Immune modulation

The research consists of three interconnected research lines:

Ex vivo analysis of antigen-specific B cells
 

Using specific antigen-probes and a dual fluorescent labelling approach we are able to identify and analyze the small fraction of antigen-specific B cells present in the overall pool of B cells in much detail by using multiparameter high-dimensional flow cytometry directly ex vivo during immunological intervention and disease. Hereby, we were able to demonstrate that so-called “atypical” CD11c⁺ B cells, previously related to autoimmunity and chronic exposure, are early precursors of a normal B cell response in humans. Furthermore, our results obtained in SARS-CoV-2 infected or vaccinated individuals suggest that a heterogenous activated B cell compartment precedes the classical MBC and ASC compartments and likely encompasses the decision point between ASC and MBC formation. These recent ex vivo antigen-specific B cell analyses reveal the complexity of the B cell compartment that arises upon antigen stimulation preceding classical MBC and ASC formation, in that it contains several CD11c⁺ B cell populations and activated B cells with multiple phenotypes. These populations partly overlap, as we identified a subset of CD11c positive activated B cells. 

We currently use integrated surface protein expression, single cell transcriptomic and B cell receptor repertoire analysis of sorted antigen-specific B cells at different time points after antigen exposure to study the relatedness of different B cell subsets. Furthermore, based on transcriptomic analysis of the B cell precursors and their differentiation trajectory, we expect to obtain insight in regulatory and metabolic networks underlying the B cell differentiation.

We specifically aim to:

• Identify specifically the B cell precursors of long-lived B cell responses

• Insights in contribution of CD11c⁺ B cells to long-lived B cell response

• Define ways to monitor and potentially irradicate recently induced B cell differentiation precursors of undesired MBC and ASC formation

• Determine phenotypic landscape of Immunoglobulin and complement receptors on human B cell subsets

• Investigate impact of immunomodulatory therapies on B cell responses

Ex vivo analysis of antigen-specific T cells

Considering the important role of CD4 T cell in the development of B cell responses, we also focus on the analyses of antigen-specific CD4 T cells ex vivo. A challenge in studying antigen-specific CD4 T cells is their very low frequency. This research lines connects very well with the experience in studying modulation of CD4 T cell responses by dendritic cells. Currently we are using several new approaches to determine frequency, but more importantly also phenotype of the antigen-specific CD4 T cells, such as a new platform for generation of MHCII multimers, an optimized activation-induced marker (AIM) assay, as well as antigen-specific TCR-fingerprinting based Thelper profiling. 

We specifically aim to:

• Optimize analysis of antigen-specific CD4 T cells responses
• Study the role of B cells in CD4 T cell differentiation
• Decipher the role of CD4 T cell responses during tolerance-inducing (cell) therapy and vaccination 

In vitro (3D lymph node organoid) B cell differentiation

We have extensive experience in studying B cell differentiation in minimalistic cultures in vitro starting from naïve B cells. Hereby we study the requirements for T cell-dependent B cell differentiation. We unravelled the first differentiation pathways from T-dependent human naïve B cells into IgG-secreting plasma cells using scRNAseq and identified a novel precursor of ASCs. In order to recapitulate GC B cell response in vitro even better as in our existing 2D in vitro culture systems, we are currently developing a more complex 3D lymph node like B cell culture system which include T and B cells to study B cell differentiation in a culture system that resembles in vivo environment and cellular interactions more closely. 

We specifically aim to:

• Establish 3D lymph node like multicellular organoid cultures that recapitulate the GC response

• Establish factors that regulate bifurcation of human B cell differentiation into MBC and ASC

• Explore ways to interfere in development of long-lived B cell responses as strategy for future targeted therapies to prevent unwanted B cell responses (eg during alloimmunization or autoimmunity) without affecting earlier established protective humoral responses.

• Study effect of antigen density and  antigen context on B cell differentiation

• Investigate the effect of immune modulating cellular (extracorporeal photopheresis (ECP) and tolerogenic dendritic cell) therapies on B cells in vitro