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Monocytic MKP-1 is a Sensor of the Metabolic Environment and Regulates Function and Phenotypic Fate of Monocyte-Derived Macrophages in Atherosclerosis

View Article: PubMed Central - PubMed

ABSTRACT

Diabetes promotes the S-glutathionylation, inactivation and subsequent degradation of mitogen-activated protein kinase phosphatase 1 (MKP-1) in blood monocytes, and hematopoietic MKP-1-deficiency in atherosclerosis-prone mice accelerates atherosclerotic lesion formation, but the underlying mechanisms were not known. Our aim was to determine the mechanisms through which MKP-1 deficiency in monocytes and macrophages promotes atherogenesis. Transplantation of MKP-1-deficient bone marrow into LDL-R−/− (MKP-1LeuKO) mice accelerated high-fat diet (HFD)-induced atherosclerotic lesion formation. After 12 weeks of HFD feeding, MKP-1LeuKO mice showed increased lesion size in both the aortic root (1.2-fold) and the aorta (1.6-fold), despite reduced plasma cholesterol levels. Macrophage content was increased in lesions of MKP-1LeuKO mice compared to mice that received wildtype bone marrow. After only 6 weeks on a HFD, in vivo chemotactic activity of monocytes was already significantly increased in MKP-1LeuKO mice. MKP-1 deficiency in monocytes and macrophages promotes and accelerates atherosclerotic lesion formation by hyper-sensitizing monocytes to chemokine-induced recruitment, predisposing macrophages to M1 polarization, decreased autophagy and oxysterol-induced cell death whereas overexpression of MKP-1 protects macrophages against metabolic stress-induced dysfunction. MKP-1 serves as a master-regulator of macrophage phenotype and function and its dysregulation by metabolic stress may be a major contributor to atherogenesis and the progression of atherosclerotic plaques.

No MeSH data available.


Metabolic priming and MKP-1 deficiency enhance macrophage M1 polarization and suppress the conversion of macrophages into an M2, inflammation-resolving phenotype.Peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages were polarized as follows: TNF-α plus IFN-γ stimulation to generate M1 macrophages, and IL-4 stimulation as a model of M2 polarization. Untreated peritoneal cells were considered unpolarized. Macrophage polarization was assessed by quantifying and comparing levels of the indicated mRNAs between wildtype and MKP-1−/−, and unpirmed and primed macrophages by RT-qPCR. Expression levels were determined using TaqMan® probes in conjunction with the BioMarkTM HD Fluidigm System and Fluidigm Real-Time PCR Analysis software. mRNA levels were normalized to a housekeeping gene (Hprt) as well as unprimed macrophages (Control), and the 2−ΔΔCt for each mRNA is reported (A+C). Results are shown as mean of 4 independent samples. STAT1 phosphorylation were assessed with peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages (B+D). Results are shown as mean ± SE (n = 3–4).
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f5: Metabolic priming and MKP-1 deficiency enhance macrophage M1 polarization and suppress the conversion of macrophages into an M2, inflammation-resolving phenotype.Peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages were polarized as follows: TNF-α plus IFN-γ stimulation to generate M1 macrophages, and IL-4 stimulation as a model of M2 polarization. Untreated peritoneal cells were considered unpolarized. Macrophage polarization was assessed by quantifying and comparing levels of the indicated mRNAs between wildtype and MKP-1−/−, and unpirmed and primed macrophages by RT-qPCR. Expression levels were determined using TaqMan® probes in conjunction with the BioMarkTM HD Fluidigm System and Fluidigm Real-Time PCR Analysis software. mRNA levels were normalized to a housekeeping gene (Hprt) as well as unprimed macrophages (Control), and the 2−ΔΔCt for each mRNA is reported (A+C). Results are shown as mean of 4 independent samples. STAT1 phosphorylation were assessed with peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages (B+D). Results are shown as mean ± SE (n = 3–4).

Mentions: In atherosclerotic lesions, as in other inflamed tissues, the microenvironment plays a key role in determining the activation state of macrophages. Cytokines are an important constituent of the inflammatory milieu and drive macrophage activation either toward a classical (M1, proinflammatory) or alternative (M2, anti-inflammatory) polarization state3. The M1 macrophage phenotype is controlled by NF-κB, signal transducer and activator of transcription 1 (STAT1) and interferon-regulatory factor 5 (IRF5)3233, but STAT1 activity is essential for macrophages to polarize into an M1 phenotype3334. Phosphorylation of STAT1 at Ser727 is required for its activation3536. P38 MAPK is required for STAT1 phosphorylation and transcriptional activation induced by interferons3537. Interestingly, STAT1 has been suggested as a physiological non-MAPK substrate for MKP-13839. We used real-time PCR in custom-designed microfluidic dynamic arrays4041 to explore how MKP-1 deficiency affects macrophage polarization into M1 or M2 activation states. We found that MKP-1 deficiency enhanced interferon gamma+ tumor necrosis factor alpha (INFγ + TNFα)-induced polarization of macrophages toward an M1 phenotype and suppressed the interleukin-4 (IL-4)-induced conversion of macrophages into an M2 phenotype (Fig. 5A). STAT1 phosphorylation was increased by 53% in MKP-1-deficient macrophages as compared to wildtype cells (Fig. 5B), confirming that STAT1, at least in macrophages, is indeed a physiological non-MAPK substrate for MKP-1.


Monocytic MKP-1 is a Sensor of the Metabolic Environment and Regulates Function and Phenotypic Fate of Monocyte-Derived Macrophages in Atherosclerosis
Metabolic priming and MKP-1 deficiency enhance macrophage M1 polarization and suppress the conversion of macrophages into an M2, inflammation-resolving phenotype.Peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages were polarized as follows: TNF-α plus IFN-γ stimulation to generate M1 macrophages, and IL-4 stimulation as a model of M2 polarization. Untreated peritoneal cells were considered unpolarized. Macrophage polarization was assessed by quantifying and comparing levels of the indicated mRNAs between wildtype and MKP-1−/−, and unpirmed and primed macrophages by RT-qPCR. Expression levels were determined using TaqMan® probes in conjunction with the BioMarkTM HD Fluidigm System and Fluidigm Real-Time PCR Analysis software. mRNA levels were normalized to a housekeeping gene (Hprt) as well as unprimed macrophages (Control), and the 2−ΔΔCt for each mRNA is reported (A+C). Results are shown as mean of 4 independent samples. STAT1 phosphorylation were assessed with peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages (B+D). Results are shown as mean ± SE (n = 3–4).
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Related In: Results  -  Collection

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f5: Metabolic priming and MKP-1 deficiency enhance macrophage M1 polarization and suppress the conversion of macrophages into an M2, inflammation-resolving phenotype.Peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages were polarized as follows: TNF-α plus IFN-γ stimulation to generate M1 macrophages, and IL-4 stimulation as a model of M2 polarization. Untreated peritoneal cells were considered unpolarized. Macrophage polarization was assessed by quantifying and comparing levels of the indicated mRNAs between wildtype and MKP-1−/−, and unpirmed and primed macrophages by RT-qPCR. Expression levels were determined using TaqMan® probes in conjunction with the BioMarkTM HD Fluidigm System and Fluidigm Real-Time PCR Analysis software. mRNA levels were normalized to a housekeeping gene (Hprt) as well as unprimed macrophages (Control), and the 2−ΔΔCt for each mRNA is reported (A+C). Results are shown as mean of 4 independent samples. STAT1 phosphorylation were assessed with peritoneal macrophages from wildtype (WT) or MKP-1−/− (KO) mice, and unprimed (Control) and metabolically primed (LDL + HG) peritoneal macrophages (B+D). Results are shown as mean ± SE (n = 3–4).
Mentions: In atherosclerotic lesions, as in other inflamed tissues, the microenvironment plays a key role in determining the activation state of macrophages. Cytokines are an important constituent of the inflammatory milieu and drive macrophage activation either toward a classical (M1, proinflammatory) or alternative (M2, anti-inflammatory) polarization state3. The M1 macrophage phenotype is controlled by NF-κB, signal transducer and activator of transcription 1 (STAT1) and interferon-regulatory factor 5 (IRF5)3233, but STAT1 activity is essential for macrophages to polarize into an M1 phenotype3334. Phosphorylation of STAT1 at Ser727 is required for its activation3536. P38 MAPK is required for STAT1 phosphorylation and transcriptional activation induced by interferons3537. Interestingly, STAT1 has been suggested as a physiological non-MAPK substrate for MKP-13839. We used real-time PCR in custom-designed microfluidic dynamic arrays4041 to explore how MKP-1 deficiency affects macrophage polarization into M1 or M2 activation states. We found that MKP-1 deficiency enhanced interferon gamma+ tumor necrosis factor alpha (INFγ + TNFα)-induced polarization of macrophages toward an M1 phenotype and suppressed the interleukin-4 (IL-4)-induced conversion of macrophages into an M2 phenotype (Fig. 5A). STAT1 phosphorylation was increased by 53% in MKP-1-deficient macrophages as compared to wildtype cells (Fig. 5B), confirming that STAT1, at least in macrophages, is indeed a physiological non-MAPK substrate for MKP-1.

View Article: PubMed Central - PubMed

ABSTRACT

Diabetes promotes the S-glutathionylation, inactivation and subsequent degradation of mitogen-activated protein kinase phosphatase 1 (MKP-1) in blood monocytes, and hematopoietic MKP-1-deficiency in atherosclerosis-prone mice accelerates atherosclerotic lesion formation, but the underlying mechanisms were not known. Our aim was to determine the mechanisms through which MKP-1 deficiency in monocytes and macrophages promotes atherogenesis. Transplantation of MKP-1-deficient bone marrow into LDL-R−/− (MKP-1LeuKO) mice accelerated high-fat diet (HFD)-induced atherosclerotic lesion formation. After 12 weeks of HFD feeding, MKP-1LeuKO mice showed increased lesion size in both the aortic root (1.2-fold) and the aorta (1.6-fold), despite reduced plasma cholesterol levels. Macrophage content was increased in lesions of MKP-1LeuKO mice compared to mice that received wildtype bone marrow. After only 6 weeks on a HFD, in vivo chemotactic activity of monocytes was already significantly increased in MKP-1LeuKO mice. MKP-1 deficiency in monocytes and macrophages promotes and accelerates atherosclerotic lesion formation by hyper-sensitizing monocytes to chemokine-induced recruitment, predisposing macrophages to M1 polarization, decreased autophagy and oxysterol-induced cell death whereas overexpression of MKP-1 protects macrophages against metabolic stress-induced dysfunction. MKP-1 serves as a master-regulator of macrophage phenotype and function and its dysregulation by metabolic stress may be a major contributor to atherogenesis and the progression of atherosclerotic plaques.

No MeSH data available.