Curcumin analog HO‐3867 triggers apoptotic pathways through activating JNK1/2 signalling in human oral squamous cell carcinoma cells

Human oral squamous cell carcinoma (OSCC) is the common head and neck malignancy in the world. While surgery, radiotherapy and chemotherapy are emerging as the standard treatment for OSCC patients, the outcome is limited to the recurrence and side effects. Therefore, patients with OSCC require alternative strategies for treatment. In this study, we aimed to explore the therapeutic effect and the mode of action of the novel curcumin analog, HO‐3867, against human OSCC cells. We analysed the cytotoxicity of HO‐3867 using MTT assay. In vitro mechanic studies were performed to determine whether MAPK pathway is involved in HO‐3867 induced cell apoptosis. As the results, we found HO‐3867 suppressed OSCC cells growth effectively. The flow cytometry data indicate that HO‐3867 induce the sub‐G1 phase. Moreover, we found that HO‐3867 induced cell apoptosis by triggering formation of activated caspase 3, caspase 8, caspase 9 and PARP. After dissecting MAPK pathway, we found HO‐3867 induced cell apoptosis via the c‐Jun N‐terminal kinase (JNK)1/2 pathway. Our results suggest that HO‐3867 is an effective anticancer agent as its induction of cell apoptosis through JNK1/2 pathway in human oral cancer cells.     

CD137-mediated pathogenesis from chronic hepatitis to hepatocellular carcinoma in hepatitis B virus-transgenic mice

Chronic hepatitis B virus (HBV) infection is characterized by sustained liver inflammation with an influx of lymphocytes, which contributes to the development of cirrhosis and hepatocellular carcinoma. The mechanisms underlying this immune-mediated hepatic pathogenesis remain ill defined. We report in this article that repetitive infusion of anti-CD137 agonist mAb in HBV-transgenic mice closely mimics this process by sequentially inducing hepatitis, fibrosis, cirrhosis, and, ultimately, liver cancer. CD137 mAb initially triggers hepatic inflammatory infiltration due to activation of nonspecific CD8(+) T cells with memory phenotype. CD8(+) T cell-derived IFN-γ plays a central role in the progression of chronic liver diseases by actively recruiting hepatic macrophages to produce fibrosis-promoting cytokines and chemokines, including TNF-α, IL-6, and MCP-1. Importantly, the natural ligand of CD137 was upregulated significantly in circulating CD14(+) monocytes in patients with chronic hepatitis B infection and closely correlated with development of liver cirrhosis. Thus, sustained CD137 stimulation may be a contributing factor for liver immunopathology in chronic HBV infection. Our studies reveal a common molecular pathway that is used to defend against viral infection but also causes chronic hepatic diseases.     

Purification and partial characterization of a procaryotic glycoprotein from the plasma membrane of Thermoplasma acidophilum

The obligate, thermophilic, acidophilic mycoplasma, Thermoplasma acidophilum, grows optimally at 56° C and pH 2.0. Its plasma membrane possessed 21–22 protein bands that were resolved by polyacrylamide gel electrophoresis. One major membrane protein, molecular weight 152 000, which stained for carbohydrate with periodic acid-Schiff reagent, accounted for 32% (w/w) of the total membrane proteins. It was isolated and further purified by concanavalin A affinity chromatography. The carbohydrate content amounted to less than 10% (w/w) compared to that of the entire glycoprotein. The carbohydrate moiety consisted mainly of mannose residues with branched α 1 → 2 linkages at the non-reducing ends of the glycopeptide as determined by permethylation followed by gas chromatography-mass spectrometry analysis. The reducing end was an N-glycosidic linkage between asparagine and N-acetylglucosamine. The amino acid composition of this glycoprotein showed 62 mol% hydrophobic residues, while the acidic amino acid content contributed 9 mol% more than that of the basic amino acids. The existence of membrane glycoproteins in the procaryotic, wall-less T. acidophilum may provide a protective coat for the plasma membrane. The stereochemistry and the conformation of the carbohydrate chains, in conjunction with water turgor, may contribute to the rigidity of the membrane and the cation binding.     

Alterations in microRNA expression linked to microcystin-LR-induced tumorigenicity in human WRL-68 Cells

Microcystin-LR (MC-LR) is a cyclic heptapeptide that acts as a potent hepatotoxin and carcinogen. However, the mechanism of its carcinogenic action remains undetermined. In this study, MC-LR was used to induce the malignant transformation of the WRL-68 cell line. Alterations in microRNA (miRNA) expression in the transformed cell were analyzed to determine the role of miRNAs in MC-LR-induced carcinogenesis. Cultured WRL-68 cells (labeled 25MC10) were continuously exposed to a low concentration (10 μg/L) of MC-LR for 25 passages. Compared with the mock-treated parental cells, the induced 25MC10 cells exhibited a higher growth rate, resistance to serum-induced terminal differentiation, and tumorigenicity in a nude mouse xenograft test. A pilot miRNA expression array analysis was conducted on the 25MC10 cells, followed by validation of select miRNAs by RT-PCR. We found that the onco-miRNAs miR-21 and miR-221 displayed upregulated expression while the liver-specific miR-122 was downregulated. These results suggest that chronic MC-LR exposure alters the miRNA expression profile of WRL-68 cells and causes phenotypic transformation. We propose that characteristic miRNA alterations could be used as molecular targets for the development of environmental water monitoring methods.     

X-ray Phase Contrast Imaging of Cell Isolation with Super-Paramagnetic Microbeads

Super-paramagnetic microbeads are widely used for cell isolation. Evaluation of the binding affinity of microbeads to cells using optical microscopy has been limited by its small scope. Here, magnetic property of microbeads was first investigated by using synchrotron radiation (SR) in-line x-ray phase contrast imaging (PCI). The cell line mouse LLC (Lewis lung carcinoma) was selected for cell adhesion studies. Targeted microbeads were prepared by attaching anti-VEGFR2 (vascular endothelial growth factor receptor-2) antibody to the shell of the microbeads. The bound microbeads were found to better adhere to LLC cells than unbound ones. PCI dynamically and clearly showed the magnetization and demagnetization of microbeads in PE-50 tube. The cells incubated with different types of microbeads were imaged by PCI, which provided clear and real-time visualization of the cell isolation. Therefore, PCI might be considered as a novel and efficient tool for further cell isolation studies.     

Identification,Expression and Activity Analyses of Five Novel Duck Beta-Defensins

In the current study, five novel avian β-defensins (AvBDs) were identified and characterized in tissues from Peking ducks (Anas platyrhynchos). The nucleotide sequences of these cDNAs comprised 198 bp, 182 bp, 201 bp, 204 bp, and 168 bp, and encoded 65, 60, 66, 67, and 55 amino acids, respectively. Homology, characterization and comparison of these genes with AvBD from other avian species confirmed that they were Apl_AvBD1, 3, 5, 6, and 16. Recombinant AvBDs were produced and purified by expressing these genes in Escherichia coli. In addition, peptides were synthesized according to the respective AvBD sequences. Investigation of the antibacterial activity of the Apl_AvBDs showed that all of them exhibited antibacterial activity against all 12 bacteria investigated (P<0.05 or P<0.01). In addition, the antibacterial activity of all of the AvBDs against M. tetragenus and P. multocida decreased significantly in the presence of 150 mM NaCl (P<0.01). None of the AvBDs showed hemolytic activity. Consistent with their broad-spectrum antibacterial activity, the five novel Apl_AvBDs inhibited replication of duck hepatitis virus (DHV) in vitro significantly (P<0.05). The mRNA expression of all five Apl_AvBD in most tissues, including immune organs and the liver, was upregulated in response to DHV infection at different time points. These findings provide evidence that these defensins activate the immune response to combat microbial infection.     

Biologically-Directed Modeling Reflects Cytolytic Clearance of SIV-Infected Cells In Vivo in Macaques

The disappointing outcomes of cellular immune-based vaccines against HIV-1 despite strong evidence for the protective role of CD8⁺ T lymphocytes (CTLs) has prompted revisiting the mechanisms of cellular immunity. Prior data from experiments examining the kinetics of Simian Immunodeficiency Virus (SIV) clearance in infected macaques with or without in vivo CD8 depletion were interpreted as refuting the concept that CTLs suppress SIV/HIV by direct killing of infected cells. Here we briefly review the biological evidence for CTL cytolytic activity in viral infections, and utilize biologically-directed modeling to assess the possibility of a killing mechanism for the antiviral effect of CTLs, taking into account the generation, proliferation, and survival of activated CD4⁺ and CD8⁺ T lymphocytes, as well as the life cycle of the virus. Our analyses of the published macaque data using these models support a killing mechanism, when one considers T lymphocyte and HIV-1 lifecycles, and factors such as the eclipse period before release of virions by infected cells, an exponential pattern of virion production by infected cells, and a variable lifespan for acutely infected cells. We conclude that for SIV/HIV pathogenesis, CTLs deserve their reputation as being cytolytic.