T-cells in human trigeminal ganglia express canonical tissue-resident memory T-cell markers

BACKGROUND: Trigeminal ganglia (TG) neurons are the main site of lifelong latent herpes simplex virus type 1 (HSV-1) infection. T-cells in ganglia contribute to long-term control of latent HSV-1 infection, but it is unclear whether these cells are bona fide tissue-resident memory T-cells (T(RM)). We optimized the processing of human post-mortem nervous tissue to accurately phenotype T-cells in human TG ex vivo and in situ. METHODS: Peripheral blood mononuclear cells (PBMC; 5 blood donors) were incubated with several commercial tissue digestion enzyme preparations to determine off-target effect on simultaneous detection of 15 specific T-cell subset markers by flow cytometry. Next, optimized enzymatic digestion was applied to ex vivo phenotype T-cells in paired PBMC, normal appearing white matter (NAWM) and TG of 8 deceased brain donors obtained < 9 h post-mortem by flow cytometry. Finally, the phenotypic and functional markers, and spatial orientation of T-cells in relation to neuronal somata, were determined in TG tissue sections of five HSV-1-latently infected individuals by multiparametric in situ analysis. RESULTS: Collagenase IV digestion of human nervous tissue was most optimal to obtain high numbers of viable T-cells without disrupting marker surface expression. Compared to blood, majority T-cells in paired NAWM and TG were effector memory T-cells expressing the canonical T(RM) markers CD69, CXCR6 and the immune checkpoint marker PD1, and about half co-expressed CD103. A trend of relatively higher T(RM) frequencies were detected in TG of latently HSV-1-infected compared to HSV-1 naïve individuals. Subsequent in situ analysis of latently HSV-1-infected TG showed the presence of cytotoxic T-cells (TIA-1(+)), which occasionally showed features of proliferation (KI-67(+)) and activation (CD137(+)), but without signs of degranulation (CD107a(+)) nor damage (TUNEL(+)) of TG cells. Whereas majority T-cells expressed PD-1, traits of T-cell senescence (p16INK4a(+)) were not detected. CONCLUSIONS: The human TG represents an immunocompetent environment in which both CD4 and CD8 T(RM) are established and retained. Based on our study insights, we advocate for T(RM)-targeted vaccine strategies to bolster local HSV-1-specific T-cell immunity, not only at the site of recurrent infection but also at the site of HSV-1 latency. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12974-022-02611-x.