Disruption of NADPH homeostasis by total flavonoids from Adinandra nitida Merr. ex Li leaves triggers ROS-dependent p53 activation leading to apoptosis in non-small cell lung cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Adinandra nitida Merr. ex Li leaves serve as a herbal tea and hold a significant role in traditional Chinese medicine, being applied to assist in tumor treatment. Flavonoids present the primary bioactive constituents in Adinandra nitida Merr. ex Li leaves. AIM OF THE STUDY: To explore the potential of total flavonoids from Adinandra nitida Merr. ex Li Leaves (TFAN) in inhibiting non-small cell lung cancer (NSCLC) and further elucidate the underlying mechanisms. MATERIALS AND METHODS: Human NSCLC cell lines and normal lung cell line were employed to assess the impact of TFAN (0-160 µg/mL for 24, 28 and 72 h) on cell proliferation in vitro. Immunofluorescence (IF) staining gauged p53 expression changes in NSCLC cells under TFAN present condition (150 µg/mL for 24 h). In vivo study utilized NSCLC cell derived xenograft tumors in nude mice, administering TFAN orally (200 and 400 mg/kg) for 14 days. Immunohistochemistry assessed Cleaved Caspase 3 expression change in A549 xenograft tumors treated with TFAN (400 mg/kg for 14 days). RNA-seq and KEGG analysis identified gene expression changes and enriched processes in A549 xenograft tumors treated with TFAN. CM-H2DCFDA and metabolomics assessed ROS level and GSH/GSSG pool changes in A549 cells under TFAN present condition. Cell viability assay and IF staining assessed A549 cell proliferation and p53 expression changes under H2O2-induced oxidative stress (0-40 µM for 24 h) and TFAN present conditions. GSEA and N-Acetyl-L-cysteine (NAC) rescue (0-1 µM for 24 h) analyzed the impact of TFAN on GSH de novo synthesis. NADPH/NADP+ pool measurement and NADPH rescue (0-10 µM for 24 h) analyzed the impact of TFAN on GSH salvage synthesis. GC-FID and HPLC-MS were utilized to detect ethanol and ethyl acetate residues, and to characterize the chemical constituents in TFAN, respectively. The total flavonoid content of TFAN was determined using a 330 nm wavelength. RESULTS: TFAN significantly inhibited A549 cells (wild-type p53) but not NCI-H1299 cells (p53-deficient), NCI-H596 cells (p53-mutant) or BEAS-2B in vitro. IF staining validated p53 genotype for the cell lines and revealed an increase in p53 expression in A549 cells after TFAN treatment. In vivo, TFAN selectively inhibited A549 xenograft tumor growth without discernible toxicity, inducing apoptosis evidenced by Cleaved Caspase 3 upregulation. RNA-seq and KEGG analysis suggested ROS biosynthesis was involved in TFAN-induced p53 activation in A549 cells. Elevated ROS level in TFAN-treated A549 cells were observed. Moreover, TFAN sensitized A549 cells to H2O2-induced oxidative stress, with higher p53 expression. Additionally, A549 cells compensated with GSH de novo synthesis under TFAN present condition, confirmed by GSEA and NAC rescue experiment. TFAN disrupted NADPH homeostasis to impair GSH salvage biosynthesis, supported by NADPH/NADP+ change and NADPH rescue experiment. The chemical constituents of TFAN, with acceptable limits for ethanol and ethyl acetate residues and a total flavonoid content of 68.87%, included Catechin, Epicatechin, Quercitroside, Camellianin A, and Apigenin. CONCLUSION: The disruption of NADPH homeostasis by TFAN triggers ROS-dependent p53 activation that leads to apoptotic cell death, ultimately suppressing NSCLC growth. These findings offer potential therapeutic implications of Adinandra nitida Merr. ex Li leaves in combating NSCLC.

Time-resolved ARPES with probe energy of 6.0/7.2 eV and switchable resolution configuration.

We present a detailed exposition of the design for time- and angle-resolved photoemission spectroscopy using a UV probe laser source that combines the nonlinear effects of ß-BaB2O4 and KBe2BO3F2 optical crystals. The photon energy of the probe laser can be switched between 6.0 and 7.2 eV, with the flexibility to operate each photon energy setting under two distinct resolution configurations. Under the fully optimized energy resolution configuration, we achieve an energy resolution of 8.5 meV at 6.0 eV and 10 meV at 7.2 eV. Alternatively, switching to the other configuration enhances the temporal resolution, yielding a temporal resolution of 72 fs for 6.0 eV and 185 fs for 7.2 eV. We validated the performance and reliability of our system by applying it to measuring two typical materials: the topological insulator MnBi2Te4 and the excitonic insulator candidate Ta2NiSe5.

Time-resolved ARPES with tunable 12-21.6 eV XUV at 400 kHz repetition rate.

Time-resolved and angle-resolved photoemission spectroscopy (trARPES) is a powerful method to detect the non-equilibrium electronic structure in solid systems. In this study, we report a trARPES apparatus with tunable photon energy selectively among 12, 16.8, and 21.6 eV at a repetition rate of 400 kHz. The energy and temporal resolutions of the three harmonics are determined as 104/111/157 meV and 276/190/154 fs, respectively. The photon flux on the sample is estimated to be 1010-1011 photons/s by using a photodiode. Finally, the performance of this setup is verified by both equilibrium and non-equilibrium ARPES measurements on topological materials Zr2Te2P and Bi2Se3. Meanwhile, the importance of the tunability of the extreme ultraviolet (XUV) source is highlighted by comparing experimental results measured with the three different photon energies.

High-flux wavelength tunable XUV source in the 12-40.8†eV photon energy range with adjustable energy and time resolution for Tr-ARPES applications.

High-order harmonic generation (HHG) has a broad spectrum covering vacuum ultraviolet to extreme ultraviolet (XUV) bands, which is useful for applications involving material analyses at different information depths. Such an HHG light source is perfect for time- and angle-resolved photoemission spectroscopy. Here, we demonstrate a high-photon flux HHG source driven by a two-color field. Applying a fused silica compression stage to reduce the driving pulse width, we obtained a high XUV photon flux of 2 × 1012 phs/s @21.6 eV on target. We designed a classical diffraction mounted (CDM) grating monochromator that can achieve a wide range of photon energy from 12 to 40.8 eV, while the time resolution is improved by reducing the pulse front tilt after the harmonic selection. We designed a spatial filtering method to adjust the time resolution using the CDM monochromator and significantly reduced the pulse front tilt of the XUV pulses. We also demonstrate a detailed prediction of the energy resolution broadening which is caused by the space charge effect.

Pathogenicity and anti-infection immunity of animal H3N2 and H6N6 subtype influenza virus cross-species infection with tree shrews.

Animal influenza viruses can spread across species and pose a fatal threat to human health due to the high pathogenicity and mortality. Animal models are crucial for studying cross-species infection and the pathogenesis of influenza viruses. Tupaia belangeri (tree shrew) has been emerging as an animal model for multiple human virus infections recently because of the close genetic relationship and phylogeny with humans. So far, tree shrew has been reported to be susceptible to human influenza virus subtype H1N1, avian influenza viruses subtype H9N2, subtype H5N1, and subtype H7N9. However, the pathogenicity, infection, and immunity of swine and land avian influenza viruses with low pathogenicity and the potential to jump to humans remain largely unexplored in the tree shrew model. Previously, our team has successfully isolated the newly emerging swine influenza virus subtype H3N2 (A/Swine/GX/NS2783/2010, SW2783) and avian influenza virus subtype H6N6 (A/CK/ZZ/346/2014, ZZ346). In this study, we observed the pathogenicity, immune characteristics, and cross-species infection potential ability of SW2783 and ZZ346 strains in tree shrew model with 50% tissue culture infective dose (TCID50), hematoxylin and eosin (HE) staining, immunohistochemistry (IHC), real-time quantitative PCR (qRT-PCR) and other experimental methods. Both animal-borne influenza viruses had a strong ability on tissue infection in the turbinate and the trachea of tree shrews in vitro, in which SW2783 showed stronger replication ability than in ZZ346. SW2783 and ZZ346 both showed pathogenic ability with infected tree shrews model in vivo without prior adaptive culture, which mainly happened in the upper respiratory tract. However, the infection ability was weak, the clinical symptoms were mild, and the histopathological changes in the respiratory tract were relatively light. Furthermore, innate immune responses and adaptive immunity were observed in the tree shrew model after the infection of SW2783 and ZZ346 strains. We observed that the unadapted SW2783 and ZZ346 virus could transmit among tree shrews by direct contact. We also observed that SW2783 virus could transmit from tree shrews to guinea pigs. These results indicated that both animal-borne influenza viruses could induce similar pathogenicity and immune response to those caused by human-common influenza viruses. Tree shrews may be an excellent animal model for studying the interaction between the influenza virus and the host and the cross-species infection mechanism of the animal influenza virus.

Mangiferin exerts neuroprotective activity against lead-induced toxicity and oxidative stress via Nrf2 pathway.

Objective: The study was designed to assess the beneficial role of mangiferin (MGN) in lead (Pb)-induced neurological damages in the activation of Nrf2-governed enzymes, genes and proteins. Methods: A total of 96 weaned Wistar rats (48 males and 48 females, 26- to 27-day-old), weighing 50-80 g were used. The experiment was performed in six groups: normal group (control, n = 16), model group (chronic Pb exposed, n = 16), Dimercaptosuccinic acid (DMSA)-treated group (positive control, Pb + DMSA, n = 16), three MGN-treated groups with different doses (Pb + MGN, n = 48). Normal group freely had access to purified water. DMSA-treated group was given DMSA, which was clinically used as the standard treatment for moderate Pb poisoning, at 50 mg/kg (2 mL suspension with purified water) by intragastric gavage (ig) 4 continual days a week for 4 weeks, MGN-treated groups were given MGN at 50, 100, or 200 mg/kg (2 mL suspension with purified water) by ig daily for 4 weeks. At the end of the treatment, all rats were sacrificed and the brain samples were collected. The haematoxylin and eosin (H&E) staining was used for observation of histopathology. Commercial kit, real-time quantitative polymerase chain reaction (RT-qPCR), Western-blot and immunohistochemistry (IHC) detection were used to detect the mRNA and protein expression. Results: Eight weeks exposure to Pb-containing water resulted in pathological alterations, anti-oxidative system disorder in the brain, all of which were blocked by MGN in a Nrf2-dependent manner. Nrf2 downstream enzymes such as HO-1, NQO1, γ-GCS were activated. Nrf2, GCLC, GCLM, HO-1 mRNA and total Nrf2, Nuclear Nrf2, γ-GCS, HO-1 protein expression were affected too. Conclusion: MGN ameliorated morphological damage in the hippocampus. Its neuroprotective effects were achieved by the activation of the Nrf2 downstream genes. The data from this in vitro study indicates that MGN targeting Nrf2 activation is a feasible approach to reduce adverse health effects associated with Pb exposure. Thus, MGN could be an effective candidate agent for the Pb-induced oxidative stress and neurotoxicity in the human body.

Mangiferin suppresses allergic asthma symptoms by decreased Th9 and Th17 responses and increased Treg response.

Mangiferin is the major bioactive ingredient in the leaves of Mangifera indica L., Aqueous extract of such leaves have been traditionally used as an indigenous remedy for respiratory diseases including cough and asthma in Traditional Chinese Medicine. Mangiferin was shown to exert its anti-asthmatic effect by modulating Th1/Th2 cytokines imbalance via STAT6 signaling pathway. However, compelling evidence indicated that subtypes of T helpers and regulatory T cells other than Th1/Th2 were also involved in the pathogenesis of asthma. In current study, we investigated the effects of mangiferin on the differentiation and function of Th9, Th17 and Treg cells in a chicken egg ovalbumin (OVA)-induced asthmatic mouse model. Mangiferin significantly attenuated the symptoms of asthma attacks, reduced the total number of leukocytes, EOS and goblet cells infiltration in lung. Simultaneously, treatment with mangiferin remarkably decreased the proportion of Th9 and Th17 cells; reduced the levels of IL-9, IL-17A; inhibited the expression of PU.1 and RORγt in lung. However, the proportion of Treg cells, the expression of IL-10, TGF-ß1 and Foxp3 were increased by mangiferin. Our data suggest that mangiferin exerted anti-asthmatic effect through decreasing Th9 and Th17 responses and increasing Treg response in OVA-induced asthmatic mouse model.

[The enhanced expression of Toll-like receptor 2 in CD8(+)T cells of chronic HBV infected patients].

Objective To detect the level of Toll-like receptor 2 (TLR2) in CD8(+)T cells of chronic HBV infected patients without antiviral treatment. Methods Forty chronic HBV infected donors without antivirus treatment and 19 healthy donors were enrolled in our study. All donors were divided into three groups: a high viral load (HBV DNA>1×10(4) copies/mL) group, a low viral load (HBV DNA>1×10(4) copies/mL) group, and a healthy control group. After the isolation and staining of peripheral blood mononuclear cells (PBMCs), the levels of TLR2, CD38, HLA-DR, CD95, programmed death-1 (PD-1) in CD8(+)T cells were detected by flow cytometry to further analyze the correlations between TLR2 and CD38, HLA-DR, CD95, PD-1. Results The expression of TLR2 in CD8(+)T cells of the HBV infected patients were significantly higher than that in the healthy donors. The expression of TLR2 in CD8(+)T cells of the low viral load group was significantly higher than that in the high viral load group. There were no correlations between the expression of TLR2 and the expressions of CD38, HLA-DR, CD95 and PD-1 in CD8(+)T cells in the chronic HBV infected patients. Conclusion The expression of TLR2 in CD8(+)T cells increases in chronic HBV infected patients.

Identification of the full-length ß-actin sequence and expression profiles in the tree shrew (Tupaia belangeri).

The tree shrew (Tupaia belangeri) diverges from the primate order (Primates) and is classified as a separate taxonomic group of mammals - Scandentia. It has been suggested that the tree shrew can be used as an animal model for studying human diseases; however, the genomic sequence of the tree shrew is largely unidentified. In the present study, we reported the full-length cDNA sequence of the housekeeping gene, ß-actin, in the tree shrew. The amino acid sequence of ß-actin in the tree shrew was compared to that of humans and other species; a simple phylogenetic relationship was discovered. Quantitative polymerase chain reaction (qPCR) and western blot analysis further demonstrated that the expression profiles of ß-actin, as a general conservative housekeeping gene, in the tree shrew were similar to those in humans, although the expression levels varied among different types of tissue in the tree shrew. Our data provide evidence that the tree shrew has a close phylogenetic association with humans. These findings further enhance the potential that the tree shrew, as a species, may be used as an animal model for studying human disorders.

Mangiferin attenuates TH1/TH2 cytokine imbalance in an ovalbumin-induced asthmatic mouse model.

Mangiferin is a major bioactive ingredient in Mangifera indica Linn. (Anacardiaceae) leaves. Aqueous extract of such leaves have been used as an indigenous remedy for respiratory diseases like asthma and coughing in traditional Chinese medicine. However, underlying molecular mechanisms of mangiferin on anti-asthma remain unclear. In our present study, we investigated the anti-asthmatic effect of mangiferin on Th1/Th2 cytokine profiles and explored its underlying immunoregulatory mechanism in mouse model of allergic asthma. Mangiferin significantly reduced the total inflammatory cell counts and eosinophil infiltration, decreased the production of ovalbumin-specific IgE in serum and PGD2 in BALF. The antibody array analysis showed that mangiferin down-regulated the levels of one group of cytokines/chemokines including Th2-related IL-4, IL-5, IL-13, and others IL-3, IL-9, IL-17, RANTES, TNF-α, but simultaneously up-regulated Th1-related IFN-γ, IL-2 and IL-10 and IL-12 expression in serum. Thus it attenuates the imbalance of Th1/Th2 cells ratio by diminishing the abnormal mRNA levels of Th1 cytokines (IFN-γ and IL-12) and Th2 cytokines (IL-4, IL-5 and IL-13). Finally, mangiferin substantially inhibited the activation and expression of STAT-6 and GATA-3 in excised lung tissues. Our results suggest that mangiferin can exert anti-asthmatic effect. The underlying mechanism may attribute to the modulation of Th1/Th2 cytokine imbalance via inhibiting the STAT6 signaling pathway.

Identification of glyceraldehyde 3-phosphate dehydrogenase sequence and expression profiles in tree shrew (Tupaia belangeri).

The tree shrews (Tupaia belangeri) diverged from the primate order (Primates) and are classified as Scandentia, a separate taxonomic group of mammals. The tree shrew has been suggested to use an animal model to study human disease but the genomic sequences of tree shrew is largely unidentified. Here we identified the full-length cDNA sequence of a housekeeping gene, Glyceraldehyde 3-phosphate Dehydrogenase (GAPDH), in tree shrew. We further constructed a phylogenetic family tree base on GAPDH molecules of various organisms and compared GAPDH sequences with human and other small experimental animals. These study revealed that tree shrew was closer to human than mouse, rat, rabbit and guinea pig. The Quantitative Reverse Transcription PCR and western blot analysis further demonstrated that GAPDH expressed in various tissues in tree shrew as a general conservative housekeeping proteins as in human. Our findings provide the novel genetic knowledge of the tree shrew and strong evidences that tree shrew can be an experimental model system to study human disorders.

Observation of non-odd order harmonics by sub-2-cycle laser pulses.

High order harmonics generation from argon gas was comprehensively investigated by using multi-cycle and few-cycle laser pulses. Non-odd order harmonics were observed for sub-5-fs pulses, compare to the normal odd-order harmonics in the multi-cycle case. Theoretic analysis shows that the new spectral structure origins from the asymmetry of laser field in few-cycle pulses. This asymmetry induced both amplitude and phase difference between attosecond pulses from consecutive half-cycle of the laser field, which change the interference property of attosecond pulses and result in complex spectrum.

[In vitro effect of Huyinling extract on Trichomonas tenax].

The active ingredient of Huyinling, a combination of Chinese traditional medicine, was extracted by five different ethanol concentrations (40%-80%). There were seven groups named as five Huyinling ethanol extract groups (40%, 50%, 60%, 70%, and 80%), metronidazole group and blank control. Each Huyinling ethanol extract group was further divided into five subgroups with final concentration of 6.25, 12.5, 25, 50, and 100 mg/ml, respectively. Metronidazole group was given 10 microg/ml of the drug. Each group had 4 wells with 125 microl T. tenax(2 x 10(5)/ml). At 12 h, 24 h and 48 h after drug treatment, the anti-T. tenax effect of Huyinling ethanol extract was tested by microscope counting method. At 24 h the effect of Huyinling on T. tenax was examined with methyl thiazolyl tetrazolium (MTl) assay. The results showed that the higher concentration of Huyinling ethanol extract, the better effect on anti-T. tenax. 60% Huyinling ethanol extract group with concentrations of 6.25 mg/ml and 12.5 mg/ml showed higher anti-T. tenax effect than other groups (P < 0.01). The ethanol extract of Huyinling granules has a remarkable effect on T. tenax, and among the groups, 60% ethanol extract shows the best anti-T. tenax activity.

Delay control in attosecond pump-probe experiments.

The time delay between the pump and probe pulses in attosecond time-resolved experiments, such as attosecond streaking, is commonly introduced by splitting and recombining the two pulses in an interferometer. This technique suffers from instability in the optical path lengths of the two arms due to mechanical vibration of the optical elements and fluctuating environmental conditions. We present a technique with which the instability of the unconventional interferometer is suppressed while at the same time the time delay is controlled to within 20 as RMS using a feedback loop. Using this scheme, the streaked spectrogram of an attosecond pulse was measured.

Extreme ultraviolet supercontinua supporting pulse durations of less than one atomic unit of time.

Double optical gating of high-harmonic generation was used to obtain supercontinuous spectra in the extreme UV (XUV) region including the water window. The spectra supported a 16 as pulse duration that is below one atomic unit of time (24 as). The dependence of the gated spectra on the carrier-envelope phase of the laser provided evidence that isolated attosecond pulses were generated. In addition, to ensure the temporal coherence of the XUV light, the pulse shape and phase of isolated 107 as XUV pulses using a portion of the spectrum were characterized by attosecond streaking.

Carrier-envelope phase stabilization and control of 1 kHz, 6 mJ, 30 fs laser pulses from a Ti:sapphire regenerative amplifier.

Carrier-envelope (CE) phase stabilization of a two-stage chirped pulse amplifier laser system with regenerative amplification as the preamplifier is demonstrated. The CE phase stability of this laser system is found to have a 90 mrad rms error averaged over 50 laser shots for a locking period of 4.5 h. The CE phase locking was confirmed unambiguously by experimental observation of the 2pi periodicity of the high-order harmonic spectrum generated by double optical gating.

Temperature feedback control for long-term carrier-envelope phase locking.

We report a double feedback loop for the improvement of the carrier-envelope phase stabilization of a chirped mirror based femtosecond laser oscillator. By combining the control of the Ti:sapphire crystal temperature and the modulation of the pump power, the carrier envelope offset frequency, fCEO, was locked for close to 20 h, which is much longer than the typical phase stabilization time with only pump power modulation.