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.

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.

Mechanistic evaluation of myristicin on apoptosis and cell cycle regulation in breast cancer cells.

The current study was focused on the anticancer activity of myristicin against MCF-7 human breast cancer (BC) cells. BC is the most common and leading malignant disease in women worldwide. Now-a-days, various conventional therapies are used against BC and still represent a chief challenge because those treatments fail to differentiate normal cells from malignant cells, and they have severe side effects also. So, there is a need develop new therapies to decrease BC-related morbidity and mortality. Myristicin, a 1­allyl­5­methoxy­3, 4­methylenedioxybenzene, is a main active aromatic compound present in various spices, such as nutmeg, mace, carrot, cinnamon, parsely and some essential oils. Myristicin has a wide range of effects, including antitumor, antioxidative and antimicrobial activity. Nevertheless, the effects of myristicin on human BC cells remain largely unrevealed. The cytotoxicity effect of myristicin on MCF­7 cells was increased dose dependently detected by (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and Lactate Dehydrogenase assays. Myristicin was found to be significantly inducing the cell apoptosis, as compared to control, using acridine orange/ethidium bromide, Hoechst stain and annexin V. Moreover, it activates cell antimigration, intracellular reactive oxygen species generation and cell cycle arrest in the G1/S phase. In addition, myristicin induces the expression of apoptosis and cell cycle genes (Caspases8, Bax, Bid, Bcl2, PARP, p53, and Cdk1) was demonstrated by quantitative polymerase chain reaction and apoptosis proteins (c-PARP, Caspase 9, Cytochrome C, PDI) expression was also analyzed with western blot. Overall, we illustrated that myristicin could regulate apoptosis signaling pathways in MCF-7 BC cells.

Transcriptomic and biochemical analyses revealed antifungal mechanism of trans-anethole on Aspergillus flavus growth.

Plant volatile compounds have great potential for preventing and controlling fungal spoilage in post-harvest grains. Recently, we have reported the antifungal effects of trans-anethole, the main volatile constituent of the Illicium verum fruit, on Aspergillus flavus. In this study, the inhibitory mechanisms of trans-anethole against the growth of A. flavus mycelia were investigated using transcriptomic and biochemical analyses. Biochemical and transcriptomic changes in A. flavus mycelia were evaluated after exposure to 0.2 µL/mL trans-anethole. Scanning electron microscopy showed that trans-anethole treatment resulted in the surface wrinkling of A. flavus mycelia, and calcofluor white staining confirmed that trans-anethole treatment disrupted the mycelial cell wall structure. Annexin V-fluorescein isothiocyanate/propidium iodide double staining suggested that trans-anethole induced apoptosis in A. flavus mycelia. Reduced mitochondrial membrane potential and DNA damage were observed in trans-anethole-treated A. flavus mycelia using 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanine and 4',6-diamidino-2-phenylindole staining, respectively. 2',7'- Dichloro-dihydro-fluorescein diacetate staining and biochemical assays demonstrated that trans-anethole treatment cause the accumulation of reactive oxygen species in the A. flavus mycelia. Transcriptome results showed that 1673 genes were differentially expressed in A. flavus mycelia exposed to trans-anethole, which were mainly associated with multidrug transport, oxidative phosphorylation, citric acid cycle, ribosomes, and cyclic adenosine monophosphate signaling. We propose that trans-anethole can inhibit the growth of A. flavus mycelia by disrupting the cell wall structure, blocking the multidrug transport process, disturbing the citric acid cycle, and inducing apoptosis. This study provides new insights into the inhibitory mechanism of trans-anethole on A. flavus mycelia and will be helpful for the development of natural fungicides. KEY POINTS: • Biochemical analyses of A. flavus mycelia exposed to trans-anethole were performed • Transcriptomic changes in trans-anethole-treated A. flavus mycelia were analyzed • An inhibitory mechanism of trans-anethole on the growth of A. flavus mycelia was proposed.

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.

Anti-amnesic and anti-cholinesterase activities of α-asarone against scopolamine-induced memory impairments in rats.

OBJECTIVE: Alzheimer's disease (AD) is a neurological ailment that causes memory loss and impairments and is linked to a drop-in acetylcholine level. Acetylcholinesterase (AChE) inhibitors are used for the management of AD. In our ongoing research to search for natural AChE inhibitors from medicinal plants, we found that the Acorus calamus possesses memory-enhancing properties. α-Asarone is the major compound isolated from the Acorus calamus and it has neuroprotective action in animal models, nonetheless, its anticholinesterase activity in different brain regions was not fully understood. The purpose of this research was to determine the anti-amnesic and anti-cholinesterase activities of α-asarone against scopolamine-induced memory impairments in rats. MATERIALS AND METHODS: The anti-cholinesterase activity of α-asarone was determined using Ellman's method in different brain areas, such as the cortex, hippocampus, and striatum. In addition, the anti-amnesic effect of α-asarone was also investigated using elevated plus-maze, passive avoidance, and active avoidance tests. RESULTS: The effect of α-asarone on memory impairment against scopolamine-induced (1 mg/kg body weight) amnesia was evaluated. Administration of α-asarone (15 and 30 mg/kg body weight) for 14 days to rats significantly ameliorated the scopolamine-induced memory impairment as measured in the elevated plus-maze, passive avoidance, and avoidance active tests compared to the scopolamine-treated group. In this study, we also show that α-asarone treatment significantly (p<0.05) reduced brain acetylcholinesterase activity in the cortex, hippocampus, and striatum brain regions of amnesic rats. CONCLUSIONS: These results confirmed that α-asarone has anti-amnesic and anti-cholinesterase potential which may be useful for the management of AD.

Anticonvulsant Activity of trans-Anethole in Mice.

Epilepsy is a chronic neurological disorder affecting 1-2% of world population, and one-third of patients are refractory to pharmacological treatment. This fact has stimulated research for new antiepileptic drugs and natural products have been an important source. trans-Anethole (TAN) is a phenylpropanoid, component of some essential oils, extracted from plants, and its effects have been little studied. Therefore, this study is aimed at investigating the TAN effect in classic seizure models and evaluate the electroencephalographic (EEG) profile of animals treated with this substance. For this, Swiss male mice (Mus musculus) were used, and the lethal dose was evaluated and subsequently submitted to the test maximal electroshock (MES), the pentylenetetrazole- (PTZ) induced seizure test, and the EEG profile. Initially, the LD50 for TAN was estimated in 1000 mg/kg (i.p.) dose and there was no sign of acute toxicity or death. In the MES test, TAN 300, i.p. (12.00 ± 2.9 s) and 400 mg/kg, i.p. (9.00 ± 4.4 s) doses was able to decrease tonic seizures duration induced by electric discharge (0.5 mA, 150 pulses/s, for 0.5 s). In the PTZ test (75 mg/kg, i.p.), TAN 400 mg/kg, i.p. increased the latency to myoclonic jerks (80.0 (56.0-134.0)), the latency totonic-clonic seizures (900.0 (861.0-900.0) and decrease seizure duration (0.0 (0.0-10.0)). No deaths were found in this groups compared to vehicle. EEG analysis showed an amplitude decrease of waves (ratio of baseline) in TAN 300 (1.82 ± 0.23) and 400 mg/kg (1.06 ± 0.16) groups. In this way, TAN at 400 mg/kg was able to inhibit and/or attenuate seizures by increasing the time for the onset of spasms and convulsions, as reducing the duration of seizures. The EEG profile corroborate with this results showing a reduction in the amplitude of waves compared to the PTZ group. Thus, TAN showed an anticonvulsant effect in all experimental models performed, behavioral and electroencephalographic.

Anethole's effects against myocardial infarction: The role of TLR4/NFκB and Nrf2/HO1 pathways.

BACKGROUND: Exploring new drugs for the management of myocardial infarction (MI) is crucial, as MI is a major contributor to mortality worldwide. Anethole, a naturally occurring essential oil component, has numerous medicinal, pharmaceutical, and cosmetic purposes. This study explored the potential action of anethole to protect myocytes against MI injure. METHODS: Wistar rats were divided into five groups: normal; anethole; and isoproterenol (ISO) groups in addition to two groups of ISO + anethole (125 and 250 mg/kg). All anethole groups were administered the oil component for 30 days, and all ISO groups were challenged with ISO on the 28th and 29th days. Parameters measured included infracted area, ECG, cardiac markers, the expression of Keap 1, nuclear Nrf2, and heme oxygenase-1, as well as the expression of TLR4 and MYD88 together with subsequent downstream oxidative stress, inflammatory, and apoptotic markers. RESULTS: Anethole reduced infarct region, degenerated cardiac indicators levels, amended ECG alterations, and diminished myocardial necrosis. Anethole reduced Keap-1, activated Nrf2/HO-1 pathway, increased mitochondrial antioxidant enzyme activities, declined the TLR4/MYD88 pathway, and ameliorated myocardial inflammation and cell death markers. CONCLUSION: Anethole may retain a cardio-protective potential by controlling myocardial oxidative stress (through Nrf2 pathway) and diminishing inflammation and apoptosis via the TLR4/MYD88 pathway.

Identification and Functional Analysis of Differentially Expressed Genes in Myzus persicae (Hemiptera: Aphididae) in Response to Trans-anethole.

Plant essential oils, with high bioactivity and biodegradability, provide promising alternatives to synthetic pesticides for pest control. Trans-anethole is the major component of essential oil from star anise, Illicium verum Hook. The compound has a strong contact toxicity against the green peach aphid, Myzus persicae (Sulzer) (Hemiptera: Aphididae), which is a major insect pest of many vegetables and crops. However, little information is known about how M. persicae responds to trans-anethole at the molecular level. We conducted a comparative transcriptome analysis of M. persicae in response to a LD50 dose of trans-anethole. A total of 559 differentially expressed genes were detected in the treated individuals, with 318 genes up-regulated, and 241 genes down-regulated. Gene ontology (GO) analysis revealed that these genes were classified into different biological processes and pathways. We also found that genes encoding ATP-binding cassette (ABC) transporters, DnaJ, and cuticle proteins were dramatically up-regulated in response to trans-anethole. To study the function of these genes, we performed RNA interference (RNAi) analysis. Knockdown of an ABC transporter gene (ABCG4) and a DnaJ gene (DnaJC1) resulted in a significantly increased mortality rate in M. persicae following trans-anethole exposure, indicating the involvement of these two genes in the toxicity response to trans-anethole. The findings provide new insights into the mechanisms of M. persicae in coping with plant essential oils.

Anti-oxidative and anti-hyperglycemic properties of Agastache foeniculum essential oil and oily fraction in hyperglycemia-stimulated and lipopolysaccharide-stimulated macrophage cells: In vitro and in silico studies.

ETHNOPHARMACOLOGICAL RELEVANCE: Hyperglycemia (HG) and lipopolysaccharide (LPS) often promote superoxide accumulation, which may increase oxidative stress. Reducing superoxide production in hyperglycemia and the inflammatory condition is an emerging way to reduce protein and lipid oxidation and diabetes complication. AIM OF STUDY: To examine the effect of Agastache foeniculum essential oil (AFEO) and oil fraction (AFoil) on HG- and LPS-stimulated oxidative stress, the pathogenicity of AFEO and AFoil on oxidative stress was assessed. METHODS: The stimulatory effects of AFEO and AFoil on the activity and expression of NADH oxide (NOX), catalase (CAT), superoxide dismutase (SOD), and the expression of nuclear respiratory factor 2 (NRF2) and nuclear factor-kappa B (NF-kB) in the stimulated macrophage cell line, J774.A1, was studied. The interaction patterns of AFEO and AFoil components with NOX, SOD, CAT, NRF2, and NF-kB proteins were also deduced using molecular docking. RESULTS: Estragole was the main ingredient in AFEO (97%). Linolenic acid (32.10%), estragole (16.22%), palmitic acid (12.62%), linoleic acid (12.04%), and oleic acid (8.73%) were the major chemical components of the AFoil. NOX activation was stimulated in macrophage cells by HG and LPS. At 20 µg/mL, AFEO and AFoil decreased NOX activity while increased SOD and CAT activities in stimulated macrophages. AFoil with estragole and omega-3 fatty acids was better than AFEO with estragole in anti-hyperglycemic and anti-oxidative activity. According to molecular docking research, estragole, linoleic acid, and linolenic acid bind to different hydrophobic pockets of NOX, SOD, CAT, NFR2, and NF-kB using hydrogen bonds, van der Waals bonds, pi-alkyl, and pi-anion interactions, with different binding energies. CONCLUSION: AFEO and AFoil showed antioxidant and anti-diabetic activity. The mechanisms in lowering oxidative stress markers depended on down-regulating superoxide-producing enzymes and up-regulating superoxide-removing enzymes at gene and protein levels. The AFoil emulsion can be used to reduce the detrimental impacts of hyperglycemia and oxidative stress.

Myristicin regulates proliferation and apoptosis in oxidized low-density lipoprotein-stimulated human vascular smooth muscle cells and human umbilical vein endothelial cells by regulating the PI3K/Akt/NF-κB signalling pathway.

CONTEXT: Atherosclerosis (AS) is a chronic inflammatory disease. Human vascular smooth muscle cell (hVSMC) accumulation and human umbilical vein endothelial cell (HUVEC) dysfunction are associated with the pathogenesis of AS. This study explores whether myristicin plays a protective role in AS. MATERIALS AND METHODS: hVSMCs and HUVECs were stimulated with 100 µg/mL oxidized low-density lipoprotein (ox-LDL) to establish a cellular model of AS. Cell viability, lactate dehydrogenase (LDH) release and cell apoptosis were evaluated using MTT, LDH and flow cytometry assays, respectively. Cell migration and inflammatory cytokine release were assessed using Transwell assay and ELISA. RESULTS: Myristicin (5, 10, 25, and 50 µM) had no obvious effect on cell viability or the activity of LDH in hVSMCs, while 100 and 200 µM myristicin markedly suppressed hVSMCs viability and increased LDH release. Myristicin had no obvious effect on cell viability or the activity of LDH in HUVECs. Myristicin inhibited viability and increased apoptosis in ox-LDL-treated hVSMCs, but was associated with increased proliferation and inhibited apoptosis in HUVECs stimulated by ox-LDL. Additionally, myristicin markedly suppressed ox-LDL-induced hVSMCs migration and the release of inflammatory cytokines, including MCP-1, IL-6, VCAM-1 and ICAM-1, in HUVECs. Results also demonstrated that the promoting effects of ox-LDL on the PI3K/Akt and NF-κB signalling pathway in both hVSMCs and HUVECs were abolished by treatment with myristicin. DISCUSSION AND CONCLUSIONS: Myristicin regulated proliferation and apoptosis by regulating the PI3K/Akt/NF-κB signalling pathway in ox-LDL-stimulated hVSMCs and HUVECs. Thus, myristicin may be used as a new potential drug for AS treatment.

The Combination of ß-Asarone and Icariin Inhibits Amyloid-ß and Reverses Cognitive Deficits by Promoting Mitophagy in Models of Alzheimer's Disease.

ß-Asarone is the main constituent of Acorus tatarinowii Schott and exhibits important effects in diseases such as neurodegenerative and neurovascular diseases. Icariin (ICA) is a major active ingredient of Epimedium that has attracted increasing attention because of its unique pharmacological effects in degenerative disease. In this paper, we primarily explored the effects of the combination of ß-asarone and ICA in clearing noxious proteins and reversing cognitive deficits. The accumulation of damaged mitochondria and mitophagy are hallmarks of aging and age-related neurodegeneration, including Alzheimer's disease (AD). Here, we provide evidence that autophagy/mitophagy is impaired in the hippocampus of APP/PS1 mice and in Aß1-42-induced PC12 cell models. Enhanced mitophagic activity has been reported to promote Aß and tau clearance in in vitro and in vivo models. Meanwhile, there is growing evidence that treatment of AD should be preceded by intervention before the formation of pathological products. The efficacy of the combination therapy was better than that of the individual therapies applied separately. Then, we found that the combination therapy also inhibited cell and mitochondrial damage by inducing autophagy/mitophagy. These findings suggest that impaired removal of defective mitochondria is a pivotal event in AD pathogenesis, and that combination treatment with mitophagy inducers represents a potential strategy for therapeutic intervention.

Pharmacological and Therapeutic Potential of Myristicin: A Literature Review.

Natural products have been used by humanity for many centuries to treat various illnesses and with the advancement of technology, it became possible to isolate the substances responsible for the beneficial effects of these products, as well as to understand their mechanisms. In this context, myristicin, a substance of natural origin, has shown several promising activities in a large number of in vitro and in vivo studies carried out. This molecule is found in plants such as nutmeg, parsley, carrots, peppers, and several species endemic to the Asian continent. The purpose of this review article is to discuss data published in the last 10 years at Pubmed, Lilacs and Scielo databases, reporting beneficial effects, toxicity and promising data of myristicin for its future use in medicine. From 94 articles found in the literature, 68 were included. Exclusion criteria took into account articles whose tested extracts did not have myristicin as one of the major compounds.

Synergistic effect of anethole and doxorubicin alleviates cell proliferation, cell cycle arrest, and ER stress and promotes ROS-mediated apoptosis in triple-negative breast cancer cells.

The heterogeneity and poor prognosis of triple-negative breast cancer (TNBC) have limited the treatment options and made clinical management challenging. This has nurtured a major effort to discover druggable molecular targets. Currently, chemotherapy is the primary treatment strategy for this disease. Doxorubicin is the most frequently used chemotherapeutic drug for TNBC and due to the fact that chemotherapeutic drugs have a lot of side effects, we evaluated the synergistic effect of the phytocompound anethole and doxorubicin. The cytotoxic effect of anethole in combination with doxorubicin on MDA-MB-231 cells was evaluated by various parameters, including apoptosis, cell cycle analysis, DNA damage, and cell proliferation. Furthermore, mitochondrial membranepotential (MMP), endoplasmic reticulum (ER) stress, and reactive oxygen species (ROS) levels were also evaluated in the cells treated with/without anethole and doxorubicin. Expression of the apoptotic proteins was evaluated by Western blot analysis. Initial evaluation of cytotoxicity of anethole on MDA-MB-231 cells demonstrated preferential suppression of cell proliferation and when treated along with doxorubicin it showed enhanced cytotoxicity with a synergistic effect. Cell cycle analysis revealed arrest at different stages of the cell cycle, such as sub G0-G1, G0-G1, S, and G2M in various treatment groups and apoptotic cell death was subsequently evident with propidium iodide (PI) staining. The synergistic action of anethole and doxorubicin effectively induced mitochondrial membrane potential loss, which, in turn, led to a burst of ROS production, which eventually produced unfolded protein response by damaging the ER. Synergistic anticancer effect was observed on exposure of MDA-MB-231 cells to anethole and doxorubicin in inducing cell death.

Anethole inhibits RANKL-induced osteoclastogenesis by downregulating ERK/AKT signaling and prevents ovariectomy-induced bone loss in vivo.

Postmenopausal osteoporosis is a chronic population health hazard systemic metabolic disease caused by excessive bone resorption and reduced bone formation. The activity between osteoblast and osteoclast, with their mutual effects, influence the procedure of normal bone remodeling. Over-activated osteoclast differentiation and function play a crucial role in excessive bone resorption. Hence, therapy strategies targeting osteoclast activity may promote the bone mass preservation and delay the osteoporosis process. Natural compound (anethole) is emerging as potential therapeutics for various metabolic diseases. The purpose of this study is to investigate the potential effects of anethole on RANKL-induced osteoclast formation and function in vitro and in vivo. Here, in vitro TRAP staining assay was performed to investigate the inhibitory effect of anethole on osteoclast differentiation. Bone pits resorption assay revealed that osteoclast-mediated bone resorption was inhibited by anethole. At mRNA and protein levels, anethole significantly reduced the expression of osteoclast-specific genes expression in a concentration- or time-dependent manner, including NFATc1, MMP-9, DC-STAMP, c-F, TRAP, CTR, Cathepsin K, and V-ATPase d2. Furthermore, intracellular signaling transduction assay indicated that anethole inhibited osteoclast formation via blocking ERK and AKT signaling. GSK3ß, the downstream signal of AKT, is simultaneously suppressed with anethole treatment. Based on ovariectomized (OVX) mice model, micro-CT and histological staining results suggested that anethole prevented estrogen deficiency-induced bone mass loss and increased osteoclast activity in vivo. In conclusion, our results show significant indications that anethole exhibits an osteoprotective effect and may be potential for the treatment of osteoporosis.

Anticancer effect of myristicin on hepatic carcinoma and related molecular mechanism.

CONTEXT: Myristicin is a natural active compound that has inflammatory, antimicrobial and anti-proliferative properties. Yet, its effect on hepatic carcinoma has not been investigated. OBJECTIVE: To explore the role and related molecular mechanism of myristicin in hepatic carcinoma in vitro. MATERIALS AND METHODS: Human hepatic carcinoma cell lines (Huh-7 and HCCLM3 cells) were treated with different concentrations of myristicin (0.5, 1 and 5 mM) for 24, 48 and 72 h. Then, (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium assay (MTT), flow cytometer (FCM) analysis and transwell assay were performed to determine cell proliferation, apoptosis and migration/invasion, respectively. Protein levels of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X (Bax), E-cadherin, N-cadherin and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signalling pathway-related proteins were detected using Western blot assay. Gene expression was determined using quantitative real time-polymerase chain reaction (qRT-PCR). RESULTS: Myristicin inhibited cell proliferation and induced apoptosis in Huh-7 and HCCLM3 cells; suppressed cell migration and invasion ability, and increased E-cadherin expression and decreased N-cadherin expression, thereby inhibiting epithelial-mesenchymal transition (EMT). Finally, the findings indicated that myristicin decreased phosphorylated (p)-mTOR and p-AKT expression at the protein level. DISCUSSION AND CONCLUSIONS: Myristicin exerts an efficient therapeutic effect on hepatic carcinoma by suppressing PI3K/Akt/mTOR signalling pathway; thus, it may be used as a new potential drug for hepatic carcinoma treatment.

Inhibition of Bacterial Efflux Pumps by Crude Extracts and Essential Oil from Myristica fragrans Houtt. (Nutmeg) Seeds against Methicillin-Resistant Staphylococcus aureus.

Myristicafragrans Houtt. (Nutmeg) is a widely known folk medicine across several parts of Asia, particularly used in antimicrobial treatment. Bacterial resistance involves the expression of efflux pump systems (chromosomal norA and mepA) in methicillin-resistant Staphylococcus aureus (MRSA). Crude extract (CE) and essential oil (EO) obtained from nutmeg were applied as efflux pump inhibitors (EPIs), thereby enhancing the antimicrobial activity of the drugs they were used in. The major substances in CE and EO, which function as EPIs, in a descending order of % peak area include elemicin, myristicin, methoxyeugenol, myristicin, and asarone. Here, we investigated whether the low amount of CE and EO used as EPIs was sufficient to sensitize MRSA killing using the antibiotic ciprofloxacin, which acts as an efflux system. Interestingly, synergy between ciprofloxacin and CE or EO revealed the most significant viability of MRSA, depending on norA and mepA, the latter being responsible for EPI function of EO. Therefore, CE and EO obtained from nutmeg can act as EPIs in combination with substances that act as efflux systems, thereby ensuring that the MRSA strain is susceptible to antibiotic treatment.

Effects of ß-Asarone on Ischemic Stroke in Middle Cerebral Artery Occlusion Rats by an Nrf2-Antioxidant Response Elements (ARE) Pathway-Dependent Mechanism.

BACKGROUND This study assessed the effects and underlying molecular mechanisms of ß-asarone on ischemic stroke model rats. MATERIAL AND METHODS Ischemic stroke was induced by middle cerebral artery occlusion (MCAO) in rats. Before and after modeling, cognitive function was evaluated via fear conditioning test and neurological deficit was determined via Longa and Bederson scores. Following treatment with ß-asarone or nuclear factor erythroid 2-related factor 2 (Nrf2) inhibitor for 20 consecutive days, the cerebral infarction was detected via TTC staining and Cresyl Violet staining in brain tissues. TUNEL staining and western blot analysis for apoptosis-related proteins were performed to assess the apoptosis of neurons. Nrf2-antioxidant response elements (ARE) pathway-related proteins were examined by RT-qPCR or western blot. RESULTS The cognitive and neurological function was defective in MCAO rats. The infarction volumes and the apoptosis of cortical neurons were significantly increased in brain tissues of model rats, which were ameliorated after treatment with ß-asarone. Meanwhile, the increase in pro-apoptotic proteins and decrease in anti-apoptotic proteins were found in brain tissues of model rats, which were markedly ameliorated by ß-asarone treatment. However, Nrf2 inhibitor worsened the cerebral infarction and the apoptosis of neurons. Western blot results showed that ß-asarone treatment activated the Nrf2-ARE pathway-related proteins in model rats, which was inhibited by Nrf2 inhibitor. CONCLUSIONS Our findings suggest that ß-asarone treatment ameliorated the cerebral infarction in MCAO rats, which could be related to activation of the Nrf2-ARE pathway.

Chemical compositions and repellent activity of Clerodendrum bungei Steud. essential oil against three stored product insects.

BACKGROUND: Several species of Verbenaceae have been widely used in medicine, and some species of Verbenaceae have been observed good insecticidal activity, such as Lantana camara and Vitex negundo. There is no report about repellent activity of Clerodendrum bungei Steud. (C. bungei) against stored product insects. The chemical composition of C. bungei essential oil (EO) were identified, repellent activity of methanol extract, EO of C. bungei and two main components of EO against T. castaneum, L. serricorne and L. bostrychophila were evaluated for the first time. RESULTS: EO of C. bungei was obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS) and GC. A total of 25 components of the C. bungei EO were identified. The principal compounds in the EO were myristicin (75.0%), 2,2,7,7-Tetramethyltricyclo[6.2.1.0(1,6)]undec-4-en-3-one (4.1%) and linalool (3.4%). Results of bioassays indicated that C. bungei EO exerted strong repellent activity against three target insects. As main constituents, myristicin and linalool also had certain repellency. CONCLUSION: This work suggests that the EO of C. bungei has promising potential to develop into botanical repellents for the control of pest damage in warehouses and grain stores.

Anethole induces anti-oral cancer activity by triggering apoptosis, autophagy and oxidative stress and by modulation of multiple signaling pathways.

Oral cancer is one of the major public health problems. The aim of this study was to evaluate the effects of anethole, 1-methoxy-4-[(E)-1-propenyl]-benzene, on growth and apoptosis of oral tumor cells, and to identify the signaling pathways involved in its interaction with these cancer cells. Cancer gingival cells (Ca9-22) were treated with different concentrations of anethole. Cell proliferation and cytotoxic effects were measured by MTT and LDH assays. Cell death, autophagy and oxidative stress markers were assessed by flow cytometry while cell migration was determined by a healing capacity assay. The effect of anethole on apoptotic and pro-carcinogenic signaling pathways proteins was assessed by immunoblotting. Our results showed that anethole selectively and in a dose-dependent manner decreases the cell proliferation rate, and conversely induces toxicity and apoptosis in oral cancer cells. This killing effect was mediated mainly through NF-κB, MAPKinases, Wnt, caspase 3, 9 and PARP1 pathways. Anethole showed an ability to induce autophagy, decrease reactive oxygen species (ROS) production and increased intracellular glutathione (GSH) activity. Finally, anethole treatment inhibits the expression of oncogenes (cyclin D1) and up-regulated cyclin-dependent kinase inhibitor (p21WAF1), increases the expression of p53 gene, but inhibits the epithelial-mesenchymal transition markers. These results indicate that anethole could be a potential molecule for the therapy of oral cancer.