A High-Methionine Diet for One-Week Induces a High Accumulation of Methionine in the Cerebrospinal Fluid and Confers Bipolar Disorder-like Behavior in Mice

Methionine (Met) is considered the most toxic amino acid in mammals. Here, we investigated biochemical and behavioral impacts of ad libitum one-week feeding of high-Met diets on mice. Adult male mice were fed the standard rodent diet that contained 0.44% Met (1×) or a diet containing 16 graded Met doses (1.2×–13×). High-Met diets for one-week induced a dose-dependent decrease in body weight and an increase in serum Met levels with a 2.55 mM peak (versus basal 53 µM) on the 12×Met diet. Total homocysteine (Hcy) levels were also upregulated while concentrations of other amino acids were almost maintained in serum. Similarly, levels of Met and Hcy (but not the other amino acids) were highly elevated in the cerebrospinal fluids of mice on the 10×Met diet; the Met levels were much higher than Hcy and the others. In a series of behavioral tests, mice on the 10×Met diet displayed increased anxiety and decreased traveled distances in an open-field test, increased activity to escape from water soaking and tail hanging, and normal learning/memory activity in a Y-maze test, which were reflections of negative/positive symptoms and normal cognitive function, respectively. These results indicate that high-Met ad libitum feeding even for a week can induce bipolar disorder-like disease models in mice.

3-Hydroxyphenylacetic Acid: A Blood Pressure-Reducing Flavonoid Metabolite

Regular intake of polyphenol-rich food has been associated with a wide variety of beneficial health effects, including the prevention of cardiovascular diseases. However, the parent flavonoids have mostly low bioavailability and, hence, their metabolites have been hypothesized to be bioactive. One of these metabolites, 3-hydroxyphenylacetic acid (3-HPAA), formed by the gut microbiota, was previously reported to exert vasorelaxant effects ex vivo. The aim of this study was to shed more light on this effect in vivo, and to elucidate the mechanism of action. 3-HPAA gave rise to a dose-dependent decrease in arterial blood pressure when administered i.v. both as a bolus and infusion to spontaneously hypertensive rats. In contrast, no significant changes in heart rate were observed. In ex vivo experiments, where porcine hearts from a slaughterhouse were used to decrease the need for laboratory animals, 3-HPAA relaxed precontracted porcine coronary artery segments via a mechanism partially dependent on endothelium integrity. This relaxation was significantly impaired after endothelial nitric oxide synthase inhibition. In contrast, the blockade of SKCa or IKCa channels, or muscarinic receptors, did not affect 3-HPAA relaxation. Similarly, no effects of 3-HPAA on cyclooxygenase nor L-type calcium channels were observed. Thus, 3-HPAA decreases blood pressure in vivo via vessel relaxation, and this mechanism might be based on the release of nitric oxide by the endothelial layer.

Insights into Cytotoxic Behavior of Lepadins and Structure Elucidation of the New Alkaloid Lepadin L from the Mediterranean Ascidian Clavelina lepadiformis

The chemical investigation of the Mediterranean ascidian Clavelina lepadiformis has led to the isolation of a new lepadin, named lepadin L, and two known metabolites belonging to the same family, lepadins A and B. The planar structure and relative configuration of the decahydroquinoline ring of lepadin L were established both by means of HR-ESIMS and by a detailed as extensive analysis of 1D and 2D NMR spectra. Moreover, microscale derivatization of the new alkaloid lepadin L was performed to assess the relative configuration of the functionalized alkyl side chain. Lepadins A, B, and L were tested for their cytotoxic activity on a panel of cancer cell lines (human melanoma [A375], human breast [MDA-MB-468], human colon adenocarcinoma [HT29], human colorectal carcinoma [HCT116], and mouse myoblast [C2C12]). Interestingly, a deeper investigation into the mechanism of action of the most cytotoxic metabolite, lepadin A, on the A375 cells has highlighted its ability to induce a strongly inhibition of cell migration, G2/M phase cell cycle arrest and a dose-dependent decrease of cell clonogenity, suggesting that it is able to impair self-renewing capacity of A375 cells.

Low Doses of Imidacloprid Induce Oxidative Stress and Neural Cell Disruption in Earthworm Eisenia fetida

Imidacloprid is a widely used pesticide that belongs to the class of neonicotinoids. There is a piece of rising evidence that neonicotinoids exert cytotoxic effects in non-target organisms including vertebrate species such as mammals. Nevertheless, dose-limiting toxicity and molecular mechanisms of neonicotinoids' deleterious effects are still poorly understood. In accord to imidacloprid fate in the environment, the most of used pesticide is absorbed in the soil. Therefore, earthworms, which are prevailing soil organisms, could be considered as a target of neonicotinoids toxicity. The earthworm’s simple nervous system is a prospective model for neurotoxicological studies. We exposed earthworms to imidacloprid in a paper contact test with a doses range of 0.1‑0.4 µg/cm2 for 14 days. In the present work, we studied the imidacloprid effect on oxidative stress generation and neuronal marker neuron-specific enolase (NSE) expression. The exposure to imidacloprid induced a dose-dependent decrease in NSE. Both reactive oxygen species production and lipid peroxidation level were upregulated as well. Observed NSE decline suggests imidacloprid-caused disturbance in earthworm neuron cells. Obtained data have shown that relatively low doses of imidacloprid are potent to induce cytotoxicity in neurons. Furthermore, neurotoxicity could be recognized as one of an individual scenario of the general imidacloprid toxicity. Thus, presented results suggest the cytotoxicity of imidacloprid low doses in non-target organisms and hypothesize that NSE downregulation could be estimated as a biomarker of neonicotinoid cytotoxicity in a nervous system of non-insect species.

Beclometasone but not fluticasone modulates PDGFD expression in the H295R adrenal cell line.

BackgroundAdrenal suppression is a clinically concerning side effect of inhaled corticosteroid (ICS) treatment in patients with asthma. Increased susceptibility to ICS-induced adrenal suppression has previously been identified in those with the rs591118 polymorphism in Platelet Derived Growth Factor D (PDGFD). The mechanism underpinning this relationship is not known.MethodsH295R cells were genotyped for rs591118 using a validated Taqman PCR allelic discrimination assay. H295R cell viability was determined after treatment with beclometasone and fluticasone (range 0-330 μM). Cortisol was measured in cell culture medium using competitive enzyme immunoassay.ResultsPDGFD protein expression in H295R cells was confirmed using Western blotting. When ACTH and forskolin were added to H295R cells, a reduction in PDGFD expression was seen which was then restored by incubation with prochloraz, a known inhibitor of steroidogenesis.A dose-dependent, decrease in PDGFD expression was observed with beclometasone (over a 24 h incubation period) but not with beclometasone incubations beyond 24 hour nor with fluticasone (at 24 or 48 hours).ConclusionsH295R cells express PDGFD protein which can be modulated by incubation with steroidogenesis agonists and antagonists and additionally with exogenous beclometasone.

The Identification of a Novel Calcium-Dependent Link Between NAD+ and Glucose Deprivation-Induced Increases in Protein O-GlcNAcylation and ER Stress

The modification of proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) is associated with the regulation of numerous cellular processes. Despite the importance of O-GlcNAc in mediating cellular function our understanding of the mechanisms that regulate O-GlcNAc levels is limited. One factor known to regulate protein O-GlcNAc levels is nutrient availability; however, the fact that nutrient deficient states such as ischemia increase O-GlcNAc levels suggests that other factors also contribute to regulating O-GlcNAc levels. We have previously reported that in unstressed cardiomyocytes exogenous NAD+ resulted in a time and dose dependent decrease in O-GlcNAc levels. Therefore, we postulated that NAD+ and cellular O-GlcNAc levels may be coordinately regulated. Using glucose deprivation as a model system in an immortalized human ventricular cell line, we examined the influence of extracellular NAD+ on cellular O-GlcNAc levels and ER stress in the presence and absence of glucose. We found that NAD+ completely blocked the increase in O-GlcNAc induced by glucose deprivation and suppressed the activation of ER stress. The NAD+ metabolite cyclic ADP-ribose (cADPR) had similar effects on O-GlcNAc and ER stress suggesting a common underlying mechanism. cADPR is a ryanodine receptor (RyR) agonist and like caffeine, which also activates the RyR, both mimicked the effects of NAD+. SERCA inhibition, which also reduces ER/SR Ca2+ levels had similar effects to both NAD+ and cADPR on O-GlcNAc and ER stress responses to glucose deprivation. The observation that NAD+, cADPR, and caffeine all attenuated the increase in O-GlcNAc and ER stress in response to glucose deprivation, suggests a potential common mechanism, linked to ER/SR Ca2+ levels, underlying their activation. Moreover, we showed that TRPM2, a plasma membrane cation channel was necessary for the cellular responses to glucose deprivation. Collectively, these findings support a novel Ca2+-dependent mechanism underlying glucose deprivation induced increase in O-GlcNAc and ER stress.

Penthorum chinense Pursh alleviates ethanol-induced hepatic oxidative impairment in zebrafish via AMPK / p62 / Nrf2 / mTOR pathway

Ethnopharmacological relevance: In China, Penthorum chinense Pursh (PCP) is renowned for its effectiveness in “promoting blood circulation” and “removing blood stasis”. It can “relieve the liver” and its application in the field of liver protection, including viral hepatitis, alcoholic liver, liver fibrosis, has been known for hundreds of years.Aim of the study: Oxidative stress is widely believed to exert a key role in the pathophysiology of alcoholic liver disease (ALD). Therefore, antioxidant therapy reflects a reasonable strategy for the prevention and treatment of ALD. Hence, this study aimed to elucidate the mechanism of PCP in ethanol-induced liver injury.Methods: Treatment of liver-specific transgenic zebrafish larvae (lfabp: EGFP) at three days post-fertilization (3 dpf) with different concentrations of PCP (100, 50, 25 μg / mL) for 48 h was followed by soaking in 350 mmol / L ethanol for 32 h. Liver function and fat accumulation were identified by phenotypic indicators and biochemical kits. The related proteins and gene expression were further estimated by western blotting and quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Finally, high performance liquid chromatography (HPLC) was adopted to analyze the chemical composition of PCP extract.Results: Firstly, PCP mediated alleviation of ethanol-induced steatosis and reduction of aspartate aminotransferase (AST), alanine transaminase (ALT), total cholesterol (TC) and triglyceride (TG) related indexes were evident. Dose-dependent decrease of intracellular reactive oxygen species (ROS) production, the activity of malondialdehyde (MDA) and increased the activity of glutathione (GSH), Superoxide dismutase (SOD) and catalase (CAT) in zebrafish substantiated the role of PCP in relieving oxidative stress. Furthermore, PCP induced downregulation of sequestosome 1 (p62 / SQSTM1, p62), Atg13 and Beclin 1 expression promoted autophagy. Meanwhile, PCP contributed to the hepatoprotective function by downregulating the expression of kelch-like ECH-associated protein 1 (Keap1) and upregulating the expression of nucleus factor-E2-related factor 2 (Nrf2), which activated cytoprotective related gene HO-1. Moreover, HPLC of PCP extract confirmed the presence of various polyphenols with potential antioxidant effects. Finally, PCP appeared to promote the activated protein kinase (AMPK) / p62 / Nrf2 / mTOR signaling pathways, which were related to oxidative stress and autophagy in zebrafish.Conclusion: This study claimed that by activating the AMPK / p62 / Nrf2 / mTOR signaling pathway, PCP could attenuate ethanol-induced liver injury in zebrafish.

Evaluation of Anxiolytic potential of various extracts of Caesalpinia pulcherrima leaves

Caesalpinia pulcherrima Swartz native to India referred to as Guletura is widely distributed in South India, and its leaves, flower, bark, and seeds are employed in Indian medicine. The plant contains many active elemental fractions like caesalpin-type diterpenoids, sitosterol, pulcherrimin, lupeol, lupeol acetate, myricetin, quercetin and rutin, flavonoids, carotenoids, glycosides, peltogynoids, phenols, and steroids. The current study was designed to gauge the anti-anxiety activity of varied extracts viz n-hexane, chloroform, ethyl acetate, and methanol of the leaves of Caesalpinia pulcherrima by using elevated plus maze (EPM) model in albino mice. Albino mice have ministered orally with different doses of the extracts (i.e. 200 and 400mg/kg) and behavior was observed on the EPM. Diazepam (2mg/kg, P.O) was used as a standard (positive control). Results indicate that the methanol extract of Caesalpinia leaves showed maximum and significant dose-dependent effect at 200 and 400mg/kg on EPM, the results were just like the standard antianxiety agent diazepam (2mg/kg). In the Actophoptometer model, two different doses of Caesalpinia pulcherrima (200 and 400mg/kg) showed a dose-dependent decrease within the locomotor activity, compared to the control animals. As the phytochemical screening of methanol extract exhibited the presence of polyphenols could be liable for the anxiolytic potential of C.pulcherrima. Hence this plant could also be developed as a potentially useful anti-anxiety agent.

Acomparative Study between Epidural Analgesia of Levobupivacaine with Fentanyl versus Dexmedetomidine for knee replacement surgeries

Background The use of epidural analgesia for the management of postoperative pain has evolved as a critical component of multimodal approach to achieve the goal of adequate analgesia with improved outcome. Epidural analgesia offers superior postoperative pain relief compared with systemic opioids. In addition to improved patient outcome. Objective To compare the effect of dexmedetomidine and fentanyl as an adjuvant to epidural levobupivacaine in knee replacement surgeries regarding duration of action and the analgesic potency of both drugs. Patients and Methods This prospective double blinded randomized clinical trial study was conducted in Maadi Military and Ain Shams University Hospitals after approval of the anesthesia department and the local ethics and research committee over 6 months and after obtaining a written informed consent. Sixty patients underwent knee replacement surgeries were included in the study their ages range between 21 and 60 years old and classified as ASA I and II. The patients were randomly divided using computer generated randomization into two groups 30 patients in each (n = 30). Results As regards sedation score intraoperative and postoperative, in the present study, we found that Ramsay sedation score was significantly higher in group ‘BD’ when compared to group ‘BF’ intraoperative and postoperative. This finding may be attributed to the sedative properties of dexmedetomidine that is far superior to fentanyl acting by dose-dependent decrease in activity of noradrenergic neurons in the brain stem via post-synaptic receptor-mediated inhibition. This increases gamma-aminobutyric acid (GABA) neurone activity, which mediates central sedative effects. Conclusion Epidural levobupivacaine with dexmedetomidine provided better sedation, adequate surgical anesthesia with prolonged postoperative analgesia for lower limb surgeries. Both adjuvants reduced the epidural dose of levobupivacaine and potentiated its efficacy.

Rutin Exerts Cytotoxic and Senescence-Inducing Properties in Human Melanoma Cells

Malignant melanoma represents the deadliest type of skin cancer with narrow treatment options in advanced stages. Herbal constituents possessing anticancer properties occupy a particular spot in melanoma research as potential chemotherapeutics. Rutin (RUT) is a natural compound exerting antioxidant, antimicrobial, anti-inflammatory, UV-filtering, and SPF-enhancing activities that are beneficial to the skin; however, its effect as an anti-melanoma agent is less investigated. The current study is focused on assessing the cytotoxic potential of RUT against two different human melanoma cell lines: RPMI-7951 and SK-MEL-28 by evaluating its impact in terms of cell viability, cells’ morphology, and nuclear aspect assessment, and senescence-inducing properties. The results indicate a dose-dependent decrease in the viability of both cell lines, with calculated IC50 values of 64.49 ± 13.27 µM for RPMI-7951 cells and 47.44 ± 2.41 µM for SK-MEL-28, respectively, accompanied by a visible reduction in the cell confluency and apoptotic features within the cell nuclei. RUT exerted a senescence-inducing property highlighted by the elevated expression of senescent-associated beta-galactosidase (SA-β-gal) in SK-MEL-28 cells. Despite the in vitro anti-melanoma effect revealed by our results, further studies are required to elucidate the mechanisms of RUT-induced cytotoxicity and senescence in melanoma cells.