Ultra-Fast Single-Nucleotide-Variation Detection Enabled by Argonaute-Mediated Transistor Platform.

"Test-and-go" single-nucleotide variation (SNV) detection within several minutes remains challenging, especially in low-abundance samples, since existing methods face a trade-off between sensitivity and testing speed. Sensitive detection usually relies on complex and time-consuming nucleic acid amplification or sequencing. Here, a graphene field-effect transistor (GFET) platform mediated by Argonaute protein that enables rapid, sensitive, and specific SNV detection is developed. The Argonaute protein provides a nanoscale binding channel to preorganize the DNA probe, accelerating target binding and rapidly recognizing SNVs with single-nucleotide resolution in unamplified tumor-associated microRNA, circulating tumor DNA, virus RNA, and reverse transcribed cDNA when a mismatch occurs in the seed region. An integrated microchip simultaneously detects multiple SNVs in agreement with sequencing results within 5 min, achieving the fastest SNV detection in a "test-and-go" manner without the requirement of nucleic acid extraction, reverse transcription, and amplification.

Phenotypic and genetic characterization of hypervirulent Klebsiella pneumoniae in patients with liver abscess and ventilator-associated pneumonia.

Ventilator-associated pneumonia (VAP) and pyogenic liver abscess (PLA) due to Klebsiella pneumoniae infection can trigger life-threatening malignant consequences, however, there are few studies on the strain-associated clinical pathogenic mechanisms between VAP and PLA. A total of 266 patients consist of 129 VAP and 137 PLA were included for analysis in this study. We conducted a comprehensive survey for the two groups of K. pneumoniae isolates, including phenotypic experiments, clinical epidemiology, genomic analysis, and instrumental analysis, i.e., to obtain the genomic differential profile of K. pneumoniae strains responsible for two distinct infection outcomes. We found that PLA group had a propensity for specific underlying diseases, especially diabetes and cholelithiasis. The resistance level of VAP was significantly higher than that of PLA (78.57% vs. 36%, P < 0.001), while the virulence results were opposite. There were also some differences in key signaling pathways of biochemical processes between the two groups. The combination of iucA, rmpA, hypermucoviscous phenotype, and ST23 presented in K. pneumoniae infection is more important and highly prudent for timely treatment. The present study may contribute a benchmark for the K. pneumoniae clinical screening, epidemiological surveillance, and effective therapeutic strategies.

Modulating Inflammation-Mediated Diseases via Natural Phenolic Compounds Loaded in Nanocarrier Systems.

The global increase and prevalence of inflammatory-mediated diseases have been a great menace to human welfare. Several works have demonstrated the anti-inflammatory potentials of natural polyphenolic compounds, including flavonoid derivatives (EGCG, rutin, apigenin, naringenin) and phenolic acids (GA, CA, etc.), among others (resveratrol, curcumin, etc.). In order to improve the stability and bioavailability of these natural polyphenolic compounds, their recent loading applications in both organic (liposomes, micelles, dendrimers, etc.) and inorganic (mesoporous silica, heavy metals, etc.) nanocarrier technologies are being employed. A great number of studies have highlighted that, apart from improving their stability and bioavailability, nanocarrier systems also enhance their target delivery, while reducing drug toxicity and adverse effects. This review article, therefore, covers the recent advances in the drug delivery of anti-inflammatory agents loaded with natural polyphenolics by the application of both organic and inorganic nanocarriers. Even though nanocarrier technology offers a variety of possible anti-inflammatory advantages to naturally occurring polyphenols, the complexes' inherent properties and mechanisms of action have not yet been fully investigated. Thus, expanding the quest on novel natural polyphenolic-loaded delivery systems, together with the optimization of complexes' activity toward inflammation, will be a new direction of future efforts.

Exploring multifunctional components from Andrographis paniculata by affinity ultrafiltration with three molecular targets.

Andrographis paniculata (Burm. F.) Nees (AP) was a typical plant resource that has the concomitant function of both foodstuff and medicine, while the action mechanisms of its immune regulation, anti-inflammatory and anti-viral effects and the specific components remain unclear. In this work, a screening approach combining bio-affinity ultrafiltration with liquid chromatography mass spectrometry (UF-LC/MS) was hired to screen potential bioactive compounds from AP. The crude extract of AP exerted COX-2 and ACE2 inhibitory effects by other bioassays. Meanwhile, a total of eleven ligands targeting COX-2, IL-6 and ACE2 were screened out. Thereinto, two compounds including andrographolide and 14-deoxy-11,12-didehydroandrographolide exhibited strong binding affinities to COX-2, IL-6 and ACE2 by UF-LC/MS and molecular docking analysis. This is the first report to apply UF-LC/MS approach to rapidly screen out multi-target ligands from AP, and further decipher corresponding mechanisms, which could be beneficial to expedite the search for new multi-target bioactive compounds in other natural products or foods.

Charge-Dependent Signal Changes for Label-Free Electrochemiluminescence Immunoassays.

Label-free electrochemiluminescence (ECL) immunoassays (lf-ECLIA), based on biomarker-induced ECL signal changes, have attracted increasing attention due to the simple, rapid, and low-cost detection of biomarkers without secondary antibodies and complicated labeling procedures. However, the interaction rule and mechanism between analytical interfaces and biomarkers have rarely been explored. Herein, the interactions between biomarkers and analytical interfaces constructed by assembly of a nanoluminophore and antibody-functionalized gold nanoparticles on an indium tin oxide electrode were studied. The nanoluminophore was synthesized by mixing Cu2+/l-cysteine chelate and N-(4-Aminobutyl)-N-ethylisoluminol-bifunctionalized gold nanoparticles with chitosan. It was found that positively charged biomarkers increased the ECL intensity, whereas negatively charged biomarkers decreased the ECL intensity. The assembly pH influenced the biomarker charges, which determined the ECL enhancement or inhibition. The detection pH only affected the ECL intensity but not the ECL changing trends. Based on the ECL signal changes, a charge-dependent lf-ECLIA was established, which exhibited inhibition responses to negatively charged human immunoglobulin G and copeptin and enhancement responses to positively charged cardiac troponin I, heart-type fatty acid binding protein, brain natriuretic peptide, and SARS-CoV-2 N protein. The linear range was 0.1-1000 pg/mL, and the detection limits were distributed in 0.024-0.091 pg/mL. Besides, a mechanism of the charge-dependent ECL enhancement and inhibition effects is proposed, which is very important for the development of new lf-ECLIA methodologies.

Flavonoids from Selaginella doederleinii Hieron and Their Antioxidant and Antiproliferative Activities.

Selaginella doederleinii Hieron. (S. doederleinii) is a traditional herb that is widely used in China to treat several ailments, but mainly cancer. Studies have been carried out to determine the phytochemicals ascribed to its pharmacological activity. However, both phytochemical and pharmacological profiles have not been fully explored as few compounds have been reported. This study evaluated the flavonoid content of the ethanol extract and its four fractions (petroleum ether, dichloromethane, ethyl acetate, and n-butanol) together with their antioxidant activity (DPPH and FRAP assays). Further, the antiproliferative activity was evaluated. Two new secondary metabolites (1 and 3) were isolated from S. doederleinii, which comprised of an apigenin skeleton with a phenyl attached at C-8 of ring A and an acetyl group. Additionally, other known metabolites 2 and 4-16 were isolated, whereby compounds 2, 4, 5, 8, 12, 15, and 16 were reported for the first time in this species. These compounds were evaluated for their antioxidative potentials by both DPPH and FRAP assays, and for their antiproliferative activities by the MTT assay on three human cancer cell lines: colon cancer (HT-29), cervical cancer (HeLa), and lung cancer (A549). Compound 7 exhibited the best activity on the three cancer cell lines (HT-29, HeLa, A549) by inhibiting the rate of growth of the cancer cells in a dose-dependent manner with IC50 values of 27.97, 35.47, and 20.71 µM, respectively. The structure-activity relationship of the pure compounds was highlighted in this study. Hence, the study enriched both the phytochemical and pharmacological profiles of S. doederleinii.

Inhibitors Targeting Multiple Janus Kinases From Zanthoxylum simulans Mediate Inhibition and Apoptosis Against Gastric Cancer Cells via the Estrogen Pathway.

Janus kinases (JAKs) play a key role in subtly regulating proliferation, apoptosis, and differentiation of cancer cells, and their inhibitors are actively sought as new drug leads. By developing JAKs based affinity ultrafiltration method coupled with LC/Q-TOF-MS in order to discover selective JAKs inhibitors from total quaternary alkaloids (QAs) from Zanthoxylum simulans, peak 19 (Berberine) and peak 21 (Chelerythrine) were revealed to exhibit notable selectivity on JAK1, JAK2, and JAK3 over Tyk2. In addition, Chelerythrine showed stronger inhibitory activity than the positive control (Cerdulatinib) on gastric cancer cells (AGS), while Berberine, with weaker inhibition. Chelerythrine and Berberine also showed obvious inhibition on human hepatocyte cells (LO2). Furthermore, molecular docking analysis revealed their discrepancies due to different interaction bonds and characteristic residues. Quaternary N was proposed as the functional group to enhance the selectivity of JAK1, and some specific moieties towards Asp1021, Leu855, and Leu828 were suggested to increase the selectivity for JAK1, JAK2, and JAK3, respectively. As the most potential inhibitor of JAKs from QAs, Chelerythrine exhibited distinct suppression of adhesion, migration, invasion, and stimulating apoptosis of AGS cells, which was consistent with the significant down-regulation of estrogen receptors (ER-α36, ER-α66, and ER-ß1) and Src expression. In conclusion, an efficient screening approach was developed to identify Berberine and Chelerythrine as potential selective candidates from Zanthoxylum simulans with significant anti-proliferative activity against gastric carcinoma. As we know, it was the first report to propose an estrogen signal pathway for Chelerythrine in anti-gastric cancer cells (AGS) study. The results supported Chelerythrine inhibitory effects on AGS by not only direct inhibiting JAKs but also down-regulating the estrogen pathway.

Potential Antioxidative Components in Azadirachta indica Revealed by Bio-Affinity Ultrafiltration with SOD and XOD.

Azadirachta indica (A. indica) has been widely used due to its diverse pharmacological activities. However, there are currently few studies on its responsible antioxidant ingredients against superoxide dismutase (SOD) and xanthine oxidase (XOD). In this study, the antioxidant activities of A. indica were evaluated by a 2,2'-azinobis-(3-ethyl-benzthiazoline)-6-sulfonic acid) and ferric-ion-reducing antioxidant power method. Meanwhile, total polyphenol and flavonoid content were determined to reveal that they were the highest in ethyl acetate (EA) fraction. Next, compounds with the most antioxidant activity were screened out from EA fraction by bio-affinity ultrafiltration liquid chromatography-mass spectrometry (UF-LC-MS) with SOD and XOD. As a result, gallic acid, protocatechuic acid and (-)-epicatechin were identified as potential SOD ligands with relative binding affinity (RBA) values of 2.15, 1.78 and 1.61, respectively. Additionally, these three ligands could effectively interact with SOD in molecular docking with binding energies (BEs) ranging from -3.84 ± 0.37 to -5.04 ± 0.01 kcal/mol. In addition, carnosic acid exhibited a strong binding affinity to XOD with an RBA value of 2.05 and BE value of -8.24 ± 0.71 kcal/mol. In conclusion, these results indicated that A. indica might have good antioxidant activity and antigout potential, and the UF-LC-MS method is suitable and efficient for screening both SOD and XOD ligands from A. indica.

Rapid and ultrasensitive electromechanical detection of ions, biomolecules and SARS-CoV-2 RNA in unamplified samples.

The detection of samples at ultralow concentrations (one to ten copies in 100 µl) in biofluids is hampered by the orders-of-magnitude higher amounts of 'background' biomolecules. Here we report a molecular system, immobilized on a liquid-gated graphene field-effect transistor and consisting of an aptamer probe bound to a flexible single-stranded DNA cantilever linked to a self-assembled stiff tetrahedral double-stranded DNA structure, for the rapid and ultrasensitive electromechanical detection (down to one to two copies in 100 µl) of unamplified nucleic acids in biofluids, and also of ions, small molecules and proteins, as we show for Hg2+, adenosine 5'-triphosphate and thrombin. We implemented an electromechanical biosensor for the detection of SARS-CoV-2 into an integrated and portable prototype device, and show that it detected SARS-CoV-2 RNA in less than four minutes in all nasopharyngeal samples from 33 patients with COVID-19 (with cycle threshold values of 24.9-41.3) and in none of the 54 COVID-19-negative controls, without the need for RNA extraction or nucleic acid amplification.

Potential hypoglycemic, hypolipidemic, and anti-inflammatory bioactive components in Nelumbo nucifera leaves explored by bioaffinity ultrafiltration with multiple targets.

Leaf of Nelumbo nucifera Gaertn. (N. nucifera) has been widely used as the main ingredient in lipid-lowering herbal teas and some prescriptions in China due to their excellent hypoglycemic and hypolipidemic effects. However, the active components responsible for these beneficial properties and their mechanisms remain unexplored. In this work, the N. nucifera leaf extracts significantly promoted the glucose consumption of HepG2 cells, and also exhibited remarkable inhibitory activities against α-glucosidase, pancreatic lipase, and COX-2. Furthermore, the top four potential active compounds (N-nornuciferine, Nuciferine, 2-Hydroxy-1-methoxyaporphine, and Isorhamnetin 3-O-glucoside) targeting the above three enzymes were screened out by bioaffinity ultrafiltration with multiple targets coupled with HPLC-MS/MS. The enzyme inhibitory activities of candidate compounds were verified by enzyme inhibition assay and molecular docking. In addition, molecular docking revealed the binding information between the candidate molecules and enzymes. The current study provided valuable information in discovering functional active ingredients from complex medicinal plant extracts.

Screening and characterisation of potential antioxidant, hypoglycemic and hypolipidemic components revealed in Portulaca oleracea via multi-target affinity ultrafiltration LC-MS and molecular docking.

INTRODUCTION: Portulaca oleracea is a commonly used nutritional vegetable and traditional herbal medicine with plenty of nutrients and manifold pharmacological activities. However, the potential active ingredients for its remarkable antioxidant, hypoglycemic and hypolipidemic activities remain unexplored. OBJECTIVES: The present study aims to systematically evaluate the antioxidant activities of different extracts of P. oleracea and screen bioactive ligands that can interact with α-glucosidase, pancreatic lipase, and superoxide dismutase (SOD). METHODS: In this research, the antioxidant activities of different parts of P. oleracea and their corresponding total phenolic content (TPC) and total flavonoid content (TFC) were systematically determined. Subsequently, a multi-target affinity ultrafiltration method was developed using affinity ultrafiltration with SOD, α-glucosidase, and pancreatic lipase coupled to liquid chromatography-mass spectrometry (UF-LC-MS). Later, molecular docking was used to further investigate the possible interaction mechanism between these ligands and target enzymes. RESULTS: Among them, the ethyl acetate (EA) fraction showed the highest antioxidant activity along with the highest TPC and TFC, and four compounds in the EA fraction were quickly retrieved as potential SOD, α-glucosidase, and pancreatic lipase ligands, respectively. Molecular docking revealed that these potential ligands exhibited strong binding ability and inhibitory activities on SOD, α-glucosidase, and pancreatic lipase. CONCLUSION: The present study revealed that P. oleracea can be used as a functional food with excellent antioxidant, hypoglycemic and hypolipidemic effects. Meanwhile, the integrated strategy based on multi-target UF-LC-MS and molecular docking also provided a powerful tool and a multidimensional perspective for further exploration of active ingredients in P. oleracea responsible for the antioxidant, hypoglycemic and hypolipidemic activities.

Stimulation of ROS Generation by Extract of Warburgia ugandensis Leading to G0/G1 Cell Cycle Arrest and Antiproliferation in A549 Cells.

Warburgia ugandensis Sprague (WU) is a traditional medicinal plant used for the treatment of various diseases, including cancer, in Africa. This study aimed to evaluate the anti-non-small cell lung cancer (NSCLC) activities of WU against A549 cells and to reveal potential molecular mechanisms. The cytotoxicity of various WU extracts was evaluated with HeLa (cervical cancer), HepG2 (liver cancer), HT-29 (colorectal cancer), and A549 (non-small cell lung cancer) cells by means of Sulforhodamine B (SRB) assay. Therein, the dimethyl carbonate extract of WU (WUD) was tested with the most potent anti-proliferative activity against the four cancer cell lines, and its effects on cell viability, cell cycle progression, DNA damage, intracellular reactive oxygen species (ROS), and expression levels of G0/G1-related proteins in A549 cells were further examined. First, it was found that WUD inhibited the proliferation of A549 cells in a time- and dose-dependent manner. In addition, WUD induced G0/G1 phase arrest and modulated the expression of G0/G1 phase-associated proteins Cyclin D1, Cyclin E1, and P27 in A549 cells. Furthermore, WUD increased the protein abundance of P27 by inhibiting FOXO3A/SKP2 axis-mediated protein degradation and also significantly induced the γH2AX expression and intracellular ROS generation of A549 cells. It was also found that the inhibitory effect of WUD on the proliferation and G0/G1 cell cycle progression of A549 cells could be attenuated by NAC, a ROS scavenger. On the other hand, phytochemical analysis of WUD with UPLC-QTOF-MS/MS indicated 10 sesquiterpenoid compounds. In conclusion, WUD exhibited remarkable anti-proliferative effects on A549 cells by improving the intracellular ROS level and by subsequently modulating the cell proliferation and G0/G1 cell cycle progression of A549 cells. These findings proved the good therapeutic potential of WU for the treatment of NSCLC.

Identification of Anti-Inflammatory and Anti-Proliferative Neolignanamides from Warburgia ugandensis Employing Multi-Target Affinity Ultrafiltration and LC-MS.

Previous reports have illustrated that the incidence and mortality of cancer are increasing year by year worldwide. In addition, the occurrence, development, recurrence and metastasis of cancer are closely related to inflammation, which is a kind of defensive response of human body to various stimuli. As an important medicinal plant in Africa, Warburgia ugandensis has been reported to have certain anti-inflammatory and anti-proliferative activities, but its specific components and mechanisms of action remain elusive. To tackle this challenge, affinity ultrafiltration with drug targets of interest coupled to high-performance liquid chromatography-mass spectrometry (AUF-HPLC-MS/MS) could be utilized to quickly screen out bioactive constituents as ligands against target enzymes from complex extracts of this plant. AUF-HPLC-MS/MS with four drug targets, i.e., cyclooxygenase-2 (COX-2), 5-lipoxygenase (5-LOX), topoisomerase I (Top I) and topoisomerase II (Top II) were used to rapidly screen and characterize the anti-inflammatory and anti-proliferative natural ligands from W. ugandensis, and the resulting potential active compounds as ligands with specific binding affinity to COX-2, 5-LOX, Top I and Top II, were isolated with modern separation and purification techniques and identified with spectroscopic method like NMR, and then their antiinflammatory and anti-proliferative activities were tested to verify the screening results from AUF-HPLC-MS/MS. Compounds 1 and 2, which screened out and identified from W. ugandensis showed remarkable binding affinity to COX-2, 5-LOX, Top I and Top II with AUF-HPLC-MS/MS. In addition, 1 new compound (compound 3), together with 5 known compounds were also isolated and identified from W. ugandensis. The structure of compound 3 was elucidated by extensive 1D, 2D NMR data and UPLC-QTOF-MS/MS. Furthermore, compounds 1 and 2 were further proved to possess both anti-inflammatory and anti-proliferative activities which are in good agreement with the screening results using AUF-HPLC-MS/MS. This work showcased an efficient method for quickly screening out bioactive components with anti-inflammatory and anti-proliferative activity from complex medicinal plant extracts using AUF-HPLC-MS/MS with target enzymes of interest, and also demonstrated that neolignanamides (compounds 1 and 2) from W. ugandensis would be the active components responsible for its anti-inflammatory and anti-proliferative activity with the potential to treat cancer and inflammation.

Phenolic Compounds from Carissa spinarum Are Characterized by Their Antioxidant, Anti-Inflammatory and Hepatoprotective Activities.

Carissa spinarum has been traditionally used for the treatment of various diseases due to its different pharmacological activities. However, the active compounds responsible for its potentially specific activities have rarely been explored. To this end, the ethyl acetate (EA) fraction was screened out and selected for further phytochemical isolation because of its promising activities in preliminary 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and COX-2 inhibition assays. As a result, 10 compounds (1-10), including a new one (5), were isolated, with eight of these being identified as phenolic compounds, as expected. Compound 9 possessed an IC50 value of 16.5 ± 1.2 µM, which was lower than that of positive control (vitamin C, 25.5 ± 0.3 µM) in the DPPH assay, and compounds 2, 6, 7 and 9 showed better total antioxidant capacity than vitamin C in the FRAP assay. Meanwhile, compounds 1-6 and 9 also had IC50 values of less than 1.0 µM, which was even better than the positive control indomethacin in the COX-2 inhibition assay. In this context, compounds 2 and 9 were further evaluated to exhibit clear hepatoprotective activities by improving the L02 cell viability and reducing ROS production using a H2O2-induced L02 cell injury model. This study provides initial evidence revealing the most potent phenolic compounds from the root bark of C. spinarum responsible for its antioxidant, anti-inflammatory and hepatoprotective activities.

Antioxidant and Antiproliferative Potentials of Ficus glumosa and Its Bioactive Polyphenol Metabolites.

Ficus glumosa Delile (Moraceae), a reputed plant that is used in herbal medicine, is of high medicinal and nutritional value in local communities primarily ascribed to its phytochemical profile. Currently, there are hardly any fine details on the chemical profiling and pharmacological evaluation of this species. In this study, the flavonoids and phenolics contents of the ethanol extracts and four extracted fractions (petroleum ether (PE), ethyl acetate (EA), n-butanol, and water) of the stem bark of Ficus glumosa were firstly quantified. Further, their antioxidant and antiproliferative potentials were also evaluated. The quantitative determination indicated that the EA and n-butanol fractions possessed the highest total flavonoids/phenolics levels of 274.05 ± 0.68 mg RE/g and 78.87 ± 0.97 mg GAE/g, respectively. Similarly, for the 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and ferric-reducing antioxidant power (FRAP) assays, the EA fraction exhibited high potency in both DPPH and ABTS+ scavenging activities with IC50 values of 0.23 ± 0.03 mg/mL, 0.22 ± 0.03 mg/mL, and FRAP potential of 2.81 ± 0.01 mg Fe2+/g, respectively. Furthermore, the EA fraction displayed high cytotoxicity against human lung (A549) and colon (HT-29) cancer cells. Additionally, the liquid chromatography coupled with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) was employed in order to characterize the chemical constituents of the EA fraction of Ficus glumosa stem bark. Our findings revealed 16 compounds from the EA fraction that were possibly responsible for the strong antioxidant and anti-proliferative properties. This study provides edge-cutting background information on the exploitation of Ficus glumosa as a potential natural antioxidant and anti-cancer remedy.

Epigenetic Landscapes of Single-Cell Chromatin Accessibility and Transcriptomic Immune Profiles of T Cells in COVID-19 Patients.

T cells play a critical role in coronavirus diseases. How they do so in COVID-19 may be revealed by analyzing the epigenetic chromatin accessibility of cis- and trans-regulatory elements and creating transcriptomic immune profiles. We performed single-cell assay for transposase-accessible chromatin (scATAC) and single-cell RNA (scRNA) sequencing (seq) on the peripheral blood mononuclear cells (PBMCs) of severely ill/critical patients (SCPs) infected with COVID-19, moderate patients (MPs), and healthy volunteer controls (HCs). About 76,570 and 107,862 single cells were used, respectively, for analyzing the characteristics of chromatin accessibility and transcriptomic immune profiles by the application of scATAC-seq (nine cases) and scRNA-seq (15 cases). The scATAC-seq detected 28,535 different peaks in the three groups; among these peaks, 41.6 and 10.7% were located in the promoter and enhancer regions, respectively. Compared to HCs, among the peak-located genes in the total T cells and its subsets, CD4+ T and CD8+ T cells, from SCPs and MPs were enriched with inflammatory pathways, such as mitogen-activated protein kinase (MAPK) signaling pathway and tumor necrosis factor (TNF) signaling pathway. The motifs of TBX21 were less accessible in the CD4+ T cells of SCPs compared with those in MPs. Furthermore, the scRNA-seq showed that the proportion of T cells, especially the CD4+ T cells, was decreased in SCPs and MPs compared with those in HCs. Transcriptomic results revealed that histone-related genes, and inflammatory genes, such as NFKBIA, S100A9, and PIK3R1, were highly expressed in the total T cells, CD4+ T and CD8+ T cells, both in the cases of SCPs and MPs. In the CD4+ T cells, decreased T helper-1 (Th1) cells were observed in SCPs and MPs. In the CD8+T cells, activation markers, such as CD69 and HLA class II genes (HLA-DRA, HLA-DRB1, and HLA-DRB5), were significantly upregulated in SCPs. An integrated analysis of the data from scATAC-seq and scRNA-seq showed some consistency between the approaches. Cumulatively, we have generated a landscape of chromatin epigenetic status and transcriptomic immune profiles of T cells in patients with COVID-19. This has provided a deeper dissection of the characteristics of the T cells involved at a higher resolution than from previously obtained data merely by the scRNA-seq analysis. Our data led us to suggest that the T-cell inflammatory states accompanied with defective functions in the CD4+ T cells of SCPs may be the key factors for determining the pathogenesis of and recovery from COVID-19.

Inhibition of Growth of Colon Tumors and Proliferation of HT-29 Cells by Warburgia ugandensis Extract through Mediating G0/G1 Cell Cycle Arrest, Cell Apoptosis, and Intracellular ROS Generation.

Warburgia ugandensis Sprague (W. ugandensis), widely distributed in Africa, is a traditional medicinal plant used for the treatment of various diseases including cancer. We intended to evaluate the anticolorectal cancer (CRC) activities of the crude extract from W. ugandensis (WUD) and reveal the underlying molecular mechanisms of its action. We found that WUD inhibited the proliferation of HT-29 and HCT116 cells in a time- and dose-dependent manner and induced intracellular ROS generation. The inhibitory effect of WUD on the proliferation of HT-29 and HCT116 cells could be attenuated by NAC (a ROS scavenger) in a dose-dependent manner. WUD induced G0/G1 phase arrest, down-regulated the protein expression of Cyclin D1 via ROS accumulation in HT-29 cells. In search of the molecular mechanism involved in WUD-induced Cyclin D1 down-regulation, it was found that WUD can suppress PI3K/Akt/GSK3ß signaling pathway in HT-29 cells. Next, it was found that WUD also activated apoptosis, poly-ADP ribose polymerase 1 (PARP1) cleavage and down-regulated pro-caspase 3 in HT-29 and HCT116 cells. Besides, WUD decreased the growth of colon tumors in vivo in the xenograft mouse model. We demonstrated for the first time that ROS and their modulation in the corresponding intracellular signaling could play a significant role in the potential activity of WUD against CRC cells.

An inter-correlated cytokine network identified at the center of cytokine storm predicted COVID-19 prognosis.

The hyper-inflammatory response is thought to be a major cause of acute respiratory distress syndrome (ARDS) in patients with COVID-19. Although multiple cytokines are reportedly associated with disease severity, the key mediators of SARS-CoV-2 induced cytokine storm and their predictive values have not been fully elucidated. The present study analyzed maximal and early (within 10 days after disease onset) concentrations of 12-plex cytokines in plasma. We found consistently elevated plasma levels of IL-6, IL-8 and IL-5 in patients who were deceased compared with those who had mild/moderate or severe disease. The early plasma concentrations of IFN-a and IL-2 positively correlated with the length of the disease course. Moreover, correlation network analysis showed that IL-6, IL-8, and IL-5 located at the center of an inter-correlated cytokine network. These findings suggested that IL-8, IL-6, IL-5 might play central roles in cytokine storms associated with COVID-19 and that the early detection of multiple plasma cytokines might help to predict the prognosis of this disease.

Comparing Three Different Extraction Techniques on Essential Oil Profiles of Cultivated and Wild Lotus (Nelumbo nucifera) Flower.

Essential oil components of Nelumbo nucifera flowers from cultivated and wild lotus samples were analyzed and compared using three different extraction techniques, i.e., headspace extraction (HE), steam distillation (SD) and solvent extraction (SE), coupled with GC-MS. Forty-two peaks in the GC-MS analysis were identified as essential oil components extracted by the three methods from N. nucifera flower. The major essential oil components extracted by SD method were found to be Z,Z-10,12-hexadecadienal and E-14-hexadecenal with relative contents of 16.3% and 16.7%, respectively, which is different from that of SE method, i.e., n-hexadecanoic acid and Z,Z-9,12-octadecadienoic acid accounting for 25.8% and 26.8%, respectively. HE method demonstrated a possibility to be used as an in situ and simplest method for extracting the essential oil components from raw materials. By adding a small amount of glycerinum onto the surface of the air-dried flower sample as a solvent trap in the HE method, the volatility of the essential oil components was found to increase by two times for the first time, which could be further utilized to improve the extraction efficiency and the recovery of the essential oil components from other materials for more applications. In addition, the comparison of essential oil components between cultivated and wild samples showed that they differed only in the chemical contents but not in chemical components. This will be a comprehensive report on the chemical information of the essential oil components of N. nucifera flower and provide guidance for its further exploration as high value-added products in the food and healthcare industries.

Effectiveness of a single treatment of photodynamic therapy using topical administration of 5-aminolevulinic acid on methicillin-resistant Staphylococcus aureus-infected wounds of diabetic mice.

Methicillin-resistant Staphylococcus aureus (MRSA) is the most important representative pathogen which causes clinically relevant infections in diabetic ulceration. We report our investigations on the efficacy of antimicrobial photodynamic therapy with topical 5-aminolevulinic acid (ALA-PDT) on diabetic murine infected wound models induced with a MRSA SA325 strain. A solution of 10 % 5-aminolevulinic acid (ALA) was placed into the wounds followed by delivery of 25 J/cm2 (635 nm). The ALA-PDT treated wounds healed earlier as compared to others (P < 0.5). A significant reduction of bacterial counts (2.05 logs) was detected in wounds after ALA-PDT on Day 2 (P < 0.5). Additionally, histological analysis revealed that wounds treated with ALA-PDT exhibited a more complete re-epithelialization, blood micro-vessels, collagen-volume fraction and considerable decrease in inflammatory cells infiltration. Immunohistochemistry assay demonstrated considerable reduction of proinflammatory cytokines interleukin-1ß (IL-1ß), earlier expression of vascular endothelial growth factor A (VEGF-A), increase of basic fibroblast growth factor (bFGF) and Ki-67 in the ALA-PDT group (P < 0.5). These data imply that the therapeutic effect of ALA-PDT revealed an accelerated diabetic wound closure rate, together with reduced hyperinflammatory response and elevated growth factors.