Butein suppresses ICAM-1 expression through the inhibition of IκBα and c-Jun phosphorylation in TNF-α- and PMA-treated HUVECs.

Butein (3,4,2',4'-tetrahydroxychalcone), a flavonoid derivative, has been reported to show several biological actions, including anti-inflammatory and anti-cancer. However, the possible molecular mechanisms involved are poorly understood. Treatment of human umbilical vein endothelial cells (HUVECs) with butein significantly inhibited cell surface intercellular adhesion molecule-1 (ICAM-1) expression, ICAM-1 protein synthesis, and mRNA expression induced by tumor necrotic factor-α (TNF-α) and/or phorbol 12-myristate 13-acetate (PMA). Electrophoretic mobility shift assay revealed that butein blocked activation of transcription factors, nuclear factor-κB (NF-κB) and activator protein-1 (AP-1), induced by TNF-α and PMA. Moreover, butein abolished TNF-α- and PMA-induced IκBα phosphorylation, which participates in NF-κB activation, and PMA-induced phosphorylation of c-Jun, a subunit composed of AP-1. In vitro, butein inhibited the phosphorylation of c-Jun, binding to GST beads, mediated by JNK isolated from PMA-treated cells. The inhibitory action of butein on the JNK-mediated in vitro c-Jun phosphorylation was abrogated in the presence of ATP. These results indicate that in HUVECs, butein suppresses the expression of ICAM-1 mRNA and protein through the inhibition of the activation of NF-κB and AP-1 induced by TNF-α and PMA, that the inhibitory action of butein on NF-κB activation results from the inhibition of IκBα phosphorylation by IκB kinase (IKK), and that the inactivation of PMA-activated AP-1 by butein is due to the blocking of JNK-mediated c-Jun phosphorylation through the inhibition of ATP binding.

Acotiamide hydrochloride (Z-338) enhances gastric motility and emptying by inhibiting acetylcholinesterase activity in rats.

In clinical trials, acotiamide hydrochloride (acotiamide: Z-338) has been reported to be useful in the treatment of functional dyspepsia. Here, we investigated the effects of acotiamide on gastric contraction and emptying activities in rats in comparison with itopride hydrochloride (itopride) and mosapride citrate (mosapride). We also examined in vitro the compound's inhibitory effect on acetylcholinesterase (AChE) activity derived from rat stomach. In in vivo studies, acotiamide (30 and 100mg/kg s.c.) and itopride (100mg/kg s.c.) markedly enhanced normal gastric antral motility in rats. In gastric motility dysfunction models, acotiamide (100mg/kg s.c.) and itopride (100mg/kg s.c.) improved both gastric antral hypomotility and the delayed gastric emptying induced by clonidine, an α(2)-adrenoceptor agonist. In contrast, mosapride (10mg/kg s.c.) had no effect on these models. Like the AChE inhibitors itopride (30 mg/kg s.c.) and neostigmine (10 µg/kg s.c.), acotiamide (10mg/kg s.c.) also clearly enhanced gastric body contractions induced by electrical stimulation of the vagus, which were abolished by atropine and hexamethonium, whereas mosapride (3 and 10mg/kg s.c.) did not. In in vitro studies, acotiamide concentration-dependently inhibited rat stomach-derived AChE activity (IC(50)=2.3 µmol/l). In addition, stomach tissue concentrations of acotiamide after administration at 10mg/kg s.c. were sufficient to produce inhibition of AChE activity in rat stomach. These results suggest that acotiamide stimulates gastric motility and improves gastric motility dysfunction in rats by inhibiting AChE activity, and may suggest a role for acotiamide in improving gastric motility dysfunction in patients with functional dyspepsia.

Gastric relaxation induced by electrical and chemical stimulation of the area postrema in the rat.

Area postrema (AP) is considered to be an important neural center for emesis in carnivores. However, it is also known that AP mediates motor responses induced by apomorphine in rats which do not have an emetic reflex. To shed more light on the possible role of AP in the control of gastric motility in physiological or pathophysiological conditions, we observed the effects of electrical or chemical (apomorphine) stimulation of AP neurons on intragastric pressure (IGP) or intragastric volume (IGV) in rat. We found that electrical stimulation (ES) reduces IGP, and this is sensitive to hexamethonium or L-NAME, and apomorphine also reduces IGP and increases IGV. In slice preparations, apomorphine (10 micromol/l) increased the frequency of spontaneous single unit discharges of AP neurons recorded extracellularly. We also succeeded retrograde labeling of AP neurons by DiI applied into the gastric corpus, for the first time. These observations indicate that rat stomach receives efferent neural input from AP and the excitation of AP neurons relaxes the stomach in rat, suggesting some functional roles of AP neurons in the regulation of gastric motility.

Immunolocalization of multispecific organic anion transporters, OAT1, OAT2, and OAT3, in rat kidney.

Recently, a family of multispecific organic anion transporters has been identified, and several isoforms have been reported. However, the physiologic and pharmacologic roles of each isoform, except OAT1, in the transepithelial transport of organic anions in the kidney remain to be elucidated. To address this issue, it is essential to determine the intrarenal distribution and membrane localization of each OAT isoform along the nephron. In this study, the intrarenal distributions of rOAT1, rOAT2, and rOAT3 were investigated by an immunofluorescence method that used frozen rat serial kidney sections. Confocal microscopic analysis showed that immunoreactivity for rOAT1 was detected exclusively in the proximal tubules (S1, S2, S3) in the cortex with basolateral membrane staining. rOAT2 was detected in the apical surface of the tubules in the medullary thick ascending limb of Henle's loop (MTAL) and cortical and medullary collecting ducts (CD). rOAT3 was localized in the basolateral digitation of the cell membrane in all the segments (S1, S2, and S3) of the proximal tubules, MTAL, cortical TAL, connecting tubules, and cortical and medullary CD. These results on the distribution of each OAT isoform will facilitate the understanding of the role of OATs in the renal processing of organic anions.