Division of Liver and Transplant Pathology , University of Pittsburgh , Pittsburgh , Pennsylvania , USA.
Pathology Unit, Department of Medical Sciences , University of Turin , Torino , Italy.
Hepatology. 2023 Feb 1;77(2):355-366. doi: 10.1002/hep.32666. Epub 2022 Aug 1.
In otherwise near-normal appearing biopsies by routine light microscopy, next-generation pathology (NGP) detected close pairings (immune pairs; iPAIRs) between lymphocytes and antigen-presenting cells (APCs) that predicted immunosuppression weaning failure in pediatric liver transplant (LTx) recipients (Immunosuppression Withdrawal for Stable Pediatric Liver Transplant Recipients [iWITH], NCT01638559). We hypothesized that NGP-detected iPAIRs enrich for true immune synapses, as determined by nuclear shape metrics, intercellular distances, and supramolecular activation complex (SMAC) formation.
Intralobular iPAIRs (CD45 high lymphocyte-major histocompatibility complex II + APC pairs; n = 1167, training set) were identified at low resolution from multiplex immunohistochemistry-stained liver biopsy slides from several multicenter LTx immunosuppression titration clinical trials (iWITH; NCT02474199 (Donor Alloantigen Reactive Tregs (darTregs) for Calcineurin Inhibitor (CNI) Reduction (ARTEMIS); Prospective Longitudinal Study of iWITH Screen Failures Secondary to Histopathology). After excluding complex multicellular aggregates, high-resolution imaging was used to examine immune synapse formation ( n = 998). By enriching for close intranuclear lymphocyte-APC distance (mean: 0.713 μm) and lymphocyte nuclear flattening (mean ferret diameter: 2.1), SMAC formation was detected in 29% of iPAIR-engaged versus 9.5% of unpaired lymphocytes. Integration of these morphometrics enhanced NGP detection of immune synapses (ai-iSYN). Using iWITH preweaning biopsies from eligible patients ( n = 53; 18 tolerant, 35 nontolerant; testing set), ai-iSYN accurately predicted (87.3% accuracy vs. 81.4% for iPAIRs; 100% sensitivity, 75% specificity) immunosuppression weaning failure. This confirmed the presence and importance of intralobular immune synapse formation in liver allografts. Stratification of biopsy mRNA expression data by immune synapse quantity yielded the top 20 genes involved in T cell activation and immune synapse formation and stability.
NGP-detected immune synapses (subpathological rejection) in LTx patients prior to immunosuppression reduction suggests that NGP-detected (allo)immune activity usefulness for titration of immunosuppressive therapy in various settings.
在常规光镜下外观正常的活检中，下一代病理学（NGP）检测到淋巴细胞和抗原呈递细胞（APC）之间的近距离配对（免疫对；iPAIRs），这些配对预测了儿科肝移植（LTx）受者的免疫抑制脱药失败（免疫抑制脱药用于稳定的儿科肝移植受者[Immunosuppression Withdrawal for Stable Pediatric Liver Transplant Recipients，iWITH]，NCT01638559）。我们假设 NGP 检测到的 iPAIRs 通过核形状度量、细胞间距离和超分子激活复合物（SMAC）形成来富集真正的免疫突触，如通过核形状度量、细胞间距离和超分子激活复合物（SMAC）形成来确定。
从几个多中心 LTx 免疫抑制滴定临床试验（iWITH；NCT02474199（供体同种抗原反应性 Tregs（darTregs）用于钙调神经磷酸酶抑制剂（CNI）减少（ARTEMIS）；前瞻性 iWITH 筛查失败的纵向研究继发于组织病理学）的多重免疫组化染色肝活检幻灯片中以低分辨率识别小叶内 iPAIR（CD45 高淋巴细胞-主要组织相容性复合体 II+APC 对；n=1167，训练集）。排除复杂的多细胞聚集物后，使用高分辨率成像检查免疫突触形成（n=998）。通过富集紧密的核内淋巴细胞-APC 距离（平均值：0.713μm）和淋巴细胞核扁平化（平均值雪貂直径：2.1），在 29%的 iPAIR 结合的淋巴细胞中检测到 SMAC 形成，而在 9.5%的未配对淋巴细胞中检测到 SMAC 形成。这些形态计量学的整合增强了 NGP 对免疫突触的检测（ai-iSYN）。使用 iWITH 预断奶活检（来自符合条件的患者，n=53；18 例耐受，35 例不耐受；测试集），ai-iSYN 准确预测（与 iPAIRs 相比，准确性为 87.3%；敏感性为 100%，特异性为 75%）免疫抑制脱药失败。这证实了免疫突触在肝同种异体移植物中的形成和重要性。根据免疫突触数量对活检 mRNA 表达数据进行分层，得到了涉及 T 细胞激活和免疫突触形成和稳定性的前 20 个基因。
在减少免疫抑制之前，NGP 检测到的 LTx 患者中的免疫突触（亚病理性排斥）提示 NGP 检测到的（同种）免疫活性可用于各种情况下免疫抑制治疗的滴定。
