Vascular endothelium plays an important role in maintaining vascular homeostasis by synthesizing and releasing several vasodilating factors, such as prostacyclin, nitric oxide (NO), and a yet unidentified endothelium-derived hyperpolarizing factor (EDHF). Possible candidates for EDHF include epoxyeicosatrienoic acids, endothelium-derived K⁺ ions, and as we have recently identified, hydrogen peroxide (H₂O₂). Electrical communication between endothelial and smooth muscle cells through gap junctions has also been suggested to be involved in endothelium-dependent hyperpolarization. Among the above candidates, the H₂O₂ hypothesis well explains the pathophysiological interactions between NO and EDHF and re-highlights the physiological roles of the reactive oxygen species in endothelium-dependent vascular responses. This brief review summarizes our current knowledge about H₂O₂ as an EDHF, with special reference to its production by the endothelium, its action on membrane potentials and its pathophysiological roles.

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Histamine, prostaglandin E₂, and catecholamines have been demonstrated to regulate the innate and acquired immune responses. In this review, we describe one of the mechanisms common to the action of these agonists; the regulation of the expression of costimulatory adhesion molecules such as ICAM-1 and B7 antigens on monocytes/macrophages. The specific receptor subtypes involved in the action of each agonist were H₂ for histamine, EP₂/EP₄ for prostaglandin E₂, and β₂ for catecholamines, all of which are coupled with adenylate cyclase via Gs protein. The regulation of the expression of adhesion molecules by these agonists in turn leads to the modulation of subsequent cytokine production mediated by cell-cell interaction under different stimuli. Histamine is synthesized in monocytes and T cells by the induction of histidine decarboxylase. The inducible histamine has different dynamics from that in storage granules of mast cells and basophils. Also, noradrenaline appears to be synthesized in lymphocytes. Thus, immune cells can produce histamine, prostaglandins, and noradrenaline by themselves and modulate the cell-cell interaction between monocytes and other cells. Some of the inhibitors of HMG-CoA reductase were shown to bind to the ICAM-1-binding domain of LFA-1, reducing the interaction mediated by ICAM-1/LFA-1. The regulation of interaction mediated by adhesion molecules may provide a new target for controlling inflammatory and immune responses.

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In the present study, we investigated whether a novel benzopyranylindol analogue, KR-31466 (KR466) (1-[(2S,3R,4S)-3,4-dihydro-2-dimethoxymethyl-3-hydroxy-2-methyl-6-nitro-2H-1-benzopyran-4-yl]-1H-indole-2-carboxylic acid ethyl ester) can attenuate hypoxic injury in heart-derived H9c2 cells and, if so, whether the protective effect of KR466 is mediated through mitochondrial ATP-sensitive potassium (mtK_ATP) opening. The treatment of H9c2 cells with KR466 (3 – 30 μM) significantly reduced hypoxia-induced cell death in a concentration-dependent manner, as shown by lactate dehydrogenase release and propidium iodide-uptake. In addition, KR466 (10 μM) significantly reduced the increase in hypoxia-induced TUNEL-positive cells, suggesting its anti-apoptotic potential in H9c2 cells. The protective effects of KR466 were abolished by 5-hydroxydecanoate, a specific blocker of the mtK_ATP channel, suggesting the involvement of the mtK_ATP channel in the protective effect of KR466. A specific inhibitor of protein kinase C (PKC), chelerythrine (3 μM), significantly attenuated the protective effect of KR466 against hypoxia-induced cardiac cell death. In conclusion, our results suggest that KR466 can protect H9c2 cells from hypoxia-induced death through mtK_ATP channel opening and PKC activation.

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We investigated the relationship between prostaglandin (PG) F_2α-induced functional luteal regression and apoptosis in the rodent ovary. Administration of PGF_2α significantly decreased the serum levels of progesterone within 12 h after treatment in pseudopregnant mice. Apparent signals were detected in luteal tissues at 24 to 72 h by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick endo labeling (TUNEL) assay. PGF_2α significantly increased the levels of the cleavage nuclear poly (ADP-ribose) polymerase fragment and transforming growth factor-β (TGF-β) mRNA within 48 h. PGF_2α (10 μM) decreased progesterone secretion 6 to 72 h after its addition in luteinized ovarian cells, whereas C2-ceramide (25 μM) decreased progesterone levels by 44% 72 h after its addition. DNA fragmentation was not observed in the cultured cells treated with PGF_2α for 24 h, although cells incubated with C2-ceramide showed fragmentation. Treatment with PGF_2α for 1 h caused a distinct decrease in luteinizing hormone (LH)-induced cyclic AMP production. Thus, the inhibitory effect of PGF_2α on progesterone secretion may be caused by both the blockage of the functional LH receptor and the activation of apoptotic signaling.

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Senescence-accelerated mouse prone 8 (SAMP8) and prone 10 (SAMP10) are useful murine model of accelerated aging. SAMP8 shows marked impairment of learning and memory, whereas SAMP10 shows brain atrophy and aging-associated depressive behavior. This study examined the expression of glial cell line-derived neurotrophic factor (GDNF) in SAMP8 and SAMP10 brains, relative to that in SAM resistant 1 (SAMR1) controls, which age normally. Hippocampal GDNF mRNA expression decreased in an age-dependent manner (10- vs 2-month-old animals) in the SAMR1, but not in the SAMP8 or SAMP10 strains. Furthermore, GDNF mRNA expression in 2-month-old SAMP8 and SAMP10 strains was less than in SAMR1 specimens of the same age. The number of surviving neurons in the CA1 region decreased with age in SAMP8 and SAMP10, and also decreased relative to the number of neurons in 10-month-old SAMR1 controls. Immunohistochemistry revealed that cells that were positive for GDNF-like activity in 10-month-old SAMP8 and SAMP10 were diffusely distributed, in part, around the pyramidal cell layer in the hippocampus. These findings suggest that low GDNF expression in young SAMP8 and SAMP10 may be involved in hippocampal dysfunctions, such as age-related learning impairment and neuronal death.

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In the present study, we examined the expression and the localization of apamin-sensitive small conductance Ca²⁺-activated K⁺ channels (SK channels) in the mouse intestine. SK3-immunoreactivity (IR) was detected in both ileum and colon. Double staining experiments showed that SK3-IR was colocalized with prolyl 4-hydroxylase (PH(α))-IR, but not with c-Kit-IR which are markers of fibroblast cells and the interstitial cells of Cajal (ICC), respectively. Although SK3-IR was colocalized with vimentin-IR, which is another marker of ICC, the reactivity of SK3-immunopositive cells was weaker than that of ICC. The SK3-immunopositive cells were similarly present in the intestine of c-Kit mutant mice (W/W^V), in which ICC were absent, and its wild-type mice. The immuno-electron microscopic analysis indicated that SK3 was localized in the cells that had some similar morphological features to ICC, but obviously different from ICC. The SK3-immunopositive cells had gap junctions with the smooth muscle cells. The gap junctions were smaller than those between ICC and smooth muscle cells. These results indicate expression of SK3 in fibroblast-like cells, but not in ICC, and suggest participation of the cells in the intestinal motility.

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The effects of cimetidine on rat adjuvant arthritis (AA) and rat osteoclast differentiation were studied. For the in vivo experiments, AA was induced by injections of Mycobacterium tuberculosis H37RA either subcutaneously into the base of the tail or into the right hind paw. The osteoclast differentiation was assessed by estimating the number of tartrate-resistant acid phosphatase-positive multinuclear cells in the bone marrow culture. Cimetidine, at the dose of 25 mg/kg body weight, reduced the paw swelling by 70% (P<0.01). Cimetidine, at 10 μM concentration, inhibited 1,25-dihydroxyvitamin D₃ (1,25[OH]₂D₃) and histamine mediated osteoclast differentiations by 40% (P<0.01) and 60% (P<0.001), respectively. Dimaprit, at 0.3 μM, stimulated the cell differentiation by 100% (P<0.01). Mepyramine reduced osteoclast differentiation, but the reduction was not statistically significant. Measurements of bone mineral density of the femur indicated that 5 mg/kg of cimetidine treated animals had 30% (P<0.01) higher mineral density in comparison with that of the AA control group that received no cimetidine. These results suggest that histamine is a potent inducer of osteoclast differentiation, at least in part, through the histamine H₂-receptor, and cimetidine has a preventive effect on articular destruction and accompanying inflammation in arthritic rats. These observations may provide critical insights into the pathogenesis of the bone pathology seen in patients with RA.

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The effect of Matricaria chamomilla (M. chamomilla) on the development of morphine dependence and expression of abstinence was investigated in rats. The frequencies of withdrawal behavioral signs (paw tremor, rearing, teeth chattering, body shakes, ptosis, diarrhea, and urination) and weight loss induced by naloxone challenge were demonstrated in morphine-dependent rats receiving M. chamomilla extract or saline. The withdrawal behavioral manifestations and weight loss were inhibited significantly by chronic co-administration of M. chamomilla extract with morphine. Administration of a single dose of M. chamomilla before the naloxone challenge in morphine-dependent animals abolished the withdrawal behavioral manifestations. The dramatic increase of plasma cAMP induced by naloxone-precipitated abstinence was prevented by chronic co-administration of M. chamomilla extract with morphine. These results suggest that M. chamomilla extract inhibits the development of morphine dependence and expression of abstinence syndrome.

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In the isolated rat aorta, a ROCKs (rhoA-dependent coiled coil serine/threonine kinases) inhibitor, Y-27632, inhibited the contractions induced not only by receptor agonists but also by high K⁺ with the similar IC₅₀ values (0.8 – 4.9 μM). However, Y-27632 did not inhibit the increment of cytosolic Ca²⁺ concentration ([Ca²⁺]_i) due to these stimulants. The Y-27632-induced inhibition of contraction was accompanied by an inhibition of myocin light chain (MLC) phosphorylation, although inhibition of contraction was stronger than that of MLC phosphorylation during the initial phase of contraction. Y-27632 had no effect on the myocin light chain kinase (MLCK) activity. This inhibitor also did not directly change the phosphatase activity. These results suggest that Y-27632 is a selective inhibitor of ROCKs with no direct inhibitory effect on [Ca²⁺]_i, calmodulin, MLCK, or phosphatase. Y-27632 disrupted the actin filament network and decreased the filamentous actin, implying that the stronger inhibition by Y-27632 on early phase of contraction than MLC phosphorylation may be explained by this effect. These results suggest that the high K⁺-induced MLC phosphorylation and contraction are mediated not only by the classical Ca²⁺/calmodulin-dependent MLCK system but also by a novel MLC phosphorylation pathway involving ROCKs. One of the possibilities is that high K⁺ activates ROCKs to inhibit myosin phosphatase resulting in an augmentation of MLC phosphorylation and contraction.

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Uncaria rhynchophylla is a medicinal herb used for convulsive disorders in Oriental medicine. In this study, the effect of the methanol extract of Uncaria rhynchophylla against N-methyl-D-aspartate (NMDA)-induced excitotoxicity was investigated. Pretreatment with the extract of Uncaria rhynchopylla reduced the degree of neuronal damage induced by NMDA exposure in cultured hippocampal slices. In the patch clamp study, Uncaria rhynchophylla significantly inhibited NMDA receptor-activated ion current in acutely dissociated hippocampal CA1 neurons. These results indicate that Uncaria rhynchophylla offers protection against NMDA-induced neuronal injury and inhibitory action on NMDA receptor-mediated ion current may be a mechanism behind the neuroprotective effect of Uncaria rhynchophylla.

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In the present work, we found that the ethyl acetate-soluble fraction of Cnidium officinale MAKINO (COEA) decreased nitric oxide (NO) production in the lipopolysaccharide (LPS)-stimulated BV-2 and primary microglia and suppressed expression of inducible nitric oxide synthase (iNOS) in BV-2 cells with the same pattern of NO production. In addition, we showed that excessive NO production played an important role in neuronal cell death in LPS-treated rat hippocampal slice cultures. Our data suggest that the COEA inhibits neuronal cell death by reduction of excessive NO production in LPS-treated rat hippocampal slice cultures. The ethyl acetate-soluble fraction of C. officinale reduced propidium iodide uptake and NO production in cultured media at the same time.

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