Aerobic exercise-induced HIF-1α upregulation in heart failure: exploring potential impacts on MCT1 and MPC1 regulation.

BACKGROUND: The terminal stage of ischemic heart disease develops into heart failure (HF), which is characterized by hypoxia and metabolic disturbances in cardiomyocytes. The hypoxic failing heart triggers hypoxia-inducible factor-1α (HIF-1α) actions in the cells sensitized to hypoxia and induces metabolic adaptation by accumulating HIF-1α. Furthermore, soluble monocarboxylic acid transporter protein 1 (MCT1) and mitochondrial pyruvate carrier 1 (MPC1), as key nodes of metabolic adaptation, affect metabolic homeostasis in the failing rat heart. Aerobic exercise training has been reported to retard the progression of HF due to enhancing HIF-1α levels as well as MCT1 expressions, whereas the effects of exercise on MCT1 and MPC1 in HF (hypoxia) remain elusive. This research aimed to investigate the action of exercise associated with MCT1 and MPC1 on HF under hypoxia. METHODS: The experimental rat models are composed of four study groups: sham stented (SHAM), HF sedentary (HF), HF short-term exercise trained (HF-E1), HF long-term exercise trained (HF-E2). HF was initiated via left anterior descending coronary artery ligation, the effects of exercise on the progression of HF were analyzed by ventricular ultrasound (ejection fraction, fractional shortening) and histological staining. The regulatory effects of HIF-1α on cell growth, MCT1 and MPC1 protein expression in hypoxic H9c2 cells were evaluated by HIF-1α activatort/inhibitor treatment and plasmid transfection. RESULTS: Our results indicate the presence of severe pathological remodelling (as evidenced by deep myocardial fibrosis, increased infarct size and abnormal hypertrophy of the myocardium, etc.) and reduced cardiac function in the failing hearts of rats in the HF group compared to the SHAM group. Treadmill exercise training ameliorated myocardial infarction (MI)-induced cardiac pathological remodelling and enhanced cardiac function in HF exercise group rats, and significantly increased the expression of HIF-1α (p < 0.05), MCT1 (p < 0.01) and MPC1 (p < 0.05) proteins compared to HF group rats. Moreover, pharmacological inhibition of HIF-1α in hypoxic H9c2 cells dramatically downregulated MCT1 and MPC1 protein expression. This phenomenon is consistent with knockdown of HIF-1α at the gene level. CONCLUSION: The findings propose that long-term aerobic exercise training, as a non- pharmacological treatment, is efficient enough to debilitate the disease process, improve the pathological phenotype, and reinstate cardiac function in HF rats. This benefit is most likely due to activation of myocardial HIF-1α and upregulation of MCT1 and MPC1.

Visual light flicker stimulation: enhancing alertness in sleep-deprived rats.

Introduction: In the evolving field of neurophysiological research, visual light flicker stimulation is recognized as a promising non-invasive intervention for cognitive enhancement, particularly in sleep-deprived conditions. Methods: This study explored the effects of specific flicker frequencies (40 Hz and 20-30 Hz random flicker) on alertness recovery in sleep-deprived rats. We employed a multidisciplinary approach that included behavioral assessments with the Y-maze, in vivo electrophysiological recordings, and molecular analyses such as c-FOS immunohistochemistry and hormone level measurements. Results: Both 40 Hz and 20-30 Hz flicker significantly enhanced behavioral performance in the Y-maze test, suggesting an improvement in alertness. Neurophysiological data indicated activation of neural circuits in key brain areas like the thalamus and hippocampus. Additionally, flicker exposure normalized cortisol and serotonin levels, essential for stress response and mood regulation. Notably, increased c-FOS expression in brain regions related to alertness and cognitive functions suggested heightened neural activity. Discussion: These findings underscore the potential of light flicker stimulation not only to mitigate the effects of sleep deprivation but also to enhance cognitive functions. The results pave the way for future translational research into light-based therapies in human subjects, with possible implications for occupational health and cognitive ergonomics.

Genome-wide investigation of UDP-Glycosyltransferase family in Tartary buckwheat (Fagopyrum tataricum).

BACKGROUND: Tartary buckwheat (Fagopyrum tataricum) belongs to Polygonaceae family and has attracted increasing attention owing to its high nutritional value. UDP-glycosyltransferases (UGTs) glycosylate a variety of plant secondary metabolites to control many metabolic processes during plant growth and development. However, there have been no systematic reports of UGT superfamily in F. tataricum. RESULTS: We identified 173 FtUGTs in F. tataricum based on their conserved UDPGT domain. Phylogenetic analysis of FtUGTs with 73 Arabidopsis UGTs clustered them into 21 families. FtUGTs from the same family usually had similar gene structure and motif compositions. Most of FtUGTs did not contain introns or had only one intron. Tandem repeats contributed more to FtUGTs amplification than segmental duplications. Expression analysis indicates that FtUGTs are widely expressed in various tissues and likely play important roles in plant growth and development. The gene expression analysis response to different abiotic stresses showed that some FtUGTs were involved in response to drought and cadmium stress. Our study provides useful information on the UGTs in F. tataricum, and will facilitate their further study to better understand their function. CONCLUSIONS: Our results provide a theoretical basis for further exploration of the functional characteristics of FtUGTs and for understanding the growth, development, and metabolic model in F. tataricum.

Identification and Biological Evaluation of a Novel Small-Molecule Inhibitor of Ricin Toxin.

The plant-derived toxin ricin is classified as a type 2 ribosome-inactivating protein (RIP) and currently lacks effective clinical antidotes. The toxicity of ricin is mainly due to its ricin toxin A chain (RTA), which has become an important target for drug development. Previous studies have identified two essential binding pockets in the active site of RTA, but most existing inhibitors only target one of these pockets. In this study, we used computer-aided virtual screening to identify a compound called RSMI-29, which potentially interacts with both active pockets of RTA. We found that RSMI-29 can directly bind to RTA and effectively attenuate protein synthesis inhibition and rRNA depurination induced by RTA or ricin, thereby inhibiting their cytotoxic effects on cells in vitro. Moreover, RSMI-29 significantly reduced ricin-mediated damage to the liver, spleen, intestine, and lungs in mice, demonstrating its detoxification effect against ricin in vivo. RSMI-29 also exhibited excellent drug-like properties, featuring a typical structural moiety of known sulfonamides and barbiturates. These findings suggest that RSMI-29 is a novel small-molecule inhibitor that specifically targets ricin toxin A chain, providing a potential therapeutic option for ricin intoxication.

The role of NaHS pretreatment in improving salt stress resistance in foxtail millet seedlings: physiological and molecular mechanisms.

Salt stress is a prevailing abiotic stress in nature, with soil salinization becoming a pressing issue worldwide. High soil salinity severely hampers plant growth and leads to reduced crop yields. Hydrogen sulfide (H2S), a gas signal molecule, is known to be synthesized in plants exposed to abiotic stress, contributing to enhanced plant stress resistance. To investigate the impact of sodium hydrosulfide hydrate (NaHS, a H2S donor) on millet's response to salt stress, millet seedlings were subjected to pretreatment with 200 µM NaHS, followed by 100 mM NaCl stress under soil culture conditions. The growth, osmotic adjustment substances, antioxidant characteristics, membrane damage, and expression levels of related genes in millet seedlings were detected and analyzed. The results showed that NaHS pretreatment alleviated the inhibition of salt stress on the growth of foxtail millet seedlings, increased the proline content and antioxidant enzyme activities, as well as the expression levels of SiASR4, SiRPLK35 and SiHAK23 genes under salt stress. These findings demonstrated that NaHS pretreatment can enhance salt tolerance in foxtail millet seedlings by regulating the content of osmotic adjustment substances and antioxidant enzyme activity, reducing electrolyte permeability, and activating the expression of salt-resistant genes.

d-Glucosamine induces circadian phase delay by promoting BMAL1 degradation through AMPK/mTOR pathway.

Circadian rhythms are closely linked to the metabolic network through circadian feedback regulation. The hexosamine biosynthetic pathway (HBP) is a branch of glucose metabolism that affects circadian rhythms through the O-linked N-acetylglucosamine modification (O-GlcNAcylation) of clock proteins. Here, we found out that, among the downstream metabolites regulated by d-glucosamine (GlcN) in HBP salvage pathway, only GlcN is able to induce circadian phase delay both in vitro and in vivo. Mechanistic studies indicated that the phase-shift induced by GlcN is independent of O-GlcNAcylation. Instead, GlcN selectively up-regulates p-AMPK activity, leading to the inhibition of mTOR signaling pathway, and thus down-regulation of p-BMAL1 both in human cell line and mouse tissues. Moreover, GlcN promoted BMAL1 degradation via proteasome pathway. These findings reveal a novel molecular mechanism of GlcN in regulating clock phase and suggest the therapeutic potential of GlcN as new use for an old drug in the future treatment of shift work and circadian misalignment.

Synthesis of hierarchical Cd-Ni-MOF micro/nanostructures and derived Cd-Ni-MOF/CdS/NiS hybrid photocatalysts for efficient photocatalytic hydrogen evolution.

Hierarchical micro/nanostructures assembled from nanorods and nanosheets have become promising candidates for photocatalysis. In this work, a series of hierarchical Cd-Ni-MOF micro/nanostructures, assembled from nanosheets and nanorods, were fabricated via a two-step solvothermal process involving the partial replacement of Ni2+ with Cd2+ in the Ni-MOF-74 structure. Different morphologies were obtained by considering different volume ratios of DMF and ethanol as the solvent during synthesis. Hierarchical Cd-Ni-MOF-T/CdS/NiS hybrid micro/nanostructures were synthesized by Ni2+ and Cd2+ exchange of Cd-Ni-MOFs with S2-. The as-prepared samples, which were composed of thin nanosheets alone, exhibited the best photocatalytic H2 evolution rate of about 40.08 mmol g-1 h-1. The p-n junction between CdS and NiS was found to be beneficial for the migration of photogenerated electrons from the conduction band (CB) of NiS to the CB of CdS. The heterojunction between CdS and Cd-Ni-MOF-T further promoted the transfer of an electron from the CB of CdS to the CB of Cd-Ni-MOF-T. Thus, this study demonstrated that hierarchical Cd-Ni-MOF-T/CdS/NiS architectures have a large specific surface area, leading to significantly improved photocatalytic activity.

Treadmill exercise promotes E3 ubiquitin ligase to remove amyloid ß and P-tau and improve cognitive ability in APP/PS1 transgenic mice.

BACKGROUND: Moderate physical exercise is conducive to the brains of healthy humans and AD patients. Previous reports have suggested that treadmill exercise plays an anti-AD role and improves cognitive ability by promoting amyloid clearance, inhibiting neuronal apoptosis, reducing oxidative stress level, alleviating brain inflammation, and promoting autophagy-lysosome pathway in AD mice. However, few studies have explored the relationships between the ubiquitin-proteasome system and proper exercise in AD. The current study was intended to investigate the mechanism by which the exercise-regulated E3 ubiquitin ligase improves AD. METHODS: Both wild type and APP/PS1 transgenic mice were divided into sedentary (WTC and ADC) and exercise (WTE and ADE) groups (n = 12 for each group). WTE and ADE mice were subjected to treadmill exercise of 12 weeks in order to assess the effect of treadmill running on learning and memory ability, Aß plaque burden, hyperphosphorylated Tau protein and E3 ubiquitin ligase. RESULTS: The results indicated that exercise restored learning and memory ability, reduced Aß plaque areas, inhibited the hyperphosphorylation of Tau protein activated PI3K/Akt/Hsp70 signaling pathway, and improved the function of the ubiquitin-proteasome system (increased UCHL-1 and CHIP levels, decreased BACE1 levels) in APP/PS1 transgenic mice. CONCLUSIONS: These findings suggest that exercise may promote the E3 ubiquitin ligase to clear ß-amyloid and hyperphosphorylated Tau by activating the PI3K/Akt signaling pathway in the hippocampus of AD mice, which is efficient in ameliorating pathological phenotypes and improving learning and memory ability.

State-dependent modulation of thalamocortical oscillations by gamma light flicker with different frequencies, intensities, and duty cycles.

Rhythmic light flickers have emerged as useful tools to modulate cognition and rescue pathological oscillations related to neurological disorders by entrainment. However, a mechanistic understanding of the entrainment for different brain oscillatory states and light flicker parameters is lacking. To address this issue, we proposed a biophysical neural network model for thalamocortical oscillations (TCOs) and explored the stimulation effects depending on the thalamocortical oscillatory states and stimulation parameters (frequency, intensity, and duty cycle) using the proposed model and electrophysiology experiments. The proposed model generated alpha, beta, and gamma oscillatory states (with main oscillation frequences at 9, 25, and 35 Hz, respectively), which were successfully transmitted from the thalamus to the cortex. By applying light flicker stimulation, we found that the entrainment was state-dependent and it was more prone to induce entrainment if the flicker perturbation frequency was closer to the endogenous oscillatory frequency. In addition, endogenous oscillation would be accelerated, whereas low-frequency oscillatory power would be suppressed by gamma (30-50 Hz) flickers. Notably, the effects of intensity and duty cycle on entrainment were complex; a high intensity of light flicker did not mean high entrainment possibility, and duty cycles below 50% could induce entrainment easier than those above 50%. Further, we observed entrainment discontinuity during gamma flicker stimulations with different frequencies, attributable to the non-linear characteristics of the network oscillations. These results provide support for the experimental design and clinical applications of the modulation of TCOs by gamma (30-50 Hz) light flicker.

Irisin, an Effective Treatment for Cardiovascular Diseases?

Irisin, as one of the myokines induced by exercise, has attracted much attention due to its important physiological functions such as white fat browning, the improvement in metabolism, and the alleviation of inflammation. Despite the positive role that irisin has been proven to play in the prevention and treatment of cardiovascular diseases, whether it can become a biomarker and potential target for predicting and treating cardiovascular diseases remains controversial, given the unreliability of its detection methods, the uncertainty of its receptors, and the species differences between animals and humans. This paper was intended to review the role of irisin in the diagnosis and treatment of cardiovascular diseases, the potential molecular mechanism, and the urgent problems to be solved in hopes of advancing our understanding of irisin as well as providing data for the development of new and promising intervention strategies by discussing the causes of contradictory results.

Synthesis of hierarchical and flower-like TiO2 nanowire microspheres as biocompatible cell carriers.

Fibrous materials are of great interest in the development of tissue regenerative matrix. However, synthesis of inorganic fibrous microspheres as cell carriers is a great challenge. In this study, we for the first time report on the synthesis of novel hierarchical and flower-like TiO2 nanowire (NW) microspheres as biocompatible cell carriers. TiO2 NW microspheres were synthesized through in situ alkali hydrothermal treatment of the TiO2 nanoparticle (NP) microspheres and their microstructure, formation mechanism and in vitro biocompatibility were evaluated. SEM observations show that the resulting TiO2 NW microspheres were constructed by a large number of NWs with the diameter of 10-20 nm and exhibited a flower-like and hierarchical morphology with the diameter of 400-600 µm. XRD patterns indicate that TiO2 NW microspheres were constructed by both rutile and anatase phase of TiO2. FT-IR spectra reveal that Ti-O-Ti bonds were involved in TiO2 NW microspheres. In vitro biocompatibility was evaluated by seeding the fibroblast L929 cells on the microspheres. A conventional MTT assay quantitatively indicates that the TiO2 NW microspheres favored adhesion and proliferation of cells and were biocompatible, while SEM observations qualitatively confirmed that the cells were well grown on the surface of TiO2 NW microspheres. Thus, the as-synthesized TiO2 NW microspheres would be applicable to novel and biocompatible cell carriers.

Triptolide and atorvastatin synergistically promote hepatotoxicity in cultured hepatocytes and female Sprague-Dawley rats by inhibiting pregnane X receptor-mediated transcriptional activation of CYP3A4.

Triptolide (TP), an active component of Tripterygium wilfordii Hook. F, has been widely used in China for treating autoimmune and inflammatory diseases, and has also been validated by modern science and developed as a candidate anti-cancer treatment. However, liver toxicity of TP has seriously hindered its use and development, the clinical features and primary toxicological mechanism have been unclear. Considering the major target regulation mechanism of TP is the suppression of global transcription regulated by RNAPII, which is closed related with the detoxification of drugs. This paper tries to verify the synergistic liver injury and its mechanism of TP when co-administered with CYP3A4 substrate drug. The experiments showed that TP dose-dependently blocked transcriptional activation of CYP3A4 in both hPXR and hPXR-CYP3A4 reporter cell lines, lowered the mRNA and protein expression of PXR target genes such as CYP3A1, CYP2B1, and MDR1, and inhibited the functional activity of CYP3A in a time- and concentration-dependent manner in sandwich-cultured rat hepatocytes (SCRH) and female Sprague-Dawley (f-SD) rats. Furthermore, TP combined with atorvastatin (ATR), the substrate of CYP3A4, synergistically enhanced hepatotoxicity in cultured HepG2 and SCRH cells (CI is 0.38 and 0.29, respectively), as well as in f-SD rats, with higher exposure levels of both drugs. These results clearly indicate that TP inhibits PXR-mediated transcriptional activation of CYP3A4, leading to a blockade on the detoxification of itself and ATR, thereby greatly promoting liver injury. This study may implies the key cause of TP related liver injury and provides experimental data for the rational use of TP in a clinical scenario.

Facile synthesis, microstructure, formation mechanism, in vitro biocompatibility, and drug delivery property of novel dendritic TiO2 nanofibers with ultrahigh surface area.

In this study, novel dendritic TiO2 nanofibers (D-Ti NFs) with ultrahigh surface area were for the first time synthesized through in situ hydrothermal treatment of the electrospun TiO2 nanofibers (E-Ti NFs) and their microstructure, formation mechanism, in vitro biocompatibility and drug delivery property were investigated. SEM observations showed that D-Ti NFs had dendritic and fibrous morphology with the controllable diameter of 899 ± 26 nm to 1955 ± 64 nm. TEM observations showed the individual D-Ti NF was assembled by numerous primary nanowires with the diameter of 7 ± 1 nm. XRD pattern showed that the D-Ti NFs displayed the characteristic peaks at 24.1°, 28.1°, and 48.3°, all of which were assigned to hydrogen titanium oxide hydrate. BET measurement indicated that D-Ti NFs had the ultrahigh specific surface area of 213.41 m2/g. D-Ti NFs showed good in vitro biocompatibility when exposed to Hela cells. After soaked in solution of tetracycline hydrochloride (TH, one of the representative antibiotics, model drug), D-Ti NFs supported the loading of TH with the loading efficiency of 63.73 ± 4.61%. TH-loaded D-Ti NFs supported a sustained release model for TH when immersed in the phosphate buffer saline. After separately incubated with two representative types of bacteria: Escherichia coli and Staphylococcus aureus, TH-loaded D-Ti NFs showed excellent antibacterial property to inhibit the growth of the bacteria, indicating that the released TH was biologically active.

CM082, a novel angiogenesis inhibitor, enhances the antitumor activity of gefitinib on epidermal growth factor receptor mutant non-small cell lung cancer in vitro and in vivo.

BACKGROUND: CM082 is a novel angiogenesis inhibitor targeting vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR). The purpose of this research was to evaluate the antitumor activity of CM082 combined with gefitinib on epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) in vitro and in vivo. METHODS: The effect of CM082 on human umbilical vein endothelial cells (HUVECs) was assessed. In vitro and in vivo efficacy of CM082 combined with gefitinib on EGFR NSCLC cell lines (HCC827 harboring E746_A750 deletion and H3255 harboring L858R) and a xenograft model was evaluated. RESULTS: CM082 inhibited VEGF-induced cell growth, phosphorylation of VEGFR and downstream signaling molecules, tube formation, and cell migration of HUVECs. Furthermore, CM082 combined with gefitinib was more effective in inhibiting growth and colony formation and inducing apoptosis of H3255 and HCC827 cells in vitro than monotherapy. Moreover, tumor growth inhibition by the combination in a H3255 xenograft model was 107.7% more than that by gefitinib (93.6%) (P < 0.0001) and CM082 (51.4%) (P < 0.0001) alone. In addition, coadministration had a better effect on inhibiting cell proliferation, inducing apoptosis, and inhibiting the expression of CD31 and VEGF-A. The combination therapy had a stronger inhibition effect on STAT3 phosphorylation than monotherapy. CONCLUSIONS: CM082, a novel angiogenesis inhibitor, enhances the antitumor activity of gefitinib on EGFR mutant NSCLC by inhibiting proliferation and angiogenesis and promoting apoptosis of tumor cells. KEY POINTS: Significant findings of the study CM082, a novel angiogenesis inhibitor, enhances the antitumor activity of gefitinib on EGFR mutant NSCLC by inhibiting proliferation and angiogenesis and promoting apoptosis of tumor cells. What this study adds These findings justify evaluation of the efficacy of CM082 combined with gefitinib in patients with EGFR mutant advanced NSCLC in clinical trials.

The iron chelator desferrioxamine synergizes with chemotherapy for cancer treatment.

BACKGROUND: Cisplatin (CDDP) resistance remains a major obstacle for treatment of ovarian cancer. Iron contributes to the growth and reproduction of malignant cells, thus iron chalators can inhibit the growth of tumor cells by depleting the intracellular iron pool. The iron chelator, desferrioxamine (DFO), has performed anticancer in previous study. The aim of our study is to determine the correlation between iron-deprivation and tumor chemosensitivity in ovarian cancer. METHODS: To investigate the prognostic value of ferritin light (FTL), ferroportin (FPN), hepcidin (HAMP) and divalent metal-ion transporter-1 (DMT1) in ovarian cancer, the Kaplan-Meier analysis and the Gene Expression Profiling Interactive Analysis (GEPIA) were used. The ovarian cancer cell lines (SKOV-3 and OVCAR-3) were exposed to a gradient concentration of DFO (10, 20, 50, 100, 200 µM) and CDDP (1, 5, 10, 50,100 µM) for 24 h. The protein expression of FTL was tested. The expression of cancer stem cell (CSC) markers, including Sox2, Nanog and C-myc, were downregulated with treatment of DFO. Also, the mamosphere formation and the plation of CD44+/high/CD133+/high and Aldehyde dehydrogenase (ALDH)+/high SKOV-3 cells were reduced after treatment for 7d. Furthermore, we detected the expression of p53, BCL-2, BAX, and caspase-8. RESULTS: The survival analysis revealed that high expression of FTL, DMT1, HAMP, showed poor overall survival (OS) in ovarian cancer patients. Our combined data found that DFO could effectively inhibit CSCs, improve the resistance to chemotherapy, and significantly enhanced the efficacy of CDDP therapy in vitro in promoting apoptosis. Besides, targeting molecular targets, including BAX, BCL-2, p53 and caspase-8 could serve as the clinical biomarkers to evaluate the effects of ovarian cancer. It is reasonable to believe that DFO adjuvant therapy in combination with CDDP chemotherapy can promote the improvement of treatment response in ovarian cancer patients. CONCLUSION: Our research suggests the experimental evidence for DFO and CDDP as a new effective combination therapy to enhance the efficacy of chemical therapy in ovarian cancer.

An Important Factor Affecting the Supercapacitive Properties of Hydrogenated TiO2 Nanotube Arrays: Crystal Structure.

Employing a suitable crystal structure can significantly modify the electrochemical performances of materials. Herein, hydrogenated TiO2 nanotube arrays with <001> orientation and different rutile/anatase ratio were fabricated via anodisation, high-temperature annealing and electrochemical hydrogenation. The crystal structure was determined by TEM and X-ray diffraction pattern refinement of whole powder pattern fitting. Combined with the model of anatase to rutile transformation and the characterisation of crystal structure, the effect of phase transition on the super capacitive properties of <001> oriented hydrogenated TiO2 nanotube arrays was discussed. The results suggested that the anatase grains were characterised by orientation in <001> direction with plate crystallite and stacking vertically to the substrate resulting in excellent properties of electron/ion transport within hydrogenated TiO2 nanotube arrays. In addition, the specific capacitance of <001> oriented hydrogenated TiO2 could be further improved from 20.86 to 24.99 mF cm-2 by the partial rutile/anatase transformation due to the comprehensive effects of lattice disorders and rutile, while the good rate performance and cyclic stability also retained.

WX20120108, a novel IAP antagonist, induces tumor cell autophagy via activating ROS-FOXO pathway.

Recently, inhibitor of apoptosis proteins (IAPs) and some IAP antagonists were found to regulate autophagy, but the underlying mechanisms remain unclear. WX20120108 is an analogue of GDC-0152 (a known IAP antagonist) and displays more potent anti-tumor and autophagy-regulating activity in tumor cells, we investigated the regulatory mechanisms underlying WX20120108-induced autophagy. Using molecular docking and fluorescence polarization anisotropy (FPA) competitive assay, we first demonstrated that WX20120108, acting as an IAP antagonist, bound to the XIAP-BIR3, XIAP BIR2-BIR3, cIAP1 BIR3, and cIAP2 BIR3 domains with high affinities. In six cancer cell lines, WX20120108 inhibited the cell proliferation with potencies two to ten-fold higher than that of GDC-0152. In HeLa and MDA-MB-231 cells, WX20120108 induced caspase-dependent apoptosis and activated TNFα-dependent extrinsic apoptosis. On the other hand, WX20120108 induced autophagy in HeLa and MDA-MB-231 cells in dose- and time-dependent manners. We revealed that WX20120108 selectively activated Foxo3, evidenced by Foxo3 nuclear translocation in both gene modified cell line and HeLa cells, as well as the upregulated expression of Foxo3-targeted genes (Bnip3, Pik3c3, Atg5, and Atg4b), which played a key role in autophagy initiation. WX20120108-induced autophagy was significantly suppressed when Foxo3 gene was silenced. WX20120108 dose-dependently increased the generation of reactive oxygen species (ROS) in HeLa cells, and WX20120108-induced Foxo3 activation was completely blocked in the presence of catalase, a known ROS scavenger. However, WX20120108-induced ROS generation was not affected by cIAP1/2 or XIAP gene silencing. In conclusion, WX20120108-induced autophagy relies on activating ROS-Foxo3 pathway, which is independent of IAPs. This finding provides a new insight into the mechanism of IAP antagonist-mediated regulation of autophagy.

High-content analysis boosts identification of the initial cause of triptolide-induced hepatotoxicity.

Triptolide (TP) has been widely used in China for more than 40 years as an immunosuppressive agent. Recently, serious concerns have been raised over TP-induced liver injury, though the real hepatotoxic mechanism is still unclear, particularly in terms of the initial cause. To our knowledge, this study is the first to screen systematically the mechanism of TP-induced toxicity through a global cytotoxicity profile high-content analysis using three independent cytotoxic assay panels with multiple endpoints of cytotoxicity, including cell loss, mitochondrial membrane potential, nuclear membrane permeability, manganese superoxide dismutase, phosphorylated gamma-H2AX, light chain 3B, lysosome, reactive oxygen species and glutathione. We assessed nine parameters and four stress response pathway models by labeling nuclear factor erythroid 2-related factor 2, activating transcription factor 6, hypoxia inducible factor 1α and nuclear factor κB and found that all testing parameters except glutathione and manganese superoxide dismutase showed concentration- and time-dependent changes, as well as increased cell loss after TP treatment. Considering that RNA polymerase II is the molecular target of TP, we quantified transcription from inducible genes, bromodeoxyuridine incorporation, and expression from transiently transfected green fluorescence protein plasmids in HepG2 cells. The results show that inhibition of global transcription by TP took place at earlier times and at lower concentrations than those observed for cell death. Therefore, global transcriptional suppression and the cell dysfunction it drives play a central role in TP-induced hepatotoxicity. This provides valuable information for the safe use of TP in the clinic.

Response of gas-exchange characteristics and chlorophyll fluorescence to acute sulfur dioxide exposure in landscape plants.

To explore the toxicity and action mechanism of acute sulfur dioxide (SO2) on urban landscape plants, a simulated SO2 stress environment by using fumigation chamber involving increasing SO2 concentration (0, 25, 50, 100, 200 mg m-3) was carried out among three species. After 72 h of exposure, SO2-induced oxidative damage indicated by electrolyte leakage increased with higher dose of SO2. Meanwhile, SO2 decreased the contents of chlorophyll a, chlorophyll b and carotenoid and increased the contents of sulfur. Net photosynthetic rate (Pn) decreased as a result of stomatal closure when SO2 dose was lower than 50 mg m-3, out of this range, non-stomatal limitation play a dominant role in the decline of Pn. Simultaneous measurements of chlorophyll fluorescence imaging (CFI) also revealed that the maximal quantum efficiency of PSII photochemistry in dark-adapted state (Fv/Fm) and the realized operating efficiency of PSII photochemistry (Fq'/Fm') was reduced by SO2 in a dose-dependent manner. In addition, the maximum quantum efficiency of PSII photochemistry in light-adapted state (Fv'/Fm') and the PSII efficiency factor (Fq'/Fv') decreased when exposure to SO2. These results implied that acute SO2 exposure induced photoinhibition of PSII reaction centers in landscape plants. Our study also indicated that different urban landscape plant species resist differently to SO2: Euonymus kiautschovicus > Ligustrum vicaryi > Syringa oblata according to gas-exchange characteristics and chlorophyll fluorescence responses.

Procaspase-3-activating compound 1 stabilizes hypoxia-inducible factor 1α and induces DNA damage by sequestering ferrous iron.

Procaspase-3-activating compound 1 (PAC-1) induces procaspase-3 activation via zinc chelation. However, whether PAC-1 employs other mechanisms remains unknown. Here we systematically screened for potent PAC-1 targets using 29 enhanced green fluorescent protein-labeled reporter cell lines and identified hypoxia-inducible factor 1α (HIF1α) and RAD51 pathways as PAC-1 targets. These results were verified in HepG2 cells and two other cancer cell lines. Mechanistically, PAC-1 specifically blocked HIF1α hydroxylation and upregulated HIF1α target genes. In addition, DNA damage, G1/S cell cycle arrest, and the inhibition of DNA synthesis were induced following PAC-1 administration. Interestingly, by using ferrozine-iron sequestration and iron titration assays, we uncovered the iron sequestering capacity of PAC-1. Additionally, the expression levels of iron shortage-related genes were also increased in PAC-1-treated cells, and iron (II) supplementation reversed all of the observed cellular responses. Thus, our results indicate that PAC-1 induces HIF1α stabilization and DNA damage by sequestering ferrous iron.