© 2000 American Heart Association, Inc.
Atherosclerosis and Lipoproteins |
Expression of Class A Scavenger Receptor Inhibits Apoptosis of Macrophages Triggered by Oxidized Low Density Lipoprotein and Oxysterol
From the Cardiovascular and Pulmonary Research Institute (H.-S.L., Y.-J.G.), Allegheny General Hospital, Pittsburgh, Pa; Molecular Biology and Medicine (T.K.), University of Tokyo, Tokyo, Japan; and the Cardiology Division (Y.-J.G.), Department of Internal Medicine, University of Texas School of Medicine at Houston.
Correspondence to Yong-Jian Geng, MD, PhD, Cardiology Division, Department of Internal Medicine, University of Texas School of Medicine at Houston, 6431 Fannin St, MSB 1.240, Houston, TX 77030. E-mail ygeng{at}heart.med.uth.tmc.edu
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Abstract |
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Abstract—The class A macrophage scavenger receptor (MSR-A) is a multifunctional trimeric glycoprotein involved in innate immune response as well as the development of lipid-laden foam cells during atherosclerosis. The MSR ligand, oxidized low density lipoprotein (oxLDL), is known to be cytotoxic to macrophages and other cell types. This study examined whether MSR mediates or modulates oxLDL-induced apoptosis. Treatment with oxLDL and its cytotoxic oxysterol, 7-ketocholesterol (7-KC), reduced viability and increased DNA fragmentation in human THP-1 cells, Chinese hamster ovary cells, and mouse peritoneal macrophages. However, cell death and DNA fragmentation were markedly diminished in the phorbol ester–differentiated MSR-expressing THP-1 cells and Chinese hamster ovary cells, with stable expression of MSR-AI after cDNA transfection when exposed to the same concentrations of oxLDL and 7-KC. Moreover, treatment with oxLDL and 7-KC induced much greater death and DNA fragmentation in MSR-A–deficient peritoneal macrophages compared with wild-type macrophages. Thus, MSR-A does not act as a receptor responsible for the apoptotic effect of oxLDL, and instead, expression of this receptor confers resistance of macrophages to the apoptotic stimulation by oxLDL and its cytotoxic lipid component. These results suggest that by preventing apoptosis, MSR-A may contribute to the long-term survival of macrophages and macrophage-derived lipid-laden foam cells in atherosclerotic lesions.
Key Words: macrophages • scavenger receptors • atherosclerosis • lipoproteins • cell death
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Introduction |
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Atherosclerosis is a chronic arterial disease with 2 life-threatening complications, myocardial and cerebral infarcts. The long-lasting process of atherogenesis involves dramatic alterations in cellularity, extracellular matrix, and lipid components of the arterial wall, resulting in intimal thickening, vessel lumen narrowing, and increased susceptibility to thrombosis. Cell proliferation has been traditionally regarded as a key factor that increases the number of cells in the arterial intima. However, analysis of proliferative markers has revealed a low degree of cell replication.1 On the other hand, recent studies have documented that deregulation of apoptosis, a form of genetically programmed cell death, occurs in atherosclerotic lesions.2 3 Imbalance between cell survival and death may contribute to dramatic alterations in cellularity of the arterial wall with atherosclerosis.
Formation of lipid-laden foam cells from macrophages and, to a less extent, from smooth muscle cells represents a landmark for atherosclerosis. The development of foam cells is mainly due to overloading of lipids, particularly cholesterol and cholesterol ester, into the cells through a scavenger receptor–mediated process. It is believed that the lipids within the foam cells largely come from the lipid-rich proteins in blood, particularly LDLs, which enter the cells after chemical modification, such as oxidation and converting to the ligand for macrophage scavenger receptors (MSRs). Three different classes of MSR have been identified.4 Among them, the class A MSR (MSR-A) is the first one to be cloned and characterized for its ligand binding specificity and capacity.5 Its expression is limited to differentiated or mature monomyeloid cells and is thus characterized for the monocyte-to-macrophage differentiation. Our previous studies have shown that circulating monocytes elaborate MSR-A at undetectable levels but that when the cells differentiate into tissue macrophages, they express high levels of MSR-A.6 7
The MSR ligand, oxidized LDL (oxLDL), has been identified as an apoptosis-promoting agent to a variety of cell types, including vascular smooth muscle cells,8 9 endothelial cells,10 11 12 and macrophages.13 14 Other modified LDLs, such as enzymatically degraded LDLs (E-LDL), can selectively induce the expression of monocyte chemotactic protein-1 and exert cytotoxic effects on human macrophages.15 16 In vitro treatment with oxLDL may activate the cysteinyl protease, caspase-3,9 which acts as an effector in the downstream caspase death cascade. However, despite the well-defined ligand-receptor relationship between oxLDL and MSR, it remains largely unclear as to whether MSR-A mediates or modulates apoptotic effects of oxLDL and associated lipids. Our recent work17 has shown that differentiated MSR-A–expressing human THP-1 macrophages become resistant to apoptosis triggered by fluoride anions, a global activator of G protein. Chinese hamster ovary (CHO) cells with stable expression of MSR-A also exhibit increased resistance to the fluoride apoptotic stimulation. Moreover, recent studies have shown that expression of a scavenger receptor–like gene, the cellular stress response gene, prevents cellular damage by UV irradiation and oxidative stress18 and that thymocytes lacking the apoptosis inhibitor of macrophages, a member of the scavenger receptor cysteine-rich domain family, are more susceptible to apoptosis.19 Thus, these data seem argue against the notion that oxLDL triggers apoptosis in an MSR-A–dependent manner.
The present study was aimed to determine (1) whether the expression of MSR-A prevents oxLDL-induced apoptosis in differentiated human THP-1 monocytic macrophages, (2) whether overexpression of MSR-A in CHO cells reduces apoptosis triggered by oxLDL and oxysterol, and (3) whether resident peritoneal macrophages from the MSR-A–null (MSR-/-) mice are more sensitive to oxLDL and oxysterol.
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Methods |
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Reagents
Free cholesterol and 7-ketocholesterol (7-KC) were purchased from Sigma Chemical Co and dissolved in dimethyl sulfoxide at a stock solution of 10 mg/mL. Native LDL and oxLDL were obtained from Perlmmune Inc. Acridine orange and ethidium bromide (nucleic acid–binding dyes) were obtained from Sigma. RPMI-1640 and F-12 medium were purchased from GIBCO. Phorbol 12-myristate 13-acetate (PMA) was obtained from Sigma and dissolved in dimethyl sulfoxide at a stock concentration of 10 mmol/L.
Cell Culture and Treatment
THP-1 and CHO cells were obtained from American Type Culture
Collection. THP-1 cells were maintained in RPMI-1640 supplemented with
10% heat-inactivated FBS, 100-U/mL penicillin, and 100
µg/mL streptomycin. CHO cells were maintained in F-12 medium
supplemented with 10% heat-inactivated FBS, 100 U/mL
penicillin, and 100 µg/mL streptomycin. THP-1 cells were pretreated
with or without 100 nmol/L of PMA, washed with PBS, and then treated
with various concentrations of oxLDL and 7-KC, respectively. Control
experiments were performed by using the same concentration of the
vehicle, dimethyl sulfoxide.
Resident Peritoneal Macrophage Isolation and
Treatment
Resident peritoneal macrophages were isolated from
wild-type (MSR+/+) and homozygous MSR-deficient
(MSR-/-) mice by
peritoneal lavage as described previously.20 Briefly, mice
injected with 0.2 mL of ketamine (100 µg/mL) underwent lavage
with 8 mL of serum-free RPMI-1640 media into the peritoneal cavity.
Peritoneal macrophages were obtained by
centrifugation of the lavage fluid collected from the
peritoneal cavity. The collected macrophages were resuspended
in RPMI-1640 with 10% heat-inactivated FBS and incubated
at 1x105 cells per milliliter in 8-well chamber
slides for 48 hours. The cells were then stimulated with oxLDL or 7-KC.
After stimulation, the cells were fixed for terminal
deoxynucleotidyl transferase (TdT)-mediated dUTP
nick end-labeling (TUNEL) staining as shown below.
Cell Viability Assay
Cell viability was determined by staining with a combination of
the nucleic acid–binding dyes acridine orange (10 µg/mL) and
ethidium bromide (10 µg/mL) for 5 minutes on ice. After staining,
viable and nonviable cells were counted on the basis of color and
appearance under a fluorescence microscope. Acridine orange and
ethidium bromide dyes can intercalate into DNA. Viable cells are
stained with green nuclei by acridine orange, whereas nonviable cells
are stained with ethidium bromide, producing a red or orange color in
nuclei. At least 200 cells were counted. The percentage of dead cells
was calculated by dividing the number of dead cells by total cell
number.
Analysis of DNA Fragmentation
DNA was extracted from the cells stimulated by oxLDL and 7-KC in
various concentrations for 24 hours in 6-well plates. After treatment,
5x106 cells were lysed in 1 mL of DNA extract
solution containing 100 mmol/L NaCl, 20 mmol/L Tris-HCl (pH.
8.0), and 10 mmol/L EDTA at room temperature for 15 minutes. The
cell lysates were incubated overnight with proteinase K (200 µg/mL)
at 37°C, extracted for DNA with phenol/chloroform isoamyl alcohol,
and precipitated with the same volume of isopropanol. Remaining RNA was
removed by digestion with RNase A (25 µg/mL) at 37°C for 30
minutes. After reextraction and precipitation, DNA concentration was
determined by spectrophotometer at a wavelength of 260 nmol/L. DNA at
20 µg per lane was loaded into 2% agarose gels with 0.5 µg/mL
ethidium bromide and separated by electrophoresis at a constant voltage
of 100 V for 1 to 2 hours at room temperature. The DNA ladder was
visualized under UV light.
Total RNA Isolation and RT-PCR
Total RNA was extracted from THP-1 cells treated with or without
PMA (100 nmol/L) for 48 hours by using the phenol/guanidinium
thiocyanate method.21 cDNA was generated by reverse
transcription (RT) from 1 µg of total RNA and a 30-cycle polymerase
chain reaction (PCR) with specific primers for MSR-A types I and II
(MSR-AI and MSR-AII, respectively), as described
previously.6 RT-PCR for ß-actin mRNA was used as an
internal control. The PCR products were analyzed by
electrophoresis on 2% agarose gels containing 0.5 µg/mL ethidium
bromide.
In Situ 3' End Labeling of DNA Fragments (TUNEL)
Mouse peritoneal macrophages treated with and without
oxLDL or 7-KC in 8-well chamber slides for 24 hours were fixed with 3%
paraformaldehyde in PBS. After they were washed in PBS,
the cells were incubated with 2%
H2O2 in methanol to
inactivate endogenous peroxidase.
Digoxigenin-conjugated dUTP was incorporated into 3' ends of DNA with
TdT by using an ApoTag in situ apoptosis detection kit
(Oncor, Inc). The presence of digoxigenin-labeled DNA fragments was
determined by using a peroxidase-conjugated antibody against
digoxigenin. The chromogenic substance diaminobenzidine was
used as a substrate for visualization of the
immunostaining. The percentage of TUNEL-positive cells
was calculated by dividing the number of positive cells by the total
cell number.
CHO Cell Line With Stable Overexpression of MSR-A
CHO cells were transfected with MSR-A type I cDNA by a standard
calcium phosphate precipitate method.17 After
transfection, CHO cells were selected with G-418 medium for 4 weeks.
The establishment of the CHO cell line with stable expression of MSR-A
was confirmed by Northern and Western blotting.
Flow Cytometry
THP-1 cells were analyzed by flow cytometry for the
expression of different isoforms of MSR, such as MSR-A, CD36, and CD68.
The cells were pretreated with PMA and then exposed to 25 µg/ml oxLDL
at 37°C for 24 hours. After they were washed in PBS, the cells were
stained with monoclonal antibodies against MSR-A, CD36, and CD68,
followed by incubating with FITC-conjugated anti-mouse IgG. The
immunostains were detected by using a FACScalibur
flow cytometer (Becton Dickinson) with Cytoquest software.
Statistic Analysis
The experimental data were analyzed by using
Excell software (Microsoft), and ANOVA was used for statistical
analysis of multiple groups. Significant difference was
established at a value of P<0.05.
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Results |
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MSR+ THP-1 Cells Resist Apoptosis Triggered by OxLDL
Human THP-1 monocytic cells expressed high levels of mRNA for MSR-AI and MSR-AII after stimulation with PMA (Figure 1
). To determine the extent of
apoptosis in the MSR+ THP-1 cells, the
cells were pretreated with PMA and then treated with various
concentrations of oxLDL for 24 hours. We observed that treatment with
oxLDL significantly reduced the viability of
MSR- and MSR+ THP-1 cells
in a concentration-dependent manner, as determined by staining with
acridine orange and ethidium bromide
(Table). In contrast, under the
same concentration, native LDL did not alter cell viability.
Interestingly, the number of nonviable MSR+ THP-1
cells appeared significantly lower than the number of nonviable
MSR- THP-1 cells ( Table). Because
internucleosomal DNA fragmentation biochemically characterizes
apoptosis, we analyzed the sizes of DNA isolated from
oxLDL-treated and untreated THP-1 cells by agarose gel electrophoresis.
As shown in Figure 1b, PMA-treated THP-1 cells showed decreased
fragmentation of DNA at the internucleosomal sizes compared with the
untreated MSR- THP-1 cells. Morphological
changes, including cell shrinkage, blebbing, chromatin condensation,
and nucleus fragmentation, also occurred, suggesting that the cells
underwent apoptosis.
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MSR+ THP-1 Cells Resist Apoptosis Triggered
by 7-KC
OxLDL contains many different lipid components, eg, lipid
peroxides, hydroxides, and oxidized cholesterol derivatives
(eg, oxysterol), many of which are potentially cytotoxic. We examined
whether 7-KC, a major oxysterol component found in
oxLDL,22 23 promotes apoptosis of THP-1 cells. We
observed that similar to oxLDL, 7-KC also reduced the viability of
THP-1 cells in a concentration-dependent fashion ( Table
).
Increasing concentrations of 7-KC from 5 µg/mL to 20 µg/mL led to
50% cell death ( Table). Thus, 7-KC appeared to exert a
cytotoxic effect on THP-1 cells. The 7-KC–induced cell death occurred
via an apoptotic mechanism that was evidenced by
internucleosomal DNA fragmentation, as shown by the appearance of DNA
ladder in agarose gel electrophoresis in the 7-KC–treated cells
(Figure 2). In control experiments, we
treated the cells with the same concentrations of free
cholesterol. We observed no effect of free
cholesterol on the cell viability, because it neither
increased the percent cell death (4.3±0.4% in the free
cholesterol–treated cells versus 3.9±0.5% in the control
cells, n=3) nor caused DNA fragmentation (data not shown). To determine
whether MSR-expressing cells resist apoptosis induced by the
oxysterol, we analyzed the differences in viability and DNA
fragmentation between MSR- and
MSR+ THP-1 cells. We found that the percentage of
cell death and the internucleosomal DNA fragmentation were diminished
in the MSR+ cells compared with the
MSR- THP-1 cells ( Table and Figure
2). These results indicate that PMA-pretreated MSR-A–expressing
THP-1 macrophages were resistant to apoptosis
induced by 7-KC.
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To determine whether other isoforms of MSR participate in the regulation of apoptosis induced by oxLDL and oxysterols, we analyzed the expression of CD36 and CD68, 2 isoforms of MSR, in the PMA-differentiated THP-1 cells by flow cytometry after stimulation with oxLDL. We observed that compared with untreated control cells, there were no significant increases in the numbers of CD36-positive cells (11% versus 12% in control cells) and CD68-positive cells (20% versus 19% in control cells) in the PMA-pretreated cultures 24 hours after stimulation with oxLDL (50 µg/mL). Similarly, we found that the 7-KC–treated cells exhibited no changes in the expression of the 2 proteins. This suggests that MSR-A other than CD36 and CD68 contributed to the resistance of the cells to apoptosis induced by oxLDL and 7-KC.
Overexpression of MSR-A by cDNA Transfection Inhibits
Apoptosis Induced by OxLDL in CHO Cells
To further confirm that expression of MSR itself prevents
apoptosis induced by oxLDL and 7-KC, we recently established a
CHO cell line with stable overexpressed MSR-AI by transfection with the
MSR-AI cDNA. Under baseline conditions, CHO cells expressed little MSR
activities, but transfection with a cDNA coding for this receptor
dramatically increased MSR expression in the cells, as shown
previously.5 24 Figure 3a
demonstrates that the CHO cells with stable expression of MSR showed
significantly higher viability than untransfected wild-type CHO cells
when treated with the same concentrations of oxLDL. Agarose gel
electrophoresis of DNA revealed a decrease in internucleosomal
fragmentation of DNA in the MSR-A–expressing CHO cells compared with
wild-type control cells (Figure 3b). These results suggest that
overexpression of MSR-AI might increase the resistance of CHO cells to
apoptosis induced by oxLDL.
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Expression of MSR by cDNA Transfection Inhibits Apoptosis
Induced by 7-KC in CHO Cells
To determine whether MSR expression confers resistance of
CHO cells to apoptosis induced by the oxysterol, 7-KC, we
examined the viability of wild-type and MSR-AI–transfected CHO cells
after treatment with 7-KC. We observed a concentration-dependent
increase in the percentage of cell death in untransfected wild-type
cells treated with 7-KC for 24 hours (Figure 4a). However, the percentage of cell
death was much lower in the transfected cells than in the wild-type
control cells. Agarose gel electrophoresis also showed a reduction in
DNA cleavage in the CHO cells with overexpressed MSR (Figure
4b). Thus, the MSR-overexpressing CHO cells were also
resistant to apoptotic stimulation by 7-KC.
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MSR-A–Null Macrophages Exhibit Increased Sensitivity to
Apoptosis Induced by OxLDL and 7-KC
To further verify the antiapoptotic effect of MSR-A on
macrophages, we treated resident peritoneal macrophages
isolated from wild-type (MSR+/+) and MSR-knockout
(MSR-/-) mice with oxLDL
and 7-KC. We observed that treatment with oxLDL led to increased
numbers of cell death in
MSR-/-
macrophages compared with wild-type cells in a
concentration-dependent manner (Figure 5), as determined by staining with
acridine orange and ethidium bromide.25 26 In situ 3' end
labeling of DNA fragments by use of the TUNEL technique demonstrated an
increase in nuclear DNA fragmentation in the treated cells (Figure 6). There were significantly higher
numbers of TUNEL-positive nuclei present in the
MSR-/- cells compared
with wild-type macrophages treated with the same concentration
of oxLDL (Figure 7a). The
MSR-/-
macrophages also exhibited higher TUNEL positivity than did the
wild-type cells when exposed to 7-KC for 24 hours (Figures 6 and
Figure 7b). Thus, the MSR-A deficiency appeared to increase the
susceptibility of the peritoneal macrophages to
apoptotic stimulation by oxLDL and the oxysterol 7-KC.
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Discussion |
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As part of the innate immune system, macrophages recognize and remove denatured macromolecules, pathogenic microorganisms, and damaged tissue or apoptotic cells through their scavenger system. Despite the high frequency of exposure to those environmental cytostatic or cytotoxic factors, the lifespan of tissue macrophages remains considerably longer. This is particularly the case in the atherosclerotic lesions, in which numerous macrophages survive an extremely harsh microenvironment that contains numerous toxic substances, such as oxLDL and E-LDL.15 16 27 This mystery has led us to explore the possibility that tissue macrophages may express a protein(s) with the potential of preventing apoptosis, a form of genetically programmed cell death.
MSR, an abundant glycoprotein expressed on the surface of macrophages, mediates the uptake of chemically modified LDL by macrophages. This raises an interesting question as to whether oxLDL exerts its cytotoxic effect through the scavenger receptor pathway. In the present study, we used 3 different in vitro models to define the role for MSR-A in the modulation of macrophage apoptosis triggered by oxLDL and its cytotoxic lipid component, 7-KC. The first model uses PMA-differentiated human THP-1 cells, which express MSR-A at high levels. The second model uses a CHO cell line with stably overexpressed MSR-A. The third or last model focuses on peritoneal macrophages from the MSR-A knockout mice. Our observations clearly indicate that MSR-A does not act as a signaling receptor for oxLDL and 7-KC to achieve their cytotoxic effects. Instead, the expression of MSR-A reduces the extent of apoptosis in macrophages exposed to oxLDL and 7-KC, whereas MSR-A deficiency increases the sensitivity to apoptosis. Thus, MSR-A may function as an inhibitor of apoptosis triggered by the oxidized lipoprotein and its toxic lipid component rather than as a signal transducer responsible for the oxLDL-induced cell death. The cytotoxicity of oxLDL appears to be nonselective or independent of MSR-A, inasmuch as it causes apoptosis in a broad range of cell types, including MSR-A–negative and –positive cells.
However, we cannot exclude other scavenger receptor families, which may serve as death-signaling receptors for oxLDL. In fact, MSR-A has a relatively lower affinity to oxLDL compared with other forms of MSR, such as CD3628 and CD 68.29 30 The major ligand used for evaluating MSR activity is acetylated LDL, an artificially modified lipoprotein that may not exist in vivo. Hence, the physiological ligand for MSR-A has not been characterized at this time point. Nevertheless, the observation that oxLDL and 7-KC induce apoptosis of THP-1 and CHO cells confirms and extends previous findings8 9 by other groups that oxLDL treatment can trigger vascular cell apoptosis. Because we observed no major change in the expression of CD36 and CD68 in the PMA-differentiated THP-1 cells after stimulation with oxLDL and because oxLDL can trigger apoptosis in MSR-negative cells, it is unlikely that CD36 and CD68 are the principal contributors to the resistance of macrophages to apoptosis induced by oxLDL and oxysterols.
The mechanism underlying the inhibitory effect of MSR on oxLDL-mediated apoptosis remains to be determined. OxLDL is composed of various cytotoxic components, including oxysterols. 7-KC is the second most abundant oxysterol found in human atherosclerotic lesions after 27-hydroxycholesterol. Whether 7-KC is a major cytotoxic component in oxLDL remains to be investigated. The intracellular domains of MSR do not contain the sequences of any known signaling factor–binding sites. Although ligand transporting to lysosomes via the MSR pathway can help to remove and degrade oxLDL, the MSR-mediated endocytosis may not be the sole or major mechanism by which MSR prevents apoptosis. This notion is supported by the fact that there is a low affinity between oxLDL and MSR-A and no 7-KC binding to MSR-A. Our recent studies17 have provided an additional interpretation for the protective effect of MSR, showing that expression of MSR-A blocks apoptosis induced by the activation of membrane GTP-binding proteins. In this regard, MSR-A may participate in G-protein–mediated cross-membrane signal transduction in macrophages.
THP-1 cells, a human monocytic leukemia cell line, can differentiate
into macrophage-like cells, after exposure to the transforming
agent, phorbol esters. The differentiated THP-1 macrophages
express high levels of MSR and behave in a manner highly similar to
that of native monocyte-derived macrophages.31 PMA
is a potent activator of protein kinase C. Regulation of
apoptotic death by the protein kinase C pathway has been
recently reported in the cells.32 33 34 35 However, in the
present study, we observed no difference in cell death between
PMA-treated and untreated THP-1 cells. Very recently, we have shown
increased expression of apoptosis-promoting genes, such as
FasL, and CPP-32 (caspase-3) in PMA-differentiated THP-1 cells, but
there is no increase in the number of cells bearing markers of
apoptosis.17 It is reasonable to expect that the
expression of MSR-A plays a role in the regulation of apoptosis
of macrophages. Nishio and Watanabe9 reported that
oxysterols induced apoptosis in cultured smooth muscle cells
through CPP-32 activation and bcl-2 protein downregulation. However, it
is unclear whether 7-KC exerts a similar effect on macrophages
and whether MSR can block the regulatory effects of 7-KC on the cells.
Munn et al36 reported that macrophage
colony-stimulating factor (CSF)–differentiated macrophages are
resistant to apoptosis, whereas
granulocyte-macrophage CSF–differentiated and
interferon-
–differentiated macrophages are sensitive to
apoptosis. Interestingly, macrophage CSF increases but
interferon- and granulocyte-macrophage CSF decrease the
expression of MSR.37 38 39
Recently, several proteins with the MSR cysteine-rich domain (SRCR), such as cellular stress response elements18 and apoptosis inhibitors of macrophages,19 have been reported to exert protective effects on apoptosis of fibroblasts and macrophages, respectively. The mechanism underlying the protective effects of the SRCR-containing proteins is unknown. The mRNA and protein levels of MSR-AI with the SRCR domain are not appreciable in most peripheral blood monocytes, but MSR-AI expression elevates dramatically when monocytes differentiate into macrophages. The increased expression of MSR-AI concurs with the development of resistance to apoptosis in the monocyte-derived macrophages. Further investigation is needed to delineate the relationship of MSR-AI expression with the macrophage resistance to apoptosis.
It has been shown that macrophages of atherosclerotic lesions express high levels of MSR-A,6 suggesting that the expression of MSR is associated with maturation of monocytes and resistance of certain macrophages to apoptosis in atherosclerosis. Most of the plaque macrophages contain substantial amounts of lipids, mainly cholesterol, that are derived from modified lipoproteins internalized via the scavenger pathway. The increased MSR-A expression may not only promote the uptake of those lipid components but also contribute to the longevity of the lipid-rich foam cells by increasing the resistance of the cells to cytotoxic effects of oxLDL and oxysterols. Recently, Kockx and colleagues40 41 reported lower levels of apoptosis in certain atherosclerotic lesions than previously thought, suggesting that nonspecific staining for the cells undergoing apoptosis or attenuation of apoptosis might occur in the lesions. Our present data appear to support the notion that in atherosclerotic plaque, there is a mechanism that is responsible for the inhibition of apoptosis and that leads to long-term accumulation of lipid-laden macrophage-derived foam cells during atherogenesis.
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Acknowledgments |
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This study was supported by National Institutes of the Health grant HL-59249–01 (Y.-J. Geng) and an American Heart Association, Pennsylvania affiliate, Grant-in-Aid award (Y.-J. Geng). We thank Dr Hiroshi Suzuki, Chugaii Pharmaceutical Co, Japan, for the generation of MSR-A knockout mice. We also thank Tamara Pittman and Qi Wu for their skilled technical assistance.
Received December 31, 1999; accepted January 17, 2000.
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