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Isolation, expansion and functional assessment of CD4+CD25+FoxP3+ regulatory T cells and Tr1 cells from uremic patients awaiting kidney transplantation
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Transplantation Surgery. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. (Gunnar Tufveson, Olle Korsgren)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Transplantation Surgery.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
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2012 (English)In: Transplant Immunology, ISSN 0966-3274, E-ISSN 1878-5492, Vol. 26, no 1, 27-33 p.Article in journal (Refereed) Published
Abstract [en]

Background: The immunosuppressive properties of regulatory T cells have emerged as an attractive tool for the development of immunotherapies in various disease contexts, e.g. to treat transplantation induced immune reactions. This paper focuses on the process of obtaining and functionally characterizing CD4+CD25+FoxP3+ regulatory T cells and Tr1 cells from uremic patients awaiting kidney transplantation.

Methods: From October 2010 to March 2011 uremic patients awaiting living donor kidney transplantation, and their corresponding kidney donors, were enrolled in the study. A total of seven pairs were included. Isolation of CD4+CD25+FoxP3+ regulatory T cells was performed by magnetic activated cell sorting of peripheral blood mononuclear cells obtained from the uremic patients. Donor specific Tr1 cells were differentiated by repetitive stimulation of immature CD4+ T cells with immature dendritic cells, with the T cells coming from the future kidney recipients and the dendritic cells from the corresponding kidney donors. Cells were then expanded and functionally characterized by the one-way mixed leukocyte reaction and assessment of IL-10 production. Phenotypic analysis was performed by flow cytometry.

Results: The fraction of CD4+CD25+FoxP3+ regulatory T cells after expansion varied from 39.1 to 50.4% and the cells retained their ability to substantially suppress the mixed leukocyte reaction in all but one patient (3.8–19.2% of the baseline stimulated leukocyte activity, p<0.05). Tr1 cells were successfully differentiated from all but one patient and produced high levels of IL-10 when stimulated with immature dendritic cells (1,275–11,038% of the baseline IL-10 secretion, pb0.05).

Conclusion: It is practically feasible to obtain and subsequently expand CD4+CD25+FoxP3+ regulatory T cells and Tr1 cells from uremic patients without loss of function as assessed by in vitro analyses. This forms a base for adoptive regulatory T cell therapy in the setting of living donor kidney transplantation.

Place, publisher, year, edition, pages
2012. Vol. 26, no 1, 27-33 p.
National Category
Immunology in the medical area
Identifiers
URN: urn:nbn:se:uu:diva-170517DOI: 10.1016/j.trim.2011.09.003ISI: 000300203800004OAI: oai:DiVA.org:uu-170517DiVA: diva2:509242
Available from: 2012-03-13 Created: 2012-03-12 Last updated: 2017-12-07Bibliographically approved
In thesis
1. Preparatory Studies to Introduce Regulatory T Cells in Clinical Transplantation
Open this publication in new window or tab >>Preparatory Studies to Introduce Regulatory T Cells in Clinical Transplantation
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Solid organ transplantation has evolved from being an experimental procedure to a life-saving treatment for patients with end-stage organ failure. The risk of losing a transplant due to acute rejection is very low with the use of modern immunosuppressive protocols and the short-term results are impressive. However, long-term outcomes are suboptimal and transplant recipients are at increased risks for severe complications such as cancers, opportunistic infections and cardiovascular events. The previous struggle to achieve short-term survival has turned into a search for new strategies to improve patient and transplant longevity.

Regulatory T cells (TRegs), a subset of T cells, occur naturally in the immune system and have the capacity to down regulate immune responses. Under normal conditions they maintain self-tolerance and prevent excessive immune activation. Functional TReg defects lead to a massive autoimmune response and are not compatible with life. Preclinical data support that TRegs can be used as a cell therapy to prevent transplant rejection, with the potential to minimize the need for traditional immunosuppression and improve the long-term outcome.

This thesis aims to enhance the translation of TReg cell therapy to clinical organ transplantation. In particular, strategies for isolation and expansion of TRegs from uremic patients awaiting kidney transplantation have been assessed. A non-invasive imaging technique to study T cell products after intravenous administration was developed, for use in future clinical trials. The performance of a novel cell purification technique was investigated to potentially improve the clinical production of TRegs.

The thesis demonstrates that TRegs can be isolated and expanded from uremic patients to display potent suppressive properties in vitro. The mode of isolation and expansion affect the functional characteristics, where cells purified with cytometry based techniques and expanded with mature dendritic cells were the most advantageous. T cells can be labeled using the radioactive tracer [111In]oxine with preserved viability and subsequently followed in vivo with SPECT/CT for more than 1 week after intravenous administration. The use of microfluidic switch technology offers a novel way of purifying TRegs at high speed, purity and viability, under conditions compatible with clinical use.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. 80 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 983
Keyword
Transplantation, Tolerance, Cell therapy, Regulatory T cell, In vivo imaging, Cell purification
National Category
Immunology in the medical area
Identifiers
urn:nbn:se:uu:diva-220873 (URN)978-91-554-8907-6 (ISBN)
Public defence
2014-05-10, Sal X, Universitetshuset, Biskopsgatan 3, Uppsala, 09:15 (English)
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Available from: 2014-04-15 Created: 2014-03-21 Last updated: 2014-04-29

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Berglund, DavidKorsgren, OlleLorant, TomasSchneider, KarinTufveson, Gunnar

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