Immune Phenotype by Flow Cytometry of Paediatric Kidney Transplant Recipients and Healthy Adult Controls

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Abstract

Introduction

The proportion of immune cell subsets in paediatric kidney transplant recipients could help monitor the best approach to allow tolerance to renal allografts.

Aim

To determine/establish the differences in the immunophenotype of pediatric kidney transplant recipients and healthy adult controls.

Methods

Absolute cell count (TruCount) and seven leukocyte-profiling panels containing 8-10 marker-antigens (46 of which were unique within the panels) consisting of Tregs, NKT, B, NK, DCs, and monocyte subsets were used to monitor the immune profiles of 9 paediatric kidney transplant and 8 adult control samples. Whole-blood samples, 100-300μl/panel, were stained and acquired on a BD-LSRFortessa and Flowjo was used for data analysis.

Results

Differences in subpopulations between pediatric and healthy adult control can be seen in Table 1. This showed a significant increase in absolute numbers of Granulocytes, CD14+Monocytes, double negative NKT cells and CD56+HICD16+ Intermediate NK cells. B cell panel showed a significant increase in naïve B cells, IgD+IgM+ B cells, and IgM+CD27- B cells. The naïve CD4+ (CD45RA+CCR7+/CD62+) and naïve CD8+ (CD45RA+CCR7+/CD62+) T cells, and naïve Tregs (CD4+CD25+hiCD127lowCD45RA+CCR7+/CD62+ and CD4+FOXP3+CD45RO-) which were higher than in adult controls. Moreover, naïve Foxp3 Tregs in pediatric transplanted patients had higher CD25 expression than naive Tregs in adult controls. Memory CD4+ T cells (CD45RA-CD4+) in pediatric patients has a similar HLA-DR expression as adult controls (Fig. 1).Further we found a significant decrease in the following cell populations in our pediatric kidney transplant patients compare to healthy adult controls: Non-classical monocytes, CD8+NKT, memory B cells (IgD-CD27+), IgD-IgM- B cells, CD27+CD38low Class-switched memory. A decreased proportion in CD27-CD28+, CD25+ of CD4+, CD57+CD27-CD28+ and CD25+ of CD8, CCR7-/CD62L-CD45RA- of CD4 and CD8, and CXCR3+CD45RO+ of CD4 and of CD8 (Fig.2), and CD127+CD45RO+ and CD25+CD45RO+ on CD4 T cells (effector Foxp3 Tregs) suggested less memory and effector memory T cells in our pediatric kidney transplant patients than controls (Fig.3).

Conclusion

Immune profiling of pediatric kidney transplant recipients demonstrated more naïve T cells, B cells and Tregs and less memory and effector memory T cells compared to healthy adult controls. Immunophenotyping by flow cytometry is a non-invasive technique requiring less than 1.5ml of whole blood and maybe a useful tool to monitor determine the differences in the immunophenotype in transplant recipients.

Conclusion

Birgit Sawitzki. Mathias Streitz.

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