European Journal of Pharmacology (v.728, #C)

Neuroinflammation is a key mechanism against infection, injury, and trauma in the central nervous system (CNS). The heartwood of Dalbergia odorifera T. Chen is an important source of traditional Korean and Chinese medicines. (2R, 3R)-Obtusafuran (1) and isoparvifuran (2) are arylbenzofuran compounds isolated from D. odorifera. This study determined the efficacy of (1) and (2) in modulating the regulation of anti-inflammatory activity through the upregulation of heme oxygenase (HO)-1 in BV2 microglia. Compound (1) inhibited the protein expression of inducible nitric oxide synthase (iNOS), iNOS-derived nitric oxide (NO), cyclooxygenase (COX)-2, and COX-2-derived prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-stimulated mouse BV2 microglia. (2R, 3R)-Obtusafuran (1) also reduced tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) production, and these anti-neuroinflammatory effects were shown to be correlated with the suppression of the phosphorylation and degradation of inhibitor of nuclear factor kappa B-α (IκB-α), and nuclear factor kappa B nuclear (NF-κB) translocation and DNA binding activity. In addition, (1) upregulated HO-1 expression via nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in mouse BV2 microglia. Using tin protoporphyrin (SnPP), an HO activity inhibitor, we verified that the inhibitory effects of (1) on the proinflammatory mediators and proteins were associated with the induction of HO-1 expression. Activated microglia-mediated cell death of mouse hippocampal HT22 cells was significantly repressed by (1). Our data suggest that (2R, 3R)-obtusafuran (1) has therapeutic potential against neurodegenerative diseases caused by neuroinflammation.
Keywords: Dalbergia odorifera; (2R, 3R)-Obtusafuran; BV2 microglia; Neuroinflammation; Neuroprotection; Heme oxygenase-1;

Curcumin regulates delta-like homolog 1 expression in activated hepatic stellate cell by Jianwei Qiu; Qian Zhou; Xuguang Zhai; Xin Jia; Yajun Zhou (9-15).
Hepatic stellate cell activation is a key cellular event in the development of liver fibrosis. Recently, Delta-like homolog 1 (DLK1) protein level has been shown to increase in HSC activation and serve as a new contributor to HSC activation and liver fibrosis. Curcumin, a natural yellow polyphenol, possesses therapeutic roles in many diseases including liver fibrosis and has long been used in traditional medicine. The present study was aimed to elucidate the effect of curcumin on DLK1 expression in HSCs in vitro and in vivo, which is still unknown. Our results demonstrated that curcumin reduced DLK1 expression in culture-activated HSCs and in rat model of liver fibrosis. The inhibitory effect of curcumin on DLK1 expression may be mediated in part by interruption of Shh signaling pathway, which contributes to the promotion effect of curcumin on the expression of PPAR-gamma, a key factor in inhibiting HSC activation. Our results in this study may reveal a new mechanisms through which curcumin exerts its inhibitory effect on HSC activation and liver fibrosis.Display Omitted
Keywords: Curcumin; Delta-like homolog 1; Hepatic stellate cell; Liver fibrosis; Sonic Hedgehog;

Cyclosporin A (CsA), one of the most fundamental immunosuppressive drugs, is routinely used in clinics for the treatment of liver and other organ rejections. However, one of the major challenges of the application of CsA is the occurrence of the serious adverse effects, namely, acute and chronic nephrotoxicity, severe hypertension and neurotoxicity. Although N-(3׳,4׳-dimethoxycinnamoyl) anthranilic acid (3,4-DAA) plays an important role in apoptosis of activated T cells, and is therapeutically used as an orally active anti-allergic drug for the treatment of allergy, it has not been tested for use in the treatment of organ rejection. In this study, we used the dark agouti (DA)-Lewis rat orthotopic liver transplantation (OLT) model to investigate whether the combination of 3,4-DAA with CsA is a promising and useful strategy to lower CsA dosage for reducing CsA side effect and preserve therapeutic effect of CsA. Here, we document that the combination treatment effectively inhibits acute liver rejection in OLT model with only half the normally suggested dosage of CsA that has much less side effect in rats than that of the full dosage. These results indicate that 3,4-DAA may serve as an effective adjunct for a CsA-based immunosuppressive regimen to treat transplant recipients for reducing CsA side effect.
Keywords: 3; 4-DAA; CsA nephrotoxicity; Liver transplantation; Rat;

Effects of (−)-epicatechin on a diet-induced rat model of cardiometabolic risk factors by Gabriel Gutiérrez-Salmeán; Pilar Ortiz-Vilchis; Claudia Maria Vacaseydel; Leticia Garduño-Siciliano; German Chamorro-Cevallos; Eduardo Meaney; Santiago Villafaña; Francisco Villarreal; Guillermo Ceballos; Israel Ramírez-Sánchez (24-30).
Overweight and obesity have been associated with increase in cardiometabolic risk. Therapeutics include lifestyle changes and/or pharmacologic agents. However, such interventions are often limited by poor compliance and/or significant side effects. The consumption of certain dietary products, such as cocoa, exerts positive effects on cardiometabolic risk factors. (−)-Epicatechin (EPI), the most abundant flavonoid in cacao has been reported to replicate such effects. However its mechanisms of action have not been fully elucidated.In a rat model of high-fat diet-induced obesity and its associated crdiometabolic risk factors, we administered 1 mg/kg of EPI, by gavage, for 2 weeks. Endpoints included weight-gain, glycemia, triglyceridemia, and systolic blood pressure. We also assessed food intake and fecal excretion. Mitochondrial function and structure related proteins were measured by Westerns.Obesity, hyperglycemia, hypertriglyceridemia, and systolic hypertension were developed after the administration of the high-fat diet for five weeks. EPI significantly decreased the rate of weight gain, glycemia and hypertriglyceridemia. The ratio between energy intake and excretion was not significantly modified by treatment. EPI restored the obesity-induced decreases in the levels of skeletal muscle and abdominal tissue sirtuins (SIRTs), peroxisome proliferator-activated receptor coactivator (PGC-1α), mitofilin, transcription factor A mitochondrial (TFAM), uncoupling protein 1 (UCP1), and deiodinase.EPI treatment yielded beneficial effects on high fat diet-induced endpoints thus may be considered as a potential agent for the treatment of obesity and its cardiometabolic associated abnormalities. Mechanism of action may be attributed to the modulation of cellular/mitochondrial function, thus improving overall metabolism.Display Omitted
Keywords: Cacao; Epicatechin; Flavonoids; Metabolic cardiovascular syndrome; Mitochondria;

Effects of the selective adenosine A2A receptor antagonist, SCH 412348, on the parkinsonian phenotype of MitoPark mice by Karen M. Smith; Susan E. Browne; Srinivasan Jayaraman; Carina J. Bleickardt; Lisa M. Hodge; Edward Lis; Leon Yao; Sunday L. Rittle; Nathalie Innocent; Deborra E. Mullins; George Boykow; Ian J. Reynolds; David Hill; Eric M. Parker; Robert A. Hodgson (31-38).
Adenosine A2A receptors are predominantly localized on striatopallidal gamma-aminobutyric acid (GABA) neurons, where they are colocalized with dopamine D2 receptors and are involved in the regulation of movement. Adenosine A2A receptor antagonists have been evaluated as a novel treatment for Parkinson׳s disease and have demonstrated efficacy in a broad spectrum of pharmacological and toxicological rodent and primate models. Fewer studies have been performed to evaluate the efficacy of adenosine A2A receptor antagonists in genetic models of hypodopaminergic states. SCH 412348 is a potent and selective adenosine A2A receptor antagonist that shows efficacy in rodent and primate models of movement disorders. Here we evaluated the effects of SCH 412348 in the MitoPark mouse, a genetic model that displays a progressive loss of dopamine neurons. The dopamine cell loss is associated with a profound akinetic phenotype that is sensitive to levodopa (l-dopa). SCH 412348 (0.3–10 mg/kg administered orally) dose dependently increased locomotor activity in the mice. Moreover, SCH 412348 retained its efficacy in the mice as motor impairment progressed (12–22 weeks of age), demonstrating that the compound was efficacious in mild to severe Parkinson׳s disease–like impairment in the mice. Additionally, SCH 412348 fully restored lost functionality in a measure of hind limb bradykinesia and partially restored functionality in a rotarod test. These findings provide further evidence of the anti-Parkinsonian effects of selective adenosine A2A receptor antagonists and predict that they will retain their efficacy in both mild and severe forms of motor impairment.
Keywords: A2A receptor antagonist; Movement disorder; MitoPark mouse; Locomotor activity; Parkinson׳s disease;

Perinatal growth restriction decreases diuretic action of furosemide in adult rats by Barent N. DuBois; Jacob Pearson; Tahir Mahmood; Duc Nguyen; Kent Thornburg; Ganesh Cherala (39-47).
Perinatal growth restriction programs higher risk for chronic disease during adulthood via morphological and physiological changes in organ systems. Perinatal growth restriction is highly correlated with a decreased nephron number, altered renal function and subsequent hypertension. We hypothesize that such renal maladaptations result in altered pharmacologic patterns for life. Maternal protein restriction during gestation and lactation was used to induce perinatal growth restriction in the current study. The diuretic response of furosemide (2 mg/kg single i.p. dose) in perinatally growth restricted rats during adulthood was investigated. Diuresis, natriuresis and renal excretion of furosemide were significantly reduced relative to controls, indicative of decreased efficacy. While a modest 12% decrease in diuresis was observed in males, females experienced 26% reduction. It is important to note that the baseline urine output and natriuresis were similar between treatment groups. The in vitro renal and hepatic metabolism of furosemide, the in vivo urinary excretion of the metabolite, and the expression of renal drug transporters were unaltered. Creatinine clearance was significantly reduced by 15% and 19% in perinatally growth restricted male and female rats, respectively. Further evidence of renal insufficiency was suggested by decreased uric acid clearance. Renal protein expression of sodium–potassium–chloride cotransporter, a pharmacodynamic target, was unaltered. In summary, perinatal growth restriction could permanently imprint pharmacokinetic processes affecting drug response.
Keywords: Fetal programming; Furosemide; In utero growth restriction; Perinatal growth restriction; Pharmacokinetics; Renal insufficiency;

Direct action and modulating effect of (+)- and (−)-nicotine on ion channels expressed in trigeminal sensory neurons by Benjamin S.P. Schreiner; Ramona Lehmann; Ulrike Thiel; Paul M. Ziemba; Leopoldo R. Beltrán; Muhammad A. Sherkheli; Philippe Jeanbourquin; Alain Hugi; Markus Werner; Günter Gisselmann; Hanns Hatt (48-58).
Nicotine sensory perception is generally thought to be mediated by nicotinic acetylcholine (nACh) receptors. However, recent data strongly support the idea that other receptors (e.g., transient receptor potential A1 channel, TRPA1) and other pathways contribute to the detection mechanisms underlying the olfactory and trigeminal cell response to nicotine flavor. This is in accordance with the reported ability of humans to discriminate between (+)- and (−)- nicotine enantiomers. To get a more detailed understanding of the molecular and cellular basis underlying the sensory perception of nicotine, we studied the activity of (+)- and (−)-nicotine on cultured murine trigeminal sensory neurons and on a range of heterologously expressed receptors.The human TRPA1 channel is activated by (−)-nicotine. In this work, we show that (+)-nicotine is also an activator of this channel. Pharmacological experiments using nicotinic acetylcholine receptors and transient receptor potential blockers revealed that trigeminal neurons express one or more unidentified receptors that are sensitive to (+)- and/or (−)-nicotine. Results also indicate that the presence of extracellular calcium ions is required to elicit trigeminal neuron responses to (+)- and (−)-nicotine. Results also show that both (+)-nicotine and (−)-nicotine can block 5-hydroxytryptamine type 3 (5-HT3) receptor-mediated responses in recombinant expression systems and in cultured trigeminal neurons expressing 5-HT3 receptors. Our investigations broaden the spectra of receptors that are targets for nicotine enantiomers and give new insights into the physiological role of nicotine.
Keywords: Nicotine isomers; Trigeminal ganglia; Receptor modulation; Chemosensation; Human chemoreceptor; Recombinant expression;

Activated microglia in the spinal cord underlies diabetic neuropathic pain by Dongmei Wang; Réjean Couture; Yanguo Hong (59-66).
Diabetes mellitus is an increasingly common chronic medical condition. Approximately 30% of diabetic patients develop neuropathic pain, manifested as spontaneous pain, hyperalgesia and allodynia. Hyperglycemia induces metabolic changes in peripheral tissues and enhances oxidative stress in nerve fibers. The damages and subsequent reactive inflammation affect structural properties of Schwann cells and axons leading to the release of neuropoietic mediators, such as pro-inflammatory cytokines and pro-nociceptive mediators. Therefore, diabetic neuropathic pain (DNP) shares some histological features and underlying mechanisms with traumatic neuropathy. DNP displays, however, other distinct features; for instance, sensory input to the spinal cord decreases rather than increasing in diabetic patients. Consequently, development of central sensitization in DNP involves mechanisms that are distinct from traumatic neuropathic pain. In DNP, the contribution of spinal cord microglia activation to central sensitization and pain processes is emerging as a new concept. Besides inflammation in the periphery, hyperglycemia and the resulting production of reactive oxygen species affect the local microenvironment in the spinal cord. All these alterations could trigger resting and sessile microglia to the activated phenotype. In turn, microglia synthesize and release pro-inflammatory cytokines and neuroactive molecules capable of inducing hyperactivity of spinal nociceptive neurons. Hence, it is imperative to elucidate glial mechanisms underlying DNP for the development of effective therapeutic agents. The present review highlights the recent developments regarding the contribution of spinal microglia as compelling target for the treatment of DNP.
Keywords: Bradykinin B1 receptor; Diabetic neuropathic pain; Hyperglycemia; Microglia; Proinflammatory cytokines; Spinal cord;

Berberine improves pressure overload-induced cardiac hypertrophy and dysfunction through enhanced autophagy by Ming-Hui Li; Yao-Jun Zhang; Yi-Hui Yu; Shao-Hua Yang; Javaid Iqbal; Qiong-Yu Mi; Bing Li; Zhi-Mei Wang; Wen-Xing Mao; Hong-Guang Xie; Shao-Liang Chen (67-76).
Cardiac hypertrophy is a maladaptive change in response to pressure overload, and is also an important risk for developing heart failure. Berberine is known to have cardioprotective effects in patients with hypertension and in animal models of cardiac hypertrophy. In the current study, we observed that transverse aortic contraction (TAC) surgery induced a marked increase in heart size, the ratio of heart weight to body weight, cardiomyocyte apoptosis, myocardial fibrosis, and hypertrophic marker brain natriuretic peptide, all of which were effectively suppressed by berberine administration. In addition, berberine enhanced autophagy in hypertrophic hearts, which was accompanied by a decrease in heart size, cardiac apoptosis, and the attenuation of cardiac dysfunction. Furthermore, use of autophagy inhibitor 3-methyladenine (3-MA) blocked berberine-induced autophagy level, and abrogated the protection of berberine against heart hypertrophy, cardiac dysfunction, and apoptosis. Berberine ameliorated TAC-induced endoplasmic reticulum stress, which was also abolished by 3-MA. Moreover, berberine significantly inhibited the upstream signaling of autophagy, such as the mammalian target of rapamycin (mTOR), extracellular signal-regulated kinase (ERK1/2), and p38 mitogen-activated protein kinase (MAPK) phosphorylation. We conclude that berberine could attenuate left ventricular remodeling and cardiomyocyte apoptosis through an autophagy-dependent mechanism in a rat model of cardiac hypertrophy, which is, at least in part, associated with enhanced autophagy through inhibition of mTOR, p38 and ERK1/2 MAPK signaling pathways.
Keywords: Berberine; Transverse aortic constriction; Hypertrophy; Autophagy; Endoplasmic reticulum stress;

Intestinal and hepatic Niemann–Pick C1L1 proteins: Future therapeutic targets for cholesterol gallstones disease? by Ibrahim Guillermo Castro-Torres; Minarda De la O-Arciniega; Gustavo Adolfo Bravo-Duarte; Janeth Gallegos-Estudillo; Miguel Ángel Domínguez-Ortíz; Mariano Martínez-Vázquez (77-81).
The formation of cholesterol gallstones is a very complex and polygenic disorder that involves an alteration of the secretion of bile lipids, cholesterol crystallization, important immunological reactions in the gallbladder tissue, formation of biliary sludge composed of mucin, and inadequate gallbladder motility. The search for a therapeutic target is oriented towards decreasing bile secretion and intestinal absorption of cholesterol, in which Niemann–Pick C1L1 (NPC1L1) proteins play an important role. In basic and clinical studies, regulating the expression of these proteins can reduce intestinal, liver, plasma and bile cholesterol levels, a therapeutic effect that would be useful not only for treating the disease, but to prevent it, given the large quantity of risk factors. We discuss these effects in this review and propose NPC1L1 proteins as future therapeutic targets of cholesterol gallstones disease.
Keywords: Cholesterol gallstones; NPC1L1 proteins; Therapeutic target; Gallbladder;

Dracorhodin perchlorate induces apoptosis in primary fibroblasts from human skin hypertrophic scars via participation of caspase-3 by Peihua Zhang; Jin Li; Xudong Tang; Junlei Zhang; Jie Liang; Guofang Zeng (82-92).
Hypertrophic scar (HS) is an abnormally proliferative disorder characterized by excessive proliferation of fibroblasts and redundant deposition of extracellular matrix. An unbalance between fibroblast proliferation and apoptosis has been assumed to play an important role in HS formation. To explore the regulative effects of dracorhodin perchlorate (Dp), one of the derivants of dracorhodin that is a major constituent in the traditional Chinese medicine, on primary fibroblasts from human skin hypertrophic scars, 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometric analysis were respectively used to evaluate the inhibitory effect of Dp on the cells and to determine cell cycle distribution. Additionally, cellular apoptosis was separately detected with Hoechst 33258 staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay. The expression levels of caspase-3 mRNA and protein were respectively measured with reverse transcription-polymerase chain reaction and western blot analysis, and caspase-3 activity were determined using a colorimetric assay kit. The results showed that Dp significantly inhibited cell growth, and induced apoptosis in fibroblasts in a dose-and time-dependent manner, arresting cell cycle at G1 phase. Additionally, Dp slightly up-regulated caspase-3 mRNA expression in fibroblasts, but significantly down-regulated caspase-3 protein expression in a dose- and time-dependent manner, and concurrently elevated caspase-3 activity. Taken together, these data indicated that Dp could effectively inhibit cell proliferation, and induced cell cycle arrest and apoptosis in fibroblasts, at least partially via modulation of caspase-3 expression and its activity, which suggests that Dp is an effective and potential candidate to develop for HS treatment.
Keywords: Dracorhodin perchlorate; Apoptosis; Caspase-3; Fibroblasts; Hypertrophic scar;

Hyperbaric oxygen enlarges the area of brain damage in MCAO rats by blocking autophagy via ERK1/2 activation by Yuxiong Lu; Jinsong Kang; Yang Bai; Yu Zhang; Hongyan Li; Xiaochun Yang; Xiyan Xiang; Xinxue Wang; Yuanping Huang; Jing Su; Ye Chen; Bingjin Li; Liankun Sun (93-99).
Hyperbaric oxygen (HBO) is emerging as a therapy for brain ischemia, although its benefits are still debated. The present study aimed to investigate the effect of HBO on brain damage in a rat model of transient focal cerebral ischemia and its underlying mechanism of action. Male Wistar rats, which had suffered 1.5 h of transient middle cerebral artery occlusion (tMCAO) and had a Longa׳s neuron score of 3, were given pure oxygen at 3.0 atm absolute, for 60 min after the third hour of reperfusion. After 24 h of reperfusion, rat brains were removed and studied. 2,3,5-triphenyltetrazolium chloride (TTC) and hematoxylin and eosin staining revealed that the infarct ratio in the HBO group increased remarkably when compared with the MCAO group. Up-regulation of extracellular signal-regulated kinase 1/2 (ERK1/2) activation was detected in the HBO group because of reactive oxygen species (ROS) generation. Autophagy appeared to be obstructed in the HBO group. Administration of the ERK1/2 inhibitor U0126 decreased the infarct ratio and improved protein clearance by autophagy in the HBO group. Collectively, these results suggest that HBO enlarges the area of brain damage via reactive oxygen species-induced activation of ERK1/2, which interrupts autophagy flux.
Keywords: Hyperbaric oxygen; Autophagy; Extracellular signal-regulated kinase 1/2; Middle cerebral artery occlusion;

Risperidone inhibits voltage-gated sodium channels by Jan M. Brauner; Sabine Hessler; Teja W. Groemer; Christian Alzheimer; Tobias Huth (100-106).
In contrast to several other antipsychotic drugs, the effects of the atypical antipsychotic risperidone on voltage-gated sodium channels have not been characterized yet, despite its wide clinical use. Here we performed whole-cell voltage-clamp recordings to analyze the effects of risperidone on voltage-dependent sodium currents of N1E-115 mouse neuroblastoma cells carried by either endogenous sodium channels or transfected NaV1.6 channels. Risperidone inhibited both endogenous and NaV1.6-mediated sodium currents at concentrations that are expected around active synaptic release sites owing to its strong accumulation in synaptic vesicles. When determined for pharmacologically isolated NaV1.6, risperidone inhibited peak inward currents with an IC50 of 49 µM. Channel block occurred in a state-dependent fashion with risperidone displaying a fourfold higher affinity for the inactivated state than for the resting state. As a consequence of the low state dependence, risperidone produced only a small, but significant leftward shift of the steady-state inactivation curve and it required concentrations≥30 µM to significantly slow the time course of recovery from inactivation. Risperidone (10 µM) gave rise to a pronounced use-dependent block when sodium currents were elicited by trains of brief voltage pulses at higher frequencies. Our data suggest that, compared to other antipsychotic drugs as well as to local anesthetics and sodium channel-targeting anticonvulsants, risperidone displays an unusual blocking profile where a rather low degree of state dependence is associated with a prominent use-dependent block.
Keywords: Risperidone; Sodium channel; NaV1.6; Use dependence; Antipsychotic drug; Schizophrenia; Chemical compounds studied in this article:Risperidone (PubChem CID: 5073);

Chrysin alleviates acute doxorubicin cardiotoxicity in rats via suppression of oxidative stress, inflammation and apoptosis by Eman M. Mantawy; Wesam M. El-Bakly; Ahmed Esmat; Amira M. Badr; Ebtehal El-Demerdash (107-118).
Doxorubicin (DOX) is one of the most effective chemotherapeutic drugs; however, its incidence of cardiotoxicity compromises its therapeutic index. Chrysin, a natural flavone, possesses multiple biological activities, such as antioxidant, anti-inflammatory and anti-cancer. The present study was designed to investigate whether chrysin could protect against DOX-induced acute cardiotoxicity; and if so, unravel the molecular mechanisms of this protective effect. Chrysin was administered to male albino rats once daily for 12 consecutive days at doses of 25 and 50 mg/kg orally. DOX (15 mg/kg; i.p.) was administered on day 12. Chrysin pretreatment significantly protected against DOX-induced myocardial damage which was characterized by conduction abnormalities, increased serum creatine kinase isoenzyme-MB (CK-MB), and lactate dehydrogenase (LDH) and myofibrillar disarrangement. As indicators of oxidative stress, DOX caused significant glutathione depletion, lipid peroxidation and reduction in activities of antioxidant enzymes; catalase (CAT) and superoxide dismutase (SOD). Chrysin pretreatment significantly attenuated DOX-induced oxidative injury. Additionally, DOX provoked inflammatory responses by increasing the expressions of nuclear factor kappa-B (NF-κB), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and the levels of tumor necrosis factor-alpha (TNF-α) and nitric oxide while chrysin pretreatment significantly inhibited these inflammatory responses. Furthermore, DOX induced apoptotic tissue damage by increasing Bax and cytochrome c expressions and caspase-3 activity while decreasing the expression of Bcl-2. Chrysin pretreatment significantly ameliorated these apoptotic actions of DOX. Collectively, these findings indicate that chrysin possesses a potent protective effect against DOX-induced acute cardiotoxicity via suppressing oxidative stress, inflammation and apoptotic tissue damage.
Keywords: Cardiotoxicity; Doxorubicin; Chrysin; Oxidative stress; Inflammation; Apoptosis;

The hypotensive agent dodoneine inhibits L-type Ca2+ current with negative inotropic effect on rat heart by Grégoire Carré; Hélène Carreyre; Maurice Ouedraogo; Frédéric Becq; Patrick Bois; Sébastien Thibaudeau; Clarisse Vandebrouck; Jocelyn Bescond (119-127).
Agelanthus dodoneifolius is one of the medicinal plants used in African pharmacopeia and traditional medicine for the treatment of cardiovascular diseases. A chemical analysis has identified one of the active principles: Dodoneine (Ddn). It is a new dihydropyranone which exerts hypotensive and vasorelaxant effects on rat. Since the mechanism of the hypotensive effect is unknown, we performed a variety of preclinical and mechanistic studies to characterize the specific cardiac effect of Ddn at tissue (ex-vivo) and cellular levels (in-vitro) in order to determine a molecular target. Ddn effects were evaluated in an isolated rat heart preparation using Langendorff retrograde perfusion and then, the effects of Ddn were characterized in freshly dissociated cardiac ventricular myocytes using the whole-cell patch-clamp configuration. Ex-vivo, Ddn produced a dose-dependent negative inotropic effect with an IC50 value of 10 µM without changed heart rate. 100 µM Ddn decreased left ventricular developed pressure of about 40%. In isolated cardiac myocytes, Ddn reduced I Ca,L density of about 30% with an IC50 value estimated at 3 µM. Ddn did not change current–voltage relation but it shifted the inactivation curve toward negative potentials and modified the half inactivation potentials. Furthermore, Ddn induced a phasic-dependent blocking on I Ca,L. This study demonstrates that the hypotensive property of dodoneine is likely associated with a negative inotropic effect and the blockade of the L-type calcium channels.Display Omitted
Keywords: Dodoneine; Natural substance; Hypertension; Isolated rat heart; L-type calcium current; Calcium channel blocking drugs;

Cardioprotective effects of magnesium valproate in type 2 diabetes mellitus by Bhoomika M. Patel; Suchi Raghunathan; Urvashi Porwal (128-134).
We have evaluated the effect of magnesium valproate (210 mg/kg/day, p.o.) in type 2 diabetes induced cardiovascular complications induced by streptozotocin (STZ, 90 mg/kg, i.p.) in neonatal wistar rats. Various biochemical, cardiovascular and hemodynamic parameters were measured at the end of 8 weeks of treatment. STZ produced significant hyperglycaemia, hypoinsulinemia and dyslipidemia, which was prevented by magnesium valproate treatment. STZ produced increase in Creatinine Kinase, C-reactive protein and lactate dehydrogenase levels and treatment with magnesium valproate produced reduction in these levels. STZ produced increase in cardiac and LV hypertrophy index, LV/RV ratio, LV collagen deposition and LV cardiomyocyte diameter which were decreased by magnesium valproate treatment. Magnesium valproate also prevented STZ induced hemodynamic alterations and oxidative stress. These results were further supported by histopathological studies in which magnesium valproate showed marked reduction in fibrosis and cardiac fiber disarray. In conclusion, our data suggests that magnesium valproate is beneficial as an anti-diabetic agent in type-2 diabetes mellitus and also prevents its cardiac complications.
Keywords: Magnesium valproate; Type 2 diabetes mellitus; HDAC; Cardiovascular complications;

Atorvastatin prevents type 2 diabetes mellitus—An experimental study by Sri Venkata Madhu; Mohammad Aslam; Vikas Galav; Swapan Kumar Bhattacharya; Aiman Abbas Jafri (135-140).
Recent reports of increased diabetes risk have raised concerns regarding the use of statins. The present study was therefore planned to clarify whether atorvastatin can prevent diabetes development in a rat model of type 2 diabetes mellitus. Eight week old male Wistar rats were randomized into three groups (n=12 each group). Group A was given standard chow diet, while group B and group C were offered high sucrose diet. In addition to high sucrose diet, group C was given atorvastatin (20 mg/kg/day) from beginning of study till 26th week. After 26 weeks, a low dose of streptozotocin (15 mg/kg, i.p.) was given to all 3 groups and further followed for 4 weeks. Oral glucose tolerance tests were done at week 4, 26 and week 30. Development of impaired glucose tolerance at week 26 (16.66% vs 100%, P=<0.001) and diabetes at week 30 (16.66% vs 81.81%, P=0.002) was significantly lower in rats pretreated with atorvastatin along with high sucrose diet viz group C compared to group B rats who received high sucrose diet only respectively. Also, metabolic indices like body weight, hypertriglyceridemia, glucose area under the curve (Gl-AUC) were significantly lower in group C compared to group B (P=<0.05) while insulin resistance (HOMA-IR) was also lower in group C (P=0.05). This study clearly demonstrates for the first time in a rat model of type 2 diabetes mellitus that atorvastatin prevents development of type 2 diabetes.
Keywords: Atorvastatin; Hypertriglyceridemia; Glucose area under the curve; Insulin resistance; Impaired glucose tolerance; Type 2 diabetes;

Role of microRNA-95 in the anticancer activity of Brucein D in hepatocellular carcinoma by Zhangang Xiao; Sheung Ching Chow; Chi Han Li; Shing Chun Tang; Stephen K.W. Tsui; Zhixiu Lin; Yangchao Chen (141-150).
Brucea javanica fruit has been used to treat amebic dysentery, malaria and various parasites and it has been applied as an anti-cancer agent in Traditional Chinese Medicine. Brucein D (BD) is a naturally occurring compound extracted from Brucea javanica fruit which shows anti-cancer activity against pancreatic cancer. Here, we further demonstrated that BD inhibited hepatocellular carcinoma (HCC) cell growth in vitro and tumor growth in vivo that were attributed to the induction of cell apoptosis. BD did not exert growth inhibition on non-tumorigenic human hepatocytes. MTT assay was used to measure cell viability. Annexin V and TUNEL assay were applied to identify apoptotic cells in cell suspension and in tissue section respectively. Downstream micro-RNA (miRNA) targets of BD were screened out by miRNA array. miRNAs and their target proteins were identified by bioinformatics analysis and luciferase reporter assay. 39 miRNAs regulated by BD in HCC were identified. miR-95 was found to be a potential drug target of BD. We further identified CUG triplet repeat RNA-binding protein 2 (CUGBP2) as the downstream target of miR-95. Our data suggested that BD exerted its anti-cancer activity against HCC through modulation of miR-95 expression.
Keywords: Brucein D; Hepatocellular carcinoma; MicroRNA-95;

Suramin inhibits hepatic tissue damage in hepatocellular carcinoma through deactivation of heparanase enzyme by Ahmed Tayel; Khaled H. Abd El Galil; Mohamed A. Ebrahim; Ahmed S. Ibrahim; Amal M. El-Gayar; Mohammed M.H. Al-Gayyar (151-160).
Hepatocellular carcinoma (HCC) is resistant to conventional chemotherapy, and is rarely amenable to radiotherapy. Heparanase, enzyme attacks heparan sulfate proteoglycans (HSPGs), is preferentially expressed in human tumors and its overexpression in low-metastatic tumor confers a highly invasive phenotype in experimental animals. Meanwhile, high doses of suramin dramatically increase tissue glycosaminoglycans due, in part, to inhibition of heparanase enzymes. Therefore, the following study was conducted to evaluate the chemopreventive and hepatoprotective effects of suramin in in-vivo model of HCC. Therefore, HCC was induced in SD rats by thioacetamide (200 mg/kg) in presence/absence of suramin (20 mg/kg). Liver impairment was assessed by measuring serum α-fetoprotein and investigating liver sections stained with Hematoxylin/Eosin. Hepatic HSPGs and heparanse were measured by ELISA. Glucosamine and glucuronic acid were measured by chemical methods. Gene expression of fibroblast growth factor (FGF)-2 and caspase-3 was measured. Apoptotic pathway was evaluated by measuring the activity of caspase-3/8/9. Suramin increased the animal survival and decreased serum α-fetoprotein. In addition, suramin ameliorated fibrosis and massive hepatic tissue breakdown. Suramin restored hepatic HSPGs and reduced the activity of hepatic heparanase leading to decreased hepatic levels of glucosamine and glucuronic acid. Moreover, suramin reduced the gene expression of FGF-2 and caspase-3. Finally, suramin blocked the elevated activity of caspase-3/8/9. In conclusion, surmain showed antitumor activity as well as hepatoprotective effects. Besides its antioxidant activity, other mechanisms are involved including restoration of HSPGs and inhibition of heparanase and FGF-2. Suramin inhibits intrinsic and extrinsic apoptotic pathway. Targeting HSPGs expression is potential therapeutic target for HCC.
Keywords: Extrinsic cell death; Fibroblast Growth Factor (FGF)-2; Glucosamine; Glucuronic acid; Heparan sulfate proteoglycans (HSPGs); Intrinsic cell death;

Effects of anatabine and unilateral maximal eccentric isokinetic muscle actions on serum markers of muscle damage and inflammation by Nathaniel D.M. Jenkins; Terry J. Housh; Kristen C. Cochrane; Haley C. Bergstrom; Daniel A. Traylor; Robert W. Lewis; Samuel L. Buckner; Richard J. Schmidt; Glen O. Johnson; Joel T. Cramer (161-166).
The purpose of this study was to examine the effects of anatabine supplementation in conjunction with unilateral, maximal eccentric isokinetic muscle actions on serum markers of muscle damage and pro-inflammatory cytokines in humans. Seventeen men (mean±S.D. age=22.4±3.2 yrs) participated in this double-blinded, placebo-controlled, crossover study. Participants were randomly assigned to two 10-day conditions (anatabine and placebo) separated by a 2–4 week washout period. After seven days of supplementation, blood was sampled immediately prior to PRE, immediately following POST, and 24, 48, and 72 h after 6 sets of 10 repetitions of unilateral, maximal eccentric isokinetic forearm flexion exercise. Concentrations of serum creatine kinase, lactate dehydrogenase, myoglobin, high sensitivity c-reactive protein, and TNF-α were measured. Creatine kinase, myoglobin, and lactate dehydrogenase increased (P<0.05), while high sensitivity c-reactive protein and TNF-α did not change (P>0.05) after the eccentric exercise during both conditions. Lactate dehydrogenase was higher (P<0.05) during the anatabine condition. The primary findings of this study were two-fold: (a) anatabine had no beneficial effects on traditional markers of muscle damage (creatine kinase, lactate dehydrogenase, and myoglobin) compared to placebo after the eccentric exercise protocol, and (b) the eccentric exercise protocol did not elicit increase in the pro-inflammatory cytokines (c-reactive protein and TNF-α). Future studies are needed to examine the effects of anatabine on naturally-occurring inflammation that is common with aging or obesity. Furthermore, additional research is needed to examine the relationship between muscle damage and inflammation after eccentric exercises of different modes, durations, and intensities.
Keywords: Anatabine; Supplementation; Eccentric muscle damage; Muscle function; Cytokines;

Modes of direct modulation by taurine of the glutamate NMDA receptor in rat cortex by Christopher Y. Chan; Herless S. Sun; Sanket M. Shah; Mervan S. Agovic; Eitan Friedman; Shailesh P. Banerjee (167-175).
Taurine is an endogenous brain substance with robust neuromodulatory and possible neuroprotective properties. Though other mechanisms of action have been reported, its interaction with the NMDA (N-methyl-d-aspartic acid) receptor is undocumented. We investigated taurine׳s interaction with the NMDA receptor using electrophysiological and receptor binding approaches. The effects of taurine on field potential responses in layer-5 of prelimbic cortex in rat brain slices evoked by single-pulse electrical stimulation of ventral medial cortex were determined. Picrotoxin (80 µM) was present in all control and drug solutions to block the Cl channels associated with the GABA-, taurine-, and strychnine sensitive glycine- receptors. A typical response consisted of an NBQX (2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo-[f]-quinoxaline-7-sulfonamide)-sensitive negative wave (N1) followed by a positive wave (P1) and a broad negativity (N2), both sensitive to dl-AP5 (dl-2-amino-5-phosphonopentanoic acid) inhibition. Taurine exerted a 41.5±8.3% (n=9) voltage reduction within the late phase of N2. This taurine action was prevented by 100 µM AP5, but not by 10 µM nifedipine, supporting a direct modulation of NMDA receptor function by taurine, without requiring the involvement of the l-type Ca2+ channel. Taurine did not alter specific [3H] MK-801 binding to rat cortical membranes in the presence of glycine or glutamate; but inhibited spermine-potentiated specific [3H] MK-801 binding to NMDA receptors by 15–20% in the presence of glycine. In addition, taurine reduced the apparent affinity of the NMDA receptor for glycine (in the presence of spermine) by 10-fold. These results show that taurine interacts directly with the NMDA receptor by multiple mechanisms.
Keywords: Taurine; NMDA receptor; Spermine; Glycine; Prefrontal cortex; l-type calcium channel;

Effect of sauchinone, a lignan from Saururus chinensis, on bacterial phagocytosis by macrophages by Kyung-Min Jeong; Jeong-Il Choi; Seong-Heon Lee; Hyun-Jung Lee; Jong-Keun Son; Chang-Seob Seo; Seung-Won Song; Sang-Hyun Kwak; Hong-Beom Bae (176-182).
AMP-activated protein kinase (AMPK) plays an important role in inflammation in various cells and increases the phagocytic ability of macrophages. In this study, we found that sauchinone increased the phosphorylation of AMPK and acetyl-CoA carboxylase (ACC), a downstream target of AMPK, in mouse peritoneal macrophages. Sauchinone increased macrophage phagocytosis of fluorescent Escherichia coli, which was blocked by compound C, an AMPK inhibitor. Sauchinone also increased the phosphorylation of p38 mitogen activated protein kinase (MAPK) in cultured macrophages in a concentration-dependent fashion, which was not blocked by compound C. However, the increase of sauchinone-induced phagocytosis was prevented by SB203580. An inhibitor of the upstream kinase TGF-beta-activated kinase (TAK1), (5z)-7-oxozeaenol, abolished the phosphorylation of ACC and p38 MAPK. Systemic administration of sauchinone to mice led to increased phosphorylation of AMPK and p38 MAPK in the lung, and enhanced phagocytosis of fluorescent E. coli in bronchoalveolar lavage fluid as compared with control mice. These results suggest sauchinone to be a useful adjunctive treatment for bacterial infection.
Keywords: AMP-activated protein kinase; Bacteria; Phagocytosis; p38 Mitogen activated protein kinase; Sauchinone;

Roles of subunit phosphorylation in regulating glutamate receptor function by John Q. Wang; Ming-Lei Guo; Dao-Zhong Jin; Bing Xue; Eugene E. Fibuch; Li-Min Mao (183-187).
Protein phosphorylation is an important mechanism for regulating ionotropic glutamate receptors (iGluRs). Early studies have established that major iGluR subtypes, including α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors and N-methyl-d-aspartate (NMDA) receptors, are subject to phosphorylation. Multiple serine, threonine, and tyrosine residues predominantly within the C-terminal regions of AMPA receptor and NMDA receptor subunits have been identified as sensitive phosphorylation sites. These distinct sites undergo either constitutive phosphorylation or activity-dependent phosphorylation induced by changing cellular and synaptic inputs. An increasing number of synapse-enriched protein kinases have been found to phosphorylate iGluRs The common kinases include protein kinase A, protein kinase C, Ca2+/calmodulin-dependent protein kinase II, Src/Fyn non-receptor tyrosine kinases, and cyclin dependent kinase-5. Regulated phosphorylation plays a well-documented role in modulating the biochemical, biophysical, and functional properties of the receptor. In the future, identifying the precise mechanisms how phosphorylation regulates iGluR activities and finding the link between iGluR phosphorylation and the pathogenesis of various brain diseases, including psychiatric and neurodegenerative diseases, chronic pain, stroke, Alzheimer's disease and substance addiction, will be hot topics and could contribute to the development of novel pharmacotherapies, by targeting the defined phosphorylation process, for suppressing iGluR-related disorders.
Keywords: Excitatory amino acid; AMPA; NMDA; PKA; PKC; CaMKII; Cdk5; Tyrosine kinase;

Erratum to “Effects of clonidine in the isolated rat testicular capsule” [Eur. J. Pharmacol. 726 (2014) 16–26] by Edilson Dantas da Silva Júnior; Bruno Palmieri de Souza; Juliano Quintella Dantas Rodrigues; Afonso Caricati-Neto; Aron Jurkiewicz; Neide H. Jurkiewicz (188-189).

Retraction notice to “PBOX-15 induces apoptosis and improves the efficacy of oxaliplatin in human colorectal cancer cell lines” [Eur. J. Pharmacol. 714(1–3) (2013) 379–387] by Giuseppina Gangemi; Patrizia Gazzerro; Donatella Fiore; Maria Chiara Proto; Stefania Butini; Sandra Gemma; Alice Casagni; Chiara Laezza; Mario Vitale; Alessia Ligresti; Vincenzo DiMarzo; Daniela M. Zisterer; Seema Nathwani; D. Clive Williams; Giuseppe Campiani; Maurizio Bifulco (190).