INFLAMMASOME SIGNALLING IN HUMAN HYPOXIC CHRONIC KIDNEY DISEASE (CKD)

K GIULIANI1,2,3, A GRIVEI2,3, P NAG2,3, X WANG2,3, J UNGERER1,3, J FORBES1,4, H HEALY1,2,3, A KASSIANOS1,2,3 

1Faculty of Medicine, University of Queensland, Herston, Australia, 2Kidney Health Service, Royal Brisbane and Women’s Hospital, Herston, Australia, 3Conjoint Internal Medicine Laboratory, Chemical Pathology, Pathology Queensland, Herston, Australia, 4Mater Research Institute, University of Queensland, Translational Research Institute, Woolloongabba, Australia

Aim: Describe how dendritic cells (DCs), key players in the innate immune response, switch to a pro-inflammatory state in hypoxic CKD. 

Background: CKD affects over 1 in 10 Australians. Hypoxia and inflammation are key mechanisms in the progression of CKD to fibrosis. We previously reported NLRP3-inflammasomes are activated in the inflammation of a human model of hypoxic CKD. Here we identify the cellular source of this NLRP3 activation.  

Methods: Human proximal tubule epithelial cells (PTEC) were isolated and cultured from healthy portions of tumour nephrectomies, before co-culture with isolated DCs. Inflammasome activation was blocked by the specific NLRP3 (MCC950) or non-specific Caspase-1 (VX-765) inhibitor in co-cultures. Culture supernatant cytokines were analysed (cytokine bead array) for release of the downstream, inflammasome activated pro-inflammatory molecules IL-1β and/or IL-18. Cells with activated inflammasomes were identified by intracellular staining for IL-1β and evaluated (FACs). Inflammasomes were geo-mapped to kidney biopsies by in situ immunofluorescence. 

Results: Flow cytometry assessment of the co-culture populations revealed a significant increase in IL-1β expression in DCs but not within PTEC. Co-culture inhibitor studies found a significant decrease in IL-1β and IL-18 expression in culture supernatants in the presence of either MCC950 or VX-765, identifying the NLRP3 inflammasome. There was a significant increase of active inflammasome complexes within DC, adjacent to PTEC in the fibrotic kidney biopsies.  

Conclusions: This data establishes the NLRP3 inflammasome as the predominant inflammasome active in this model of hypoxic CKD.  The likely source of NLRP3 activation in CKD is not intrinsic kidney cells but the DCs of the innate immune response. Further studies of the target/s of the downstream pro-inflammatory cytokines will be analysed for therapeutic and diagnostic efficacy.  


Biography: 

Kurt is a third and final year PhD student, based in the Conjoint Internal Medicine Laboratory – Kidney Research Group. Kurt is passionate about understanding the systems biology and signalling networks between the kidney and immune system. Specifically, Kurt is interested in the role that inflammasomes, innate-immune signal detection and response platforms, play in the progression of kidney disease.

 

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