PD-L1+ neutrophils at the invasive margin drive contact-dependent CD8+ T cell exhaustion in hepatic alveolar echinococcosis

Abstract

Introduction Hepatic alveolar echinococcosis (HAE), a lethal chronic helminth infection caused by Echinococcus multilocularis, is characterized by pronounced local immune evasion and progressive CD8+ T cell exhaustion at the parasite?host invasive margin. However, the spatial cellular network and core regulatory subsets driving this localizedimmunosuppression remainpoorly understood. Methods We performed high-resolution spatial immune profiling of the lesion invasive margin (collagenous layer tissue, CLT) and paired distal normal liver tissues (DLT) from 12 HAE patients by integrating imaging mass cytometry, single-cell RNA sequencing, and multiplex immunofluorescence. The underlying molecular mechanism was validated via in vitro co-culture assays with or without Transwell physical separation, and the therapeutic potential was further confirmed in a delayed?treatment HAE mouse model using neutrophil depletion(anti-Ly6G) or PD-L1 blockade (anti-PD-L1). Results Spatial multi-omics identified a specific subset of APC-like PD-L1 neutrophils exclusively enriched at the HAE invasive margin, which showed striking spatial co-localization and frequent membrane-to-membrane contacts with PD-1 CD8 T cells. In vitro, activated CD8 Tcells potently induced PD-L1 upregulation on neutrophils; reciprocally, PD-L1 neutrophils significantly suppressed T cell effector function (downregulating GZMB, IFN-γ, and TNF-α) and upregulated exhaustion-associated transcription factors TOX and NR4A1 via a contact-dependent PD-L1/PD-1 pathway, effects largely abolished by Transwell separation. In the HAE mouse model, both neutrophil depletion and PD-L1 blockade significantly reduced parasite burden, alleviated liver inflammation and fibrosis, and restored CD8 T cell cytotoxicity; notably, neutrophil depletion achieved superior therapeutic efficacy compared with PD-L1 monotherapy. Conclusion Collectively, our findings demonstrate that PD-L1-expressing neutrophils at the HAE invasivefront contribute importantly to localized CD8 T cell exhaustion via direct cell-cell contact, anddisrupting this crosstalk represents a promising targeted strategy to restore host protective immunity against parasitic infection.