Ceramide-dependent PP2A regulation of TNFalpha-induced IL-8 production in respiratory epithelial cells.
IL-8 is a key mediator in the pathophysiology of acute lung injury. TNFalpha stimulates IL-8 production in respiratory epithelial cells by activating both the NF-kappaB and MAP kinase pathways. The precise mechanism by which these pathways are downregulated to terminate IL-8 production remains unclear. We studied the regulatory role of the serine/threonine phosphatase, PP2A, on the signaling pathways involved in IL-8 production from respiratory epithelial cells. Inhibition of PP2A using okadaic acid or gene knockdown using siRNA resulted in an augmentation of TNFalpha-induced IL-8 production. We also found that PP2A inhibition resulted in prolonged activation of JNK, p38, and ERK resulting in both increased transcriptional activation of the IL-8 promoter and posttranscriptional stabilization of IL-8 mRNA. Because TNFalpha had been shown to activate ceramide accumulation, and separate studies had linked ceramide with activation of PP2A, we hypothesized the pathway of TNFalpha-inducing ceramide to activate PP2A comprised an endogenous regulatory pathway. Inhibition of the immediate sphingomyelinase-dependent pathway as well as the de novo synthesis pathway of ceramide production reduced serine/threonine phosphatase activity and augmented IL-8 production. These data suggest that ceramide plays a role in activating PP2A to terminate ongoing IL-8 production. In summary, our data suggest that in respiratory epithelium, TNFalpha induces ceramide accumulation, resulting in subsequent activation of PP2A, which targets those kinases responsible for transcriptional activation of IL-8.
Cornell
Timothy T
TT
Division of Pediatric Critical Care Medicine, C. S. Mott Children's Hospital, Ann Arbor, MI 48109-0243, USA. ttcornel@med.umich.edu
Hinkovska-Galcheva
Vania
V
Sun
Lei
L
Cai
Qing
Q
Hershenson
Marc B
MB
Vanway
Susan
S
Shanley
Thomas P
TP
D002460:Cell Line; D002518:Ceramides / Q000096:biosynthesis / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects* / Q000201:enzymology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008168:Lung / Q000166:cytology*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D055550:Protein Stability / Q000187:drug effects; D034741:RNA, Small Interfering / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D002518:Ceramides; D016209:Interleukin-8; D016328:NF-kappa B; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D054648:Protein Phosphatase 2