Ouisse, L.-H. | bioRxiv Science

bioRxiv2021

CD4+ and CD8+ regulatory T cells characterization in the rat using a unique transgenic Foxp3-EGFP model.

BackgroundCD4+ and CD8+ regulatory T cells (Treg) in diverse species include different subsets from different origins. In all species, CD8+ Treg have been poorly characterized. CD4+ and CD8+ Treg in rats have only partially been characterized and there is no rat model in which FOXP3+ Treg are genetically tagged. ResultsWe generated a rat transgenic line using the CRISPR/Cas9 system in which EGFP was inserted in frame on the 3 end of the Foxp3 gene using a 2A self-cleaving peptide. EGFP was exclusively expressed by CD4+ and CD8+ T cells in similar proportion as observed with anti-FOXP3 antibodies. CD4+EGFP+ Treg were 5-10 times more frequent than CD8+EGFP+ Treg. CD4+ and CD8+ EGFP+ Treg expressed both the CD25highCD127lowCD45RClow/- markers. The suppressive activity of CD4+ and CD8+ Treg was largely confined to EGFP+ cells. RNAseq analyses showed similarities but also differences among CD4+ and CD8+ EGFP+ cells and provided the first description of the natural FOXP3+ CD8+ Treg transcriptome. In vitro culture of CD4+ and CD8+ EGFP- cells with TGFbeta and IL-2 resulted in the induction of EGFP+ Treg. Preferential expansion of CD4+ and CD8+ EGFP+ Treg could be detected upon in vivo administration of a low dose of IL-2. ConclusionsThis new and unique Foxp3-EGFP rat line constitutes a useful model to identify and isolate viable natural and induced CD4+ and CD8+ Treg. Additionally, it allows to identify new molecules expressed in CD8+ Treg that may allow to better define their phenotype and function not only in rats but also in other species.

immunology↗

bioRxiv2018

Immunological characterization of a rat model of Duchenne’s disease and demonstration of improved muscle strength after anti-CD45RC antibody treatment

Duchenne muscular dystrophy (DMD) has as standard pharmacological therapy with corticoisteroids (CS) that decrease inflammation and immune responses present in patients and animal models. CS have however limited efficacy and important and numerous side effects. Therefore, there is a need for new anti-inflammatory and pro-tolerogenic treatments that could replace or decrease doses of CS. We first assessed the status of immune system of dystrophin-deficient rats (Dmdmdx) that closely reproduce the phenotype of DMD patients. Dmdmdx rats showed increased leukocyte infiltration in skeletal and cardiac muscles, containing mostly macrophages but also T cells, and increased expression of several cytokines. Anti-CD45RC Monoclonal antibody (Mab) treatment induced immune tolerance in models of organ transplantation and GVHD (Graft Versus Host Disease). We observed that muscles and blood of DMD patients contained T CD4+ and CD8+ expressing high levels of CD45RChigh cells. Treatment of young Dmdmdx rats with anti-CD45RC MAb corrected skeletal muscle strength associated to a depletion of effectors CD45RChigh T cells with no obvious side-effects. Prednisolone treatment of Dmdmdx rats similarly increased skeletal muscle strength and was also associated to a depletion of effectors CD45RChigh cells but resulted in severe weight loss.\n\nOverall, Dmdmdx rats display important immune inflammatory response and thus represent a useful model to analyze new anti-inflammatory and tolerogenic treatments for DMD. As an example, a new treatment with anti-CD45RC antibodies improved muscle strength in Dmdmdx rats as prednisolone did but without side effects. Anti-CD45RC therapy could complement other therapies in DMD patients.

immunology↗