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Isolation and Characterization of Dendritic Cells and Macrophages from the Mouse Intestine
Published on: May 21, 2012
Corticotropin-releasing Factor Changes the Phenotype and Function of Dendritic Cells in Mouse Mesenteric Lymph Nodes
Li Meng1, Zhang Lu1, Wang Xiaoteng1
1Department of Gastroenterology, First Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, China.
Insights
Corticotropin-releasing factor (CRF) influences intestinal dendritic cells (DCs) via CRF-R1 and CRF-R2 receptors. CRF-R1 activation reduces DC activation and T cell stimulation, while CRF-R2 activation has opposite effects.
Area of Science:
- Immunology
- Cell Biology
- Gastroenterology
Background:
- Dendritic cells (DCs) play a key role in chronic inflammatory autoimmune diseases.
- The impact of Corticotropin-releasing factor (CRF) on intestinal DCs is not well understood.
Purpose of the Study:
- To investigate the role of CRF in modulating the phenotype and function of intestinal DCs.
- To determine the specific roles of CRF receptor type 1 (CRF-R1) and CRF receptor type 2 (CRF-R2) in these alterations.
Main Methods:
- Mouse mesenteric lymph node dendritic cells (MLNDCs) were isolated and characterized for CRF receptor expression using immunofluorescence and qPCR.
- MLNDCs were treated with CRF in the presence or absence of CRF-R1 and CRF-R2 antagonists.
- DC activation markers (MHC-I, MHC-II, CD80, CD86) and T cell stimulatory capacity were assessed via flow cytometry, Western blot, and mixed lymphocyte reaction.
Main Results:
- Both CRF-R1 and CRF-R2 are expressed on MLNDCs.
- CRF exposure enhanced MLNDC expression of MHC-II and T cell stimulatory capacity.
- CRF-R1 antagonist treatment led to reduced DC activation and T cell stimulation, whereas CRF-R2 antagonist treatment showed opposite effects.
Conclusions:
- CRF directly alters intestinal DC phenotype and function through CRF-R1 and CRF-R2.
- Activation of CRF-R1 and CRF-R2 pathways results in opposing functional outcomes for intestinal DCs.
Background/Aims:
Dendritic cells (DCs) are a significant contributor to the pathology of numerous chronic inflammatory autoimmune disorders; however, the effects of Corticotropin-releasing factor (CRF) on intestinal DCs are poorly understood. In this study, we investigated the role of CRF in alterations of intestinal dendritic cell phenotype and function.
Methods:
Mouse mesenteric lymph node dendritic cells (MLNDCs) were obtained using magnetic bead sorting. Surface expression of CRF receptor type 1 (CRF-R1) and CRF-R2 was determined by double-labeling immunofluorescence and quantitative polymerase chain reaction (qPCR) and MLNDCs were subsequently exposed to CRF in the presence or absence of CRF-R1 and CRF-R2 antagonists. Expression of surface molecules (MHC-I and MHC-II) and co-stimulatory molecules (CD80 and CD86) was determined by flow cytometric and western blot analyses, and the T cell stimulatory capacity of MLNDCs was evaluated by mixed lymphocyte reaction.
Results:
Immunofluorescent staining and quatitative polymerase chain reaction indicated that both the CRF receptors (CRF-R1 and CRF-2) are expressed on the surface of MLNDCs. Exposure to CRF increased the expression of MHC-II on MLNDCs as well as their capacity to stimulate T cell proliferation. MLNDCs treated with CRF-R1 antagonist exhibited a phenotype characterized by a less activated state and reduced surface expression of MHC-II, and consequently showed reduced capacity to stimulate T cells. In contrast, treatment of MLNDCs with CRF-R2 antagonist yielded an opposite result.
Conclusions:
CRF can alter the phenotype and function of intestinal DCs through direct action on CRF-R1 and CRF-R2, and activation of the CRF-R1 and CRF-R2 pathways yields opposing outcomes.
