Sodium Polyoxotungstate Inhibits the Replication of Influenza Virus by Blocking the Nuclear Import of vRNP.

Both pandemic and seasonal influenza are major health concerns, causing significant mortality and morbidity. Current influenza drugs primarily target viral neuraminidase and RNA polymerase, which are prone to drug resistance. Polyoxometalates (POMs) are metal cation clusters bridged by oxide anions. They have exhibited potent anti-tumor, antiviral, and antibacterial effects. They have remarkable activity against various DNA and RNA viruses, including human immunodeficiency virus, herpes simplex virus, hepatitis B and C viruses, dengue virus, and influenza virus. In this study, we have identified sodium polyoxotungstate (POM-1) from an ion channel inhibitor library. In vitro, POM-1 has been demonstrated to have potent antiviral activity against H1N1, H3N2, and oseltamivir-resistant H1N1 strains. POM-1 can cause virion aggregation during adsorption, as well as endocytosis. However, the aggregation is reversible; it does not interfere with virus adsorption and endocytosis. Our results suggest that POM-1 exerts its antiviral activity by inhibiting the nuclear import of viral ribonucleoprotein (vRNP). This distinct mechanism of action, combined with its wide range of efficacy, positions POM-1 as a promising therapeutic candidate for influenza treatment and warrants further investigation.

[Reversal Effect of NVP-BEZ235 on Doxorubicin-Resistance in Burkitt Lymphoma RAJI Cell Line].

OBJECTIVE: To study the reversal effect of NVP-BEZ235 on doxorubicin resistance in Burkitt lymphoma RAJI cell line. METHODS: The doxorubicin-resistant cell line was induced by treating RAJI cells with a concentration gradient of doxorubicin. The levels of Pgp, p-AKT, and p-mTOR in cells were detected by Western blot. Cell viability was detected by MTT assay. IC50 was computed by SPSS. RESULTS: The doxorubicin-resistant Burkitt lymphoma cell line, RAJI/DOX, was established successfully. The expression of Pgp and the phosphorylation levels of AKT and mTOR in RAJI/DOX cell line were both higher than those in RAJI cell line. NVP-BEZ235 downregulated the phosphorylation levels of AKT and mTOR in RAJI/DOX cell line. NVP-BEZ235 inhibited the proliferation of RAJI/DOX cell line, and the effect was obvious when it was cooperated with doxorubicin. CONCLUSION: The constitutive activation of PI3K/AKT/mTOR pathway of RAJI/DOX cell line was more serious than RAJI cell line. NVP-BEZ235 reversed doxorubicin resistance of RAJI/DOX cell line by inhibiting the PI3K/AKT/mTOR signal pathway.

A panel of janus kinase inhibitors identified with anti-inflammatory effects protect mice from lethal influenza virus infection.

Influenza remains a significant threat to public health. In severe cases, excessive inflammation can lead to severe pneumonia or acute respiratory distress syndrome, contributing to patient morbidity and mortality. While antivirals can be effective if administered early, current anti-inflammatory drugs have limited success in treating severe cases. Therefore, discovering new anti-inflammatory agents to inhibit influenza-related inflammatory diseases is crucial. Herein, we screened a drug library with known targets using a human monocyte U937 infected with the influenza virus to identify novel anti-inflammatory agents. We also evaluated the anti-inflammatory effects of the hit compounds in an influenza mouse model. Our research revealed that JAK inhibitors exhibited a higher hit rate and more potent inhibition effect than inhibitors targeting other drug targets in vitro. Of the 22 JAK inhibitors tested, 15 exhibited robust anti-inflammatory activity against influenza virus infection in vitro. Subsequently, we evaluated the efficacy of 10 JAK inhibitors using an influenza mouse model and observed that seven provided protection ranging from 40% to 70% against lethal influenza virus infection. We selected oclacitinib as a representative compound for an extensive study to further investigate the in vivo therapeutic potential of JAK inhibitors for severe influenza-associated inflammation. Our results revealed that oclacitinib effectively suppressed neutrophil and macrophage infiltration, reduced pro-inflammatory cytokine production, and ultimately mitigated lung injury in mice infected with lethal influenza virus without impacting viral titer. These findings suggest that JAK inhibitors can modulate immune responses to influenza virus infection and may serve as potential treatments for influenza.IMPORTANCEAntivirals exhibit limited efficacy in treating severe influenza when not administered promptly during the infection. Current steroidal and nonsteroidal anti-inflammatory drugs demonstrate restricted effectiveness against severe influenza or are associated with significant side effects. Therefore, there is an urgent need for novel anti-inflammatory agents that possess high potency and minimal adverse reactions. In this study, 15 JAK inhibitors were identified through a screening process based on their anti-inflammatory activity against influenza virus infection in vitro. Remarkably, 7 of the 10 selected inhibitors exhibited protective effects against lethal influenza virus infection in mice, thereby highlighting the potential therapeutic value of JAK inhibitors for treating influenza.

BMS-265246, a Cyclin-Dependent Kinase Inhibitor, Inhibits the Infection of Herpes Simplex Virus Type 1.

Herpes simplex virus type 1 (HSV-1) infections are prevalent illnesses that can cause mucocutaneous ulcerative disease, keratitis, and genital herpes. In patients with compromised immune systems, the infection can lead to serious problems, such as encephalitis. Additionally, neonatal infections can cause brain problems and even death. Current first-line antiviral drugs are nucleoside analog inhibitors that target viral polymerase, and resistant strains have emerged. As a result, new drugs with distinct action modes are needed. Recent research indicates that cyclin-dependent kinases (CDKs) are prospective antiviral targets. Thus, CDK inhibitors may be effective antiviral agents against HSV-1 infection. In this study, we examined a panel of CDK inhibitors that target CDKs in the present study. BMS-265246 (BMS), a CDK 1/2 inhibitor, was found to effectively limit HSV-1 multiplication in Vero, HepG2, and Hela cells. A mechanism of action study suggested that BMS inhibits the early stages of viral replication when added early in the viral infection. The suppression of multiple steps in viral replication by BMS was revealed when HSV-1 infected cells were treated at different time periods in the viral life cycle. Our results suggest that BMS is a potent anti-HSV-1 agent and unique in that it may interfere with multiple steps in HSV-1 replication.

Modified Cap-Assisted Endoscopic Mucosal Resection Versus Endoscopic Submucosal Dissection for the Treatment of Rectal Neuroendocrine Tumors ≤10 mm: A Randomized Noninferiority Trial.

INTRODUCTION: Although recent guidelines recommend endoscopic resection of rectal neuroendocrine tumors (NET) ≤10 mm, there is no consensus on which endoscopic modality should be performed. We aimed to compare the safety and efficacy of modified cap-assisted endoscopic mucosal resection (mEMR-C) and endoscopic submucosal dissection (ESD) methods for the treatment of rectal NET ≤10 mm. METHODS: A randomized noninferiority trial comparing mEMR-C and ESD was conducted. The primary outcome was the histological complete resection rate; the secondary outcomes included en bloc resection rate, operation time, complications, and so on. Subgroup analyses and follow-up were also performed. RESULTS: Ninety patients were enrolled, and 79 patients with pathologically confirmed rectal NET were finally analyzed, including 38 cases of mEMR-C and 41 cases of ESD. Histological complete resection rate was 97.4% in the mEMR-C group and 92.7% in the ESD group. The noninferiority of mEMR-C compared with that of ESD was confirmed because the absolute difference was 4.7% (2-sided 90% confidence interval, -3.3% to 12.2%; P = 0.616). En bloc resection and successful removal of rectal NET were achieved in all patients. Advantages of mEMR-C over ESD included shorter operation time (8.89 ± 4.58 vs 24.8 ± 9.14 minutes, P < 0.05) and lower hospitalization cost ($2,233.76 ± $717.70 vs $2,987.27 ± $871.81, P < 0.05). Postoperative complications were recorded in 4 patients who received mEMR-C and 2 patients in the ESD group (11.5% vs 4.9%, P = 0.509), which were all well managed using endoscopy. Similar findings were observed when subgroup analysis was performed. DISCUSSION: mEMR-C is noninferior to ESD with a similar complete resection rate. In addition, mEMR-C had shorter procedure duration time and lower hospitalization costs. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03982264.

Identification and Validation of a Novel Prognostic Gene Model for Colorectal Cancer.

Aims: Colon cancer (CRC), with high morbidity and mortality, is a common and highly malignant cancer, which always has a bad prognosis. So it is urgent to employ a reasonable manner to assess the prognosis of patients. We developed and validated a gene model for predicting CRC risk. Methods: The Gene Expression Omnibus (GEO) database was used to extract the gene expression profiles of CRC patients (N = 181) from GEO to identify genes that were differentially expressed between CRC patients and controls and then stable signature genes by firstly using both robust likelihood-based modeling with 1000 iterations and random survival forest variable hunting algorithms. Cluster analysis using the longest distance method was drawn out, and Kaplan-Meier (KM) survival analysis was used to compare the clusters. Meanwhile, the risk score was evaluated in three independent datasets including the GEO and Illumina HiSeq sequencing platforms. The corresponding risk index was calculated, and samples were clustered into high- and low-risk groups according to the median. And survival ROC analysis was used to evaluate the prognostic model. Finally, the Gene Set Enrichment Analysis (GSEA) was performed for further functional enrichment analyses. Results: A 10-gene model was obtained, including 7 negative impact factors (SLC39A14, AACS, ERP29, LAMP3, TMEM106C, TMED2, and SLC25A3) and 3 positive ones (CNPY2, GRB10, and PBK), which related with several important oncogenic pathways (KRAS signaling, TNF-α signaling pathway, and WNT signaling pathway) and several cancer-related cellular processes (epithelial mesenchymal transition and cellular apoptosis). By using colon cancer datasets from The Cancer Genome Atlas (TCGA), the model was validated in KM survival analysis (P ≤ 0.001) and significant analysis with recurrence time (P = 0.0018). Conclusions: This study firstly developed a stable and effective 10-gene model by using novel combined methods, and CRC patients might be able to use it as a prognostic marker for predicting their survival and monitoring their long-term treatment.

Aromatic profiles and enantiomeric distributions of chiral odorants in baked green teas with different picking tenderness.

Suitable picking tenderness is an essential prerequisite for manufacturing tea. However, the influence of picking tenderness of fresh tea leaves on the aromatic components is still unclear. In this study, aromatic profiles and chiral odorants in fresh tea leaves and corresponding baked green teas with five levels of tenderness of two representative cultivars were analysed using stir bar sorptive extraction-gas chromatography-mass spectrometry. cis-Linalool oxide (furanoid) and methyl salicylate exhibited significantly increasing trends as samples of all series matured. The content of most chiral odorants was significantly high in the mature samples, and significant content variations of all enantiomers during baked green tea processing could be observed with different trends according to their precursors. In particular, the enantiomeric ratios of most chiral odorants were less influenced by the picking tenderness and processing, while drying (limonene), spreading and fixation (α-terpineol), and spreading (dihydroactinidiolide) influenced the chiral distribution of the aforementioned odorants.

Effect of Emulsion Particle Size on the Encapsulation Behavior and Oxidative Stability of Spray Microencapsulated Sweet Orange Oil (Citrus aurantium var. dulcis).

Three different feed emulsions of different particle sizes were mixed with a modified starch and maltodextrin and spray dried to make a large (LP), small (SP), and nano-size encapsulated powder (NP), respectively. Emulsion size, oil content, loading capacity (LC), encapsulation efficiency (EE), water content, aw, scanning electron microscopy (SEM), glass transition temperature (Tg), as well as d-limonene release characteristic and limonene oxide formation rate during 37 °C and various aw storage were determined. With the increase of the feed emulsion size, the reconstituted emulsion size of the LP tended to increase and change to a bimodal distribution. The surface oil content increased with the increasing size of the reconstituted emulsion, and the opposite was true for EE. The smaller the reconstituted emulsion size, the higher Tg during a low aw condition. The Tg of the LP, SP and NP were 62, 88, and 100 °C, respectively, and NP > SP > LP. The release and the oxidative rate of d-limonene was the lowest for the NP and then increased for the SP and LP. The release and oxidative rates increased with the elevation of aw and peaked at 0.33. The powder surface morphological structure was intact, the spray-dried powder was more stable, and microstructure changed from a glass state to a rubbery state during storage.

Assessment of the contribution of chiral odorants to aroma property of baked green teas using an efficient sequential stir bar sorptive extraction approach.

Chiral volatile compounds are known to be distributed in teas at various enantiomeric ratios. However, the performance of each enantiomer, including aroma characteristics, aroma intensities, and contribution to the overall flavor of tea, is still unclear. In this study, aroma characteristics and intensities of 38 volatile enantiomers in standards and baked green teas with chestnut-like aroma and clean aroma were evaluated by an efficient sequential headspace-stir bar sorptive extraction (seq-HS-SBSE) approach combined with the enantioselective gas chromatography-olfactometry/mass spectrometry (Es-GC-O/MS) technique. Moreover, aroma recombination results for the two types of baked green teas using 14 chiral odorants and four achiral odorants indicated that the combinations of the detected odorants mainly contributed to the "floral", "sweet", and "chestnut-like" aromas. R-Linalool simultaneously enhanced the "floral", "sweet", and "chestnut-like" aromas; R-limonene mainly contributed to the "sweet" and "clean" aromas; and S-α-terpineol promoted the "sweet" and "floral" aromas of baked green tea.

Blue laser imaging combined with JNET (Japan NBI Expert Team) classification for pathological prediction of colorectal laterally spreading tumors.

BACKGROUND: Blue laser imaging (BLI) can provide useful information on colorectal laterally spreading tumors (LSTs) by visualizing the surface and vessel patterns in detail. The present research aimed to evaluate the diagnostic performance of BLI-combined JNET (Japan NBI Expert Team) classification for identifying LSTs. METHODS: This retrospective, multicenter study included 172 LSTs consisted of 6 hyperplastic polyps/sessile serrated polyps, 94 low-grade dysplasias (LGD), 60 high-grade dysplasias (HGD), 6 superficial submucosal invasive (m-SMs) carcinomas, and 4 deep submucosal invasive carcinomas. The relationship between the JNET classification and the histologic findings of these lesions were then analyzed. RESULTS: For all LSTs, non-experts and experts had a 79.7% and 90.7% accuracy for Type 2A (P = 0.004), a sensitivity of 94.7% and 96.8% (P = 0.718), and a specificity of 61.5% and 83.3% (P = 0.002) for prediction of LGD, respectively. The results also demonstrated 80.8% and 91.3% accuracy for Type 2B (P = 0.005), a sensitivity of 65.2% and 83.3% (P = 0.017), and a specificity of 90.6% and 96.2% (P = 0.097) for predicting HGD or m-SMs. For LST-granular (LST-G) lesions, Type 2A in experts had higher specificity (65.6% vs. 83.6%, P = 0.022) and accuracy (81.8% vs. 91.2%, P = 0.022). Type 2B in experts only had higher accuracy (82.5% vs. 92.0%, P = 0.019). However, no significant differences were noted for any comparisons between non-experts and experts for LST-non-granular (LST-NG) lesions. CONCLUSIONS: BLI combined with JNET classification was an effective method for the precise prediction of pathological diagnosis in patients with LSTs. Diagnostic performance of JNET classification by experts was better than that by non-experts for all examined LST or LST-G lesions when delineating between Type 2A and 2B, but there was no difference for the identification of LST-NG lesions by these two groups.

Eradication of Posterior Capsule Opacification: Documentation of a Marked Decrease in Nd:YAG Laser Posterior Capsulotomy Rates Noted in an Analysis of 5416 Pseudophakic Human Eyes Obtained Postmortem.

OBJECTIVE: (1) To report the neodymium:yttrium-aluminum-garnet (Nd:YAG) laser posterior capsulotomy rate (%) of eight rigid and foldable intraocular lens (IOL) designs in a series of 5416 pseudophakic human eyes obtained postmortem, accessioned in our center between January 1988 and January 2000. (2) To identify factors that are instrumental in reducing the incidence of posterior capsule opacification, (PCO, secondary cataract) and hence the need for Nd:YAG laser posterior capsulotomy. DESIGN: Comparative autopsy tissue analysis. PARTICIPANTS: A total of 5416 globes with posterior chamber intraocular lens (PC-IOLs) obtained postmortem received from Lions Eye Banks between 1988 and 2000. METHODS: Miyake-Apple posterior photographic technique. Special reference was given to the presence or absence of Nd:YAG laser posterior capsulotomy orifice on the posterior capsule of each eye. MAIN OUTCOME MEASURES: The Nd:YAG laser posterior capsulotomy rate (%) as of January 2000 was documented. In addition, the Nd:YAG laser posterior capsulotomy rate for each lens was plotted on a monthly basis for the same period, creating a computerized trend or "timeline" for each IOL style. RESULTS: Relatively high Nd:YAG laser posterior capsulotomy rates ranging from 20.3% to 33.4% were noted with four relatively older designs (high incidence of implantation between 1988 and the early 1990s). Four modern foldable lOLs manufactured from silicone and acrylic materials had significantly lower Nd:YAG laser posterior capsulotomy rates ranging from 0.9% (Alcon Acrysof) to 17.1%. The difference in Nd:YAG rates among the eight IOL designs was found to be significant (P < 0.0001, chi-square test). Comparing foldable versus rigid designs, the foldable IOLs were associated with a much lower Nd:YAG laser posterior capsulotomy rate (14.1% vs. 31.1%). CONCLUSIONS: By use of the six factors regarding surgical technique and IOL choice described in this article, we strongly believe that the overall incidence of PCO and hence the incidence of Nd:YAG laser posterior capsulotomy is now rapidly decreasing from rates as high as 50% in the 1980s to early 1990s. Surgical tools and IOLs are now available to bring these rates down to single digits. Careful application and use of these tools by surgeons can genuinely lead in the direction of virtual eradication of secondary cataract, the second most common cause of visual loss worldwide.

Determination of Kaurenoic Acid in Acanthopanax trifoliatus by Ultra-High Performance Liquid Chromatography Coupled with Tandem Mass Spectrometry (UHPLC-MS/MS).

Acanthopanax trifoliatus (L.) Merr. (A. trifoliatus) belongs to the family Araliaceae, which is called "Le Cai", and is an indigenous plant to Guangdong Province that has been prevalently planted for years. A. trifoliatus is used in folk medicine and has ginseng-like activity. Kaurenoic acid ((-)-kaur-16-en-19-oic acid, KA) is a kaurane-type diterpenoid that is regarded as a major compound in A. trifoliatus. Early studies have reported the determination of KA by HPLC capillary electrophoresis. However, KA could not be completely separated from other components in the plant extract by HPLC because of their similar molecular structures and physical and chemical properties. UHPLC-MS/MS could be a useful tool to identify and quantify KA. In the present work, a UHPLC-ESI-MS/MS method for determining KA in A. trifoliatus was developed and validated. KA was extracted from lyophilized A. trifoliatus leaves by ultrasound-assisted extraction and further purified by solid phase extraction (SPE). KA was quantified and separated on an Accucore C18 LC column. Mass spectrometry with multi-reaction monitoring (MRM) and quantitative fragment ion/product ion (m/z: 301.3/301.3) in ESI negative mode was used for quantification. The intra-assay and inter-assay relative standard deviation (R.S.D.) were 2.8% and 3.2%, respectively. The inter-person R.S.D. on the same day was 3.6%. The inter-instrument R.S.D. with the same model on the same day was 2.9%. The recoveries evaluated upon spiking three different concentrations of KA were above 97%. A minor matrix effect of 94% was observed. This method has been applied successfully for the determination of KA in A. trifoliatus leaves.

The protective and pathogenic roles of IL-17 in viral infections: friend or foe?

Viral infections cause substantial human morbidity and mortality, and are a significant health burden worldwide. Following a viral infection, the host may initiate complex antiviral immune responses to antagonize viral invasion and replication. However, proinflammatory antiviral immune responses pose a great threat to the host if not properly held in check. Interleukin (IL)-17 is a pleiotropic cytokine participating in a variety of physiological and pathophysiological conditions, including tissue integrity maintenance, cancer progression, autoimmune disease development and, more intriguingly, infectious diseases. Abundant evidence suggests that while IL-17 plays a crucial role in enhancing effective antiviral immune responses, it may also promote and exacerbate virus-induced illnesses. Accumulated experimental and clinical evidence has broadened our understanding of the seemingly paradoxical role of IL-17 in viral infections and suggests that IL-17-targeted immunotherapy may be a promising therapeutic option. Herein, we summarize current knowledge regarding the protective and pathogenic roles of IL-17 in viral infections, with emphasis on underlying mechanisms. The various and critical roles of IL-17 in viral infections necessitate the development of therapeutic strategies that are uniquely tailored to both the infectious agent and the infection environment.

Identification and quantification of key odorants in the world's four most famous black teas.

Keemun, Assam, Darjeeling and Ceylon black teas are honored as the world's four most famous black teas, and their excellent aroma qualities are well received by people around the world. In this study, aroma components in these four types of teas were analyzed by comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC × GC-TOFMS) and gas chromatography-olfactometry (GC-O) technologies. A total of 42 aroma-active compounds were ultimately identified, especially benzeneacetaldehyde, geraniol, (Z)-3-hexen-1-yl hexanoate, trans-ß-ionone, cis-linalool oxide (pyranoid), hotrienol, and methyl salicylate presented the strongest aroma strengths with pleasant scents in all tested teas. The quantification results indicated that 19 compounds including (Z)-3-hexenol, 1-octen-3-ol, linalool, phenylethyl alcohol, hexanal, benzeneacetaldehyde, limonene, heptanoic acid, (Z)-3-hexen-1-ol, acetate, benzyl alcohol, trans-linalool oxide (furanoid), hotrienol, 1-octen-3-one, 2-nonanone, (E)-2-octenal, nonanal, ß-myrcene, 2-pentylfuran, and methylpyrazine were identified as the key compounds with odor activity values (OAVs) higher than 1.0 in the world's four most famous black teas. Notably, the comparison of GC-O and OAV calculation results showed that methyl salicylate (Ceylon), (E)-2-octenal (Assam), benzeneacetaldehyde (Keemun) and linalool and trans-linalool oxide (furanoid) (Darjeeling) might be the most definitive odorants in the corresponding tea categories.

Arenobufagin induces MCF-7 cell apoptosis by promoting JNK-mediated multisite phosphorylation of Yes-associated protein.

BACKGROUND: It has been demonstrated that bufadienolides exert potent anti-cancer activity in various tumor types. However, the mechanisms that underlie their anti-cancer properties remain unclear. Yes-associated protein, a key effector of Hippo signaling, functions as a transcription coactivator, plays oncogenic and tumor suppressor roles under different conditions. Here, we report that arenobufagin (ABF), a representative bufadienolide, induced breast cancer MCF-7 cells to undergo apoptosis, which occurred through the JNK-mediated multisite phosphorylation of YAP. METHODS: Cytotoxicity was examined using an MTT assay. ABF-induced apoptosis was measured with a TUNEL assay and Annexin V-FITC/PI double staining assay. Western blotting, immunofluorescence, qRT-PCR and coimmunoprecipitation were employed to assess the expression levels of the indicated molecules. Lose-of-function experiments were carried out with siRNA transfection and pharmacological inhibitors. ABF-induced phosphopeptides were enriched with Ti4+-IMAC chromatography and further subjected to reverse-phase nano-LC-MS/MS analysis. RESULTS: ABF significantly reduced the viability of MCF-7 cells and increased the percentage of early and late apoptotic cells in a concentration- and time-dependent manner. Following ABF treatment, YAP accumulated in the nucleus and bound to p73, which enhanced the transcription of the pro-apoptotic genes Bax and p53AIP1. YAP knock-down significantly attenuated ABF-induced apoptotic cell death. Importantly, we found that the mobility shift of YAP was derived from its phosphorylation at multiple sites, including Tyr357. Moreover, mass spectrometry analysis identified 19 potential phosphorylation sites in YAP, with a distribution of 14 phosphoserine and 5 phosphothreonine residues. Furthermore, we found that the JNK inhibitor SP600125 completely diminished the mobility shift of YAP and its phosphorylation at Tyr357, the binding of YAP and p73, the transcription of Bax and p53AIP1 as well as the apoptosis induced by ABF. These data indicate that ABF induced YAP multisite phosphorylation, which was associated with p73 binding, and that apoptosis was mediated by the JNK signaling pathway. CONCLUSIONS: Our data demonstrate that ABF suppresses MCF-7 breast cancer proliferation by triggering the pro-apoptotic activity of YAP, which is mediated by JNK signaling-induced YAP multisite phosphorylation as well as its association with p73. The present work not only provides additional information on the use of ABF as an anti-breast cancer drug, but also offers evidence that the induction of the tumor suppressor role of YAP may be a therapeutic strategy.

Hybrids of arenobufagin and benzoisoselenazol reducing the cardiotoxicity of arenobufagin.

Arenobufagin is a naturally occurring bufadienolide showing promising antitumor activity accompanied however with apparent cardiac toxicity. Following the recent discovery that oxidative damage possibly be an important cause of the cardiac toxicity of cardenolides, a strategy fusing the antitumor agent arenobufagin with a benzoisoselenazol fragment, a reactive oxygen species (ROS) scavenger, has been developed. Six novel hybrids were synthesized and their ROS scavenging activities as well as their in vitro cytotoxicity against the human hepatocellular carcinoma cell line HepG2, an adriamycin-resistant subline HepG2/ADM, and the human myocardial cell line AC16 were evaluated. The results indicate that the hybrids exhibit various degrees of in vitro ROS scavenging activities, and weaker cytotoxicity than that of arenobufagin against the myocardial cell line AC16. These findings suggest the feasibility of a strategy in which the cardiotoxicity of the potential antitumor agent arenobufagin is reduced.

The enantiomeric distributions of volatile constituents in different tea cultivars.

Although the enantiomeric distribution of chiral volatiles presents great potential in discrimination of tea cultivars and their geographic origins, this area has received little attention. Thus, we herein aimed to determine the relationships between tea cultivars and the enantiomeric distributions of their chiral volatile constituents. Headspace solid-phase microextraction (HS-SPME) and enantioselective gas chromatography-mass spectrometry (Es-GC-MS) were employed to quantify 15 volatile components in 22 tea cultivars from different locations within China. The tea cultivars were successfully differentiated by their geographical origins, and the concentrations of R-linalool, S-citronellol, S-E-nerolidol, (1R, 2R)-methyl jasmonate, S-α-ionone, and the two enantiomers of linalool oxide A differed significantly among the different groups. It should also be noted that tea processing methods greatly influenced the formation of volatile enantiomers. Our results demonstrated that the enantiomeric distribution of volatile constituents closely correlates with the geographical origins, leaf types, and manufacturing suitabilities of the tea cultivars examined herein.

Identification of key odorants responsible for chestnut-like aroma quality of green teas.

A chestnut-like aroma is widely considered an important indicator of an excellent-quality green tea; however, the key odorants responsible for chestnut-like aroma have never been systematically studied and remain unknown. In this study, the aroma components of green teas and Chinese chestnuts were analyzed using comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC × GC-TOFMS), and 58 compounds were identified as common aroma components among green teas, boiled Chinese chestnuts, roasted Chinese chestnuts and raw Chinese chestnuts. Subsequently, 17 volatiles, including 3-methylbutanal, (E)-3-penten-2-one, ethylbenzene, heptanal, benzaldehyde, 2-pentylfuran, octanal, benzeneacetaldehyde, (E)-2-octenal, (E,E)-3,5-octadien-2-one, linalool, nonanal, (E)-2-nonenal, decanal, (Z)-hex-3-en-1-yl hexanoate, trans-ß-ionone and (E)-nerolidol, were identified as the key odorants responsible for chestnut-like aroma based on the odor activity value (OAV) calculation method. Besides, the comparison of OAVs of key odorants between fresh tea leaves and finished teas indicated that all key odorants were present in fresh tea leaves and that their contents increased or decreased during tea processing. Moreover, the comparison between results of OAV and gas chromatography-olfactometry (GC-O) methods showed that ethylbenzene, heptanal, benzaldehyde, 2-pentylfuran, (E,E)-3,5-octadien-2-one, linalool, (Z)-hex-3-en-1-yl hexanoate and trans-ß-ionone were the common identified compounds between the two methods. The identification of chestnut-like aroma in green teas will provide a theoretical basis for further research on the directional adjustment and control of tea aroma quality.

Enantiomeric and quantitative analysis of volatile terpenoids in different teas (Camellia sinensis).

Volatile terpenoids play important roles in the formation of tea aroma quality due to their pleasant scents and low odor thresholds. Most volatile terpenoids contain stereogenic centers, which results in various stereo distributions of their enantiomers and diastereoisomers in different types of tea. However, the distribution characteristics of terpenoid enantiomers in teas were still unclear, which poses an obstacle to the scientific understanding of tea aroma. In this work, a new and efficient analysis approach based on headspace solid phase microextraction (HS-SPME)-chiral gas chromatography-mass spectrometry (GC-MS) was established to analyze 12 pairs of familiar terpenoid enantiomers in different teas. The extraction efficiency of the HS-SPME method to extract volatile terpenoids in teas was the greatest when using CAR-DVB-PDMS (50/30µm) fibers and 1:10 proportions between tea and boiling water at a 50°C extraction temperature for 40min, and the stability observation of enantiomeric ratios of the terpenoids well proved the feasibility of the extraction method. The favorable limits of detection, limits of quantitation, repeatability, linearity, and concentration ranges of each terpenoid enantiomer demonstrated the repeatability and reliability of the analytical approach. The enantiomeric and quantitative analyses indicated that S-limonene, S-linalool, (2S, 5S)-linalool oxide A, (2S, 5R)-linalool oxide B, R-4-terpineol, (2S, 5R)-linalool oxide C, (2S, 5S)-linalool oxide D, S-α-terpineol, R-α-ionone, peak 1 of theaspirane A and peak 2 of theaspirane B were the major terpenoid components in most Chinese teas; instead, higher proportions of the opposite enantiomers of the above terpenoids were frequently detected in black teas with large leaf origin and Indonesia white teas. Besides, great diversities of enantiomeric ratios and concentrations among different teas were observed. Furthermore, partial least-squares discriminant analyses were performed to distinguish the concentration differences of the terpenoid enantiomers among different teas; the analysis results indicated that highly significant concentration differences existed between large and small leaf origins of black teas, and significant differences of the concentrations of linalool oxides A-C were observed between green, white and dark teas. The successful application of this chiral analysis technique of tea aroma will lay a scientific foundation for further quality assessment, botanical origin determination and authenticity assessment of teas.

Arenobufagin intercalates with DNA leading to G2 cell cycle arrest via ATM/ATR pathway.

Arenobufagin, a representative bufadienolide, is the major active component in the traditional Chinese medicine Chan'su. It possesses significant antineoplastic activity in vitro. Although bufadienolide has been found to disrupt the cell cycle, the underlying mechanisms of this disruption are not defined. Here, we reported that arenobufagin blocked the transition from G2 to M phase of cell cycle through inhibiting the activation of CDK1-Cyclin B1 complex; The tumor suppressor p53 contributed to sustaining arrest at the G2 phase of the cell cycle in hepatocellular carcinoma (HCC) cells. Moreover, arenobufagin caused double-strand DNA breaks (DSBs) and triggered the DNA damage response (DDR), partly via the ATM/ATR-Chk1/Chk2-Cdc25C signaling pathway. Importantly, we used a synthetic biotinylated arenobufagin-conjugated chemical probe in live cells to show that arenobufagin accumulated mainly in the nucleus. The microscopic thermodynamic parameters measured using isothermal titration calorimetry (ITC) also demonstrated that arenobufagin directly bound to DNA in vitro. The hypochromicity in the UV-visible absorption spectrum, the significant changes in the circular dichroism (CD) spectrum of DNA, and the distinct quenching in the fluorescence intensity of the ethidium bromide (EB)-DNA system before and after arenobufagin treatment indicated that arenobufagin bound to DNA in vitro by intercalation. Molecular modeling suggested arenobufagin intercalated with DNA via hydrogen bonds between arenobufagin and GT base pairs. Collectively, these data provide novel insights into arenobufagin-induced cell cycle disruption that are valuable for the further discussion and investigation of the use of arenobufagin in clinical anticancer chemotherapy.