Regulatory T cells in multiple sclerosis: good guys go bad at the gateway to the brain

Abstract

Multiple sclerosis (MS) is the most common neurological disorder in young adults. Worldwide, 2.8 million patients suffer from the disease. Most patients are women and are about 30 years old when symptoms start to occur. MS is a chronic, inflammatory disease of the brain and spinal cord. The central nervous system is attacked by infiltrating immune cells, more specifically by T cells. This causes a disturbed signal transduction between nerves and organs. As a consequence, symptoms like visual issues, loss of muscle strength, fatigue and emotional and cognitive problems appear. In healthy individuals, these autoreactive T cells are kept in check by regulatory T cells (Tregs). In addition, the blood-brain barrier limits infiltration into the central nervous system. Previous research shows that Tregs of MS patients are no longer capable to suppress autoreactive T cells. Moreover, the blood-brain barrier no longer forms a hurdle for infiltrating T cells. Current therapies for treatment of MS are general and immunosuppressive drugs that can cause many side effects. Indeed, patients are more sensitive to infections like the flu or COVID-19. Unfortunately, these therapies do not cure MS. Therefore, patients require lifelong treatment while the disease still progresses. Eventually, inflammation diminishes, but already inflicted damage, and thus symptoms, persist. Currently, only one treatment is available for this progressive phase of MS. Accordingly, there is a high need for new therapies for MS patients. I believe that Tregs form the holy grail for MS therapy to treat both early and progressive MS patients. These cells are currently being investigated as a cell-based therapy in clinical trials for other autoimmune diseases and organ transplantation. Results of Treg-based cell therapy are summarized in Chapter 2 of this thesis. As mentioned before, healthy Tregs are anti-inflammatory. In addition, they have been shown to be regenerative in the central nervous system. However, it has become clear from previous research that Tregs isolated from MS patients are no longer suppressive. Even more, animal models of MS show that Tregs in the central nervous system even contribute to the inflammatory response. The goal of my research is to investigate how Tregs lose their suppressive function and whether they eventually add up to the disease. The focus of this study lies on the gateway to the brain, the blood-brain barrier. It consists of endothelial cells, pericytes and astrocytes collaborating to limit immune cell infiltration into the central nervous system while controlled exchange of nutrients and waste products is mediated. However, in MS, these immune cells can freely enter the brain. In addition, the blood-brain barrier add to the inflammatory environment and can even promote activation of passing immune cells. In this thesis, I study whether the blood-brain barrier has a similar pathogenic influence on transmigrating Tregs. Finally, I investigate how this effect can be reversed or even prevented. In Chapter 3, I use both an animal model of MS as well as a human cellular model of the blood-brain barrier. Using genetically edited mice, I track Tregs in different tissues of the mouse during a MS-like disease. Results show that Tregs in the central nervous system indeed lose their typical characteristics compared to peripheral Tregs. These exTregs accumulate in the central nervous system during the course of the disease. Next, I study this mechanism using human cells in a model of the bloodbrain barrier, which had not been done before. This model only consists of endothelial cells as they are the first and most inflamed layer that immune cells have to cross. These endothelial cells are cultured into an inflamed cell layer to mimic a MS-like situation. Next, Tregs are isolated from healthy donors and early MS patients and added to this endothelial cell layer. Due to the inflamed state of the endothelial cells, Tregs migrate across this cell layer like they would over the blood-brain barrier in MS patients. My results show that Tregs are indeed negatively influenced by mere contact with the endothelial cells, an effect which is incrementally worsened by transmigration through the cell layer. Next to the lost suppressive function of migrated Tregs, these cells also appear to be highly inflammatory. For the first time, I report that human Tregs passing the blood-brain barrier not only fail to suppressive the immune response but also acquire an inflammatory profile. Even more, Tregs from MS patients are more sensitive to this pathogenic switch. I found that one specific mechanism correlates to transmigrated Tregs. It is known that blocking this mechanism using the drug rapamycin boosts Treg suppressive capacity and keeps them in an anti-inflammatory state. Rapamycin is already being used in clinical trials to treat patients with MS and other autoimmune diseases. Indeed, I found that treatment of endothelial cell-transmigrated Tregs restores and even augments their lost suppressive capacity. Finally, these results were validated in early MS patients. During MS diagnosis, patients undergo a lumbar puncture to collect cerebrospinal fluid. Cells in these samples represent the blood-brain barrier-transmigrated immune cells. Comparing Tregs from the cerebrospinal fluid to Tregs from the blood reveals that pathogenic Tregs indeed accumulate in the central nervous system of early MS patients. This research shows that Tregs that infiltrate the brain are not capable of controlling the ongoing immune response and even contribute to the autoreactive immune response. The identified pathogenic mechanisms can be translated into new treatments for both early and progressive MS patients. I suggest to use Tregs as a cell-based therapy by genetically editing the Tregs to get them safely and functionally across the blood-brain barrier into the central nervous system. In Chapter 4, I propose different strategies with the eye on a possible cure for MS patients.