Supplementary Material for: The balance of CD8-positive T cells and PD-L1 expression in the myocardium predicts prognosis in lymphocytic fulminant myocarditis

Introduction: The clinical significance and prognostic value of T cell involvement and programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) have not been established in lymphocytic fulminant myocarditis (FM). We investigated the prognostic impact of the number of CD4+, CD8+, FoxP3+, and PD-1+ T cells, as well as PD-L1 expression, in cardiomyocytes in lymphocytic FM. Methods: This is a single-center observational cohort study. Myocardial tissue was obtained from 16 consecutive patients at lymphocytic FM onset. The median follow-up was 140 days. Cardiac events were defined as a composite of cardiac death and left ventricular-assist device implantation. CD4, CD8, FoxP3, PD-1, and PD-L1 immunostaining was performed on myocardial specimens. Results: The median age of the patients was 52 years (seven men and nine women). There was no significant difference in the number of CD4+ cells. The number of CD8+ cells and the CD8+/CD4+ T cell ratio were higher in the cardiac event group (Event+) than in the group without cardiac events (Event−) (P = 0.048 and P = 0.022, respectively). The number of FoxP3+ T cells was higher in the Event+ group (P = 0.049). Although there was no difference in the number of PD-1+ cells, cardiomyocyte PD-L1 expression was higher in the Event+ group (P = 0.112). Event-free survival was worse in the group with a high CD8+ cell count (P = 0.012) and high PD-L1 expression (P = 0.049). When divided into three groups based on the number of CD8+ cells and PD-L1 expression (CD8highPD-L1high [n = 8], CD8lowPD-L1high [n = 1], and CD8lowPD-L1low [n = 7]), the CD8highPD-L1high group demonstrated the worst event-free survival, while the CD8lowPD-L1high group had a favorable prognosis without cardiac events (P = 0.041). Conclusion: High myocardial expression of CD8+ T cells and PD-L1 may predict a poor prognosis in lymphocytic FM.

引言：淋巴细胞性暴发性心肌炎（lymphocytic fulminant myocarditis, FM）中，T细胞浸润以及程序性死亡受体1（programmed cell death-1, PD-1）/程序性死亡配体1（programmed cell death ligand-1, PD-L1）的临床意义与预后价值尚未明确。本研究旨在探讨淋巴细胞性FM患者心肌组织中CD4+、CD8+、FoxP3+及PD-1+ T细胞数量，以及心肌细胞PD-L1表达的预后影响。方法：本研究为单中心观察性队列研究。纳入16例淋巴细胞性FM发病时的连续患者，获取其心肌组织样本；中位随访时间为140天。心脏不良事件定义为心源性死亡与左心室辅助装置植入的复合终点。对心肌标本进行CD4、CD8、FoxP3、PD-1及PD-L1免疫组化染色。结果：患者中位年龄为52岁，其中男性7例、女性9例。CD4+ T细胞数量在两组间无显著差异。心脏不良事件组（Event+组）的CD8+ T细胞数量及CD8+/CD4+ T细胞比值均高于无不良事件组（Event−组），差异具有统计学意义（分别为P=0.048、P=0.022）。Event+组的FoxP3+ T细胞数量更高（P=0.049）。尽管PD-1+ T细胞数量在两组间无差异，但Event+组的心肌细胞PD-L1表达水平更高（P=0.112）。CD8+ T细胞计数高组与PD-L1高表达组的无事件生存率更差（分别为P=0.012、P=0.049）。根据CD8+ T细胞数量与PD-L1表达水平将患者分为三组：CD8highPD-L1high组（n=8）、CD8lowPD-L1high组（n=1）及CD8lowPD-L1low组（n=7）；其中CD8highPD-L1high组的无事件生存率最差，而CD8lowPD-L1high组未发生心脏不良事件，预后良好（P=0.041）。结论：心肌组织中CD8+ T细胞高浸润与PD-L1高表达可预测淋巴细胞性FM患者的不良预后。