Dr. Henk-Jan van Manen Profile

Dr. Henk-Jan van Manen

at Univ Twente

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Proceedings Article | 27 February 2006 Paper

Raman microscopy of phagocytosis: shedding light on macrophage foam cell formation

Henk-Jan van Manen, Aart van Apeldoorn, Dirk Roos, Cees Otto

Proceedings Volume 6093, 60930I (2006) https://doi.org/10.1117/12.674406

KEYWORDS: Raman spectroscopy, Foam, Microscopy, Confocal microscopy, Spectroscopy, Microorganisms, Proteins, Electron transport, Iron, Imaging spectroscopy

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The phagocyte NADPH oxidase is a crucial enzyme in the innate immune response of leukocytes against invading microorganisms. The superoxide (O₂^-) that is generated by this enzyme upon infection is directly and indirectly used in bacterial killing. The catalytic subunit of NADPH oxidase, the membrane-bound protein heterodimer flavocytochrome b₅₅₈, contains two heme moieties. Here, we first briefly discuss our recent confocal resonant Raman (RR) spectroscopy and microscopy experiments on flavocytochrome b₅₅₈ in both resting and phagocytosing neutrophilic granulocytes. Such experiments allow the determination of the redox state of flavocytochrome b₅₅₈ inside the cell, which directly reflects the electron transporting activity of NADPH oxidase. Subsequently, we report that incubation of murine RAW 264.7 macrophages with PolyActive microspheres for 1 week in culture medium leads to morphological and biochemical changes in the macrophages that are characteristic for the generation of macrophage-derived foam cells. Lipid-laden foam cells are the hallmark of early atherosclerotic lesions. Using nonresonant Raman spectroscopy and microscopy, we demonstrate that the numerous intracellular droplets in macrophages exposed to microspheres are rich in cholesteryl esters. The finding that phagocytic processes may trigger foam cell formation reinforces the current belief that (chronic) infection and inflammation are linked to the initiation and progression of atherosclerotic lesions. The study of such a connection may reveal new therapeutic targets for atherosclerosis treatment or prevention.

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