Wenqi Dong

Acute myeloid leukemia (AML) is a deadly hematologic malignancy. Studies have shown that miroRNAs (miRNAs), evolutionarily conserved small noncoding RNAs, contribute to AML disease development, suggesting that therapeutical manipulation of miRNAs may benefit disease treatment in AML. In this study, we found miR-361-3p was significantly upregulated in the blood of AML patients, whereas the expression of its direct target, B-cell translocation gene (BTG2, a tumor suppressor), was markedly lower compared with the expression in the blood of healthy controls. Inhibition of miR-361-3p significantly increased BTG2, leading to inhibition of cell proliferation and promotion of cell apoptosis in AML cell line, HL-60. In contrast, increasing of miR-361-3p expression exerted opposite effects to miR-361-3p inhibitor on BTG2 expression, cell proliferation and apoptosis. We further showed 9s-Hydroxyoctadecadienoic acid (9s-HODE), a major active derivative of linoleic acid, exerted an anti-AML effect through, at least partly, targeting miR-361-3p/BTG2 axis and its associated pathways. These findings not just demonstrate an oncogenic role of miR-361-3p in AML but also provide novel insights to develop a potential therapeutic for AML treatment by manipulating its expressions.

Cytoglobin (Cygb) is a globin molecule that is ubiquitously expressed in all tissues and has a protective role under oxidative stress. It has also been demonstrated to be effective in the treatment of alcoholic fatty liver disease (AFLD). In order to study the molecular mechanisms underlying its beneficial effects for the treatment of alcoholic liver, two‑dimensional electrophoresis and mass spectrometric analysis were performed on serum and liver tissues from an in vivo rat model of AFLD. A total of 26 differentially expressed proteins were identified in the serum and 20 differentially expressed proteins were identified in liver specimens. Using online bioinformatics tools, it was indicated that these differentially expressed proteins were primarily associated with pathways including binding and uptake of ligands by scavenger receptors, response to corticosteroid, plasma lipoprotein remodeling, regulation of complement cascade, hydrogen peroxide catabolic process, as well as response to nutrient and monosaccharide. The present results suggested that recombinant human Cygb exerts its role in the treatment of AFLD primarily through affecting nutrient metabolism, monocarboxylic acid biosynthesis, regulation of glutathione expression, plasma lipoprotein remodeling and removal of metabolic waste from the blood.

Objective To construct eukaryotic expression vectors of human IgE heavy chain 2-4 region (IgECepsilon2-4) and purify the recombinant protein, and then capture its interacted proteins by surface plasmon resonance (SPR). Methods Three recombinant eukaryotic expression vectors of IgECepsilon2-4 containing different signal peptides were constructed and transiently transfected into HEK293FT suspension cells separately. The recombinant plasmid with the highest-level expression was selected to express the recombinant protein in a huge amount, and then the recombinant protein was purified by Ni-NTA affinity chromatography. The interaction between high-affinity IgE receptor (FcepsilonR I) of KU812 cell surface and IgECepsilon2-4 was identified by immunofluorescence cytochemistry. The unknown proteins that specifically interacted with IgECepsilon2-4 were captured from human serum by SPR technique. Results The recombinant plasmid containing the signal peptide III showed the highest expression (6.2 mg/L). Highly purified recombinant protein IgECepsilon2-4 was obtained by affinity purification. Immunofluorescence cytochemistry showed that the recombinant protein IgECepsilon2-4 could be combined with the surface receptor of KU812 cells. Thirty-nine kinds of proteins which were likely to interact with IgECepsilon2-4 were captured from human serum by SPR. Conclusion We obtained the purified recombinant protein IgECepsilon2-4 that could be combined with KU812 cell surface receptor. Target fishing experiment revealed that the recombinant protein IgECepsilon2-4 was likely to interact with 39 kinds of proteins in human serum.

Atherosclerosis is a chronic inflammatory vascular disease characterized by lipid accumulation and endothelial dysfunction. Cytoglobin has been shown to exert protective effects under oxidative stress conditions. The aim of this study was to determine whether recombinant human cytoglobin (rhCYGB) has protective effects against atherosclerosis. We intraperitoneally injected rhCYGB (0, 4, or 7 mg/kg BW) into the atherosclerotic rats daily for 60 days. The rhCYGB injections reduced low-density lipoprotein cholesterol (LDL-C) levels and increased high-density lipoprotein cholesterol levels in a dose-dependent manner, rhCYGB (7 mg/kg) significantly attenuated atherosclerosis. Blood proteins were separated by 2-dimensional electrophoresis and analyzed by mass spectrometry, and the majority of the proteins in question were participated in oxidative stress pathways and cardiovascular diseases. Human hepatocellular liver carcinoma cell line (HepG2) cells were treated with oleic acid (0.3 mmol/L), and Human acute monocytic leukemia cell line (THP-1) cells were incubated with oxidized LDL (ox-LDL; 50 µg/mL) to induce foam cell (FC) formation in vitro. Treatment with different concentrations of rhCYGB (0, 5, 10, and 15 μg/mL) significantly decreased the lipid droplet levels in HepG2 cells and cholesterol ester levels in FCs. Moreover, rhCYGB significantly increased superoxide dismutase and glutathione peroxidase activity and decreased malondialdehyde and nicotinamide adenine dinucleotide phosphate oxidase activity in cells. In addition, rhCYGB decreased NO and Reactive oxygen species (ROS) levels in FCs by functioning as an NO dioxygenase enzyme and ROS scavenger. Taken together, our findings indicate that rhCYGB prevented atherosclerosis by regulating lipid metabolism and oxidative stress. Our study provides insights into the possible usefulness of rhCYGB as an antiatherosclerosis agent.

9s-hydroxy-octadecadienoic acid (9S-HOD), one of the natural products of linoleic acid oxygenation by 15-lipoxygenase (15-LOX), has been found to have anti-tumor properties in vitro and in vivo. The present study therefore investigated whether 9S-HOD affects acute leukemia HL-60 cells. The cytotoxicity of 9S-HOD in HL-60 with or without the presence of fetal bovine serum (FBS) in the culture media was tested using cell viability assays and flow cytometry. To explore the mechanism of its anti-tumor activity by 9S-HOD, we used a proteomic analysis to identify HL-60 cells protein profiles, based on two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) identification. 9S-HOD exerted cytotoxicity efficacy and induced apoptosis in HL-60 cells, and the cytotoxicity was largely attenuated by the presence of FBS in culture media. The proteomic results revealed that 9S-HOD remarkably altered the abundance of 23 proteins that were involved in mRNA metabolic process, protein binding, DNA replication and apoptosis. Our results indicated that 9S-HOD exerts cytotoxicity in HL-60 cells by affecting several pathways.

Cytoglobin (CYGB), a member of the globin family, is thought to protect cells from reactive oxygen and nitrogen species and deal with hypoxic conditions and oxidative stress. However, its molecular mechanisms of action are not clearly understood. Through immunoprecipitation combined with a two-dimensional electrophoresis–mass spectrometry assay, we identified a CYGB interactor: aldo-keto reductase family 7 member A2 (AKR7A2). The interaction was further confirmed using yeast two-hybrid and co-immunoprecipitation assays. Our results show that AKR7A2 physically interacts with CYGB.

This study aims to investigate whether the use of recombinant human cytoglobin (rhCygb) impact on hepatic fibrogenesis caused by CCl₄. SD (n = 150) rats were randomly divided into three groups of normal, CCl₄model and rhCygb groups. After model establishment, rats in rhCygb groups were administered daily with rhCygb (2 mg/kg, s.c.). Histological lesions were staged according to metavir. Serum parameters including ALT, AST, HA, LN, Col III and Col IV were determined. The liver proteins were separated by 2-DE and identified. As a result, the stage of hepatic damage and liver fibrosis in rhCygb groups were significantly milder than that in CCl₄model groups. Meanwhile, rhCygb dramatically reversed serum levels of ALT and AST, and also markedly decreased the liver fibrosis markers levels of LN, HA, Col III and Col IV. In 2-DE, 33 proteins among three groups with the same changing tendency in normal and rhCygb treated groups compared with CCl₄model group were identified. GO analysis showed that several identified proteins involved in oxidative stress pathway. The study provides new insights and data for administration of rhCygb reversing CCl₄-induced liver fibrosis suggesting that rhCygb might be used in the treatment of liver fibrosis.

A sensitive, rapid and novel measurement method for cytokeratin 19 fragment (CYFRA 21-1) in human serum by magnetic particle-based time-resolved fluoroimmunoassay (TRFIA) is described. Built on a sandwich-type immunoassay format, analytes in samples were captured by one monoclonal antibody coating onto the surface of magnetic beads and "sandwiched" by another monoclonal antibody labeled with europium chelates. The coefficient variations of the method were lower than 7%, and the recoveries were in the range of 90-110% for serum samples. The lower limit of quantitation of the present method for CYFRA 21-1 was 0.78 ng/ml. The correlation coefficient of CYFRA 21-1 values obtained by our novel TRFIA and CLIA was 0.980. The present novel TRFIA demonstrated high sensitivity, wider effective detection range and excellent reproducibility for determination of CYFRA 21-1 can be useful for early screening and prognosis evaluation of patients with non-small cell lung cancer.

Angiostrongylus cantonensis (A. cantonensis) infection causes eosinophilic meningitis in humans. Eosinophilia and a Th2-type immune response are the crucial immune mechanisms for eosinophilic meningitis. CD4+CD25+ regulatory T cells (Treg) are involved in the pathogenesis of A. cantonensis. Diammonium glycyrrhizinate (DG) is a compound related to glycyrrhizin (GL), a triterpene glycoside extracted from liquorice root. We investigated the curative effects and probable mechanisms of therapy involving a combination of albendazole and DG in BALB/c mice infected with A. cantonensis, and compared these with therapy involving albendazole and dexamethasone. We analysed survival time, body weight, signs, eosinophil numbers, immunoglobulin E (IgE), interleukin-5 (IL-5), and eotaxin concentrations, numbers and Foxp3 expression of CD4+CD25+ Treg, worm recovery and histopathology. The present results demonstrated that the combination of albendazole and DG could increase survival time more efficiently and relieve neurological dysfunction; decrease weight loss, eosinophil numbers, concentrations of IgE, IL-5 and eotaxin, the number and expression of Foxp3 of CD4+CD25+ Treg; and improve worm recovery and histopathology changes in treated animals, compared with the combination of albendazole and dexamethasone. The observations presented here suggest that the albendazole and dexamethasone combination could be replaced by the combination of albendazole and DG.