Colocalization of prostaglandin F(2alpha) receptor FP and prostaglandin F synthase-I in the spinal cord.

Prostaglandin F(2alpha) is synthesized by prostaglandin F synthase, which exists in two types, prostaglandin F synthase I (PGFS I) and prostaglandin F synthase II (PGFS II). Prostaglandin F(2alpha) binds to its specific receptor, FP. Our previous immunohistochemical study showed the distinct localization of prostaglandin F synthases in rat spinal cord. PGFS I exists in neuronal somata and dendrites in the gray substance, and PGFS II exists in ependymal cells and tanycytes surrounding the central canal. Both enzymes are also present in endothelial cells of blood vessels in the white and gray substances of the spinal cord. In this study, we found that FP localizes in neuronal somata and dendrites but not in ependymal cells, tanycytes, or endothelial cells. Immunohistochemical analysis of serial sections showed the colocalization of FP and PGFS I. FP immunoreactivity was intense in spinal laminae I and II of the dorsal horn, a connection site of pain transmission, and was similar to that of PGFS I in neuronal elements. These findings suggest that prostaglandin F(2alpha) synthesized in the neuronal somata and dendrites exert an autocrine action there.

Immunocytochemical localization of prostaglandin F synthase II in the rat spinal cord.

Prostaglandin F synthase has at least two isozymes, i.e. prostaglandin F synthase I and II. Recently, we demonstrated immunocytochemically that prostaglandin F synthase I was localized in neuronal dendrites and somata, and in endothelial cells of blood vessels in the whole area of rat spinal cord. In the present study, we immunocytochemically localized prostaglandin F synthase II in ependymal cells and tanycytes surrounding the central canal and in endothelial cells of blood vessels, but not in any neuronal elements at all segmental levels of the rat spinal cord. Immunoelectron microscopy and confocal laser scanning microscopy confirmed these findings and further revealed that strong immunoreactivity was found in the basal processes of the tanycytes. Our present and recent studies using antibodies against the two isozymes of prostaglandin F synthase clearly indicated that they were localized differentially in ependymal (prostaglandin F synthase II) and neuronal elements (prostaglandin F synthase I), but were co-localized in blood vessels in the rat spinal cord. The distinct localization of the two isozymes suggests that prostaglandin F(2) has different transcellular biological actions via different cell groups.

Immunocytochemical localization of lung-type prostaglandin F synthase in the rat spinal cord.

Prostaglandin F synthase, producing prostaglandin F(2 alpha) and 9 alpha,11 beta-prostaglandin F(2), has at least two isozymes, lung-type and liver-type ones. The present study including double immunolabelling with microtubule-associated protein 2 indicated that the lung-type isozyme was present in neuronal dendrites and somata of gray matter (relatively intense in lamina I and II in dorsal horn, and IX in ventral horn) and vascular endothelial cells in the rat spinal cord at all segmental levels.

cDNA cloning, expression and characterization of human prostaglandin F synthase.

A cDNA clone of prostaglandin F synthase (PGFS) was isolated from human lung by using cDNA of bovine lung-type PGFS as a probe and its protein expressed in Escherichia coli was purified to apparent homogeneity. The human PGFS catalyzed the reduction of prostaglandin (PG) D2, PGH2 and phenanthrenequinone (PQ), and the oxidation of 9alpha,11beta-PGF2 to PGD2. The kcat/Km values for PGD2 and 9alpha,11beta-PGF2 were 21000 and 1800 min(-1) mM(-1), respectively, indicating that the catalytic efficiency for PGD2 and 9alpha,11beta-PGF2 was the highest among the various substrates, except for PQ. The PGFS activity in the cytosol of human lung was completely absorbed with antihuman PGFS antiserum. Moreover, mRNA of PGFS was expressed in peripheral blood lymphocytes and the expression in lymphocytes was markedly suppressed by the T cell mitogen concanavalin A. These results support the notion that human PGFS plays an important role in the pathogenesis of allergic diseases such as asthma.

Identification of prostaglandin F-producing cells in the liver.

Prostaglandin (PG) F synthase forms PGF(2alpha) and 9alpha, 11beta-PGF2 from PGH2 and PGD2, respectively. PGH2 is synthesized from arachidonic acid by cyclooxygenase (COX) and then metabolized to various PGs and thromboxane by specific enzymes. PGD2 is synthesized from PGH2 by PGD synthase. To identify PGF2-producing cells in the rat liver, the occurrence and localization of PGF synthase and COX were studied with immunochemical and immunohistochemical techniques using anti-liver-type PGF synthase and anti-COX antibodies. In Western blot analyses, positive bands of liver-type PGF synthase and constitutive COX-1 were observed at positions approximately 37 kDa and 70-72 kDa, respectively. However, inducible COX-2 was not detected. In the immunohistochemical study, PGF synthase was present in the cytoplasm of the sinusoidal endothelial cells. COX-1 was present on the membranes of the nucleus and endoplasmic reticulum of the endothelial cells and Kupffer cells. Double immunostaining for PGF synthase and COX-1 showed that both enzymes were present in the same endothelial cells. These results suggest that the main site of PGF2 synthesis in the liver is the sinusoidal endothelial cell.