AbstractMacrophages are key inflammatory cells, which are involved in atherosclerotic lesion formation, evolution and resorption (Glass and Witztum, 2001; Pluddemann et al., 2007). Since about 15 years now, the concept of monocyte-macrophage polarization has been intensively studied in vitro (Hirose et al., 2011) and in vivo (Khallou-Laschet et al., 2010). Indeed, characterization of macrophage phenotype and functions can also help to characterize the inflammatory state of the lesions. M1 macrophages are pro-inflammatory, while M2 macrophages are anti-inflammatory. The macrophage phenotype can switch from M2 to M1 or vice-versa, because of changes of the microenvironment within the lesion (Khallou-Laschet et al., 2010). This switch can happen through the regulation of the expression of pro-inflammatory and anti-inflammatory genes via transcriptional and epigenetic mechanisms (Ivashkiv, 2013; Lawrence and Natoli, 2011; Takeuch and Akira, 2011).
In this study, we choose to work with the RAW 264.7 murine cell line, often used in culture in the context of atherosclerosis. We have started the study by setting up, evaluating and characterizing the polarization of RAW 264.7 macrophages by monitoring the expression of M1 and M2 marker genes at the mRNA and protein levels. Then, we checked whether M1 and M2 polarized cells were able to form foam cells, by incubating them with native and modified LDLs. We have shown that LDLs oxidized with myeloperoxidase (Mox-LDLs) seem to favour the M2 anti-inflammatory phenotype as well as the antioxidant phenotype, while also favouring the expression of some pro-inflammatory M1 genes. Moreover, these LDLs were the most efficiently engulfed by (non)-polarized macrophages. All these data have been validated on macrophages derived from bone marrow cells of C57BL/6 mice.
Afterwards, macrophage polarization has been studied within atherosclerotic lesions of the in vivo ApoE-/- x P2y6-/- mouse model. This work was carried out in collaboration with the laboratory of B. Robaye (IRIBHM, ULB, Gosselies). From this study we concluded that in this model, although lesions are clearly reduced compared to ApoE-/- mice, with a significant overall reduction in macrophages, there was no change in the ratio of M1 to M2 cells in the lesions.
Finally, in a more complex model, we have showed that patients undergoing haemodialysis, prone to accelerated atherosclerosis, display higher levels of M2 circulating monocytes as compared to healthy individuals. Furthermore, this increase in the percentage of M2 circulating monocytes is coupled with an increase of the M-CSF plasmatic concentration in these patients.
Further experiments are needed to identify the molecular actors involved in the modulation of monocyte-macrophage polarization, for instance in the presence of oxidized LDLs, within the atherosclerotic lesions.
|Date of Award||17 May 2017|
|Supervisor||Martine Raes (Supervisor), Karim Zouaoui Boudjeltia (Co-Supervisor), Yves Poumay (President), Carine MICHIELS (Jury), Philippe Lesnik (Jury) & Muriel Moser (Jury)|
Attachment to an Research Institute in UNAMUR