Small Molecule Activation of NAPE-PLD Enhances Efferocytosis by Macrophages.

N-Acyl-phosphatidylethanolamine hydrolyzing phospholipase D (NAPE-PLD) is a zinc metallohydrolase that hydrolyzes N-acyl-phosphatidylethanolamines (NAPEs) to form N-acyl-ethanolamines (NAEs) and phosphatidic acid. Several lines of evidence suggest that reduced NAPE-PLD activity could contribute to cardiometabolic diseases. For instance, NAPEPLD expression is reduced in human coronary arteries with unstable atherosclerotic lesions, defective efferocytosis is implicated in the enlargement of necrotic cores of these lesions, and NAPE-PLD products such as palmitoylethanolamide and oleoylethanolamide have been shown to enhance efferocytosis. Thus, enzyme activation mediated by a small molecule may serve as a therapeutic treatment for cardiometabolic diseases. As a proof-of-concept study, we sought to identify small molecule activators of NAPE-PLD. High-throughput screening followed by hit validation and primary lead optimization studies identified a series of benzothiazole phenylsulfonyl-piperidine carboxamides that variably increased activity of both mouse and human NAPE-PLD. From this set of small molecules, two NAPE-PLD activators (VU534 and VU533) were shown to increase efferocytosis by bone-marrow derived macrophages isolated from wild-type mice, while efferocytosis was significantly reduced in Napepld-/- BMDM or after Nape-pld inhibition. Together, these studies demonstrate an essential role for NAPE-PLD in the regulation of efferocytosis and the potential value of NAPE-PLD activators as a strategy to treat cardiometabolic diseases.

Small Molecule Activation of NAPE-PLD Enhances Efferocytosis by Macrophages.

N -acyl-phosphatidylethanolamine hydrolyzing phospholipase D (NAPE-PLD) is a zinc metallohydrolase that hydrolyzes N -acyl-phosphatidylethanolamine (NAPEs) to form N -acyl-ethanolamides (NAEs) and phosphatidic acid. Several lines of evidence suggest that reduced NAPE-PLD activity could contribute to cardiometabolic diseases. For instance, NAPEPLD expression is reduced in human coronary arteries with unstable atherosclerotic lesions, defective efferocytosis is implicated in the enlargement of necrotic cores of these lesions, and NAPE-PLD products such as palmitoylethanolamide and oleoylethanolamide have been shown to enhance efferocytosis. Thus, enzyme activation mediated by a small molecule may serve as a therapeutic treatment for cardiometabolic diseases. As a proof-of-concept study, we sought to identify small molecule activators of NAPE-PLD. High-throughput screening followed by hit validation and primary lead optimization studies identified a series of benzothiazole phenylsulfonyl-piperidine carboxamides that variably increased activity of both mouse and human NAPE-PLD. From this set of small molecules, two NAPE-PLD activators (VU534 and VU533) were shown to increase efferocytosis by bone-marrow derived macrophages isolated from wild-type mice, while efferocytosis was significantly reduced in Napepld -/- BMDM or after Nape-pld inhibition. Together these studies demonstrate an essential role for NAPE-PLD in the regulation of efferocytosis and the potential value of NAPE-PLD activators as a strategy to treat cardiometabolic diseases.

Melatonin salvages lead-induced neuro-cognitive shutdown, anxiety, and depressive-like symptoms via oxido-inflammatory and cholinergic mechanisms.

INTRODUCTION: Lead is the most used nonphysiological neurotoxic heavy metal in the world that has been indicated to interfere with the cognitive and noncognitive processes via numerous mechanisms. The neuroprotective effect of melatonin is well known, but the effect of its interaction with lead in the brain remains inconclusive. OBJECTIVE: To assess the therapeutic role of melatonin on cognitive deficit, anxiety and depressive-like symptoms in matured male Wistar rats exposed to a subchronic lead chloride (PbCl2 ). METHODS: Twenty male Wistar rats were blindly randomized into four groups (n = 5/group): group 1 to 4 underwent intragastric administration of physiological saline (10 ml/kg; vehicle), PbCl2 (50 mg/kg), melatonin (10 mg/kg) and PbCl2 + melatonin respectively for a period of 4 weeks during which neurobehavioral data were extracted, followed by neurochemical and histopathological evaluations. RESULTS: Exposure to PbCl2 reduced cognitive performance by increasing the escape latency and average proximity to the platform zone border, decreasing average path length in the platform zone, cognitive score, and time spent in probing. It raised the thigmotaxis percentage, time spent in rearing, number of pellet-like feces, and time spent in the dark compartment of a bright/dark box which are predictors of anxiety. It also induced depressive-like behavior as immobility time was enhanced. PbCl2 deranged neurochemicals; malondialdehyde, interlukin-1ß, and tumor necrotic factor-α were increased while superoxide dismutase and acetylcholinesterase were decreased without remarkable alteration in reduced glutathione and nitric oxide. Administration of PbCl2 further disrupted neuronal settings of hippocampal proper and dentate gyrus. In contrast, the supplementation of melatonin reversed all the neurological consequences of PbCl2 neurotoxicity by eliciting its properties against oxidative and nonoxidative action of PbCl2 . CONCLUSION: These findings suggest that melatonin down-regulates neurotoxicant interplays in the brain systems. Therefore, this study suggests the use of melatonin as an adjuvant therapy in neuropathological disorders/dysfunctions.

Telfairia occidentalis stimulates hepatic glycolysis and pyruvate production via insulin-dependent and insulin-independent mechanisms.

BACKGROUND: Telfairia occidentalis (TO), a plant consumed for its nutritional and medicinal values, exhibits hypoglycaemic effect. However, the metabolic fate of the glucose following TO-induced insulin secretion and consequent hypoglycaemia is not clear. OBJECTIVE: This study determined the effect of ethyl acetate and n-hexane fractions of TO leaf extracts on some biochemical parameters in the glucose metabolic pathway to explain the possible fate of blood glucose following TO-induced hypoglycaemia. METHODS: Eighteen male Wistar rats (180-200 g) divided into control, n-hexane TO fraction- and ethyl acetate TO fraction-treated groups (n = 6/group) were used. The control animals received normal saline while the treated groups received TO at 100 mg/kg for seven days. After 24 h following the last dose, the animals were anaesthetised using ketamine; blood samples were collected and livers harvested to determine some biochemical parameters. RESULTS: Ethyl acetate TO fraction significantly increased plasma insulin, liver glucokinase activity and plasma pyruvate concentration, but significantly decreased plasma glucose and liver glycogen, without significant changes in plasma lactate, glucose-6-phosphate, liver glucose-6-phosphatase and lactate dehydrogenase activities when compared with control. N-hexane TO fraction significantly reduced liver glucose-6-phosphatase activity and glycogen but significantly increased plasma pyruvate, without significant changes in plasma glucose, insulin, glucose-6-phosphate and lactate concentrations; and liver glucokinase and lactate dehydrogenase activities. CONCLUSION: The present study showed that insulin-mediated TO-induced hypoglycaemia resulted in the stimulation of glycolysis and pyruvate production via insulin-dependent and insulin-independent mechanisms.

Neurobehavioral, neurochemical and synaptic plasticity perturbations during postnatal life of rats exposed to chloroquine in-utero.

Despite reports that quinoline antimalarials including chloroquine (Chq) exhibit idiosyncratic neuropsychiatric effects even at low doses, the drug continues to be in widespread use during pregnancy. Surprisingly, very few studies have examined the potential neurotoxic action of Chq exposure at different points of gestation or how this phenomenon may affect neurophysiological well-being in later life. We therefore studied behavior, and the expression of specific genes and neurochemicals modulating crucial neural processes in offspring of rats exposed to prophylactic dose of Chq during different stages of gestation. Pregnant rats were injected 5 mg/kg/day (3 times) of Chq either during early- (first week), mid- (second week), late- (third week), or throughout- (all weeks) gestation, while controls received PBS injection. Behavioral characterization of offspring between postnatal days 15-20 in the open field, Y-maze, elevated plus and elevated zero mazes revealed that Chq evoked anxiogenic responses and perturbed spatial memory in rats, although locomotor activity was generally unaltered. In the prefrontal cortex (PFC), hippocampus and cerebellum of rats prenatally exposed to Chq, RT-qPCR analysis revealed decreased mRNA expression of presynaptic marker synaptophysin, which was accompanied by downregulation of postsynaptic marker PSD95. Synaptic marker PICK1 expression was also downregulated in the hippocampus but was unperturbed in the PFC and cerebellum. In addition to recorded SOD downregulation in cortical and hippocampal lysates, induction of oxidative stress in rats prenatally exposed to Chq was corroborated by lipid peroxidation as evinced by increased MDA levels. Offspring of rats infused with Chq at mid-gestation and weekly treatment throughout gestation were particularly susceptible to neurotoxic changes, especially in the hippocampus. Interestingly, Chq did not cause histopathological changes in any of the brain areas. Taken together, our findings causally link intrauterine exposure to Chq with postnatal behavioral impairment and neurotoxic changes in rats.

Frequent exposure to varied home cage sizes alters pain sensitivity and some key inflammation-related biomarkers.

BACKGROUND: Nature and size of rodent cages vary from one laboratory or country to another. Little is however known about the physiological implications of exposure to diverse cage sizes in animal-based experiments. METHOD: Here, two groups of male Swiss mice (Control group - Cage stationed, and Test group - Cage migrated) were used for this study. The cage-migrated mice were exposed daily to various cage sizes used across laboratories in Nigeria while the cage-stationed mice exposed daily to different but the same cage size and shape. At the end of the 30 days exposure, top-rated paradigms were used to profile changes in physiological behaviours, and this was followed by evaluation of histological and biochemical metrics. RESULTS: The study showed a significant (p < 0.05) decrease in blood glucose levels (at 60 and 120 min of oral glucose tolerance test) in the cage-migrated mice compared to cage-stationed mice. Strikingly, peripheral oxidative stress (plasma malondialdehyde) and pain sensitivity (formalin test, hot-and-cold plate test, and von Frey test) decreased significantly in cage-migrated mice compared to cage-stationed animals. Also, the pro-inflammation mediators (IL-6 and NF-κB) increased significantly in cage-migrated mice compared to cage-stationed mice. However, emotion-linked behaviours, neurotransmitters (serotonin, noradrenaline and GABA), brain and plasma electrolytes were not significantly difference in cage-migrated animals compared to cage-stationed mice. CONCLUSION: Taken together, these results suggest that varied size cage-to-cage exposure of experimental mice could affect targeted behavioural and biomolecular parameters of pain and inflammation, thus diminishing research reproducibility, precipitating false negative/positive results and leading to poor translational outcomes.

Curcumin induces secretion of glucagon-like peptide-1 through an oxidation-dependent mechanism.

Curcumin shows antiglycemic effects in animals. Curcumin is chemically unstable at physiological pH, and its oxidative degradation products were shown to contribute to its anti-inflammatory effects. Since the degradation products may also contribute to other effects, we analyzed their role in the antiglycemic activity of curcumin. We quantified curcumin-induced release of glucagon-like peptide 1 (GLP-1) from mouse STC-1 cells that represent enteroendocrine L-cells as a major source of this anti-diabetic hormone. Curcumin induced secretion of GLP-1 in a dose-dependent manner. Two chemically stable analogues of curcumin that do not readily undergo degradation, were less active while two unstable analogues were active secretagogues. Chromatographically isolated spiroepoxide, an unstable oxidative metabolite of curcumin with anti-inflammatory activity, also induced secretion of GLP-1. Stable compounds like the final oxidative metabolite bicyclopentadione, and the major plasma metabolite, curcumin-glucuronide, were inactive. GLP-1 secretion induced by curcumin and its oxidative degradation products was associated with activation of PKC, ERK, and CaM kinase II. Since activity largely correlated with instability of curcumin and the analogues, we tested the extent of covalent binding to proteins in STC-1 cells and found it occurred with similar affinity as N-ethylmaleimide, indicating covalent binding occurred with nucleophilic cysteine residues. These results suggest that oxidative metabolites of curcumin are involved in the antiglycemic effects of curcumin. Our findings support the hypothesis that curcumin functions as a pro-drug requiring oxidative activation to reveal its bioactive metabolites that act by binding to target proteins thereby causing a change in function.

Oral Ingestion of Cannabis sativa: Risks, Benefits, and Effects on Malaria-Infected Hosts.

Background: The emergence of a multidrug-resistant strain of Plasmodium falciparum (Pf Pailin) raises concern about malaria control strategies. Unfortunately, the role(s) of natural plants/remedies in curtailing malaria catastrophe remains uncertain. The claims of potential antimalarial activity of Cannabis sativa in vivo have not been well established nor the consequences defined. This study was, therefore, designed to evaluate the effects of whole cannabis consumption on malaria-infected host. Methods: Thirty mice were inoculated with dose of 1×107 chloroquine-resistant Plasmodium berghei ANKA-infected erythrocyte and divided into six treatment groups. Cannabis diet formulations were prepared based on weighted percentages of dried cannabis and standard mice diet and the study animals were fed ad libitum. Chemosuppression of parasitemia, survival rates, parasite clearance, and recrudescence time were evaluated. Histopathological studies were performed on the prefrontal cortex (PFC) and hippocampus of the animals after 14 days' consumption of cannabis diet formulation by naive mice. Results: There was a significant difference (p<0.05) in the day-4 chemosuppression of parasitemia between the animals that were fed C. sativa and chloroquine relative to the untreated controls. There was also a significant difference in the survival rate (p<0.05) of animals fed C. sativa diet (40%, 20%, 10%, and 1%) in contrast to control animals on standard mice diet. A parasite clearance time of 2.18±0.4 was recorded in the chloroquine treatment group, whereas recrudescence in chloroquine group occurred on day 7. There were slight histomorphological changes in the PFC and cell densities of the dentate gyrus of the hippocampus of animals that were fed C. sativa. Conclusions: C. sativa displayed mild antimalarial activity in vivo. There was evident reduction in symptomatic manifestation of malaria disease, though unrelated to levels of parasitemia. This disease tolerance status may be beneficial, but may also constitute a transmission burden through asymptomatic carriage of parasites by habitual cannabis users.