Enhancement of Solubility, Stability, Cellular Uptake, and Bioactivity of Curcumin by Polyvinyl Alcohol.

The biological activities and related mechanisms of curcumin, a major polyphenolic compound in turmeric, the rhizome of Curcuma longa, have been extensively investigated. Due to its poor solubility in water, the analysis of curcumin's biological activities is limited in most aqueous experimental systems. In the present study, the effects of polyvinyl alcohol (PVA), a dietary-compatible vehicle, on the solubility, stability, cellular uptake, and bioactivities of curcumin were investigated. Curcumin solubility was improved significantly by PVA; the color intensity of curcumin aqueous solution in the presence of PVA increased concentration-dependently with its peak shift to a shorter wavelength. Improved suspension stability and photostability of curcumin in an aqueous solution were also observed in the presence of PVA, even at 62.5 µg/mL. The scavenging activities of curcumin against DPPH, ABTS, AAPH radicals, and nitric oxide were enhanced significantly in the presence of PVA. PVA at 250 µg/mL also significantly enhanced the cytotoxic activity of curcumin against both HCT 116 colon cancer and INT 407 (HeLa-derived) embryonic intestinal cells by reducing the IC50 from 16 to 11 µM and 25 to 15 µM, respectively. PVA improved the cellular uptake of curcumin in a concentration-dependent manner in INT 407 cells; it increased the cellular levels more effectively at lower curcumin treatment concentrations. The present results indicate that PVA improves the solubility and stability of curcumin, and changes in these chemical behaviors of curcumin in aqueous systems by PVA could enhance the bioavailability and pharmacological efficacy of curcumin.

Development of a Convenient and Quantitative Method for Evaluating Photosensitizing Activity Using Thiazolyl Blue Formazan Dye.

Photosensitizers cause oxidative damages in various biological systems under light. In this study, the method for analyzing photosensitizing activity of various dietary and medicinal sources was developed using 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (thiazolyl blue formazan; MTT-F) as a probe. Significant and quantitative decolorization of MTT-F was observed in the presence of photosensitizers used in this study under light but not under dark conditions. The decolorization of MTT-F occurred irradiation time-, light intensity-, and photosensitizer concentration-dependently. The decolorized MTT-F was reversibly reduced by living cells; the LC-MS/MS results indicated the formation of oxidized products with -1 m/z of base peak from MTT-F, suggesting that MTT-F decolorized by photosensitizers was its corresponding tetrazolium. The present results indicate that MTT-F is a reliable probe for the quantitative analysis of photosensitizing activities, and the MTT-F-based method can be an useful tool for screening and evaluating photosensitizing properties of various compounds used in many industrial purposes.

Interaction of Thiol Antioxidants with α,ß-Unsaturated Ketone Moiety: Its Implication for Stability and Bioactivity of Curcuminoids.

Many biological functions of curcumin have been reported. As certain bioactivities of curcumin are eliminated by antioxidants, reactive oxygen species generated by curcumin have been suggested as a relevant mechanism. In the present study, the effects of different types of antioxidants on the stability and bioactivities of curcumin were analyzed. High concentrations (>4 mM) of thiol antioxidants, including N-acetylcysteine (NAC), glutathione (GSH), and ß-mercaptoethanol, accelerated the decomposition of curcumin and other curcuminoids; the submillimolar levels (<0.5 mM) of GSH and NAC rather improved their stability. Ascorbic acid or superoxide dismutase also stabilized curcumin, regardless of their concentration. The cellular levels and bioactivities of curcumin, including its cytotoxicity and the induction of heme oxygenase-1, were significantly reduced in the presence of 8 mM of GSH and NAC. The effects were enhanced in the presence of submillilmolar GSH and NAC, or non-thiol antioxidants. The present results indicate that antioxidants with a reduced thiol group could directly interact with the α,ß-unsaturated carbonyl moiety of curcuminoids and modulate their stability and bioactivity.

Targeted Writing and Deleting of Magnetic Skyrmions in Two-Terminal Nanowire Devices.

Controllable writing and deleting of nanoscale magnetic skyrmions are key requirements for their use as information carriers for next-generation memory and computing technologies. While several schemes have been proposed, they require complex fabrication techniques or precisely tailored electrical inputs, which limits their long-term scalability. Here, we demonstrate an alternative approach for writing and deleting skyrmions using conventional electrical pulses within a simple, two-terminal wire geometry. X-ray microscopy experiments and micromagnetic simulations establish the observed skyrmion creation and annihilation as arising from Joule heating and Oersted field effects of the current pulses, respectively. The unique characteristics of these writing and deleting schemes, such as spatial and temporal selectivity, together with the simplicity of the two-terminal device architecture, provide a flexible and scalable route to the viable applications of skyrmions.

Interfering with Color Response by Porphyrin-Related Compounds in the MTT Tetrazolium-Based Colorimetric Assay.

Porphyrin compounds are widely distributed in various natural products and biological systems. In this study, effects of porphyrin-related compounds including zinc protoporphyrin (ZnPP), protoporphyrin IX (PPIX), cyanocobalamin (CBL), hemin, and zinc phthalocyanine (ZnPC) were analyzed on color response of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) tetrazolium-based assay, a commonly-used method for analyzing cell viability. Color responses of MTT formazan formed in cells treated with ZnPP, PPIX, or ZnPC were significantly reduced even at submicromolar concentrations without affecting cell viability, whereas hemin and CBL did not. ZnPP, PPIX, and ZnPC rapidly induced degradation of MTT formazan already-produced by cells when exposed to light, but not under a dark condition. Photosensitizing properties of the three compounds were also verified through extensive generation of reactive oxygen species under light. The porphyrins did not affect the stability of water-soluble formazans including XTT, WST-1, WST-8, and MTS formazans. Several factors including different light sources and antioxidants modulated the degradation process of MTT formazan by the porphyrins. The results suggest that certain porphyrin compounds could cause a severe artifact in the MTT assay through rapid degradation of formazan dye due to their photosensitizing property, which needs to be considered carefully in the related assays.

Inhibition of Glutamine Uptake Resensitizes Paclitaxel Resistance in SKOV3-TR Ovarian Cancer Cell via mTORC1/S6K Signaling Pathway.

Ovarian cancer is a carcinoma that affects women and that has a high mortality rate. Overcoming paclitaxel resistance is important for clinical application. However, the effect of amino acid metabolism regulation on paclitaxel-resistant ovarian cancer is still unknown. In this study, the effect of an amino acid-deprived condition on paclitaxel resistance in paclitaxel-resistant SKOV3-TR cells was analyzed. We analyzed the cell viability of SKOV3-TR in culture conditions in which each of the 20 amino acids were deprived. As a result, the cell viability of the SKOV3-TR was significantly reduced in cultures deprived of arginine, glutamine, and lysine. Furthermore, we showed that the glutamine-deprived condition inhibited mTORC1/S6K signaling. The decreased cell viability and mTORC1/S6K signaling under glutamine-deprived conditions could be restored by glutamine and α-KG supplementation. Treatment with PF-4708671, a selective S6K inhibitor, and the selective glutamine transporter ASCT2 inhibitor V-9302 downregulated mTOR/S6K signaling and resensitized SKOV3-TR to paclitaxel. Immunoblotting showed the upregulation of Bcl-2 phosphorylation and a decrease in Mcl-1 expression in SKOV3-TR via the cotreatment of paclitaxel with PF-4708671 and V-9302. Collectively, this study demonstrates that the inhibition of glutamine uptake can resensitize SKOV3-TR to paclitaxel and represents a promising therapeutic target for overcoming paclitaxel resistance in ovarian cancer.

Inhibition of mTORC1 through ATF4-induced REDD1 and Sestrin2 expression by Metformin.

BACKGROUND: Although the major anticancer effect of metformin involves AMPK-dependent or AMPK-independent mTORC1 inhibition, the mechanisms of action are still not fully understood. METHODS: To investigate the molecular mechanisms underlying the effect of metformin on the mTORC1 inhibition, MTT assay, RT-PCR, and western blot analysis were performed. RESULTS: Metformin induced the expression of ATF4, REDD1, and Sestrin2 concomitant with its inhibition of mTORC1 activity. Treatment with REDD1 or Sestrin2 siRNA reversed the mTORC1 inhibition induced by metformin, indicating that REDD1 and Sestrin2 are important for the inhibition of mTORC1 triggered by metformin treatment. Moreover, REDD1- and Sestrin2-mediated mTORC1 inhibition in response to metformin was independent of AMPK activation. Additionally, lapatinib enhances cell sensitivity to metformin, and knockdown of REDD1 and Sestrin2 decreased cell sensitivity to metformin and lapatinib. CONCLUSIONS: ATF4-induced REDD1 and Sestrin2 expression in response to metformin plays an important role in mTORC1 inhibition independent of AMPK activation, and this signalling pathway could have therapeutic value.

Lysine is required for growth factor-induced mTORC1 activation.

Mechanistic target of rapamycincomplex 1 (mTORC1) integrates various environmental signals to regulate cell growth and metabolism. mTORC1 activity is sensitive to changes in amino acid levels. Here, we investigated the effect of lysine on mTORC1 activity in non-small cell lung cancer (NSCLC) cells. Lysine deprivation suppressed mTORC1 activity and lysine replenishment restored the decreased mTORC1 activity in lysine-deprived cells. Supplementing growth factors, such as insulin growth factor-1 or insulin restored the decreased mTORC1 activity in serum-deprived cells. However, in serum/lysine-deprived cells, supplementing growth factors was not sufficient to restore mTORC1 activity, suggesting thatgrowth factors could not activate mTORC1 efficiently in the absence of lysine. General control nonderepressible 2 and AMP-activated protein kinase were involved in lysine deprivation-mediated inhibition of mTORC1. Taken together, these results suggest that lysine might play role in the regulation of mTORC1 activation in NSCLC cells.

Dibenzoylmethane, a Component of Licorice, Suppresses Monocyte-to-Macrophage Differentiation and Inflammatory Responses in Human Monocytes and Mouse Macrophages.

The objective of this study was to investigate the effect of dibenzoylmethane (DBM) on monocyte-to-macrophage differentiation, the inflammatory response, and the resulting signaling in human monocytes and murine macrophage. DBM effectively inhibited the monocyte-to-macrophage differentiation induced by phorbol 12-myristate 13-acetate (PMA) through a reduction in adhesion of THP-1 cells. Cluster of differentiation molecule ß (CD11ß) and CD36, which are surface markers of macrophage differentiation, were downregulated by 80 and 74%, respectively. DBM also significantly inhibited lipopolysaccharide (LPS)-induced nitrite (NO) production through the downregulation of inducible oxide synthase (iNOS) in RAW264.7 cells. The abundance of cyclooxygenase-2 (COX-2), a pro-inflammatory protein, was also effectively decreased by DBM in a dose-dependent manner. DBM (50 µM) reduced the levels of COX-2 and iNOS by 81 and 78%, respectively. DBM significantly inhibited the translocation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), an inflammatory transcription factor, into the nucleus. DBM-mediated increase of NF-κB translocation resulted from the DBM-induced suppression of the phosphorylation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα). In contrast, DBM effectively increased the expression of nuclear factor E2-related factor 2 (Nrf2) and its target protein, hemeoxygenase-1 (HO-1). Nrf2 translocation into the nucleus was also significantly enhanced by DBM. Furthermore, DBM effectively inhibited the expression of pro-inflammatory cytokines such as tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1ß), IL-6, and monocyte chemoattractant protein-1 (MCP-1). These results indicated that the DBM-mediated differential regulation of NF-κB and Nrf2, which are major transcription factors involved in inflammation, inhibited the expression of inflammatory cytokines.

Dibenzoylmethane Suppresses Lipid Accumulation and Reactive Oxygen Species Production through Regulation of Nuclear Factor (Erythroid-Derived 2)-Like 2 and Insulin Signaling in Adipocytes.

The aim of this study was to investigate the effects of dibenzoylmethane (1,3-diphenyl-1,3-propanedione, DBM) from licorice roots on lipid accumulation and reactive oxygen species (ROS) production in 3T3-L1 cells. DBM effectively inhibited lipid accumulation during adipogenesis, and its inhibitory effect was shown to be due to the down-regulation of adipogenic factors such as CCAAT-enhancer-binding protein-α (C/EBPα), peroxisome proliferator-activated receptor γ (PPARγ), and fatty acid-binding protein 4 (FABP4). DBM was observed to exert its inhibitory effect on lipid accumulation in the early adipogenic stage (days 0-2) by regulating early adipogenic factors including CCAAT-enhancer-binding protein-ß (C/EBPß) and Krueppel-like factor (KLF) 2. DBM significantly increased the translocation of nuclear factor (erythroid-derived 2)-like 2(Nrf2) into the nucleus, promoting the protein expression of its target gene, heme oxygenase-1 (HO-1). DBM significantly suppressed the insulin-mediated activation of phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt), which are components of insulin signaling. In addition, intracellular ROS production was effectively reduced by DBM treatment, which upregulated antioxidant genes such as glutathione peroxidase (Gpx), catalase (CAT), and superoxide dismutase 1 (SOD1). Furthermore, DBM significantly regulated the expression of the adipokines, resistin and adiponectin. This DBM-mediated regulation of lipid accumulation, ROS production, and adipokine production was shown to be involved in the regulation of the Nrf2 and insulin signaling.

TRAIL restores DCA/metformin-mediated cell death in hypoxia.

Previous studies have shown that hypoxia can reverse DCA/metformin-induced cell death in breast cancer cells. Therefore, targeting hypoxia is necessary for therapies targeting cancer metabolism. In the present study, we found that TRAIL can overcome the effect of hypoxia on the cell death induced by treatment of DCA and metformin in breast cancer cells. Unexpectedly, DR5 is upregulated in the cells treated with DCA/metformin, and sustained under hypoxia. Blocking DR5 by siRNA inhibited DCA/metformin/TRAIL-induced cell death, indicating that DR5 upregulation plays an important role in sensitizing cancer cells to TRAIL-induced cell death. Furthermore, we found that activation of JNK and c-Jun is responsible for upregulation of DR5 induced by DCA/metformin. These findings support the potential application of combining TRAIL and metabolism-targeting drugs in the treatment of cancers under hypoxia.

δ-Tocopherol inhibits receptor tyrosine kinase-induced AKT activation in prostate cancer cells.

The cancer preventive activity of vitamin E is suggested by epidemiological studies and supported by animal studies with vitamin E forms, γ-tocopherol and δ-tocopherol (δ-T). Several recent large-scale cancer prevention trials with high dose of α-tocopherol, however, yielded disappointing results. Whether vitamin E prevents or promotes cancer is a serious concern. A better understanding of the molecular mechanisms of action of the different forms of tocopherols would enhance our understanding of this topic. In this study, we demonstrated that δ-T was the most effective tocopherol form in inhibiting prostate cancer cell growth, by inducing cell cycle arrest and apoptosis. By profiling the effects of δ-T on the cell signaling using the phospho-kinase array, we found that the most inhibited target was the phosphorylation of AKT on T308. Further study on the activation of AKT by EGFR and IGFR revealed that δ-T attenuated the EGF/IGF-induced activation of AKT (via the phosphorylation of AKT on T308 induced by the activation of PIK3). Expression of dominant active PIK3 and AKT in prostate cancer cell line DU145 in which PIK3, AKT, and PTEN are wild type caused the cells to be reflectory to the inhibition of δ-T, supporting that δ-T inhibits the PIK3-mediated activation of AKT. Our data also suggest that δ-T interferes with the EGF-induced EGFR internalization, which leads to the inhibition of the receptor tyrosine kinase-dependent activation of AKT. In summary, our results revealed a novel mechanism of δ-T in inhibiting prostate cancer cell growth, supporting the cancer preventive activity δ-T. © 2015 Wiley Periodicals, Inc.

Prevention of chronic diseases by tea: possible mechanisms and human relevance.

Tea, made from leaves of the plant Camellia sinensis, Theaceae, has been used by humans for thousands of years, first as a medicinal herb and then as a beverage that is consumed widely. For the past 25 years, tea has been studied extensively for its beneficial health effects, including prevention of cancer, reduction of body weight, alleviation of metabolic syndrome, prevention of cardiovascular diseases, and protection against neurodegenerative diseases. Whether these effects can be produced by tea at the levels commonly consumed by humans is an open question. This review examines these topics and elucidates the common mechanisms for these beneficial health effects. It also discusses other health effects and possible side effects of tea consumption. This article provides a critical assessment of the health effects of tea consumption and suggests new directions for research in this area.

Duloxetine enhances the sensitivity of non-small cell lung cancer cells to EGFR inhibitors by REDD1-induced mTORC1/S6K1 suppression.

Although epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) have been effective targeted therapies for non-small cell lung cancer (NSCLC), most advanced NSCLC inevitably develop resistance to these therapies. Combination therapies emerge as valuable approach to preventing, delaying, or overcoming disease progression. Duloxetine, an antidepressant known as a serotonin-noradrenaline reuptake inhibitor, is commonly prescribed for the treatment of chemotherapy-induced peripheral neuropathy. In the present study, we investigated the combined effects of duloxetine and EGFR-TKIs and their possible mechanism in NSCLC cells. Compared with either monotherapy, the combination of duloxetine and EGFR-TKIs leads to synergistic cell death. Mechanistically, duloxetine suppresses 70-kDa ribosomal protein S6 kinase 1 (p70S6K1) activity through mechanistic target of rapamycin complex 1 (mTORC1), and this effect is associated with the synergistic induction of cell death of duloxetine combined with EGFR-TKIs. More importantly, activating transcription factor 4 (ATF4)-induced regulated in development and DNA damage response 1 (REDD1) is responsible for the suppression of mTORC1/S6K1 activation. Additionally, we found that the combination effect was significantly attenuated in REDD1 knockout NSCLC cells. Taken together, our findings reveal that the ATF4/REDD1/mTORC1/S6K1 signaling axis, as a novel mechanism, is responsible for the synergistic therapeutic effect of duloxetine with EGFR-TKIs. These results suggest that combining EGFR-TKIs with duloxetine appears to be a promising way to improve EGFR-TKI efficacy against NSCLC.

Downregulation of TRIB3 enhances the sensitivity of lung cancer cells to amino acid deprivation by suppressing AKT activation.

Tribbles pseudokinase 3 (TRIB3), a member of the mammalian Tribbles family, is implicated in multiple biological processes. This study aimed to investigate the biological functions of TRIB3 in lung cancer and its effect on amino acid-deprived lung cancer cells. TRIB3 mRNA expression was elevated in lung cancer tissues and cell lines compared to normal lung tissues and cells. TRIB3 knockdown markedly reduced the viability and proliferation of H1299 lung cancer cells. Deprivation of amino acids, particularly arginine, glutamine, lysine, or methionine, strongly increased TRIB3 expression via ATF4 activation in H1299 lung cancer cells. Knockdown of TRIB3 led to transcriptional downregulation of ATF4 and reduced AKT activation induced by amino acid deprivation, ultimately increasing the sensitivity of H1299 lung cancer cells to amino acid deprivation. Additionally, TRIB3 knockdown enhanced the sensitivity of H1299 cells to V-9302, a competitive antagonist of transmembrane glutamine flux. These results suggest that TRIB3 is a pro-survival regulator of cell viability in amino acid-deficient tumor microenvironments and a promising therapeutic target for lung cancer treatment.

Magnetism in dopant-free ZnO nanoplates.

It is known that bulk ZnO is a nonmagnetic material. However, the electronic band structure of ZnO is severely distorted when the ZnO is in the shape of a very thin plate with its dimension along the c-axis reduced to a few nanometers while keeping the bulk scale sizes in the other two dimensions. We found that the chemically synthesized ZnO nanoplates exhibit magnetism even at room temperature. First-principles calculations show a growing asymmetry in the spin distribution within the distorted bands formed from Zn (3d) and O (2p) orbitals with the reduction of thickness of the ZnO nanoplates, which is suggested to be responsible for the observed magnetism. In contrast, reducing the dimension along the a- or b-axes of a ZnO crystal does not yield any magnetism for ZnO nanowires that grow along c-axis, suggesting that the internal electric field produced by the large {0001} polar surfaces of the nanoplates may be responsible for the distorted electronic band structures of thin ZnO nanoplates.

Nebivolol Sensitizes BT-474 Breast Cancer Cells to FGFR Inhibitors.

BACKGROUND/AIM: The fibroblast growth factor receptor (FGFR) signaling pathway is abnormally activated in human cancers, including breast cancer. Therefore, targeting the FGFR signaling pathway is a potent strategy to treat breast cancer. The purpose of this study was to find drugs that could increase sensitivity to FGFR inhibitor effects in BT-474 breast cancer cells, and to investigate the combined effects and underlying mechanisms of these combinations for BT-474 breast cancer cell survival. MATERIALS AND METHODS: Cell viability was measured by MTT assay. Protein expression was determined by western blot analysis. mRNA expression was detected by Real-time PCR. Drug synergy effect was determined by isobologram analysis. RESULTS: Nebivolol, a third generation ß1-blocker, synergistically increased the sensitivity of BT-474 breast cancer cells to the potent and selective FGFR inhibitors erdafitinib (JNJ-42756493) and AZD4547. A combination of nebivolol and erdafitinib markedly reduced AKT activation. Suppression of AKT activation using specific siRNA and a selective inhibitor further enhanced cell sensitivity to combined treatment with nebivolol and erdafitinib, whereas SC79, a potent activator of AKT, reduced cell sensitivity to nebivolol and erdafitinib. CONCLUSION: Enhanced sensitivity of BT-474 breast cancer cells to nebivolol and erdafitinib was probably associated with down-regulation of AKT activation. Combined treatment with nebivolol and erdafitinib is a promising strategy for breast cancer treatment.

Roles of anti- and pro-oxidant potential of cinnamic acid and phenylpropanoid derivatives in modulating growth of cultured cells.

Cinnamic acid (CiA) and phenylpropanoid derivatives are widely distributed in plant foods. In this study, anti- and pro-oxidant properties of the derivatives and their roles in modulating cell growth were investigated. Ferulic acid, sinapinic acid, caffeic acid (CaA), and 3,4-dihydroxyhydrocinnamic acid (DHC) showed strong radical scavenging activities. They, except DHC, also performed considerable inhibitory effects on lipid peroxidation and reduced levels of intracellular reactive oxygen species (ROS). CaA and DHC, however, produced substantial amount of H2O2 with oxidative degradation in culture conditions. CaA and DHC (> 400 µM) showed potent cytotoxic effects which were abolished by superoxide dismutase/catalase; they significantly enhanced cell growth ROS-dependently at low levels (~ 100 µM). CiA derivatives without bearing hydroxyl group did not show any appreciable antioxidant activities. The results indicate that CiA derivatives with ortho-dihydroxyl group had strong anti- and pro-oxidant properties, which also play an important role in modulating cell growth. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-022-01042-x.

Cellular uptake of anthocyanins extracted from black soybean, grape, and purple sweet potato using INT-407 cells.

This study combined in vitro digestion and INT-407 cells to evaluate the bioaccessibility of anthocyanins in the small intestinal epithelial cells. Black soybean, grape, and purple sweet potato were chosen as they have a different anthocyanin composition. After the aqueous extract was digested under in vitro gastric and intestinal conditions, the digested mixture was incubated in the media of INT-407 for 2 h at 37 °C. Low proportion (< 0.3%) of anthocyanins in black soybean and grape passed through cell membranes. Cyanidin-3-O-glucoside and pelargonidin-3-O-glucoside in black soybean and cyanidin-3-O-(6-O-p-coumaroyl)-5-O-diglucoside and delphinidin-3-O-(6-O-p-coumaroyl)-5-O-diglucoside in grape were found inside the cell. However, acylated anthocyanins containing three sugar moieties in purple sweet potato were not detected inside the cell. p-Coumaric acid was detected in the cells incubated with grape, but not in the media. These indicate that chemical structure of anthocyanins affected their cellular uptake and antioxidant activity in INT-407 cells. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10068-021-00976-y.