Abstract
Coronary artery disease, resulting from atherosclerosis, is the leading cause of death in the Western world. Most previous studies have subjected atherosclerotic arteries, a tissue of mixed cellular composition, to homogenization in order to identify the factors in plaque development, thereby obscuring information relevant to specific cell types. Because macrophage foam cells are critical mediators in atherosclerotic plaque advancement, we reasoned that performing gene analysis on those cells would provide specific insight in novel regulatory factors and potential therapeutic targets. We demonstrated for the first time in vascular biology that foam cell-specific RNA can be isolated by laser capture microdissection (LCM) of plaques. As expected, compared to whole tissue, a significant enrichment in foam cell-specific RNA transcripts was observed. Furthermore, because regression of atherosclerosis is a tantalizing clinical goal, we developed and reported a transplantation-based mouse model. This involved allowing plaques to form in apoE−/− mice and then changing the plaque’s plasma environment from hyperlipidemia to normolipidemia. Under those conditions, rapid regression ensued in a process involving emigration of plaque foam cells to regional and systemic lymph nodes. Using LCM, we were able to show that under regression conditions, there was decreased expression in foam cells of inflammatory genes, but an up-regulation of cholesterol efflux genes. Interestingly, we also found that increased expression of chemokine receptor CCR7, a known factor in dendritic cell migration, was required for regression. In conclusion, the LCM methods described in this chapter, which have already lead to a number of striking findings, will likely further facilitate the study of cell type-specific gene expression in animal and human plaques during various stages of atherosclerosis, and after genetic, pharmacologic, and environmental perturbations.
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References
Glass CK, Witztum JL (2001) Atherosclerosis: the road ahead. Cell 104:503–516
Smith JD, Trogan E, Ginsberg M, Grigaux C, Tian J, Miyata M (1995) Decreased atherosclerosis in mice deficient in both macrophage colony-stimulating factor (op) and apolipoprotein E. Proc Natl Acad Sci USA 92:8264–8268
Chong PH, Bachenheimer BS (2000) Current, new and future treatments in dyslipidaemia and atherosclerosis. Drugs 60:55–93
Brewer HB Jr (2000) The lipid-laden foam cell: an elusive target for therapeutic intervention. J Clin Invest 105:703–705
Plutzky J (1999) Atherosclerotic plaque rupture: emerging insights and opportunities. Am J Cardiol 84:15J–20J
Emmert-Buck MR, Bonner RF, Smith PD, Chuaqui RF, Zhuang Z, Goldstein SR et al (1996) Laser capture microdissection. Science 274:998–1001
Bonner RF, Emmert-Buck M, Cole K, Pohida T, Chuaqui R, Goldstein S, Liotta LA (1997) Laser capture microdissection: molecular analysis of tissue. Science 278:1481–1483
Ramprasad MP, Fischer W, Witztum JL, Sambrano GR, Quehenberger O, Steinberg D (1995) The 94- to 97-kDa mouse macrophage membrane protein that recognizes oxidized low density lipoprotein and phosphatidylserine-rich liposomes is identical to macrosialin, the mouse homologue of human CD68. Proc Natl Acad Sci USA 92:9580–9584
Ramprasad MP, Terpstra V, Kondratenko N, Quehenberger O, Steinberg D (1996) Cell surface expression of mouse macrosialin and human CD68 and their role as macrophage receptors for oxidized low density lipoprotein. Proc Natl Acad Sci USA 93:14833–14838
Trogan E, Choudhury RP, Dansky HM, Rong JX, Breslow JL, Fisher EA (2002) Laser capture microdissection analysis of gene expression in macrophages from atherosclerotic lesions of apolipoprotein E-deficient mice. Proc Natl Acad Sci USA 99:2234–2239
Reis ED, Li J, Fayad ZA, Rong JX, Hansoty D, Aguinaldo JG, Fallon JT, Fisher EA (2001) Dramatic remodeling of advanced atherosclerotic plaques of the apolipoprotein E-deficient mouse in a novel transplantation model. J Vasc Surg 34:541–547
Chereshnev I, Trogan E, Omerhodzic S, Itskovich V, Aguinaldo JG, Fayad ZA, Fisher EA, Reis ED (2003) Mouse model of heterotopic aortic arch transplantation. J Surg Res 111:171–176
Trogan E, Feig JE, Dogan S, Rothblat GH, Angeli V, Tacke F, Randolph GJ, Fisher EA (2006) Gene expression changes in foam cells and the role of chemokine receptor CCR7 during atherosclerosis regression in ApoE-deficient mice. Proc Natl Acad Sci USA 103:3781–3786
Breslow JL (1996) Mouse models of atherosclerosis. Science 272:685–688
Acknowledgments
This work was supported by NIH grant HL-084312 (E.A.F.) and by an NIH predoctoral fellowship AG-029748 (J.E.F.).
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Feig, J.E., Fisher, E.A. (2013). Laser Capture Microdissection for Analysis of Macrophage Gene Expression from Atherosclerotic Lesions. In: Freeman, L. (eds) Lipoproteins and Cardiovascular Disease. Methods in Molecular Biology, vol 1027. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60327-369-5_5
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DOI: https://doi.org/10.1007/978-1-60327-369-5_5
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