Transcriptome-wide analysis of filarial extract-primed human monocytes reveal changes in LPS-induced PTX3 expression levels.

Filarial nematodes modulate immune responses in their host to enable their survival and mediate protective effects against autoimmunity and allergies. In this study, we examined the immunomodulatory capacity of extracts from the human pathogenic filaria Brugia malayi (BmA) on human monocyte responses in a transcriptome-wide manner to identify associated pathways and diseases. As previous transcriptome studies often observed quiescent responses of innate cells to filariae, the potential of BmA to alter LPS driven responses was investigated by analyzing >47.000 transcripts of monocytes from healthy male volunteers stimulated with BmA, Escherichia coli LPS or a sequential stimulation of both. In comparison to ~2200 differentially expressed genes in LPS-only stimulated monocytes, only a limited number of differentially expressed genes were identified upon BmA priming before LPS re-stimulation with only PTX3↓ reaching statistical significance after correcting for multiple testing. Nominal significant differences were reached for metallothioneins↑, MMP9↑, CXCL5/ENA-78↑, CXCL6/GCP-2↑, TNFRSF21↓, and CCL20/MIP3α↓ and were confirmed by qPCR or ELISA. Flow cytometric analysis of activation markers revealed a reduced LPS-induced expression of HLA-DR and CD86 on BmA-primed monocytes as well as a reduced apoptosis of BmA-stimulated monocytes. While our experimental design does not allow a stringent extrapolation of our results to the development of filarial pathology, several genes that were identified in BmA-primed monocytes had previously been associated with filarial pathology, supporting the need for further research.

Caspase-8 activity has an essential role in CD95/Fas-mediated MAPK activation.

Stimulation of CD95/Fas/APO-1 results in the induction of both apoptotic and non-apoptotic signaling pathways. The processes regulating these two opposing pathways have not been thoroughly elucidated to date. In this study, using quantitative immunoblots, imaging, and mathematical modeling, we addressed the dynamics of the DED proteins of the death-inducing signaling complex (DISC), procaspase-8, and cellular FLICE inhibitory proteins (c-FLIPs) to the onset of CD95-mediated ERK1/2 and p38 mitogen-activated protein kinase (MAPK) activation. We found that CD95 DISC-induced caspase-8 activity is important for the initiation of ERK1/2 and p38 MAPK activation. The long c-FLIP isoform, c-FLIP(L), and the short c-FLIP isoform, c-FLIP(R), inhibited MAPK induction by blocking caspase-8 processing at the DISC. Furthermore, we built a mathematical model describing CD95 DISC-mediated MAPK activation and apoptosis. The model quantitatively defined the dynamics of DED proteins, procaspase-8, and c-FLIP, which lead to caspase-8 activation and induction of apoptotic and non-apoptotic signaling pathways. In conclusion, the combination of biochemical analysis with mathematical modeling provides evidence for an important role of caspase-8 in CD95-mediated activation of MAPKs, while c-FLIP exerts a regulatory function in this process.