing GH. However,
these changes differ in several respects from the effects of acromegaly
(29, 37, 48, 55, 59) on lipoprotein metabolism. In Table
4, the effects of overexpression of bGH in mice on certain aspects of lipoproteins are compared with the effects of acromegaly. The differences between acromegaly and bGH-transgenic mice may have several reasons. In humans, increased activity of cholesteryl ester transfer protein (55) and
absence of editing of apoB mRNA in the liver may contribute to
differences in HDL and LDL plus VLDL metabolism. Moreover, LPL is
differently regulated in acromegaly and in bGH-transgenic mice. The
tendency toward higher serum triglycerides in bGH-transgenic mice
compared with control mice after fat feeding indicates that the
different amount of dietary fat in laboratory chow and human diet may
at least partly contribute to the observed differences between
acromegaly and bGH-transgenic mice.
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ACKNOWLEDGEMENTS |
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We thank Birgitta Odén and Britt-Marie Larsson for excellent technical assistance.
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FOOTNOTES |
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This study was supported by grants from the Swedish Medical Research Council (8269), Novo Nordisk Foundation, Handledaren Hjalmar Svensson Foundation, Magnus Bergvall Foundation, Åke Wiberg Foundation, King Gustav V's and Queen Victoria's Foundation, The Swedish Medical Society, The Swedish Cancer Society, and AstraZeneca R&D, Molndal, Sweden.
Address for reprint requests and other correspondence: F. Frick, Dept. of Physiology, Endocrinology unit, Göteborg Univ., Box 434, S-405 30 Goteborg, Sweden (E-mail: Fredrik.Frick{at}fysiologi.gu.se).
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.
Received 5 March 2001; accepted in final form 17 July 2001.
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