Chemical composition, antioxidant, antimicrobial and antiviral activities of the leaf extracts of Syzygium myrtifolium.

This study was conducted to evaluate the chemical composition and biological activities of the leaf extracts of Syzygium myrtifolium Walp. (Myrtaceae). The results indicate that the leaf extracts of S. myrtifolium contain various classes of phytochemicals (alkaloids, anthraquinones, flavonoids, phenolics, saponins, tannins and triterpenoids) and possess antioxidant, antibacterial, antifungal and antiviral activities. Ethyl acetate, ethanol, methanol, and water extracts exhibited significantly higher (p < 0.05) oxygen radical absorbance capacity and ferric-reducing antioxidant power than the hexane and chloroform extracts. However, all extracts exhibited stronger inhibitory activity against four tested species of yeasts (minimal inhibitory concentration: 0.02-0.31 mg mL-1) than against six tested species of bacteria (minimal inhibitory concentration: 0.16-1.25 mg mL-1). The ethanolic extract offered the highest protection of Vero cells (viability > 70 %) from the cytopathic effect caused by the Chikungunya virus while the ethyl acetate extract showed significant replication inhibitory activity against the virus (p < 0.001) using the replicon-enhanced green fluorescent protein reporter system.

Hericium erinaceus Mycelium and Its Isolated Compound, Erinacine A, Ameliorate High-Fat High-Sucrose Diet-Induced Metabolic Dysfunction and Spatial Learning Deficits in Aging Mice.

Aging and lifestyle factors, including high-sugar and high-fat diets, promote a systemic metabolic imbalance that promotes neurodegeneration. Hericium erinaceus has long been used in traditional Chinese medicine. Recently, its functional activities, such as antimetabolic dysfunction, antineuroinflammatory activities, and stimulation of nerve growth factor (NGF) synthesis, have been revealed. This study demonstrated that Hericium erinaceus mycelium (HEM) and an isolated diterpenoid derivative, erinacine A (EA), may reverse spatial learning disabilities in aging mice (15 months old) fed with a high-fat and high-sucrose diet (HFSD). Aging mice were randomly assigned to one of four treatment groups: (1) a chow diet (control), (2) an HFSD, and an HFSD supplemented with either (3) HEM or (4) EA for 18 weeks. The Morris water maze (MWM) and Y-maze were used for behavioral assessments. Both HEM- and EA-treated mice had shorter mean daily escape latencies than HFSD-treated mice in the MWM. In addition, HEM-treated mice had a slightly increased exploratory time and frequency in the novel arm in the Y-maze. Quantitative PCR revealed that both HEM- and EA-treated mice exhibited reduced messenger RNA (mRNA) expression of tumor necrosis factor-α, interleukin-1ß, and HEM-treated mice exhibited increased mRNA expression of NGF and NeuN in the hippocampus. Moreover, HEM and EA also decreased body weight, abdominal fat, plasma glucose, serum and liver total cholesterol, and liver triacylglycerol. Thus, HEM may be a potential health-promoting supplement for minimizing the progression of aging and obesity-induced neurodegeneration by reducing metabolic abnormalities and neuroinflammatory cytokines and increasing neurogenesis factors.

Using Bicistronic Baculovirus Expression Vector System to Screen the Compounds That Interfere with the Infection of Chikungunya Virus.

Chikungunya virus (CHIKV) is the etiologic agent of Chikungunya fever and has emerged in many countries over the past decade. There are no effective drugs for controlling the disease. A bicistronic baculovirus expression system was utilized to co-express CHIKV structural proteins C (capsid), E2 and E1 and the enhanced green fluorescence protein (EGFP) in Spodoptera frugiperda insect cells (Sf21). The EGFP-positive Sf21 cells fused with each other and with uninfected cells to form a syncytium is mediated by the CHIKV E1 allowing it to identify chemicals that can prevent syncytium formation. The compounds characterized by this method could be anti-CHIKV drugs.

Potential natural products for Alzheimer's disease: targeted search using the internal ribosome entry site of tau and amyloid-ß precursor protein.

Overexpression of the amyloid precursor protein (APP) and the hyperphosphorylation of the tau protein are vital in the understanding of the cause of Alzheimer's disease (AD). As a consequence, regulation of the expression of both APP and tau proteins is one important approach in combating AD. The APP and tau proteins can be targeted at the levels of transcription, translation and protein structural integrity. This paper reports the utilization of a bi-cistronic vector containing either APP or tau internal ribosome entry site (IRES) elements flanked by ß-galactosidase gene (cap-dependent) and secreted alkaline phosphatase (SEAP) (cap-independent) to discern the mechanism of action of memantine, an N-methyl-D-aspartate (NMDA) receptor antagonist. Results indicate that memantine could reduce the activity of both the APP and tau IRES at a concentration of ~10 µM (monitored by SEAP activity) without interfering with the cap-dependent translation as monitored by the ß-galactosidase assay. Western blot analysis of the tau protein in neuroblastoma (N2A) and rat hippocampal cells confirmed the halting of the expression of the tau proteins. We also employed this approach to identify a preparation named NB34, extracts of Boussingaultia baselloides (madeira-vine) fermented with Lactobacillus spp., which can function similarly to memantine in both IRES of APP and Tau. The water maze test demonstrated that NB34 could improve the spatial memory of a high fat diet induced neurodegeneration in apolipoprotein E-knockout (ApoE-/-) mice. These results revealed that the bi-cistronic vector provided a simple, and effective platform in screening and establishing the mechanistic action of potential compounds for the treatment and management of AD.

Magnesium lithospermate B and rosmarinic acid, two compounds present in Salvia miltiorrhiza, have potent antiviral activity against enterovirus 71 infections.

The aim of this study was to identify the active ingredients responsible for the anti-EV71 activity produced by Salvia miltiorrhiza extracts. A pGS-EV71 IRES-based bicistronic reporter assay platform was used for rapid analysis of compounds that could specifically inhibit EV71 viral IRES-mediated translation. The analysis identified 2 caffeic acid derivatives, magnesium lithospermate B (MLB) and rosmarinic acid (RA), which suppressed EV71 IRES-mediated translation at concentrations of 30µg/ml. We also found that MLB and RA inhibited EV71 infection when they were added to RD cells during the viral absorption stage. MLB had a low IC50 value of 0.09mM and a high TI value of 10.52. In contrast, RA had an IC50 value of 0.50mM with a TI value of 2.97. MLB and RA (100µg/ml) also reduced EV71 viral particle production and significantly decreased VP1 protein production. We propose that these two derivatives inhibit EV71 viral entry into cells and viral IRES activity, thereby reducing viral particle production and viral RNA expression and blocking viral VP1 protein translation. This study provides useful information for the development of anti-EV71 assays and reagents by demonstrating a convenient EV71 IRES-based bicistronic assay platform to screen for anti-EV71 IRES activity, and also reports 2 compounds, MLB and RA, which are responsible for the anti-EV71 activity of S. miltiorrhiza.

Enhanced protein secretion from insect cells by co-expression of the chaperone calreticulin and translation initiation factor eIF4E.

Host protein synthesis is shut down in the lytic baculovirus expression vector system (BEVS). This also affects host proteins involved in routing secretory proteins through the endoplasmic reticulum (ER)-Golgi system. It has been demonstrated that a secretory alkaline phosphatase-EGFP fusion protein (SEFP) can act as a traceable and sensitive secretory reporter protein in BEVS. In this study, a chaperone, calreticulin (CALR), and the translation initiation factor eIF4E were co-expressed with SEFP using a bicistronic baculovirus expression vector. We observed that the intracellular distribution of SEFP in cells co-expressing CALR was different from co-expressing eIF4E. The increased green fluorescence emitted by cells co-expressing CALR had a good correlation with the abundance of intracellular SEFP protein and an unconventional ER expansion. Cells co-expressing eIF4E, on the other hand, showed an increase in extracellular SEAP activity compared to the control. Utilization of these baculovirus expression constructs containing either eIF4E or CALR offers a significant advantage for producing secreted proteins for various biotechnological and therapeutic applications.

Curcuminoids and resveratrol as anti-Alzheimer agents.

Alzheimer disease (AD) is by far the most common cause of dementia globally. This neurodegenerative disorder of the brain is chronic and progressive, characterized clinically by the deterioration in the key symptoms of behavioral and cognitive abilities. Treatment options for this disease currently are limited. Deposition of amyloid-ß and tau hyperphosphorylation are cardinal pathologic features of AD that lead to the formation of neuronal plaques and neurofibrillary tangles, respectively. In addition to mounting research on herbal compounds for the treatment of AD, curcuminoids and resveratrol appear to be beneficial as anti-AD agents. Curcuminoids (curcumin and demethoxycurcumin) and resveratrol possess unique properties that make them especially worthy of further studies. This review article revisits and presents the current research done on the potential of the curcuminoids curcumin and demethoxycurcumin and the polyphenolic compound resveratrol as anti-AD compounds.

Effects of curcumin and demethoxycurcumin on amyloid-ß precursor and tau proteins through the internal ribosome entry sites: a potential therapeutic for Alzheimer's disease.

OBJECTIVE: This study aims to determine the effects of curcumin and demethoxycurcumin on the internal ribosome entry site of the amyloid-ß precursor protein (APP) and tau protein through a bi-cistronic reporter assay for screening of anti-Alzheimer's disease agents. MATERIALS AND METHODS: A bi-cistronic assay was performed wherein the expression of the first cistron, a ß-galactosidase gene under the control of a cytomegalovirus promoter, represents the canonical cap-dependent mechanism of translation initiation; while the second cistron involves the utilization of the APP or the tau IRES elements to drive the expression of secreted alkaline phosphatase (SEAP) under a cap-independent mechanism. Bioactive natural products reported to have therapeutic potential for AD such as curcumin and demethoxycurcumin were screened in an murine neuroblastoma (N2A) cell model. Western blot analyses for the expression of APP C-terminal protein, human tau-1, and phosphorylated tau at Serine 262 (p(262)) and Serine 396 (pS(396)) were done after treatment of N2A cells with the test compounds. RESULTS: The bi-cistronic reporter assay revealed that curcumin was more effective than demethoxycurcumin, a structural analog of curcumin, in inhibiting both APP and tau IRES-dependent translation initiation. This result was further confirmed by Western blot analysis for the expression of APP C-terminal protein, human tau-1, pS(262) and pS(396) suggesting that curcumin may play a role in AD pathology alleviation through the inhibition of the APP and tau IRES-mediated translation mechanism. On the other hand, demethoxycurcumin was observed to inhibit the phosphorylation of both tau pS(262) and pS(396). CONCLUSION: A novel assay system using the bi-cistronic reporter constructs for the identification of compounds with activity against the translation directed by APP and tau IRES was developed. The results provide novel suggestive insights for the potential use of the mentioned compounds as prophylactic and therapeutic anti-AD agents.

Traditional Chinese medicines and Alzheimer's disease.

Traditional Chinese medicines have been widely investigated for the treatment of Alzheimer's disease (AD) because none of the current therapies-either the cholinesterase inhibitors or antagonist of N-methyl-d-aspartate receptors-has profound effects on halting the progression of AD. In recent years, scientists have isolated many active compounds from herbs, which can alleviate dementia and neurodegenerative syndrome with fewer side effects than conventional drugs and, thus, are regarded as promising drug candidates for AD therapy. In this review, we summarize the latest research progress on six herbs for AD therapy-Huperzia serrata, Amaryllidaceae family, Ginkgo biloba, Uncaria rhynchophylla, Polygala tenuifolia, and Salvia officinalis-and focus on the analysis of their active components and possible mechanisms of pharmacological actions on AD.

Amantadine as a regulator of internal ribosome entry site.

AIM: Studies of eukaryotes have yielded 2 translation initiation mechanisms: a classical cap-dependent mechanism and a cap-independent mechanism proceeding through the internal ribosomal entry site (IRES). We hypothesized that it might be possible to identify compounds that may distinguish between cap-dependent translation and cap-independent IRES-mediated translation. METHODS: To facilitate compound screening, we developed bicistronic reporter constructs containing a beta-galactosidase gene (beta-gal) and a secreted human placental alkaline phosphatase (SEAP) reporter gene. Following transcription, the beta-gal gene is translated by a cap-dependent mechanism, while SEAP expression is controlled by the IRES derived from either enterovirus 71 (EV-71) or encephalomyocarditis virus (EMCV). This assay could potentially identify compounds that inhibit SEAP expression (cap-independent) without affecting beta-gal activity (cap-dependent). RESULTS: Using a bicistronic plasmid-based transient transfection assay in the COS-1 cells, we identified amantadine, a compound that inhibited the IRES of EV71- and EMCV-mediated cap-independent translation but did not interfere with cap-dependent translation when the dose of amantadine was lower than 0.25 mg/mL. CONCLUSION: These results imply that amantadine may distinguish between cap-dependent translation and cap-independent IRES-mediated translation and can be used to regulate gene expression at a translational level.