Shunting of prostanoid biosynthesis in microsomal prostaglandin E synthase-1 null embryo fibroblasts: regulatory effects on inducible nitric oxide synthase expression and nitrite synthesis.
Microsomal prostaglandin (PG) E synthase (mPGES)-1 is an inducible enzyme that acts downstream of cyclooxygenase (COX) and specifically catalyzes the conversion of prostaglandin (PG)H2 to PGE2, most prominently in inflammatory conditions. Specific inhibitors of mPGES-1 are not yet available, however, mice with genetic deletion of mPGES-1 have been generated that have given insight into the specific role of mPGES-1 in eicosanoid biosynthesis in vivo and in peritoneal macrophages. We created mouse embryo fibroblast (MEF) cell lines that would facilitate investigation of the effect of mPGES-1 genetic deletion on prostanoid biosynthesis in fibroblast lineage cells and its subsequent effect on the expression of inducible NOS (iNOS) and nitrite biosynthesis using cells derived from mPGES-1 wild-type (WT), heterozygous (Het), and null mice. The results show that genetic deletion of mPGES-1 results in a dramatic decrease in PGE2 production in Het and null MEFs under basal conditions and after stimulation with interleukin (IL)-1beta, suggesting that mPGES-1 is critically important for PGE2 production. Furthermore, we show that mPGES-1 gene deletion results in diversion of prostanoid production from PGE2 to 6-keto PGF1alpha (the stable metabolic product of PGI2; prostacyclin) in a gene dose-dependent manner in Het and null MEFs compared with their WT counterparts, suggesting a shunting phenomenon within the arachidonic acid (AA) metabolic pathway. In addition, we show that mPGES-1 gene deletion and subsequent decrease in PGE2 levels results in a differential induction profile of iNOS and nitrite levels (the stable breakdown product of nitric oxide (NO) in mPGES-1 WT MEFs compared with null MEFs. These results provide important information regarding the therapeutic potential for pharmacologic inhibition of mPGES-1 in inflammatory conditions.
Kapoor
Mohit
M
Department of Internal Medicine, Rheumatology Division, Kentucky Clinic, University of Kentucky, Lexington, KY 40536-0284, USA.
Kojima
Fumiaki
F
Qian
Min
M
Yang
Lihua
L
Crofford
Leslie J
LJ
D000818:Animals; D019746:Intramolecular Oxidoreductases / Q000172:deficiency* / Q000235:genetics*; D051379:Mice; D008811:Mice, Inbred DBA; D018345:Mice, Knockout; D008861:Microsomes / Q000201:enzymology*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics*; D009573:Nitrites / Q000378:metabolism*; D000071876:Prostaglandin-E Synthases; D011453:Prostaglandins / Q000096:biosynthesis*
D009573:Nitrites; D011453:Prostaglandins; D052247:Nitric Oxide Synthase Type II; D019746:Intramolecular Oxidoreductases; D000071876:Prostaglandin-E Synthases; C000605865:Ptges protein, mouse