Download IGF-1 induces rat glomerular mesangial cells to accumulate

Am J Physiol Renal Physiol 290: F138 –F147, 2006.
First published August 2, 2005; doi:10.1152/ajprenal.00054.2005.
IGF-1 induces rat glomerular mesangial cells to accumulate triglyceride
Anne K. Berfield,1 Alan Chait,2 John F. Oram,2
Richard A. Zager,3 Ali C. Johnson,3 and Christine K. Abrass1
1
Department of Medicine, Veterans Affairs Puget Sound Health Care System and University of Washington
School of Medicine, 2Division of Endocrinology and Metabolism, Department of Medicine, University of
Washington School of Medicine, and 3The Fred Hutchison Cancer Research Center, Seattle, Washington
Submitted 8 February 2005; accepted in final form 25 July 2005
chronic kidney disease; cholesterol; peroxisome proliferator-activated
receptor; foam cells
MESANGIAL CELLS (MC) ARE SPECIALIZED glomerular pericytes that
share many properties with vascular smooth muscle cells
(VSMC) (77) and macrophages (81). Like VSMC, MC provide
structural support through production of extracellular matrix
(2) and regulate glomerular blood flow and intracapillary
pressure via contraction (39). MC properties typical of macrophages include phagocytosis and production of reactive oxygen
species (7). Recent studies of progression of chronic kidney
disease indicate that, like atherosclerosis, lipid accumulation
can contribute to glomerular injury (53). In some cases, lipid
accumulates in MC, and they take on the appearance of foam
cells (46, 63). MC metabolize lipids similar to macrophages
and VSMC in that they express low-density lipoprotein (LDL)
and scavenger receptors (73, 74) and secrete increased amounts
of extracellular matrix (10) and monocyte chemoattractant
protein 1 (35, 52, 72) in response to exposure to oxidized LDL.
Under normal conditions, the uptake, metabolism, and efflux of
cholesterol and triglycerides (TG) are tightly controlled to meet
Address for reprint requests and other correspondence: C. K. Abrass,
Univ. of Washington School of Medicine and Veterans Affairs Puget Sound
Health Care System, 1660 South Columbian Way, Seattle, WA 98108 (e-mail:
[email protected]).
F138
the metabolic needs of the cell and to protect the cell from the
toxic effects of elevated levels of intracellular free cholesterol
(38, 74). Current work in atherosclerosis demonstrates that
inflammatory cytokines cause dysregulation of cellular lipid
metabolism, leading to foam cell formation (81). Recent data
suggest that dysregulation of lipid metabolism may accompany
immune-mediated glomerular diseases and thereby contribute
to glomerulosclerosis (45).
IGF-1 has been implicated as a cytokine that contributes to
the development of glomerulosclerosis (56). Mice that overexpress IGF-1 develop glomerulosclerosis (28), and MC from
diabetic mice that develop nephropathy secrete increased
amounts of IGF-1 (28). MC express IGF-1 receptors (3) and
synthesize IGF-1 in vitro (23). Exposure to IGF-1 induces an
increase in the synthesis of extracellular matrix, indicating that
IGF-1 might contribute to mesangial matrix accumulation in
glomerular disease (6). IGF-1 is thought to contribute to
VSMC proliferation in vessel walls and participate in lesion
progression in atherosclerosis (Ref. 76 and references therein).
In prior studies, we observed that rat MC exposed to IGF-1 for
several days become lipid laden, developing a foam cell
appearance (11). We demonstrated that lipid uptake was primarily through increased endocytosis and that lipid accumulation could be increased by supplementing the medium with
cholesterol ester (12). As lipid accumulated, the actin cytoskeleton became disordered, and the membrane could no longer
reorganize sufficiently to engulf Escherichia coli particles (11).
Furthermore, lipid-laden MC were unable to migrate in response to insulin-like growth factor binding protein-5 (IGFBP-5) or to contract in response to angiotensin II (12). These
studies indicate that IGF-1-mediated foam cell formation in
MC is associated with impairment of important MC functions.
The present studies were undertaken to characterize the lipid
content of MC exposed to IGF-1 and to define the mechanisms
responsible for IGF-1-induced lipid accumulation.
MATERIALS AND METHODS
Materials. The following reagents were purchased: human recombinant IGF-1 (Collaborative Research, Waltham, MA); IGFBP-5
peptide (AA201–218, RKGFYKRKQCKPSRGRKR, Fred Hutchinson Cancer Research Center, Seattle, WA); Lucifer yellow (Sigma, St.
Louis, MO); 125I (DuPont-New England Nuclear, Boston, MA); and
RPMI 1640 tissue culture medium (GIBCO, Grand Island, NY).
Experimental design. MC were cultured in routine medium (20%
FCS-RPMI 1640) with or without IGF-1 (100 nM) for 1–7 days. The
lipid content of this medium was (in mg/dl) 10 cholesterol, 2 high-
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.
http://www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
Berfield, Anne K., Alan Chait, John F. Oram, Richard A.
Zager, Ali C. Johnson, and Christine K. Abrass. IGF-1 induces rat
glomerular mesangial cells to accumulate triglyceride. Am J Physiol
Renal Physiol 289: F138 –F147, 2006. First published August 2, 2005;
doi:10.1152/ajprenal.00054.2005.—Rat glomerular mesangial cells
(MC) become lipid-laden foam cells when they are exposed to IGF-1.
IGF-1 increased accumulation of triglyceride (TG) 2.5-fold in MC
after 7 days. TG accumulation resulted from enhanced macropinocytosis and decreased efflux secondary to a 40 –50% reduction in
peroxisome proliferator-activated receptor (PPAR)-␦ (PPAR␦). There
was no evidence of primary or secondary changes in cholesterol or TG
synthesis, increased uptake by LDL or scavenger receptors, or reduced efflux via ATP-binding cassette A-1. Although the lipid moiety
taken up can be influenced by the concentration of cholesterol or TG
in the medium, in standard medium MC preferentially accumulate
TG. TG-rich MC foam cells fail to contract in response to angiotensin
II (Berfield AK, Andress DL, and Abrass CK. Kidney Int 62: 1229 –
1237, 2002); however, their migratory response to IGF binding
protein-5 is unaffected. This differs from cholesterol loading, which
impairs both phagocytosis and migration. These findings have important implications for understanding the mechanisms that contribute to
lipid accumulation in MC and the functional consequences of different forms of foam cells. These observations are relevant to understanding vascular disease and progressive renal diseases that are
accelerated by abnormalities in lipid metabolism.
IGF-1, LIPIDS, AND MESANGIAL CELLS
AJP-Renal Physiol • VOL
P. A. Edwards, UCLA, Los Angeles, CA); or rabbit PPAR␣, PPAR␥,
and PPAR␦ (Santa Cruz Biotechnology) followed by horseradish
peroxidase-labeled donkey anti-rabbit or mouse IgG (Amersham
Pharmacia, Piscataway, NJ) (41). Staining was enhanced with an ECL
chemiluminescence kit (Amersham Pharmacia). Negative controls
included the secondary antibody alone. Semiquantitative analysis was
by optical density scanning of the relevant bands (Kodak).
Endocytic assay. MCs were grown for 7 days with or without
IGF-1 in chamber slides, then incubated in media containing Lucifer
yellow dipotassium salt (1 ␮g/ml) for 1 h at 37°C and other slides at
4°C. The 4°C control was included to determine the degree of
nonspecific surface binding of antigen to the cells. Following incubation, cells were washed twice with PBS to remove excess Lucifer
yellow. Slides were examined by fluorescence microscopy.
Electron microscopy. MC were plated and treated as above, then
subsequently fixed in 2% glutaraldehyde for 2 h. Cells were treated
with 1% osmium tetroxide before sequentially dehydrated to Medcast.
Thin sections were examined with a JEOL S100 electron microscope.
MC migration. MC migration was measured with and without
IGFBP-5201-218 (30 ␮g/ml) in a wounding assay as described (12). In
brief, MC were plated in 60-mm tissue culture dishes, grown to near
confluence, growth arrested in 2% FCS-RPMI, and scraped with a
sterile razor blade to create a linear wound across the dish. IGFBP5201–218 peptide was added to the cultures, and migration was examined 48 h later. Controls were maintained in 2% FCS-RPMI medium.
Cultures were stained with toluidine blue. Migrating cells along a
1-mm length of the wound edge were counted in 0.1-mm increments.
Five areas were examined per sample. Experiments were repeated
three times. Data were analyzed as the total number of migrating cells
and were expressed as a percentage of control.
Calculations and statistics. Unless otherwise stated, results were
calculated on triplicate experiments, done on two to four separate
occasions. For variables, means and SE were computed. An unpaired
Student’s t-test was used to compare means between two experimental
groups, whereas for multiple comparisons ANOVA was used, followed
by Tukey’s test. Statistical significance was defined as P ⬍ 0.05.
RESULTS
Previously, we showed that MC cultured with IGF-1 for ⬎3
days develop morphological changes characteristic of foam
cells (11). Compared with controls, IGF-1-treated MC show
increased accumulation of intracellular neutral lipids as evidenced by Nile red staining. The presence of neutral lipid
droplets indicates accumulation of either cholesterol or TG. As
we previously showed that cholesterol uptake was enhanced in
IGF-1-treated MC when the medium was supplemented with
cholesterol, we expected that the foam cells that developed
after chronic exposure to IGF-1 in standard medium would be
cholesterol rich. Furthermore, because oxidized LDL accumulation is thought to be an important contributor to foam cell
formation, we postulated that IGF-1 might enhance uptake of
modified lipids. To address these possibilities, we systematically evaluated the potential contribution of cholesterol synthesis and uptake of LDL and oxidized LDL to IGF-1-induced
foam cell formation.
IGF-1 has modest effects on receptor-mediated cholesterol
uptake. Most cholesterol and some TG uptake occur via LDL
receptor-mediated endocytosis (38, 74). As shown in Fig. 1,
there were no differences in the specific binding of LDL in the
first day in untreated compared with IGF-1-treated MC. After
3 and 7 days of exposure to IGF-1, there was a minimal
reduction (25%) in LDL binding. This suggests that IGF-1 had
no direct effect on LDL receptor expression and that the
modest fall in LDL binding at days 3 and 7 may have occurred
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
density lipoprotein (HDL), 5 LDL, 15 TG, and 8 phospholipids.
Medium was changed every 2–3 days. After designated times, MC
were assessed for cholesterol and TG content; LDL and acetylated
LDL (AcLDL) binding and degradation; and levels of protein expression of sterol-regulatory element binding protein-1c (SREBP-1c), the
peroxisome proliferator-activated receptors (PPAR), PPAR␣, PPAR␥
and PPAR␦, scavenger receptor (SR) SR-B1, CD36, ATP-binding
cassette A-1 (ABCA-1), and 3-hydroxy-3-methylglutaryl (HMG)CoA reductase. Macroendocytosis was visualized with Lucifer yellow
staining and electron microscopic examination. To measure the effects
of lipid accumulation on IGFBP-5-mediated migration, MC propagated with and without IGF-1 (100 nM) were incubated in standard
medium or loaded with cholesteryl ester (2.5 ␮g/ml) or linoleic and
oleic acids conjugated to albumin (9.3 ␮g/ml, Sigma) for 7 days.
Migration was measured as described elsewhere (12).
Cell culture. Rat glomerular MC were prepared by modification (4,
5) of routine methods (47). In brief, minced rat kidney cortex was
sieved. Isolated glomeruli were plated in medium containing a 1:1 mix
of 20% FCS-RPMI 1640 and previously collected glomerular conditioned medium. Insulin routinely added to supplement MC cultures
was omitted. MC outgrowths were harvested and passed in this
medium for an additional week, after which the conditioned medium
was omitted. MC were cloned and studied at passages 8 –12.
Cholesterol and TG analysis. MC were plated (5 ⫻ 104 cells/
35-mm dish) and grown for 7 days with and without IGF-1 (100 nM).
Lipids were extracted in hexane/isopropyl alcohol (3:2) or chloroform-methanol and isopropanol (13). Cholesterol and cholesterol ester
were measured by mass spectrometer (PerkinElmer) as previously
described (9). TG were assayed with a G GPO trinder kit (Sigma)
using the manufacturer’s instructions and a modification of McGowan
(61). Protein was measured by the method of Lowry (55). Cell counts
were performed using a Coulter counter; n ⫽ 3– 4/condition and
experiments were repeated on four separate occasions.
LDL and AcLDL binding and degradation. LDL was prepared from
human plasma as described previously (9). LDL was acetylated by
repeated additions of acetic anhydride to LDL (10 mg/ml) diluted with
saturated ammonium acetate. LDL and AcLDL were radiolabeled by
the iodine monochloride method as modified for lipoproteins as
described (43). MC (3 ⫻ 106 cells/75-cm2 flask) were cultured for 1,
3, or 7 days with and without IGF-1 (100 nM). To measure LDL and
AcLDL binding, cells were washed in warm RPMI 1640 tissue culture
medium and 0.5 mg/ml fatty acid-free albumin, chilled, and incubated
on ice for 3 h with 125I-LDL or 125I-AcLDL [250 cpm (counts/min)/
ng] in the presence and absence of unlabeled lipoproteins. The cells
were washed with PBS, dissolved in 0.1 N NaOH, and bound counts
were determined. For measurements of lipoprotein degradation, cultures were prepared as above. After 125I-LDL and 125I-AcLDL binding, replicate samples were washed and warmed to 37°C. After
incubation for 4 h, the supernatants were collected, precipitated with
trichloroacetic acid, and measured in a gamma counter. Cell-free
degradation, which is ⬍5% of total radioactivity, is subtracted from total
degradation (43). Protein was measured by the method of Lowry (55).
Western blot analysis. MC (5 ⫻ 106 cells/75-cm2 flask) were
extracted using Triton-glycerol lysis buffer (1% Triton X-100, 10%
glycerol, 20 mM HEPES, 100 mM NaCl) containing protease inhibitors (Complete Protease Inhibitor Cocktail tablets, Roche, Indianapolis, IN). The concentrated samples were assayed for total protein by
BCA (Pierce, Rockford, IL). Samples (10 –20 ␮g) were subjected to
electrophoresis using 10% (HMG-CoA), 12% (SR-B1), or 4 –12%
(ABCA-1) SDS precast gels (Bio-Rad, Richmond, CA). PPARs were
immunoprecipitated from cell lysates with respective antibodies and
subjected to electrophoresis on 7% SDS gel. Proteins were transferred
to nitrocellulose and incubated with the following antibodies: monoclonal mouse anti-mouse CD36 (MAB1258, Chemicon International,
Temecula, CA); rabbit anti-SR-B1, mouse anti-SREBP1c (Santa Cruz
Biotechnology, Santa Cruz, CA); rabbit anti-ABCA-1 (Novus Biologicals, Littleton, CO); rabbit anti-HMG-CoA reductase (gift from
F139
F140
IGF-1, LIPIDS, AND MESANGIAL CELLS
as a result of downregulation by the intracellular accumulation
of lipid. As shown in Fig. 1B, 1 day of exposure to IGF-1 was
associated with a very modest increase in the amount of LDL
internalized and degraded by MC. This increase was not
sustained with longer exposure to IGF-1; thus changes in the
rates of uptake and metabolism of LDL do not account for our
findings.
Altered foam cell function has traditionally been attributed
to intracellular accumulation of cholesterol and oxidized LDL
by heightened scavenger receptor action. Scavenger receptor
expression was assessed by measuring receptor binding, internalization, and degradation of AcLDL (9). As shown in Fig.
1C, no significant differences in AcLDL binding were detected
over a period of 7 days. Similarly, in Fig. 1D, although a slight
decrease in degradation occurred the first day, this was not
sustained. This is in agreement with our previous data in which
we found no change in the uptake of fluorescent-labeled
AcLDL (11). These data indicate that IGF-1 did not significantly influence uptake of lipids by scavenger receptors, but it
does not exclude the possibility that IGF-1 modulates other
scavenger receptors, including SR-B1 and CD36 (67).
IGF-1 has little effect on intracellular synthesis, transfer, or
efflux of cholesterol. MC express HMG-CoA reductase, which
regulates intracellular synthesis of cholesterol (1). In most
cells, intracellular accumulation of cholesterol is followed by a
AJP-Renal Physiol • VOL
decrease in expression of HMG-CoA reductase (74), which
permits cholesterol levels to return to baseline. As shown in
Fig. 2, HMG-CoA reductase protein content was unchanged in
MC after 7 days of exposure to IGF-1; thus there is no evidence
for enhanced cholesterol synthesis.
SR-B1, the HDL receptor, serves as both a scavenger receptor for uptake of oxidized LDL, uptake of cholesterol and
cholesterol ester from HDL, and as a passive cholesterol efflux
mechanism by transferring cholesterol to HDL (22, 69). There
were no differences in expression of SR-B1 in IGF-1-treated
MC to account for the lipid accumulation in these cells (Fig. 2).
This correlates with the results of the scavenger receptor
and suggests that there is little lateral transfer of cholesterol
to HDL.
A second scavenger receptor, CD36, has received considerable attention because of its role in uptake of oxidized LDL and
long-chain fatty acids, its regulation in inflammatory states,
and demonstration that null mutations in this receptor protect
against the development of atherosclerosis (62). As shown in
Fig. 2, chronic IGF-1 was associated with a marked decrease in
CD36 content in MC. Such a prominent decrease in CD36
expression would be expected to reduce lipid accumulation and
foam cell formation. Given the development of foam cells in
the face of a decrease in CD36 expression, it argues that IGF-1
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
Fig. 1. LDL or acetylated LDL (AcLDL) binding and degradation. For the binding assay, I125-labeled LDL (A) and AcLDL (C) bound to untreated control and
IGF-1-treated mesangial cells (MC) were determined. For the degradation assay, cultures were incubated for 4 h, and the released isotope in the supernatant was
counted LDL (B) and AcLDL (D). Values are means ⫾ SE expressed as ng/␮g cell protein. *P ⬍ 0.05, ANOVA.
F141
IGF-1, LIPIDS, AND MESANGIAL CELLS
mediates lipid accumulation in MC by mechanisms that do not
involve CD36.
ABCA-1, expressed in the plasma membrane and Golgi,
mediates apo-A1-associated cholesterol efflux from cells (79).
Its expression is influenced by both SREBP activation and
PPAR␥. Expression in MC has been variably detected, and its
expression is low compared with expression in other cell types
(71, 79). Under cholesterol loading of normal cells, expression
of this efflux pathway is increased. Some studies have suggested that it is particularly important for the efflux of TG.
Similar to other reports, we found that ABCA-1 expression in
MC is very low, and furthermore, IGF-1 had no affect on its
expression (Fig. 2). Failure of ABCA-1 to increase in the face
of elevated intracellular lipids may contribute to lipid accumulation in our cells; yet, the very low expression of ABCA1 in
MC suggests that other undefined transport mechanisms might
be more important.
In summary, these studies fail to show a mechanism for
IGF-1-induced intracellular cholesterol accumulation other
than the enhanced nonreceptor endocytosis in cholesterolsupplemented medium that we showed previously (12). In the
absence of an explanation for cholesterol accumulation in
IGF-1-treated MC grown in routine medium, we specifically
measured both cholesterol and TG content in control and
IGF-1-treated MC to define the neutral lipid identified by Nile
red staining.
Table 1. Cholesterol and triglyceride content of IGF-1-treated mesangial cells
Cholesterol Content
Control
IGF-1
TG Content
␮g/sample*
␮g/mg protein
pg/cell
␮g/sample*
␮g/mg protein*
pg/cell*
8.4⫾1.6
12.7⫾2.5*
22.4⫾4.9
21.3⫾3.9
3.6⫾1.3
4.5⫾1.4
2.9⫾1.8
18.9⫾7.2*
14.8⫾1.6
44.2⫾11.6*
2.1⫾1.7
5.3⫾2.5*
Values are means ⫾ SD. MC, mesengral cells; TG, triglyceride. TG cholesterol and levels were assessed by mass spectrometry and a triglyceride kit.
*P ⬍ 0.05, ANOVA.
AJP-Renal Physiol • VOL
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
Fig. 2. Cholesterol synthesis, transfer, and efflux. Expression of 3-hydroxy3-methylglutaryl (HMG)-CoA reductase, scavenger receptor (SR-B1), ATPbinding cassette A-1 (ABCA1), and CD36 protein was determined by Western
blot analysis in triplicate samples of MC untreated (black bars) or treated with
IGF-1 (grey bars) for 7 days. Densitometric analysis of bands is expressed in
arbitrary units, means ⫾ SD, and calculated as a percentage of control. Inset:
representative blots. P ⬎ 0.05, except for CD36, which is P ⬍ 0.01, by t-test
(n ⫽ 3–5).
The primary lipid that accumulates in IGF-1-treated MC is
TG. As shown in Table 1, IGF-1 treatment was associated with
a modest increase in the total cholesterol measured per sample;
however, this difference disappears when the cholesterol measurement for cellular protein or cell number is factored in. In
contrast to the measurement of cholesterol, TG content was
significantly increased even when the measurements were
expressed per cell. These results indicate that in routine medium (in mg/dl: 10 cholesterol, 2 HDL, 5 LDL, 15 TG, 8
phospholipids), IGF-1-treated MC preferentially accumulate
TG. This is a novel and unexpected finding.
For many years, foam cell formation has been attributed to
intracellular accumulation of cholesterol and oxidized LDL
(33). Recent studies have implicated TG accumulation in foam
cell formation and implicated TG in some of the aberrant
functions of foam cells (8, 34); yet, little is known about the
factors that lead to TG accumulation, the subsequent change in
lipid metabolism, or the mechanisms that mediate changes for
cell function.
TG uptake is enhanced by macropinocytosis. Fluid and
small, soluble antigens can be taken up by micro (up to ⬃0.1
␮m)- and macrocytosis (from 0.5 to 3 ␮m). We previously
showed that IGF-1 induces an increase in fluid-phase endocytosis (12). By uptake of low-molecular-weight FITC-dextran
and electron microscopy, we revealed increased internalization
by microendocytosis via caveoli and coated pits. Accumulation
of Lucifer yellow has been shown to be a marker of macroendocytosis (66), and now in Fig. 3A we show that IGF-1 also
increases lipid uptake by macroendocytosis. This is verified by
the electron micrograph in Fig. 3B showing not only caveoli
and endosomes of micropinocytosis but also, more specifically,
many ruffles of macropinosomes. The circular ruffles of macroendosomes are heterogeneous in size, closed by purse-string
movement, nonselectively enclosing bulk-fluid and macromolecules. These distinctive features of IGF-1-treated MC have
not been previously described. These data indicate that IGF-1
induces lipid uptake by both micro- and macroendocytosis.
Intracellular mechanisms of IGF-1-treated MC show that
TG accumulation occurs not by increased biosynthesis but by
decreased efflux. SREBP-1 plays an important role in TG
biosynthesis, whereas SREBP-2 mediates cholesterol synthesis. As shown in Fig. 4, there was no change in the expression
of SREBP-1 in IGF-1-treated MC to account for TG accumulation in these cells, reinforcing the concept that external lipids
are endocytosed.
PPAR are nuclear hormone ligand-activated transcription
factors, which play important roles in cholesterol and TG
efflux through regulation of ABCA-1 activity and other less
well-defined transporters (50). PPAR␣ is primarily found in
the liver and promotes fatty acid oxidation to generate energy
F142
IGF-1, LIPIDS, AND MESANGIAL CELLS
for peripheral tissues (36). PPAR␥ potentiates adipocyte differentiation and modulates lipid storage and glucose homeostasis (36, 50). PPAR␦ activates TG uptake and efflux, has
recently shown to regulate and mediate VLDL signaling in
macrophages (19, 50), and exerts both pro- and anti-inflammatory activity (49). As shown in Fig. 5, MC express protein
for all three PPAR isoforms. By density analysis, we found that
although exposure of MC to IGF-1 for 7 days had no affect on
expression of PPAR␣ or ␥, expression of PPAR␦ was markedly reduced. These findings have been repeated in triplicate
on three separate occasions, with similar reductions in PPAR␦
expression. Given that PPAR␦ is the major determinant of
long-chain fatty acid and TG efflux, it is likely that this change
is a significant contributor to TG accumulation. This is a new
and novel observation.
Differences in intracellular lipid composition influence MC
function. In prior studies, we showed that MC migratory
response to IGFBP-5 is affected by treatment with IGF-1,
particularly following the addition of cholesteryl esters to the
culture medium. At that time, we presumed that the absence of
an effect without supplementation of the medium reflected a
dose effect of cholesterol. In light of our findings described
above showing that the lipid that accumulates in IGF-1-treated
MC grown in standard medium is TG, we wondered what the
AJP-Renal Physiol • VOL
effects of additional TG loading would be. In Table 2, we
confirmed prior studies showing that chronic exposure to
IGF-1 alone has no effect on IGFBP-5-mediated MC migration
and that supplementation with cholesteryl esters blunts the
migratory response. Consistent with our present data showing
that MC exposed to IGF-1 in standard medium accumulate TG
and have a normal migratory response, MC that were TG
loaded by the addition of TG to the medium also had a normal
migratory response. Phagocytosis and endocytosis, which were
impaired by chronic IGF-1 treatment, were still impaired with
supplemental TG. These data indicate that the nature of the
lipid that accumulates determines the consequences to specific
MC functions. In this case, cholesterol, but not TG, accumulation interferes with IGFBP-5-mediated migration. The data in
Table 2 summarize the different functional modification of
supplemental lipids in growth media and reaffirm that IGF-1treated MC are TG loaded.
DISCUSSION
IGF-1-treated MC become lipid-laden foam cells, which are
unable to phagocytose or contract in response to physiological
stimuli (11, 12). Lipid uptake occurs through IGF-1-mediated
enhancement of endocytosis and is not associated with in-
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
Fig. 3. Triglyceride (TG) uptake by macroendocytosis. A: Lucifer yellow staining of untreated MC (a) and IGF-1-treated MC (b).
Bar ⫽ 10 ␮m. B: electron micrograph of endocytic vesicles (arrow) and macrocytotic ruffle extensions (arrowheads). Bar ⫽ 1 ␮m.
F143
IGF-1, LIPIDS, AND MESANGIAL CELLS
creases in surface concentration of lipid transporters. The type
of lipid that accumulates can be influenced by the concentration of specific lipid moieties in the medium; however, TG
preferentially accumulates in IGF-1-treated MC grown in standard serum-containing medium. We noted a 40% reduction in
expression of PPAR␦; thus TG may accumulate as a result of
failure of PPAR␦-dependent downregulation of the VLDL
receptor and/or reduced TG efflux (19, 49). The role of TG in
foam cell formation, the mechanisms of TG accumulation, the
influence of TG on cellular lipid metabolism, and the functional consequences of TG to foam cells are just beginning to
be examined (70). Data presented in this study complement our
previous studies and show that the composition of the lipid that
accumulates determines the effect on MC function (Table 2).
Cholesterol accumulation interferes with phagocytosis, contraction, and IGFBP-5-mediated migration, whereas TG accumulation alters contraction but has no affect on migration.
Considerable data show that intracellular cholesterol content
is tightly regulated and under normal circumstances synthesis
and uptake are appropriately modulated to maintain stable
cholesterol content. Current concepts of the pathogenesis of
atherosclerosis (30) and progressive renal disease (74) indicate
that inflammatory cytokines disrupt the feedback control mechanisms that maintain normal intracellular cholesterol levels,
thereby allowing for intracellular lipid accumulation and foam
cell formation. Oxidized lipoproteins have a propensity to
accumulate in vessel walls through binding to matricellular
proteins (14, 18). Uptake of oxidized lipids stimulates macrophages (25, 59) and MC (15, 60) to release inflammatory
cytokines, which perpetuate progression of the vascular disease
and extracellular matrix accumulation (51). Until recently, TG
were thought to play little role in this process; however, there
is growing evidence that TG also contribute to the pathogenesis
of vascular and renal diseases (34, 57). Additional studies are
needed to define the mechanisms whereby TG accumulation
interferes with MC function.
Little is known about changes that occur in lipid metabolism
in the presence of TG accumulation. In macrophages, VSMC,
AJP-Renal Physiol • VOL
Fig. 5. Peroxisome proliferator-activated receptor-␣ (PPAR␣; A), PPAR␥ (B),
and PPAR␦ (C) protein expression. PPAR protein expression was determined
by Western blot analysis of MC untreated (black bars) or treated with IGF-1
(grey bars) for 7 days. Densitometry analysis of the bands is expressed in
arbitrary units, means ⫾ SD. Inset: representative blots. *P ⬍ 0.05 by t-test
(n ⫽ 3).
and MC, cytokines such as TNF-␣ and IL-1␤ dysregulate
expression of LDL and scavenger receptors, which contribute
to lipid accumulation. In the present studies, we found no
significant differences in receptor binding or degradation of
LDL or AcLDL in MC chronically exposed to IGF-1. These
observations are consistent with the unchanged levels of cholesterol and indicate that neither IGF-1 nor intracellular TG
levels directly influence expression of these receptors. As we
previously showed that IGF-1 enhances endocytosis and that
cholesteryl esters can be taken up by these mechanisms and
accumulate when the medium is supplemented with cholesteryl
ester, it is possible that cholesterol metabolism is abnormal in
that setting. Additional studies would be required to determine
the role of IGF-1 in dysregulation of cholesterol homeostasis in
the cholesterol-loaded MC.
In the present studies, we show that HMG-CoA reductase
levels were equal in untreated and IGF-1-treated MC. Not only
does this correspond to the normal levels of cholesterol in the
cell, but it also indicates that neither IGF-1 nor TG modulate
HMG-CoA reductase levels in MC. CD36 and SR-B1/CLA-1
are members of a family of receptors that influence lipid
transport. SR-B1 receptors contribute to VLDL uptake; however, their major role is in cholesterol efflux, which is influenced by the external concentration of HDL (40). As prior
studies had shown that IGF-1 downregulates expression of this
Table 2. Functions of TG- and cholesterol-loaded MC
Phagocytosis
Migration
Contraction
Cholesterol content
TG content
Control
IGF-1
IGF-1⫹CE
IGF-1⫹TG
Normal
Normal
Normal
Normal
Normal
Impaired
Normal
Impaired
Normal
Elevated
Impaired
Impaired
Impaired
Elevated
Elevated
Impaired
Normal
Impaired
Normal
Elevated
The table provides a summary of data obtained from various experiments
provided herein, as well as those published previously (12). CE, cholesteryl
esters. These studies indicate that cellular migration is most impaired by
changes in intracellular cholesterol content, whereas the contractile response to
angiotensin II is affected by TG accumulation.
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
Fig. 4. Intracellular synthesis of TG. Sterol-regulatory element binding protein-1c (SREBP-1c) protein expression was determined by Western blot
analysis of MC untreated or treated with IGF-1 for 7 days. Densitometry
analysis of bands is expressed in arbitrary units, means ⫾ SD. Inset: representative blots. P ⬎ 0.05 by t-test (n ⫽ 3).
F144
IGF-1, LIPIDS, AND MESANGIAL CELLS
AJP-Renal Physiol • VOL
through activation of the ABCA-1 transporter (20). We found
no evidence through measurements of PPAR␣, PPAR␥, or
ABCA-1 that they played a role in the TG accumulation in
IGF-1-treated MC. As ABCA-1 activity is very low in MC,
other transporters may be more important to lipid efflux in MC.
In a recent report, Davies et al. (24) demonstrated that adipogenic conditions induce VSMC to enhance TG synthesis,
leading to lipid accumulation in intracellular vacuoles as a
result of liver X receptor (LXR)/SREBP1c activation. As
VSMC in atherosclerotic plaques lack a scavenger receptor
phenotype but express adipocyte markers (e.g., fatty acid
synthase, SREBP-1, LXR␣, and adipsin), this would suggest
that they become foam cells by mechanisms that differ from
those of macrophage foam cells. Similar to their reports, we
found no evidence that IGF-1-treated MC primarily developed
lipid or cholesterol accumulation via enhanced expression and
activity of LDL or scavenger receptors. Although we did not
examine LXR␣ function directly, we did not find an increase in
the expression of SREBP1c or ABCA1 as evidence of LXR␣
activity (24, 82); thus IGF-1 might mediate foam cell formation by yet another mechanism. Although the effects of PPAR␣
and PPAR␥ involve LXR␣ in the development of macrophage
foam cells, PPAR␦ does not (54). The reduced expression of
PPAR␦ may contribute to TG accumulation because of reduced
fatty acid oxidation, as occurs in cardiac muscle of animals
with targeted deletion of PPAR␦ (21). Furthermore, IGF-1
increases gene expression for fatty acid synthase in adipose
tissue (78). Effects in smooth muscle cells have not been
examined.
PPAR␦ is ubiquitously expressed but is most abundant in
muscles, which rely heavily on fatty acids for energy (50).
PPAR␦ functions primarily as a sensor for TG in the VLDL
particle and to promote fatty acid oxidation (19, 49). TG
participates in macrophage differentiation by PPAR␦-dependent induction of transcription of adipocyte differentiationrelated protein (19). In PPAR␦ null cells, the VLDL receptor
gene is induced, indicating that activation of PPAR␦ reduces
TG uptake by this receptor. Furthermore, TG loading downregulates the VLDL receptor in a PPAR␦-dependent fashion
(19). Thus our findings that PPAR␦ expression is significantly
decreased in IGF-1-treated MC indicate that TG accumulation
may have occurred as a result of impaired suppression of
VLDL receptor expression and reduced TG efflux. These
alterations in PPAR␦ expression are particularly interesting
given the known increased expression of IGF-1 in MC from
animals with type 2 diabetes (17, 28), the elevated levels of TG
that are typical of dyslipidemia in diabetes, and recent studies
showing the importance of this receptor in mediating insulin
resistance (80). Furthermore, two fatty acids common to TG,
oleate and linolate, enhance the growth-promoting effects of
IGF-1 in SMC (8), which have been implicated in vascular (76)
and renal disease progression (27, 57). Additional studies are
needed to define the mechanisms responsible for IGF-1-mediated alterations in TG metabolism and PPAR␦-mediated effects.
Considerable evidence implicates lipids in the progression of
renal diseases, including diabetic nephropathy (reviewed in
Refs. 44 and 1). Hypercholesterolemia accelerates the rate of
progression of kidney disease and leads to macrophage infiltration and foam cell formation in rats (37). Genetic abnormal-
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
receptor in fibroblasts (65) and HepG2 cells (16), we expected
it to be reduced. No changes in protein content of this receptor
were observed; however, we examined it after 7 days, and
studies in fibroblasts and hepatic cells were done after shortterm exposure to IGF-1. Additional studies would be needed to
determine the appropriate expression of this receptor in TGloaded cells; however, our results provide no evidence for
increased expression of these receptors as a mechanism for TG
accumulation in IGF-1-treated MC. We found that IGF-1
markedly decreased the protein content of CD36 in MC, a
finding that has not been reported previously. CD36 was
originally identified as a platelet membrane glycoprotein and as
a receptor for thrombospondin 1, and it is the primary receptor
that mediates uptake of oxidized LDL and long-chain fatty
acids (reviewed in Refs. 22 and 62). Uptake of oxidized LDL
by CD36 leads to induction of CD36 and further lipid uptake,
thereby aggravating macrophage foam cell formation. Null
mutations in CD36 actually protect against the development of
atherosclerosis; thus it is of note that we found such a marked
decrease in expression of this receptor in IGF-1-treated MC.
Although additional studies are needed to understand the importance of this observation in the net effects on lipid metabolism in vivo, they exclude increased CD36 activity as a
contributor to lipid accumulation in our studies.
In our previous studies and the ones described herein, we
found that IGF-1 increased both micro- and macropinocytosis
(11, 12), processes that have been implicated in foam cell
formation (48). Micropinocytosis can occur in clathrin-associated, caveolin-associated, or other vesicles, which involve
lipid-raft domains of the plasma membrane, whereas macropinocytosis is an actin-dependent process that involves larger
regions of the cell membrane (48). Caveolin-1-rich caveolae
can influence lipid uptake by fluid-phase endocytosis/transcytosis and receptor-mediated endocytosis, as in the case of
CD36 (31). Lipid rafts lacking caveolin-1 also participate in
long-chain fatty acid uptake, as many members of the fatty acid
translocases, including SR-B1 and CD36, can attach to the rafts
directly (68). Although caveolin-1 enhances CD36 function,
the influence of caveolin-1 on SR-B1 function in endothelial
cells and macrophages is less clear as studies have produced
variable results (31). The role of caveolae in lipid handling by
smooth muscle cells has been less well studied, but caveolin-1
appears to play little or no role in lipid uptake in these cells
(31). Similar to vascular smooth muscle cells, caveolin-1
suppresses MC proliferation (32), but no studies have evaluated the role of caveolin-1 in MC handling of lipids or the
effects of IGF-1 on caveolar-dependent lipid uptake. Understanding the pinocytotic uptake of lipids is complicated, as
different lipid moieties are processed differently by the rafts,
with variable contributions by caveolin-1, and there are differences in different cell types and in the response to different
stimuli and with different translocases. Additional studies are
needed to specifically define the role of caveolin-1 and lipid
rafts in mediating IGF-1-induced changes in cholesterol and
TG uptake in MC.
PPARs are fatty acid receptors that play important roles in
fatty acid catabolism, lipid storage, adipocyte differentiation,
inflammation, and glucose homeostasis (50). PPAR␣ stimulates hepatic fatty acid oxidation and ketogenesis. PPAR␥
responds to oxidized LDL and promotes cholesterol efflux
IGF-1, LIPIDS, AND MESANGIAL CELLS
GRANTS
These studies were supported by the Medical Research Service of the
Department of Veterans Affairs (C. K. Abrass) and the National Institute of
Diabetes and Digestive and Kidney Diseases (R01-DK-9771– 05 to C. K.
Abrass; DK-02456 to A. Chait and J. F. Oram; R37-DK-38432 to R. A. Zager).
REFERENCES
1. Abrass CK. Cellular lipid metabolism and the role of lipids in progressive
renal disease. Am J Nephrol 24: 46 –53, 2004.
2. Abrass CK, Peterson CV, and Raugi GJ. Phenotypic expression of
collagen types in mesangial matrix of diabetic and nondiabetic rats.
Diabetes 37: 1695–1702, 1988.
3. Abrass CK, Raugi GJ, Gabourel LS, and Lovett DH. Insulin and
insulin-like growth factor I binding to cultured rat glomerular mesangial
cells. Endocrinology 123: 2432–2439, 1988.
4. Abrass CK, Spicer D, and Raugi GJ. Insulin induces a change in
extracellular matrix glycoproteins synthesized by rat mesangial cells in
culture. Kidney Int 46: 613– 620, 1994.
5. Abrass CK, Spicer D, and Raugi GJ. Induction of nodular sclerosis by
insulin in rat mesangial cells in vitro: studies of collagen. Kidney Int 47:
25–37, 1995.
6. Abrass CK, Zawadzki I, and Raugi GJ. Insulin(I), insulin-like growth
factor I (IGF-I) and growth hormone (GH) treatment of cultured rat
mesangial cells (RMC) is associated with changes in mRNA expression
for collagen I, III & IV (Abstract). J Am Soc Nephrol 2: 570, 1991.
AJP-Renal Physiol • VOL
7. Akiba S, Chiba M, Mukaida Y, and Sato T. Involvement of reactive
oxygen species and SP-1 in fibronectin production by oxidized LDL.
Biochem Biophys Res Commun 310: 491– 497, 2003.
8. Askari B, Carroll MA, Capparelli M, Kramer F, Gerrity RG, and
Bornfeldt KE. Oleate and linoleate enhance the growth-promoting effects
of insulin-like growth factor-I through a phospholipase D-dependent
pathway in arterial smooth muscle cells. J Biol Chem 277: 36338 –36344,
2002.
9. Aviram M, Bierman EL, and Chait A. Modification of low density
lipoprotein by lipoprotein lipase or hepatic lipase induces enhanced uptake
and cholesterol accumulation in cells. J Biol Chem 263: 15416 –15422,
1988.
10. Bayes-Genis A, Conover CA, and Schwartz RS. The insulin-like growth
factor axis: a review of atherosclerosis and restenosis. Circ Res 86:
125–130, 2000.
11. Berfield AK and Abrass CK. IGF-1 induces foam cell formation in rat
glomerular mesangial cells. J Histochem Cytochem 50: 395– 403, 2002.
12. Berfield AK, Andress DL, and Abrass CK. IGF-1-induced lipid accumulation impairs mesangial cell migration and contractile function. Kidney Int 62: 1229 –1237, 2002.
13. Bligh EG and Dyer WJ. A rapid method of total lipid extraction and
purification. Can J Med Sci 37: 911–917, 1959.
14. Borén J, Olin K, Lee I, Chait A, Wight TN, and Innerarity TL.
Identification of the principal proteoglycan-binding site in LDL. A single
point mutation in apo-B100 severely affects proteoglycan interaction
without affecting LDL receptor binding. J Clin Invest 101: 2658 –2664,
1998.
15. Bruneval P, Bariety J, Belair MF, Mandet C, Heudes D, and Nicoletti
A. Mesangial expansion associated with glomerular endothelial cell activation and macrophage recruitment is developing in hyperlipidemic apoE
null mice. Nephrol Dial Transplant 17: 2099 –2107, 2002.
16. Cao WM, Murao K, Imachi H, Yu X, Dobashi H, Yoshida K,
Muraoka T, Kotsuna N, Nagao S, Wong NCW, and Ishida T. Insulinlike growth factor-I regulation of hepatic scavenger receptor class BI.
Endocrinology 145: 5540 –5547, 2004.
17. Chan W, Wang M, Martin RJ, Trachtman H, Hisano S, and Chan JC.
mRNA expression for insulin-like growth factor 1, receptors of growth
hormone and IGF-1 and transforming growth factor-beta in the kidney and
liver of Zucker rats. Nutr Res 21: 1015–1023, 2001.
18. Chang MY, Potter-Perigo S, Wight TN, and Chait A. Oxidized LDL
bind to nonproteoglycan components of smooth muscle extracellular
matrices. J Lipid Res 42: 824 – 833, 2001.
19. Chawla A, Lee CH, Barak Y, He W, Rosenfeld J, Liao D, Han J, Kang
H, and Evans RM. PPAR␦ is a very low-density lipoprotein sensor in
macrophages. Proc Natl Acad Sci USA 100: 1268 –1273, 2003.
20. Chawla A, Repa JJ, Evans RM, and Mangelsdorf DJ. Nuclear receptors
and lipid physiology: opening the X-files. Science 294: 1866 –1870, 2001.
21. Cheng L, Ding G, Qin Q, Huang Y, Lewis W, He N, Evans RM,
Schneider MD, Brako FA, Xiao Y, Chen YE, and Yang Q. Cardiomyocyte-restricted peroxisome proliferator-activated receptor-delta deletion perturbs myocardial fatty acid oxidation and leads to cardiomyopathy.
Nat Med 10: 1245–1250, 2004.
22. Connelly MA and Williams DL. Scavenger receptor BI: a scavenger
receptor with a mission to transport high density lipoprotein lipids. Curr
Opin Lipidol 15: 287–295, 2004.
23. Conti F, Striker L, Elliot S, Andreani D, and Striker GE. Synthesis and
release of insulin-like growth factor-I by mesangial cells in culture. Am J
Physiol Renal Fluid Electrolyte Physiol 255: F1214 –F1219, 1988.
24. Davies JD, Carpenter KLH, Challis IR, Figg NL, McNair R, Proudfoot
D, Weissberg PL, and Shanahan CM. Adipocytic differentiation and liver
X receptor pathways regulate the accumulation of triacylglycerols in human
vascular smooth muscle cells. J Biol Chem 280: 3911–3919, 2005.
25. De Villiers WJ and Smart EJ. Macrophage scavenger receptors and
foam cell formation. J Leukoc Biol 66: 740 –746, 1999.
26. Doi T, Striker LJ, Gibson CC, Agodoa LYC, Brinster RL, and Striker
GE. Glomerular lesions in mice transgenic for growth hormone and
insulin-like growth factor-I. I. Relationship between increased glomerular
size and mesangial sclerosis. Am J Pathol 137: 541–552, 1990.
27. Dominquez JH, Tang N, Xu W, Evan AP, Siakotos AN, Agarwal R,
Walsh J, Deeg M, Pratt JH, March KL, Monnier VM, Weiss MF,
Baynes JW, and Peterson R. Studies of renal injury III: Lipid-induced
nephropathy in type II diabetes. Kidney Int 57: 92–104, 2000.
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
ities in VLDL handling and apoE lead to MC foam cell
formation and progressive glomerulosclerosis (75). Obese
Zucker rats have significant glomerulosclerosis, which is ameliorated by correction of hypertriglyceridemia (42). In nephrotic syndrome, reduced VLDL clearance enhances interstitial injury (64). The potential role of IGF-1 in glomerulosclerosis and progression of diabetic nephropathy is well
established (17, 26). Given the increased expression of IGF-1
by MC in diabetes mellitus (28), and the TG-rich plasma
typical of uncontrolled diabetes mellitus, our findings have
important implications for understanding the pathogenesis of
diabetic nephropathy. Definition of the mechanisms of lipid
accumulation in MC may lead to interventions that could
modify the outcome of diseases such as diabetic nephropathy
and focal and segmental glomerulosclerosis in which mesangial foam cells are prominent (58, 63).
In summary, IGF-1 leads to an increase in MC lipid accumulation primarily as a result of increased endocytosis (11,
12). The lipid moiety that accumulates, in part, is dependent on
the concentration of lipids in the medium. When the medium is
not supplemented with specific lipids, IGF-1-treated MC preferentially accumulate TG. Our findings that these MC have
reduced expression of PPAR␦ suggest that chronic exposure to
IGF-1 might limit the normal TG-mediated downregulation of
the VLDL receptor and reduce TG efflux, both of which would
favor TG accumulation. The specificity of these effects was
further supported by the absence of alterations in HMG-CoA
reductase, LDL, or scavenger receptors that might have increased lipid accumulation. The abnormal function of mesangial foam cells is dependent on the lipid moiety that accumulates as cholesterol-loaded cells fail to contract, phagocytose,
or migrate normally, whereas TG-loaded cells migrate normally in response to IGFBP-5 but are unable to contract in
response to angiotensin II. Additional studies are needed to
further define the specific effects of IGF-1 on fatty acid
synthase, VLDL receptors, and PPAR␦ expression.
F145
F146
IGF-1, LIPIDS, AND MESANGIAL CELLS
AJP-Renal Physiol • VOL
53. Lee HS, Lee JS, Koh HI, and Ko KW. Intraglomerular lipid deposition
in routine biopsies. Clin Nephrol 36: 67–75, 1991.
54. Li AC, Binder CJ, Gutierrez A, Brown KK, Plotkin CR, Pattison JW,
Valledor AF, Davis RA, Willson TM, Witztum JL, Palinski W, and
Glass CK. Differential inhibition of macrophage foam-cell formation and
atherosclerosis in mice by PPAR␣, ␤/␦, and ␥. J Clin Invest 114:
1564 –1576, 2004.
55. Lowry OH, Rosebrough NH, and Farr AL. Protein measurement with
the folin phenol reagent. J Biol Chem 193: 265–275, 1951.
56. Lupia E, Elliot SJ, Lenz O, Zheng F, Mattori M, Striker GE, and
Striker LJ. IGF-1 decreases collagen degradation in diabetic NOD
mesangial cells: implications for diabetic nephropathy. Diabetes 8: 1638 –
1644, 1999.
57. Lynn EG, Siow YL, and OK. Very low-density lipoprotein stimulates the
expression of monocyte chemoattractant protein-1 in mesangial cells.
Kidney Int 57: 1472–1483, 2000.
58. Magil AB. Interstitial foam cells and oxidized lipoprotein in human
glomerular disease. Mod Pathol 12: 33– 40, 1999.
59. Malden LT, Chait A, Raines EW, and Ross R. The influence of
oxidatively modified low density lipoproteins on expression of plateletderived growth factor by human monocyte-derived macrophages. J Biol
Chem 266: 13901–13907, 1991.
60. Massy ZA, Kim Y, Guijarro C, Kasiske B, Keane WF, and O’Donnell
MP. Low-density lipoprotein-induced expression of interleukin-6, a
marker of human mesangial cell inflammation: effects of oxidation and
modulation by lovastatin. Biochem Biophys Res Commun 267: 536 –540,
2000.
61. McGowan MW, Artiss JD, Strandbergh DR, and Zak B. A peroxidasecoupled method for the colorimetric determination of serum triglycerides.
Clin Chem 29: 538 –541, 1983.
62. Nicholson AC and Hajjar DP. CD36, oxidized LDL and PPAR␥:
pathological interactions in macrophages and atherosclerosis. Vasc Pharm
41: 139 –146, 2004.
63. Noel LH. Morphological features of primary focal and segmental glomerulosclerosis. Nephrol Dial Transplant 14, Suppl 3: 53–57, 1999.
64. O’Donnell MP. Mechanisms and clinical importance of hypertriglyceridemia in the nephrotic syndrome. Kidney Int 59: 380 –382, 2001.
65. Oppenheimer MJ, Sunquist K, and Bierman EL. Downregulation of
high-density lipoprotein receptors in human fibroblasts by insulin and
IGF-1. Diabetes 38: 117–122, 1989.
66. Piemonti L, Monti P, Allaventa P, Leone BE, Caputo A, and Carlo
VD. Glucocorticoids increase the endocytic activity of human dendritic
cells. Int Immunol 11: 1519 –1526, 1999.
67. Podrez EA, Febbraio M, Sheibani N, Schmitt D, Silverstein RL,
Hajjar DP, Cohen PA, Frazier WA, Hoff HF, and Hazen SL. Macrophage scavenger receptor CD36 is the major receptor for LDL modified by
monocyte-generated reactive nitrogen species. J Clin Invest 105: 1095–
1108, 2000.
68. Pohl J, Ring A, Ehehalt R, Herrmann T, and Stremmel W. New
concepts of cellular fatty acid uptake: role of fatty acid transport proteins
and of caveolae. Proc Nutr Soc 63: 259 –262, 2004.
69. Rothblat GH, de la Llera-Moya M, Atger V, Kellner-Weibel G, Williams
DL, and Phillips MC. Cell cholesterol efflux: integration of old and new
observations provides new insights. J Lipid Res 40: 781–796, 1999.
70. Rowe AH, Argmann CA, Edwards JY, Sawyez CG, Morand OH,
Hegele RA, and Huff MW. Enhanced synthesis of the oxysterol 24(S),25epoxycholesterol in macrophages by inhibitors of 2,3-oxidosqualene:
lanosterol cyclase: a novel mechanism for the attenuation of foam cell
formation. Circ Res 93: 717–725, 2003.
71. Ruan XZ, Moorhead JF, Fernando R, Wheeler DC, Powis SH, and
Varghese Z. PPAR antagonists protect mesangial cells from interleukin
1␤-induced intracellular lipid accumulation by activating the ABCA1
cholesterol efflux pathway. J Am Soc Nephrol 14: 593– 600, 2003.
72. Ruan XZ, Varghese Z, Powis SH, and Moorhead JF. Human mesangial
cells express inducible macrophage scavenger receptors, an Ap-1 and
ects-mediated response. Kidney Int 56: 5163–5166, 1991.
73. Ruan XZ, Varghese Z, Powis SH, and Moorhead JF. Human mesangial
cells express inducible macrophage scavenger receptor. Kidney Int 56:
440 – 451, 1999.
74. Ruan XZ, Varghese Z, Powis SH, and Moorhead JF. Dysregulation of
LDL receptor under the influence of inflammatory cytokines: a new
pathway for foam cell formation. Kidney Int 60: 1716 –1725, 2001.
290 • JANUARY 2006 •
www.ajprenal.org
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
28. Elliot SJ, Striker L, Hattori M, Yang CW, He CJ, Peten EP, and
Striker GE. Mesangial cells from diabetic NOD mice constitutively
secrete increased amounts of insulin-like growth factor-I. Endocrinology
133: 1783–1788, 1993.
30. Febbraio M, Hajjar DP, and Silverstein RL. CD36: a calls B scavenger
receptor involved in angiogenesis, atherosclerosis, inflammation, and lipid
metabolism. J Clin Invest 108: 785–791, 2001.
31. Frank PG and Lisanti MP. Caveolin-1 and caveolae in atherosclerosis:
differential roles in fatty streak formation and neointimal hyperplasia.
Curr Opin Lipidol 15: 523–529, 2004.
32. Fujita Y, Maruyama S, Kogo H, Matsuo S, and Fujimoto T. Caveolin-1
in mesangial cells suppresses MAP kinase activation and cell proliferation
induced by bFGF and PDGF. Kidney Int 66: 1794 –1804, 2004.
33. Gerrity RG. The role of monocytes in atherosclerosis: transition of
blood-borne monocytes into foam cells in fatty lesions. Am J Pathol 103:
181–190, 1981.
34. Ginsberg HN. New perspectives on atherogenesis: role of abnormal triglyceride-rich lipoprotein metabolism. Circulation 106: 2137–2142, 2002.
35. Grone EF, Abboud HE, Hohne M, Walli AK, Grone HJ, Stuker D,
Robenek H, Weiland E, and Seidel D. Actions of lipoproteins in cultured
human mesangial cells. Am J Physiol Renal Fluid Electrolyte Physiol 263:
F686 –F696, 1992.
36. Guan Y and Breyer MD. Peroxisome proliferator-activated receptors
(PPARs): novel therapeutic targets in renal disease. Kidney Int 60: 14 –30,
2001.
37. Hattori M, Nikolic-Paterson DJ, Miyazaki K, Isbel NM, Lan HY, Atkin
C, Kawaguchi H, and Ito K. Mechanisms of glomerular macrophage
infiltration in lipid-induced renal injury. Kidney Int 71: S47–S50, 1999.
38. Hussain MM, Strickland DK, and Bakillah A. The mammalian
low-density lipoprotein receptor family. Annu Rev Nutr 19: 141–172,
1999.
39. Inokuchi S, Kimura K, Sugaya T, Yoneda H, Shirato I, Murakami K,
and Sakai T. Angiotensin II maintains the structure and function of
glomerular mesangium via type 1a receptor. What we have learned from
null mutant mice minus the angiotensin II type 1a receptor gene. Virchows
Arch 433: 349 –357, 1998.
40. Jian B, de la Llera-Moya M, Ji Y, Wang N, Phillip MC, Swaney JB,
Tall AR, and Rothblat GH. Scavenger receptor class B type 1 as a
mediator of cellular cholesterol efflux to lipoproteins and phospholipid
acceptors. J Biol Chem 273: 5599 –5606, 1998.
41. Johnson AC, Yabu JM, Hanson S, Shah VO, and Zager RA. Experimental glomerulopathy alters renal cortical cholesterol, SR-B1, ABCA1,
and HMG CoA reductase expression. Am J Pathol 162: 283–291, 2003.
42. Kasiske BL, O’Donnell MD, Cleary MP, and Keane WF. Treatment of
hyperlipidemia reduces glomerular injury in obese Zucker rats. Kidney Int
33: 667– 672, 1988.
43. Kawamura M, Heinecke JW, and Chait A. Increased uptake of a-hydroxy aldehyde-modified low density lipoprotein by macrophage scavenger receptors. J Lipid Res 41: 1054 –1059, 2000.
44. Keane WF. The role of lipids in renal disease: future challenges. Kidney
Int 57: S27–S31, 2003.
45. Keane WF, Kasiske BL, and O’Donnell MP. Lipids and progressive
glomerulosclerosis: a model analogous to atherosclerosis. Am J Nephrol 8:
261–271, 1988.
46. Kodama N, Otani H, Yamada Y, and Mune MYS. Involvement of
MCP-1 and M-CSF in glomerular foam cell formation in ExHC rats.
Kidney Int 71: S174 –S177, 1999.
47. Kreisberg JI and Karnovsky MJ. Glomerular cells in culture. Kidney Int
23: 439 – 447, 1983.
48. Kruth HS, Jones NL, Huang W, Zhao B, Ishii I, Chang J, Combs CA,
Malide D, and Zhang WY. Macropinocytosis is the endocytic pathway
that mediates macrophage foam cell formation with native low density
lipoprotein. J Biol Chem 280: 2352–2360, 2005.
49. Lee CH, Chawla A, Urbiztondo N, Liao D, Boisvert WA, and Evans
RM. Transcriptional repression of atherogenic inflammation: modulation
by PPAR␦. Science 302: 453– 457, 2003.
50. Lee CH, Olson P, and Evans RM. Minireview: lipid metabolism,
metabolic diseases, and peroxisome proliferator-activated receptors. Endocrinology 144: 2201–2207, 2003.
51. Lee HS. Oxidized LDL, glomerular mesangial cells and collagen. Diabetes Res Clin Pract 45: 117–122, 2000.
52. Lee HS and Kim YS. Identification of oxidized low density lipoproteins
in human renal biopsies. Kidney Int 54: 848 – 856, 1998.
IGF-1, LIPIDS, AND MESANGIAL CELLS
75. Sato T, Liang K, and Vaziri ND. Protein restriction and AST-120
improve lipoprotein lipase and VLDL receptor in focal glomerulosclerosis. Kidney Int 64: 1780 –1786, 2003.
76. Scheidegger KJ, James RW, and Delafontaine P. Differential effects of
low density lipoproteins on insulin-like growth factor-1 (IGF-1) and IGF-1
receptor expression in vascular smooth muscle cells. J Biol Chem 275:
26864 –26869, 2000.
77. Schlondorff D. The glomerular mesangial cell: an expanding role for a
specialized pericyte. FASEB J 1: 272–281, 1987.
78. Teruel T, Valverde AM, Benito M, and Lorenzo M. Insulin-like growth
factor I and insulin induce adipogenic-related gene expression in fetal
brown adipocyte primary cultures. Biochem J 319: 627– 632, 1996.
F147
79. Wang N, Silver DL, Costet P, and Tall AR. Specific binding of
ApoA-1 enhanced cholesterol efflux, and altered plasma membrane
morphology in cell expressing ABCA-1. J Biol Chem 275: 33053–
33058, 2000.
80. Wang YX, Lee CH, Tiep S, Yu RT, Ham J, Kang H, and Evans RM.
Peroxisome-proliferator-activated receptor ␦ activates fat metabolism to
prevent obesity. Cell 113: 159 –170, 2003.
81. Wheeler DC and Chana RS. Interactions between lipoproteins, glomerular cells and matrix. Miner Electrolyte Metab 19: 149 –164, 1993.
82. Wu J, Zhang Y, Wang N, Davis L, Yang G, Wang X, Zhu Y, Breyer MD,
and Guan Y. Liver X receptor-␣ mediates cholesterol efflux in glomerular
mesangial cells. Am J Physiol Renal Physiol 287: F886 –F895, 2004.
Downloaded from http://ajprenal.physiology.org/ by 10.220.33.6 on August 2, 2017
AJP-Renal Physiol • VOL
290 • JANUARY 2006 •
www.ajprenal.org