Application of a heat- and steam-generating sheet increases peripheral blood flow and induces parasympathetic predominance.

To promote the practical application of a Japanese traditional medical treatment, such as hot compresses, we developed a plaster-type warming device consisting of a heat- and steam-generating sheet (HSG sheet). First, we tested its effects when applied to the anterior abdominal wall or lumbar region of women complaining of a tendency towards constipation. Application of the sheet to either region produced a feeling of comfort in the abdomen, as assessed by a survey of the subjects. The significant increases in the total hemoglobin observed in these regions suggested an increase in peripheral blood flow, and significant increases in the HF component on ECG and in the amplitude of gastric motility suggested parasympathetic predominance. We concluded that application of the HSG sheet improves the peripheral hemodynamics and autonomic regulation, induces a feeling of comfort in the abdomen, and provides a beneficial environment for the improvement of gastrointestinal movements.

Prostaglandin I2 plays a key role in zymosan-induced mouse pleurisy.

Zymosan, the cell wall of Saccharomyces cerevisiae, induces innate immune responses involving prostanoid production and complement activation. However, the roles of prostanoids in zymosan-induced inflammation and their interaction with the complement system remain to be determined. To clarify these issues, we examined zymosan-induced pleurisy in mice lacking receptors for prostaglandin (PG) E(2) (EP(-/-) mice) or PGI(2) (IP(-/-) mice). Zymosan-induced exudate formation was significantly reduced in IP(-/-) mice compared with wild-type (WT) mice, whereas none of the EP(-/-) mice (EP(1)(-/-), EP(2)(-/-), EP(3)(-/-), and EP(1)(-/-)(4) mice) showed any significant difference from WT mice. Furthermore, indomethacin, an inhibitor of prostanoid biosynthesis, suppressed exudate formation in WT mice to almost the same level as that of IP(-/-) mice. Accordingly, significant production of PGI(2) in the pleural cavity, suggested to be cyclooxygenase-2-dependent, was observed after zymosan injection. Complement activation in the pleural cavity after zymosan injection was confirmed, and preinjection of cobra venom factor (CVF), to deplete blood complement C3, was significantly suppressed after zymosan-induced exudate formation in WT mice. Simultaneous treatment with indomethacin and CVF further suppressed exudate formation in WT mice compared with each treatment alone. Because, some degree of exudate formation was still observed, other factor(s) seem to be involved. However, platelet-activating factor, a promising candidate as one such factor, was not involved in zymosan-induced exudate formation. These results clearly indicate that the PGI(2)-IP system together with the complement system plays a key role in exudate formation in zymosan-induced pleurisy.

Prostaglandin receptors EP2, EP3, and IP mediate exudate formation in carrageenin-induced mouse pleurisy.

The roles of prostaglandins (PGs) as mediators of inflammation have been extensively studied, and production of PGI2 and PGE2 at inflammatory sites has been reported. However, it has not yet been clarified which type of PG receptors has a major role in inflammatory exudation. To examine in vivo role of PG receptors in inflammatory exudation, we induced pleurisy in PG receptors (IP, EP1, EP2, EP3, or EP4) knockout mice by intrapleural injection of carrageenin. Pleural exudate accumulation in wild-type (WT) mice at 1 to 5 h, but not at 24 h, was significantly attenuated by the pretreatment with indomethacin, indicating that PGs are responsible for exudate formation at the early phase of pleurisy. Pleural exudation at 1 to 5 h in IP, EP2, or EP3 knockout mice, but not in EP1 and EP4 knockout, was significantly reduced compared with in WT mice. In the exudates, 6-keto-PGF1alpha and PGE2 were detected as the major PGs, each with its peak concentration at 3 h. In addition, involvement of bradykinin in the phenomenon was suggested by the fact that captopril, a kininase inhibitor, enhanced the exudate formation and increased the amount of 6-keto-PGF1alpha and PGE2 and that a bradykinin B2-receptor antagonist inhibited the exudate formation. In contrast, leukocyte migration into pleural cavity was not influenced by indomethacin-treatment nor by these receptor deficiencies. These results demonstrate participation of EP2 and EP3 along with IP in pleural exudate formation but not in leukocyte migration in carrageenin-induced mouse pleurisy.

Reduced pain hypersensitivity and inflammation in mice lacking microsomal prostaglandin e synthase-1.

We examined the in vivo role of membrane-bound prostaglandin E synthase (mPGES)-1, a terminal enzyme in the PGE2-biosynthetic pathway, using mPGES-1 knockout (KO) mice. Comparison of PGES activity in the membrane fraction of tissues from mPGES-1 KO and wild-type (WT) mice indicated that mPGES-1 accounted for the majority of lipopolysaccharide (LPS)-inducible PGES in WT mice. LPS-stimulated production of PGE2, but not other PGs, was impaired markedly in mPGES-1-null macrophages, although a low level of cyclooxygenase-2-dependent PGE2 production still remained. Pain nociception, as assessed by the acetic acid writhing response, was reduced significantly in KO mice relative to WT mice. This phenotype was particularly evident when these mice were primed with LPS, where the stretching behavior and the peritoneal PGE2 level of KO mice were far less than those of WT mice. Formation of inflammatory granulation tissue and attendant angiogenesis in the dorsum induced by subcutaneous implantation of a cotton thread were reduced significantly in KO mice compared with WT mice. Moreover, collagen antibody-induced arthritis, a model for human rheumatoid arthritis, was milder in KO mice than in WT mice. Collectively, our present results provide unequivocal evidence that mPGES-1 contributes to the formation of PGE2 involved in pain hypersensitivity and inflammation.

Induction of distinct sets of secretory phospholipase A(2) in rodents during inflammation.

Although the expression of the prototypic secretory phospholipase A(2) (sPLA(2)), group IIA (sPLA(2)-IIA), is known to be up-regulated during inflammation, it remains uncertain if other sPLA(2) enzymes display similar or distinct profiles of induction under pathological conditions. In this study, we investigated the expression of several sPLA(2)s in rodent inflammation models. In lipopolysaccharide (LPS)-treated mice, the expression of sPLA(2)-V, and to a lesser extent that of sPLA(2)-IID, -IIE, and -IIF, were increased, whereas that of sPLA(2)-X was rather constant, in distinct tissues. 12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced mouse ear edema, in which the expression of sPLA(2)-IID, -IIF and -V was increased, was significantly reduced by YM-26734, a competitive sPLA(2)-IIA inhibitor that turned out to inhibit sPLA(2)-IID, -IIE, -V and -X as well. In contrast, sPLA(2)-IIA was dominant in carageenin-induced pleurisy in rats, where the accumulation of exudate fluids and leukocytes was significantly ameliorated by YM-26734. These results indicate that distinct sPLA(2)s can participate in inflammatory diseases according to tissues, animal species, and types of inflammation.

Roles for the kallikrein-kinin system in inflammatory exudation and pain: lessons from studies on kininogen-deficient rats.

Roles for the kallikrein-kinin system in inflammation have been investigated extensively, and many reviews on this topic have been published during the 50 years since the discovery of bradykinin in 1949. Recent progress in the field has been remarkable with the help of experiments using gene-targetted transgenic or knockout mice, which have added further valuable information in addition to previous results obtained from pharmacological and biochemical studies using purified and isolated components of the system. Furthermore, much knowledge has been accumulated as a result of the development of various bradykinin agonists and antagonists. In this review, we focused on the data obtained from the kininogen-deficient rat, which is a natural mutant, and discuss the results in comparison with those from bradykinin receptor knockout mice. These data have clarified that endogenous bradykinin exerts a most important role in inflammatory exudation along with prostanoids, preferentially to histamine, serotonin, or neuropeptides. In inflammatory pain perception also, bradykinin produced in the local perivascular spaces stimulates polymodal pain receptors in conjunction with co-helpers such as prostanoids, vanilloids, and neuropeptides. These important roles are concluded based on consistent results obtained from experiments using several antagonists of bradykinin, kininogen-deficient rats, and bradykinin receptor knockout mice.

Regulation of cytosolic prostaglandin E2 synthase by 90-kDa heat shock protein.

Cytosolic prostaglandin (PG) E(2) synthase (cPGES) is constitutively expressed in a wide variety of cells and converts cyclooxygenase (COX)-1-derived PGH(2) to PGE(2). Given the fact that cPGES is identical to p23, a heat shock protein 90 (Hsp90)-binding protein, we herein examined the effect of Hsp90 on PGE(2) generation by cPGES. Incubation of cPGES with Hsp90 resulted in a significant increase in PGES activity in vitro. Association of cPGES with Hsp90 was increased in cells stimulated with A23187 or bradykinin, accompanied by concomitant increases in cPGES activity and PGE(2) production. Moreover, treatment of cells with Hsp90 inhibitors, which destabilized the cPGES/Hsp90 complex, reduced cPGES activity and PGE(2) production to basal levels. These results suggest that the regulation of cPGES activity in cells depends on its association with Hsp90 and provide the first line of evidence that eicosanoid biosynthesis is under the control of the molecular chaperone.

Effects of dexamethasone and protein kinase C inhibitors on the induction of bradykinin B1 mRNA and the bradykinin B1 receptor-mediated contractile response in isolated rat ileum.

We detected the expression of inducible bradykinin (BK) B1 receptor mRNA in the rat ileum by the reverse transcriptase-polymerase chain reaction (RT-PCR) method, when the isolated ileum was suspended for at least 1 hr in an aerated Tyrode's solution at 37 degrees. The induction of this mRNA was both time- and temperature-dependent, and was followed by a contractile response to des-Arg9-BK at around 3 hr of incubation; this response increased in magnitude with time and was maximal at 6 hr. In contrast, the contraction in response to BK and the expression of B2 receptor mRNA were constant throughout this 6-hr incubation period. The contraction due to des-Arg9-BK was selectively suppressed by B1 receptor antagonists, i.e. des-Arg9[Leu8]-BK and des-Arg10-HOE140, but not by the B2 antagonists D-Arg-[Hyp3,Thi5,8,D-Phe7]-BK and HOE140. The inducible des-Arg9-BK contractile response was suppressed by continuous in vitro exposure of the ileum to cycloheximide or actinomycin D, but neither inhibitor affected the contraction induced by BK, suggesting that the B1 receptor could be induced de novo. In vitro and ex vivo treatment of the ileum with dexamethasone suppressed the induction of the contractile response to des-Arg9-BK, but had no significant effect on the expression of B1 receptor mRNA. Some protein kinase C inhibitors, i.e. H7 and calphostin C, suppressed the expression of B1 receptor mRNA and diminished the contractile response to des-Arg9-BK. These results suggest that the de novo synthesis of the B1 receptor in the ileum preparation can be up-regulated at the transcriptional level (a process in which a specific isoform of protein kinase C may be involved). Additionally, these data suggest that the contractile response to des-Arg9-BK involves a process sensitive to some post-transcriptional action of dexamethasone.

Transcriptional regulation of the membrane-associated prostaglandin E2 synthase gene. Essential role of the transcription factor Egr-1.

Membrane-associated prostaglandin (PG) E2 synthase (mPGES) is an inducible terminal enzyme in the biosynthetic pathway for prostaglandin E2, which participates in many biological processes. In this study, we investigated the molecular mechanism controlling the inducible expression of mPGES. The mouse mPGES gene consisted of three exons, and its 5'-proximal promoter contained consensus motifs for the binding of several transcription factors. Transgenic expression in mice of the mouse mPGES promoter flanked by a reporter gene resulted in stimulus-dependent induction of the reporter in tissues where mPGES was intrinsically induced. Deletion and site-specific mutation analyses of the 5'-flanking region demonstrated that stimulus-inducible expression of mouse and human mPGES required tandem GC boxes adjacent to the initiation site. The stimulus-induced GC box binding activity was present in nuclear extracts of cells, in which the proximal GC box was essential for binding. An 80-kDa stimulus-inducible nuclear protein that bound to this GC box was identified as the transcription factor Egr-1 (for early growth response-1). These results suggest that Egr-1 is a key transcription factor in regulating the inducible expression of mPGES.

Critical roles for bradykinin and prostanoids in acute inflammatory reactions: a search using experimental animal models.

Searches for chemical mediators of inflammation underlying classical signs of inflammation i.e. heat, redness, swelling, and pain have been performed and various experimental models for evaluation of new agents to manage these inflammatory signs have been developed extensively during the last century. Now, at the beginning of the 21st-century, after great progress in gene technology, the necessity of in vivo animal study is being reconsidered. Therefore, this review introduces and describes findings obtained by the use of various experimental animal models. We have compared the inflammatory characteristics among species using reported animal models such as dye exudation in the skin, paw edema, pleurisy, and writhing reaction; then we have precisely examined mediators involved in these inflammatory reactions. In the process of plasma exudation and pain perception in the earlier phases of acute inflammation, involvement of the kallikrein-kinin system and prostanoids was demonstrated. Precisely, bradykinin, and PGI(2) among the prostanoids, are major mediators for exudation and pain perception of the initial acute phase of inflammation; both mediators collaborate to enhance these effects. PGE(2), perhaps produced by cyclooxygenase-2, was involved in induction of plasma exudation and pain perception in a later phase than the timing of involvement of PGI(2). Precise roles of various prostanoids will hopefully be clarified by the research projects in progress.