HYPEROSMOLARITY STIMULATES PROSTAGLANDIN SYNTHESIS AND CYCLOOXYGENASE-2 EXPRESSION IN ACTIVATED RAT-LIVER MACROPHAGES

The effect of aniso-osmotic exposure on the level of inducible cycloox ygenase (Cox-2)and on prostanoid synthesis was studied in cultured rat liver macrophages (Kupffer cells). In lipopolysaccharide (LPS)- or ph orbol 12-myristate 13-acetate-stimulated Kupffer cells, hyperosmotic ( 355 mosmol/l) exposure, due to addition of NaCl or impermeant sugars, markedly increased prostaglandin (PG) E(2), D-2 and thromboxane B-2 sy nthesis in a time- and osmolarity-dependent manner. Increased prostano id production was observed about sh after exposure to LPS in hyperosmo tic medium compared to Kupffer cells treated with LPS under normotonic (305 mosmol/l) conditions. A similar stimulatory effect of hyperosmol arity on PGE(2) production was also seen when arachidonate was added e xogenously. Hyperosmotic stimulation of PGE(2) production was accompan ied by a strong induction of Cox-2 mRNA levels and an increase in immu noreactive Cox-2, whereas the levels of immunoreactive phospholipase A (2) and cyclooxygenase-1 did not change significantly. Dexamethasone, indomethacin and the selective Cox-2 inhibitor, NS-398, abolished the hypertonicity-induced stimulation of PGE(2) formation; dexamethasone a lso prevented the increase in Cox-2 mRNA and protein. The increase of immunoreactive Cox-2 lasted for about 24 h and was also blocked by act inomycin D or cycloheximide, but not by brefeldin A. Tunicamycin or tr eatment with endoglucosidase H reduced the molecular mass of hypertoni city-induced Cox-2 by 5kDa. Tunicamycin treatment also suppressed the hypertonicity-induced stimulation of PGE(2) production. The hyperosmol arity/ LPS-induced stimulation of prostaglandin formation was partly s ensitive to protein kinase C inhibition but was not accompanied by an increase in the cytosolic free Ca2+ concentration. The data suggest th at osmolarity may be a critical factor in the regulation of Cox-2 expr ession and prostanoid production in activated rat liver macrophages.