Cox-2 and osteopontin in cocultured platelets and mesangial cells: role of glucocorticoids.
BACKGROUND
Glomerular inflammation is characterized by a consecutive infiltration of immunoreactive cells. To mimic the early phase of glomerular injury, a coculture system of platelets and rat renal mesangial cells was established. As prototypes, the inflammation-related proteins cyclooxygenase-2 (Cox-2) and the chemotactic protein osteopontin (OPN) were investigated.


METHODS
The expression of OPN and Cox-2 mRNA and protein was determined by Northern and Western blot analyses.


RESULTS
Coincubation of platelets and mesangial cells led to a rapid, transient induction of Cox-2 mRNA, which peaked at two hours, whereas OPN and monocyte chemoattractant protein-1 (MCP-1) were induced at later time points. The induction of Cox-2 mRNA was concentration dependent and highly reproducible when platelets of different donors were investigated. Partial Cox-2 induction was observed when supernatants of preactivated platelets were incubated with mesangial cells. The inhibition of the signaling pathways of platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) or interference with Gi-protein signaling partially inhibited platelet-induced Cox-2 expression. Down-regulation of protein kinase C (PKC), which is a common signaling module in many pathways leading to Cox-2 induction, almost completely abrogated platelet-induced Cox-2 expression. The time pattern of Cox-2 and OPN expression suggested that Cox-2 might play a role in OPN induction. The up-regulation of OPN was dependent on de novo protein synthesis and was induced by high levels of exogenous prostaglandin E2 (PGE2; 10 micromol/L). Endogenous PGE2, however, proved not to be essential for OPN mRNA expression, because inhibition of Cox activity did not change OPN mRNA levels. Dexamethasone inhibited Cox-2 mRNA induction but increased OPN mRNA and protein expression.


CONCLUSIONS
These data indicate that Cox-2 and OPN are independently up-regulated upon interaction of platelets and mesangial cells.
Goppelt-Struebe
M
M
Medizinische Klinik IV, Universität Erlangen-Nürnberg, Erlangen, and Institut für Anatomie II, Albert-Ludwigs Universität Freiburg, Freiburg, Germany. Goppelt-Struebe@rzmail.uni-erlangen.de
Wiedemann
T
T
Heusinger-Ribeiro
J
J
Vucadinovic
M
M
Rehm
M
M
Pröls
F
F
D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D018920:Coculture Techniques; D051546:Cyclooxygenase 2; D003907:Dexamethasone / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000502:physiology; D005920:Glomerular Mesangium / Q000166:cytology / Q000201:enzymology* / Q000502:physiology; D005938:Glucocorticoids / Q000494:pharmacology / Q000502:physiology*; D006133:Growth Substances / Q000502:physiology; D006801:Humans; D007527:Isoenzymes / Q000097:blood* / Q000235:genetics; D008565:Membrane Proteins; D053495:Osteopontin; D011451:Prostaglandin-Endoperoxide Synthases / Q000097:blood* / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D012795:Sialoglycoproteins / Q000097:blood* / Q000235:genetics; D015854:Up-Regulation
D005938:Glucocorticoids; D006133:Growth Substances; D007527:Isoenzymes; D008565:Membrane Proteins; D012333:RNA, Messenger; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; C506897:Spp1 protein, rat; D053495:Osteopontin; D003907:Dexamethasone; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases