309 - Distinct MAPK and NF-B Signaling Dynamics in Native and LPS-Activated Neonatal and Adult Monocytes upon Infection with Ureaplasma Species

Poster Number: 309
Publication Number: 309.124

Kirsten Glaser, University Medical Center Leipzig, Leipzig, Sachsen, Germany; Nicolas Schlegel, University of Würzburg, Würzburg, Bayern, Germany; Ulrich H. Thome, University of Leipzig, Leipzig, Sachsen, Germany; Christian Speer, University Children`s Hospital, Höchberg, Bayern, Germany

Background: Ureaplasma species have been increasingly recognized as etiological agents of prenatal and postnatal inflammation contributing to preterm labor and neonatal morbidity. Backing clinical trials, our group confirmed a profound pro-inflammatory capacity of Ureaplasma in neonatal monocytes. Data on virulence factors, host receptors and signaling pathways involved are scarce. Experimental studies in immortalized cell lines exposed to extracted or recombinant Ureaplasma membrane proteins indicate a role of toll-like receptors (TLR) and nuclear factor k B (NF-k B) signaling.

Objective: The present study addressed involvement of TLR-associated signaling in Ureaplasma-driven monocyte activation.

Design/Methods: Cord blood and adult monocytes were incubated ex vivo with viable U. parvum and U. urealyticum in the presence or absence of E. coli LPS. Using flow cytometry and western blotting we assessed phosphorylation of the key signaling proteins p65/NF-k B, and the mitogen-activated protein kinases p38, ERK1/2 (extracellular signal-regulated kinase) and JNK (c-Jun N-terminal kinase), and interferon regulatory transcription factor (IRF) 3. Bead-based immunoassays allowed for quantitative analyses including phosphorylated p70 and signal transducer and activator of transcription (STAT) 3.

Results: Both Ureaplasma isolates induced phosphorylation of p38 and ERK1/2. While borderline activation was detected at 30min, 60min and 90min, we found significant changes in phosphorylated p38 and ERK1/2 at 12h and 18h. Stimulation with viable Ureaplasma did not affect phosphorylation of p65, neither in neonatal nor in adult cells. On the contrary, LPS stimulation resulted in significant phosphorylation of p65 and p38 as well as transcription factor STAT3 and – to a lesser extent – ERK1/2, JNK and p70 in neonatal and adult samples at 15min and 30min. Of note, phosphorylation of p38 and p65, and less pronounced ERK1/2, was significantly enhanced in monocytes co-stimulated with Ureaplasma and LPS compared to LPS alone. Ureaplasma-induced pH shift, a potential virulence factor, did not affect any of the signaling proteins.Conclusion(s): Our data demonstrate Ureaplasma-driven activation of p38 and ERK1/2 but not p65/NF-k B in neonatal and adult monocytes. Dynamics and duration of phosphorylation significantly differed from LPS-mediated effects – a finding that might promote more targeted therapy. Notably, Ureaplasma enhanced phosphorylation of p38 and p65 in LPS-stimulated monocytes. This effect might account for immunomodulatory capacities of Ureaplasma with underlying mechanisms to be elucidated.