Phagosome-regulated mTOR Signalling during Sarcoidosis Granuloma Biogenesis.
Sarcoidosis and tuberculosis are granulomatous pulmonary diseases characterised by heightened immune reactivity to Mycobacterium tuberculosis (M.tb.) antigens. We hypothesised that an unsupervised analysis comparing the molecular characteristics of granulomas formed in response to M.tb. antigens in patients with sarcoidosis or latent tuberculosis infection (LTBI) would provide novel insights into the pathogenesis of sarcoidosis.A genomic analysis identified differentially expressed (DE) genes in granuloma-like cell aggregates formed by sarcoidosis (n=12) or LTBI patients (n=5) in an established in vitro human granuloma model wherein peripheral blood mononuclear cells (PBMCs) were exposed to M.tb. antigens (beads coated with purified protein derivative, PPD) and cultured for 7 days. Pathway analysis of DE genes identified canonical pathways, most notably antigen processing and presentation via phagolysosomes, as a prominent pathway in sarcoidosis granuloma formation. The phagolysosomal pathway promoted mTORc1/STAT3 signal transduction. Thus, granuloma formation and related immune mediators were evaluated in the absence or presence of various pre-treatments known to prevent phagolysosome formation (chloroquine) or phagosome acidification (bafilomycin A1) or directly inhibit mTORc1 activation (rapamycin).In keeping with genomic analyses indicating enhanced phagolysosomal activation and predicted mTORc1 signalling, it was determined that sarcoidosis granuloma formation and related inflammatory mediator release was dependent upon phagolysosome assembly and acidification and mTORc1/S6/STAT3 signal transduction.Sarcoidosis granulomas exhibit enhanced and sustained intracellular antigen processing and presentation capacities, and related phagolysosome assembly and acidification are required to support mTORc1 signalling to promote sarcoidosis granuloma formation.