Microsomal prostaglandin E synthase-2 is not essential for in vivo prostaglandin E2 biosynthesis

Prostaglandin E₂ (PGE₂) plays an important role in the normal physiology of many organ systems. Increased levels of this lipid mediator are associated with many disease states, and it potently regulates inflammatory responses. Three enzymes capable of in vitro synthesis of PGE₂ from the cyclooxygenase metabolite PGH₂ have been described. Here, we examine the contribution of one of these enzymes to PGE₂ production, mPges-2, which encodes microsomal prostaglandin synthase-2 (mPGES-2), by generating mice homozygous for the null allele of this gene. Loss of mPges-2 expression did not result in a measurable decrease in PGE₂ levels in any tissue or cell type examined from healthy mice. Taken together, analysis of the mPGES-2 deficient mouse lines does not substantiate the contention that mPGES-2 is a PGE₂ synthase.     

Impaired bone resorption to prostaglandin E2 in prostaglandin E receptor EP4-knockout mice

Prostaglandin E(2) (PGE(2)) acts as a potent stimulator of bone resorption. In this study, we first clarified in normal ddy mice the involvement of protein kinase A and induction of matrix metalloproteinases (MMPs) in PGE(2)-induced bone resorption, and then identified PGE receptor subtype(s) mediating this PGE(2) action using mice lacking each subtype (EP1, EP2, EP3, and EP4) of PGE receptor. In calvarial culture obtained from normal ddy mice, both PGE(2) and dibutyryl cyclic AMP (Bt(2)cAMP) stimulated bone resorption and induced MMPs including MMP-2 and MMP-13. Addition of an inhibitor of protein kinase A, H89, or an inhibitor of MMPs, BB94, significantly suppressed bone-resorbing activity induced by PGE(2.) In calvarial culture from EP1-, EP2-, and EP3-knockout mice, PGE(2) stimulated bone resorption to an extent similar to that found in calvaria from the wild-type mice. On the other hand, a marked reduction in bone resorption to PGE(2) was found in the calvarial culture from EP4-knockout mice. The impaired bone resorption to PGE(2) was also detected in long bone cultures from EP4-knockout mice. Bt(2)cAMP greatly stimulated bone resorption similarly in both wild-type and EP4-knockout mice. Induction of MMP-2 and MMP-13 by PGE(2) was greatly impaired in calvarial culture from EP4-knockout mice, but Bt(2)cAMP stimulated MMPs induction similarly in the wild-type and EP4-knockout mice. These findings suggest that PGE(2) stimulates bone resorption by a cAMP-dependent mechanism via the EP4 receptor.     

Up-regulation of prostaglandin E2 binding and of prostanoid release in rabbits producing antibodies against prostaglandin E2

German Giant rabbits successfully immunized against prostaglandin (PG) E2 as shown by a rise in antibody titers developed gastric mucosal lesions. Enzymatically dispersed gastric mucosal cells of these animals had a significantly enhanced production of PG E2 and PG I2 as measured by specific radioimmunoassays. This may be explained by an increased supply with endogenous arachidonic acid (as indicated by an enhanced phospholipase A2/LAT ratio) and by a higher activity of the subsequent PG forming enzymes (as indicated by a more effective stimulation of PG production by exogenous arachidonic acid). Gastric mucosal plasma membranes of immunized rabbits had significantly higher PG E2 binding capacity (108 +/- 9 fmol/mg protein) than those of nonimmunized rabbits (72 +/- 5 fmol/mg protein). The ligand affinity was not affected by immunization. Neither histamine-stimulated 14C-amino-pyrine uptake of isolated parietal cells as a marker for acid production nor its inhibition by PG E2 were influenced by receptor up-regulation. The increased eicosanoid release can be regarded as an endogenous defense mechanism against increased mucosal vulnerability caused by PG E2 scavenging. The potential role of PG E2 receptor up-regulation in support of this process remains to be established.     

Microglia-specific expression of microsomal prostaglandin E2 synthase-1 contributes to lipopolysaccharide-induced prostaglandin E2 production

Microsomal prostaglandin E2 synthase (mPGES)-1 is an inducible protein recently shown to be an important enzyme in inflammatory prostaglandin E2 (PGE2) production in some peripheral inflammatory lesions. However, in inflammatory sites in the brain, the induction of mPGES-1 is poorly understood. In this study, we demonstrated the expression of mPGES-1 in the brain parenchyma in a lipopolysaccharide (LPS)-induced inflammation model. A local injection of LPS into the rat substantia nigra led to the induction of mPGES-1 in activated microglia. In neuron-glial mixed cultures, mPGES-1 was co-induced with cyclooxygenase-2 (COX-2) specifically in microglia, but not in astrocytes, oligodendrocytes or neurons. In microglia-enriched cultures, the induction of mPGES-1, the activity of PGES and the production of PGE2 were preceded by the induction of mPGES-1 mRNA and almost completely inhibited by the synthetic glucocorticoid dexamethasone. The induction of mPGES-1 and production of PGE2 were also either attenuated or absent in microglia treated with mPGES-1 antisense oligonucleotide or microglia from mPGES-1 knockout (KO) mice, respectively, suggesting the necessity of mPGES-1 for microglial PGE2 production. These results suggest that the activation of microglia contributes to PGE2 production through the concerted de novo synthesis of mPGES-1 and COX-2 at sites of inflammation of the brain parenchyma.     

Cellular prostaglandin E2 production by membrane-bound prostaglandin E synthase-2 via both cyclooxygenases-1 and -2

Current evidence suggests that two forms of prostaglandin (PG) E synthase (PGES), cytosolic PGES and membrane-bound PGES (mPGES) -1, preferentially lie downstream of cyclooxygenase (COX) -1 and -2, respectively, in the PGE2 biosynthetic pathway. In this study, we examined the expression and functional aspects of the third PGES enzyme, mPGES-2, in mammalian cells and tissues. mPGES-2 was synthesized as a Golgi membrane-associated protein, and spontaneous cleavage of the N-terminal hydrophobic domain led to the formation of a truncated mature protein that was distributed in the cytosol with a trend to be enriched in the perinuclear region. In several cell lines, mPGES-2 promoted PGE2 production via both COX-1 and COX-2 in the immediate and delayed responses with modest COX-2 preference. In contrast to the marked inducibility of mPGES-1, mPGES-2 was constitutively expressed in various cells and tissues and was not increased appreciably during tissue inflammation or damage. Interestingly, a considerable elevation of mPGES-2 expression was observed in human colorectal cancer. Collectively, mPGES-2 is a unique PGES that can be coupled with both COXs and may play a role in the production of the PGE2 involved in both tissue homeostasis and disease.     

Regulation of prostaglandin E2 biosynthesis by inducible membrane-associated prostaglandin E2 synthase that acts in concert with cyclooxygenase-2

Here we report the molecular identification of membrane-bound glutathione (GSH)-dependent prostaglandin (PG) E(2) synthase (mPGES), a terminal enzyme of the cyclooxygenase (COX)-2-mediated PGE(2) biosynthetic pathway. The activity of mPGES was increased markedly in macrophages and osteoblasts following proinflammatory stimuli. cDNA for mouse and rat mPGESs encoded functional proteins that showed high homology with the human ortholog (microsomal glutathione S-transferase-like 1). mPGES expression was markedly induced by proinflammatory stimuli in various tissues and cells and was down-regulated by dexamethasone, accompanied by changes in COX-2 expression and delayed PGE(2) generation. Arg(110), a residue well conserved in the microsomal GSH S-transferase family, was essential for catalytic function. mPGES was functionally coupled with COX-2 in marked preference to COX-1, particularly when the supply of arachidonic acid was limited. Increased supply of arachidonic acid by explosive activation of cytosolic phospholipase A(2) allowed mPGES to be coupled with COX-1. mPGES colocalized with both COX isozymes in the perinuclear envelope. Moreover, cells stably cotransfected with COX-2 and mPGES grew faster, were highly aggregated, and exhibited aberrant morphology. Thus, COX-2 and mPGES are essential components for delayed PGE(2) biosynthesis, which may be linked to inflammation, fever, osteogenesis, and even cancer.     

Influence of a type 2 diabetes associated prostaglandin E synthase 2 polymorphism on blood prostaglandin E2 levels

In this study we tested whether the type 2 diabetes mellitus associated prostaglandin E synthase 2 arginine to histidine polymorphism at position 298 (R298H) influences prostaglandin E2 levels in humans. Fasting prostaglandin E2 was determined in the blood of subjects carrying different genotypes of the R298H polymorphism. Subjects were matched by sex, age, and body mass index. No differences in prostaglandin E2 levels were found with respect to genotypes when considering the whole group. Male homozygous histidine carriers showed elevated prostaglandin E2 levels compared to heterozygous carriers and homozygous arginine carriers (188.2±42.4 vs. 80.4±26.5 pg/ml, p=0.021; and vs. 92.9±15.3 pg/ml, p=0.11). These differences were not evident in female subjects. In contrast, 6-keto-prostaglandin F1alpha levels as independent marker of arachidonic acid metabolism showed ambiguous results. Nevertheless, preliminary evidence of the prostaglandin E synthase 2 R298H polymorphism possibly influencing prostaglandin E2 blood levels in a gender-specific manner was obtained.     

Urinary excretion of prostaglandin E2 and prostaglandin F2alpha in potassium-deficient rats

Potassium-deficiency was induced in rats by dietary deprivation of potassium. The animals became polyuric and urine osmolality decreased more then three-fold compared to controls. Urinary excretion of prostaglandin E2 (PGE2) and prostaglandin F2alpha (PGF2alpha) did not increase during 2 weeks of potassium depletion. Partial inhibition of renal prostaglandin synthesis by meclofenamate did not increase the urine osmolality after water deprivation. These results make unlikely the hypothesis that the polyuria of potassium-deficiency, is the result of enhanced renal synthesis of prostaglandins with subsequent antagonism of the hydro-osmotic effect of vasopressin. Male animals consistently excreted less PGE2 than female animals.     

Amniotic fluid prostaglandin E2 in preterm labor

These studies were designed to determine amniotic fluid concentrations of prostaglandin E2 (PGE) in women with preterm labor. Amniotic fluid was retrieved by transabdominal amniocentesis from 68 women with preterm labor (less than 37 weeks). Patients were divided into three groups according to the response to tocolysis and the presence or absence of an intraamniotic infection. Amniotic fluid concentrations of PGE2 were significantly greater in women with preterm labor and intraamniotic infection than in women without infection. Patients unresponsive to tocolysis without intraamniotic infection had a significantly greater concentration of PGE2 in amniotic fluid than those responsive to tocolysis.     

Dietary fat and salivary prostaglandin E2

A study of the relation between dietary fat intake and salivary prostaglandin E2 was undertaken in Transkei, South Africa. Samples of saliva were obtained from (1) Transkeians on a very low fat diet, (2) Transkeians on a low fat diet, (3) Transkeians on a medium fat diet, (4) British patients. Salivary PGE2 means for the groups were (1) 2357 pg/ml, (2) 2020 pg/ml, (3) 733 pg/ml, (4) 312.5 pg/ml. Differences between groups 1 and 3, 1 and 4, and 2 and 4 were significant. Rural Transkeians on a low-fat diet have an elevated level of PGE2 in saliva. As fat increases in the diet, PGE2 in saliva tends towards the level found in those who eat a western diet. An increase in the level of PGE2 production in the upper gastrointestinal tract and in the tissues of the body may be a factor in promoting cancer of the esophagus and diseases favored by Th1 immune dysfunction.     

Theoretical study of prostaglandin E2 conformation

The theoretical conformational analysis of PGE2 has been performed using semiempirical approach and fixed values of the bond length and valence angles. A set of low-energy conformations was obtained for the alpha-chain and the whole PGE2 molecule. Unlike the PGE1 molecule, PGE2 alpha-chain prefers semifolded conformations. The hairpin structures with the parallel orientation of alpha- and omega-chains are the lowest-energy conformations of PGE2 conformations of PGE1 and PGE2. Sterically similar explain the similar biological activity of the two molecules.     

Regulation of agonist-induced prostaglandin E1 versus prostaglandin E2 production. A mass analysis.

Prostaglandin E1 (PGE1) and prostaglandin E2 (PGE2), derived by enzymatic oxidation of cellular dihomogammalinolenic acid (DHLA) and arachidonic acid (AA), respectively, have diverse and, at times, distinct biological actions. It has been suggested that PGE1 specifically inhibits a variety of inflammatory processes, and, in light of the potential therapeutic benefit of PGE1 and its fatty acid precursor in inflammatory disorders, there is growing interest in the biochemical mechanisms which determine the balance between PGE1 and PGE2 synthesis. Metabolic studies in this area have been hampered by the difficulties in measuring the extremely small masses of these prostaglandins which are generated in cell culture systems. We studied the regulation of PGE1 versus PGE2 synthesis using an essential fatty acid-deficient, PGE-producing, mouse fibrosarcoma cell line, EFD-1. Because EFD-1 cells contain no endogenous AA or DHLA, we were able to replete the cells with AA and DHLA of known specific activities; thus, the mass of both cellular AA and DHLA, and synthesized PGE1 and PGE2, could be accurately determined. The major finding of this study is that production of PGE2 was highly favored over production of PGE1 due to preferential incorporation of AA versus DHLA into, and release from, the total cellular phospholipid pool. Further, we correlated the selective release of AA versus DHLA from total cellular phospholipids with the selective incorporation of AA versus DHLA into specific phospholipid pools. In addition, we showed that conversion of DHLA to AA by delta 5 desaturase was enhanced by increasing the cellular mass of n-6 fatty acids and by increasing the cell proliferative activity. Together, these results indicate that the relative abundance of PGE2 versus PGE1 in vivo is not merely a function of the relative abundance of AA versus DHLA in tissues, but also relates to markedly different cellular metabolism of these two fatty acids.     

Treatment of vasospastic disease with prostaglandin E1.

Prostaglandin E1, a vasodilator and potent inhibitor of platelet aggregation, was administered to 26 patients with severe vasospastic disease of the hands. Patients tolerated infusions well and reported appreciable symptomatic improvement. Five of eight ischaemic ulcers healed in six weeks. Non-invasive studies of blood flow were used to observe haemodynamic changes during and after infusions. The Doppler-derived radial artery pulsatility index fell from 8.8 +/- 0.6 to 4.6 +/0 0.5 (mean +/- SEM), indicating hand vasodilatation. This fall was maintained 24 hours after infusion (5.9 +/- 0.9), but the index had returned to normal values at two weeks. The amplitude of finger pulse volume recordings increased (5.6 +/- 0.7 mm to 23.8 +/- 3.4 mm) and was raised two and six weeks after infusion (13.5 +/- 2.1 mm). Hand temperature measured by infrared radiometry also increased (27.4 +/- 0.7 degrees C to 31.2 +/- 1.2 degrees C). Intensity of digital vasospasm induced by cold water challenge was not objectively affected by prostaglandin E1 despite an increased finger temperature after infusion. Nevertheless, patients reported less frequent and severe attacks. Prostaglandin E1 given by central venous infusion is a safe new vasoactive agent that can produce appreciable symptomatic improvement by increasing digital perfusion, which may last for several weeks after treatment. Further study will define its mode of action and its place in the management of peripheral vascular disease.     

Urinary prostaglandin E2 in the newborn and infant

Prostaglandin E(2) (PGE(2)) belongs to a family of biologically active lipids derived from the 20-carbon essential fatty acids. Renal PGE(2) is involved in the development of the kidney; it also contributes to regulate renal perfusion and glomerular filtration rate, and controls water and electrolyte balance. Furthermore, this mediator protects the kidney against excessive functional changes during the transition from fetal to extrauterine life, when it counteracts the vasoconstrictive effects of high levels of angiotensin II and other mediators. There is evidence that PGE(2) plays an important pathophysiological role in neonatal conditions of renal stress, and in congenital or acquired nephropaties. Thus, measurement of urinary PGE(2) as an index of renal synthesis of this primary prostaglandin may represent a non-invasive and sensitive method of investigating the homeostatic function of the kidney in early life. The aim of this literature review is to examine urinary PGE(2) as a non-invasive marker of renal homeostasis in the newborn and infant under both physiological and pathological conditions, or during treatments with widely used, potentially toxic drugs.