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Monitor Foxp3 cells in vivo without interfering with regulatory T cell function |
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Foxp3 is a regulatory T cell-specific transcription factor that functions as the master regulator of the development and function of regulatory T cells.
The Foxp3-IRES-mRFP (FIR) mouse model:
- displays a normal expression of the endogenous Foxp3 gene
- displays an intact function of regulatory T cells
- allows an optimum monitoring of regulatory T cells trafficking and cell sorting
Features of the Foxp3-IRES-mRFP (FIR) mouse model:
- The reporter gene was inserted in the 3’UTR of the endogenous Foxp3 locus
- Endogenous Foxp3 coding sequence is not altered
- Reporter gene is expressed when Foxp3 is expressed
- Reporter gene minors Foxp3 expression pattern and level
- Foxp3 expressing cells are detectable by monitoring mRFP expression
mRFP expression correlates with Foxp3-expressing T cells:
Figure 2: mRFP marker faithfully marks Foxp3-expressing T cells without compromising their regulatory activity. (A) Foxp3 expression was detected in peripheral lymphocytes from Foxp3-IRES-mRFP mice harvested and stained with fluorophore-conjugated anti-CD4 and anti-CD25 antibodies. mRFP expression in CD4 T cells was monitored by flow cytometry (B) CD4+mRFP+ suppressor (S) and CD4+CD25-mRFP- responder (R) T cells were purified by FACS. Suppressor and responder cells were either cultured alone or mixed at indicated ratios (R:S), whereas the number of responder cells remained the same. T cells were activated by soluble anti-CD3 and anti-CD28 antibodies in the presence of irradiated APCs. Three days after stimulation, T cell proliferation was measured by a [3H]thymidine incorporation assay.
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mRFP expression correlates with Foxp3-expressing T cells in vivo:
Figure 3: mRFP marker allows the monitoring of Foxp3-expressing cells generated in vivo: CD4+Foxp3- T cells are not converted into CD4+Foxp3+ cells in the immunodeficient hosts after adoptive transfer. CD25-Foxp3- (CD4) peripheral T cells and Foxp3- bone marrow cells (BM) were purified by FACS and then transferred into multiple sublethally irradiated Rag-deficient syngeneic hosts. At various time points after transfer, recipient mice were killed, and the peripheral T cells were harvested and stained with fluorophore-conjugated anti-CD4 antibodies; the percentage of Foxp3+ cells among CD4 T cells was determined by flow cytometry, and combined results were plotted with each dot represents one recipient mouse.
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- Genetic background is C57BL/6
- Mice delivered are SOPF / VAF+ (Specific and Opportunistic Pathogen Free/Virus and Antibody Free conditions) certified by Charles River Laboratories
- Worldwide delivery using the logistic network of Charles River Laboratories
- Wan et al. PNAS 2005
- Kamanaka et al, Immunity 2006
- Li et al, Immunity 2007
- Wan et al, Nature 2007
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