The receptors for endothelins (ETs) are classified into the ETA and ETB types. ETB receptors are highly expressed in astrocytes, but pharmacological usages of this receptor are not clarified. In this article, recent studies on the pathophysiological roles of astrocytic ETB receptors in the brain are reviewed. The administration of ETB agonists and antagonists in nerve injury models showed that several astrocytic functions are regulated by ETB receptors. The activation of ETB receptors causes morphological alterations and proliferation of cultured astrocytes. Astrocytes produce various bio-active substances that can affect damage to nerve tissues. ETs stimulate the production of neurotrophic factors by astrocytes. This action improves impaired brain functions. On the other hand, the production of matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF), which induce brain edema, also are stimulated by ETs. These findings indicate that astrocytic functions are effectively regulated by modulations of ETB receptors. In brain insults and neurodegenerative diseases, these functions of astrocytes affect the protection and repair of damaged nerve tissues. Thus, astrocytic ETB receptors could be a target for novel types of neuroprotective drugs.
β-Arrestins (β-arrestin-1 and β-arrestin-2) were first identified as proteins that have the ability to desensitize G protein-coupled receptors (GPCRs). However, it has recently been found that β-arrestins can activate signaling pathways independent of G protein activation. The diversity of these signaling pathways has also been recognized. This leads to an appreciation of β-arrestin-biased agonists, which is a new class of drugs that selectively activate β-arrestin-mediated signaling without G protein activation. In this review, we will discuss the recent advance of β-arrestin-mediated signaling pathways, including a brief account of different biased agonists, their pharmacological applications, and novel β-arrestin research.
Higher impulsivity is thought to be a risk factor for drug addiction, criminal involvement, and suicide. Excessive levels of impulsivity are often observed in several psychiatric disorders including attention-deficit/hyperactivity disorder and schizophrenia. Previous studies have demonstrated that nicotinic acetylcholine receptors (nAChRs) are involved in impulsive behavior. Here, we introduce recent advances in this field and describe the role of the following nAChR-related brain mechanisms in modulating impulsive behavior: dopamine release in the ventral striatum; α4β2 nAChRs in the infralimbic cortex, which is a ventral part of the medial prefrontal cortex (mPFC); and dopamine release in the mPFC. We also suggest several potential therapeutic drugs to address these mechanisms in impulsivity-related disorders and explore future directions to further elucidate the roles of central nAChRs in impulsive behavior.
SU11274, a small molecule inhibitor of c-Met, was reported to induce apoptosis in human non-small-cell lung cancer (NSCLC) cells. However, SU11274-mediated autophagy in NSCLC cells has rarely been reported. The aim of this study was to elucidate the molecular mechanisms mediating SU11274-induced autophagy in NSCLC A549 cells. Here we reported that SU11274-induced autophagy was accompanied with an increase in the conversion of LC3-I to LC3-II and up-regulation of Beclin-1 expression. Subsequently, we also found that small interfering RNA against c-Met induced A549 cell autophagy while promotion of c-Met by hepatocyte growth factor (HGF) suppressed A549 cell autophagy. Inhibition of autophagy by 3-methyladenine (3-MA) suppressed SU11274-induced cell death, suggesting that SU11274-induced autophagy caused cell death. Further study showed that ERK and p53 were activated after SU11274 treatment. Interruption of ERK and p53 activities decreased SU11274-induced autophagy, and blocking of ERK by the specific inhibitor PD98059 suppressed SU11274-induced p53 activation. Moreover, ERK activation upregulated Beclin-1 expression through induction of Bcl-2 phosphorylation, but p53 did not induce Bcl-2 phosphorylation. In conclusion, inhibition of c-Met induced autophagic cell death, which was associated with ERK–p53 activation and ERK-mediated Bcl-2 phosphorylation in A549 cells.
Multi-glycoside, one of the extracted compounds from Tripterygium wilfordii HOOK. f. (GTW), has been shown to be clinically effective in suppressing glomerular inflammation in chronic kidney disease. However, its clinical application is often limited by its cytotoxic actions on the liver. This study was performed to contrast the dose–effects of GTW on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis (GN). Rats with acute or chronic anti-Thy1.1 GN were either left untreated (the Vehicle group) or treated with a high or low dose of GTW and sacrificed on day 7 or day 45. GTW was administrated 3 days before or at the same time as the antibody injection and lasted until sacrifice. GTW at high dose ameliorated glomerular macrophage accumulation, mesangial proliferation, proteinuria, and interleukin-2 expression in the acute anti-Thy1.1 GN model, but caused structural and functional lesions in the liver. In contrast, GTW at low dose improved activated macrophage and T lymphocyte infiltration, mesangial injury, proteinuria, and interleukin-2 and interferon-γ expressions without hepatic toxicity in the chronic model of GN induced by anti-Thy1.1 antibody. In conclusion, GTW at low dose not only effectively inhibits glomerular inflammation but also avoids severe injuries to the liver.
The nuclear factor-κB (NF-κB) transcription factors control many physiological processes including inflammation, immunity, and apoptosis. In our search for NF-κB inhibitors from natural resources, we identified yangonin from Piper methysticum as an inhibitor of NF-κB activation. In the present study, we demonstrate that yangonin potently inhibits NF-κB activation through suppression of the transcriptional activity of the RelA/p65 subunit of NF-κB. This compound significantly inhibited the induced expression of the NF-κB-reporter gene. However, this compound did not interfere with tumor necrosis factor-α (TNF-α)-induced inhibitor of κBα (IκBα) degradation, p65 nuclear translocation, and DNA-binding activity of NF-κB. Further analysis revealed that yangonin inhibited not only the induced NF-κB activation by overexpression of RelA/p65, but also transactivation activity of RelA/p65. Moreover, yangonin did not inhibit TNF-α-induced activation of p38, but it significantly impaired activation of extracellular signal-regulated kinase 1/2 and stress-activated protein kinase/c-Jun NH2-terminal kinase. We also demonstrated that pretreatment of cells with this compound prevented TNF-α-induced expression of NF-κB target genes, such as interleukin 6, interleukin 8, monocyte chemotactic protein 1, cyclooxygenase-2 and inducible nitric oxide. Taken together, yangonin could be a valuable candidate for the intervention of NF-κB-dependent pathological conditions such as inflammation.
We studied the antiallodynic effect of gabapentin (GBP) in the mouse model of neuropathic pain, aiming at clarifying the underlying mechanism. The L5 spinal nerve ligation induced tactile allodynia, an increase of CD11b expression, and an increase in the protein expression level of the voltage-dependent Ca2+ channel α2/δ-1 subunit in the spinal dorsal horn on the injured side. The chronic intrathecal administration of GBP (100 μg/body per day) as well as ω-conotoxin MVIIA, an N-type Ca2+-channel blocker, completely suppressed allodynia, but did not attenuate the CD11b expression. The antiallodynic effect of GBP lasted for several days after the termination of the drug, while that of ω-conotoxin MVIIA disappeared immediately after the termination. GBP suppressed the elevation of the protein level of the α2/δ-1 subunit in the spinal dorsal horn, although it did not affect its mRNA level in the L5 DRG. These results suggest that GBP inhibits the development of allodynia by suppressing the up-regulation of N-type Ca2+ channels, through normalization of the protein level of the α2/δ-1 subunit at the primary afferent nerve terminal via the inhibition of its anterograde transport. In addition, we propose that the nerve injury enhances the expression level of α2/δ-1 in the downstream of the activation of microglia.
In this study, we describe the antitumor activity of QO-1, one of the new 2-aryl-1,4-naphthoquinone-1-oxime methyl ether derivatives. QO-1 is a derivative of macarpine, a natural occurring product from Rutaceae plant. It could potently inhibit cell growth when tested on 19 cancer cell lines. To investigate its mechanism, two cell lines (HeLa and MCF-7) sensitive to QO-1 were selected. Based on flow cytometry, it was found to induce G2/M-phase arrest. Moreover, it could cause microtubule depolymerization both in vitro and in vivo. On the other hand, QO-1 activated spindle assembly checkpoint (SAC) proteins. Expression of Bub1, one of the SAC, was gradually increased, reaching a peak after 16 – 20 h, and then gradually decreased. Instead, QO-1 increased the sub-G1 population, which suggested a cell death population. Actually, expression of Bcl-2 family proteins and activation of caspase-3/7 were evidences of apoptosis. Consistent with these results, cells with DNA fragmentation and multinucleated cells were increased time-dependently after QO-1 exposure. In conclusion, QO-1 has promising antitumor effects via microtubule depolymerization.
Osteoclasts (OCLs) are multinucleated bone resorbing cells whose differentiation is regulated by receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). It is known that inflammatory cytokines and oxidative stress stimulate differentiation of OCLs. Here we evaluated the effects of kahweol, a coffee-specific diterpene, which has been reported to possess anti-oxidant and anti-inflammatory properties, on the differentiation of bone marrow–derived macrophages (BMMs) or murine monocytic cell line RAW-D cells into OCLs. Kahweol dose-dependently inhibited the formation of tartrate-resistant acid phosphatase staining–positive OCLs from both BMMs and RAW-D cells. In addition, kahweol prevented the bone resorbing activity of OCLs. Kahweol completely abolished RANKL-stimulated phosphorylation of extracellular signal-regulated kinase and impaired phosphorylation of Akt. Moreover, the protein levels of nuclear factor of activated T cells cytoplasmic-1 (NFATc1), a master regulator for OCL differentiation; and OCL markers transcriptionally regulated by NFATc1 such as Src and cathepsin K were down-regulated by kahweol treatment. As one of the molecular mechanisms for the inhibitory effects of kahweol, we also showed that kahweol up-regulated heme oxygenase-1 and inhibited high mobility group box 1 release. Thus, kahweol in coffee is a useful constituent for inhibition of OCL differentiation.
We attempted to establish and validate an in vivo pharmacodynamic (PD) rabbit model to screen tachykinin NK2 receptor (NK2-R) antagonists using pharmacological and pharmacokinetic (PK)/PD analyses. Under urethane anesthesia, changes in intracolonic pressure associated with intravenous (i.v.) administration of a selective NK2-R agonist, βAla8-neurokinin A(4-10) (βA-NKA), was monitored as a PD marker. The analgesic effects of NK2-R antagonists were evaluated by monitoring visceromotor response (VMR) to colorectal distension in a rabbit model of visceral hypersensitivity induced by intracolonic treatment of acetic acid. Intravenous administration of βA-NKA induced transient colonic contractions dose-dependently, which were inhibited by the selective NK2-R antagonists in dose- and/or plasma concentration–dependent manners. The correlation between PD inhibition and plasma concentration normalized with the corresponding in vitro binding affinity was relatively high (r2 = 0.61). Furthermore, the minimum effective doses on the VMR and ID50 values calculated in the PD model were highly correlated (r2 = 0.74). In conclusion, we newly established and validated a rabbit model of agonist-induced colonic contractions as a screening tool for NK2-R antagonists. In a drug discovery process, this PD model could enhance the therapeutic candidate selection for irritable bowel syndrome, pharmacologically connecting in vitro affinity for NK2-R with in vivo therapeutic efficacy.
An endogenous cannabinoid anandamide (N-arachidonoylethanolamide) has been shown to cause vasodilatation in vitro and a brief vasoconstriction followed by prolonged depressor response in vivo. This study investigated the vascular effects of anandamide and underlying mechanisms in rat mesenteric vascular beds. In preparations with an intact endothelium and active tone, anandamide at low concentrations (0.1 – 1 nM) caused a concentration-dependent decrease in perfusion pressure due to vasodilatation, but at high concentrations (10 nM – 1 μM) elicited an initial and sharp increase in perfusion pressure due to vasoconstriction followed by long-lasting vasodilatation in a concentration-dependent manner. Treatment with SR141716A [cannabinoid-1 (CB1)-receptor antagonist] blunted both the vasoconstrictor and vasodilator responses. Also, removal of the endothelium and indomethacin (cyclooxygenase inhibitor), but not adrenergic denervation with 6-hydoxydopamine (adrenergic neurotoxin), markedly inhibited the vasoconstrictor response to anandamide, while these treatments did not affect vasodilatation. The vasodilatation, but not vasoconstriction, in response to anandamide was markedly attenuated by capsazepine [selective antagonist for transient receptor potential vanilloid-1 (TRPV1)], pretreatment with capsaicin [calcitonin gene–related peptide (CGRP)ergic-nerve depletor], or cold-storage denervation. These results suggest that in rat mesenteric vascular beds, anandamide causes CB1-receptor– and prostanoid-mediated endothelium-dependent vasoconstriction and perivascular capsaicin-sensitive CGRPergic nerve–mediated vasodilatation.
The pulmonary vein is known as an important source of ectopic beats, initiating frequent paroxysms of atrial fibrillation. We compared effects of the class Ic antiarrhythmic drug pilsicainide on the electrophysiological parameters in the isolated pulmonary vein preparation from guinea pigs with those in the left atrium. Three pairs of bipolar electrodes were attached to the left atrium, pulmonary vein, and junctional region of the left atrium and pulmonary vein to measure intra-atrial and intra-pulmonary vein conduction velocity and effective refractory period. Pilsicainide (10 μM) decreased the conduction velocity in the pulmonary vein as well as the left atrium, whose effect on the pulmonary vein was relatively greater than that on the left atrium. The drug prolonged the effective refractory period in the pulmonary vein as well as the left atrium, and the effect of the drug on the pulmonary vein was less than that on the left atrium. The currently observed electrophysiological property of pilsicainide suggests that its effects on reentry within the pulmonary vein are estimated to be weaker than within the left atrium, which may be one of the key considerations for understanding its antiarrhythmic mechanisms in the atrium and pulmonary vein.
Cystic fibrosis (CF) is the most common lethal inherited disorder and is caused by mutations in the gene encoding the CF transmembrane regulator (CFTR). The CF lung expresses a profound proinflammatory phenotype that appears to be related to a constitutive hypersecretion of interleukin (IL)-8 from airway epithelial cells in response to microbial infection. Since overproduction of IL-8 in CF contributes to massive bronchial infiltrates of neutrophils, identification of the pathways underlying IL-8 induction could provide novel drug targets for treatment of neutrophil-dominated inflammatory diseases such as CF. Here, we show that IL-17A synergistically increases IL-8 production induced by a toll-like receptor (TLR) 2 agonist, peptidoglycan (PGN), or TLR4 agonist, lipopolysaccharide (LPS), in a human CF bronchial epithelial cell line (CFBE41o-). A strong synergism was also observed in primary human CF bronchial epithelial cells, but not in human non-CF cell lines and primary cells. Notably, despite the induction of nuclear factor-κB and MAP kinases during TLR2 or TLR4 activation in CFBE41o-, IL-17A-dependent synergism appears to be the result of enhanced PGN- or LPS-induced phosphorylation of p38. Taken together, these studies provide evidence that IL-17A is a critical factor in increasing IL-8 expression in bacteria-infected CF airways via a pathway that regulates p38 phosphorylation.
Vasorelaxant properties of N-2-(ferulamidoethyl)-nitrate (ferulate nitrate, FLNT), a newly synthesized nitrate, were compared with those of isosorbide dinitrate, nicorandil, nitroglycerin, and 8-bromoguanosine 3,5-cyclic monophosphate (8-Br-cGMP) in rat aorta pre-contracted by phenylephrine. FLNT produced vasorelaxation in a concentration-dependent manner (0.1 – 100 μM). The degree of relaxation induced by FLNT was similar to that induced by isosorbide dinitrate. In addition, removal of endothelium did not affect the relaxant effect of FLNT. FLNT caused a rightward shift of the cumulative concentration–response curves of phenylephrine and reduced the maximal efficacy of contraction. 1H-[1,2,4]Oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 μM) and K+-channel blockers charybdotoxin (CHT, 0.1 μM) and BaCl2 (1 μM) reduced the relaxant effect of FLNT in the endothelium-denuded arteries, whereas glibenclamide (1 μM) and 4-aminopyridine (1 mM) failed to influence FLNT-induced vasorelaxation. Furthermore, in the presence of ODQ, both CHT (0.1 μM) and BaCl2 (1 μM) still significantly reduced the relaxation evoked by FLNT. Pretreatment of vessels with hydroxocobalamin, a nitric oxide scavenger, abolished the FLNT effect. These findings demonstrate that FLNT induces relaxation of the rat aorta rings endothelium-independently. Furthermore, we demonstrated that FLNT-induced vasorelaxation is related to its stimulation of soluble guanylate cyclase and activation of K+ channels.
Sauchinone is a lignan isolated from Saururus chinensis known to suppress nitric oxide (NO) activity. Previous studies demonstrate that NO plays a key role in methamphetamine-induced neurotoxicity. Thus, we hypothesized that sauchinone could have a suppressive effect on the neurotoxicity induced by methamphetamine. Repeated injections of methamphetamine cause degeneration of dopaminergic nerve terminals, whereas sauchinone treatment significantly prevented this degeneration. Sauchinone treatment also inhibited the methamphetamine-induced activation of glia cells and the production of NO via a blockade of inducible NO synthase protein expression. Our results suggest that sauchinone can prevent methamphetamine-induced neurotoxicity through the suppression of NO production.
The present study investigated whether histamine was taken up by perivascular adrenergic nerves and released by periarterial nerve stimulation (PNS) to induce vascular responses. In rat mesenteric vascular beds treated with capsaicin to eliminate calcitonin gene–related peptide (CGRP)ergic vasodilation and with active tone, PNS (1 – 4 Hz) induced only adrenergic nerve–mediated vasoconstriction. Histamine treatment for 20 min induced PNS-induced vasoconstriction followed by vasodilation without affecting CGRP-induced vasodilation. Chlorpheniramine, guanethidine, combination of histamine and desipramine, and endothelium-removal abolished PNS-induced vasodilation in histamine-treated preparations. These results suggest that histamine taken up by and released from adrenergic nerves by PNS causes endothelium-dependent vasodilation in rat mesenteric arteries.
We have recently shown that prenatal valproic acid (VPA) exposure causes autism spectrum disorders–like behavioral abnormalities and Nissl-positive cell loss in both prefrontal and somatosensory cortices in male mice. We have also found that VPA-induced social interaction deficits are observed in male but not female offspring. This study demonstrated that the exposure to VPA at embryonic day 12.5 significantly decreased Nissl-positive cell numbers in the prefrontal cortex, but not in the somatosensory cortex, in female offspring. These findings suggest that VPA-induced morphological abnormalities in the somatosensory cortex may be involved in the sex-dependent social interaction deficits.
Oxaliplatin is a key drug in the treatment of colorectal cancer, but it causes acute and chronic neuropathies in patients. Amitriptyline has widely been used in patients with painful neuropathy. In this study, we investigated the effect of amitriptyline on the oxaliplatin-induced neuropathy in rats. Repeated administration of amitriptyline (5 and 10 mg/kg, p.o., once a day) reduced the oxaliplatin-induced mechanical allodynia but not cold hyperalgesia and reversed the oxaliplatin-induced increase in the expression of NR2B protein and mRNA in rat spinal cord. These results suggest that amitriptyline is useful for the treatment of oxaliplatin-induced neuropathy clinically.