Involvement of ERK, p38 and NFκB Signalling in the Maturation Defects of Monocyte Derived Dendritic Cells in Patients with Myelodysplastic Syndrome.
Authors: Micheva Ilina , Ziros Panos , Pherson Lorna , Giannakoulas Nikolaos , Symeonidis Argiris , Zoumbos Nicholas 
Published: October 2019

Abstract
Dendritic cells (DC) are professional antigen-presenting cells involved in the initiation of T-cell dependent immune responses. The maturation process is central for the DC and enables them to perform different functions sequentially. We have recently demonstrated that in patients with myelodysplastic syndrome (MDS) monocyte-derived DC (MoDC) exhibit some phenotypic and functional abnormalities with a maturation failure toward TNF-α but not LPS. However, the mechanisms underlying the differential response to those stimuli have not yet been clarified. In the present study three different signalling pathways, ERK, p38K and NFκB, known to be implicated in MoDC maturation, were examined on TNF-α or LPS stimulated MoDC from 7 patients with MDS and 5 healthy controls. We also analyzed DC functions such as migration and cytokine secretion controlled by the coordinated actions of those signal transduction pathways. In agreement with our previous report stimulation of MoDC with LPS but not with TNF-α induced up-regulation of CD83, CD40, CD80 expression and allostimulatory activity of MoDC in MDS patients. Activation of MDS MoDC with LPS but not TNF-α was accompanied by nuclear translocation of NFκB. Accordingly, in 6/7 patients the NFκB binding activity following stimulation with TNF-α for 48 hours was extremely low, whereas LPS stimulated NFκB activity in MDS MoDC, although at lower levels compared to control MoDC. This was accompanied by lower than control migratory capacity (0.73±0.6 vs.3.75±0.6 x 103) and lack of IL-12p70 secretion by LPS-stimulated MoDC from patients. Interestingly, in one patient with similar to control NFκB activity the migration and IL-12p70 production were comparable to the controls. TNF-α stimulation resulted in MoDC with migratory capacity but no IL-12p70 production even after 48 hours stimulation in both patients and controls. However, the migration of MDS MoDC toward CCL21 was significantly lower (0.55±0.5 vs.2.4±1.7 x 103). Both TNF-α and LPS induced phosphorylation of p38 and ERK after 48 hours stimulation at similar levels in MoDC from controls, whereas in MDS MoDC the pattern was heterogeneous with predominant activation of ERK over p38K. The kinetics of p38K and ERK phosphorylation differed in TNF-α activated MDS MoDC. Rapid phosphorylation of p38K and ERK 30 min after stimulation was followed by loss of p38 activity and persistent activation of ERK after 48 hours. Our results provide strong evidence that NFκB is responsible for the differences in the phenotype and allostimulatory capacity of MDS MoDC after TNF-α and LPS stimulation, since LPS induced NFkB activity, although at lower levels compared to control. The low NFκB activity in MDS MoDC shows that the maturation failure of MDS MoDC, including functions such as migration and IL-12p70 secretion, is NFκB dependent. In addition, predominant activation of ERK pathway is probably also involved in the negative regulation of these MDS MoDC functions.