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Prostaglandin D(2) (PGD(2)) is the most abundant prostaglandin produced in the brain. It is a metabolite of arachidonic acid and synthesized by prostaglandin D(2) synthases (PGDS) via the cyclooxygenase pathway. Two distinct types of PGDS have been identified: hematopoietic prostaglandin D synthase (H-PGDS) and lipocalin-type prostaglandin D synthase (L-PGDS). Because relatively little is known about the role of L-PGDS in the CNS, here we examined the outcomes in L-PGDS knockout and wild-type (WT) mice after two different cerebral ischemia models, transient middle cerebral artery (MCA) occlusion (tMCAO) and permanent distal middle cerebral artery occlusion (pMCAO). In the tMCAO model, the MCA was occluded with a monofilament for 90 min and then reperfused for 4 days. In the pMCAO model, the distal part of the MCA was permanently occluded and the mice were sacrificed after 7 days. Percent corrected infarct volume and neurological score were determined after 4 and 7 days, respectively. L-PGDS knockout mice had significantly greater infarct volume and brain edema than did WT mice after tMCAO (P

Prostaglandin D 2 is one of the major prostanoids formed from [ 14 C]arachidonic acid by the central nervous system. The aim of the present study is to specify the prostaglandin D 2 biosynthetic capacity in the chick spinal cord and to identify the cell type involved in this synthesis. A highly specific and sensitive enzyme immunoassay allowed us to demonstrate that the amount of newly formed prostaglandin D 2 increases proportionally with the concentration of free arachidonic acid of either exogenous or endogenous origin and reaches concentration values ranging from 10 −9 to 10 −6 M. The sites of prostaglandin D 2 synthesis were localized in Vibratome sections of spinal cord after incubation with antibodies raised against glutathione-independent prostaglandin D synthase; controls were performed with anti-glutathione-dependent prostaglandin D synthase antibodies and non-immune rabbit or goat serum. After immunoprocessing, electron microscope examination revealed that the specific immunoreactivity was confined to small neurons of laminae II and III in the dorsal horn and to motoneurons in the ventral horn of the spinal cord. The immunodeposits were associated with rough endoplasmic reticulum profiles distributed throughout the dorsal horn neurons or restricted to limited subsurface areas of perikarya and dendrites in motoneurons. Since the immunoreactive neurons in the dorsal horn were closely related to blood capillaries, prostaglandin D 2 may be suspected to play a role in the regulation of the microcirculation. The accumulation of prostaglandin D synthase in motoneuron areas facing astrocytic membrane stacks suggests that prostaglandin D 2 could interact with astrocytic functions.

Cerebral ischemia/reperfusion injury is characterized by the development of inflammatory response, in which vascular macrophages and endogenous microglia are involved. Recent studies showed marked induction of hematopoietic prostaglandin D synthase (HPGDS) after ischemic/reperfusion injury and its localization in microglia, but the molecular mechanism(s) of HPGDS actions in cerebral ischemia is not clear. To clarify the role of HPGDS in cerebral ischemia, C57BL/6 mice and bone marrow chimera mice with cerebral ischemia/reperfusion injury were treated with (4-benzhydryloxy-(1) {3-(1H-tetrazol-5-yl)-propyl}piperidine (HQL-79), a specific inhibitor of HPGDS. The bone marrow chimera mice exhibit expression of enhanced green fluorescent protein (EGFP) in bone marrow/blood-derived monocytes/macrophages. Mice were subjected to ischemia/reperfusion and either treated with HQL-79 (n=44) or vehicle (n=44). Brain sections prepared at 72 h and 7 days after reperfusion were analyzed for neuronal nuclei (NeuN), HPGDS, ionized calcium-binding adapter molecule 1 (Iba1), inducible NO synthase (iNOS), nitrotyrosine, nuclear factor kappa B (NF-kB) and cyclooxygenase-2 (COX-2). The mortality rate (80%) and infarct size were larger in HQL-79- than vehicle-treated mice (58.7+/-8.5 versus 45.2+/-4.9 mm(3); mean+/-SEM, P0.0001) at 7 days after reperfusion. HQL-79 reduced NeuN expression in the transition area and Iba1 expression (P0.0001) in the ischemic peri- and penumbra area, but increased COX-2 (P0.05) and NF-kB expression (P0.05) in ischemic penumbra and increased formation of nitrotyrosine (P0.0001) and iNOS (P0.0001) in the ischemic core area at 72 h and 7 days after reperfusion. In EGFP chimera mice, HQL-79 increased the migration of Iba1/EGFP-positive bone marrow-derived monocytes/macrophages, and simultaneously upregulated iNOS expression in the ischemic core area (P0.0001), but increased intrinsic microglia/macrophages in ischemic peri-area and penumbra (P0.0001) at 72 h and 7 days after reperfusion, suggesting involvement of monocytes/macrophages in HQL-79-induced expansion of ischemic injury. Our results demonstrated that the neuroprotective effects of HPGDS in our model are mediated by suppression of activation and infiltration of inflammatory cells.

Hematopoietic prostaglandin D synthase is a key enzyme in synthesis of prostaglandin D. Hematopoietic prostaglandin D synthase is expressed in microglia of the developing mouse brain. This study determined the serial changes and cellular localization of hematopoietic prostaglandin D synthase, and its role in cerebral ischemia/reperfusion injury using C57BL/6 mice (n=84) and bone marrow chimera mice (n=16). The latter mice were selected based on their expression of enhanced green fluorescent protein in bone marrow/blood-derived monocytes/macrophages. The middle cerebral artery was occluded for 60 min, followed by reperfusion. Hematopoietic prostaglandin D synthase expression was examined by immunohistochemistry and Western blotting. Hematopoietic prostaglandin D synthase-positive cells were mainly expressed in the peri-ischemic area at 12 h (P

Peripheral inflammation causes upregulation of cyclooxygenase in the spinal cord and subsequent increase in prostaglandin biosynthesis. However, prostaglandin synthases, which are downstream of cyclooxygenase control the type of prostaglandin that is formed predominantly. Since there is little known about the regulation of prostaglandin synthases, the present study was conducted in order to determine the effect of endotoxin treatment on the expression of messenger RNA encoding interleukin 1β, cyclooxygenase-2, and prostaglandin synthases mediating the formation of prostaglandin E2 (membrane bound prostaglandin E synthase) and prostaglandin D2 (lipocalin prostaglandin D synthase) in spinal cord, dorsal root ganglia and skin of rats. Endotoxin (2 mg/kg i.p.) induced the expression of interleukin-1β, cyclooxygenase-2, and membrane bound prostaglandin E synthase messenger RNA in spinal cord, dorsal root ganglia, and skin as determined by reverse transcription polymerase chain reaction. In contrast, basal expression of lipocalin prostaglandin D synthase messenger RNA in spinal cord and dorsal root ganglia was not significantly altered by endotoxin. Dexamethasone (1 mg/kg s.c. at −18 h and −1 h) attenuated the effect endotoxin on the expression of interleukin-1β, cyclooxygenase-2, and membrane bound prostaglandin E synthase messenger RNA in all tissues investigated, but did not significantly influence expression of lipocalin prostaglandin D synthase mRNA in spinal cord and dorsal root ganglia. In situ hybridisation histochemistry showed endotoxin-induced expression of cyclooxygenase-2 and membrane bound prostaglandin E synthase messenger RNA throughout gray and white matter of spinal cord sections. In dorsal root ganglia, expression of membrane bound prostaglandin E synthase seemed primarily located to non-neuronal cells, while cyclooxygenase-2 messenger RNA was not detectable. The results show that the immune response elicited by endotoxin induced cyclooxygenase-2 and membrane bound prostaglandin E synthase, but not lipocalin prostaglandin D synthase messenger RNA in spinal cord and dorsal root ganglia of rats. The distribution of cyclooxygenase-2 and membrane bound prostaglandin E synthase messenger RNA expressing cells suggests major involvement of non-neuronal cells in spinal prostaglandin biosynthesis. Determination of the regulation of enzymes downstream of cyclooxygenase at the messenger RNA level may represent a valuable tool to investigate effects of analgesic/anti-inflammatory drugs on the regulation of spinal prostaglandin biosynthesis.