An analysis of regulatory T-cell and Th-17 cell dynamics during cytomegalovirus replication in solid organ transplant recipients

Abstract

Background: CMV-specific T-cells are crucial to control CMV-replication post-transplant. Regulatory T-cells (T-regs) are associated with a tolerant immune state and may contribute to CMV-replication. However, T-cell subsets such as T-regs and IL-17 producing T-cells (Th-17) are not well studied in this context. We explored T-regs and Th-17 frequencies during CMV-replication after transplantation.

Methods: We prospectively evaluated 30 transplant patients with CMV-viremia. We quantified CMV-specific CD4(+) and CD8(+) T-cells, T-regs (CD4(+)CD25(+)FoxP3(+)) and Th-17 frequencies using flow-cytometry and followed patients requiring anti-viral treatment. Two subsets were compared: anti-viral treatment requirement (n = 20) vs. spontaneous clearance of viremia (n = 10).

Results: Higher initial CMV-specific CD4(+) T-cells and lower T-regs were observed in patients with spontaneous clearance (p = 0.043; p = 0.021 respectively). Using a ratio of CMV-specific CD4(+) T-cells to T-regs allowed prediction of viral clearance with 80% sensitivity and 90% specificity (p = 0.001). One month after stop of treatment, the same correlation was observed in patients protected from CMV-relapse. The ratio of CMV-specific CD4(+) T-cells to T-regs allowed prediction of relapse with 85% sensitivity and 86% specificity (p = 0.004). Th-17 responses were not correlated with virologic outcomes.

Conclusions: This study provides novel insights into T-regs and Th-17 subpopulations during CMV-replication after transplantation. These preliminary data suggest that measurement of CMV-specific CD4(+) T-cells together with T-regs has value in predicting spontaneous clearance of viremia and relapse.

Figure 1. CMV-specific CD4⁺ and CD8⁺ T-cell, T-reg and Th-17 dynamics in patients with different viral outcomes.

Square (□) indicates CMV-specific CD4⁺ T-cell response. Circle (○) indicates CMV-specific CD8⁺ T-cell response. Triangle (▵) indicates T-reg response. Diamond (⋄) indicates Th-17 response. Black bar indicates median value, whiskers indicate inter-quartile range. (A), patients with spontaneous clearance of viremia compared with patients with progressive viral-loads who commenced treatment. Initial immune response at treatment commencement was determined. (B), patients with relapse compared to patients without relapse. Immune responses as determined 1 month after treatment discontinuation.

Figure 2. Receiver operating characteristic (ROC) curve analysis of CMV-specific CD4⁺ T-cells and T-regs to predict virus clearance and development of relapse.

ROC curves were generated by analyzing the sensitivity and specificity of different cut-points for defining a positive test result and their ability to predict the outcome of interest. For Figure 2A, 2B, and 2C, the outcome of interest is spontaneous clearance of viremia [vs. progression]; n = 30. For Figure 2D, 2E, and 2F, the outcome is relapse of CMV after completion of treatment [n = 20]. The following three tests were analyzed: CMV-specific CD4⁺ T-cell response [Figure 2A and 2D], T-reg response [Figure 2B and 2E] and the ratio of the CMV-specific CD4⁺ T-cells to T-reg response [Figure 2C and 2F]. To determine the sensitivity and specificity for predicting clearance of viremia we used baseline immune measurements (2A, 2B, and 2C); for prediction of relapse we used measurements one month after completing treatment (2D, 2E, and 2F).