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Background and purpose Alzheimer's disease (AD) is the most prevalent disease associated with cognitive dysfunction. Current AD therapeutic agents have several gastrointestinal or psychological adverse effects and therefore, novel therapeutic agents with fewer adverse effects must be developed. Previously, we demonstrated that oleanolic acid, which is similar in chemical structure to maslinic acid, ameliorates cognitive impairment through the activation of tropomyosin receptor kinase (TrkB)-ERK-cAMP response element-binding protein (CREB) phosphorylation and increased levels of brain-derived neurotrophic factor (BDNF). In the present study, we investigate the effect of maslinic acid on cholinergic blockade-induced memory impairment in mice. Methods and key results Maslinic acid reversed scopolamine-induced memory impairment, as determined by the Y-maze, passive avoidance and Morris water maze tests. In addition, we also observed that ERK-CREB, PI3K and PKB (Akt) phosphorylation levels were increased by maslinic acid administration in the mouse hippocampus. Moreover, we determined that the effects of maslinic acid on scopolamine-induced memory impairment in the passive avoidance test were abolished by a specific TrkB receptor antagonist (ANA-12). Additionally, we observed similar temporal changes in the expression levels between BDNF and tissue plasminogen activator in the hippocampus. Conclusion and implications These findings suggest that maslinic acid enhances cognitive function through the activation of BDNF and its downstream pathway signalling in the hippocampus and that it might be a potential therapeutic agent for cognitive decline, such as that observed in AD.

As a heterogeneous disorder, schizophrenia is known to be associated with neuroinflammation. A recent study showed that several cytokines are higher in the plasma and cerebrospinal fluid of schizophrenia patients. Lansoprazole, a proton pump inhibitor used for treating erosive esophagitis, has been reported to reduce INF-γ-induced neurotoxicity and decrease inflammatory cytokines including IL-1β, IL-6, and TNF-α. These findings persuaded us to examine whether lansoprazole ameliorates schizophrenia-like symptoms. The schizophrenia mouse model was induced by the acute administration of MK-801, an NMDA receptor antagonist. Sensorimotor gating, Barnes maze, and social novelty preference tests were conducted to evaluate schizophrenia-like behaviors. We found that lansoprazole (0.3, 1, or 3 mg/kg) ameliorated sensorimotor gating deficits, spatial learning, and social deficits caused by MK-801 treatment (0.2 mg/kg). The catalepsy test, balance beam test, and rotarod test were performed to reveal the adverse effects of lansoprazole on motor coordination. The behavioral results indicated that lansoprazole did not result in any motor function deficits. Moreover, lansoprazole decreased inflammatory cytokines including IL-6 and TNF-α only in the cortex, but not in the hippocampus. Collectively, these results suggest that lansoprazole could be a potential candidate for treating schizophrenia patients who suffer from sensorimotor gating deficits or social disability without any motor-related adverse effects.

Current antipsychotics have limited effects on the cognitive deficits of schizophrenia patients, therefore, cognitive remediation has been applied to schizophrenia patients to ameliorate cognitive dysfunction. However, the neurobiological mechanisms of cognitive training programs have not been well studied because established animal models are not suitable or because repetitive training has not been introduced in such animal models. In the present study, we employed Toll-like receptor 2 knockout (TLR2 KO) mouse as a schizophrenia mouse model and evaluated the effects of repetitive training as cognitive remediation therapy for schizophrenia. TLR2 KO mice could fully learn the Barnes maze paradigm through repetitive training to improve memory retrieval and reversal learning ability, although the learning speed was slower than that of wild-type (WT) animals. In addition, highly repetitive training activated the neuronal cells in the prefrontal cortex, hippocampal CA3 and hippocampal DG regions of TLR2 KO mice, similar to WT mice. These results indicated that TLR2 KO mouse would be a useful tool for studying the neurobiological mechanisms of cognitive remediation in schizophrenia.

The leaves of Aster glehni Fr. Schm. (Asteraceae) have been used to treat insomnia in Korea. Insomnia is a common adverse effect of therapeutic agents for Alzheimer's disease (AD), and the control of sleep disturbance may prevent dementia. We hypothesized that the leaves of A. glehni can attenuate cognitive dysfunctions observed in AD. We observed the ameliorating effects of the ethanolic extract of leaves of A. glehni (AG-D) on memory dysfunction through the Morris water maze test, the passive avoidance test, and the Y-maze test. We performed acetylcholinesterase (AChE) activity assay and Western blotting to determine the mechanism of action of AG-D. AG-D significantly attenuated memory dysfunction observed in the above behavior studies and inhibited the activity of AChE. AG-D also increased the levels of phosphorylation extracellular signal-regulated kinase (ERK), cAMP response element-binding protein (CREB), phosphatidylinositol 3-kinase (PI3K), protein kinase B (Akt), and glycogen synthase kinase 3β (GSK-3β) and the expression levels of brain-derived neurotrophic factor (BDNF) in the hippocampi. These results suggest that AG-D ameliorates memory impairments by AChE inhibition and activation of ERK-CREB-BDNF and PI3K-Akt-GSK-3β signaling pathways. Taken together, this study suggests that AG-D could be used as a potential treatment for cognitive dysfunction.

As the elderly population is increasing, Alzheimer's disease (AD) has become a global issue and many clinical trials have been conducted to evaluate treatments for AD. As these clinical trials have been conducted and have failed, the development of new theraphies for AD with fewer adverse effects remains a challenge. In this study, we examined the effects of Theracurmin on cognitive decline using 5XFAD mice, an AD mouse model. Theracurmin is more bioavailable form of curcumin, generated with submicron colloidal dispersion. Mice were treated with Theracurmin (100, 300 and 1,000 mg/kg) for 12 weeks and were subjected to the novel object recognition test and the Barnes maze test. Theracurmin-treated mice showed significant amelioration in recognition and spatial memories compared those of the vehicle-treated controls. In addition, the antioxidant activities of Theracurmin were investigated by measuring the superoxide dismutase (SOD) activity, malondialdehyde (MDA) and glutathione (GSH) levels. The increased MDA level and decreased SOD and GSH levels in the vehicle-treated 5XFAD mice were significantly reversed by the administration of Theracurmin. Moreover, we observed that Theracurmin administration elevated the expression levels of synaptic components, including synaptophysin and post synaptic density protein 95, and decreased the expression levels of ionized calcium-binding adapter molecule 1 (Iba-1), a marker of activated microglia. These results suggest that Theracurmin ameliorates cognitive function by increasing the expression of synaptic components and by preventing neuronal cell damage from oxidative stress or from the activation of microglia. Thus, Theracurmin would be useful for treating the cognitive dysfunctions observed in AD.

The importance of alterations in bidirectional communication between gut and brain has become obvious in neuropsychiatric disorders. Gastrointestinal (GI) disturbances are very common in autism spectrum disorders (ASD), and the GI microbiota profiles in children with ASD are significantly different from those in the general population. Fragile X syndrome (FXS) is an inheritable developmental disability in humans, and patients with FXS exhibit autistic behaviors such as mental retardation and impaired social communication or interaction. We hypothesized that an increase in specific gut microbiota by fecal microbiota transplantation (FMT) would mitigate autistic-like behaviors. To test this hypothesis, we measured the effects of FMT from normal mice to Fmr1 KO mice on autistic-like behaviors using several behavioral tests. Because the amounts of A. muciniphila in Fmr1 KO mice was very low, we assessed A. muciniphila population, tested the expression of MUC2, and analyzed goblet cells in the gut after the FMT. We found that FMT ameliorated autistic-like behaviors, especially memory deficits and social withdrawal, and we observed that the levels of A. muciniphila were normalized to wild-type levels. In addition, FMT attenuated the increased levels of TNFα and Iba1 in the brains of Fmr1 KO mice. These results suggest that FMT could be a useful tool for the treatments of cognitive deficits and social withdrawal symptoms observed in FXS or ASD because it increases the population of A. muciniphila and decreases TNFα and Iba1 levels.

Ethnopharmacological relevance Scrophularia buergeriana has been used for traditional medicine as an agent for reducing heat in the blood and for nourishing kidney ‘Yin’. Therefore, S. buergeriana might be a potential treatment for mental illness, especially schizophrenia, which may be attenuated by supplying kidney Yin and reducing blood heat. In a pilot study, we found that S. buergeriana alleviated sensorimotor gating dysfunction induced by MK-801. Aim of the study In the present study, we attempted to reveal the active component(s) of S. buergeriana as a candidate for treating sensorimotor gating dysfunction, and we identified 4-methoxycinnamic acid. We explored whether 4-methoxycinnamic acid could affect schizophrenia-like behaviors induced by hypofunction of the glutamatergic neurotransmitter system. Materials and methods Mice were treated with 4-methoxycinnamic acid (3, 10, or 30 mg/kg, i.g.) under MK-801-induced schizophrenia-like conditions. The effect of 4-methoxycinnamic acid on schizophrenia-like behaviors were explored using several behavioral tasks. We also used Western blotting to investigate which signaling pathway(s) is involved in the pharmacological activities of 4-methoxycinnamic acid. Results 4-Methoxycinnamic acid ameliorated MK-801-induced prepulse inhibition deficits, social interaction disorders and cognitive impairment by regulating the phosphorylation levels of PI3K, Akt and GSK-3β signaling in the prefrontal cortex. And there were no adverse effects in terms of catalepsy and motor coordination impairments. Conclusion Collectively, 4-methoxycinnamic acid would be a potential candidate for treating schizophrenia with fewer adverse effects, especially the negative symptoms and cognitive dysfunctions.

Present work aimed to introduce non-peptidic ABAD loop D (LD) hot spots mimetics as ABAD-Aβ inhibitors. A full length atomistic model of ABAD-Aβ complex was built as a scaffold to launch the lead design and its topology later verified by crosschecking the computational mutagenesis results with that of in vitro data. Thereafter, the interactions of prime Aβ-binding LD residues- Tyr101, Thr108, and Thr110 were translated into specific pharmacophore features and this hypothesis subsequently used as a virtual screen query. ELISA-based screening of 20 hits identified two promising lead candidates, VC15 and VC19 with an IC50 of 4.4±0.3 and 9.6±0.1 μM respectively. They productively reversed Aβ-induced mitochondrial dysfunctions such as mitochondrial membrane potential loss (JC-1 assay), toxicity (MTT assay), and ATP reduction (ATP assay) in addition to increased cell viabilities. This is the first reporting of LD hot spots-centric in silico scheme to discover novel compounds with promising ABAD-Aβ inhibitory potential. These chemotypes are proposed for further structural optimization to derive novel Alzheimer's disease (AD) therapeutics. This article is protected by copyright. All rights reserved.

Ethnopharmacological relevance Green tea has been used as a traditional medicine to control brain function and digestion. Recent works suggest that drinking green tea could prevent cognitive function impairment. During tea manufacturing processes, such as brewing and sterilization, green tea catechins are epimerized. However, the effects of heat-epimerized catechins on cognitive function are still unknown. To take this advantage, we developed a new green tea extract, high temperature processed-green tea extract (HTP-GTE), which has a similar catechin composition to green tea beverages. Aim of the study This study aimed to investigate the effect of HTP-GTE on scopolamine-induced cognitive dysfunction and neuronal differentiation, and to elucidate its underlying mechanisms of action. Materials and methods The neuronal differentiation promoting effects of HTP-GTE in SH-SY5Y cells was assessed by evaluating neurite length and the expression level of synaptophysin. The DNA methylation status at the synaptophysin promoter was determined in differentiated SH-SY5Y cells and in the hippocampi of mice. HTP-GTE was administered for 10 days at doses of 30, 100 and 300 mg/kg (p.o.) to mice, and its effects on cognitive functions were measured by Y-maze and passive avoidance tests under scopolamine-induced cholinergic blockade state. Results HTP-GTE induced neuronal differentiation and neurite outgrowth via the upregulation of synaptophysin gene expression. These beneficial effects of HTP-GTE resulted from reducing DNA methylation levels at the synaptophysin promoter via the suppression of DNMT1 activity. The administration of HTP-GTE ameliorated cognitive impairments in a scopolamine-treated mouse model. Conclusions These results suggest that HTP-GTE could alleviate cognitive impairment by regulating synaptophysin expression and DNA methylation levels. Taken together, HTP-GTE would be a promising treatment for the cognitive impairment observed in dysfunction of the cholinergic neurotransmitter system.

Objectives A botanical drug derived from the ethanolic extract composed of Clematis chinensis Osbeck (Ranunculaceae), Trichosanthes kirilowii Maximowicz (Cucurbitaceae) and Prunella vulgaris Linné (Lamiaceae) has been used to ameliorate rheumatoid arthritis as an ethical drug in Korea. In our study, we investigated the effect of this herbal complex extract (HCE) on schizophrenia-like behaviours induced by MK-801. Methods HCE (30, 100 or 300 mg/kg, p.o) was orally administered to male ICR mice to a schizophrenia-like animal model induced by MK-801. We conducted an acoustic startle response task, an open-field task, a novel object recognition task and a social novelty preference task. Key findings We found that a single administration of HCE (100 or 300 mg/kg) ameliorated MK-801-induced abnormal behaviours including sensorimotor gating deficits and social or object recognition memory deficits. In addition, MK-801-induced increases in phosphorylated Akt and GSK-3β expression levels in the prefrontal cortex were reversed by HCE (30, 100 or 300 mg/kg). Conclusions These results imply that HCE ameliorates MK-801-induced dysfunctions in prepulse inhibition, social interactions and cognitive function, partly by regulating the Akt and GSK-3β signalling pathways.

In this study, we designed a library of compounds based on the structures of well-known ligands of the 18 kDa translocator protein (TSPO), one of the putative components of the mPTP. We performed diverse mitochondrial functional assays to assess their ability to restore cells from Aβ-induced toxicity in vitro and in vivo. Among tested compounds, compound 25 effectively improved cognitive function in animal models of AD. Given the excellent in vitro and in vivo activity and a favorable pharmacokinetic profile of compound 25, we believe that it can serve as a promising lead compound for a potential treatment option for AD.

TREK1 (Twik-RElated Potassium (K+ ) channel 1), although a well-characterized target for several neuropsychiatric disorders, underwent very few explorations for prototypic inhibitors. This study aimed to find diverse chemotypes by an in silico means. Homology-built TREK1 on docking with high-affinity quaternary ammonium compounds (QAs) corroborated the previous findings by recreating the binding mode with proximally positioned key residues: Thr157, Thr266, Ile182, Leu189, and Leu304. Physical interactions between TREK1 and known antagonists were modeled to compensate the lack of ligand-bound protein crystal structures. A common feature hypothesis (Hypo1) was deduced from the chemical features of six active compounds. Validated Hypo1 and the most potent compound in the data set were employed as pharmacophore- and similarity-based virtual screening queries, respectively. Thirty-three hit compounds were tested for their ability to block TREK1 currents in HEK-293-transfected cells using whole-cell patch-clamp recording. Eleven candidates displayed dose-dependent inhibition of channel currents; among these, NC30 possessing a 4-((1H-pyrrolo[2,3-b]pyridin-1-yl)methyl)piperidin-4-ol heterocyclic core was the most potent one with an IC50 of 4.7 μm. These results form a rational basis to design future drugs, and this is the first report of novel TREK1 antagonists delineated by a synergistic application of structure- and ligand-based approaches.

Ethnopharmacological relevance The fruit of Vitex rotundifolia L. (Verbenaceae) has been used in traditional medicine as sedative or analgesic agent for headache. Recent population-based cohort studies have shown that headache including migraines is a risk factor for dementia. Thus, the fruit of V. rotundifolia may be useful for treating cognitive dysfunction observed in dementia. Aim of the study We had previously found that the ethanolic extract of the fruit of V. rotundifolia ameliorated cognitive dysfunction and isolated casticin as an active compound. In the present study, we studied the effect of casticin on a mouse model of cognitive impairment induced by scopolamine. Materials and methods Mice were treated with the ethanolic extract of the fruit of V. rotundifolia (EEVR; 30, 100 or 300 mg/kg, p.o.) or casticin (0.3, 1 or 3 mg/kg, p.o.). We examined the effect of casticin or EEVR using the passive avoidance test, the Morris water maze test and the novel object recognition test. Scopolamine (1 mg/kg, i.p.) was used to induce cognitive impairment by blocking cholinergic neurotransmitter system. We investigated the effects of casticin on acetylcholinesterase (AchE) activity and the phosphorylation levels of extracellular signal-regulated kinase (ERK), cAMP response element binding protein (CREB), and the expression levels of brain-derived neurotrophic factor (BDNF). Results EEVR (100 and 300 mg/kg, p.o.) significantly ameliorated the latency in the passive avoidance test, and casticin (1 and 3 mg/kg, p.o.) also significantly improved the latency in the passive avoidance test, novel object preference in the novel object recognition test, and swimming time in the target quadrant of the Morris water maze test. Casticin also decreased AChE activity in ex vivo analysis and increased the phosphorylation levels of memory-related signaling molecules, such as ERK, CREB and BDNF in the cortex. Conclusion These results suggest that casticin ameliorates cholinergic blockade-induced cognitive impairment, in part, through the inhibition of AChE and the activation of the ERK-CREB-BDNF signaling pathway. Taken together, the results suggest that casticin may be useful for treating the cognitive dysfunction observed during cholinergic impairment.

The stem of Opuntia ficus-indica var. saboten is edible and has been used as a medicinal herb on Jeju Island in Korea. We previously reported that the butanolic extract of O. ficus-indica var. saboten exerts the enhancement of long-term memory in mice. However, the antiamnesic effects of O. ficus-indica var. saboten and its mode of action has not been clearly elucidated. In the present study, we explored the effects of the ethanolic extract of stems of O. ficus-indica var. saboten (EOFS) on cognitive performance in mouse and attempted to delineate its mechanism of action. We used the passive avoidance, Y-maze, and novel object recognition tests to assess its effects on cognitive functions in scopolamine-induced memory-impaired mice. We observed that EOFS (100, 200, and 400 mg/kg) ameliorated scopolamine-induced cognitive dysfunction. We also explored its mechanism of action by conducting an acetylcholinesterase (AChE) activity assay using the mouse whole brain and Western blot using the mouse hippocampal tissue. Western blot analysis and the ex vivo study revealed that EOFS increased the levels of phosphorylated extracellular signal-regulated kinase and cAMP response element-binding protein (CREB) and the levels of brain-derived neurotrophic factor (BDNF) expression in the hippocampus. It also inhibited AChE activity in the brain. Our findings suggest that EOFS would be useful for the treatment of cholinergic blockade-induced cognitive dysfunction.

Cyclophilin D (CypD) is a mitochondria-specific cyclophilin that is known to play a pivotal role in the formation of the mitochondrial permeability transition pore (mPTP).The formation and opening of the mPTP disrupt mitochondrial homeostasis, cause mitochondrial dysfunction and eventually lead to cell death. Several recent studies have found that CypD promotes the formation of the mPTP upon binding to β amyloid (Aβ) peptides inside brain mitochondria, suggesting that neuronal CypD has a potential to be a promising therapeutic target for Alzheimer’s disease (AD). In this study, we generated an energy-based pharmacophore model by using the crystal structure of CypD—cyclosporine A (CsA) complex and performed virtual screening of ChemDiv database, which yielded forty-five potential hit compounds with novel scaffolds. We further tested those compounds using mitochondrial functional assays in neuronal cells and identified fifteen compounds with excellent protective effects against Aβ-induced mitochondrial dysfunction. To validate whether these effects derived from binding to CypD, we performed surface plasmon resonance (SPR)—based direct binding assays with selected compounds and discovered compound 29 was found to have the equilibrium dissociation constants (KD) value of 88.2 nM. This binding affinity value and biological activity correspond well with our predicted binding mode. We believe that this study offers new insights into the rational design of small molecule CypD inhibitors, and provides a promising lead for future therapeutic development.

Recovery of mitochondrial dysfunction has gained increasing attention as an alternative therapeutic strategy for Alzheimer's disease (AD). Recent studies suggested that the 18 kDa mitochondrial translocator protein (TSPO) has the potential to serve as a drug target for the treatment of AD. In this study, we generated a structure-based pharmacophore model and virtually screened a commercial library, identifying SVH07 as a virtual hit, which contained a tricyclic core structure, thieno[2',3':4,5]pyrrolo[1,2-d][1,2,4]triazine group. A series of SVH07 analogues were synthesized and their effects on the mitochondrial membrane potential and ATP production were determined by using neuronal cells under Aβ-induced toxicity. Among these analogues, compound 26 significantly recovered mitochondrial membrane depolarization and ATP production. In vitro binding assays indicated that SVH07 and 26 showed high affinities to TSPO with the IC50 values in a nanomolar range. We believe that compound 26 is a promising lead compound for the development of TSPO-targeted mitochondrial functional modulators with therapeutic potential in AD.

Present work aimed to introduce non-peptidic ABAD loop D (L

Stroke is the third leading cause of death in most industrialized countries, and the most important source of chronic disability, among which cerebral ischemic stroke represents about 85% of all. Ischemic stroke occurs by thrombotic or embolic blockade of an artery to the brain resulting in a deficiency in blood flow and causing a brain infarction. During ischemic stroke, diminished blood flow initiates a series of events such as inducing excessive release of excitatory amino acids and subsequent several receptor activations leading to elevated calcium influx that may result in damage to brain cells. Oxidative stress is one of the primary factors that exacerbate damage in cerebral ischemia. Moreover, the reactive oxygen species can create a secondary source of extensive cell injury and death since re-supply of oxygen to the brain through the reperfusion is relevant during the ischemic phase. Therefore, antioxidants that can scavenge oxygen free radicals have the therapeutic potential for the treatment of neuronal injury following ischemia and reperfusion. Many natural antioxidants are reported to reduce reactive oxygen species and protect neuronal cells in animal models of cerebral ischemia. Previously, we reported protective effects of quercetin 3O-methyl ether (1) and its related compounds (2, 3) isolated from Opuntia ficus-indica var. saboten against oxidative neuronal injuries induced in primary cultured rat cortical cells (Fig. 1). Quercetin (2) was found to inhibit H2O2or xanthine (X)/xanthine oxidase (XO)-induced oxidative neuronal cell injury with an estimated IC50 value of 4-5 μg/ mL. (+)-Dihydroquercetin (3) inhibited oxidative neuronal injuries concentration-dependently, but it was 2-3 fold less potent than 2. On the other hand, quercetin 3-O-methyl ether (1) potently inhibited H2O2and X/XO-induced neuronal injuries with IC50 values of 0.6 and 0.7 μg/mL, respectively, indicating that 1 appeared to be the most potent neuroprotectant among three flavonoids isolated from this plant. Recently, antioxidative activities in cell-free systems of 1 have also been reported. However, neuroprotective effects of 1 in ischemic animal models have not yet been reported. The present study, therefore, examined neuroprotective effects of 1 in a transient focal cerebral ischemic rat model and compared with those of related flavonoids 2 and (±)-3. The middle cerebral artery occlusion (MCAO) model has usually been used for assessing neuroprotective effects since most ischemic strokes occur in the territory of the middle cerebral artery. Transient focal cerebral ischemia was induced by occlusion of the right middle cerebral artery for 2 h with a silicone-coated 4-0 nylon monofilament in male Sprague-Dawley rats. The antioxidative flavonoids dissolved in 0.9% saline vehicle were administered intraperitoneally at a dose of 10 mg/kg 30 min after onset of ischemia. Infarct size and % edema were measured 24 h after onset of ischemia using a 2,3,5-triphenyltetrazolium chloride (TTC) staining method. Neurological scoring was performed 30 min and 24 h after MCAO. The neurobehavioral tests consisted of scoring the degree of left forelimb flexion (0 to 3), the duration of left forelimb flexion (0 to 4) and symmetry of movement/forepaw outstretching (0 to 3). Rats were scored on a ranking scale of 0 to 10, which reflects the cumulative score of the individual tests with a score of 10 reflecting normal behavior. Representative TTC staining is shown in Figure 2 to illustrate neuroprotective effects of 1, 2 and (±)3. Area stained red with TTC was considered normal, while area not stained red with TTC was considered infarcted.

Opuntia ficus-indica var. saboten Makino (Cactaceae) is used to treat burns, edema, dyspepsia, and asthma in traditional medicine. The present study investigated the beneficial effects of the n-butanolic extract of O. ficus-indica var. saboten (BOF) on memory performance in mice and attempts to uncover the mechanisms underlying its action. Memory performance was assessed with the passive avoidance task, and western blotting and immunohistochemistry were used to measure changes in protein expression and cell survival. After the oral administration of BOF for 7 days, the latency time in the passive avoidance task was significantly increased relative to vehicle-treated controls (P

Calpains are involved in a variety of calcium-regulated cellular processes, such as signal transduction, cell proliferation, differentiation, and apoptosis. Excessive calpain activation contributes to serious cellular damage and has been reported in many pathological conditions. 4-Quinolinone 2-carboxamide derivatives were prepared and evaluated for μ-calpain inhibitory activities. Of the compounds synthesized, 3a and 3k, which possess a primary amide and 4-methoxyphenethyl amide at P1′ region, were found to most potently inhibit μ-calpain with IC50 values of 0.71 ± 0.07 and 0.73 ± 0.23 μM, respectively. On the other hand, the incorporation of pyridine-containing amides decreased inhibitory activity.

Reactive oxygen species have been implicated in several diseases, particularly in ischemia-reperfusion injury. Quercetin 3-O-methyl ether has been reported to show potent antioxidant and neuroprotective activity against neuronal damage induced by reactive oxygen species. Several aminoethyl-substituted derivatives of quercetin 3-O-methyl ether have been synthesized to increase water solubility while retaining antioxidant and neuroprotective activity. Among such derivatives, compound 3a shows potent and well-balanced antioxidant activity in three types of cell-free assay systems and has in vivo neuroprotective effects on transient focal ischemic injury induced by the occlusion of the middle cerebral artery in rats.

New chromone carboxamide derivatives like (I) are synthesized by conventional procedures to provide alternative μ-calpain inhibitors to other conformationally constrained small peptide variants.

The excessive calpain activation causes serious cellular damage or even cell death in neurological disorders such as stroke and Alzheimer’s disease. Oxidative stress has also been implicated in the initiation or progression of neurodegenerative diseases. In the present studies, a series of cinnamoyl ketoamides 4a–4j were synthesized as hybrid structures of antioxidants and calpain inhibitors. Cinnamoyl ketoamides, possessing an alkyl chain at the α-position, showed potent μ-calpain inhibitory activities indicating that the cinnamoyl skeleton can be regarded as an acyclic variant of calpain inhibitory chromone carboxamide 2. Among synthesized, compound 4e was the most potent inhibitor of μ-calpain (IC50 = 0.13 μM) and also exhibited strong antioxidant activities in DPPH and superoxide anion radical scavenging and lipid peroxidation inhibition assay systems.

The overactivation of μ-calpain can cause serious cell damage in several diseases. Furthermore, cell death in a number of neurodegenerative disorders is linked to the overproduction of reactive oxygen species. Therefore, antioxidants and μ-calpain inhibitors could have the therapeutic potentials to treat cell death related diseases. New chromone carboxamide derivatives 3 were synthesized to provide alternative μ-calpain inhibitors to compound 2, a conformationally constrained structural variant of MDL 28,170. Compounds 3h and 3l exhibited the most potent μ-calpain inhibitory activities (IC50=0.09-0.10 μM), and were comparable to 2 in this respect (IC50=0.07 μM). Compound 3i showed both potent μ-calpain inhibitory activity (IC50=0.28 μM) and antioxidant activities in DPPH scavenging and lipid peroxidation inhibition assays.

Ischemic preconditioning can provide protection to neurons from subsequent lethal ischemia. The molecular mechanisms of neuronal ischemic tolerance, however, are still not well-known. The present study, therefore, examined the role of MAPK and PI3K/Akt pathways in ischemic tolerance induced by preconditioning with sublethal oxygen-glucose deprivation (OGD) in cultured rat cortical neurons. Ischemic tolerance was simulated by preconditioning of the neurons with sublethal 1-h OGD imposed 12 h before lethal 3-h OGD. The time-course studies of relative phosphorylation and expression levels of ERK1/2, JNK and p38 MAPK showed lack of their involvement in ischemic tolerance. However, there were significant increases in Akt phosphorylation levels during the reperfusion period following preconditioned lethal OGD. In addition, Bcl-2 associated death promoter (Bad) and GSK-3β were also found to be inactivated during that reperfusion period. Finally, treatment with an inhibitor of PI3K, wortmannin, applied from 15 min before and during lethal OGD abolished not only the preconditioning-induced neuroprotection but also the Akt activation. Concomitant with blockade of the Akt activation, PI3K inhibition also resulted in activation of Bad and GSK-3β. The results suggest that ischemic tolerance induced by sublethal OGD preconditioning is primarily mediated through activation of the PI3K/Akt pathway, but not the MAPK pathway, in rat cortical neurons.

To search for secondary metabolites of Alnus hirsuta (Betulaceae), various chromatographic separations of the ethyl acetate soluble fraction of the stem bark of A. hirsuta led to the isolation of a new diarylheptanoid glycoside, (3R)-1,7-bis-(4-dihydroxyphenyl)-3-heptanol 3-O-β-D-glucopyranosyl(1→3)-β-D-xylopyranoside (13) and twelve diarylheptanoid derivatives, namely, oregonin (1), rubranoside A (2), hirsutanonol 5-O-β-D-glucopyranoside (3), rubranoside B (4), rubranoside C (5), hirsutanonol (6), hirsutenone (7), (5S)-O-methylhirsutanonol (8), platyphylloside (9), platyphyllonol 5-O-β-D-xylopyranoside (10), aceroside VII (11) and platyphyllenone (12). Isolates were assessed for their hepatoprotective effects against tert-butylhydroperoxide (t-BHP)-induced toxicity in HepG2 cells. Of these isolates, compounds 1—8 showed significant hepatoprotective effects on t-BHP-induced damage to HepG2 cells, with 8 exhibiting the greatest protective effect (50.7±3.7% at a concentration of 10 μM).

To search for secondary metabolites of Alnus hirsuta (Betulaceae), various chromatographic separations of the ethyl acetate soluble fraction of the stem bark of A. hirsuta led to the isolation of a new diarylheptanoid glycoside, (3R)-1,7-bis-(4-dihydroxyphenyl)-3-heptanol 3-O-beta-D-glucopyranosyl(1--3)-beta-D-xylopyranoside (13) and twelve diarylheptanoid derivatives, namely, oregonin (1), rubranoside A (2), hirsutanonol 5-O-beta-D-glucopyranoside (3), rubranoside B (4), rubranoside C (5), hirsutanonol (6), hirsutenone (7), (5S)-O-methylhirsutanonol (8), platyphylloside (9), platyphyllonol 5-O-beta-D-xylopyranoside (10), aceroside VII (11) and platyphyllenone (12). Isolates were assessed for their hepatoprotective effects against tert-butylhydroperoxide (t-BHP)-induced toxicity in HepG2 cells. Of these isolates, compounds 1-8 showed significant hepatoprotective effects on t-BHP-induced damage to HepG2 cells, with 8 exhibiting the greatest protective effect (50.7 + or - 3.7% at a concentration of 10 microM).

The complex molecular cascades of ischemic tolerance in brain cells remain unclear. Recently, sphingolipid-related metabolite ceramide has been implicated as a second messenger in many biological functions, including neuronal survival and death. The present study, therefore, examined the roles of ceramide (Cer) in ischemic tolerance induced by preconditioning with sublethal oxygen-glucose deprivation (OGD) using primary cultured cortical neurons of rats. Preconditioning of the neurons with sublethal 1-h OGD produced robust neuroprotection against cell death induced by lethal 3-h OGD imposed 12 h after preconditioning when measured by the MTT assay. Analysis of sphingolipids using LC-MS/MS showed that the ischemic preconditioning resulted in significant increases in the levels of C(16 : 0) Cer, C(18 : 0) Cer, C(20 : 0) Cer, C(24 : 0) Cer, C(24 : 1) Cer and the total ceramide contents compared with the sham-washed control group. However, sphingomyelin contents were not significantly changed by the ischemic preconditioning, suggesting that ceramides were increased through the de novo synthetic pathway. In the case of severe OGD paradigm, levels of ceramide and sphingomyelin in the lethal OGD group were not significantly different from those of the control group or the lethal OGD group with preconditioning at any time points studied. Treatment with an inhibitor of de novo ceramide synthesis, fumonisin B(1), during the ischemic preconditioning period completely blocked preconditioning-induced ischemic tolerance. Moreover, application of a non-cytotoxic concentration of exogenous cell-permeable ceramide produced neuroprotection against lethal OGD. The results suggest that ceramides increased by sublethal OGD preconditioning play an important role in induction of ischemic tolerance.

In the present study, we assessed the effects of gluco-obtusifolin, isolated from the seeds of Cassia obtusifolia L., and its aglycone, obtusifolin, on the learning and memory impairments induced by scopolamine using the passive avoidance and the Morris water maze tasks in mice. Gluco-obtusifolin (1, 2, and 4 mg/kg, p.o.) and obtusifolin (0.25, 0.5, 1, and 2 mg/kg, p.o.) significantly reversed scopolamine-induced cognitive impairments in the passive avoidance test (P