Abstract
BackgroundMacrophages play pivotal roles in the progression of atherosclerosis (AS) and their heterogeneous differentiation patterns have been studied extensively. The classical subtype of activated macrophage, M1, promotes the progression of AS. Conversely, the alternative subtype of activated macrophage, M2, is regarded as a repressor of AS. Homocysteine (Hcy) may influence macrophage subtype polarization both in vivo and in vitro. Homocysteinemia (HHcy) is an independent risk factor in coronary heart disease and the effect of Hcy on macrophage differentiation has not been studied until now.MethodsDifferent concentrations of Hcy in combination with a fixed concentration of lipopolysaccharide (LPS, 200 ng/mL) were used to treat RAW264.7 macrophages. Real-time PCR was used to detect and quantify RNA transcripts indicative of M1 and M2 differentiation. The efficacy and specificity for each chemical stimulant in inducing macrophage differentiation were also investigated. The M2 macrophages (anti-inflammatory subtype) induced using classical methods (IL-4, 10 ng/mL) were also treated with different concentrations of Hcy complemented with LPS. The synergistic effect of Hcy and LPS in the converting the M2 subtype to M1 was also studied.ResultsMacrophages can be induced to differentiate towards M1 by a combination of Hcy with LPS, with the strongest effect observed at an Hcy concentration of 50 μmol/L. After inducing macrophages to the M2 subtype using IL-4, treatment with both Hcy and LPS could elicit conversion from the M2 to M1 subtype.ConclusionCombined treatment with Hcy and LPS can induce the polarization of cultured RAW264.7 macrophages into the pro-inflammatory subtype, as well as promote subtype conversion from anti-inflammatory to pro-inflammatory.
Highlights
Macrophages play pivotal roles in the progression of atherosclerosis (AS) and their heterogeneous differentiation patterns have been studied extensively
The mRNA levels of M1 markers (IL-6, TNF-α, and inducible nitric oxide synthase (iNOS)) were up-regulated by the 12-hour treatment with different concentrations of Hcy combined with LPS. mRNA expression levels of these genes reached their maxima at an Hcy concentration of 50 μmol/L (Figure 1)
When cells were treated with 50 μmol/L Hcy and 200 ng/mL LPS, mRNA expression of IL-6, TNF-α, and iNOS peaked after 12, 24, and 24 hours, respectively (Figure 2)
Summary
Macrophages play pivotal roles in the progression of atherosclerosis (AS) and their heterogeneous differentiation patterns have been studied extensively. The classical subtype of activated macrophage, M1, promotes the progression of AS. The alternative subtype of activated macrophage, M2, is regarded as a repressor of AS. Homocysteine (Hcy) may influence macrophage subtype polarization both in vivo and in vitro. Homocysteinemia (HHcy) is an independent risk factor in coronary heart disease and the effect of Hcy on macrophage differentiation has not been studied until now. Mechanisms governing AS continue to draw extensive research interest. Macrophages are important in both the initiation and progression of AS [4,5]. Macrophages can differentiate into two subtypes (M1 and M2) in response to microenvironmental changes.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
