RESUMO
OBJECTIVES: Inflammatory cytokines that signal through the Janus kinases-signal transducer and activator of transcription (JAK-STAT) pathway, especially interferons (IFNs), are implicated in Sjögren's disease (SjD). Although inhibition of JAKs is effective in other autoimmune diseases, a systematic investigation of IFN-JAK-STAT signalling and the effect of JAK inhibitor (JAKi) therapy in SjD-affected human tissues has not been fully investigated. METHODS: Human minor salivary glands (MSGs) and peripheral blood mononuclear cells (PBMCs) were investigated using bulk or single-cell (sc) RNA sequencing (RNAseq), immunofluorescence (IF) microscopy and flow cytometry. Ex vivo culture assays on PBMCs and primary salivary gland epithelial cell (pSGEC) lines were performed to model changes in target tissues before and after JAKi. RESULTS: RNAseq and IF showed activated JAK-STAT pathway in SjD MSGs. Elevated IFN-stimulated gene (ISGs) expression associated with clinical variables (eg, focus scores, anti-SSA positivity). scRNAseq of MSGs exhibited cell type-specific upregulation of JAK-STAT and ISGs; PBMCs showed similar trends, including markedly upregulated ISGs in monocytes. Ex vivo studies showed elevated basal pSTAT levels in SjD MSGs and PBMCs that were corrected with JAKi. SjD-derived pSGECs exhibited higher basal ISG expressions and exaggerated responses to IFN-ß, which were normalised by JAKi without cytotoxicity. CONCLUSIONS: SjD patients' tissues exhibit increased expression of ISGs and activation of the JAK-STAT pathway in a cell type-dependent manner. JAKi normalises this aberrant signalling at the tissue level and in PBMCs, suggesting a putative viable therapy for SjD, targeting both glandular and extraglandular symptoms. Predicated on these data, a phase Ib/IIa randomised controlled trial to treat SjD with tofacitinib was initiated.
Assuntos
Inibidores de Janus Quinases , Janus Quinases , Leucócitos Mononucleares , Fatores de Transcrição STAT , Glândulas Salivares Menores , Transdução de Sinais , Síndrome de Sjogren , Humanos , Síndrome de Sjogren/tratamento farmacológico , Síndrome de Sjogren/imunologia , Inibidores de Janus Quinases/farmacologia , Inibidores de Janus Quinases/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Janus Quinases/metabolismo , Fatores de Transcrição STAT/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Glândulas Salivares Menores/imunologia , Feminino , Interferons , Piperidinas/farmacologia , Piperidinas/uso terapêutico , Pessoa de Meia-Idade , Masculino , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Adulto , Inflamação , Pirróis/farmacologia , Pirróis/uso terapêutico , Células Epiteliais/efeitos dos fármacosRESUMO
OBJECTIVE: Lysosome-associated membrane protein 3 (LAMP3) misexpression in salivary gland epithelial cells plays a causal role in the development of salivary gland dysfunction and autoimmunity associated with Sjögren's disease (SjD). This study aimed to clarify how epithelial LAMP3 misexpression is induced in SjD. METHODS: To explore upstream signaling pathways associated with LAMP3 expression, we conducted multiple RNA sequencing analyses of minor salivary glands from patients with SjD, submandibular glands from a mouse model of SjD, and salivary gland epithelial cell lines. A hypothesis generated by the RNA sequencing analyses was further tested by in vitro and in vivo assays with gene manipulation. RESULTS: Transcriptome analysis suggested LAMP3 expression was associated with enhanced type I interferon (IFN) and IFNγ signaling pathways in patients with SjD. In vitro studies showed that type I IFN but not IFNγ stimulation could induce LAMP3 expression in salivary gland epithelial cells. Moreover, we discovered that LAMP3 overexpression could induce ectopic Toll-like receptor 7 (TLR-7) expression and type I IFN production in salivary gland epithelial cells both in vitro and in vivo. TLR-7 knockout mice did not develop any SjD-related symptoms following LAMP3 induction. CONCLUSION: Epithelial LAMP3 misexpression can be induced through enhanced type I IFN response in salivary glands. In addition, LAMP3 can promote type I IFN production via ectopic TLR-7 expression in salivary gland epithelial cells. This positive feedback loop can contribute to maintaining LAMP3 misexpression and amplifying type I IFN production in salivary glands, which plays an essential role in the pathophysiology of SjD.
Assuntos
Células Epiteliais , Interferon Tipo I , Proteínas de Membrana Lisossomal , Glândulas Salivares , Síndrome de Sjogren , Receptor 7 Toll-Like , Síndrome de Sjogren/imunologia , Síndrome de Sjogren/genética , Síndrome de Sjogren/metabolismo , Animais , Camundongos , Interferon Tipo I/metabolismo , Humanos , Células Epiteliais/metabolismo , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Glândulas Salivares/metabolismo , Glândulas Salivares/imunologia , Proteínas de Membrana Lisossomal/genética , Proteínas de Membrana Lisossomal/metabolismo , Transdução de Sinais , Feminino , Interferon gama/metabolismo , Linhagem Celular , Glândulas Salivares Menores/imunologia , Glândulas Salivares Menores/metabolismo , Proteínas de Neoplasias , Proteína 3 de Membrana Associada ao LisossomoRESUMO
Sjögren's Disease (SjD) is a systemic autoimmune disease without a clear etiology or effective therapy. Utilizing unbiased single-cell and spatial transcriptomics to analyze human minor salivary glands in health and disease we developed a comprehensive understanding of the cellular landscape of healthy salivary glands and how that landscape changes in SjD patients. We identified novel seromucous acinar cell types and identified a population of PRR4+CST3+WFDC2- seromucous acinar cells that are particularly targeted in SjD. Notably, GZMK+CD8 T cells, enriched in SjD, exhibited a cytotoxic phenotype and were physically associated with immune-engaged epithelial cells in disease. These findings shed light on the immune response's impact on transitioning acinar cells with high levels of secretion and explain the loss of this specific cell population in SjD. This study explores the complex interplay of varied cell types in the salivary glands and their role in the pathology of Sjögren's Disease.