Plasmacytoid dendritic cells (pDCs) play a key role in the activation of the autoimmune response in LN "G. De Palma - 2011 [1], " N. Fiore - 2008 " [2]. However, the effects of pDCs, the major producer of IFN-alpha, on the renal tubular epithelial cells (RPTEC) is unknown. The aim of the study was to investigate the effects of IFN-alpha in RPTEC, in vitro and ex vivo.
Microarray analysis showed 108 significantly up-regulated genes and only 7 down-regulated genes with a fold-change >2 (false discovery rate, FDR<0.05). Gene set enrichment analysis identified 123 processes differentially regulated in IFN-α treated cells. The most represented processes resulted interferon response pathway, antigen presentation and inflammatory pathways. Among over-expressed genes, contributing to these processes, we identified the HLA-I, the ubiquitin (in particular FBXO6 and DTX3L) and the immunoproteasome subunits LMP7(PSMB8 gene)(Figura 1). After validation by RT-PCR, flow cytometry experiments on RPTEC cells confirmed a significant increase of the antigen presentation pathway (HLA I : 75%±2/MnI 7.8±3 basal vs 90%±3/MnI 11±2 48h 100 U/ml IFN-alpha) and the inflammatory signaling (LMP7 49.75%±2 basal vs 72.4%±2 48h 100 U/ml IFN-alpha) (Figura 2). IH analysis on renal biopsies of patients with LN showed a significant increase of LMP7 expression, at tubular interstitial level, (LMP7 5% ± 2 NL class I vs 16%± 5 class IV, p <0.0001) proportional to MXA protein, a specific marker of IFN-alpha (MXA 0% ± 1.0 class I vs 4.5% ± 1.2 class IV, P <0.0001)(Figura 3). Confocal microscopy confirmed the colocalization of LMP7-MXA proteins expression at tubular epithelial cells (Figura 4) and the activation of inflammatory signaling via NF-kB(p65) in the MXA+ tubules (Figura 5).
Our data demonstrate a critical role of pDC-derived IFN-alpha in the activation of RPTEC. Therefore we hypothesize that inhibition of IFN-alpha pathway may be a novel therapeutic strategy to reduce renal tubular damage in patients with LN.
[1] De Palma G, Castellano G, Del Prete A et al. The possible role of ChemR23/Chemerin axis in the recruitment of dendritic cells in lupus nephritis. Kidney international 2011 Jun;79(11):1228-35
[2] Fiore N, Castellano G, Blasi A et al. Immature myeloid and plasmacytoid dendritic cells infiltrate renal tubulointerstitium in patients with lupus nephritis. Molecular immunology 2008 Jan;45(1):259-65
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