B cells and regulatory T cells in Graft versus Host Disease: a clinicopathological study in humanized mice

Abstract

Morbidity and mortality are major concerns in graft-versus-host disease (GvHD), a response often seen after hematologic cell transplantation. Treatment of GvHD remains difficult and more knowledge is necessary. In this thesis we tried to contribute a small piece to the puzzle of GvHD by analyzing the possible relation between B cells and regulatory T cells (Tregs), from a primarily histological perspective. Humanized mice may serve as promising models in studies to GvHD. Designing the right model for the topic of interest is however subject to many choices. Based on our thorough review of the pearls and pitfalls from different mouse strains, transfer routes and sources of material, we decided that RAG2-/-γc-/- mice with intravenous transfer of huPBMC’s would be a sensible choice for our experiments. A thorough analysis was performed to evaluate the basic kinetics and pathology of our model. The induced GvH response in our model appeared to show many similarities with the sclerotic variant of human GvHD. Our analysis continued with an evaluation of our model with additional modifications: macrophage depletion and early B cell depletion. Macrophage depletion accelerated the induction and morbidity of the clinical GvH response and histologically increased the number of B-cells in the infiltrating cell signature, with only limited changes to the amount of Tregs. In early B cell depletion however, a clear improvement on survival was seen. Despite clinical improvement though, our histology only showed a significant effect on liver fibrosis and human TGF-β production within the liver. Other affected organs did not show signs of improvement or alterations. Our next exploration analyzed the effect of B cell activation on Tregs, using a combination of multiple in vitro studies with an in vivo study. Although based on our in vitro studies additional Tregs should temper the GvH reaction (even after B-cell activation), our in vivo study with myeloma was suprising. Clinical symptoms and survival of our mice did improve with additional Tregs, but no significant improvement was shown on a histological level. This contradiction between clinical symptoms and histology is known from human GvHD, as we continued to show with our last evaluation in this thesis, regarding human cutaneous GvHD. In this last analysis however, we’ve found that nowadays neither clinical grading nor histological grading seems to accurately predict a patients survival. With regard to the aim of these thesis, three major conclusions/considerations come forward. First, B cells and Tregs in GvHD have a complex relation, which is subject to the activation of B cells. Second, the lack of correlation between histology and clinical symptoms in GvHD might be explained by cytokines. Last but not least, we need to consider that Rituximab might serve best as a treatment for GvHD when used as an early B cell depletion therapy. However, since there might be a huge contrast between clinical symptoms and histology, we need to decide on our goal of therapy first: do we treat the patient or the disease? The answer to this, will ultimately decide whether our therapy really works.