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|Title:||The effect of MPO modified LDL on M1/M2 THP-1 macrophage repolarization in vitro||Authors:||Frangie, Christian||Advisors:||Daher, Jalil||Keywords:||Atherosclerosis, macrophages, oxidized LDL, Mox-LDL, THP-1, PMA-induced differentiation, repolarization, CD11b, CD11c CD80, CD209, IL-6, IL-10||Subjects:||Atherosclerosis
University of Balamand--Dissertations
Background: Atherosclerosis is a chronic inflammatory disorder characterized by the accumulation of fibrous elements, lipids, and inflammatory cells inside the intimal layer of arteries. Foam cells which are macrophages that have ingested modified LDL particles play a major role in the development of atherosclerosis. Oxidized low density lipoprotein (ox-LDL) is one of most renowned post translational modifications of LDL. Myeloperoxidase targets and oxidizes LDL in vivo to form myeloperoxidase-modified LDL(Mox-LDL). Studies suggest that Mox‑LDLs is a pathophysiological model of LDL oxidation and it may play a key role in the development of atherosclerosis. Classically activated (M1) and alternatively activated (M2) macrophages are both implicated in the process of atherogensis. Macrophages are known to have the ability of repolarization, which is characterized by their capability to change phenotypic polarized sates. Ox-LDL was shown to have either a pro or anti- inflammatory effects on macrophages. However, the relationship between Mox-LDL and macrophage polarization and repolarization in vitro is still poorly understood. Aim: Our study intended to explore the effects of Mox-LDL on the repolarization of macrophages in vitro, by making use of the THP-1 cell line model.
Methods: THP-1 monocytes were cultured and differentiated into macrophages by treatment with PMA in vitro. CD11c and CD11b surface expression were analyzed to confirm PMA-induced differentiation. Afterwards resting macrophages(M0) were polarized toward M1 and M2 macrophages(MΦs), and then treated with Mox-LDL. ELISA assays for IL-6 and IL-10 cytokines were performed to assess their levels in the culture supernatants of the different conditions. Surface expression of CD80 and CD209 on the different THP-1 derived MΦ types was determined by flow cytometry analysis.
Results: The surface expression of CD11b and CD11c was significantly upregulated upon differentiation of THP-1 monocytes into MΦs (p<0.01), thereby confirming differentiation. Regarding CD80 surface expression there was no statistically significant difference among all the conditions when compared with each other; however, there was a trend toward significance when comparing M1-MΦ vs. M2-MΦ, M1-MΦ vs. M2/Mox-MΦ, M1/Mox-MΦ vs. M2-MΦ, M1/Mox-MΦ vs. M2/Mox-MΦ, M0-MΦ vs. M1-MΦ, M0-MΦ vs. M1/Mox-MΦ (p ~ 0.05). Meanwhile and as expected, stimulation of THP-1 M0-MΦs with M2 polarizing conditions resulted in a significant increase in the surface expression of CD209 (p<0.01). Mox-LDL treatment did not significantly alter the surface expression of CD80 and CD209. Stimulation of THP-1 M0-MΦs with M1 polarizing conditions resulted in a significant increase in IL-6 secretion (p<0.0001). Unexpectedly, IL-10 secretion was also significantly upregulated in M1-MΦs in comparison with M0-MΦs, M1/Mox-MΦs, M2-MΦs, and M2/Mox-MΦ (p<0.01). Overall, treatment of THP-1 M1 MΦs with Mox-LDL had a significant negative effect on IL-10 cytokine secretion.
Conclusion: Our data point to a potential role for Mox-LDL in increasing the pro-inflammatory state in macrophages by reducing the anti-inflammatory cytokine IL-10. However, further studies need to be conducted in order to gain a better insight onto this phenomenon; this would hopefully pave the way for future investigations that could provide us with a better understanding of the role of Mox-LDL in macrophage biology and the development of atherosclerosis.
Includes bibliographical references (p. 57-70)
|URI:||https://scholarhub.balamand.edu.lb/handle/uob/5057||Rights:||This object is protected by copyright, and is made available here for research and educational purposes. Permission to reuse, publish, or reproduce the object beyond the personal and educational use exceptions must be obtained from the copyright holder||Type:||Thesis|
|Appears in Collections:||UOB Theses and Projects|
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