Gammaherpesvirus infection impairs lung fibrosis resolution through PD-L1+ fibroblasts and PD-1+ CD8+ T cell interaction

Pulmonary fibrosis is a complex disease with poorly understood, multifactorial triggers. Gammaherpesviruses (γHVs), including Epstein-Barr virus in humans and Murid herpesvirus 4 (MuHV-4) in mice, have been linked to pulmonary fibrosis exacerbation, although the underlying mechanisms remain unclear. This study explores how γHV infection modulates bleomycin (bleo)-induced lung fibrosis in mice, using flow cytometry and multiplex spectral immunofluorescence (MSI) to analyze cell dynamics, localization, and intercellular crosstalk. Despite the substantial contribution of monocytes (MOs) to the alveolar macrophage (AM) pool following MuHV-4 infection, these cells do not exhibit overt profibrotic properties upon bleo treatment. In contrast, MuHV-4 infection led to the accumulation of activated, IFNγ-producing memory CD8+ T cells in the lungs of bleo-treated mice, with MSI revealing their enrichment and colocalization with fibroblasts. Notably, fibroblasts showed increased IFNγ-induced PD-L1 expression, while T cells upregulated PD-1, co-localizing in fibrosis-rich regions. In vivo neutralization of the PD-L1/PD-1 axis after bleo treatment significantly improved the resolution of fibrosis in MuHV-4-infected mice, highlighting the key pathogenic role of this interaction in sequelae persistence. Compared to fibroblasts cocultured with T cells from naïve mice, those cocultured with MuHV-4-induced T cells exhibited significantly increased collagen expression, while this effect was abrogated by PD-1 blockade. Similar results were observed in cocultures of human fibroblasts and CD8+ T cells isolated from EBV-infected patients, underscoring the translational potential of these findings. Collectively, our findings identify interactions between PD-1+ CD8+ T cells and PD-L1+ fibroblasts as central drivers of persistent lung fibrosis associated with γHV infection.