In vitro and in vivo Repair Activities of Undifferentiated and Classically and Alternatively Activated Macrophages

Although it is well-established that macrophages can occur in distinct activation states, the molecular characteristics of differentially activated macrophages, and particularly those of alternatively activated macrophages (aaMφ), are still poorly unraveled. Recently, we demonstrated that the expression of FIZZ1 and Ym is induced in aaMφ as compared with classically activated macrophages (caMφ), elicitedin vitroor developedin vivoduring infection withTrypanosoma brucei brucei. In the present study, we analyzed the expression of FIZZ1 and Ym in caMφ and aaMφ elicited duringTrypanosoma congolenseinfection and show that the use of FIZZ1 and Ym for the identification of aaMφ is not limited toT. b. bruceiinfection and is independent of the organ sources from which macrophages are obtained. We also demonstrate that FIZZ1 can be used to discriminate between different populations of aaMφ. Furthermore, we studied the effects of various stimuli, and combinations thereof, on the expression of FIZZ1 and Ym in macrophages from different mouse strains and demonstrate that regulation of the expression of FIZZ1 and Ym in macrophages is not dependent on the mouse strain. Finally, we show that these genes can be used to monitor the macrophage activation status without the need to obtain pure macrophage populations.

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Interdependence between Chromogranin-A, Alternatively Activated Macrophages, Tight Junction Proteins and the Epithelial Functions. A Human and In-Vivo/In-Vitro Descriptive Study

International Journal of Molecular Sciences ◽

10.3390/ijms21217976 ◽

2020 ◽

Vol 21 (21) ◽

pp. 7976

Author(s):

Nour Eissa ◽

Hayam Hussein ◽

Diane M. Tshikudi ◽

Geoffrey N. Hendy ◽

Charles N. Bernstein ◽

...

Keyword(s):

Epithelial Cells ◽

Mrna Expression ◽

Chromogranin A ◽

Intestinal Inflammation ◽

Collagen Deposition ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Alternatively Activated

Background: Ulcerative colitis (UC) is characterized by altered chromogranin-A (CHGA), alternatively activated macrophages (M2) and intestinal epithelial cells (IECs). We previously demonstrated that CHGA is implicated in colitis progression by regulating the macrophages. Here, we investigated the interplay between CHGA, M2, tight junctions (TJ) and IECs in an inflammatory environment. Methods: Correlations between CHGA mRNA expression of and TJ proteins mRNA expressions of (Occludin [OCLN], zonula occludens-1 [ZO1], Claudin-1 [CLDN1]), epithelial associated cytokines (interleukin [IL]-8, IL-18), and collagen (COL1A2) were determined in human colonic mucosal biopsies isolated from active UC and healthy patients. Acute UC-like colitis (5% dextran sulphate sodium [DSS], five days) was induced in Chga-C57BL/6-deficient (Chga−/−) and wild type (Chga+/+) mice. Col1a2 TJ proteins, Il-18 mRNA expression and collagen deposition were determined in whole colonic sections. Naïve Chga−/− and Chga+/+ peritoneal macrophages were isolated and exposed six hours to IL-4/IL-13 (20 ng/mL) to promote M2 and generate M2-conditioned supernatant. Caco-2 epithelial cells were cultured in the presence of Chga−/− and Chga+/+ non- or M2-conditioned supernatant for 24 h then exposed to 5% DSS for 24 h, and their functional properties were assessed. Results: In humans, CHGA mRNA correlated positively with COL1A2, IL-8 and IL-18, and negatively with TJ proteins mRNA markers. In the experimental model, the deletion of Chga reduced IL-18 mRNA and its release, COL1A2 mRNA and colonic collagen deposition, and maintained colonic TJ proteins. Chga−/− M2-conditioned supernatant protected caco-2 cells from DSS and oxidative stress injuries by improving caco-2 cells functions (proliferation, viability, wound healing) and by decreasing the release of IL-8 and IL-18 and by maintaining the levels of TJ proteins, and when compared with Chga+/+ M2-conditioned supernatant. Conclusions: CHGA contributes to the development of intestinal inflammation through the regulation of M2 and epithelial cells. Targeting CHGA may lead to novel biomarkers and therapeutic strategies in UC.

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113 Type I and III Interferon Are Attenuated in a Human In Vitro Model of Alternatively Activated Macrophages

World Allergy Organization Journal ◽

10.1097/01.wox.0000411858.75911.af ◽

2012 ◽

Vol 5 ◽

pp. S37-S38

Author(s):

Ashraf EL Fiky

Keyword(s):

In Vitro Model ◽

Type I ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Vitro Model ◽

Alternatively Activated

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Mesenchymal Stem Cells Convert Human Macrophages to a Novel Type of Alternatively Activated Macrophages.

Blood ◽

10.1182/blood.v114.22.3632.3632 ◽

2009 ◽

Vol 114 (22) ◽

pp. 3632-3632 ◽

Cited By ~ 1

Author(s):

Jaehyup Kim ◽

Peiman Hematti

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Cell Surface ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Tnf Α ◽

Low Levels ◽

Alternatively Activated

Abstract Abstract 3632 Poster Board III-568 Mesenchymal stem cells (MSCs) are capable of modulating the immune system through interaction with a wide range of immune cells. This study investigates the hypothesis that interaction of MSCs with macrophages could play a significant role in their anti-inflammatory/immune modulatory effects. All studies were approved by IRB of University of Wisconsin School of Medicine and Public Health. MSCs were culture expanded from discarded bone marrow filters after bone marrow harvest from normal healthy sibling HLA-matched donors. We used passages -35 for our experiments. Ex vivo culture expanded MSCs were characterized by their cell surface phenotype (positive for MSC markers: CD29, CD44, CD73, CD90, and CD105; and negative for hematopoietic markers: CD31, CD34, and CD45), and their differentiation potential into bone, fat and cartilage. Monocytes were isolated from peripheral blood mononuclear cell fraction of healthy donors using CD14+ Miltenyi magnetic bead cell separation method. We cultured human CD14+ monocytes for seven days without any added cytokines to generate macrophages, and then co-cultured them for three more days with culture-expanded MSCs. We used cell surface antigen expression and intracellular cytokine expression patterns to study the immunophenotype of macrophages at the end of this co-culture period, and phagocytic assays to investigate their functional activity in vitro. Macrophages co-cultured with MSCs consistently showed high level expression of CD206, a marker of alternatively activated macrophages, in addition to being positive fro CD14 marker. Using CD1a and CD209 staining we did not detect presence of any dendritic cells either at the end of seven days culture of monocyte-derived macrophages or at the end of co-culture period. Furthermore, macrophages that were co-cultured with MSCs expressed high levels of IL-10 and low levels of IL-12, as determined by intracellular staining, typical of alternatively activated macrophages. However, macrophages co-cultured with MSCs also expressed high levels of IL-6 and low levels of TNF-α, compared to controls. Functionally, macrophages co-cultured with MSCs showed a higher level of phagocytic activity using Alexa 488-conjugated E. coli phagocytic assay. In summary we describe a novel type of human macrophage generated in vitro after co-culture with MSCs that assume an immunophenotype defined as IL-10 high, IL-12 low, IL-6 high and TNF-α low secreting cells. These MSC-educated macrophages may be a unique and novel type of alternatively activated macrophages with potentially significant role in tissue repair. Disclosures: No relevant conflicts of interest to declare.

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Pancreastatin Reduces Alternatively Activated Macrophages, Disrupts the Epithelial Homeostasis and Aggravates Colonic Inflammation. A Descriptive Analysis

Biomedicines ◽

10.3390/biomedicines9020134 ◽

2021 ◽

Vol 9 (2) ◽

pp. 134

Author(s):

Nour Eissa ◽

Omar Elgazzar ◽

Hayam Hussein ◽

Geoffrey N. Hendy ◽

Charles N. Bernstein ◽

...

Keyword(s):

Epithelial Cells ◽

Mrna Expression ◽

Descriptive Analysis ◽

Rectal Administration ◽

Mrna Levels ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Epithelial Homeostasis ◽

Alternatively Activated

Ulcerative colitis (UC) is characterized by modifying alternatively activated macrophages (AAM) and epithelial homeostasis. Chromogranin-A (CHGA), released by enterochromaffin cells, is elevated in UC and is implicated in inflammation progression. CHGA can be cleaved into several derived peptides, including pancreastatin (PST), which is involved in proinflammatory mechanisms. Previously, we showed that the deletion of Chga decreased the onset and severity of colitis correlated with an increase in AAM and epithelial cells’ functions. Here, we investigated PST activity in colonic biopsies of participants with active UC and investigated PST treatment in dextran sulfate sodium (DSS)-induced colitis using Chga−/− mice, macrophages, and a human colonic epithelial cells line. We found that the colonic protein expression of PST correlated negatively with mRNA expression of AAM markers and tight junction (TJ) proteins and positively with mRNA expression of interleukin (IL)-8, IL18, and collagen in human. In a preclinical setting, intra-rectal administration of PST aggravated DSS-induced colitis by decreasing AAM’s functions, enhancing colonic collagen deposition and disrupting epithelial homeostasis in Chga+/+ and Chga−/− mice. This effect was associated with a significant reduction in AAM markers, increased colonic IL-18 release, and decreased TJ proteins’ gene expression. In vitro, PST reduced Chga+/+ and Chga−/− AAM polarization and decreased anti-inflammatory mediators’ production. Conditioned medium harvested from PST-treated Chga+/+ and Chga−/− AAM reduced Caco-2 cell migration, viability, proliferation, and mRNA levels of TJ proteins and increased oxidative stress-induced apoptosis and proinflammatory cytokines release. In conclusion, PST is a CHGA proinflammatory peptide that enhances the severity of colitis and the inflammatory process via decreasing AAM functions and disrupting epithelial homeostasis.

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In Vitro-Derived Alternatively Activated Macrophages Reduce Colonic Inflammation in Mice

Gastroenterology ◽

10.1053/j.gastro.2009.12.041 ◽

2010 ◽

Vol 138 (4) ◽

pp. 1395-1405 ◽

Cited By ~ 182

Author(s):

Meaghan M. Hunter ◽

Arthur Wang ◽

Kuljit S. Parhar ◽

Michael J.G. Johnston ◽

Nico Van Rooijen ◽

...

Keyword(s):

Activated Macrophages ◽

Colonic Inflammation ◽

Alternatively Activated Macrophages ◽

Alternatively Activated

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Alternatively activated macrophages actively inhibit proliferation of peripheral blood lymphocytes and CD4 + T cells in vitro

Immunology ◽

10.1046/j.1365-2567.1997.00371.x ◽

1997 ◽

Vol 92 (4) ◽

pp. 478-486 ◽

Cited By ~ 86

Author(s):

C. SCHEBESCH ◽

V. KODELJA ◽

C. MÜLLER ◽

N. HAKIJ ◽

S. BISSON ◽

...

Keyword(s):

T Cells ◽

Peripheral Blood ◽

Cd4 T Cells ◽

Peripheral Blood Lymphocytes ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Blood Lymphocytes ◽

Alternatively Activated

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Identification of Caspase-6 as a New Regulator of Alternatively Activated Macrophages

Journal of Biological Chemistry ◽

10.1074/jbc.m116.717868 ◽

2016 ◽

Vol 291 (33) ◽

pp. 17450-17466 ◽

Cited By ~ 7

Author(s):

Yongfang Yao ◽

Qian Shi ◽

Bing Chen ◽

Qingsong Wang ◽

Xinda Li ◽

...

Keyword(s):

Cell Invasion ◽

Protein Composition ◽

Alternative Activation ◽

Activated Macrophages ◽

Alternatively Activated Macrophages ◽

Protein Levels ◽

Caspase 6 ◽

Alternatively Activated

Alternatively activated macrophages (AAMs) play essential roles in the promotion of tissue remodeling, vasculogenesis, and tumor progression; however, the detailed mechanisms underlying the activation of AAMs remain largely unknown. Here, by using quantitative proteomic analysis, we identified 62 proteins that were up-regulated in IL-4-induced macrophages. Among these, Caspase-6 was increased significantly. Caspase-6 is important in the apoptotic signaling pathway; however, its role in non-apoptosis is also reported. Here, we first examined the non-apoptotic role of Caspase-6 in the alternative activation of macrophages after administration of IL-4, 4T1 tumor conditional medium, or co-culture with 4T1 cells. Both treatments promoted alternative activation of RAW264.7 cells and primary macrophages, whereas disruption of caspase-6 expression and activity could markedly suppress the biomarker levels of AAMs. Overexpression of Caspase-6 could significantly promote the activation of AAMs. Importantly, we further present evidence that caspase-6 could regulate breast cancer cell invasion by modulating MMP-2 and MMP-9 expression in 4T1 tumor-associated macrophages, as ablation of protein levels or activity of caspase-6 suppressed tumor cell invasion in vitro. In conclusion, the observed results markedly expanded our views of the dynamic changes in protein composition during alternative activation of macrophages, and they revealed a critical new role of caspase-6 in regulating this cellular biological process, which suggested that caspase-6 might be a key nod molecule to regulate immunological steady-state and be a therapeutic candidate for tumor immunotherapy.

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