An ex vivo method to evaluate vasoactivity induced by hemoglobin-based oxygen carriers in resistance vessels.

Red blood cell substitutes offer a solution to the problem of blood shortage and side effects of blood transfusion. Hemoglobin-based oxygen carriers (HBOCs) are one of the promising substitutes for red blood cells. Vasoactivity, which refers to the side effect of HBOCs that causes vasoconstriction and subsequent hypertension, limits the clinical application of HBOCs. In this study, an ex vivo method for the evaluation of vasoactivity induced by HBOCs was established based on isolated rat mesenteric artery vessels and the DMT120CP system. The DMT120CP system, equipped with a flowmeter, permits the control of intravascular pressure, pressure gradient, and flow conditions with high accuracy, simulating the physiological conditions for isolated vessels. The concentration of noradrenaline was optimized to 1 × 10-6∼3 × 10-6 M. PEGylated bovine hemoglobin (PEG-bHb) was synthesized and perfused into the vessel for vasoactivity evaluation, with bHb as the positive control and PSS buffer solution as the negative control. PEG-bHb showed a hydration diameter of 15.5 ± 1.4 nm and a P50 value of 6.99 mmHg. PEG-bHb exhibited a colloid osmotic pressure of 64.1 mmHg and a viscosity of 1.73 cp at 40 mg/mL. The established vasoactivity evaluation method showed significant differences in samples (bHb or PEG-bHb) with different vasoactivity properties. The vasoconstriction percentage induced by PEG-bHb samples synthesized in different batches showed coefficients of variation less than 5%, indicating good applicability and repeatability. The established evaluation method can be applied to study the vasoactivity induction and elimination strategies, promoting the clinical application of HBOCs.

Effect of roxadustat on red blood cell lifespan in patients with long-term haemodialysis: a single-centre, prospective, single-arm study.

Background: Reduced survival of red blood cells (RBCs) in patients with chronic kidney disease (CKD) is thought to contribute to renal anaemia. Although renal anaemia improved greatly because of the wide use of erythropoiesis-stimulating agents (ESAs) and the advancement of dialysis techniques, RBC longevity seems not to be obviously ameliorated. Methods: In this single-centre, single-arm trial, patients who had been undergoing haemodialysis and ESA therapy with epoetin alfa for at least 12 weeks changed their anti-anaemia drugs from epoetin alfa to oral roxadustat three times per week for 24 weeks. The primary endpoint was the change in RBC lifespan from baseline at week 24. The change in the circulating percentage of eryptotic RBCs, RBC deformability and RBC oxygen transport ability were also assessed. Results: A total of 27 patients were enrolled, with 26 completing the full course of intervention. At baseline, the average RBC lifespan was 60.1 days [standard deviation (SD) 14.4; n = 27]. At the end of the study period, 26 patients had an RBC lifespan measurement (83.9 days on average; SD 21.9). The RBC lifespan increased by 22.8 days on average [95% confidence interval (CI) 15.5-30.0, P < .001]. This equated to an average RBC lifespan increase of 39.2% (95% CI 27.8-50.6). The percentage of circulating eryptotic RBCs, erythrocyte filtration index and the pressure at which haemoglobin is 50% saturated decreased significantly from baseline to week 24 (1.39 ± 0.44% versus 0.89 ± 0.25%, P < .0001; 0.29 ± 0.12 versus 0.16 ± 0.08, P < .0001 and 32.54 ± 4.83 versus 28.40 ± 2.29, P < .001, respectively). Conclusion: Roxadustat prolonged RBC lifespan in patients with long-term haemodialysis.

Cefmetazole sodium as an allosteric effector that regulates the oxygen supply efficiency of adult hemoglobin.

Allosteric effectors play an important role in regulating the oxygen supply efficiency of hemoglobin for blood storage and disease treatment. However, allosteric effectors that are approved by the US FDA are limited. In this study, cefmetazole sodium (CS) was found to bind adult hemoglobin (HbA) from FDA library (1338 compounds) using surface plasmon resonance imaging high-throughput screening. Using surface plasmon resonance (SPR), the interaction between CS and HbA was verified. The oxygen dissociation curve of HbA after CS interaction showed a significant increase in P50 and theoretical oxygen-release capacity. Acid-base sensitivity (SI) exhibited a decreasing trend, although not significantly different. An oxygen dissociation assay indicated that CS accelerated HbA deoxygenation. Microfluidic modulated spectroscopy showed that CS changed the ratio of the alpha-helix to the beta-sheet of HbA. Molecular docking suggested CS bound to HbA's ß-chains via hydrogen bonds, with key amino acids being N282, K225, H545, K625, K675, and V544.The results of molecular dynamics simulations (MD) revealed a stable orientation of the HbA-CS complex. CS did not significantly affect the P50 of bovine hemoglobin, possibly due to the lack of Valß1 and Hisß2, indicating that these were the crucial amino acids involved in HbA's oxygen affinity. Competition between the 2,3-Diphosphoglycerate (2,3-DPG) and CS in the HbA interaction was also determined by SPR, molecular docking and MD. In summary, CS could interact with HbA and regulate the oxygen supply efficiency via forming stable hydrogen bonds with the ß-chains of HbA, and showed competition with 2,3-DPG.Communicated by Ramaswamy H. Sarma.

Resveratrol, a New Allosteric Effector of Hemoglobin, Enhances Oxygen Supply Efficiency and Improves Adaption to Acute Severe Hypoxia.

Acute altitude hypoxia represents the cause of multiple adverse consequences. Current treatments are limited by side effects. Recent studies have shown the protective effects of resveratrol (RSV), but the mechanism remains unknown. To address this, the effects of RSV on the structure and function of hemoglobin of adult (HbA) were preliminarily analyzed using surface plasmon resonance (SPR) and oxygen dissociation assays (ODA). Molecular docking was conducted to specifically analyze the binding regions between RSV and HbA. The thermal stability was characterized to further validate the authenticity and effect of binding. Changes in the oxygen supply efficiency of HbA and rat RBCs incubated with RSV were detected ex vivo. The effect of RSV on the anti-hypoxic capacity under acute hypoxic conditions in vivo was evaluated. We found that RSV binds to the heme region of HbA following a concentration gradient and affects the structural stability and rate of oxygen release of HbA. RSV enhances the oxygen supply efficiency of HbA and rat RBCs ex vivo. RSV prolongs the tolerance times of mice suffering from acute asphyxia. By enhancing the oxygen supply efficiency, it alleviates the detrimental effects of acute severe hypoxia. In conclusion, RSV binds to HbA and regulates its conformation, which enhances oxygen supply efficiency and improves adaption to acute severe hypoxia.

PEG-conjugated bovine haemoglobin enhances efficiency of chemotherapeutic agent doxorubicin with alleviating DOX-induced splenocardiac toxicity in the breast cancer.

Doxorubicin (DOX) is an effective chemotherapeutic agent widely used for cancer treatment. However, hypoxia in tumour tissue and obvious adverse effects particularly cardiotoxicity restricts the clinical usage of DOX. Our study is based on the co-administration of haemoglobin-based oxygen carriers (HBOCs) and DOX in a breast cancer model to investigate HBOCs' ability to enhance chemotherapeutic effectiveness and its capabilities to alleviate the side effects induced by DOX. In an in-vitro study, the results suggested the cytotoxicity of DOX was significantly improved when combined with HBOCs in a hypoxic environment, and produced more γ-H2AX indicating higher DNA damage than free DOX did. Compared with administration of free DOX, combined therapy exhibited a stronger tumour suppressive effect in an in-vivo study. Further mechanism studies showed that the expression of various proteins such as hypoxia-inducible factor-1α (HIF-1α), CD31, CD34, and vascular endothelial growth factor (VEGF) in tumour tissues was also significantly reduced in the combined treatment group. In addition, HBOCs can significantly reduce the splenocardiac toxicity induced by DOX, according to the results of the haematoxylin and eosin (H&E) staining and histological investigation. This study suggested that PEG-conjugated bovine haemoglobin may not only reduce the hypoxia in tumours and increase the efficiency of chemotherapeutic agent DOX, but also alleviate the irreversible heart toxicity caused by DOX-inducted splenocardiac dysregulation.

Insights into the Composition and Antibacterial Activity of Amomum tsao-ko Essential Oils from Different Regions Based on GC-MS and GC-IMS.

Chemical components are one of the most significant traits and attributes of plant tissues, and lead to their different functions. In this study, the composition of Amomun tsao-ko essential oils (AEOs) from different regions was first determined by a combination of gas chromatography-mass spectrometry (GC-MS) and gas chromatography-ion mobility spectrometry (GC-IMS). In total, 141 compounds were identified, of which terpenes and aldehydes were the main groups. Orthogonal partial least square discriminant analysis (OPLS-DA) distinguished the samples from different regions clearly, and the main differences were terpenes, aldehydes, and esters. Meanwhile, AEOs showed strong antibacterial activity against Staphylococcus aureus (S. aureus), and the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) reached 0.20 mg/mL and 0.39-0.78 mg/mL, respectively. From correlation analysis, 1,8-cineole, (E)-dec-2-enal, citral, α-pinene, and α-terpineol were determined to be the potential antibacterial compounds. This study provides the basis for the variety optimization of A. tsao-ko and its application as a natural food preservative.

[The Genome-Wide Changes in Expression Profile of CML T Cells After Up-regulation of TCRζ Chain Expression].

OBJECTIVE: To analyze the changes in the gene expression profile of T cells in CML patients after TCRζ up-regulation expression, and to explore the molecular mechanism of T cell reactivation after transgenic up-regulation of TCRζ. METHODS: The peripheral blood mononuclear cells(PBMCs) from 3 newly untreated chronic-stage CML patients were collected, and the CD3+ T cells were obtained by MACS method. The TCRζ-IRES2-EGFP (experimental group) and pIRES2-EGFP (control group) plasmids were transfected into T cells by nuclear transfection technique. The gene expression profiles of CML T cells up-regulated TCRζ chain and control cells were detected by Affymetrix GeneChip Human Gene 2.0 ST Array. The differentially expressed genes were analyzed by GO functional annotation analysis and KEGG pathway enrichment analysis. RESULTS: A total of 2248 differentially-expressed genes were obtained, including 553 up-regulated genes and 1695 down-regulated genes in experimental group as compared with those in control group (P<0.05) . The GO and KEGG enrichment analyses showed that differentially expressed genes involved in the biological processes related to T cell immune function, such as TCR signaling pathway, T cell proliferation and activation. Some of core genes involved in promoting the TCR signaling pathway, T cell proliferation, activation and apoptosis pathways were significantly up-regulated, while some core genes involved in inhibiting T cell activation were significantly down-regulated. CONCLUSION: The molecular mechanism of the significantly improved T cell activation and proliferation ability in CML patients after TCRζ up-regulation may be related to the differential transcripts mediated signaling pathways of T cell activation, proliferation and apoptosis.

Monitoring of adsorption and transfer of organochlorines in soybean seeds and sprouts with mass spectrometric imaging.

Development of analytical techniques that can monitor the adsorption, transfer and in-situ distribution of environmental pollutants in agricultural products is essential to ensure the implementation of stringent food safety standards for consumer protection. A mass spectrometric imaging approach is described herein to investigate the dynamic changes and spatial distributions of 4, 4'-DDT (dichlorodiphenyltrichloroethane) in soybean seeds and sprouts during the growth. Soy beans seeds incubated in DDT containing water were sliced in every 20 µm and directly blotted on the surface of a compressed thin film of (Bi2O3)0.07(CoO)0.03(ZnO)0.9 nanoparticles. Endogenous molecules and exogenous DDT compounds in soy bean seeds were ionized and dissociated by photoelectrons that are generated on surfaces of semiconductor nanoparticles upon the irradiation of the 3rd harmonic (355 nm) of Nd3+:YAG laser. Structural identification is achieved by the interpretation of fragment ions resulting from electron-initiated specific bond cleavages or hole oxidization. Mass spectrometric images reveal increased quantities of DDT residues in soy bean seeds and sprouts during the growth. It provides an in situ way without extensive sample preparation to monitor the transfer and distribution of exogenous pollutants as well as the possible impacts on plant growth.

Real-time laser induced chemical derivatizations of peptide N-Terminus for in-situ mass spectrometric sequencing at sub-picomole and nanosecond scale.

Distinguishing b- and y-ions is essential to compute amino acid sequences from either N- or C-terminus in mass spectrometry. We described herein a solvent free and real time on-plate derivatization approach that can tag N-terminus of peptides at microliter level with p-chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde for matrix assisted laser desorption ionization mass spectrometry (MALDI MS). Less than 1 µL of sample solutions can be directly mixed with equal volumes of p-chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde and α-cyano-4-hydroxycinnamic acid (CHCA), a matrix compound to co-crystalize with analytes for efficient absorption of laser energy and peptide ionization. When the mixture spotted on the sample plate is irradiated with the 3rd harmonic (355 nm) of Nd3+:YAG laser pulses (3 ns width), N-terminal amine groups of peptides instantly react with carbonyl groups of chlorobenzaldehyde or 2-hydroxy-5-methylisophthalaldehyde. Resultant peptides carrying with on-plate formed azomethine group (-CN-) are simultaneously protonated and isolated as precursor ions for subsequent collision-activated dissociation. The mass shift with unique Cl isotopic signature unambiguously distinguishes b ions from y ions and other ions. This method does not need extensive sample preparation and is useful for those samples with limited quantities down to sub-picomole level in sub-microliter volumes. The efficiency was demonstrated with synthetic peptides and tryptic peptides of model proteins. It was found that 2-hydroxy-5-methylisophthalaldehyde provides improved yield for peptides containing lysine residues. Unknown proteins of human saliva and bovine milk as well as phosphopeptides have been identified.

Enhanced Electrocatalytic Activity of Trace Pt in Ternary CuCoPt Alloy Nanoparticles for Hydrogen Evolution.

We report an enhanced high electrocatalytic hydrogen evolution activity of trace Pt and Co diluted in ternary CuCoPt alloy nanoparticles with Cu as the substrate. Using only 10% Pt atoms can display even better activity and stability in hydrogen evolution reactions than using pure Pt nanoparticles.

Electron Acceptive Mass Tag for Mass Spectrometric Imaging-Guided Synergistic Targeting to Mice Brain Glutamate Receptors.

Dysfunctional glutamate receptors (GluRs) have been implicated in neurological disorders and injuries. Hetero-tetrameric assemblies of different GluR subunits or splicing variants have distinct spatiotemporal expression patterns and pharmacological properties. Mass spectrometric imaging of GluRs-targeted small molecules is important for determining the regional preferences of these compounds. We report herein the development of a mass tag covalently bonded with glutamate or N-methyl-d-aspartate that functions as both an electron acceptor to generate mass spectrometric signals on irradiated (Bi2O3)0.07(CoO)0.03(ZnO)0.9 nanoparticles with the third harmonic (355 nm) of Nd3+:YAG laser and as the core component to target bilobed clamshell-like structures of GluRs. In this approach, different molecules produce the same tag ion. It provides a new avenue for quantitative assessment of spatial densities of different compounds, which cannot be achieved with well-established stable isotope labeling technique due to different ionization efficiency of different compounds. Various coexisting endogenous molecules are also simultaneously detected for investigation of overall physiological changes induced by these compounds. Because semiconductors do not generate background peaks, this method eliminates interferences from organic matrix materials that are used in regular MALDI (matrix assisted laser desorption ionization). The localized ionization provides high spatial resolution that can be down to sub-micrometers.

Heteroatom tri-doped porous carbon derived from waste biomass as Pt-free counter electrode in dye-sensitized solar cells.

Strategies for environmentally friendly reutilization of waste biomass are highly desirable nowadays. Meanwhile, seeking Pt-free electrocatalysts for triiodide reduction with both high catalytic activity and low cost is critical for the development of dye-sensitized solar cells (DSCs). In the study, heteroatom tri-doped porous carbons (TPCs) were prepared via carbonization of a typical food waste (fish waste) and explored as a counter electrode (CE) in a DSC. The as-prepared carbon materials possessed a porous structure with a large BET surface area of 2933 m2 g-1, while being simultaneously naturally doped by three heteroatoms (N, P and S). More importantly, the resultant N, P, S-tri-doped porous carbon exhibited outstanding electrochemical activity towards triiodide reduction with good stability. Moreover, the DSC with the optimized TPC electrode showed a power conversion efficiency of 7.83%, which is comparable to the device with a costly Pt-based CE (8.34%), measured under one sun illumination (AM 1.5G). This work demonstrates that carbonization of fish waste offers a cost-effective approach to prepare multifunctional carbon materials for advanced energy applications.

A Cost-Effective Approach to Optimizing Microstructure and Magnetic Properties in Ce17Fe78B6 Alloys.

Optimizing fabrication parameters for rapid solidification of Re-Fe-B (Re = Rare earth) alloys can lead to nanocrystalline products with hard magnetic properties without any heat-treatment. In this work, we enhanced the magnetic properties of Ce17Fe78B6 ribbons by engineering both the microstructure and volume fraction of the Ce2Fe14B phase through optimization of the chamber pressure and the wheel speed necessary for quenching the liquid. We explored the relationship between these two parameters (chamber pressure and wheel speed), and proposed an approach to identifying the experimental conditions most likely to yield homogenous microstructure and reproducible magnetic properties. Optimized experimental conditions resulted in a microstructure with homogeneously dispersed Ce2Fe14B and CeFe2 nanocrystals. The best magnetic properties were obtained at a chamber pressure of 0.05 MPa and a wheel speed of 15 m·s-1. Without the conventional heat-treatment that is usually required, key magnetic properties were maximized by optimization processing parameters in rapid solidification of magnetic materials in a cost-effective manner.