Rapamycin rescues loss-of-function in blood-brain barrier transmigrated regulatory T cells

Insights can help in significantly improving the efficacy of autologous Treg therapy of MS.

In many autoimmune diseases, FOXP3+ regulatory T cells (Tregs) skew towards a pro-inflammatory and non-suppressive phenotype and are therefore unable to control the exaggerated autoimmune responses. This may largely impact the success of autologous Treg therapy which is currently under investigation for treatment of autoimmune diseases, including multiple sclerosis (MS). Thus, there is a need to ensure in vivo stability of Tregs before successful Treg therapy can be applied. In this study, we postulate that the inflamed blood brain barrier (BBB) is an important driver in Treg loss of function. Using a murine genetic FOXP3 fate-mapping model, we demonstrate that inflammatory exFOXP3 T cells accumulate in the central nervous system in experimental autoimmune encephalomyelitis over time. This suggests that migration into the brain parenchyma destabilizes Tregs. To elucidate the mechanism, a human in vitro model of the BBB, using the hCMEC/D3 endothelial cell (EC) line seeded in Thincerts, was employed. Tregs of healthy donors (HD) and MS patients were allowed to migrate for 24h. Transcriptome analysis indicated that migrated human Tregs are less suppressive, have a pro-inflammatory Th1/17 signature and upregulate the mTOR signaling pathway. In MS-derived Tregs specifically, Th17-related pathways were increased after migration, suggesting a pre-existing susceptibility to Th17 skewing of Tregs in MS patients. In addition, downregulation of amphiregulin (AREG) in MS-derived migrated Tregs suggests a loss of regenerative capacity after BBB transmigration. Using functional in vitro assays, we found that the suppressive capacity of migrated Tregs was affected.  In vitro treatment of migrated Tregs with the clinically-approved mTOR inhibitor rapamycin completely restored the loss of suppressive function. Flow cytometric analysis indicated an enrichment of inflammatory, less suppressive CD49d+ Tregs in the cerebrospinal fluid of MS patients, thereby underscoring the relevance of our findings for human disease. In sum, our findings provide for the first time firm evidence that the inflamed BBB affects human Treg stability, which can be restored using an mTOR inhibitor. These insights can help in significantly improving the efficacy of autologous Treg therapy of MS.

Authors

Paulien Baeten(1,2), Ibrahim Hamad(1,3), Cindy Hoeks(1,2), Michael Hiltensperger(4), Bart Van Wijmeersch(5,6), Veronica Popescu(5,6), Lilian Aly(4), Thomas Korn(4,7), Markus Kleinewietfeld(1,3), Niels Hellings(1,2,#), Bieke Broux(1,2,8)

Organisations

Universitair MS Centrum, Campus Diepenbeek, Belgium(1), Neuro-Immune Connections and Repair Lab, Department of Immunology and Infection, Biomedical Research Institute, UHasselt, Diepenbeek, Belgium(2), VIB Laboratory of Translational Immunomodulation, Center for Inflammation Research (IRC), Department of Immunology and Infection, Biomedical Research Institute, UHasselt, Diepenbeek, Belgium(3), Klinikum Rechts der Isar, Institute for Experimental Neuroimmunology, Technische Universität München, Munich, Germany(4), Universitair MS Centrum, Campus Pelt, Belgium(5), Noorderhart, Revalidatie & MS centrum, Pelt, Belgium(6), Munich Cluster for Systems Neurology (SyNergy), Munich, Germany(7), Cardiovascular Research Institute Maastricht, Department of Internal Medicine, Maastricht University, Maastricht, The Netherlands(8), Equally contributing senior authors(#)

Presenting author

Paulien Baeten, PhD Student, Biomedical Research Institute, UHasselt
paulien.baeten@uhasselt.be
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