Integrating Molecular Networking and 1H NMR To Target the Isolation of Chrysogeamides from a Library of Marine-Derived Penicillium Fungi.

A challenging problem in natural product discovery is to rapidly dereplicate known compounds and expose novel ones from complicated components. Herein, integrating the LC-MS/MS-dependent molecular networking and 1H NMR techniques efficiently and successfully enabled the targeted identification of seven new cyclohexadepsipeptides, chrysogeamides A-G (1-7), from the coral-derived fungus Penicillium chrysogenum (CHNSCLM-0003) which was targeted from a library of marine-derived Penicillium fungi. Compound 4 features a rare 3-hydroxy-4-methylhexanoic acid (HMHA) moiety which was first discovered from marine-derived organisms. Interestingly, isotope-labeling feeding experiments confirmed that 13C1-l-Leu was transformed into 13C1-d-Leu moiety, indicating that d-Leu could be isomerized from l-Leu. Compounds 1 and 2 obviously promoted angiogenesis in zebrafish at 1.0 µg/mL with nontoxic to embryonic zebrafish at 100 µg/mL. Combining molecular networking with 1H NMR as a discovery tool will be implemented as a systematic strategy, not only for known compounds dereplication but also for untapped reservoir discovery.

Schisantherin A attenuates ischemia/reperfusion-induced neuronal injury in rats via regulation of TLR4 and C5aR1 signaling pathways.

Toll-like receptor 4 (TLR4) and C5aR1 (CD88) have been recognized as potential therapeutic targets for the reduction of inflammation and secondary damage and improvement of outcome after ischemia and reperfusion (I/R). The inflammatory responses which induce cell apoptosis and necrosis after I/R brain injury lead to a limited process of neural repair. To further comprehend how these targets function in I/R state, we investigated the pathological changes and TLR4 and C5aR1 signaling pathways in vitro and in vivo models of I/R brain injury in this study. Meanwhile, we explored the roles of schisantherin A on I/R brain injury, and whether it exerted neuroprotective effects by regulating the TLR4 and C5aR1 signaling pathways or not. The results showed that schisantherin A significantly reduced the neuronal apoptosis induced by oxygen and glucose deprivation and reperfusion (OGD/R) injury in primary culture of rat cortical neurons. Also, schisantherin A alleviated neurological deficits, reduced infarct volume, attenuated oxidation stress, inflammation and apoptosis in ischemic parietal cortex of rats after middle cerebral artery occlusion and reperfusion (MCAO/R) injury. Moreover, the activated TLR4 and C5aR1 signaling pathways were inhibited by schisantherin A treatment. In conclusion, TLR4 and C5aR1 played a vital role during I/R brain injury in rats, and schisantherin A exhibited neuroprotective effects by TLR4 and C5aR1 signaling pathways. These findings also provided new insights that would aid in elucidating the effect of schisantherin A against cerebral I/R and support the development of schisantherin A as a potential treatment for ischemic stroke.

Neuroprotective Effects of Loganin on MPTP-Induced Parkinson's Disease Mice: Neurochemistry, Glial Reaction and Autophagy Studies.

Parkinson's disease (PD) is a progressive neurodegenerative disease, involving resting tremor and bradykinesia, for which no recognized therapies or drugs are available to halt or slow progression. In recent years, natural botanic products have been considered relatively safe, with limited side effects, and are expected to become an important source for clinical mediation of PD in the future. Our study focuses on the ability of loganin, a compound derived from fruits of cornus, to mediate neuroprotection in a mouse model of PD. Mice were administered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) with a dosage of 30 mg/kg daily for 5 days to establish a subacute PD model and treated with loganin. Locomotor activity was assessed by a pole test, then mice were euthanized at 1 and 3 days after the last treatment, and brain tissue was prepared for subsequent assays. Loganin rescued decrease of dopamine levels and tyrosine hydroxylase (TH) expression in the striatum, and shortened total locomotor activity (TLA) time of mice. Furthermore, loganin alleviated microglia and astrocyte activation, and suppressed TNF-α and caspase-3 expression through a c-Abl-p38-NFκB pathway. Loganin also downregulated LC3-II and Drp1 expression, and decreased the level of acidic vesicular organelles (AVOs). Loganin exerts neuroprotective effects on MPTP-induced PD mice by decreasing inflammation, autophagy, and apoptosis, suggesting that loganin could serve as a therapeutic drug to ameliorate PD. J. Cell. Biochem. 118: 3495-3510, 2017. © 2017 Wiley Periodicals, Inc.

Isoquercetin Ameliorates Cerebral Impairment in Focal Ischemia Through Anti-Oxidative, Anti-Inflammatory, and Anti-Apoptotic Effects in Primary Culture of Rat Hippocampal Neurons and Hippocampal CA1 Region of Rats.

Ischemic stroke is a major disability and cause of death worldwide due to its narrow therapeutic time window. Neuroprotective agent is a promising strategy to salvage acutely ischemic brain tissue and extend the therapeutic time window for stroke treatment. In this study, we aimed to evaluate the neuroprotective effects of isoquercetin in (1) primary culture of rat hippocampal neurons exposure on oxygen and glucose deprivation and reperfusion (OGD/R) injury and (2) rats subjected to transient middle cerebral artery occlusion and reperfusion (MCAO/R) injury. The results showed that isoquercetin post-treatment reduced the infarct size, number of apoptotic cells, oxidative stress, and inflammatory response after ischemia and reperfusion injury. The underlying mechanism study indicated that the neuroprotective effects of isoquercetin were elicited via suppressing the activation of toll-like receptor 4 (TLR4), nuclear factor-kappa B (NF-κB) and caspase-1; the phosphorylation of ERK1/2, JNK1/2, and p38 mitogen-activated protein kinase (MAPK); and the secretion of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6. In addition, isoquercetin also effectively alleviated hippocampus neuron apoptosis by regulation of cyclic AMP responsive element-binding protein (CREB), Bax, Bcl-2, and caspase-3. Our report provided new considerations into the therapeutic action and the underlying mechanisms of isoquercetin to improve brain injury in individuals who have suffered from ischemic stroke. As a potent anti-inflammatory and anti-oxidative compound with neuroprotective capacities, the beneficial effects of isoquercetin when used to treat ischemic stroke and related diseases in humans warrant further studies.

Combretastatin A-4 efficiently inhibits angiogenesis and induces neuronal apoptosis in zebrafish.

Cis-stilbene combretastatin A-4 (CA-4) and a large group of its derivant compounds have been shown significant anti-angiogenesis activity. However the side effects even the toxicities of these chemicals were not evaluated adequately. The zebrafish model has become an important vertebrate model for evaluating drug effects. The testing of CA-4 on zebrafish is so far lacking and assessment of CA-4 on this model will provide with new insights of understanding the function of CA-4 on angiogenesis, the toxicities and side effects of CA-4. We discovered that 7-9 ng/ml CA-4 treatments resulted in developmental retardation and morphological malformation, and led to potent angiogenic defects in zebrafish embryos. Next, we demonstrated that intraperitoneal injection of 5, 10 and 20 mg/kg CA-4 obviously inhibited vessel plexus formation in regenerated pectoral fins of adult zebrafish. Interestingly, we proved that CA-4 treatment induced significant cell apoptosis in central nervous system of zebrafish embryos and adults. Furthermore, it was demonstrated that the neuronal apoptosis induced by CA-4 treatment was alleviated in p53 mutants. In addition, notch1a was up-regulated in CA-4 treated embryos, and inhibition of Notch signaling by DAPT partially rescued the apoptosis in zebrafish central nervous system caused by CA-4.

Neuroprotective effect of schizandrin A on oxygen and glucose deprivation/reperfusion-induced cell injury in primary culture of rat cortical neurons

Brain ischemia appears to be associated with innate immunity. Recent reports showed that C3a and C5a, as potent targets, might protect against ischemia induced cell death. In traditional Chinese medicine, the fruit of Schizandra chinesis Baill (Fructus schizandrae) has been widely used as a tonic. In the present study, we sought to evaluate the neuroprotective effects of schizandrin A, a composition of S. chinesis Baill, against oxygen and glucose deprivation followed by reperfusion (OGD/R)-induced cell death in primary culture of rat cortical neurons, and to test whether C3a and C5a affected cortical neuron recovery from ischemic injury after schizandrin A treatment. The results showed that schizandrin A significantly reduced cell apoptosis and necrosis, increased cell survival, and decreased intracellular calcium concentration ([Ca2+]i) and lactate dehydrogenase (LDH) release in primary culture of rat cortical neurons after OGD/R. Mechanism studies suggested that the modulation of extracellular-regulated kinase (ERK), c-Jun NH2-terminal kinases (JNK), and p38, as well as caspase-3 activity played an important role on the progress of neuronal apoptosis. C5aR participated in the neuroprotective effect of schizandrin A in primary culture of rat cortical neurons after OGD/R. Our findings suggested that schizandrin A might act as a candidate therapeutic target drug used for brain ischemia and related diseases

Neuroprotective effect of schizandrin A on oxygen and glucose deprivation/reperfusion-induced cell injury in primary culture of rat cortical neurons.

Brain ischemia appears to be associated with innate immunity. Recent reports showed that C3a and C5a, as potent targets, might protect against ischemia induced cell death. In traditional Chinese medicine, the fruit of Schizandra chinesis Baill (Fructus schizandrae) has been widely used as a tonic. In the present study, we sought to evaluate the neuroprotective effects of schizandrin A, a composition of S. chinesis Baill, against oxygen and glucose deprivation followed by reperfusion (OGD/R)-induced cell death in primary culture of rat cortical neurons, and to test whether C3a and C5a affected cortical neuron recovery from ischemic injury after schizandrin A treatment. The results showed that schizandrin A significantly reduced cell apoptosis and necrosis, increased cell survival, and decreased intracellular calcium concentration ([Ca(2+)]i) and lactate dehydrogenase (LDH) release in primary culture of rat cortical neurons after OGD/R. Mechanism studies suggested that the modulation of extracellular-regulated kinase (ERK), c-Jun NH2-terminal kinases (JNK), and p38, as well as caspase-3 activity played an important role on the progress of neuronal apoptosis. C5aR participated in the neuroprotective effect of schizandrin A in primary culture of rat cortical neurons after OGD/R. Our findings suggested that schizandrin A might act as a candidate therapeutic target drug used for brain ischemia and related diseases.

Mulberroside A protects against ischemic impairment in primary culture of rat cortical neurons after oxygen-glucose deprivation followed by reperfusion.

Mulberroside A is a natural polyhydroxylated stilbene compound present at relatively high abundance in the roots and twigs of Morus alba L. It is known for its nephroprotective, hypoglycemic, and antidiabetic effects. Because its metabolite, oxyresveratrol, possessed purported anti-inflammatory and neuroprotective effects, we proposed that mulberroside A may elicit neuroprotective effects that can be used in the treatment of brain ischemic injury. Therefore, we decided to investigate the pharmacological properties of mulberroside A in primary culture of rat cortical neurons after oxygen-glucose deprivation followed by reperfusion (OGD/R), evaluating its ability to counteract the hypoxia-ischemia impairment. The results showed that mulberroside A elicited neuroprotective effects comparable to nimodipine. The mechanistic studies showed that mulberroside A decreased the expressions of tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, and IL-6 and inhibited the activation of NALP3, caspase-1, and nuclear factor-κB and the phosphorylation of extracellular signal-regulated protein kinases, the c-Jun N-terminal kinase, and p38, exhibiting anti-inflammatory antiapoptotic effects. Our results also further demonstrate that the proinflammatory cytokines of IL-1ß, IL-6, and TNF-α are promising targets for treatment of cerebral ischemic injury. Although further investigation is required for its development, all of these findings led us to speculate that mulberroside A is a candidate for the treatment of ischemic stroke, which would act as a multifactorial neuroprotectant.

Morin reduces hepatic inflammation-associated lipid accumulation in high fructose-fed rats via inhibiting sphingosine kinase 1/sphingosine 1-phosphate signaling pathway.

SphK1/S1P signaling pathway is involved in the development of hepatic inflammation and injury. But its role in high fructose-induced NAFLD has not yet been reported. The aim of this study was to elucidate the crucial role of SphK1/S1P signaling pathway in high fructose-induced hepatic inflammation and lipid accumulation in rats. Moreover, the hepatoprotective effects of morin, a flavonoid with anti-inflammatory and anti-hyperlipedimic activities, on these hepatic changes in rats were investigated. High fructose-fed rats were orally treated with morin (30 and 60mg/kg) and pioglitazone (4mg/kg) for 8 weeks, respectively. Fructose feeding induced hyperlipidemia, and activated SphK1/S1P signaling pathway characterized by the elevation of SphK1 activity, S1P production as well as SphK1, S1PR1 and S1PR3 protein levels, which in turn caused NF-κB signaling activation to produce IL-1ß, IL-6 and TNF-α and inflammation in the liver of rats. Subsequently, hepatic insulin and leptin signaling impairment and lipid metabolic disorder were observed in this animal model, resulting in liver lipid accumulation. Morin restored high fructose-induced the activation of hepatic SphK1/S1P signaling pathway in rats. Subsequently, the reduced NF-κB signaling activation by morin decreased inflammatory cytokine production, recovered insulin and leptin signaling impairment to reduce lipid accumulation and injury in the rat liver. These effects of morin were confirmed in Buffalo rat liver (BRL3A) cell model stimulated with 5mM fructose. Thus, the inhibition of hepatic SphK1/S1P signaling pathway may be a novel mechanism by which morin exerts hepatoprotection in high fructose-fed rats, possibly involving liver inflammation inhibition and lipid accumulation recovery.

Uricosuric and nephroprotective properties of Ramulus Mori ethanol extract in hyperuricemic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Ramulus Mori, the branch of Morus alba, is widely used in traditional Chinese medicine prescriptions to treat gout and hyperuricemia. AIM OF THIS STUDY: To evaluate the uricosuric and nephroprotective effects of ethanol extract of Ramulus Mori (ERM) and explore its possible mechanisms in hyperuricemic mice. MATERIALS AND METHODS: HPLC analysis was employed to determine the main constituents. Hyperuricemia was induced by potassium oxonate (250 mg/kg) in male mice. ERM (10, 20 and 40 mg/kg) was orally administered to hyperuricemic and normal mice for 7 days. Serum and urine levels of uric acid, creatinine and blood urea nitrogen (BUN) were measured. Simultaneously, renal mRNA and protein levels of mouse urate transporter 1 (mURAT1), glucose transporter 9 (mGLUT9), organic anion transporter 1 (mOAT1) and organic cation/carnitine transporters (mOCT1/2, mOCTN1/2) were analyzed by RT-PCR and Western blotting methods. RESULTS: ERM mainly contained mulberroside A, oxyresveratrol, 4-hydroxycinnamic acid, resveratrol, 7-hydroxycumarin and morin. ERM significantly reduced serum urate levels and increased 24h-urine urate excretion and fractional excretion of uric acid in hyperuricemic mice. It effectively restored oxonate-induced expression alteration of renal mURAT1, mGLUT9 and mOAT1, resulting in urate excretion enhancement. Moreover, ERM decreased serum creatinine and BUN levels and increased creatinine clearance, and up-regulated expression of mOCT1/2 and mOCTN1/2, contributing to kidney function improvement in this model. CONCLUSION: These results suggest that ERM exerts the uricosuric and nephroprotective actions by the regulation of these renal organic ion transporters in hyperuricemic mice, and provide scientific support for the empirical use of Ramulus Mori.

Antihyperuricemic and nephroprotective effects of resveratrol and its analogues in hyperuricemic mice.

SCOPE: Stilbenes, of which, resveratrol is a representative compound in foods and plants, possess a variety of bioactivities including antioxidation, anti-inflammation, chemoprevention, and cardioprotection. This study was conducted to evaluate the antihyperuricemic and nephroprotective effects of resveratrol and its analogues and explore the possible mechanisms. The structure-activity relationships were analyzed. METHODS AND RESULTS: Potassium oxonate-induced hyperuricemic mice were dosed by gavage with eight stilbenes. Uric acid, creatinine, and blood urea nitrogen (BUN) levels in serum and urine, clearance rate of creatinine and BUN, 24-h urate excretion, and fractional excretion of uric acid, uromodulin levels in urine and kidney were determined to evaluate renal urate handling and function. Renal protein levels of organic ion transporters were detected to elucidate the possible mechanisms. Resveratrol, trans-4-hydroxystilbene, pterostilbene, polydatin, and mulberroside A were found to have antihyperuricemic activities. These compounds together with trans-2-hydroxystilbene provided nephroprotection. Trans-3,4',5-trimethoxystilbene and cis-combretastatin A-4 had no effects. CONCLUSION: The uricosuric and nephroprotective actions of resveratrol and its analogues were mediated by regulating renal organic ion transporters in hyperuricemic mice, supporting their beneficial effects for the prevention of hyperuricemia. The number and position, methoxylation and glycosylation of hydroxyl groups in these trans-stilbenes were required for their effects.

Furocoumarins affect hepatic cytochrome P450 and renal organic ion transporters in mice.

Furocoumarins are a group of natural products with many biological activities. Clinical evidences have demonstrated the important contribution of furocoumarins to the toxicity of some foods and herbs. In order to assess liver and kidney toxicity of furocoumarins, male mice were orally administrated with psoralen, isopsoralen, imperatorin, isoimperatorin and xanthotoxin at 20 and 40 mg/kg once daily for 28 days, respectively. No changes of food or water intake were observed in furocoumarins-treated mice. Only 40 mg/kg isopsoralen reduced body weight. 40 mg/kg furocoumarins altered serum activities of alanine transaminase, aspartate aminotransferase, alkaline phosphatase, and/or levels of albumin, showing hepatotoxicity. Furthermore, furocoumarins increased activity and protein expression of hepatic microsomal cytochrome P450 (CYP450) 3A11. CYP 2E1 activity and protein expression were suppressed by psoralen and isopsoralen and increased by xanthotoxin. Renal protein levels of organic cation/carnitine transporters (OCT1, OCT2 and OCTN2) and organic anion transporter 3 were increased by most furocoumarins. Renal urate transporter 1, glucose transporter 9 and multidrug resistance protein 4 were influenced by furocoumarins. These findings suggest that furocoumarins may interfere in metabolism, excretion and bioavailability of endogenous and exogenous compounds to impair liver and kidney functions mediated by affecting hepatic CYP450 and renal organic ion transport system.

Morin improves urate excretion and kidney function through regulation of renal organic ion transporters in hyperuricemic mice.

PURPOSE: Morin (2',3,4',5,7-pentahydroxyflavone), a plant-derived flavonoid, has beneficial effects on hyperuricemia and renal dysfunction in animals. Since the decreased renal excretion of uric acid is the hallmark of hyperuricemia, here we studied the effects of oral morin administration on renal organic ion transporters in potassium oxonate-induced hyperuricemic mice. METHODS: Hyperuricemia in mice was induced by potassium oxonate. Uric acid and creatinine concentrations in urine and serum, and fractional excretion of uric acid (FEUA) were performed to evaluate renal urate handling. Changes in expression levels of renal organic ion transporters were detected by Western blotting and semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) methods. RESULTS: Morin treatment significantly increased urinary uric acid/creatinine ratio and FEUA, resulting in reduction of serum uric acid levels in hyperuricemic mice. And kidney conditions were also improved after morin treatment in this model. Protein and mRNA levels of glucose transporter 9 (mGLUT9) and urate transporter 1 (mURAT1) were significantly decreased, and of organic anion transporter 1 (mOAT1) were remarkably increased in the kidney of morin-treated hyperuricemic mice. Morin treatment also blocked down-regulations of renal organic cation and carnitine transporters (mOCT1, mOCT2, mOCTN1 and mOCTN2) in hyperuricemic mice. CONCLUSION: These results suggest that morin exhibits the uricosuric effects via suppressing urate reabsorption and promoting urate secretion in the kidney of hyperuricemic mice and may help to attenuate deleterious effects of hyperuricemia with renal dysfunction.