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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2000;20:2506.)
© 2000 American Heart Association, Inc.

Editorial

Scavenger Receptors in Atherosclerosis

New Answers, New Questions

Theodore Mazzone

From the Departments of Medicine and Biochemistry, Rush Medical College, Chicago, Ill.

Correspondence to Dr Theodore Mazzone, Rush Medical Center, 1653 W Congress Pkwy, Chicago, IL 60612. E-mail tmazzone{at}rush.edu

Key Words: Editorials • atherosclerosis • macrophages • scavenger receptors • transgenic mice

The existence of a cell surface receptor, now known as the macrophage scavenger receptor A I/II (MSR-A), was inferred from early studies based on ligand binding analyses. Description of this binding activity, with acetylated low density lipoprotein (LDL) used as a ligand, was important because it provided a mechanism for the formation of foam cells in vivo and because the activity of this receptor was not downregulated by expanding cellular cholesterol stores as had been recently shown for the LDL receptor. The presence of acetylated LDL receptor binding activity, predominantly on macrophage-type cells, helped to underscore the importance of the macrophage in atherogenesis and stimulated further investigation of macrophage function in the vessel wall. The description of this activity also helped to fuel investigative efforts to identify a potential in vivo ligand, and the various forms of oxidized LDL were soon suggested as physiological ligands for this receptor. Thus, the MSR-A has already shaped investigative efforts into the mechanisms of atherosclerosis. Now that MSR-A has been cloned, attempts to further understand its role in atherogenesis with the use of mice deficient in its expression have been undertaken.

An early report by Suzuki et al¹ demonstrated that compared with control apolipoprotein E (apoE)–knockout mice, mice with macrophage scavenger receptor A I/II (MSR-A) deficiency, on an apoE-deficient background, exhibited a 60% decrease in lesions; this observation was consistent with a proatherogenic role for this receptor in vivo. There were a number of surprising findings in their study. Even though acetylated LDL degradation by macrophages was reduced substantially, there was no difference in the metabolism of acetylated LDL in vivo. In spite of this, the plasma cholesterol level in double-knockout mice was higher than in apoE-knockout mice. These double-knockout mice were also found to have increased susceptibility to infection, and the macrophages from these mice displayed defective adhesion in vitro. Finally, it was noted that the lesions of double-knockout mice still contained macrophage-derived foam cells, indicating the involvement of other macrophage scavenger receptors in foam cell formation.

In a subsequent report, the effect of eliminating MSR-A expression was examined for atherosclerosis in an LDL receptor–deficient background.² Double-knockout mice in that study showed a significant decrease in lesions; however, this decrease was smaller than that reported previously (20%). In still another study, the effect of MSR-A deficiency was examined on atherosclerosis in the apoE Leiden transgenic mouse fed a high fat diet.³ In that study, there was no decreased lesion formation as a result of MSR-A deficiency. In fact, lesions actually tended to be larger and somewhat more complex in the apoE3 Leiden/MSR-A–deficient animals. As a result of the above reports, it has been suggested that an antiatherogenic role of the MSR-A depends on the mouse model used to provide the atherogenic background and that an interaction between MSR-A and apoE, perhaps in modulating macrophage cholesterol homeostasis, is a modifying factor.

A recent study by Babaev et al⁴ in this issue of Arteriosclerosis, Thrombosis, and Vascular Biology provides new and important information regarding MSR-A involvement in atherogenesis. In their study, the authors used MSR-A–deficient mice on a C57BL/6 background challenged with a butterfat diet. The MSR-A–deficient mice had 60% less atherosclerosis. The authors then repeated this observation with use of the C57BL/6 diet-induced model but with the transplantation of MSR-A^-/- or MSR-A^+/+ fetal liver cells into lethally irradiated recipients. They again noted a substantial and significant decrease in atherogenesis in the animals receiving the MSR-A–deficient cells. In yet another approach, the authors used fetal liver cells transplanted into LDL receptor–deficient mice on a high fat diet. Using this model, they again demonstrated a substantial and significant decrease (again 60%) in lesion formation in the mice receiving the MSR-A–deficient cells. On the basis of these results, the authors conclude that MSR-A expression in macrophages significantly contributes to atherosclerotic lesion formation and that a mixed genetic background in animals may have contributed to the smaller effect on atherosclerosis previously reported in LDL receptor^-/- mice and to the absence of effect previously reported in apoE Leiden transgenic mice. More specifically, they point out that the mating of MSR-A–deficient mice (129/ICR) with LDL receptor–deficient mice (129/C57BL/6) or apoE Leiden mice (C57BL/6) produced hybrids that likely differed in the segregation of loci for atherosclerosis susceptibility.^{4 5}

There are several issues to be considered in integrating these new observations. As the authors suggest,⁴ the issue of genetic background can be an important source of unpredictable and/or inconsistent results in studies using transgenic or gene-targeted mice. This issue has been recently reviewed in Arteriosclerosis, Thrombosis, and Vascular Biology.⁶ For atherosclerosis studies, the issue of genetic variability can be separated into distinct considerations. Because mice do not spontaneously develop atherosclerosis, a common study design begins with a mouse strain genetically manipulated to produce atherosclerosis. Atherosclerosis-prone models are produced on mice of different genetic backgrounds, and these variable backgrounds are a potential source of inconsistent results. To investigate the involvement of a specific gene in atherogenesis, these atherosclerosis-prone mice are usually mated to another strain of mice, deficient in the gene of interest. An important question for the generalization of the results of such studies is whether the experiment has been designed so that (single knockout or transgenic) control mice (ie, the proatherogenic model) are genetically identical to the experimental mice (bearing the new deletion on the atherogenic background) in every way except for deletion of the gene of interest. The next issue is the nature of the proatherogenic manipulation itself. It is entirely conceivable that manipulating the expression of a specific gene might have different effects on atherosclerosis depending on the mechanism and potency of the proatherogenic milieu selected for study. Again, the study by Babaev et al⁴ helps to deal with some of these complexities, as noted by the authors. An important advantage of their present study is the examination of 2 different proatherogenic models, a diet-induced C57/BL6 and an LDL receptor–deficient model. Consistent results were obtained between these models. Furthermore, the use of the transplant approach provides potential advantage with respect to eliminating genetic background variability. By transplanting cells from engineered donor mice into genetically homogeneous hosts for experimental comparisons, potentially confounding genetic factors expressed in the donor animals (but not expressed in the transplanted cells) are eliminated. Therefore, the authors convincingly demonstrate the involvement of MSR-A in atherogenesis.

As the present study of Babaev et al⁴ shows, many confounding issues that could contribute to unpredictable and/or variable results in mouse atherosclerosis studies are amenable to experimental elimination. In fact, with a systemic approach, the use of well-characterized and genetically distinct lines of mice can be used to advantage to identify modifier genes for atherosclerosis. Such an approach has been proposed for investigating the large differences in atherosclerosis observed in apoE-deficient C57BL/6J mice compared with apoE-deficient FVB/NJ mice.⁷ Furthermore, the use of multiple proatherogenic models, when the influence of a specific gene of interest on atherogenesis is evaluated, facilitates the generalization of conclusions if the results are consistent across models. Results that differ between models can be mechanistically informative and lead to the generation of hypotheses dealing with more subtle and complex issues regarding the effect of a specific genetic manipulations on atherogenesis in the mouse.

One of these more complex issues relates to the temporal effects of genetic manipulation on atherogenesis. Babaev et al⁴ also raise this issue by suggesting that MSR-A may assume a greater role in atherosclerosis with prolonged exposure to a high fat diet, which was used for 30 weeks in their study. Temporal issues could also interact with the potency of the proatherogenic model to influence results. These considerations could perhaps be related to recent observations regarding deletion of the expression of another scavenger receptor—the class B scavenger receptor, CD36. Mice deficient in both CD36 and apoE expression were shown to have significantly less atherosclerosis than mice deficient in apoE expression alone, when they were examined at 12 weeks.⁸ That important study clearly demonstrated a significant role for CD36 in in vivo foam cell formation. However, the accompanying editorial pointed out that CD36 deficiency in a human Japanese population appears to be associated with an increased risk of coronary heart disease.⁹ Although the latter observation cannot be definitive for a cause-and-effect relationship in humans, how could a potential discrepancy such as this be accommodated? As suggested by de Winther and Hofker,⁹ it remains plausible that scavenger receptor activity and uptake of modified lipoproteins by macrophages may be beneficial in the setting of less potent and acute atherogenic influences.

Finally, even with appropriate precautions and controls, the pleiotropic effects of altering the expression of specific genes may always give rise to unexpected phenotypes in genetically engineered mice. For example, in the present study of Babaev et al,⁴ there was the unexpected observation that mice reconstituted with MSR-A^-/- macrophages had significant increases in body weight compared with those reconstituted with MSR-A^+/+ macrophages in response to a high fat diet. The authors hypothesize that this could be due to the redirection of modified LDL (shown to be a ligand for the peroxisome proliferator–activated receptor- [PPAR-]) to adipocytes, where it could stimulate adipogenesis. However, this very reasonable hypothesis leads to another hypothesis that directly bears on the reduction of atherosclerosis shown in the present study of Babaev et al. It has recently been shown that treatment of LDL receptor–deficient mice on a high fat diet with PPAR- ligands substantially suppresses atherosclerosis.¹⁰ Approximately 60% to 80% reductions in lesion area were observed in male mice, and a direct effect of PPAR- ligands on the artery wall was considered a potential mechanism for this substantial antiatherogenic result. Could accumulation of ligands for PPAR- in the vessel wall of MSR-A^-/- mice, or even CD36^-/- mice, have contributed to the suppression of atherosclerosis seen in these animals?

References

1. Suzuki H, Kurihara Y, Takeya M, Kamada N, Kataoka M, Jishage K, Ueda O, Sakaguchi H, Higashi T, Suzuki T, et al. A role for macrophage scavenger receptors in atherosclerosis and susceptibility to infection. Nature. 1997;386:292–286.[Medline] [Order article via Infotrieve]
2. Sakaguchi H, Takeya M, Suzuki H, Hakamata H, Kodama T, Horiuchi S, Gordon S, van der Laan LJW, Kraal G, Ishibashi S, et al. Role of macrophage scavenger receptors in diet-induced atherosclerosis in mice. Lab Invest. 1998;78:423–434.[Medline] [Order article via Infotrieve]
3. de Winther MPJ, Gijbels MJJ, van Dijk KW, van Gorp PJJ, Suzuki H, Kodama T, Frants RR, Havekes LM, Hofker MH. Scavenger receptor deficiency leads to more complex atherosclerotic lesions in APOE3Leiden transgenic mice. Atherosclerosis. 1999;144:315–321.[Medline] [Order article via Infotrieve]
4. Babaev VR, Gleaves LA, Carter KJ, Suzuki H, Kodama T, Fazio S, Linton MF. Reduced atherosclerotic lesions in mice deficient for total or macrophage-specific expression of scavenger receptor A. Arterioscler Thromb Vasc Biol. 2000;20:2593–2599.[Abstract/Free Full Text]
5. Paigen B, Morrow A, Brandon C, Mitchell D, Holmes P. Variation in susceptibility to atherosclerosis among inbred strains of mice. Atherosclerosis. 1985;57:65–73.[Medline] [Order article via Infotrieve]
6. Sigmund CD. Viewpoint: are studies in genetically altered mice out of control? Arterioscler Thromb Vasc Biol. 2000;20:1425–1429.[Abstract/Free Full Text]
7. Dansky HM, Charlton SA, Sikes JL, Heath SC, Simantov R, Levin LF, Shu P, Moore KJ, Breslow JL, Smith JD. Genetic background determines the extent of atherosclerosis in apoE-deficient mice. Arterioscler Thromb Vasc Biol. 1999;19:1960–1968.[Abstract/Free Full Text]
8. Febbraio M, Podrez EA, Smith JD, Hajjar DP, Hazen SL, Hoff HF, Sharma K, Silverstein RL. Targeted disruption of the class B scavenger receptor CD36 protects against atherosclerotic lesion development in mice. J Clin Invest. 2000;105:1049–1056.[Medline] [Order article via Infotrieve]
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10. Li AC, Brown KK, Silvestre MJ, Willson TM, Palinski W, Glass CK. Peroxisome proliferator-activated receptor ligands inhibit development of atherosclerosis in LDL receptor-deficient mice. J Clin Invest. 2000;106:523–531.[Medline] [Order article via Infotrieve]

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