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Case report: Mafb promoter activity may define the alveolar macrophage dichotomy
1Department of Comparative Biomedical Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA, United States.
Abstract:
Cre-LoxP system has been widely used to induce recombination of floxed genes of interest. Currently available macrophage promoter-specific Cre recombinase mice strains have various limitations that warrants the testing of additional Cre strains. V-maf musculoaponeurotic fibrosarcoma oncogene family, protein b -Cre (Mafb-Cre) mice label macrophages in most organs such as spleen, small intestine, lung, bone marrow, and peritoneal cavity. However, whether Mafb-Cre recombinase targets the gene recombination in alveolar macrophage remains untested. Here, we utilized Mafb strain that expresses mTOM protein in all the cells of mouse body except for those that express Mafb-Cre-regulated mEGFP. We performed fluorescent microscopy and flow cytometry to analyze mTOM and mEGFP expression in alveolar macrophages from Mafb mice. Our analyses revealed that the Mafb-Cre is active in only ~40% of the alveolar macrophages in an age-independent manner. While Mafb- (mTOM+/mEGFP-) and Mafb+ (mEGFP+) alveolar macrophages exhibit comparable expression of CD11b and CD11c surface markers, the surface expression of MHCII is elevated in the Mafb+ (mEGFP+) macrophages. The bone marrow-derived macrophages from Mafb mice are highly amenable to Cre-LoxP recombination in vitro. The bone marrow depletion and reconstitution experiment revealed that ~98% of alveolar macrophages from Mafb → WT chimera are amenable to the Mafb-Cre-mediated recombination. Finally, the Th2 stimulation and ozone exposure to the Mafb mice promote the Mafb-Cre-mediated recombination in alveolar macrophages. In conclusion, while the Mafb-/Mafb+ dichotomy thwarts the use of Mafb-Cre for the induction of floxed alleles in the entire alveolar macrophage population, this strain provides a unique tool to induce gene deletion in alveolar macrophages that encounter Th2 microenvironment in the lung airspaces.
Insights
The Mafb-Cre mouse line shows activity in approximately 40% of alveolar macrophages, offering a tool for targeted gene recombination in specific lung environments. This strain allows for conditional gene deletion in alveolar macrophages responding to Th2 stimulation.
Area of Science:
- Immunology
- Genetics
- Cell Biology
Background:
- The Cre-loxP system is crucial for conditional gene manipulation.
- Existing macrophage promoter-specific Cre recombinase mouse strains have limitations.
- The Mafb-Cre mouse strain labels macrophages across various organs, but its efficacy in alveolar macrophages was untested.
Observation:
- Mafb-Cre activity was assessed in alveolar macrophages using a reporter system (mTOM/mEGFP).
- Approximately 40% of alveolar macrophages showed Mafb-Cre activity, independent of age.
- Mafb-Cre+ alveolar macrophages exhibited elevated MHCII surface expression compared to Mafb-Cre- cells.
Findings:
- Bone marrow-derived macrophages from Mafb mice are amenable to in vitro Cre-LoxP recombination.
- Chimeric mouse experiments indicated high Mafb-Cre recombination potential in alveolar macrophages (~98%).
- Th2 stimulation and ozone exposure enhanced Mafb-Cre recombination in alveolar macrophages.
Implications:
- The Mafb-Cre strain allows for targeted gene deletion in a subset of alveolar macrophages.
- This tool is valuable for studying gene function in alveolar macrophages within specific lung microenvironments, particularly those associated with Th2 responses.
- The heterogeneity in Mafb-Cre activity within the alveolar macrophage population limits its use for targeting the entire population but enables specific subset analysis.
