ß-asarone induces viability and angiogenesis and suppresses apoptosis of human vascular endothelial cells after ischemic stroke by upregulating vascular endothelial growth factor A.

Ischemic stroke (IS) is a disease with a high mortality and disability rate worldwide, and its incidence is increasing per year. Angiogenesis after IS improves blood supply to ischemic areas, accelerating neurological recovery. ß-asarone has been reported to exhibit a significant protective effect against hypoxia injury. The ability of ß-asarone to improve IS injury by inducing angiogenesis has not been distinctly clarified. The experimental rats were induced with middle cerebral artery occlusion (MCAO), and oxygen-glucose deprivation (OGD) model cells were constructed using human microvascular endothelial cell line (HMEC-1) cells. Cerebral infarction and pathological damage were first determined via triphenyl tetrazolium chloride (TTC) and hematoxylin and eosin (H&E) staining. Then, cell viability, apoptosis, and angiogenesis were assessed by utilizing cell counting kit-8 (CCK-8), flow cytometry, spheroid-based angiogenesis, and tube formation assays in OGD HMEC-1 cells. Besides, angiogenesis and other related proteins were identified with western blot. The study confirms that ß-asarone, like nimodipine, can ameliorate cerebral infarction and pathological damage. ß-asarone can also upregulate vascular endothelial growth factor A (VEGFA) and endothelial nitric oxide synthase (eNOS) and induce phosphorylation of p38. Besides, the study proves that ß-asarone can protect against IS injury by increasing the expression of VEGFA. In vitro experiments affirmed that ß-asarone can induce viability and suppress apoptosis in OGD-mediated HMEC-1 cells and promote angiogenesis of OGD HMEC-1 cells by upregulating VEGFA. This establishes the potential for ß-asarone to be a latent drug for IS therapy.

Effects of ß-asarone on spatial learning and memory impairment by exhaustive exercise-induced fatigue: Role of NR-CaMKII-ERK/CREB signal in hippocampus of rats.

OBJECTIVE: It is to investigate the effects of ß-asarone on learning and memory, hippocampal morphology, synaptophysin (SYP) and postsynaptic density 95(PSD95) protein expression, N-methyl-D-aspartic acid receptor 2B (NR2B)- Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) - Extracellular signal-regulated kinase (ERK) / Cyclic-AMP response element binding protein (CREB) signal in hippocampus of rats with exhaustive exercise-induced fatigue. METHODS: Fifty Sprague-Dawley male rats were randomly divided into five groups: normal group, exercise group, exercise and ß-asarone (2.5, 10, 40 mg/kg)-treated groups. The learning and memory in rats were tested by Morris water maze experiment. We measured the hippocampal morphology by Nissl staining. The levels of SYP, PSD95, NR2B, CaMKII, ERK1/2, CREB, p-NR2B, p-CaMKII, p-ERK1/2 and p-CREB expression were measured by western blot analysis. RESULTS: The results demonstrated that ß-asarone (10, 40 mg/kg) treatment significantly decreased the latency to find the platform, increased the time spent in the target quadrant and the number of crossing the platform of rats with exhaustive exercise-induced fatigue. ß-asarone (10, 40 mg/kg) treatment increased the cell density in the hippocampus CA1 region, significantly up-regulated NR2B-CaMKII-ERK/CREB signal and improved the protein expression levels of SYP and PSD95 in hippocampus of rats with exhaustive exercise-induced fatigue. CONCLUSIONS: It suggests that ß-asarone could improve learning and memory of rats with exhaustive exercise-induced fatigue. The mechanism might be related to ß-asarone protecting the morphology of hippocampus, increasing the protein expression levels of SYP and PSD95 and up-regulating NR2B-CaMKII-ERK/CREB signal in hippocampus of rats with exhaustive exercise-induced fatigue.

Acorus calamus L. rhizome extract and its bioactive fraction exhibits antibacterial effect by modulating membrane permeability and fatty acid composition.

ETHNOPHARMACOLOGICAL RELEVANCE: India's ancient texts, the Charak Samhita and Sushruta Samhita, make reference to the traditional medicinal usage of Acorus calamus L. In India and China, it has long been used to cure stomach aches, cuts, diarrhea, and skin conditions. This ability of the rhizome is attributed to its antimicrobial properties. Research studies to date have shown its antimicrobial properties. However, scientific evidence on its mode of action is still lacking. AIM OF THE STUDY: Acorus calamus L. rhizome extract and its bioactive fraction exhibits antibacterial effect by modulating membrane permeability and fatty acid composition. MATERIAL AND METHOD: The secondary metabolites in the rhizome of A. calamus L. were extracted in hexane using Soxhlet apparatus. The ability of the extract to inhibit multidrug resistant bacterial isolates, namely Bacillus cereus, Escherichia coli, Acinetobacter baumannii, and Pseudomonas aeruginosa were evaluated using checkerboard assay. Further, the extract was purified using thin layer chromatography, gravity column chromatography, and combiflash chromatography. Structure elucidation of the active compound was done using GC-MS, FT-IR, and UV-Vis spectral scan. The mode of action of the bioactive fraction was determined. Bacterial membrane damage was analyzed using SEM, membrane permeability was determined using SYBR green I and PI dye, leakage of cytoplasmic contents were analyzed using Bradford assay and Fehling's reagent. The ability to inhibit efflux pump of A. baumannii was determined using EtBr accumulation assay and ß-lactamase inhibition was analyzed using nitrocefin as substrate. Also, the biofilm inhibition of B. cereus was determined using crystal violet dye. Moreover, the effect of the bioactive fraction on the fatty acid profile of the bacterial membrane was determined by GC-FAME analysis using 37 component FAME mix as standard. RESULTS: Acorus calamus L. rhizome hexane extract (AC-R-H) demonstrated broad-spectrum antibacterial activity against all the isolates tested. AC-R-H extract also significantly reduced the MIC of ampicillin against all tested bacteria, indicating its bacterial resistance modulating properties. The assay guided purification determined Asarone as the major compound present in the bioactive fraction (S-III-BAF). S-III-BAF was found to reduce the MIC of ampicillin against Escherichia coli (100-25 mg/mL), Pseudomonas aeruginosa (15-3.25 mg/mL), Acinetobacter baumannii (12.5-1.56 mg/ml), and Bacillus cereus (10-1.25 mg/mL). Further, it recorded synergistic activity with ampicillin against B. cereus (FICI = 0.365), P. aeruginosa (FICI = 0.456), and A. baumannii (FICI = 0.245). The mode of action of S-III-BAF can be attributed to its ability to disturb the membrane integrity, enhance membrane permeability, reduce biofilm formation, and possibly alter the fatty acid composition of the bacterial cell membranes. CONCLUSION: The bioactive fraction of AC-R-H extract containing Asarone as the active compound showed antibacterial activity and synergistic interactions with ampicillin against the tested bacterial isolates. Such activity can be attributed to the modulation of fatty acids present in bacterial membranes, which enhances membrane permeability and causes membrane damage.

The novel anti-fibrillary effects of volatile compounds α-asarone and ß-caryophyllene on tau protein: Towards promising therapeutic agents for Alzheimer's disease.

The abnormal deposition of tau protein is one of the critical causes of tauopathies including Alzheimer's disease (AD). In recent years, there has been great interest in the use of essential oils and volatile compounds in aromatherapy for treating AD, since volatile compounds can directly reach the brain through intranasal administration. The volatile compounds α-asarone (ASA) and ß-caryophyllene (BCP) have revealed various important neuroprotective properties, useful in treating AD. In this study, the volatile compounds ASA and BCP were assessed for their effectiveness in preventing tau fibrillation, disassembly of pre-formed tau fibrils, and disaggregation of tau aggregates. SDS-PAGE and AFM analyses revealed that ASA and BCP inhibited tau fibrillation/aggregation and decreased the mean size of tau oligomers. Tau samples treated with ASA and BCP, showed a reduction in ThT and ANS fluorescence intensities, and a decrease in the ß-sheet content. Additionally, ASA and BCP disassembled the pre-formed tau fibrils to the granular and linear oligomeric intermediates. Treatment of neuroblastoma SH-SY5Y cells with tau samples treated with ASA and BCP, revealed protective effects as shown by reduced toxicity of the cells, due to the inhibition of tau fibrillation/aggregation. Overall, ASA and BCP appeared to be promising therapeutic candidates for AD.

ß-asarone inhibits autophagy by activating the PI3K/Akt/mTOR pathway in a rat model of depression in Parkinson's disease.

OBJECTIVE: It is unclear whether ß-asarone has a good antidepressant effect and what is the main mechanism in Depression in Parkinson's disease (DPD) model rats. METHODS: In this study, DPD model rats were screened from 6-OHDA induced rats by sucrose preference test (SPT) and forced swimming test (FST). DPD model rats were divided into eight groups: model group, pramipexole group, ß-asarone low-dose group (ß-asarone 7.5 group), ß-asarone medium-dose group (ß-asarone 15 group), ß-asarone high-dose group (ß-asarone 30 group), 3-MA group, rapamycin group, and PI3K inhibitor group. 28 days after the end of treatment, open field test (OFT), SPT and FST were conducted in rats. The level of α-synuclein (α-syn) in the striatum was determined by enzyme-linked immunosorbent assay (ELISA). The expression of Beclin-1, p62 in the striatum was determined by western blot. The expression of PI3K, p-PI3K, Akt, p-Akt, mTOR, p-mTOR, Beclin-1, and p62 in the hippocampus was determined by western blot. The spine density of neurons in the hippocampus was detected by golgi staining. RESULTS: The results showed that 4-week oral administration of ß-asarone improve the motor and depressive symptoms of DPD model rats, and decrease the content of α-syn in the striatum. ß-asarone inhibited the expression of autophagy in the striatum of DPD model rats. Furthermore, ß-asarone decreased the levels of Beclin-1 protein, increased the expression of p62, p-PI3K, p-AKT, and p-mTOR, and improved the density of neuron dendritic spine in the hippocampus. CONCLUSIONS: We concluded that ß-asarone might improve the behavior of DPD model rats by activating the PI3K/Akt/mTOR pathway, inhibiting autophagy and protecting neuron.

Inhibitory effects of ß-asarone on lncRNA BACE1-mediated induction of autophagy in a model of Alzheimer's disease.

The primary aim of this study was to examine the correlation between the formation of Aß plaques and autophagy, which is regulated by ß-asarone and the lncRNA BACE1-AS. Additionally, the study sought to explore potential targets of the drug in inhibiting the deposition of toxic AD-related proteins and restoring impaired mitochondrial and autophagic functions. SHY5Y cells were utilized to construct a stable Alzheimer's disease (AD) model, followed by the utilization of interference and overexpression lentiviruses targeting BACE1-AS to establish a cell model. The cells were categorized into five groups, including a normal group, siRNA/BACE1 group, and ß-asarone group. The fluorescence quantitative PCR technique was employed to assess the disparity in BACE1 mRNA expression, while changes in immunofluorescence (IF) were observed to determine the stable interference titre and action time of the lentiviruses. Additionally, western blotting (WB) and fluorescence quantitative PCR were employed to evaluate the expression of proteins and mRNAs associated with AD and autophagy. The findings demonstrated a significant elevation in BACE1 expression levels in brain tissue among individuals with AD compared to those without the condition. Moreover, the results indicated that the introduction of ß-asarone led to an increase in the expression of the BACE1-AS gene in the cell group transfected with plasmid H12732. Furthermore, it was observed that ß-asarone enhanced the expression levels of shRNA and BACE1 after 72 h. In contrast, ß-asarone suppressed the expression of PS1, Aß, BACE1, APP, and p62, while promoting the expression of syn, LC3 I/II, and Beclin-1. Based on these findings, it can be concluded that ß-Asarone exerts a comprehensive influence on the expression of proteins associated with AD and synaptic function. ß-Asarone exhibits the potential to mitigate Aß deposition by impeding the expression of lncBACE1, thereby facilitating autophagy through the suppression of BACE1's inhibitory impact on autophagy. This complements the self-enhancing effect of autophagy.

Alpha-Asarone Ameliorates Neurological Dysfunction of Subarachnoid Hemorrhagic Rats in Both Acute and Recovery Phases via Regulating the CaMKII-Dependent Pathways.

Early brain injury (EBI) is the leading cause of poor prognosis for patients suffering from subarachnoid hemorrhage (SAH), particularly learning and memory deficits in the repair phase. A recent report has involved calcium/calmodulin-dependent protein kinase II (CaMKII) in the pathophysiological process underlying SAH-induced EBI. Alpha-asarone (ASA), a major compound isolated from the Chinese medicinal herb Acorus tatarinowii Schott, was proven to reduce secondary brain injury by decreasing CaMKII over-phosphorylation in rats' model of intracerebral hemorrhage in our previous report. However, the effect of ASA on SAH remains unclear, and the role of CaMKII in both acute and recovery stages of SAH needs further investigation. In this work, we first established a classic SAH rat model by endovascular perforation and intraperitoneally administrated different ASA doses (10, 20, and 40 mg/kg) 2 h after successful modeling. Then, the short- and long-term neurobehavioral performances were blindly evaluated to confirm ASA's efficacy against SAH. Subsequently, we explored ASA's therapeutic mechanism in both acute and recovery stages using histopathological examination, TUNEL staining, flow cytometry, Western-blot, double-immunofluorescence staining, and transmission electron microscopy (TEM) observation. Finally, KN93, a selective CaMKII inhibitor, was applied in oxyhemoglobin-damaged HT22 cells to explore the role of CaMKII in ASA's neuroprotective effect. The results demonstrated that ASA alleviated short- and long-term neurological dysfunction, reduced mortality and seizure rate within 24 h, and prolonged 14-day survival in SAH rats. Histopathological examination showed a reduction of neuronal damage and a restoration of the hippocampal structure after ASA treatment in both acute and recovery phases of SAH. In the acute stage, the Western-blot and flow cytometer analyses showed that ASA restored E/I balance, reduced calcium overload and CaMKII phosphorylation, and inhibited mitochondrion-involved apoptosis, thus preventing neuronal damage and apoptosis underlying EBI post-SAH. In the recovery stage, the TEM observation, double-immunofluorescence staining, and Western-blot analyses indicated that ASA increased the numbers of synapses and enhanced synaptic plasticity in the ipsilateral hippocampi, probably by promoting NR2B/CaMKII interaction and activating subsequent CREB/BDNF/TrkB signaling pathways. Furthermore, KN93 notably reversed ASA's neuroprotective effect on oxyhemoglobin-damaged HT22 cells, confirming CaMKII a potential target for ASA's efficacy against SAH. Our study confirmed for the first time that ASA ameliorated the SAH rats' neurobehavioral deterioration, possibly via modulating CaMKII-involved pathways. These findings provided a promising candidate for the clinical treatment of SAH and shed light on future drug discovery against SAH.

ß-asarone improves cognitive impairment and alleviates autophagy in mice with vascular dementia via the cAMP/PKA/CREB pathway.

BACKGROUND: Vascular dementia (VD) is the second most common type of dementia after Alzheimer's disease. ß-asarone, a major component of Acorus tatarinowii Schott, is important in neurodegenerative and neurovascular diseases. Studies have confirmed that ß-asarone can mitigate autophagy and reduce damage in hypoxic cells. We also reported that ß-asarone improves learning and memory. This study further clarifies whether ß-asarone attenuates cerebral ischaemic injury by acting through the cAMP/PKA/CREB pathway in VD model mice. METHODS: Here, genes and potential pathways that may be targeted by ß-asarone for the treatment of transient cerebral ischaemia (TCI) and cognitive impairment (CI) were obtained using network pharmacology. The two-vessel occlusion method was used to establish the VD model. The Morris water maze test was used to evaluate the effects on memory. Then, the protein levels of mitofusin-2 (Mfn2), brain-derived neurotrophic factor (BDNF), optic atrophy 1 (OPA1), cyclic adenosine monophosphate (cAMP), myelin basic protein (MBP), matrix metalloproteinase-9 (MMP9) and neuron specific enolase (NSE) were determined by ELISA. The levels of superoxide dismutase (SOD) and malonaldehyde (MDA) were measured using commercial kits. Then, qRT-PCR was employed to investigate the expression of the candidate genes screened from the protein-protein interaction (PPI) network. Furthermore, the expression of the autophagy-related proteins Beclin-1, (microtubule-associated protein light chain 3) LC3, p62, postsynaptic density protein 95 (PSD95), protein kinase A (PKA), pPKA, cyclic-AMP response binding protein (CREB), and pCREB was determined by western blotting. The expression of autophagy-related proteins, PSD95 and translocase of outer mitochondrial membrane 20 (TOM20) was determined by immunofluorescence analyses. RESULTS: The network pharmacological analysis showed 234 targets related to ß-asarone, 1,118 genes related to TCI and 2,039 genes associated with CI. Our results confirm that ß-asarone treatment not only alleviated brain damage in the VD model by improving mitochondrial and synaptic function, reducing neuronal injury and upregulating the expression of antioxidants but also effectively improved the cognitive behaviour of VD model mice. Moreover, ß-asarone downregulated VD-induced RELA and CCND1 mRNA expression. In addition, we validated that ß-asarone increased the phosphorylation of PKA and CREB and upregulated cAMP protein expression. The results showed that the cAMP/PKA/CREB signalling pathway was upregulated. Moreover, ß-asarone administration decreased the protein expression levels of Beclin-1 and LC3 and increased the expression levels of p62 in VD model mice. CONCLUSIONS: ß-asarone inhibits Beclin-1-dependent autophagy and upregulates the cAMP/PKA/CREB signalling pathway to attenuate mitochondrial and synaptic damage from cerebral ischaemia and improve learning and cognitive abilities in VD model mice.

Inhibitors of 3-Hydroxy-3-methylglutaryl Coenzyme A Reductase Decrease the Growth, Ergosterol Synthesis and Generation of petite Mutants in Candida glabrata and Candida albicans.

Candida glabrata and Candida albicans, the most frequently isolated candidiasis species in the world, have developed mechanisms of resistance to treatment with azoles. Among the clinically used antifungal drugs are statins and other compounds that inhibit 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), resulting in decreased growth and ergosterol levels in yeasts. Ergosterol is a key element for the formation of the yeast cell membrane. However, statins often cause DNA damage to yeast cells, facilitating mutation and drug resistance. The aim of the current contribution was to synthesize seven series of compounds as inhibitors of the HMGR enzyme of Candida ssp., and to evaluate their effect on cellular growth, ergosterol synthesis and generation of petite mutants of C. glabrata and C. albicans. Compared to the reference drugs (fluconazole and simvastatin), some HMGR inhibitors caused lower growth and ergosterol synthesis in the yeast species and generated fewer petite mutants. Moreover, heterologous expression was achieved in Pichia pastoris, and compounds 1a, 1b, 6g and 7a inhibited the activity of recombinant CgHMGR and showed better binding energy values than for α-asarone and simvastatin. Thus, we believe these are good candidates for future antifungal drug development.

Purple perilla frutescens extracts containing α-asarone inhibit inflammatory atheroma formation and promote hepatic HDL cholesterol uptake in dyslipidemic apoE-deficient mice.

BACKGROUND/OBJECTIVES: Dyslipidemia causes metabolic disorders such as atherosclerosis and fatty liver syndrome due to abnormally high blood lipids. Purple perilla frutescens extract (PPE) possesses various bioactive compounds such as α-asarone, chlorogenic acid and rosmarinic acid. This study examined whether PPE and α-asarone improved dyslipidemia-associated inflammation and inhibited atheroma formation in apolipoprotein E (apoE)-deficient mice, an experimental animal model of atherosclerosis. MATERIALS/METHODS: ApoE-deficient mice were fed on high cholesterol-diet (Paigen's diet) and orally administrated with 10-20 mg/kg PPE and α-asarone for 10 wk. RESULTS: The Paigen's diet reduced body weight gain in apoE-deficient mice, which was not restored by PPE or α-asarone. PPE or α-asarone improved the plasma lipid profiles in Paigen's diet-fed apoE-deficient mice, and despite a small increase in high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein (LDL)-cholesterol, and very LDL were significantly reduced. Paigen's diet-induced systemic inflammation was reduced in PPE or α-asarone-treated apoE-deficient mice. Supplying PPE or α-asarone to mice lacking apoE suppressed aorta atherogenesis induced by atherogenic diet. PPE or α-asarone diminished aorta accumulation of CD68- and/or F4/80-positive macrophages induced by atherogenic diet in apoE-deficient mice. Treatment of apoE-deficient mice with PPE and α-asarone resulted in a significant decrease in plasma cholesteryl ester transfer protein level and an increase in lecithin:cholesterol acyltransferase reduced by supply of Paigen's diet. Supplementation of PPE and α-asarone enhanced the transcription of hepatic apoA1 and SR-B1 reduced by Paigen's diet in apoE-deficient mice. CONCLUSIONS: α-Asarone in PPE inhibited inflammation-associated atheroma formation and promoted hepatic HDL-C trafficking in dyslipidemic mice.

Ethnic, Botanic, Phytochemistry and Pharmacology of the Acorus L. Genus: A Review.

The genus Acorus, a perennial monocotyledonous-class herb and part of the Acoraceae family, is widely distributed in the temperate and subtropical zones of the Northern and Southern Hemispheres. Acorus is rich in biological activities and can be used to treat various diseases of the nervous system, cardiovascular system, and digestive system, including Alzheimer's disease, depression, epilepsy, hyperlipidemia, and indigestion. Recently, it has been widely used to improve eutrophic water and control heavy-metal-polluted water. Thus far, only three species of Acorus have been reported in terms of chemical components and pharmacological activities. Previously published reviews have not further distinguished or comprehensively expounded the chemical components and pharmacological activities of Acorus plants. By carrying out a literature search, we collected documents closely related to Acorus published from 1956 to 2022. We then performed a comprehensive and systematic review of the genus Acorus from different perspectives, including botanical aspects, ethnic applications, phytochemistry aspects, and pharmacological aspects. Our aim was to provide a basis for further research and the development of new concepts.

Plant Extracts from the Yucatan Peninsula in the In Vitro Control of Curvularia lunata and Antifungal Effect of Mosannona depressa and Piper neesianum Extracts on Postharvest Fruits of Habanero Pepper.

Plant extracts are a valuable alternative for the control of phytopathogenic fungi in horticultural crops. In the present work, the in vitro antifungal effect of ethanol and aqueous extracts from different vegetative parts of 40 native plants of the Yucatan Peninsula on Curvularia lunata ITC26, a pathogen of habanero pepper (Capsicum chinense), and effects of the most active extracts on postharvest fruits were investigated. Among these, the ethanol extracts of Mosannona depressa (bark from stems and roots) and Piper neesianum (leaves) inhibited 100% of the mycelial growth of C. lunata. The three extracts were partitioned between acetonitrile and n-hexane. The acetonitrile fraction from M. depressa stem bark showed the lowest mean inhibitory concentration (IC50) of 188 µg/mL against C. lunata. The application of this extract and its active principle α-asarone in the postharvest fruits of C. chinense (500 µg/mL) was shown to inhibit 100% of the severity of the infection caused by C. lunata after 11 days of contact. Both samples caused the distortion and collapse of the conidia of the phytopathogen when observed using electron microscopy at 96 h. The spectrum of M. depressa enriched antifungal action is a potential candidate to be a botanical fungicide in the control of C. lunata in cultivating habanero pepper.

Nose to brain delivery of Astragaloside IV by ß-Asarone modified chitosan nanoparticles for multiple sclerosis therapy.

Multiple sclerosis (MS), an autoimmune disease, has been considered an inflammatory disorder of the central nervous system (CNS) with demyelination and axonal damage. Although there are certain first-line therapies to treat MS, their unsatisfactory efficacy is partly due to the limited CNS access after systemic administration. Besides, there is an urgent need to treat MS by enhancing remyelination or neuroprotection, or dampen the activity of microglia. Astragaloside IV (ASI) bears anti-inflammatory, antioxidant, remyelination and neuroprotective activity. While its poor permeability, relatively high molecular weight and low lipophilicity restrict it to reach the brain. Therefore, ß-asarone modified ASI loaded chitosan nanoparticles (ASI-ßCS-NP) were prepared to enhance the nose-to-brain delivery and therapeutic effects of ASI on EAE mice. The prepared ASI-ßCS-NP showed mean size of about 120 nm, and zeta potential from +19 to +25 mV. DiR-ßCS-NP was confirmed with good nose-to-brain targeting ability. After intranasal administration, the ASI-ßCS-NP significantly reduced behavioral scores, decreased weight loss, suppressed inflammatory infiltration and astrocyte/microglial activation, reduced demyelination and increased remyelination on a mice EAE model. Our findings indicate that ASI-ßCS-NP may be a potent treatment for MS after nose-to-brain drug delivery.

Synergetic effect of ß-asarone and cannabidiol against Aß aggregation in vitro and in vivo.

Alzheimer's disease (AD) is a complex and multifactorial neurodegenerative disorder, and it is unlikely that any single drug or intervention will be very successful. The pathophysiology of Alzheimer's disease involves a range of complicated biological processes, including the accumulation of beta-amyloid protein and tau protein. Given the complexity of AD and amyloid accumulation, a combination of interventions remains to be further explored. Here, we investigated the potential of combining ß-asarone and cannabidiol (CBD) as a treatment for AD. The study analyzed the combined effects of these two phytochemicals on beta-amyloid (Aß) protein aggregation and toxicity in bulk solution, in cells as well as in C.elegans. We detailed the morphological and size changes of Aß40 aggregates in the presence of ß-asarone and cannabidiol. More importantly, the presence of both compounds synergistically inhibited apoptosis and downregulated relative gene expression in cells, and that it may also slow aging, decrease the rate of paralysis, enhance learning capacity, and boost autophagy activity in C.elegans. Our studies suggest that multiple drugs, like ß-asarone and CBD, may be potentially developed as a medicinal adjunct in the treatment of AD, although further clinical trials are needed to determine the efficacy and safety of this combination treatment in humans.

ß-Asarone Alleviates High-Glucose-Induced Oxidative Damage via Inhibition of ROS Generation and Inactivation of the NF-κB/NLRP3 Inflammasome Pathway in Human Retinal Pigment Epithelial Cells.

Diabetic retinopathy (DR) is the leading cause of vision loss and a major complication of diabetes. Hyperglycemia-induced accumulation of reactive oxygen species (ROS) is an important risk factor for DR. ß-asarone, a major component of volatile oil extracted from Acori graminei Rhizoma, exerts antioxidant effects; however, its efficacy in DR remains unknown. In this study, we investigated whether ß-asarone inhibits high-glucose (HG)-induced oxidative damage in human retinal pigment epithelial (RPE) ARPE-19 cells. We found that ß-asarone significantly alleviated cytotoxicity, apoptosis, and DNA damage in HG-treated ARPE-19 cells via scavenging of ROS generation. ß-Asarone also significantly attenuated the excessive accumulation of lactate dehydrogenase and mitochondrial ROS by increasing the manganese superoxide dismutase and glutathione activities. HG conditions markedly increased the release of interleukin (IL)-1ß and IL-18 and upregulated their protein expression and activation of the nuclear factor-kappa B (NF-κB) signaling pathway, whereas ß-asarone reversed these effects. Moreover, expression levels of the NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome multiprotein complex molecules, including thioredoxin-interacting protein, NLRP3, apoptosis-associated speck-like protein containing a caspase-recruitment domain, and cysteinyl aspartate-specific proteinase-1, were increased in ARPE-19 cells under HG conditions. However, their expression levels remained similar to those in the control group in the presence of ß-asarone. Therefore, ß-asarone protects RPE cells from HG-induced injury by blocking ROS generation and NF-κB/NLRP3 inflammasome activation, indicating its potential as a therapeutic agent for DR treatment.

Concept of dementia (Nisy a n) in Unani system of medicine and scientific validation of an important Unani pharmacopoeial preparation 'Majoon Vaj' for its management: a review.

OBJECTIVES: This review focused on the concept of dementia in the Unani system of medicine and comprehensive, updated information on Majoon Vaj about the phytochemistry, nootropic, CNS activities and provide insights into potential opportunities for future research. METHODS: The classical literature on Majoon Vaj for its anti-dementic properties, and therapeutic uses were gathered from nearly thirteen classical Unani books including Unani Pharmacopoeia. The information of pharmacognosy, phytochemical and pharmacological activities of Majoon Vaj and its ingredient was collected by browsing the Internet (PubMed, ScienceDirect, Wiley online library, Google Scholar, ResearchGate). The relevant primary sources were probed, analysed, and included in this review. The keywords used to browse were Majoon Vaj, Dementia, Nootropic, Acorus calamus, Piper nigram, Zingiber officinalis, Nigella sativa, Carum carvi, Plumbago zeylanica, and ß-asarone. Relevant Sources were gathered up to July 2021, and the chemical structures were drawn using ACD/ChemSketch software. The species name and synonyms were checked with WFO (2021): World Flora online (http://www.worldfloraonline.org) an updated version of 'The Plant List.' RESULTS: Majoon Vaj contains an excess of bioactive compounds e.g., alkaloids, phenols, flavonoids, tannins, diterpenes, coumarins, carbohydrates, and fixed oils and its ingredients possess broad pharmacological properties, including cognitive-enhancing, neuroprotective, anti-inflammatory, antioxidant and antimicrobial properties. CONCLUSIONS: The literature of Unani medicine is quite rich in discussing the pathophysiological basis of memory disorders. It argues that memory, retention, and retrieval are regulated by a complex process involving various faculties. Majoon Vaj seems to have great potential for therapeutic applications in the treatment of dementia and thus encourage more preclinical and clinical trials in this field.

ß-asarone inhibits the migration, invasion, and EMT of bladder cancer through activating ER stress.

BACKGROUND: ß-asarone (ß-as), a compound extracted from Acorus calamus, has been found to have anticancer effects on a variety of human cancers. However, the potential effect of ß-as on bladder cancer (BCa) remains unknown. METHODS: After exposure to ß-as, migration, invasion, and epithelial-mesenchymal transition (EMT) of BCa were determined by wound healing, transwell, and Western blot assays. Expression of proteins involved in the EMT and ER stress were explored by Western blot assays. Nude mouse xenograft model was served as the model system in vivo. RESULTS: The migration, invasion, and EMT of BCa were significantly inhibited after ß-as treatment. Further experiments revealed that endoplasmic reticulum (ER) stress is involved in ß-as-mediated metastasis inhibition. In addition, ß-as significantly up-regulated activating transcription factor 6 (ATF6), a branch of ER stress, and promoted its Golgi cleavage and nuclear localization. ATF6 silencing attenuated ß-as-mediated metastasis and EMT inhibition in BCa cells. CONCLUSION: Our data suggests that ß-as inhibits migration, invasion, and EMT of BCa by activating the ATF6 branch of ER stress. Thus, ß-as represents a potential candidate for BCa treatment.

Isolation, identification, and characterization of α- asarone, from hydromethanolic leaf extract of Acorus calamus L. and its apoptosis-inducing mechanism in A549 cells.

Due to the presence of several active secondary metabolites, the traditional Indian and Chinese medicinal herb Acorus calamus L. has been utilized for both medical and culinary purposes since ancient times. A recent report has underscored the promising cytotoxic effect of A. calamus leaves extract against non-small cell lung cancer A549 cells. Thus, we want to separate the bioactive substance from the hydromethanolic extract of A. calamus leaves in the current investigation. Thin-layer chromatography was used to separate the compounds and different spectroscopic methods (UV, FTIR, NMR, and LCMS/MS) were used for the structure prediction. α-asarone was found to be the main bioactive compound present and it was isolated from A. calamus leaves extract. It exerted a good cytotoxic effect with an IC50 value of 21.43 ± 1.27 µM against A549 cells and IC50 value of 324.12 ± 1.32 µM against WI-38 cells. The induction of apoptosis in A549 cells by α-asarone was reaffirmed by the diverse differential staining methods including DAPI, Acridine Orange/Ethidium Bromide, and Giemsa staining. Additionally, α-asarone induced mitochondrial membrane potential (ΔΨm) dissipation with a concomitant increase in the production of ROS. Furthermore, it also increased expressions of caspase-3, caspase-9, caspase-8, DR4, and DR5 genes in A549 cells. In conclusion, α-asarone-induced apoptotic cell death in non-small lung cancer cells (A549) as a result of loss of mitochondrial function, increased ROS production, subsequent activation of an internal and extrinsic caspase pathway, and altered expression of genes controlling apoptosis. As a whole, α-asarone is a plausible therapeutic agent for managing lung cancer. HIGHLIGHTSIsolation of bioactive compound from hydromethanolic leaves extract of Acorus calamus L. by thin layer chromatography.Structural elucidation of the bioactive compound was carried out using different methods like UV analysis, FTIR, NMR, and LC-MS/MS analysis.A plausible mode of action revealed that α-asarone can induce apoptosis in lung cancer cells (A549).Communicated by Ramaswamy H. Sarma.

ß-asarone prolongs sleep via regulating the level of glutamate in the PVN.

People of all ages could suffer from sleep disorders, which are increasingly recognized as common manifestations of neurologic disease. Acorus tatarinowii is a herb that has been used in traditional medicine to promote sleep. ß-asarone, as the main component of volatile oil obtained from Acorus tatarinowii, may be the main contributor to the sleeping-promoting efficacy of Acorus tatarinowii. In the study, adult male C57BL/6 mice were administered ß-asarone at 12.5 mg/kg, 25 mg/kg, and 50 mg/kg. Behavioral experiments showed that ß-asarone at 25 mg/kg could significantly improve sleep duration. It was also observed that the proportion of NREM (Non-Rapid Eye Movement) sleep increased considerably after administration of ß-asarone. In the PVN (paraventricular nucleus of hypothalamus) region of the hypothalamus, it was observed that the glutamate content decreased after ß-asarone treatment. At the same time, the expression of VGLUT2 (vesicular glutamate transporters 2) decreased while the expression of GAD65 (glutamic acid decarboxylase 65) and GABARAP (GABA Type A Receptor-Associated Protein) increased in the hypothalamus, suggesting that ß-asarone may suppress arousal by reducing glutamate and promoting transformation of glutamate to the inhibitory neurotransmitter GABA (γ-aminobutyric acid). This study is the first to focus on the association between ß-asarone and sleep, shedding perspectives for pharmacological applications of ß-asarone and providing a new direction for future research.

Network pharmacology-based prediction and validation of the active ingredients and potential mechanisms of the Huangxiong formula for treating ischemic stroke.

ETHNOPHARMACOLOGICAL RELEVANCE: Huangxiong Formula (HXF) is composed of four herbs: Rheum palmatum L., Ligusticum striatum DC., Curcuma aromatica Salisb., and Acorus gramineus Aiton. HXF is clinically used for the treatment of ischemic stroke (IS). However, its molecular mechanism remains unclear. AIM OF THE STUDY: A network pharmacology-based strategy combined with experimental study in vivo and in vitro to were used to investigate the bioactive components, potential targets, and molecular mechanisms of HXF in the treatment of IS. MATERIALS AND METHODS: The components of HXF were detected by ultra-performance liquid chromatography (UPLC). The potential active ingredients of HXF were acquired from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) and literature, and corresponding targets were discerned through the Swiss TargetPrediction database. IS-related targets were obtained from Genecards, Online Mendelian Inheritance in Man (OMIM), Therapeutic Target Database (TTD), and DisGeNET. The intersection of ingredient and disease targets was screened, and a herbal-compound-target network was constructed. A protein-protein interaction (PPI) network was created, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed. Based on these analyses, we established a compound-target-pathway (C-T-P) network. A cerebral ischemia-reperfusion (I/R) animal model was established, and the cerebral protective effect of HXF was assessed. The accuracy of the predicted targets was verified by real-time quantitative polymerase chain reaction (RT-qPCR). Hippocampal neuronal injury cell model induced by oxygen-glucose deprivation and reperfusion (OGD/R) was used to evaluate the protective effect of α-Asarone. Furthermore, molecular docking, drug affinity responsive target stability (DARTS) assay, and cellular thermal shift assay (CETSA) were performed to verify whether α-Asarone can bind to PI3K. RESULTS: A total of 44 active ingredients and 795 gene targets were identified through network pharmacology. Network analysis showed that naringenin, eupatin, kaempferol, and α-Asarone were possible drug candidates. SRC, AKT1, TP53, MAPK3, STAT3, HRAS, CTNNB1, EGFR, VEGFA, PIK3R1 could serve as potential drug targets. KEGG analysis implied that the PI3K/AKT signaling pathway might play an important role in treating IS by HXF. Moreover, HXF significantly reduced neurological impairment, cerebral infarct volume, brain index, and brain histopathological damage in I/R rats. The mRNA expression of the top 10 potential targets was verified in the brain tissue. The C-T-P network and UPLC analysis suggested that α-Asarone might be an important component of HXF and can inhibit oxidative stress and apoptosis in HT22 cells by activating the PI3K/AKT signaling pathway. Molecular docking, DARTS, and CETSA assay analysis confirmed that there were direct interactions between α-Asarone and PI3K. CONCLUSION: HXF had a therapeutic effect in IS with multi-component, multi-target, and multi-approach features. α-Asarone, identified as one of the major active components of HXF, could alleviate oxidative stress and apoptosis by targeting PI3K/AKT pathway.