Sub-Chronic Restraint Stress Suppresses Sexual Potency and Erection Efficiency by Targeting the Hypothalamic-Pituitary-Testicular Axis and the Nitric Oxide/Cyclic Guanosine Monophosphate/Phosphodiesterase 5α Pathway in Adult Rats.

INTRODUCTION: Male sexual potency and vigor are a complex neuroendocrine process and an important component of well-being. Psychological stress is one of the leading causes of male impotence worldwide. Therefore, to better understand the effects of psychological stress on male sexual potency, vigor, and the physiology of erection, we used the rat restraint stress (RS) model, which can most aptly simulate psychological stress. METHODS: Adult male SD rats were exposed to RS for 1.5 or 3 h/day for 30 days. Neuromodulators and hormones of sexual potency and penile erection were quantified using ELISA kit. The histoarchitecture of the penis was examined using Masson trichrome staining. Immunoblotting and immunofluorescence were used to assess the expression and immunolocalization patterns of penile erection markers. To assess sexual potency and vigor, a noncontact erection and a copulatory test were performed. RESULTS: RS exposure decreased the circulatory levels of gonadotropins and testosterone while increasing the serum corticosterone level. RS exposure altered the histomorphology of the penis by decreasing the smooth muscle/collagen ratio and increasing oxidative stress in penile tissue. Furthermore, RS adversely affected NO availability for penile erection by decreasing the neurotransmitter acetylcholine and other erection facilitatory markers such as p-Akt, nNOS, eNOS, and cGMP, while increasing the inhibitory marker PDE5α in the penis. RS exposure significantly reduced the frequencies of mount, intromission, and ejaculation, whereas it prolonged sexual exhaustion by increasing latencies of postejaculatory mount, intromission, and ejaculation. CONCLUSION: The current findings suggest that psychological stressors, such as RS, cause erectile dysfunction in adult male rats by modulating the hypothalamic-pituitary-testicular axis, oxidative balance, penile fibrosis, and the NO/cGMP/PDE5α pathway of penile erection.

Zhenwu decoction ameliorates cardiac hypertrophy through activating sGC (soluble guanylate cyclase) - cGMP (cyclic guanosine monophosphate) - PKG (protein kinase G) pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Zhenwu Decoction (ZWD) is a traditional Chinese medicine (TCM) formula which has wide scope of indications related to Yang deficiency and dampness retention in TCM syndrome. Cardiac hypertrophy can induce similar symptoms and signs to the clinical features of Yang deficiency and dampness retention syndrome. ZWD can increase the left ventricular ejection fraction, reduce cardiac hypertrophy of patients with chronic heart failure. However, its underlying pharmacological mechanism remains unclear. AIM OF THE STUDY: The study aimed to confirm the protective effects of ZWD on cardiac hypertrophy and explore the underlying mechanisms. MATERIALS AND METHODS: The potential targets and pathways of ZWD in cardiac hypertrophy were highlighted by network pharmacology and validated by mechanistic and functional studies. RESULTS: Our network pharmacology analysis suggests that the protective effects of ZWD on cardiac hypertrophy are related to cyclic guanosine monophosphate (cGMP) - protein kinase G (PKG) pathway. Subsequent animal studies showed that ZWD significantly ameliorated cardiac function decline, cardiac hypertrophy, cardiac fibrosis and cardiomyocyte apoptosis. To explore the underlying mechanisms of action, we performed Western blotting, immunohistochemical analysis, and detection of inflammatory response and oxidative stress. Our results showed that ZWD activated the soluble guanylate cyclase (sGC) - cGMP - PKG signaling pathway. The sGC inhibitor ODQ that blocks the sGC-cGMP-PKG signaling pathway in zebrafish abolished the protective effects of ZWD, suggesting sGC-cGMP-PKG is the main signaling pathway mediates the protective effect of ZWD in cardiac hypertrophy. In addition, three major ingredients from ZWD, poricoic acid C, hederagenin and dehydrotumulosic acid, showed a high binding energy with prototype sGC. CONCLUSION: ZWD reduces oxidative stress and inflammation and exerts cardioprotective effects by activating the sGC-cGMP-PKG signaling pathway.

Shunxin decoction improves diastolic function in rats with heart failure with preserved ejection fraction induced by abdominal aorta constriction through cyclic guanosine monophosphate-dependent protein kinase Signaling Pathway.

OBJECTIVE: To determine whether Shunxin decoction improves diastolic function in rats with heart failure with preserved ejection fraction (HFpEF) by regulating the cyclic guanosine monophosphate-dependent protein kinase (cGMP-PKG) signaling pathway. METHODS: Except for control group 8 and sham surgery group 8, the remaining 32 male Sprague-Dawlay rats were developed into HFpEF rat models using the abdominal aorta constriction method. These rats in the HFpEF model were randomly divided into the model group, the Shunxin high-dose group, the Shunxin low-dose group, and the Qiliqiangxin capsule group. The three groups received high-dose Shunxin decoction, low-dose Shunxin decoction, and Qiliqiangxin capsule by gavage, respectively, for 14 d. After the intervention, the diastolic function of each rat was evaluated by testing E/A, heart index, hematoxylin-eosin staining, Masson, myocardial ultrastructure, and N-terminal pro-brain natriuretic peptide (NT-proBNP). The Bioinformatics Analysis Tool for Molecular Mechanism of Traditional Chinese Medicine (BATMAN-TCM) software was used to predict targets for which Shunxin decoction acts on the cGMP-PKG pathway. Natriuretic peptide receptor A (NPRA) and guanylate cyclase (GC) were detected by immunohistochemistry, and eNOS, phosphodiesterase 5A (PDE5A), and cGMP-dependent protein kinase 1(PKG I) were determined by Western blotting. RESULTS: Compared to the model group, the thickness of the interventricular septum at the end of diastole (IVSd) and the thickness of the posterior wall at the end of diastole (PWd) of the Shunxin decoction high-dose group, Shunxin decoction low-dose group, and Qiliqiangxin capsule group were all significantly reduced ( < 0.01). Furthermore, Shunxin decoction high-dose group E/A value was decreased ( < 0.01). Compared to the model group, the expression of NPRA and GC increased in the Shunxin decoction low-dose group and the Qiliqiangxin capsule group ( < 0.01). Compared to the model group, the expressions of eNOS and PKG I increased ( < 0.05) in the Shunxin decoction high-dose group. The expression of PDE5A expression decreased in the myocardium of the Shunxin decoction high-dose group, Shunxin decoction low-dose group, and Qiliqiangxin capsule group compared to the model group ( < 0.01). CONCLUSIONS: Shunxin decoction can improve diastolic function in rats with HFpEF. It increases the expression of NPRA, GC, and eNOS in the myocardial cell cGMP-PKG signaling pathway, upregulates cGMP expression, decreases PDE5A expression to reduce the cGMP degradation. Thus, the cGMP continually stimulates PKG I, reversing myocardial hypertrophy and improving myocardial compliance in HFpEF rats.

Double-blind controlled dietary cross-over intervention with differentially fertilised intact lettuce leaves shows acute reduction in blood pressure in young adults, associated with faster uptake of nitrate than of phenolics.

PURPOSE: To compare acute effects on blood pressure (BP) of ingestion of visually similar lettuce with controlled high and low content of either nitrate or phenolic compounds. METHODS: In a randomised cross-over design, 19 healthy participants (22-31 years) received 50 g of lettuce containing either 530 mg (8.4 mmol) nitrate + 11 mg (0.03 mmol) phenolic compounds (HNLP); or 3 mg nitrate (0.05 mmol) + 77 mg (0.2 mmol) phenolic compounds (LNHP), obtained by differential fertilisation. Ambulatory BP was recorded along with plasma, salivary and urinary nitrate and nitrite and plasma concentrations of cyclic guanosine monophosphate (cGMP), phenolic metabolites, Trolox equivalent antioxidant capacity (TEAC) and ferric reducing antioxidant power (FRAP). RESULTS: Compared with LNHP, 3 h post ingestion of HNLP, plasma nitrate increased 0.31 ± (95%CI) 0.12 mM (+ 240%), and salivary nitrate 5.5 ± 1.4 mM (+ 910%); accumulated urinary nitrate excretion increased 188 ± 72 mg (+ 296%) (all P < 0.001). Systolic BP was reduced 4.9 ± 4.2 mmHg (P = 0.031) between 3 and 6 h after ingestion of HNLP compared with LNHP; systolic BP differences were negatively correlated (P = 0.004) with differences in saliva nitrate concentrations. LNHP increased plasma phenolics at 6 h, predominantly 3'-methoxycinnamic acid-4'-glucuronide (ferulic acid-4'-glucuronide), 116%, 204 ± 138 nM more than HNLP (P = 0.001); increased cGMP 14% (P = 0.019); and reduced FRAP 3.1% (P = 0.009). CONCLUSION: The acute BP difference within 6 h of consumption matched the plasma/saliva nitrate peak, not the slower changes of plasma phenolics. This is the first double-blind controlled dietary intervention demonstrating differential effects on human physiology by consumption of an intact plant food, where compositional differences were obtained by controlling growing conditions, indicating potential opportunities for health claims relating to precision/vertical farming. CLINICAL TRIAL REGISTRATION: The trial was retrospectively registered on ClinicalTrials.gov, with identifier NCT02701959, on March 8, 2016.

Probable mechanisms involved in the antiepileptic activity of Clerodendrum polycephalum Baker (Labiatae) leaf extract in mice exposed to chemical-induced seizures.

The leaves of Clerodendrum polycephalum Baker (Labiatae) are used as a dietary legume supplement and applied ethnomedicinally for the management of epilepsy, convulsion, and spasms. This study is aimed at evaluating the effects of Clerodendrum polycephalum (CP) leaf extract on chemical-induced seizures in mice and the possible mechanisms of action. Swiss albino mice were pretreated with CP (50, 100, or 500 mg/kg, p.o.) prior to intraperitoneal injection of picrotoxin (PTX) or pentylenetetrazole (PTZ). However, the most effective dose was used to elucidate the role of GABAergic and nitric oxide-cyclic guanosine monophosphate (NO-cGMP) signaling mechanisms in mice brains. Accordingly, we evaluated the preventive and reversal effects of CP on kainic acid (KA)-induced temporal lobe epilepsy (TLE), oxidative stress, and neuroinflammatory in mice. The pretreatment of mice with CP delayed the latencies to PTX and PTZ-induced seizures and decrement in the period of tonic-clonic attacks. Interestingly, CP (100 mg/kg) completely prevented PTZ-induced tonic-clonic seizures. Contrastingly, flumazenil (benzodiazepine receptor antagonist), NG -nitro-L-Arginine (L-NNA) (10 mg/kg., neuronal nitric oxide synthase inhibitor), and methylene blue (MB) (2 mg/kg, a soluble guanylyl cyclase inhibitor) but not L-arginine (150 mg/kg., nitric oxide precursor) reversed CP-induced anticonvulsant-like effect in PTZ model. Furthermore, KA-elicited TLE was prevented by CP treatment. CP also attenuated KA-induced oxidative stress, cyooxygenase-2 (COX-2), and nuclear factor kappa-B (NF-κB) elevated expressions in the hippocampus. The study revealed that the ethanolic leaf extract of CP produced anticonvulsant actions through enhancement of antioxidant defense, GABAergic, and NO-cGMP signaling pathways as well as attenuation of inflammatory processes. PRACTICAL APPLICATIONS: The leaves of Clerodendrum polycephalum Baker (Labiatae) are used as a dietary legume supplement and applied ethnomedicinally for the management of epilepsy, convulsion, and spasms. For this reason, we believe that supplementation of the Clerodendrum polycephalum leaf extract would prevent epileptic-related disorders in mice induced with epileptic conditions using kainic acid and other behavioral phenotypic models. Here, our findings clearly revealed that Clerodendrum polycephalum leaf extract protects against conditions of epileptic-related disorders and thus might be relevant as a dietary supplement in the prevention or delay of the onset of seizures and epileptic behavior.

The NO-cGMP-K+ Channel Pathway Participates in Diuretic and Cardioprotective Effects of Blutaparon portulacoides in Spontaneously Hypertensive Rats.

Blutaparon portulacoides is a Brazilian plant species that is widely used in folk medicine. The present study investigated the role of an aqueous extract of B. portulacoides against hypertension in spontaneously hypertensive rats. The aqueous extract of B. portulacoides was obtained from the whole plant. Its chemical profile was analyzed by ultraperformance liquid chromatography-tandem mass spectrometry. The acute toxicity of the aqueous extract of B. portulacoides was evaluated in female Wistar rats. Male 6-month-old spontaneously hypertensive rats then received the aqueous extract of B. portulacoides (30, 100, and 300 mg/kg), hydrochlorothiazide (25 mg/kg), or vehicle once daily for 28 days. On days 1, 14, and 28, the diuretic effects of the aqueous extract of B. portulacoides were evaluated. The role of prostaglandins and the nitric oxide-cyclic guanosine monophosphate-potassium channel pathway in the diuretic activity of the aqueous extract of B. portulacoides was also investigated. At the end of the treatment, hepatic and renal biochemical markers, serum nitrotyrosine, malondialdehyde, nitrite, and aldosterone levels, and angiotensin-converting enzyme activity were measured. The electrocardiographic profile, blood pressure, and renal vascular reactivity were also assessed. The heart, kidneys, and liver were collected to determine relative organ weight, histopathology, and cardiac morphometry. Caffeic acid, ferulic acid, and several flavonoids were identified in the aqueous extract of B. portulacoides. No signs of toxicity were observed. Prolonged treatment with the aqueous extract of B. portulacoides (300 mg/kg) induced significant diuretic activity by activating the nitric oxide-cyclic guanosine monophosphate-potassium channel pathway. These effects reduced blood pressure and oxidative stress and prevented renal vascular dysfunction and left ventricular hypertrophy that was induced by hypertension. Overall, the present data suggest that the aqueous extract of B. portulacoides has important diuretic and cardioprotective effects by activation of the nitric oxide-cyclic guanosine monophosphate-potassium channel pathway.

Glucosyl-hesperidin enhances the cyclic guanosine monophosphate-inducing effect of a green tea polyphenol EGCG.

Animal and clinical studies have revealed that (-)-epigallocatechin-3-O-gallate (EGCG), one of the major bioactive polyphenols in green tea, showed several pharmacological effects including anti-obesity effect and anti-inflammatory effect. We previously reported that the second messenger cyclic guanosine monophosphate (cGMP) mediates its anti-inflammatory and anti-cancer properties. Here we demonstrated that glucosyl-hesperidin, enhances the cGMP-inducing effects of green tea extract in vivo. Moreover, glucosyl-hesperidin intake potentiated the green tea-elicited upregulation of the anti-inflammatory factor, toll-interacting protein.

Protective effects of 17-ß-oestradiol and phytoestrogen on age-induced oxidative stress and inhibition of surfactant synthesis in rat type II pneumocytes.

Epidemiological data suggest protective effects of oestrogen and phytoestrogen on lung tissue. This study aimed to elucidate the role of 17-ß-oestradiol and phytoestrogen in age-related inhibition of surfactant synthesis and oxidative stress in rat type II pneumocytes. Forty male and 66 female Wistar rats were used. Female rats were randomly kept intact or ovariectomized at age 12 months. At age 22 months, ovariectomized rats received 17-ß-oestradiol, soy extract, or no treatment. Oxidative stress markers CO, NO, cGMP and lipid peroxide (LPO), antioxidant enzymes and phosphatidylcholine (PC) were measured in cultured type II pneumocytes isolated at ages 2, 14, 18, 22 and 24 months. Old, male and ovariectomized rats showed significantly higher CO, NO, cGMP and LPO and lower PC content and antioxidant enzymes. 17-ß-oestradiol and phytoestrogen significantly reversed these effects. In conclusion, aging and oestrogen deprivation decreased PC synthesis and altered the redox status in type II pneumocytes, which were partially restored by 17-ß-oestradiol or soy supplementation.

6'-ß-Fluoro-Homoaristeromycin and 6'-Fluoro-Homoneplanocin A Are Potent Inhibitors of Chikungunya Virus Replication through Their Direct Effect on Viral Nonstructural Protein 1.

Alphaviruses are arthropod-borne, positive-stranded RNA viruses capable of causing severe disease with high morbidity. Chikungunya virus (CHIKV) is an alphavirus that causes a febrile illness which can progress into chronic arthralgia. The current lack of vaccines and specific treatment for CHIKV infection underscores the need to develop new therapeutic interventions. To discover new antiviral agents, we performed a compound screen in cell culture-based infection models and identified two carbocyclic adenosine analogues, 6'-ß-fluoro-homoaristeromycin (FHA) and 6'-fluoro-homoneplanocin A (FHNA), that displayed potent activity against CHIKV and Semliki Forest virus (SFV) with 50% effective concentrations in the nanomolar range at nontoxic concentrations. The compounds, designed as inhibitors of the host enzyme S-adenosylhomocysteine (SAH) hydrolase, impeded postentry steps in CHIKV and SFV replication. Selection of FHNA-resistant mutants and reverse genetics studies demonstrated that the combination of mutations G230R and K299E in CHIKV nonstructural protein 1 (nsP1) conferred resistance to the compounds. Enzymatic assays with purified wild-type (wt) SFV nsP1 suggested that an oxidized (3'-keto) form, rather than FHNA itself, directly inhibited the MTase activity, while a mutant protein with the K231R and K299E substitutions was insensitive to the compound. Both wt nsP1 and the resistant mutant were equally sensitive to the inhibitory effect of SAH. Our combined data suggest that FHA and FHNA inhibit CHIKV and SFV replication by directly targeting the MTase activity of nsP1, rather than through an indirect effect on host SAH hydrolase. The high potency and selectivity of these novel alphavirus mRNA capping inhibitors warrant further preclinical investigation of these compounds.

Nitric Oxide and Cyclic Guanosine Monophosphate Signaling Mediates the Antidepressant Effects of Acupuncture in the Rat Model of Chronic Unpredictable Mild Stress.

BACKGROUND Depression is a major mood disorder. Some patients have been reported to improve following acupuncture. This study aimed to investigate the effects of acupuncture on behaviors associated with depression in the chronic unpredictable mild stress (CUMS) rat model. The expression of signaling pathway components of nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) in the rat hippocampus and plasma were also measured. MATERIAL AND METHODS Male Sprague-Dawley rats (N=40) were divided into the control group (N=10), the model group (N=10), the acupuncture group (N=10), and the non-acupuncture group (N=10). The rat model was established by orphaning combined with chronic unpredictable mild stress (CUMS) for six weeks. The acupuncture group was given 21 days of treatment using acupoints (AP) or non-acupoints (NP). Rat behaviors associated with depression were tested using the sucrose preference test (SPT), the open field test (OFT), and the elevated plus maze (EPM) test. Enzyme-linked immunoassay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect the expression of inducible nitric oxide synthase (iNOS), neuronal nitric oxide synthase (nNOS), and the N-methyl-D-aspartate (NMDA) receptor subunits, NR1, NR2A, and NR2B in the rat plasma and hippocampus. RESULTS Acupuncture reversed the behaviors associated with depression in the CUMS rat model and reduced the expression of components of the NO and cGMP pathway in the rat hippocampus and plasma. CONCLUSIONS In the CUMS rat model, treatment with acupuncture reduced behaviors associated with depression, and these effects were associated with changes in the NO and cGMP signaling pathway.

Antiallodynic effect induced by [6]-gingerol in neuropathic rats is mediated by activation of the serotoninergic system and the nitric oxide-cyclic guanosine monophosphate-adenosine triphosphate-sensitive K+ channel pathway.

The present study evaluated the possible antiallodynic effect induced by [6]-gingerol in rats with L5-L6 spinal nerve ligation (SNL). Moreover, we determined the possible mechanism underlying the antiallodynic effect induced by [6]-gingerol in neuropathic rats. The animals underwent L5-L6 SNL for the purpose of developing tactile allodynia. Tactile allodynia was measured with von Frey filaments. Intrathecal administration of [6]-gingerol reversed SNL-induced tactile allodynia. The [6]-gingerol-induced antiallodynic effect was prevented by the intrathecal administration of methiothepin (30 µg per rat; nonselective 5-hydroxytryptamine [5-HT] antagonist), WAY-100635 (6 µg per rat; selective 5-HT1A receptor antagonist), SB-224289 (5 µg per rat; selective 5-HT1B receptor antagonist), BRL-15572 (4 µg per rat; selective 5-HT1D receptor antagonist), and SB-659551 (6 µg per rat; selective 5-HT5A receptor antagonist), but naloxone (50 µg per rat; nonselective opioid receptor antagonist) did not prevent the [6]-gingerol-induced antiallodynic effect. Moreover, intrathecal administration of Nω-nitro-l-arginine methyl ester (100 µg per rat; nonselective nitric oxide [NO] synthase inhibitor), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (10 µg per rat; inhibitor of guanylate cyclase), and glibenclamide (50 µg per rat; channel blocker of adenosine triphosphate [ATP]-sensitive K+ channels) prevented the [6]-gingerol-induced antiallodynic effect. These data suggest that the antiallodynic effect induced by [6]-gingerol is mediated by the serotoninergic system involving the activation of 5-HT1A/1B/1D/5A receptors, as well as the NO-cyclic guanosine monophosphate-ATP-sensitive K+ channel pathway but not by the opioidergic system.

Effects of Schisandra chinensis fruit extract and gomisin A on the contractility of penile corpus cavernosum smooth muscle: a potential mechanism through the nitric oxide - cyclic guanosine monophosphate pathway

BACKGROUND/OBJECTIVES: This study evaluated the effects and molecular mechanisms of the Schisandra chinensis fruit extract (SC) and its major compound gomisin A (GA), on the contractility of rabbit penile corpus cavernosum smooth muscle (PCCSM). MATERIALS/METHODS: PCCSM was exposed to SC or GA after appropriate pretreatment with nitric oxide synthase (NOS) blocker, guanylate cyclase blocker, adenylyl cyclase blocker or protein kinase A blocker. Subsequently, we evaluated the cyclic nucleotide in the perfusate by radioimmunoassay, protein expression level of neuronal NOS (nNOS) and endothelial NOS (eNOS) by western blot, and the interaction of SC or GA with udenafil and rolipram. RESULTS: Both SC and GA induce PCCSM relaxations in a concentration-dependent manner. Pretreatment with NOS blocker, guanylate cyclase blocker, adenylyl cyclase blocker or protein kinase A blocker result in significantly decreased relaxation. SC and GA also induce the levels of cyclic nucleotide in the perfusate in a concentration-dependent manner. Perfusion with GA also showed significantly higher levels of eNOS protein. Furthermore, the udenafil and rolipram induced relaxations of PCCSM were enhanced after exposure to SC and GA. Our results indicate that SC and GA induce the relaxation of PCCSM via the nitric oxide (NO)-cGMP and cAMP signaling pathways. CONCLUSIONS: The SC and GA are potential alternative treatments for men who want to consume natural products to ameliorate erectile function, or who do not respond to the commercially available medicines.

Dietary substitution of fishmeal by alternative protein with guanosine monophosphate supplementation influences growth, digestibility, blood chemistry profile, immunity, and stress resistance of red sea bream, Pagrus major.

We determined the effects of complete fishmeal (FM) replacement by alternative protein (soy protein concentrate, SPC) with guanosine monophosphate (GMP) supplementation on growth, digestibility, immunity, blood chemistry profile, and stress resistance of juvenile red sea bream, Pagrus major. FM protein of a FM-based control diet (FM0) was replaced with 33.3 (FM33.3), 66.6 (FM66.7), and 100% (FM100) by SPC protein, and each replacement group was supplemented with 0.4% GMP to formulate four experimental diets. Each diet was randomly allocated to triplicate groups of fish (4.8 g) for 56 days. Results demonstrated that fish fed diet group FM33.3 had the significantly highest final weight, weight gain-specific growth rate, and feed intake. Meanwhile, in comparison to control, growth performance and feed utilization did not significantly differ with 66.7% FM replacement by SPC with GMP supplementation. Apparent digestibility coefficient of protein and lipid also followed a similar trend. All growth, feed utilization, and digestibility parameters were significantly lower in FM100 diet group. Blood urea nitrogen (BUN) and triglycerides (TG) increased (P < 0.05) with increasing FM replacement level by SPC. Interestingly, total cholesterol level reduces with the increasing level of FM replacement by SPC with GMP supplementation. Fish fed FM0 diet group showed the best condition of both oxidative and freshwater stress resistance. Meanwhile, FM33.3 and FM66.7 diet groups showed acceptable conditions. Innate immune responses enhanced with the increasing FM replacement level by SPC with GMP supplementation. In conclusion, FM could be replaced ≤66.7% by SPC with GMP supplementation in diets for red sea bream without any adverse effects on fish performances.

Guanosine 5'-monophosphate-chelated calcium and iron feed additives maintains egg production and prevents Salmonella Gallinarum in experimentally infected layers.

We evaluated the effects of guanosine 5'-monophosphate (GMP)-chelated calcium and iron (CaFe-GMP) on health and egg quality in layers experimentally infected with Salmonella Gallinarum. In this study, a CaFe-GMP feed additive was added to a commercial layer feed and fed to layers over a four-week period. All were inoculated with Salmonella Gallinarum. Body weight, mortality, clinical symptoms, and poultry production including feed intake, egg production, egg loss, and feed conversion rate were observed, and Salmonella Gallinarum was re-isolated from the liver, spleen, and cecum of the layers. All tested internal organs for the CaFe-GMP additive group exhibited significantly lower re-isolation numbers of Salmonella Gallinarum and less severe pathological changes than those in the control group, indicating that the CaFe-GMP feed supplement induced bacterial clearance and increased resistance to Salmonella Gallinarum. Additionally, due to the inhibitory action of CaFe-GMP on the growth of Salmonella Gallinarum, the CaFe-GMP additive group exhibited better egg production, including a higher laying rate and fewer broken eggs. The results suggest that a 0.16% CaFe-GMP additive may help prevent salmonellosis in the poultry industry.

The Effect of Alcohol Administration on the Corpus Cavernosum

PURPOSE: We studied the effects of alcohol administration on the corpus cavernosum (CC) using an animal model. MATERIALS AND METHODS: CC sections and the aortic ring of rabbits were used in an organ bath study. After acute alcohol administration, changes in blood alcohol concentration and electrical stimulation induced intracavernosal pressure/mean arterial pressure (ICP/MAP) percentage were compared in rats. Cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) levels in the CC were measured using immunoassays. After chronic alcohol administration, ICP/MAP percentage, cAMP and cGMP were compared in rats. Histological changes were examined using the Masson trichrome stain and the Sircol collagen assay. Endothelial nitric oxide synthase (eNOS) expression was examined using immunohistochemistry and Western blotting. RESULTS: Alcohol relaxed the CC in a dose-dependent manner, and the relaxation response was suppressed when pretreated with propranolol, indomethacin, glibenclamide, and 4-aminopyridine. In rats with acute alcohol exposure, the cAMP level in the CC was significantly greater than was observed in the control group (p<0.05). In rats with chronic alcohol exposure, however, changes in cAMP and cGMP levels were insignificant, and the CC showed markedly smaller areas of smooth muscle, greater amounts of dense collagen (p<0.05). Immunohistochemical analysis of eNOS showed a less intense response, and western blotting showed that eNOS expression was significantly lower in this group (p<0.05). CONCLUSIONS: Acute alcohol administration activated the cAMP pathway with positive effects on erectile function. In contrast, chronic alcohol administration changed the ultrastructures of the CC and suppressed eNOS expression, thereby leading to erectile dysfunction.

Penile Erection Induced by Scoparone from Artemisia capillaris through the Nitric Oxide-Cyclic Guanosine Monophosphate Signaling Pathway

PURPOSE: The objective of this study was to evaluate the relaxant effect of scoparone from Artemisia capillaris on rabbit penile corpus cavernosum smooth muscle (PCCSM) and to elucidate the mechanism of action of scoparone for the treatment of erectile dysfunction (ED). MATERIALS AND METHODS: PCCSM that had been precontracted with phenylephrine was treated with 3 Artemisia herbs (A. princeps, A. capillaris, and A. iwayomogi) and 3 fractions (n-hexane, ethyl acetate, and n-butanol) with different concentrations (0.1, 0.5, 1.0, and 2.0 mg/mL). Four components (esculetin, scopoletin, capillarisin, and scoparone) isolated from A. capillaris were also evaluated. The PCCSM was preincubated with Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME) and 1H-[1,2,4] oxadiazolo [4,3-a]quinoxalin-1-one (ODQ). Cyclic nucleotides in the perfusate were measured by a radioimmunoassay. The interactions of scoparone with udenafil and rolipram were also evaluated. RESULTS: A. capillaris extract relaxed PCCSM in a concentration-dependent manner. Scoparone had the highest relaxant effect on PCCSM among the 4 components (esculetin, scopoletin, capillarisin, and scoparone) isolated from the ethyl acetate fraction. The application of scoparone on PCCSM pretreated with L-NAME and ODQ led to significantly less relaxation. Scoparone also increased the cyclic guanosine monophosphate (cGMP) levels in the perfusate in a concentration-dependent manner. Furthermore, scoparone enhanced udenafil- and rolipram-induced relaxation of the PCCSM. CONCLUSIONS: Scoparone relaxed the PCCSM mainly by activating the nitric oxide-cGMP signaling pathway, and it may be a new promising treatment for ED patients who do not completely respond to udenafil.

Effects of dietary administration of guanosine monophosphate on the growth, digestibility, innate immune responses and stress resistance of juvenile red sea bream, Pagrus major.

The present study explored the dietary administration effects of guanosine monophosphate (GMP) on growth, digestibility, innate immune responses and stress resistance of juvenile red sea bream, Pagrus major. A semi-purified basal diet supplemented with 0% (Control), 0.1% (GMP-0.1), 0.2% (GMP-0.2), 0.4% (GMP-0.4) and 0.8% (GMP-0.8) purified GMP to formulate five experimental diets. Each diet was randomly allocated to triplicate groups of fish (mean initial weight 3.4 g) for 56 days. The obtained results clearly indicated that, growth performance of red sea bream enhanced by dietary GMP supplementation compared to control and significantly higher final weight was found in fish fed diet group GMP-0.4. Specific growth rate (SGR) and percent weight gain (%WG) also significantly higher in diet group GMP-0.4 in compared to control and it was not differed (P > 0.05) with diet group GMP-0.8. Feed intake significantly increased with the supplementation of GMP. Feed conversion efficiency (FCE) and protein efficiency ratio (PER) also improved (P < 0.05) when fish fed the diets containing GMP and diet group GMP-0.4 showed the significantly higher value in compared to control. The Apparent digestibility coefficients (dry matter, protein and lipid) also improved by GMP supplementation and the significantly higher protein digestibility was observed in fish fed diet groups GMP-0.2, GMP-0.4 and GMP-0.8. Among the measured non specific immune parameters peroxidase activity (PA), respiratory burst activity (NBT), Bactericidal activity (BA) were significantly affected by dietary supplementation and highest value obtained in diet group GMP-0.4. Total serum protein, lysozyme activity (LA), and agglutination antibody titer also increased (P > 0.05) by GMP supplementation. In contrast, catalase activity decreased with GMP supplementation. In terms of oxidative stress GMP-0.2 showed best condition with low oxidative stress and high antioxidant level. Moreover, the fish fed GMP supplemented diets had better improvement (P < 0.05) in body protein contents, hepatosomatic index, hematocrit content and glutamyl oxaloacetic transaminase (GOT) and glutamic-pyruvate transaminase (GPT) level than the control group. Supplementation also improved (P < 0.05) freshwater stress resistances. Quadratic regression analysis of WG and LA revealed that, the optimal levels of dietary GMP were 0.45 and 0.48%, respectively, for juvenile red sea bream, which is also in line with the most of the growth performance and health parameters of the fish.

A novel multi-biofunctional protein from brown rice hydrolysed by endo/endo-exoproteases.

Brown rice, which is a less allergenic food grain and contains essential amino acids, was hydrolysed by bromelain and PROTEASE FP51® to improve its functionalities and taste for food applications. The hydrolysate prepared by bromelain (eb-RPH) had high protein solubility, surface hydrophobicity, low molecular weight peptides, hydrophobic amino acids (leucine, valine and glycine) and flavor amino acids (glutamic acid and aspartic acid). The eb-RPH exhibited higher 1,1-diphenyl-2-picrylhydrazyl (DPPH˙) and 2,2'-azino-bis 3-ethylbenzthiazoline-6-sulfonic (ABTS˙(+)) radical-scavenging activities of 76.62% and 52.96%, respectively, and possessed a better foaming capacity (221.76%) and emulsifying capacity (32.34%) than the hydrolysate prepared by PROTEASE FP51® (ep-RPH) did. The eb-RPH gave the desired taste, which is attributed to volatile flavor compounds (benzaldehyde, benzeneacetaldehyde and 2-acetyl-1-pyrroline) and non-volatile flavor compounds, such as monosodium glutamate, 5'-guanosine monophosphate and 5'-inosine monophosphate (0.07, 0.03 and 0.05 mg mL(-1), respectively). Brown rice peptides generated by bromelain were novel bioactive peptides with multifunctional properties.

Toxicological effects of pet food ingredients on canine bone marrow-derived mesenchymal stem cells and enterocyte-like cells.

We developed an in vitro method to assess pet food ingredients safety. Canine bone marrow-derived mesenchymal stem cells (BMSC) were differentiated into enterocyte-like cells (ELC) to assess toxicity in cells representing similar patterns of exposure in vivo. The toxicological profile of clove leave oil, eugenol, guanosine monophosphate (GMP), GMP + inosine monophosphate, sorbose, ginger root extract, cinnamon bark oil, cinnamaldehyde, thyme oil, thymol and citric acid was assessed in BMSC and ELC. The LC50 for GMP + inosine monophosphate was 59.42 ± 0.90 and 56.7 ± 3.5 mg ml(-1) for BMSC and ELC; 56.84 ± 0.95 and 53.66 ± 1.36 mg ml(-1) for GMP; 0.02 ± 0.001 and 1.25 ± 0.47 mg ml(-1) for citric acid; 0.077 ± 0.002 and 0.037 ± 0.01 mg ml(-1) for cinnamaldehyde; 0.002 ± 0.0001 and 0.002 ± 0.0008 mg ml(-1) for thymol; 0.080 ± 0.003 and 0.059 ± 0.001 mg ml(-1) for thyme oil; 0.111 ± 0.002 and 0.054 ± 0.01 mg ml(-1) for cinnamon bark oil; 0.119 ± 0.0004 and 0.099 ± 0.011 mg ml(-1) for clove leave oil; 0.04 ± 0.001 and 0.028 ± 0.002 mg ml(-1) for eugenol; 2.80 ± 0.11 and 1.75 ± 0.51 mg ml(-1) for ginger root extract; > 200 and 116.78 ± 7.35 mg ml(-1) for sorbose. Lemon grass oil was evaluated at 0.003-0.9 in BMSC and .03-0.9 mg ml(-1) in ELC and its mechanistic effect was investigated. The gene toxicology studies showed regulation of 61% genes in CYP450 pathway, 37% in cholestasis and 33% in immunotoxicity pathways for BMSC. For ELC, 80% for heat shock response, 69% for beta-oxidation and 65% for mitochondrial energy metabolism. In conclusion, these studies provide a baseline against which differential toxicity of dietary feed ingredients can be assessed in vitro for direct effects on canine cells and demonstrate differential toxicity in differentiated cells that represent gastrointestinal epithelial cells.

Ginsenoside Re Enriched Fraction (GS-F3K1) from Ginseng Berries Ameliorates Ethanol-Induced Erectile Dysfunction via Nitric Oxide-cGMP Pathway

Erectile dysfunction (ED) is a highly prevalent disorder that affects millions of men and considered to be an early symptom of atherosclerosis and a precursor of various systemic vascular disorders. The aim of the present study was to prepare ginsenoside Re enriched fraction (GS-F3K1, ginsenoside Re 10%, w/w) from ginseng berries flesh and to investigate the enhanced activities of GS-F3K1 on alcohol-induced ED. GS-F3K1 was prepared by the continuous liquid and solid separating centrifugation and circulatory ultrafiltration from ginseng berries flesh. GS-F3K1 was administered for 5 weeks in ethanol-induced ED rat by oral administration of 20% ethanol. To investigate the effects of GS-F3K1 on ED model, the levels of nitrite expression, cyclic guanosine monophosphate (cGMP) and erectile response of the penile corpus cavernosum of rat were measured. The erectile response of the corpus cavernosum was restored after GS-F3K1 administration, to a level similar to the normal group. The level of nitrite and cGMP expression in the corpus cavernosum of GS-F3K1-administered male rats was increased significantly compared to positive control group. GS-F3K1 from ginseng berries should effectively restore ethanol-induced ED in male rats and could be developed as a new functional food for the elderly men.