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Hao H.,Northwest University, China | Zhen Y.,Northwest University, China | Wang Z.,Peking University | Chen F.,Northwest University, China | And 2 more authors.
Cell Biology International | Year: 2013

Colon cancer is a type of malignant tumor that causes considerable mortality worldwide. Epithelial cellular adhesion molecule (EpCAM), a tumor-associated antigen of colon tumors, is a target for colon cancer therapy. EpCAM-specific monoclonal antibodies (mAbs) have been applied in human colon cancer since the 1990s; however, the therapeutic effects are limited. EpCAM activates nuclear signaling pathways by releasing its intracellular domain (EpICD). The released EpICD stimulates the Wnt/b-catenin signaling pathway, which is also strongly associated with tumorigenesis. EpCAM is also a target gene of the Wnt/ b-catenin signaling pathway. EpCAM and the Wnt/b-catenin signaling pathway form a functional interaction cycle in colon cancer. Thus, we propose a new therapeutic drug for colon cancer: an EpCAM single-chain fragment variable antibody (scFv)- truncated protamine-siRNA. EpCAM scFv can recognize and bind colon cancer cells through its EpCAM antigen activity. Furthermore, the specific siRNA transferred into colon cancer cells specifically inhibits Wnt/b-catenin signal transmission. Therefore, this new drug may efficiently interrupt the functional cycle between EpCAM and Wnt/b-catenin signaling and be an effective therapeutic strategy for colon cancer. © 2013 International Federation for Cell Biology.


Xu Z.,Institute of Integrated Medical Information | Chen H.,Institute of Integrated Medical Information | Wang X.,Xi'an Jiaotong University | Zhang P.,Institute of Integrated Medical Information | And 2 more authors.
Monoclonal Antibodies in Immunodiagnosis and Immunotherapy | Year: 2014

PMEL, also known as Pmel17 or gp100, is a melanocyte-specific glycoprotein that is essential for the formation of stage II melanosomes. As it has a highly restricted expression pattern in normal tissues and a transient presence on the cell surface, PMEL is believed to be a potential target for antibody drug conjugate therapy in some pigmentary diseases. The production of a high specificity and high affinity monoclonal antibody against human PMEL was helpful for the antibody drug conjugate therapy study. In the present study, monoclonal antibodies (MAbs) against PMEL were obtained by immunizing BALB/c mice with the recombinant PMEL-GST fusion protein. Three mAbs (A3F, G11B, and J7E) with a titer of 1:6000, 1:10,000, and 1:3000, respectively, were obtained. Immunoglobulin subclass assay revealed that A3F was IgG2b, G11B was IgG1, and J7E was IgG2a. Specificity analysis by Western blotting demonstrated that A3F and J7E cross-reacted with GPNMB or LAMP; however, G11B reacted with PMEL only. Immunohistochemistry experiments showed that G11B could bind human PMEL antigen in normal skin. Flow cytometry assay demonstrated that G11B could bind to the surface of PMEL positive melanoma cells but not PMEL negative cells. Taken together, these results show that this G11B provides a useful tool for the antibody drug conjugate therapy study in some pigmentary diseases. © Copyright 2014, Mary Ann Liebert, Inc. 2014.


Sun L.,Shaanxi Provincial Peoples Hospital | Pan S.,Huanghuai University | Yang Y.,Centers for Disease Control and Prevention | Sun J.,Shaanxi Provincial Peoples Hospital | And 5 more authors.
Experimental Biology and Medicine | Year: 2016

Toll-like receptors play essential roles in the modulation of melanogenesis, which has been implicated in the pathogenesis of hyper- or hypopigmentation-related diseases. However, little is currently known regarding the role of TLR9 in human melanocytes. TLR9 recognizes unmethylated cytosine-phosphate-guanine motif-containing oligodeoxynucleotides, and cytosine-phosphate-guanine ODN2006 acts as an hTLR9 agonist. The aim of the present study was to investigate the effect of cytosine-phosphate-guanine ODN2006 on melanogenesis in the human melanocyte cells. MTT assay and enzyme-linked immunosorbent assay indicated that ODN2006 stimulation (0, 1, 5, 10 µM) dose-dependently reduced cell viability and promoted the production of TNF-α, IL-6, and IL-8 in PIG1 melanocytes. The mRNA and protein levels of PMEL and TYRosinase were elevated at 6 h, and then decreased 24 h later, but were significantly augmented 72 h later following ODN2006 stimulation; whereas, TLR9 expressions were time-dependently increased in PIG1 melanocytes. Moreover, ultraviolet B irradiation combined with ODN2006 stimulation induced much more significant enhancement of PMEL, TYRosinase, and TLR9 mRNA and protein after three days in PIG1 melanocytes, and the similar results were obtained using the primary human melanocytes. The expression of TLR9 protein was down-regulated by TLR9 siRNA transfection. ODN2006 had an additive effect on ultraviolet B-induced melanogenesis and PMEL expression, as well as NF-κB activation, which could be blocked by TLR9 knockdown, the NF-κB specific inhibitor PDTC, or the TBK1 inhibitor BX795. Collectively, we concluded that TLR9 regulates melanogenesis through NF-κB activation, suggesting that TLR9 may play a role in microbial-induced melanogenesis. © 2016, © 2016 by the Society for Experimental Biology and Medicine.


Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | Wang C.,The Tumor Research Institute of Jilin Province | Xie Y.,Northwest University, China | And 7 more authors.
Journal of Immunological Methods | Year: 2013

CD306, also known as soluble leukocyte-associated immunoglobulin-like receptor-2 (LAIR-2), is a member of an immunoglobulin superfamily with the shared characteristic of an immunoglobulin-like C2-type domain. CD306 is speculated to be secretory and has 84% similarity with the extracellular domain of CD305, which binds to the same ligands as CD306. However, data on its distribution are absent due to the lack of an efficient method to detect it. In this study, we successfully cloned the cDNA of CD306 from the peripheral blood mononuclear cells (PBMCs) of patients with hemorrhagic fever with renal syndrome. The fusion proteins were expressed and purified, and three strains of monoclonal antibodies (mAbs) against CD306 were prepared and characterized. The sandwich ELISA for detecting CD306 was established and optimized with sensitivity up to 15. pg/ml, and the assay showed high specificity for the detection of CD306. With this method, a right skewed frequency distribution of CD306 in the sera of healthy subjects was determined. The concentrations of CD306 in sera and urine were detected in patients with different diseases. Aberrantly high levels of CD306 were found in the sera of pregnant women and patients with inflammation and rheumatic heart disease and in the urine of pregnant women. Meanwhile, there was a positive correlation between CD306 and soluble CD305 expression and secretion levels in sera and urine samples from patients, and both proteins inhibited CD305-mediated immunosuppressive functions. Our results demonstrate that CD306 represents a potentially useful predictor for disease diagnosis and that the method developed has potential for clinical application. © 2013 Elsevier B.V.


Bai J.,Xi'an Jiaotong University | Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | Lei Y.,Xi'an Jiaotong University | And 3 more authors.
Cell Biology International | Year: 2014

Efforts to get a strong and sustained anti-tumour immune response induced by a tumour specific antigen have failed, but sipuleucel-T has been approved by the US Food and Drug Administration (FDA).We noticed that exosomes secreted by tumour cells or immune cells may be crucially involved in the tumour immune response, whereas others have had inconsistent findings on exosome involvement. Based on immune network theory, we summarise research advances of exosomes and speculate that in the tumour immune response exosomes follow the immune response curve hypothesis. Exosomes activate simultabeously both immune activation and immune tolerance, but at different intensities. To obtain a desired anti-immune response, the initial point of immunity should be determined to achieve the strongest anti-tumour response, and repeated in vitro to extend and enhance this response. As a result, our hypothesis proposes that studies should now be directed at determining the exact time of exosome activity in maintaining a viable anti-tumour immune response in vivo. © 2013 International Federation for Cell Biology.


Bai J.,Xi'an Jiaotong University | Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | Lei Y.,Xi'an Jiaotong University | And 3 more authors.
Molecular Medicine Reports | Year: 2014

Malignant melanoma has the highest risk of mortality among all types of skin cancer due to its highly metastatic potential. The ocular albinism type 1 (OA1) protein is a pigment cell-specific glycoprotein, which shares significant structural and functional features with G protein-coupled receptors. However, the role of OA1 in melanoma has yet to be elucidated. The present study aimed to investigate whether OA1 is involved in melanoma cell migration. OA1 was found to stimulate cell migration in a dose-dependent manner in cultured human melanoma cells. Furthermore, knockdown of OA1 using small interfering RNA was observed to significantly inhibit melanoma cell migration. In addition, the mechanism underlying OA1-induced melanoma cell migration was investigated. Stimulation of the RAS/RAF/mitogen activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway using growth factors enhanced OA1 expression and melanoma cell migration, whereas inhibition of this pathway using U0126 was observed to markedly decrease OA1 expression and the number of migrated cells. These findings indicate that OA1 is involved in melanoma cell migration and that OA1-induced melanoma cell migration is mediated through the RAS/RAF/MEK/ERK signaling pathway. Therefore, OA1 may serve as a novel therapeutic target for melanoma.


Guo C.,Xi'an Jiaotong University | Guo C.,Key Laboratory of Infection and Immunity Disease of Shaanxi Province | Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | And 18 more authors.
Experimental and Molecular Pathology | Year: 2015

Influenza A virus infection is a persistent threat to public health worldwide due to hemagglutinin (HA) variation. Current vaccines against influenza A virus provide immunity to viral isolates similar to vaccine strains. Antibodies against common epitopes provide immunity to diverse influenza virus strains and protect against future pandemic influenza. Therefore, it is vital to analyze common HA antigenic epitopes of influenza virus. In this study, 14 strains of monoclonal antibodies with high sensitivity to common epitopes of influenza virus antigens identified in our previous study were selected as the tool to predict common HA epitopes. The common HA antigenic epitopes were divided into four categories by ELISA blocking experiments, and separately, into three categories according to the preliminary results of computer simulation. Comparison between the results of computer simulations and ELISA blocking experiments indicated that at least two classes of common epitopes are present in influenza virus HA. This study provides experimental data for improving the prediction of HA epitopes of influenza virus (H1 subtype) and the development of a potential universal vaccine as well as a novel approach for the prediction of epitopes on other pathogenic microorganisms. © 2014 Elsevier Inc.


Zheng J.,General Hospital of Guangzhou Military Command | Zhou J.,General Hospital of Guangzhou Military Command | Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | And 4 more authors.
DNA and Cell Biology | Year: 2014

Few studies have referred to the implication of anoikis processes following hormonal treatment. No data are available on the influence of estrogen in ovarian cancer anoikis. To gain insights into the effects and mechanism of estrogen in ovarian cancer cells, we have carried out studies on the anoikis of ovarian cancer cells treated with estrogen and on the pathways involved. We observed an anti-anoikis role of E2 in suspended Caov-3 cells, and this was mainly due to the decreasing of Bit1 level in cytosol. We also found that estrogen receptor α (ERα) was the main mediator involved in this process. To study the signaling pathways well, phosphatidylinositol 3-kinase (PI3K)/AKT were further investigated. Results demonstrated that the decreasing of the Bit1 level in cytosol mediated by E2 binding to ERα was mainly through PI3K/AKT pathways. Overall, these findings disclose a new perspective for estrogen on ovarian cancer therapy. © Mary Ann Liebert, Inc. 2014.


Xie X.,Northwest University, China | Xie X.,Institute of Integrated Medical Information | Wu X.,Shanghai Tenth Peoples Hospital | Cui J.,Northwest University, China | And 2 more authors.
Brain Research | Year: 2013

Subarachnoid hemorrhage (SAH) is a frequent occurrence in cerebrovascular accidents, and inflammation occurs in the subarachnoid space after SAH. Arachnoid cells have the capability to present antigens and active T-lymphocytes after stimulation by cerebrospinal fluid (CSF). However, the effect of CSF on T-lymphocytes and arachnoid cell adhesion was not clearly understood. In this study, we used ELISA to detected tumor necrosis factor-α (TNF-α) content in CSF of SAH patients. CSF or recombinant TNF-α were applied on arachnoid cells and T-lymphoctes, and RT-PCR and western blotting were performed to determine the expression of intercellular adhesion molecule-1 (ICAM-1) in arachnoid cells and Lymphocyte Function-Associated Antigen-1 (LFA-1) in T-lymphocytes, respectively. Meanwhile, the Matrix Metal Proteinase-9 (MMP-9) expression in these cells was also determined. We found that the content of TNF-α in the CSF was significantly increased in the CSF of SAH patients (from 22±8 pg/mL of healthy people to 436-450 pg/mL of SAH patients). Treatement with CSF could increase the expression of ICAM-1in arachnoid cells and that of LFA-1 in T-lymphocytes, mainly through the increased levels of TNF-α. We also found that the co-culture of arachnoid cells and T-lymphocytes increased the expression of MMP-9 in both cells through the interaction of ICAM-1 of and LFA-1. All of these results suggested that arachnoid cells are involved in the T-lymphocytes invasion in the subarachnoid space after SAH. © 2013 Elsevier B.V.

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