http://hdl.handle.net/10379/362 2017-10-29T23:03:25Z 2017-10-29T23:03:25Z Islam, Md Nahidul Bradley, Benjamin A. Ceredig, Rhodri http://hdl.handle.net/10379/6289 2017-02-16T02:00:57Z 2016-04-29T00:00:00Z

Sterile post-traumatic immunosuppression Islam, Md Nahidul; Bradley, Benjamin A.; Ceredig, Rhodri After major trauma, the human immune system initiates a series of inflammatory events at the injury site that is later followed by suppression of local inflammation favoring the repair and remodeling of the damaged tissues. This local immune response involves complex interactions between resident cells such as macrophages and dendritic cells, soluble mediators such as cytokines and chemokines, and recruited cells such as neutrophils, monocytes and mesenchymal stromal cells. If of sufficient magnitude, these initial immune responses nevertheless have systemic consequences resulting in a state called post-traumatic immunosuppression (PTI). However, controversy exists regarding the exact immunological changes occurring in systemic compartments triggered by these local immune responses. PTI is one of the leading causes of post-surgical mortality and makes patients vulnerable to hospital-acquired infections, multiple organ failure and many other complications. In addition, hemorrhage, blood transfusion, immunesenescence and immunosuppressant drugs aggravate PTI. PTI has been intensively studied, but published results are frequently cloudy. The purpose of this review is to focus on the contributions made by different responsive modalities to immunosuppression following sterile trauma and to try to integrate these into an overall scheme of PTI.

2016-04-29T00:00:00Z Ribeiro, Andreia Ritter, Thomas Griffin, Matthew Ceredig, Rhodri http://hdl.handle.net/10379/6288 2017-02-16T02:00:54Z 2016-07-20T00:00:00Z

Development of a flow cytometry-based potency assay for measuring the in vitro immunomodulatory properties of mesenchymal stromal cells Ribeiro, Andreia; Ritter, Thomas; Griffin, Matthew; Ceredig, Rhodri Human bone marrow-derived mesenchymal stromal/stem cells (MSC) have well-documented modulatory effects on multiple immune cell types. Although these effects are linked to their therapeutic benefit in diverse diseases, a reliable, quantitative assay of the immunomodulatory potency of individual human MSC preparations is lacking. The aims of this study were to develop an optimised rapid turnaround, flow cytometry-based whole-blood assay to monitor MSC potency and to validate its application to MSC immunomodulation. A protocol for short-term LPS stimulation of anti-coagulated whole blood samples followed by combined surface CD45/CD14 and intracellular TNF-alpha staining was initially developed for analysis on a 4 colour desktop cytometer. Optimal monocyte activation was dependent on the presence of extracellular calcium ions thereby precluding the use of EDTA and sodium citrate as anticoagulants. Optimal assay conditions proved to be 1 ng/mL ultrapure-LPS added to 10-fold diluted, heparin anti coagulated whole blood incubated for 6h at 37 degrees C. Under these conditions, addition of human bone marrow-derived MSC (hBM-MSC) from multiple donors resulted in a reproducible, dose-dependent inhibition of LPS-stimulated monocyte TNF-a expression. We conclude that this protocol represents a practical, quantitative assay of a clinically relevant functional effect of hBM-MSCs as well as other immunomodulatory agents. (C) 2016 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

2016-07-20T00:00:00Z von Muenchow, Lilly Tsapogas, Panagiotis Albertí-Servera, Llucia Capoferri, Giuseppina Doelz, Marianne Rolink, Hannie Bosco, Nabil Ceredig, Rhodri Rolink, Antonius G. http://hdl.handle.net/10379/6285 2017-02-09T02:00:49Z 2016-12-28T00:00:00Z

Pro-B cells propagated in stromal cell-free cultures reconstitute functional B-cell compartments in immunodeficient mice von Muenchow, Lilly; Tsapogas, Panagiotis; Albertí-Servera, Llucia; Capoferri, Giuseppina; Doelz, Marianne; Rolink, Hannie; Bosco, Nabil; Ceredig, Rhodri; Rolink, Antonius G. Up to now long-term in vitro growth of pro-B cells was thought to require stromal cells. However, here we show that fetal liver (FL) and bone marrow (BM) derived pro-B cells can be propagated long-term in stromal cell-free cultures supplemented with IL-7, stem cell factor and FLT3 ligand. Within a week, most cells expressed surface CD19, CD79A, λ5, and VpreB antigens and had rearranged immunoglobulin D-J heavy chain genes. Both FL and BM pro-B cells reconstituted the B-cell compartments of immuno-incompetent Rag2-deficient mice, with FL pro-B cells generating follicular, marginal zone (MZB) and B1a B cells, and BM pro-B cells giving rise mainly to MZB cells. Reconstituted Rag2-deficient mice generated significant levels of IgM and IgG antibodies to a type II T-independent antigen; mice reconstituted with FL pro-B cells generated surprisingly high IgG1 titers. Finally, we show for the first time that mice reconstituted with mixtures of pro-B and pro-T cells propagated in stromal cell-free in vitro cultures mounted a T-cell-dependent antibody response. This novel stromal cell-free culture system facilitates our understanding of B-cell development and might be applied clinically.

2016-12-28T00:00:00Z von Muenchow, Lilly Alberti-Servera, Llucia Klein, Fabian Capoferri, Giuseppina Finke, Daniela Ceredig, Rhodri Rolink, Antonius Tsapogas, Panagiotis http://hdl.handle.net/10379/6279 2017-02-03T02:01:02Z 2016-11-04T00:00:00Z

Permissive roles of cytokines Interleukin-7 and Flt3-ligand in mouse B cell lineage commitment von Muenchow, Lilly; Alberti-Servera, Llucia; Klein, Fabian; Capoferri, Giuseppina; Finke, Daniela; Ceredig, Rhodri; Rolink, Antonius; Tsapogas, Panagiotis Hematopoietic cells are continuously generated throughout life from hematopoietic stem cells, thus making hematopoiesis a favorable system to study developmental cell lineage commitment. The main factors incorporating environmental signals to developing hematopoietic cells are cytokines, which regulate commitment of hematopoietic progenitors to the different blood lineages by acting either in an instructive or a permissive manner. Fms-like tyrosine kinase-3 (Flt3) ligand (FL) and Interleukin-7 (IL-7) are cytokines pivotal for B-cell development, as manifested by the severely compromised B-cell development in their absence. However, their precise role in regulating B-cell commitment has been the subject of debate. In the present study we assessed the rescue of B-cell commitment in mice lacking IL-7 but simultaneously overexpressing FL. Results obtained demonstrate that FL overexpression in IL-7–deficient mice rescues B-cell commitment, resulting in significant Ebf1 and Pax5 expression in Ly6D+CD135+CD127+CD19− precursors and subsequent generation of normal numbers of CD19+ B-cell progenitors, therefore indicating that IL-7 can be dispensable for commitment to the B-cell lineage. Further analysis of Ly6D+CD135+CD127+CD19− progenitors in IL-7– or FL-deficient mice overexpressing Bcl2, as well as in IL-7 transgenic mice suggests that both FL and IL-7 regulate B-cell commitment in a permissive manner: FL by inducing proliferation of Ly6D+CD135+CD127+CD19− progenitors and IL-7 by providing survival signals to these progenitors.

2016-11-04T00:00:00Z