Preferential Th2 Polarization at the Onset of Thrombotic Microangiopathy (TMA): Immunological Comparison of TMA Versus Acute GVHD in Terms of Peripheral T Cell Subsets, Dendritic Cell Subsets and Cytokine Profiles.
Authors: Koh Ki-Ryang , Nakamae Hirohisa , Ohta Kensuke , Koh Hideo , Nakane Takahiko , Nakamae Mika , Kanashima Hiroshi , Hagihara Kiyoyuki , Sakamoto Erina , Terada Yoshiki , Takeoka Yasunobu , Hirose Asao , Yamane Takahisa , Hino Masayuki 
Published: October 2019

Abstract
Thrombotic microangiopathy (TMA) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a serious complication with a high mortality. Acute GVHD (aGVHD) is one of risk factors for TMA and often overlaps it. In particular, gastrointestinal endothelium is a common target of aGVHD and TMA, which makes clinical diagnosis of TMA difficult, leading to delay early and appropriate treatment for it. In this study, to gain more insight into differences between TMA and aGVHD, comprehensive immunological analysis was performed.
Methods: We determined kinetics of peripheral T cell subsets (CD4, CD8, Th1, Th2, γδ-T, NKT) and dendritic cell (DC) subsets (CD11c+DC and CD123+DC), serum 17 different cytokines (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-17, TNF-α, IFN-γ, G-CSF, GM-CSF, MIP-1β, MCP-1), and C-reactive protein (CRP) at the onsets of aGVHD or TMA in 25 patients undergoing allo-HSCT. T cell subsets including CD4 (CD3+CD4+CD8−), CD8 (CD3+CD4−CD8+), Th1 (CD4+CXCR3+CCR4−), Th2 (CD4+CXCR3−CCR4+), γδ-T (CD3+TCR-Vδ2+), and NKT (CD3+CD161+) and DC subsets were determined with a flow cytometer. TMA was diagnosed, following Iacopino’s criteria (Iacopino et al, Bone Marrow Tranplant. 24: 47, 1999). Data of aGVHD or TMA were compared with those of control without either aGVHD nor TMA between on 30 days and 60 days after allo-HSCT.
Results: There was a significant decrease in the percentage of Th1 cells in CD4+T cells in TMA (9.1%, n=10), compared to in aGVHD (24.9%, n=9, p=0.003) or in control (21.6%, n=12, p=0.009). In contrast, the percentage of Th2 cells in CD4+T cells was higher in TMA (20.2%) than in aGVHD (9.3%, p<0.001) or in control (12.1%, p=0.003). Accordingly, a significant increase in Th2/Th1 ratio was observed in TMA (4.0), compared to in aGVHD (0.4, p<0.001) or in control (0.7, p<0.001). In addition, a significant increase in the percentage of CD4 cells in CD3+T cells in TMA (59.2%, n=9) was found, compared to in aGVHD (31.2%, n=8, p=0.005) or in control (34.8%, n=11, p=0.007). The percentage of CD8+ cells in CD3+T cells was lower in TMA (26.3%) than in aGVHD (45.8%, p=0.03) or in control (54.4%, p=0.002). On the other hand, there was a significant increase in the ratio of CD11c+DC/CD123+DC in aGVHD (3.6, n=8), compared to in TMA (1.4, n=5, p=0.02) or in control (n=1.8, p=0.02). γδ-T and NKT did not show any significant changes among aGVHD, TMA and control. Moreover, no significant changes were observed in either 17 different cytokines among aGVHD, TMA and control. Of note, in TMA but not in aGVHD nor in control, positive correlations of Th2/Th1 ratio were found with IL-6 (p=0.01, r=0.91, n=6), IL-10 (p=0.03, r=0.86, n=6), and CRP (p<0.001, r=0.93, n=9).
Conclusion: Preferential Th2 and CD4 polarizations were observed at the onset of TMA. Thus, simultaneous monitoring of Th1, Th2, CD4 and CD8 was suggested to become a useful immunological parameter for differentiating TMA from aGVHD after allo-HSCT.