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.

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.

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.

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.

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.

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.

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.

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.

Electrical Generation and Deletion of Magnetic Skyrmion-Bubbles via Vertical Current Injection.

The magnetic skyrmion is a topologically protected spin texture that has attracted much attention as a promising information carrier because of its distinct features of suitability for high-density storage, low power consumption, and stability. One of the skyrmion devices proposed so far is the skyrmion racetrack memory, which is the skyrmion version of the domain-wall racetrack memory. For application in devices, skyrmion racetrack memory requires electrical generation, deletion, and displacement of isolated skyrmions. Despite the progress in experimental demonstrations of skyrmion generation, deletion, and displacement, these three operations have yet to be realized in one device. Here, a route for generating and deleting isolated skyrmion-bubbles through vertical current injection with an explanation of its microscopic origin is presented. By combining the proposed skyrmion-bubble generation/deletion method with the spin-orbit-torque-driven skyrmion shift, a proof-of-concept experimental demonstration of the skyrmion racetrack memory operation in a three-terminal device structure is provided.

Metagenomic, Metabolomic, and Functional Evaluation of Kimchi Broth Treated with Light-Emitting Diodes (LEDs).

The light-emitting diode (LED) has been widely used in the food industry, and its application has been focused on microbial sterilization, specifically using blue-LED. The investigation has been recently extended to characterize the biotic and abiotic (photodynamic) effects of different wavelengths. Here, we investigated LED effects on kimchi fermentation. Kimchi broths were treated with three different colored-LEDs (red, green, and blue) or kept in the dark as a control. Multiomics was applied to evaluate the microbial taxonomic composition using 16S rRNA gene amplicon sequencing, and the metabolomic profiles were determined using liquid chromatography-Orbitrap mass spectrometry. Cell viability was tested to determine the potential cytotoxicity of the LED-treated kimchi broths. First, the amplicon sequencing data showed substantial changes in taxonomic composition at the family and genus levels according to incubation (initial condition vs. all other groups). The differences among the treated groups (red-LED (RLED), green-LED (GLED), blue-LED (BLED), and dark condition) were marginal. The relative abundance of Weissella was decreased in all treated groups compared to that of the initial condition, which coincided with the decreased composition of Lactobacillus. Compositional changes were relatively high in the GLED group. Subsequent metabolomic analysis indicated a unique metabolic phenotype instigated by different LED treatments, which led to the identification of the LED treatment-specific and common compounds (e.g., luteolin, 6-methylquinoline, 2-hydroxycinnamic acid, and 9-HODE). These results indicate that different LED wavelengths induce characteristic alterations in the microbial composition and metabolomic content, which may have applications in food processing and storage with the aim of improving nutritional quality and the safety of food.

Programmable Dynamics of Exchange-Biased Domain Wall via Spin-Current-Induced Antiferromagnet Switching.

Magnetic domain wall (DW) motion in perpendicularly magnetized materials is drawing increased attention due to the prospect of new type of information storage devices, such as racetrack memory. To augment the functionalities of DW motion-based devices, it is essential to improve controllability over the DW motion. Other than electric current, which is known to induce unidirectional shifting of a train of DWs, an application of in-plane magnetic field also enables the control of DW dynamics by rotating the DW magnetization and consequently modulating the inherited chiral DW structure. Applying an external bias field, however, is not a viable approach for the miniaturization of the devices as the external field acts globally. Here, the programmable exchange-coupled DW motion in the antiferromagnet (AFM)/ferromagnet (FM) system is demonstrated, where the role of an external in-plane field is replaced by the exchange bias field from AFM layer, enabling the external field-free modulations of DW motions. Interestingly, the direction of the exchange bias field can also be reconfigured by simply injecting spin currents through the device, enabling electrical and programmable operations of the device. Furthermore, the result inspires a prototype DW motion-based device based on the AFM/FM heterostructure, that could be easily integrated in logic devices.

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.

Inhibition of AKT Enhances the Sensitivity of NSCLC Cells to Metformin.

BACKGROUND/AIM: Metformin is an antidiabetic drug that has been reported to have antitumor activity in many cancer types. This study investigated the molecular mechanisms underlying the antitumor effect of metformin. MATERIALS AND METHODS: We investigated the molecular mechanism of the antitumor effect of metformin alone and in combination with AKT serine/threonine kinase (AKT) inhibition via cell viability and western blot analyses. RESULTS: Notably, metformin increased the phosphorylation of AKT at serine 473 using protein array screening. Metformin-induced AKT activation was markedly suppressed by siRNA targeting activating transcription factor 4 (ATF4) but not AMP-activated protein kinase α. These results indicate that AKT activation by metformin was induced in an ATF4-dependent and AMPKα-independent manner. Treatment using metformin combined with MK-2206, an AKT inhibitor, or a siRNA for AKT markedly reduced the viability of cells compared with those cells treated with these agents alone. In addition, MK-2206 increased cell sensitivity to the combination of metformin with ionizing radiation or cisplatin. CONCLUSION: Inhibition of AKT can enhance the antitumor effect of metformin and would be a promising strategy to sensitize non-small-cell lung cancer to a combination of metformin with radiation or cisplatin.

Antioxidant Properties and Protective Effects of Aerial Parts from Cnidium officinale Makino on Oxidative Stress-Induced Neuronal Cell Death.

The rhizomes of Cnidium officinale Makino have been used as a traditional medicine for many purposes, however, use of its aerial parts is very limited. We investigated the antioxidant properties and protective effects of the aerial parts (leaves and stems) from C. officinale on H2O2-induced toxicity in SH-SY5Y neuroblastoma. C. officinale methanol extracts (70%) were sequentially fractionated using hexane (non-polar fraction, NF), ethyl acetate (intermediate polar fraction, IF), and water (polar fraction, PF). Total phenolics and flavonoids contents were highest in IF, followed by PF. IF also showed the strongest radical scavenging activities against 2,2-diphenyl-2-picrylhydrazyl and 2,2'-azinobis(3-ethylbenzo-thiazoline-6-sulfonic acid), as well as superoxide, with the half maximal inhibitory concentrations of 13.2, 23.2, and 12.8 mg/mL, respectively. Furthermore, all fractions significantly inhibited linoleic acid peroxidation induced by the Fenton reaction or by UV irradiation. Both PF and IF protected against H2O2-induced SH-SY5Y neuronal cell death by increasing the cell survival by 22.1∼47.7 and 35.9∼50.3% at concentrations of 25∼100 and 25∼400 µg/mL, respectively, whereas NF was toxic to the cells at these concentrations. IF also significantly decreased intracellular levels of reactive oxygen species by 7.72∼47.47% at a concentration of 25∼200 µg/mL. Our results indicate that compounds from the aerial parts of C. officinale have potent antioxidant activities, which may help rescue neuronal cells from oxidative stress-induced injury. Therefore, the aerial parts, as well as the rhizomes, of C. officinale may have medicinal applications.

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.

Enhancing the conductivity of PEDOT:PSS films for biomedical applications via hydrothermal treatment.

This paper reports a new biocompatible conductivity enhancement of poly (3,4-ethylenedioxythiophene):poly (styrene sulfonate) (PEDOT:PSS) films for biomedical applications. Conductivity of PEDOT:PSS layer was reproducibly from 0.495 to 125.367 S cm-1 by hydrothermal (HT) treatment. The HT treatment employs water (relative humidity > 80%) and heat (temperature > 61 °C) instead of organic solvent doping and post-treatments, which can leave undesirable residue. The treatment can be performed using the sterilizing conditions of an autoclave. Additionally, it is possible to simultaneously reduce the electrical resistance, and sterilize the electrode for practical use. The key to conductivity enhancement was the structural rearrangement of PEDOT:PSS, which was determined using atomic force microscopy, X-ray diffraction, Raman spectroscopy, X-ray photoelectron spectroscopy, and ultraviolet-visible spectroscopy. It was found that PEDOT inter-bridging occurred as a result of the structural rearrangement. Therefore, the conductivity increased on account of the continuous conductive pathways of the PEDOT chains. To test the biocompatible enhancement technique for biomedical applications, certain demonstrations, such as the monitoring of joint movements and skin temperature, and measuring electrocardiogram signals were conducted with the hydrothermal-treated PEDOT:PSS electrode. This simple, biocompatible treatment exhibited significant potential for use in other biomedical applications as well.

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.

Universal field-tunable terahertz emission by ultrafast photoinduced demagnetization in Fe, Ni, and Co ferromagnetic films.

We report a universal terahertz (THz) emission behavior from simple Ni, Fe, and Co metallic ferromagnetic films, triggered by the femtosecond laser pulse and subsequent photoinduced demagnetization on an ultrafast time scale. THz emission behavior in ferromagnetic films is found to be consistent with initial magnetization states controlled by external fields, where the hysteresis of the maximal THz emission signal is observed to be well-matched with the magnetic hysteresis curve. It is experimentally demonstrated that the ultrafast THz emission by the photoinduced demagnetization is controllable in a simple way by external fields as well as pump fluences.

Role of non-thermal electrons in ultrafast spin dynamics of ferromagnetic multilayer.

Understanding of ultrafast spin dynamics is crucial for future spintronic applications. In particular, the role of non-thermal electrons needs further investigation in order to gain a fundamental understanding of photoinduced demagnetization and remagnetization on a femtosecond time scale. We experimentally demonstrate that non-thermal electrons existing in the very early phase of the photoinduced demagnetization process play a key role in governing the overall ultrafast spin dynamics behavior. We simultaneously measured the time-resolved reflectivity (TR-R) and the magneto-optical Kerr effect (TR-MOKE) for a Co/Pt multilayer film. By using an extended three-temperature model (E3TM), the quantitative analysis, including non-thermal electron energy transfer into the subsystem (thermal electron, lattice, and spin), reveals that energy flow from non-thermal electrons plays a decisive role in determining the type I and II photoinduced spin dynamics behavior. Our finding proposes a new mechanism for understanding ultrafast remagnetization dynamics.