Diosgenin restores memory function via SPARC-driven axonal growth from the hippocampus to the PFC in Alzheimer's disease model mice.

Central nervous system axons have minimal capacity to regenerate in adult brains, hindering memory recovery in Alzheimer's disease (AD). Although recent studies have shown that damaged axons sprouted in adult and AD mouse brains, long-distance axonal re-innervation to their targets has not been achieved. We selectively visualized axon-growing neurons in the neural circuit for memory formation, from the hippocampus to the prefrontal cortex, and showed that damaged axons successfully extended to their native projecting area in mouse models of AD (5XFAD) by administration of an axonal regenerative agent, diosgenin. In vivo transcriptome analysis detected the expression profile of axon-growing neurons directly isolated from the hippocampus of 5XFAD mice. Secreted protein acidic and rich in cysteine (SPARC) was the most expressed gene in axon-growing neurons. Neuron-specific overexpression of SPARC via adeno-associated virus serotype 9 delivery in the hippocampus recovered memory deficits and axonal projection to the prefrontal cortex in 5XFAD mice. DREADDs (Designer receptors exclusively activated by designer drugs) analyses revealed that SPARC overexpression-induced axonal growth in the 5XFAD mouse brain directly contributes to memory recovery. Elevated levels of SPARC on axonal membranes interact with extracellular rail-like collagen type I to promote axonal remodeling along their original tracings in primary cultured hippocampal neurons. These findings suggest that SPARC-driven axonal growth in the brain may be a promising therapeutic strategy for AD and other neurodegenerative diseases.

Alkannin Attenuates Amyloid ß Aggregation and Alzheimer's Disease Pathology.

Alzheimer's disease (AD) is a neurodegenerative disease that is accompanied by memory decline and cognitive dysfunction. Aggregated amyloid ß formation and accumulation may be one of the underlying mechanisms of the pathophysiology of AD. Therefore, compounds that can inhibit amyloid ß aggregation may be useful for treatment. Based on this hypothesis, we screened plant compounds used in Kampo medicine for chemical chaperone activity and identified that alkannin had this property. Further analysis indicated that alkannin could inhibit amyloid ß aggregation. Importantly, we also found that alkannin inhibited amyloid ß aggregation after aggregates had already formed. Through the analysis of circular dichroism spectra, alkannin was found to inhibit ß-sheet structure formation, which is an aggregation-prone toxic structure. Furthermore, alkannin attenuated amyloid ß-induced neuronal cell death in PC12 cells, ameliorated amyloid ß aggregation in the AD model of Caenorhabditis elegans (C. elegans), and inhibited chemotaxis observed in AD C. elegans, suggesting that alkannin could potentially inhibit neurodegeneration in vivo. Overall, these results suggest that alkannin may have novel pharmacological properties for inhibiting amyloid ß aggregation and neuronal cell death in AD. SIGNIFICANCE STATEMENT: Aggregated amyloid ß formation and accumulation is one of the underlying mechanisms of the pathophysiology of Alzheimer's disease. We found that alkannin had chemical chaperone activity, which can inhibit ß-sheet structure formation of amyloid ß and its aggregation, neuronal cell death, and Alzheimer's disease phenotype in C. elegans. Overall, alkannin may have novel pharmacological properties for inhibiting amyloid ß aggregation and neuronal cell death in Alzheimer's disease.

The roles of natural triterpenoid saponins against Alzheimer's disease.

The aging of the world population and increasing stress levels in life are the major cause of the increased incidence of neurological disorders. Alzheimer's disease (AD) creates a huge burden on the lives and health of individuals and has become a big concern for society. Triterpenoid saponins (TS), representative natural product components, have a wide range of pharmacological bioactivities such as anti-inflammation, antioxidation, antiapoptosis, hormone-like, and gut microbiota regulation. Notably, some natural TS exhibited promising neuroprotective activity that can intervene in AD progress, especially in the early stage. Recently, studies have indicated that TS play a pronounced positive role in the prevention and treatment of AD. This review discusses the recent research on the neuroprotection of TS and proceeds to detail the action mechanisms of TS against AD, hoping to provide a reference for drug development for anti-AD.

Shati/Nat8l Overexpression Improves Cognitive Decline by Upregulating Neuronal Trophic Factor in Alzheimer's Disease Model Mice.

Alzheimer's disease (AD) is a type of dementia characterized by the deposition of amyloid ß, a causative protein of AD, in the brain. Shati/Nat8l, identified as a psychiatric disease related molecule, is a responsive enzyme of N-acetylaspartate (NAA) synthesis. In the hippocampi of AD patients and model mice, the NAA content and Shati/Nat8l expression were reported to be reduced. Having recently clarified the involvement of Shati/Nat8l in cognitive function, we examined the recovery effect of the hippocampal overexpression of Shati/Nat8l in AD model mice (5XFAD). Shati/Nat8l overexpression suppressed cognitive dysfunction without affecting the Aß burden or number of NeuN-positive neurons. In addition, brain-derived neurotrophic factor mRNA was upregulated by Shati/Nat8l overexpression in 5XFAD mice. These results suggest that Shati/Nat8l overexpression prevents cognitive dysfunction in 5XFAD mice, indicating that Shati/Nat8l could be a therapeutic target for AD.

Natural Medicines and Their Underlying Mechanisms of Prevention and Recovery from Amyloid Β-Induced Axonal Degeneration in Alzheimer's Disease.

In Alzheimer's disease (AD), amyloid ß (Aß) induces axonal degeneration, neuronal network disruption, and memory impairment. Although many candidate drugs to reduce Aß have been clinically investigated, they failed to recover the memory function in AD patients. Reportedly, Aß deposition occurred before the onset of AD. Once neuronal networks were disrupted by Aß, they could hardly be recovered. Therefore, we speculated that only removal of Aß was not enough for AD therapy, and prevention and recovery from neuronal network disruption were also needed. This review describes the challenges related to the condition of axons for AD therapy. We established novel in vitro models of Aß-induced axonal degeneration. Using these models, we found that several traditional medicines and their constituents prevented or helped recover from Aß-induced axonal degeneration. These drugs also prevented or helped recover from memory impairment in in vivo models of AD. One of these drugs ameliorated memory decline in AD patients in a clinical study. These results indicate that prevention and recovery from axonal degeneration are possible strategies for AD therapy.

Naringenin promotes microglial M2 polarization and Aß degradation enzyme expression.

Two kinds of microglia are known, classical M1 and alternative M2 phenotypes. Amyloid ß (Aß), a critical cause of Alzheimer's disease (AD), promotes M1 microglial polarization, leading to neuroinflammation and neuronal death. M2 microglia play important roles in anti-inflammatory effects, Aß clearance, and memory recovery in AD. Therefore, increasing of M2 microglia is expected to recover from AD. We previously found that naringenin, a blood-brain barrier penetrating compound, decreased Aß deposits and recovers memory function in transgenic AD model mice. Naringenin reportedly showed anti-inflammatory properties. Here, we aim to investigate potential effects of naringenin on microglial polarization and to reveal the underlying mechanisms of Aß reduction. Primary cultured cortical microglia were treated with Aß1-42 , following administration of naringenin. Naringenin remarkably promoted M2 microglia polarization and inhibited Aß1-42 -induced M1 microglia activation. Because microglia reportedly played a critical role in cerebral Aß clearance through Aß degradation enzymes after phagocytosis, we investigated the expression of Aß degradation enzymes, such as neprilysin and insulin degradation enzyme. After naringenin treatment, these Aß degradation enzymes were downregulated in M1 microglia and upregulated in M2 microglia. Taken together, our results showed that naringenin increased Aß degradation enzymes in M2 microglia, probably leading to Aß plaque reduction.

A Novel Rac1-GSPT1 Signaling Pathway Controls Astrogliosis Following Central Nervous System Injury.

Astrogliosis (i.e. glial scar), which is comprised primarily of proliferated astrocytes at the lesion site and migrated astrocytes from neighboring regions, is one of the key reactions in determining outcomes after CNS injury. In an effort to identify potential molecules/pathways that regulate astrogliosis, we sought to determine whether Rac/Rac-mediated signaling in astrocytes represents a novel candidate for therapeutic intervention following CNS injury. For these studies, we generated mice with Rac1 deletion under the control of the GFAP (glial fibrillary acidic protein) promoter (GFAP-Cre;Rac1flox/flox). GFAP-Cre;Rac1flox/flox (Rac1-KO) mice exhibited better recovery after spinal cord injury and exhibited reduced astrogliosis at the lesion site relative to control. Reduced astrogliosis was also observed in Rac1-KO mice following microbeam irradiation-induced injury. Moreover, knockdown (KD) or KO of Rac1 in astrocytes (LN229 cells, primary astrocytes, or primary astrocytes from Rac1-KO mice) led to delayed cell cycle progression and reduced cell migration. Rac1-KD or Rac1-KO astrocytes additionally had decreased levels of GSPT1 (G1 to S phase transition 1) expression and reduced responses of IL-1ß and GSPT1 to LPS treatment, indicating that IL-1ß and GSPT1 are downstream molecules of Rac1 associated with inflammatory condition. Furthermore, GSPT1-KD astrocytes had cell cycle delay, with no effect on cell migration. The cell cycle delay induced by Rac1-KD was rescued by overexpression of GSPT1. Based on these results, we propose that Rac1-GSPT1 represents a novel signaling axis in astrocytes that accelerates proliferation in response to inflammation, which is one important factor in the development of astrogliosis/glial scar following CNS injury.

Extracellular Cytosolic Aspartate Aminotransferase Promotes Axonal Growth and Object Recognition Memory.

Previous proteome analysis studies from other groups have shown that cAST might be secreted from neurons and that cAST was detected in human cerebrospinal fluid. However, none of these studies focused on its role or significance. We therefore investigated the role of extracellular cAST for neurons. cAST was detected in conditioned medium from cultured cortical neurons, but not in fresh medium. Recombinant cAST treatment of cortical neurons significantly extended axonal length. Continuous intracerebroventricular administration of recombinant cAST in normal mice for 14 days significantly enhanced object recognition ability. In the brains of those mice, axonal densities and c-Fos expression levels were enhanced, especially in the perirhinal cortex, which mainly relates to object recognition memory. The present study found, for the first time, that extracellular cAST promoted axonal growth function in neurons and activated memory function. These findings indicate a new function of extracellular cAST and may drive the establishment of new therapeutic strategies for cognitive dysfunction. The present study found, for the first time, that extracellular cAST promotes axonal growth in neurons.

Effects of Oleanane-Type Triterpene Saponins from the Leaves of Eleutherococcus senticosus in an Axonal Outgrowth Assay.

An aqueous extract of Eleutherococcus senticosus leaves exerted a beneficial effect in restoring the neurite outgrowth from Aß25-35-induced degeneration using an axonal density assay. Subsequent bioassay-guided fractionation afforded seven new oleanane-type triterpene saponins, ezoukoginosides A-G (1-7), along with nine known analogues. The structures of 1-7 were elucidated through chemical and spectroscopic approaches, and their effects on restoring the neurite outgrowth from Aß25-35-induced degeneration were investigated. The results revealed that hydrophilic oleanane-type saponins substituted with a free carboxylic acid, hydroxy, or formyl group in the aglycone, especially when the oxidation occurred at C-29, not only restrained Aß25-35-induced degeneration but also restored axonal outgrowth significantly. Compounds 2 (-COOH at C-29) and 3 (-CH2OH at C-29) showed the most potent bioactivity among the isolates.

New Age Therapy for Alzheimer's Disease by Neuronal Network Reconstruction.

Alzheimer's disease (AD) is a recognized incurable neurodegenerative disorder. Clinically prescribed medicines for AD are expected to bring about only slight symptomatic improvement or a delay of its progression. Another strategy, amyloid ß (Aß) lowing agents, has not been successful at memory improvement. We have hypothesized that an improvement in cognitive function requires the construction of neuronal networks, including neurite regeneration and synapse formation; therefore, we have been exploring candidates for radical anti-AD drugs that can restore Aß-induced neurite atrophy and memory impairment. Our studies found several promising drug candidates that may improve memory dysfunction in AD model mice. The main activity of these drugs is the restoration of damaged axons. Focusing on candidates based on the recovery of neurite atrophy in vitro certainly leads to positive effects on memory improvement also in vivo. This suggests that neuronal network reconstruction may importantly relate to functional recovery in the brain. When identifying the signaling mechanisms of exogenous compounds like natural medicine-derived constituents, molecules directly activated by the compound are hard to be identified. However, the drug affinity responsive target stability (DARTS) analysis may pave the way to an approach to determine the initial molecule of the signaling pathway. Exploring new drug candidates and clarifying their signaling pathways directly relating to neuronal network reconstruction may provide promising therapeutic strategies with which to overcome AD.

Active Constituents from Drynaria fortunei Rhizomes on the Attenuation of Aß(25-35)-Induced Axonal Atrophy.

Axonal regeneration might contribute to the restoration of damaged neuronal networks and improvement of memory deficits in a murine Alzheimer's disease (AD) model. A search for axonal regenerative drugs was performed to discover novel therapeutic options for AD. In this study, an aqueous extract of Drynaria fortunei rhizomes reversed Aß25-35-induced axonal atrophy in cultured cortical neurons of mice. Bioassay-guided fractionation of this extract led to the isolation and identification of compounds 1-5. Among them, (2S)-neoeriocitrin (2) and caffeic acid 4-O-glucoside (4) showed significant axonal elongation effects on Aß25-35-induced atrophy.

Comparing the effects of kamikihito in Japan and kami-guibi-tang in Korea on memory enhancement: working towards the development of a global study.

Traditional medicine is widely used in East Asia, and studies that demonstrate its usefulness have recently become more common. However, formulation-based studies are not globally understood because these studies are country-specific. There are many types of formulations that have been introduced to Japan and Korea from China. Establishing whether a same-origin formulation has equivalent effects in other countries is important for the development of studies that span multiple countries. The present study compared the effects of same-origin traditional medicine used in Japan and Korea in an in vivo experiment. We prepared drugs that had the same origin and the same components. The drugs are called kamikihito (KKT) in Japan and kami-guibi-tang (KGT) in Korea. KKT (500 mg extract/kg/day) and KGT (500 mg extract/kg/day) were administered to ddY mice, and object recognition and location memory tests were performed. KKT and KGT administration yielded equivalent normal memory enhancement effects. 3D-HPLC showed similar, but not identical, patterns of the detected compounds between KKT and KGT. This comparative research approach enables future global clinical studies of traditional medicine to be conducted through the use of the formulations prescribed in each country.

Synthesis of long-chain fatty acid derivatives as a novel anti-Alzheimer's agent.

In order to develop new drugs for Alzheimer's disease, we prepared 17 fatty acid derivatives with different chain lengths and different numbers and positions of double bonds by using Wittig reaction and stereospecific hydrogenation of triple bonds as key reactions. Among them, (4Z,15Z)-octadecadienoic acid (10) and (23Z,34Z)-heptatriacontadienoic acid (16) showed the most potent neurite outgrowth activities on Aß(25-35)-treated rat cortical neurons, which activities were comparable to that of a positive control, NGF. Both fatty acids 10 and 16 possess two (Z)-double bonds at the n-3 and n-14 positions, which might be important for the neurite outgrowth activity.

Effects of Ashwagandha (roots of Withania somnifera) on neurodegenerative diseases.

Neurodegenerative diseases commonly induce irreversible destruction of central nervous system (CNS) neuronal networks, resulting in permanent functional impairments. Effective medications against neurodegenerative diseases are currently lacking. Ashwagandha (roots of Withania somnifera Dunal) is used in traditional Indian medicine (Ayurveda) for general debility, consumption, nervous exhaustion, insomnia, and loss of memory. In this review, we summarize various effects and mechanisms of Ashwagandha extracts and related compounds on in vitro and in vivo models of neurodegenerative diseases such as Alzheimer's disease and spinal cord injury.

Structure-activity-relationship studies on dihydrofuran-fused perhydrophenanthrenes as an anti-Alzheimer's disease agent.

As an extended study on development of anti-Alzheimer's disease agent, we newly synthesized various dihydrofuran-fused perhydrophenanthrenes via o-quinodimethane chemistry. This study revealed that the introduction of carbon side-chain on 8-position or removal of the acetal moiety on 3-position arose a cytotoxicity on rat cortical neurons. On the other hand, the ethereal or thio-ethereal substituent on 8-position enhanced the elongation effect on Aß-damaged neurons. The necessity of the cyano group on 10b position was also proved in this structure-activity-relationship study.

Axonal Regeneration Mediated by a Novel Axonal Guidance Pair, Galectin-1 and Secernin-1.

Galectin-1 (Gal-1), a member of the Galectin family, is expressed in various tissues and responsible for multiple biological activities. Previous studies reported that extracellular Gal-1 participated in axonal growth and repair, and Gal-1 knockout mice exhibited memory impairment. However, no study has demonstrated the direct contribution of intracellular Gal-1 upregulation in neurons to promoting axonal regeneration in the brain and recovering memory function. In the present study, we found that axonal growth is promoted by overexpression of Gal-1 via adeno-associated virus serotype 9 delivery in primary cultured hippocampal neurons. Moreover, Gal-1 was expressed on the membranes of growth cones in hippocampal neurons and interacted with a novel axonal guidance molecule, Secernin-1, which was secreted from prefrontal cortex (PFC) neurons. Gal-1-overexpression-driven axonal growth was enhanced when recombinant (extracellular) Secernin-1 was treated to the axonal site in a neuron device chamber. Direct binding of extracellular Secernin-1 with Gal-1 was detected through immunoprecipitation and immunocytochemistry, demonstrating that Gal-1 possibly works as an axonal guidance receptor for Secernin-1 in hippocampal neurons. In the PFC, the expression of Gal-1 in axonal shafts and terminals of hippocampal neurons was decreased in the 5XFAD mouse model of Alzheimer's disease (AD). Overexpression of Gal-1 in hippocampal neurons recovered memory deficits and induced axonal regeneration toward the PFC in 5XFAD mice. This study suggests that the enhanced interaction of Secernin-1 and Gal-1 can be harnessed as a therapeutic strategy for long-distance and direction-specific axonal regeneration in AD.

Causal Relationships between Daily Physical Activity, Physical Function, and Cognitive Function Ultimately Leading to Happiness.

Frailty is a common age-related condition linked with mobility disorders, long-term care, and death. To prevent frailty, physical activities are considered effective. Several studies have indicated that physical activity can influence mental health as well as body function. Physical activity, cognitive function, and subjective mental health must relate to each other. However, most studies only focus on one-to-one interactions. This observational study aims to clarify the overall relationship and causality between subjective mental health, daily physical activity, and physical and cognitive functions. We recruited 45 people (24 males and 21 females) over 65 years old. Participants visited the university twice and were subjected to activity measurements at home. To examine the causal relationships and related structures between the indicators, structural equation modeling was performed. The results suggest that daily physical activity explains physical function, physical function explains cognitive function, and cognitive function explains subjective mental health, quality of life, and happiness. This study is the first to clarify interactive relationships as an axis that start from daily physical activity to happiness in older adults. Upregulating daily physical activity may improve physical and cognitive functions as well as mental health; this might protect and ameliorate physical, mental, and social frailties.

Synthesis of dihydrofuran-fused perhydrophenanthrenes having a phenolic hydroxyl group as a novel anti-Alzheimer's disease agent.

As a part of our research program on developing novel anti-Alzheimer's disease medicines, several dihydrofuran-fused perhydrophenanthrenes (DFs) possessing a phenolic hydroxyl group were found to exhibit potent dendritic and axonal regeneration activities. Introduction of a methoxy group into the perhydrophenanthrene skeleton was successfully achieved via a PhI(OAc)(2)-mediated phenolic oxidation of a benzocyclobutene nucleus and subsequent tandem intramolecular electrocyclic reactions based on o-quinodimethane chemistry. We could reveal that a new methoxy derivative having a phenolic hydroxyl group exerted the most significant effects on the dendritic and axonal extensions in the damaged neurons, among DFs examined in this study.

Epimedium koreanum Extract and Its Constituent Icariin Improve Motor Dysfunction in Spinal Cord Injury.

Although cell transplantation strategies for spinal cord injury (SCI) using sources such as iPS cells and neural stem cells are focused as expectative therapies for SCI, the possibility of medication as more accessible and practical way should not be given up. We, therefore, aimed to develop medical sources for SCI. In this paper, we evaluated effects of a famous tonic herb, Epimedium koreanum, on motor dysfunction in spinal cord injury (SCI). The spinal cord was injured by contusion after laminectomy at T10 level. Oral administration of the methanol extract of E. koreanum significantly enhanced hindlimb function in SCI mice by short period treatment (for initial 3 days) and chronic treatment (21 days), although chronic treatment recovered the function more potently. Since it is well known that icariin is the major constituent in E. koreanum, icariin was administered orally to SCI mice for initial 3 days. Motor dysfunction was ameliorated by icariin treatment similarly to the methanol extract of E. koreanum. This paper is the first report to indicate E. koreanum is effective for recovery of motor function in SCI, and at least icariin is an active constituent.

Memory enhancement effect of saponins from Eleutherococcus senticosus leaves and blood-brain barrier-permeated saponins profiling using a pseudotargeted monitoring strategy.

Dried Eleutherococcus senticosus leaves (ESL), also known as Siberian ginseng tea, are beneficial for human neural disorders. Our previous studies showed that the aqueous extract of ESL enhanced memory in mice, and its saponin fraction (ESL-SAP) exhibited promising neuroprotective activities in vitro; however, the in vivo neurally related effect, bioactive material basis, and possible mechanism of action of ESL-SAP have not been investigated. Here, a series of memory and learning tests were carried out, and the results evidenced a significant enhancement effect of ESL-SAP. Furthermore, an in vivo saponin library-guided pseudotargeted strategy was established to support the rapid monitoring of 26 blood-brain barrier (BBB)-permeated saponins from ESL-SAP-administered rats. A further network pharmacology analysis was conducted on BBB-permeated compounds, which indicated that the in vivo mechanism of ESL-SAP might be effective through multiple targets and pathways, such as the AGE-RAGE signaling pathway and PI3K-Akt signaling pathway, to exert neuroprotective effects. Moreover, the molecular docking experiments demonstrated that key BBB-transferred saponins primarily interacted with targets HRAS, MAPK1, and MAPK8 to produce the neuroprotective effect.