Acute graft-versus-host disease (aGVHD) is an immune cell-driven, potentially lethal complication of allogeneic hematopoietic stem cell transplantation affecting diverse organs, including the skin,... Show moreAcute graft-versus-host disease (aGVHD) is an immune cell-driven, potentially lethal complication of allogeneic hematopoietic stem cell transplantation affecting diverse organs, including the skin, liver, and gastrointestinal (GI) tract. We applied mass cytometry (CyTOF) to dissect circulating myeloid and lymphoid cells in children with severe (grade III-IV) aGVHD treated with immune suppressive drugs alone (first-line therapy) or in combination with mesenchymal stromal cells (MSCs; second-line therapy). These results were compared with CyTOF data generated in children who underwent transplantation with no aGVHD or age-matched healthy control participants. Onset of aGVHD was associated with the appearance of CD11b(+)CD163(+) myeloid cells in the blood and accumulation in the skin and GI tract. Distinct T-cell populations, including TCR gamma delta(+) cells, expressing activation markers and chemokine receptors guiding homing to the skin and GI tract were found in the same blood samples. CXCR3(+) T cells released inflammation-promoting factors after overnight stimulation. These results indicate that lymphoid and myeloid compartments are triggered at aGVHD onset. Immunoglobulin M (IgM) presumably class switched, plasma-blasts, and 2 distinct CD11b(-) dendritic cell subsets were other prominent immune populations found early during the course of aGVHD in patients refractory to both first- and second-line (MSC-based) therapy. In these nonresponding patients, effector and regulatory T cells with skin- or gut-homing receptors also remained proportionally high over time, whereas their frequencies declined in therapy responders. Our results underscore the additive value of high-dimensional immune cell profiling for clinical response evaluation, which may assist timely decision-making in the management of severe aGVHD. Show less
Barnhoorn, M.C.; Meulen-de Jong, A.E. van der; Schrama, E.C.L.M.; Plug, L.G.; Verspaget, H.W.; Fibbe, W.E.; ... ; Schepers, K. 2022
Locally applied mesenchymal stromal cells (MSCs) have the capacity to promote the healing of perianal fistulas in Crohn's disease (CD) and are under clinical development for the treatment of... Show moreLocally applied mesenchymal stromal cells (MSCs) have the capacity to promote the healing of perianal fistulas in Crohn's disease (CD) and are under clinical development for the treatment of proctitis in ulcerative colitis (UC). Despite these clinical advances, the mechanism of action of local MSC therapy in inflammatory bowel disease (IBD) is largely unknown. We hypothesized that the local cytokine environment in IBD patients affects the immunomodulatory properties of MSCs. To evaluate this, 11 cytokines were analyzed in inflamed tissues obtained from CD and UC patients. Based on the identified cytokine profiles 4 distinct cytokine mixtures that mimic various inflammatory IBD environments were established. Next, MSCs were cultured in the presence of either of these 4 cytokine mixtures after which the expression of immunomodulatory and tissue regenerative molecules and the capacity of MSCs to modulate T-cell proliferation and dendritic cell (DC) differentiation were assessed. Our data show that MSCs respond, in a cytokine-specific manner, by upregulation of immunomodulatory and tissue regenerative molecules, including cyclooxygenase-2, indoleamine 2,3-dioxygenase, and transforming growth factor-beta 1. Functional studies indicate that MSCs exposed to a cytokine profile mimicking one of the 2 UC cytokine milieus were less effective in inhibition of DC differentiation. In conclusion, our data indicate that cytokine mixes mimicking the local cytokine milieus of inflamed UC colonic or CD fistulas tissues can differentially affect the immunomodulatory and tissue regenerative characteristics of MSCs. These data support the hypothesis that the local intestinal cytokine milieu serves as a critical factor in the efficacy of local MSC treatment. Show less
Ruiz, A.R.; Dicks, A.; Tuerlings, M.; Schepers, K.; Pel, M. van; Nelissen, R.G.H.H.; ... ; Ramos, Y.F.M. 2021
Cartilage has little intrinsic capacity for repair, so transplantation of exogenous cartilage cells is considered a realistic option for cartilage regeneration. We explored whether human-induced... Show moreCartilage has little intrinsic capacity for repair, so transplantation of exogenous cartilage cells is considered a realistic option for cartilage regeneration. We explored whether human-induced pluripotent stem cells (hiPSCs) could represent such unlimited cell sources for neo-cartilage comparable to human primary articular chondrocytes (hPACs) or human bone marrow-derived mesenchymal stromal cells (hBMSCs). For this, chondroprogenitor cells (hiCPCs) and hiPSC-derived mesenchymal stromal cells (hiMSCs) were generated from two independent hiPSC lines and characterized by morphology, flow cytometry, and differentiation potential. Chondrogenesis was compared to hBMSCs and hPACs by histology, immunohistochemistry, and RT-qPCR, while similarities were estimated based on Pearson correlations using a panel of 20 relevant genes. Our data show successful differentiations of hiPSC into hiMSCs and hiCPCs. Characteristic hBMSC markers were shared between hBMSCs and hiMSCs, with the exception of CD146 and CD45. However, neo-cartilage generated from hiMSCs showed low resemblances when compared to hBMSCs (53%) and hPACs (39%) characterized by lower collagen type 2 and higher collagen type 1 expression. Contrarily, hiCPC neo-cartilage generated neo-cartilage more similar to hPACs (65%), with stronger expression of matrix deposition markers. Our study shows that taking a stepwise approach to generate neo-cartilage from hiPSCs via chondroprogenitor cells results in strong similarities to neo-cartilage of hPACs within 3 weeks following chondrogenesis, making them a potential candidate for regenerative therapies. Contrarily, neo-cartilage deposited by hiMSCs seems more prone to hypertrophic characteristics compared to hPACs. We therefore compared chondrocytes derived from hiMSCs and hiCPCs with hPACs and hBMSCs to outline similarities and differences between their neo-cartilage and establish their potential suitability for regenerative medicine and disease modelling. Show less
T cells recognize pathogen-derived antigens and are crucial for fighting pathogens such as viruses and bacteria. In addition, T cells are able to recognize and attack certain types of tumors, in... Show moreT cells recognize pathogen-derived antigens and are crucial for fighting pathogens such as viruses and bacteria. In addition, T cells are able to recognize and attack certain types of tumors, in particular virally induced tumors. In this thesis we aimed 1) to obtain more insight into antigen-specific T cell responses and 2) to study how antigen-specific T cell responses can be improved. For the first aim we generated new tools that by enabling the visualization of antigen-specific CD4+ and CD8+ T cells allow the study of the dynamics of antigen-specific T cell responses in time throughout an ongoing immune response (chapter 2). In addition, we developed a novel technique that enables the study of family relationships between different T cell populations. This technique for instance allows us to determine whether two different types of effector T cell populations arise from the same or different pool(s) of na_ve T cells (chapter 5). For the second aim, we analyzed whether antigen-specific T cell responses can be manipulated by providing increased costimulation in the form of constitutive triggering of CD27 (chapter 3) or by generating CD4+ T cells that are modified by the introduction of MHC class I restricted TCRs (chapter 4). Show less
