Harnessing ultrasound-derived hydroxyl radicals for the selective oxidation of aldehyde functions.

Ultrasonic irradiation holds potential for the selective oxidation of non-volatile organic substrates in the aqueous phase by harnessing hydroxyl radicals as chemical initiators. Here, a mechanistic description of hydroxyl radical-initiated glyoxal oxidation is constructed by gleaning insights from photolysis and radiation chemistry to explain the yields and kinetic trends for oxidation products. The mechanistic description and kinetic measurements reported herein reveal that increasing the formation rate of hydroxyl radicals by changing the ultrasound frequency increases both the rates of glyoxal consumption and the selectivity towards C2 acid products over those from C-C cleavage. Glyoxal consumption also occurs more rapidly and with greater selectivity towards C2 acids under acidic conditions, which favor the protonation of carboxylate intermediates into their less reactive acidic forms. Leveraging such pH and frequency effects is crucial to mitigating product degradation by secondary reactions with hydroxyl radicals and oxidation products (specifically hydrogen peroxide and superoxide). These findings demonstrate the potential of ultrasound as a driver for the selective oxidation of aldehyde functions to carboxylic acids, offering a sustainable route for valorizing biomass-derived platform molecules.

Elucidating the Strain-Vacancy-Activity Relationship on Structurally Deformed Co@CoO Nanosheets for Aqueous Phase Reforming of Formaldehyde.

Lattice strain modulation and vacancy engineering are both effective approaches to control the catalytic properties of heterogeneous catalysts. Here, Co@CoO heterointerface catalysts are prepared via the controlled reduction of CoO nanosheets. The experimental quantifications of lattice strain and oxygen vacancy concentration on CoO, as well as the charge transfer across the Co-CoO interface are all linearly correlated to the catalytic activity toward the aqueous phase reforming of formaldehyde to produce hydrogen. Mechanistic investigations by spectroscopic measurements and density functional theory calculations elucidate the bifunctional nature of the oxygen-vacancy-rich Co-CoO interfaces, where the Co and the CoO sites are responsible for CH bond cleavage and OH activation, respectively. Optimal catalytic activity is achieved by the sample reduced at 350 °C, Co@CoO-350 which exhibits the maximum concentration of Co-CoO interfaces, the maximum concentration of oxygen vacancies, a lattice strain of 5.2% in CoO, and the highest aqueous phase formaldehyde reforming turnover frequency of 50.4 h-1 at room temperature. This work provides not only new insights into the strain-vacancy-activity relationship at bifunctional catalytic interfaces, but also a facile synthetic approach to prepare heterostructures with highly tunable catalytic activities.

Strong Metal-Support Interaction for 2D Materials: Application in Noble Metal/TiB2 Heterointerfaces and their Enhanced Catalytic Performance for Formic Acid Dehydrogenation.

Strong metal-support interaction (SMSI) is a phenomenon commonly observed on heterogeneous catalysts. Here, direct evidence of SMSI between noble metal and 2D TiB2 supports is reported. The temperature-induced TiB2 overlayers encapsulate the metal nanoparticles, resulting in core-shell nanostructures that are sintering-resistant with metal loadings as high as 12.0 wt%. The TiOx -terminated TiB2 surfaces are the active sites catalyzing the dehydrogenation of formic acid at room temperature. In contrast to the trade-off between stability and activity in conventional SMSI, TiB2 -based SMSI promotes catalytic activity and stability simultaneously. By optimizing the thickness and coverage of the overlayer, the Pt/TiB2 catalyst displays an outstanding hydrogen productivity of 13.8 mmol g-1 cat h-1 in 10.0 m aqueous solution without any additive or pH adjustment, with >99.9% selectivity toward CO2 and H2 . Theoretical studies suggest that the TiB2 overlayers are stabilized on different transition metals through an interplay between covalent and electrostatic interactions. Furthermore, the computationally determined trends in metal-TiB2 interactions are fully consistent with the experimental observations regarding the extent of SMSI on different transition metals. The present research introduces a new means to create thermally stable and catalytically active metal/support interfaces for scalable chemical and energy applications.

[Establishment of A Novel Method to Activate and Expand Human NK Cells Ex Vivo].

OBJECTIVE: To develop a new method to activate and expand human NK cells ex vivo by using sodium hyaluronate as a major activating agent and to explore its related mechanism. METHODS: Mononuclear cells were isolated from 3 samples of peripheral blood from three healthy donors. New NK cell culture method and the control method were used to culture NK cells from each samples separately for 14 days. Flow cytometry was used to analyze the ratio of NK cells and CD69 expression. To measure the in vitro cytotoxicity of NK cells cultured by the two methods, the K562 cells were used as the targeting cells and flow cytometry combined with CFSE marker was used as the testing method. RESULTS: After culturing for 14 days, the number of NK cells obtained by new culture method for NK cells expanded by 188.63±3.83 times while the number of NK cells cultured by control method expanded by 152.77±5.77 times. The ratio of NK cells in new cell culture method was above 90%, while the ratio of NK cells in control method was about 70%. The ratio of CD69+ NK cells in new cell culture method was 32.37%±3.22%, while the ratio of CD69+ NK cells in control method was 17.29%±3.79%. The results of cytotoxicity experiment in vitro showed that NK cells cultured by the new method had a higher killing ability to the target cells as compared with NK cells cultured by the control method. CONCLUSION: New NK cell culture method using sodium hyaluronate as a major activating agent can expand NK cells more efficiently as compared with the cells cultured by control method, which may be related to the direct and/or indirect activation of sodium hyaluronate to NK cells, further causing the dominant expansion of the NK cells.

Challenges and Research in Managing Blood Supply in China.

Although considerable progress has been made in improving the blood service system in China over the last 2 decades, many challenges remain. A number of issues have received public attentions; however, others continue to be underacknowledged and controversial. This article describes 3 of these important and less emphasized issues: first, the ambiguity of the definition of voluntary nonremunerated blood donation and its relationship to an adequate blood supply; second, the current inadequacies of cost recovery from the blood service system; and third, the lack of a universally implemented program of hemovigilance. Currently, there is controversy regarding these challenges. Open recognition and discussion offers the prospect of bringing solutions closer to reality.

Biochemical characterization of a thermostable HNH endonuclease from deep-sea thermophilic bacteriophage GVE2.

His-Asn-His (HNH) proteins are a very common family of small nucleic acid-binding proteins that are generally associated with endonuclease activity and are found in all kingdoms of life. Although HNH endonucleases from mesophiles have been widely investigated, the biochemical functions of HNH endonucleases from thermophilic bacteriophages remain unknown. Here, we characterized the biochemical properties of a thermostable HNH endonuclease from deep-sea thermophilic bacteriophage GVE2. The recombinant GVE2 HNH endonuclease exhibited non-specific cleavage activity at high temperature. The optimal temperature of the GVE2 HNH endonuclease for cleaving DNA was 60-65 °C, and the enzyme retained its DNA cleavage activity even after heating at 100 °C for 30 min, suggesting the enzyme is a thermostable endonuclease. The GVE2 HNH endonuclease cleaved DNA over a wide pH spectrum, ranging from 5.5 to 9.0, and the optimal pH for the enzyme activity was 8.0-9.0. Furthermore, the GVE2 HNH endonuclease activity was dependent on a divalent metal ion. While the enzyme is inactive in the presence of Cu(2+), the GVE2 HNH endonuclease displayed cleavage activity of varied efficiency with Mn(2+), Mg(2+), Ca(2+), Fe(2+), Co(2+), Zn(2+), and Ni(2+). The GVE2 HNH endonuclease activity was inhibited by NaCl. This study provides the basis for determining the role of this endonuclease in life cycle of the bacteriophage GVE2 and suggests the potential application of the enzyme in molecular biology and biotechnology.

[The role of third-party tolerogenic dendritic cells in the prevention of acute graft-versus-host-disease following allogeneic bone marrow transplantation in mice].

OBJECTIVE: To explore the biological characteristic of third-party-derived tolerogenic DC(tDC) and the influence of third-party-derived tDC on acute graft-versus-host-disease (aGVHD) following allogeneic bone marrow transplantation (allo-BMT) in mice. METHODS: tDC from bone marrow cells of D1 mice was cultured with low doses of GM-CSF, IL-10 and TGF-ß1D1. The phenotype, expression of cytokines and function associated molecules were identified with FACS and RT-PCR. Mixed lymphocyte reaction was applied to analyze the influence of third-party-derived tDC on allo-CD4(+)T cells proliferation in vitro. Different doses of D1-tDC were adoptive transferred in the aGVHD model in allogeneic BMT which B6 mice as donors and D2 mice as recipients. Survival time, clinical GVHD score and the levels of Th1/2 cytokines in serum were monitored after allo-BMT using the aGVHD model as control. RESULTS: tDC expressed lower levels of MHC II and co-stimulatory molecules, such as CD80, CD86 and CD40, even when stimulated by LPS. The results by RT-PCR indicated that tDC expressed low levels of IL-12p40 and high levels of immunosuppressive molecules, such as IL-10, TGF-ß, Fas Ligand, indoleamine 2, 3-dioxygenase (IDO) and arginase. In the allogeneic MLR, third-party tDC suppressed allo-CD4(+)T cells proliferation, which was relative to the dose of tDC. In the B6→D2 mouse model, all aGVHD mice died within 18 days. Remarkably, if 10(4) third-party tDC were transferred, 60% mice survived at least 60 days. When the doses of tDC were reduced to 10(3) cells, only 20% of mice survived day 60, and when increased tDC to 10(5), all of the mice died within day 37 after allo-BMT. The cytokine levels in serum indicated that 10(4) tDC-treated mice secreted in vivo high level of IL-10 21d after BMT (P < 0.05), the levels of IL-10 in 10(3), 10(4) and 10(5) tDC-treated mice were (114.23 ± 7.78), (646.18 ± 212.02), (121.97 ± 10.47) ng/L, respectively. CONCLUSION: Third-party tDC could suppress allo-CD4(+)T cells proliferation in vitro and prevent aGVHD in allogeneic BMT mode, which may be mediated by modulating tolerogenic cytokines secretion, such as IL-10. And this effect was associated with the dose of tDC. Adoptive therapy by transfusing third-party tDC cultured with low doses of GM-CSF, IL-10 and TGF-ß1 could significantly prolong the survival of recipients and prevent aGVHD in allogeneic BMT.

Photochemically inactivated hepatitis B virus promotes upregulation of Th1-type cytokines.

Photochemical virus inactivation technology is widely used to improve the safety of blood products. However, the process by which this inactivation occurs and the resulting immunogenicity of treated viruses remain to be elucidated. This study aimed to explore the effects of two photochemical inactivation methods (methylene and riboflavin, MP and RP) on hepatitis B virus (HBV) immunogenicity. Inactivated HBV were incubated with PBMC from six healthy donors. Culture supernatants were collected at 0, 24 and 72 h for the analysis of HBsAg and HBeAg expression using ELISA. Cytokine expression was analyzed at 72 h using ELISA. Costimulatory and cell adhesion molecule mRNA expression was analyzed at 24 h by RT-PCR. No significant changes in HBsAg and HBeAg were detected following MP. However, the secretion of TNF-α and IFN-γ was upregulated. Expression of CD80, CD86, ICAM2 and LFA3 mRNA was also upregulated. In contrast, although RP did not significantly alter HBsAg expression, a reduction in HBeAg expression was observed. Furthermore, no upregulation of cytokines and intracellular molecule expression was observed following RP. These data indicate that the immunogenicity of HBV is retained following MP, and the inactivation of HBV could upregulate the Th1-type cellular immune responses, which may play significant roles in the antiviral process.

Amelioration of acute graft-versus-host disease by adoptive transfer of ex vivo expanded human cord blood CD4+CD25+ forkhead box protein 3+ regulatory T cells is associated with the polarization of Treg/Th17 balance in a mouse model.

BACKGROUND: Human cord blood (CB) is a superior source of regulatory T cells (Tregs) compared with peripheral blood. Initial studies have shown that CB-derived Tregs can be effectively expanded ex vivo. However, in vitro suppressor activity of expanded CB-Tregs and their efficacy in the prevention of acute graft-versus-host disease (aGVHD) in vivo are poorly understood. STUDY DESIGN AND METHODS: In vitro, human CB CD4+CD25+ T cells expanded with anti-CD3/CD28 beads plus interleukin (IL)-2 and the phenotypes, expression of cytokines, and suppression of expanded cells were analyzed after two cycles of stimulation. In vivo, the addition of human CB-Tregs was transferred in the major histocompatibility complex-mismatched aGVHD mouse model. Survival, body weight, GVHD scoring, histopathologic specimens, serum cytokines, and Th subsets were analyzed in CB-Treg-treated mice and untreated controls. RESULTS: After being expanded ex vivo, human CB-derived Tregs with potent suppressor function could meet clinical demands. Up to 85% of mice with CB-Tregs treatment survived beyond Day 63 after bone marrow transplantation; however, all aGVHD mice died within 18 days. In the serum of the CB-Treg-treated mice, the production of transforming growth factor-ß increased continuously, as opposed to IL-17, which decreased quickly. Consistent with the changes of cytokines, the percentage of mouse CD4+ forkhead box protein 3+ Tregs increased while that of Th17 cells decreased. CONCLUSION: Ex vivo expanded human CB-Tregs significantly prevented allogeneic aGVHD in vivo by modulating various cytokine secretion and polarizing the Treg/Th17 balance toward Treg, which suggests the potential use of expanded CB-Tregs as a therapeutic approach for GVHD.

Detection of nucleic acid lesions during photochemical inactivation of RNA viruses by treatment with methylene blue and light using real-time PCR.

The mechanism of bacteriophage photoinactivation by methylene blue and light (MB+L) involves genomic RNA damage. In this study, two RNA viruses, Sindbis virus (SINV) and hepatitis C virus were treated by MB+L and their nucleic acids were amplified to show that RNA lesions occurred during inactivation. During MB+L inactivation, the viral load of both viruses was significantly reduced as MB+L exposure increased. The nucleic acid amplification of treated viral RNA was inhibited in a time-dependent manner and the percentage inhibition of amplification reached about 99% after 30 min of treatment. Furthermore, as compared to SINV viral infectivity detected by quantification of the 50% tissue culture infective dose (TCID(50)), the inhibition of SINV RNA amplification strongly correlated with a decrease in in vitro infectivity (R(2) > 0.94), suggesting that RNA serves as the main target during MB+L inactivation.

Exosomal-like vesicles with immune-modulatory features are present in human plasma and can induce CD4+ T-cell apoptosis in vitro.

BACKGROUND: Exosomes are small membrane vesicles that are secreted from many cell types into various body fluids. These vesicles are thought to play a role in cell-cell interactions. STUDY DESIGN AND METHODS: Vesicles were isolated from human plasma of healthy donors by differential ultracentrifugation and ultrafiltration. The vesicles were identified by transmission electron microscopy, and their biochemical characteristics were analyzed by Western blot and flow cytometry. The immune-modulatory ability of exosomal-like vesicles was examined by incubating them with CD4+ T cells for CD4+ T-cell proliferation and apoptosis assays in vitro. RESULTS: Vesicles purified from human plasma displayed shapes and sizes similar to those of previously described exosomes and contained exosomes marker proteins CD63 and CD81. They also expressed molecules such as MHC Class II molecules, CD80, CD86, and the cell signal transduction molecules Wnt3a, Wnt5a, and FasL. Furthermore, functional analysis showed that allogeneic plasma exosomes restrained the survival of CD4+ T cells. Plasma exosomes can induce dose-dependent suppression of proliferation of activated CD4+ T cells, with the strongest responses induced by 500 µg/mL exosomes in vitro. Antibodies against exosomes FasL can block the activity of exosomes on CD4+ T-cell apoptosis. Moreover, three different concentrations of CD4+ T cells were inhibited by plasma exosomes and the suppressive function was not dependent on interleukin-2. CONCLUSION: Exosomes present in human plasma contain immunity-associated molecules and can induce CD4+ T-cell apoptosis in vitro. Plasma exosomes have the capacity to influence immune responses.

Adoptive therapy by transfusing expanded donor murine natural killer T cells can suppress acute graft-versus-host disease in allogeneic bone marrow transplantation.

BACKGROUND: Invariant natural killer T cells (iNKT cells) may suppress graft-versus-host disease (GVHD) after allogeneic transplantation. The purpose of this study was to investigate the therapeutic potential of iNKT cells from major histocompatibility complex (MHC)-mismatched donors for preventing GVHD after allogeneic bone marrow transplantation (BMT). STUDY DESIGN AND METHODS: In vitro, mouse iNKT cells were expanded with alpha-galactosylceramide and interleukin (IL)-2 treatment. In the NKT-treated group, lethally irradiated DBA/2(H-2K(d)) mice were adoptively transferred with expanded iNKT, bone marrow (BM), and spleen cells (SCs) from C57BL/6 (H-2K(b)) mice. Recipients in the control group were transferred only BM and SCs. The two groups were compared in survival, weight, histopathologic specimens, and serum cytokine analysis. RESULTS: In the iNKT-treated group, 80% of mice survived past Day 60 after BMT, but all died within 38 days in the control group. The mice treated with iNKT did not exhibit signs of GVHD after Day 42 except for a change in fur color. There were higher IL-4 levels by Day 7 in serum of mice that received iNKT compared to those without iNKT treatment, while the interferon-gamma levels showed no significant difference between two groups. Levels of IL-2 and IL-5 increased by Day 21 only in iNKT-treated mice. CONCLUSION: The results suggest that donor iNKT cells could alleviate GVHD symptoms and prolong survival after MHC-mismatched allogeneic BMT, which may be associated with the maintenance in IL-4 levels. These findings indicate that the therapy based on iNKT cells from MHC-mismatched donors has great potential in protection against GVHD after allogeneic hematopoietic stem cell transplantation.

[Using real-time PCR to evaluate the effect of viral inactivation by Methylene Blue with visible light].

To investigate the feasibility of using Real-Time PCR to evaluate the effectiveness of Sindbis virus inactivation by Methylene Blue with visible light. Sindbis virus was treated by Methylene Blue with different intensity of visible light and the transcribed cDNA was quantified by Real-Time PCR. Residual infectivity of treated virus was tested by cell infection method as parallel control at the same time. The residual infectivity of virus decreased from 6.50 lgTCID50/mL to under the limit of detection as light intensity increased. Meanwhile, the quantity of virus cDNA decreased significantly (P < 0.05), which correlated to the decline of virus infectivity (R2 > 0.98). Methylene Blue with visible light could cause lesion to nucleic acid of Sindbis virus, the extent of which was light intensity-dependent and correlated to the decrease of virus infectivity. The results demonstrated that Real-Time PCR can be a useful tool for evaluating effect of virus inactivation after Methylene Blue treatment with light.

TAP1 and TAP2 polymorphisms associated with ankylosing spondylitis in genetically homogenous Chinese Han population.

Human leukocyte antigen (HLA)-B27 is strongly associated with the autoimmune disease ankylosing spondylitis (AS). Other autoimmune disease-associated genes, such as transporter associated with antigen processing (TAP) genes, could also influence AS susceptibility. In this study, we investigated the association of TAP1 and TAP2 polymorphisms in genetically homogenous Chinese AS patients. Six TAP1 single nucleotide polymorphisms (SNPs) and three TAP2 SNPs sites were analyzed in B27-positive AS cases, healthy B27-negative controls, and healthy B27-positive controls. In the allele and genotype analysis, the results indicated that TAP1 site 1910 allele G, genotype AG and TAP2 site 1693 genotype AA were associated with increased AS risk in a case-B27 negative control (p < 0.05). In the haplotype analysis, TAP1 SNP haplotype (GGGGGG, TAP1*020101) and TAP1-TAP2 SNP haplotypes (GGGGGG-GGG, TAP1*020101-TAP2*0101, and GGAAGG-GAG, TAP1*0101-TAP2*0102) increased AS risk in case-B27 negative control (p < 0.05). In contrast, TAP1-TAP2 SNP haplotype GGGGGG-GAG (TAP1*020101-TAP2*0102) was less common in cases than in B27-negative controls (p < 0.05). Moreover, TAP1-TAP2 SNP haplotype GGGAGG-GGG (TAP1*0301-TAP2*0101) was less common in cases than in B27-positive controls. The two haplotypes appeared to confer protection in AS (p < 0.05). These results suggest a potential mechanism of altered antigen-peptide selection and transport in AS pathogenesis.

Inactivation of lymphocytes in blood products using riboflavin photochemical treatment with visible light.

Immunocompetent lymphocytes present in blood products are an important cause of immune reactions following blood transfusion such as transfusion-associated graft-versus-host disease (TA-GVHD). In this study the effects of riboflavin photochemical treatment (RPT) on lymphocyte proliferation and cytokine production in vitro were measured to establish whether RPT of blood products can be used to prevent immune reactions resulting from transfused lymphocytes. Lymphocytes and riboflavin were added together in medical PVC transparent bags and then exposed to visible light. Control lymphocytes were exposed to light in the absence of riboflavin. Lymphocytes exposed to riboflavin photochemical treatment (RPT-lymphocytes) and control lymphocytes were tested for the proliferative ability and the production of several cytokines upon stimulation with antigens. Upon stimulation by phytohemagglutinin (PHA) the proliferation of RPT-lymphocytes was inhibited. Using flow cytometry it was shown that RPT-lymphocytes were unable to enter the S-phase of the cell cycle following PHA stimulation. The level of cytokines present in the supernatant of RPT-lymphocytes after stimulation by antigens was significantly lower than those present in the corresponding supernatant of control lymphocytes. RPT with visible light inactivates lymphocytes, inhibits lymphocyte proliferation and the production of cytokines. It appears to be a promising method to prevent immune reactions such as TA-GVHD.

HLA-A, -B, -DR haplotype frequencies from DNA typing data of 26,266 Chinese bone marrow donors.

HLA phenotypes of 26,266 Chinese individuals who were recruited as potential hematopoietic stem cell donors by the Shanghai Red Cross Marrow Donor Registry, part of the China Marrow Donor Program, were determined for HLA-A, -B, and -DRB1 alleles at low to intermediate resolution using DNA-based typing methods. The large sample size of the study allowed accurate calculation of the Chinese HLA haplotype frequencies. The observed alleles correspond to 19 HLA-A, 44 -B, and 13 -DR split antigens. The serologic equivalents of HLA-A36, -A80, -B78, and -DR18 alleles were not observed. A total of 2,241 distinct HLA-A, -B, -DRB1 haplotypes were identified. Three-locus haplotype frequency was estimated using the maximum likelihood method. The lowest haplotype frequency that can be reliably estimated at a 95% confidence level was 0.000057. Using this cutoff value, 1,220 haplotypes (54%) were statistically reliable and their cumulative haplotype frequency was 0.9730. The cumulative haplotype frequency of the remaining 1,021 haplotypes (46%) was 0.0270. A regression equation of p = 0.192 log N - 0.576 was derived to estimate the probability (p) of finding an HLA-A, -B, -DR split antigens-matched donor in a pool of N Chinese donors.

Impact of transfection with total RNA of K562 cells upon antigen presenting, maturation, and function of human dendritic cells from peripheral blood mononuclear cells.

BACKGROUND: Vaccination of dendritic cells (DCs) with tumor antigens is a potential strategy to induce tumor-specific immunity in tumor-bearing patients. The purpose of this study was to investigate whether human monocyte-derived DCs were able to present P210(Bcr-Ab1) protein and induce antigen-specific cytotoxic T lymphocyte (CTL) responses in vitro after transfected with total RNA of K562 cells (K562-RNA). STUDY DESIGN AND METHODS: DCs derived from human peripheral blood mononuclear cells were transfected with K562-RNA with electroporation or DOTAP lipofection. The successful transfection was determined by reverse transcription-polymerase chain reaction and Western blot. The phenotypes of the DCs were analyzed by flow cytometry (FCM), and cytotoxicity of CTL was assessed by propidium iodide staining followed by FCM analysis. The CD1a expression and purity of DCs were measured by FCM. RESULTS: The Bcr-Abl fusion gene was detected in the DCs with 24 hours after the transfection. The transfected cell expressed increased levels of CD80, CD83, CD86, and HLA-DR. Moreover, the transfected DCs strongly stimulated the T lymphocytes to gain cytotoxic activity against K562 cells. Culture medium containing 1 percent human plasma was the most effective for DC growth. CONCLUSION: Human DCs transfected with K562-RNA effectively induce specific immune responses. This method can be used to induce tumor-specific immune response and may have potential application in immunotherapy of tumors.