Re-evaluation of thin layer chromatography as an alternative method for the quantification of prostaglandins from rat Kupffer cells.

In contrast to conventionally used immunoassays, thin layer chromatography (TLC)--by prelabeling of cells with radioactive arachidonic acid (AA)--allows to differentiate between cellularly built and added prostanoids and thus to investigate feedback effects of prostanoids on their own release. PGD2, TXB2 and PGE2 released from zymosan-stimulated Kupffer cells were separated with distinct RF-values, corresponding to those of the pure substances. Quantification of PGD2 and PGE2 gave comparable results with TLC and immunoassays, but measurement in the presence of added prostanoids was only possible with TLC.

Modulation of glucokinase expression by hypoxia-inducible factor 1 and upstream stimulatory factor 2 in primary rat hepatocytes.

Glucokinase (GK) is the key enzyme of glucose utilization in liver and is localized in the less aerobic perivenous area. Until now, the O2-responsive elements in the liver-specific GK promoter are unknown, and therefore the aim of this study was to identify the O2-responsive element in this promoter. We found that the GK promoter sequence -87/-80 matched the binding site for hypoxia inducible factor 1 (HIF-1) and upstream stimulatory factor (USF).

A Fenton reaction at the endoplasmic reticulum is involved in the redox control of hypoxia-inducible gene expression.

It has been proposed that hydroxyl radicals (.OH) generated in a perinuclear iron-dependent Fenton reaction are involved in O(2)-dependent gene expression. Thus, it was the aim of this study to localize the cellular compartment in which the Fenton reaction takes place and to determine whether scavenging of.

Differences in the involvement of prostanoids from Kupffer cells in the mediation of anaphylatoxin C5a-, zymosan-, and lipopolysaccharide-dependent hepatic glucose output and flow reduction.

Various inflammatory stimuli such as anaphylatoxin C5a, zymosan, and lipopolysaccharides (LPSs) have been reported both to enhance glucose output in the perfused rat liver and to induce prostanoid (ie, prostaglandin and thromboxane) release from Kupffer cells, the resident liver macrophages. Because prostanoids can enhance glucose output from hepatocytes, it was the aim of this study to compare the possible roles of prostanoids released after C5a, zymosan, and LPS in the mediation of hepatic glucose output. In perfused livers both C5a and zymosan immediately enhanced glucose output, reduced flow, and induced prostanoid overflow into the hepatic vein, but with different quantities and kinetics.

Inhibition of prostaglandin D2 clearance in rat hepatocytes by the thromboxane receptor antagonists daltroban and ifetroban and the thromboxane synthase inhibitor furegrelate.

Prostanoids, i.e. prostaglandins and thromboxane, regulate liver-specific functions both in homeostasis and during defense reactions.

Reactive oxygen species modulate HIF-1 mediated PAI-1 expression: involvement of the GTPase Rac1.

The hypoxia-inducible transcription factor HIF-1 mediates upregulation of plasminogen activator inhibitor-1 (PAI-1) expression under hypoxia. Reactive oxygen species (ROS) have also been implicated in PAI-1 gene expression. However, the role of ROS in HIF-1-mediated regulation of PAI-1 is not clear.

Regulation of the hypoxia-dependent plasminogen activator inhibitor 1 expression by MAP kinases.

Mitogen-activated protein kinases (MAPKs) and protein kinase B (PKB) mediate growth and stress signals and have been implicated in the hypoxic response. Under hypoxic conditions, the expression of plasminogen activator inhibitor-1 (PAI-1) is mainly controlled by the hypoxia-inducible factor HIF-1. However, the role of MAPKs and PKB in HIF-1-mediated PAI-1 regulation is not clear.

Hypoxia-inducible factor-1 and hypoxia response elements mediate the induction of plasminogen activator inhibitor-1 gene expression by insulin in primary rat hepatocytes.

The expression of the plasminogen activator inhibitor-1 (PAI-1) gene is enhanced by insulin both in vivo and in various cell types. Because insulin exerts a number of its biologic activities via the phosphatidylinositol 3-kinase and protein kinase B (PI3K/PKB) signaling pathway, it was the aim of the present study to investigate the role of the PI3K/PKB pathway in the expression of the PAI-1 gene and to identify the insulin responsive promoter sequences. It was shown that the induction of PAI-1 mRNA and protein expression by insulin and mild hypoxia could be repressed by the PI3K inhibitor wortmannin.

Signaling cross-talk between hypoxia and glucose via hypoxia-inducible factor 1 and glucose response elements.

The substrates oxygen and glucose are important for the appropriate regulation of metabolism, angiogenesis, tumorigenesis and embryonic development. The knowledge about an interaction between these two signals is limited. We demonstrated that the regulation of glucagon receptor, insulin receptor and L-type pyruvate kinase (L-PK) gene expression in liver is dependent upon a cross-talk between oxygen and glucose.

Inhibition by prostaglandin E(2) of anaphylatoxin C5a- but not zymosan-induced prostanoid release from rat Kupffer cells.

The proinflammatory anaphylatoxin C5a induces the release of prostanoids, ie, prostaglandins (PG) and thromboxane (TX), from the resident liver macrophages (Kupffer cells [KC]). Because KC themselves express prostanoid receptors, prostanoids--besides having paracrine functions--might regulate their own release in an autocrine loop. So far, such a possible feedback regulation has not been investigated systematically, probably because of methodological difficulties to measure newly synthesized prostanoids in the presence of added prostanoids.

Activation of glucokinase gene expression by hepatic nuclear factor 4alpha in primary hepatocytes.

Glucokinase (GK) is a key enzyme for glucose utilization in liver and shows a higher expression in the perivenous zone. In primary rat hepatocytes, the GK gene expression was activated by HNF (hepatic nuclear factor)-4alpha via the sequence -52/-39 of the GK promoter. Venous pO2 enhanced HNF-4 levels and HNF-4 binding to the GK-HNF-4 element.

Induction of anaphylatoxin C5a receptors in rat hepatocytes by lipopolysaccharide in vivo: mediation by interleukin-6 from Kupffer cells.

Background & Aims: In normal rat liver, anaphylatoxin C5a induces glucose output from hepatocytes indirectly via prostanoids released from Kupffer cells. Correspondingly, it was found that hepatocytes, in contrast to Kupffer cells, did not express C5a receptors. Lipopolysaccharide (LPS) has been reported to enhance C5a receptor expression in murine livers.