Exploring differentiation-dependent responses to 532 nm green laser photobiomodulation in SHSY5Y neuroblastoma cells.

Photobiomodulation (PBM) holds promise as a therapy modality, but its applicability is hindered by the lack of a quantitative model to predict the optimal dose for all forms of PBM. This study investigated the optimal PBM parameters for 532 nm green laser irradiation on SHSY5Y neuroblastoma cells, a commonly used in vitro model for neurodegenerative disease studies. A two-tailed, two sample t-test with equal variance was used to obtain the p-values and statistical significance. There are 3 sets of parameters showing significant ( p < 0 . 01 ) positive percentage biostimulation. 160 m W , 15 m i n produce a percentage biostimulation of ( 9 ± 10 ) % ; 180 m W , 5 m i n produce a percentage biostimulation of ( 19 ± 7 ) % ; and ( 200 m W , 5 m i n ) produce a percentage biostimulation of ( 9 ± 2 ) % . The highest significant ( p < 0 . 01 ) percentage bioinhibition observed is for 220 m W , 15 m i n (dose: 1008 J / c m 2 ) producing a bioinhibition of ( 54 ± 1 ) % . After identifying several parameters that produce noticeable photobiological effects (biostimulation and bioinhibition), this study compared the reaction of undifferentiated and differentiated SHSY5Y cells to laser irradiation and found that undifferentiated SHSY5Y cells shows greater photobiological effect from 532 nm laser irradiation ( p < 0 . 01 ) . This study demonstrated the differentiation-dependant photobiological effect of SHSY5Y in 532 nm laser PBM. This shows that considerations on the differentiation state of cells is important in PBM studies. The hypothesis of difference in intracellular reactive oxygen species (ROS) accumulation from laser irradiation can serve as a versatile explanation of the observed difference in photobiological effect. Further investigation into the role of ROS as a mediator of various photobiological effects from laser of different wavelengths is warranted.

Optimization of the CYP inhibition assay using LC-MS/MS.

The data presented in this article are related to the research article entitled "Cytochrome P450 inhibition activities of non-standardized botanical products" [1], in which the possible CYP inhibitory properties of botanical products were investigated. This article describes the optimization and bioanalytical method validation of the CYP (Cytochrome P450 inhibition assay) inhibition assays, namely, phenacetin O-deethylase assay, testosterone 6ß-hydroxylase assay, felodipine dehydrogenase assay and midazolam 1'-hydroxylase assay using LC-MS/MS.

Cytochrome P450 inhibition activities of non-standardized botanical products.

ETHNOPHARMACOLOGICAL RELEVANCE: R-tab, H-tab and E-cap botanical products are used for the treatment of various ailments. R-tab is traditionally prescribed for improving urination, H-tab is for relieving piles, hemorrhoids, fissures, and rectal inflammation and E-cap is for regulating menstruation. AIMS OF THE STUDY: To extract the botanical products and determine their potential interaction with the cytochrome P450 (CYP1A2, CYP2D6 and CYP3A4) enzymes. MATERIALS AND METHODS: R-tab, H-tab and E-cap botanical products were first extracted using solvents and analyzed using HPLC and LC-MS/MS. The effects of methanol extracts on the cytochrome induction and inhibition activities were determined using a series of in vitro assays, including multiplex RT-qPCR, CYP activity assays (P450-Glo™) and LC-MS/MS-based assays. For the CYP induction assay, omeprazole, rifampicin and dexamethasone were used as CYP1A2, CYP2D6 and CYP3A4 inducers, respectively. Ketoconazole and acetaminophen were used as positive and negative controls for the CYP3A4 inhibition assay, whereas furafylline and ketoconazole were used as positive and negative controls for the CYP1A2 inhibition assay. RESULTS: All three botanical products did not show any significant induction in CYP1A2, CYP2D6 and CYP3A4 mRNA expression. By contrast, R-tab inhibited the mRNA expression of CYP1A2 significantly from the lowest concentration of 0.01 µg/mL, while, H-tab inhibited the mRNA expression of CYP1A2 and CYP3A4 from 0.1 µg/mL. Based on the P450 Glo assays, E-cap extract inhibited the metabolic activity of CYP1A2 with an IC50 value of 37.24 µg/mL. On the other hand, R-tab, H-tab and E-cap showed inhibitory effects on the CYP3A4 enzymatic activity with IC50 values of 17.42, 18.20 and 20.60 µg/mL, respectively. However, using the LC-MS/MS-based methods, the concentration-dependent effects of R-tab and H-tab extracts on the metabolism of testosterone appeared to be more prominent, with IC50 values of 51.90 and 56.90 µg/mL as compared with the rest of the results, which were all above 100 µg/mL CONCLUSION: The CYP3A4 mRNA and enzymatic activity were moderately inhibited by R-tab and H-tab. Methanol extract of botanical products in solid dosage forms can be evaluated for their herb-drug interaction risks using in vitro assays and may provide the minimum data required for safety labeling.

Ketogenesis and SIRT1 as a tool in managing obesity.

Obesity is a serious chronic disease and a public health concern in both developing and developed countries. Managing obesity has been a great challenge for both health care professionals and patients alike. Among the various diet programs aimed at promoting weight loss, the ketogenic diet, a diet high in fat and low in carbohydrates, has been at the forefront recently and its mechanism in weight loss is much debated. Activation of Sirtuin 1 or SIRT1 is able to circumvent various diseases, including metabolic syndrome and obesity and is thought to be a potentially reliable treatment target for both of them. Augmentation of SIRT1 may be carried out using dietary means such as nicotinamide adenine dinucleotide (NAD) supplementation and/or ketogenic diet. Although ketogenic diet may augment SIRT1 activation in people affected by obesity, recent studies have indicated that the relationship between SIRT1 and ketogenesis is unpredictable. The exact circumstances and mechanisms of SIRT1, NAD and ketogenesis in the clinical setting as an intervention tool in managing obesity remained uncertain. Although several recent literatures have documented significant weight-loss following ketogenic diet interventions, there were limitations with regards to duration of trial, choice and the number of trial subjects. Studies investigating the safety of ketogenic diet in the long term, beyond 46 weeks and related mechanism and pathways are still lacking and the sustainability of this diet remains to be determined. This review explores the recent progress on ketogenic diet and its relationships with SIRT1 as a tool in managing obesity and relevant clinical implications.

Development of a NS2B/NS3 protease inhibition assay using AlphaScreen® beads for screening of anti-dengue activities.

BACKGROUND: Dengue infection is an endemic infectious disease and it can lead to dengue fever, dengue hemorrhagic fever, and/or dengue shock syndromes. Dengue NS2B/NS3 protease complex is essential for viral replication and is a primary target for anti-dengue drug development. In this study, a NS2B/NS3 protease inhibition assay was developed using AlphaScreen® beads and was used to screen compounds for their protease inhibition activities. METHODS: The assay system utilized a known NS2B/NS3 peptide substrate, a recombinant of NS2B/NS3 protease with proprietary StrepTactin® donor and nickel chelate acceptor beads in 384-well format. RESULTS: The optimized assay to screen for NS2B/NS3 protease inhibitors was demonstrated to be potentially useful with reasonable z' factor, coefficient variance and signal to background ratio. However, screening of synthesized thioguanine derivatives using the optimized AlphaScreen® assay revealed weak NS2B/NS3 inhibition activities. CONCLUSION: The AlphaScreen® assay to screen for NS2B/NS3 protease inhibitors is potentially applicable for high throughput screening.

Data on the construction of a recombinant HEK293 cell line overexpressing hERG potassium channel and examining the presence of hERG mRNA and protein expression.

The data presented in this article are related to the research article entitled "The effects of deoxyelephantopin on the cardiac delayed rectifier potassium channel current (IKr) and human ether-a-go-go-related gene (hERG) expression" (Y.F. Teah, M.A. Abduraman, A. Amanah, M.I. Adenan, S.F. Sulaiman, M.L. Tan) [1], which the possible hERG blocking properties of deoxyelephantopin were investigated. This article describes the construction of human embryonic kidney 293 (HEK293) cells overexpressing HERG potassium channel and verification of the presence of hERG mRNA and protein expression in this recombinant cell line.

The effects of deoxyelephantopin on the cardiac delayed rectifier potassium channel current (IKr) and human ether-a-go-go-related gene (hERG) expression.

Elephantopus scaber Linn and its major bioactive component, deoxyelephantopin are known for their medicinal properties and are often reported to have various cytotoxic and antitumor activities. This plant is widely used as folk medicine for a plethora of indications although its safety profile remains unknown. Human ether-a-go-go-related gene (hERG) encodes the cardiac IKr current which is a determinant of the duration of ventricular action potentials and QT interval. The hERG potassium channel is an important antitarget in cardiotoxicity evaluation. This study investigated the effects of deoxyelephantopin on the current, mRNA and protein expression of hERG channel in hERG-transfected HEK293 cells. The hERG tail currents following depolarization pulses were insignificantly affected by deoxyelephantopin in the transfected cell line. Current reduction was less than 40% as compared with baseline at the highest concentration of 50 µM. The results were consistent with the molecular docking simulation and hERG surface protein expression. Interestingly, it does not affect the hERG expression at both transcriptional and translational level at most concentrations, although higher concentration at 10 µM caused protein accumulation. In conclusion, deoxyelephantopin is unlikely a clinically significant hERG channel and Ikr blocker.