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ADAM17 flow cytometry

Ivana Vidlickova, Franck Dequiedt, Lenka Jelenska, Olga Sedlakova, Michal Pastorek, Stanislav Stuchlik, Jaromir Pastorek, Miriam Zatovicova, Silvia Pastorekova
BACKGROUND: Carbonic anhydrase IX (CA IX) is a tumor-associated, highly active, transmembrane carbonic anhydrase isoform regulated by hypoxia and implicated in pH control and adhesion-migration-invasion. CA IX ectodomain (ECD) is shed from the tumor cell surface to serum/plasma of patients, where it can signify cancer prognosis. We previously showed that the CA IX ECD release is mediated by disintegrin and metalloproteinase ADAM17. Here we investigated the CA IX ECD shedding in tumor cells undergoing apoptosis in response to cytotoxic drugs, including cycloheximide and doxorubicin...
2016: BMC Cancer
F Ren, J Li, X Jiang, K Xiao, D Zhang, Z Zhao, J Ai, C Hou, Y Jia, G Han, L Xie
Immune dysfunction is the main characteristic of sepsis. T cell Ig and mucin domain protein 3 (Tim-3) on the monocytes has been reported to promote immune homeostasis during sepsis, but the influences of plasm soluble Tim-3 (sTim-3) on the immune system during sepsis remain unknown. Here, 100 patients with different severities of sepsis (40 sepsis, 42 severe sepsis, and 18 septic shock) were enrolled in this study. The Tim-3 and human leukocyte antigen-DR (HLA-DR) on the circulating monocytes were detected using flow cytometry...
November 2015: Tissue Antigens
Jing Wu, Li Yin, Ning Jiang, Wen-Jie Guo, Jia-Jia Gu, Meng Chen, You-You Xia, Jian-Zhong Wu, Dan Chen, Jian-Feng Wu, De-Jun Wang, Dan Zong, Nan Zhang, Kai Ding, Teng Huang, Xia He
MiR-145 is downregulated and functions as a tumor suppressor in many malignancies. In this study, the biological function, molecular mechanism, and direct target genes of miR-145 in nasopharyngeal carcinoma (NPC) cells were investigated. Cell survival was detected by cell viability assay, and cell cycle was determined through flow cytometry. Invasion and migration of NPC cells were examined using cell invasion and wound healing assays, respectively. A disintegrin and metalloproteinase 17 (ADAM17) was verified as the target of miR-145 through luciferase reporter assay, qRT-PCR, and Western blot analysis...
November 1, 2015: Experimental Cell Research
Kensaku Okada, Hiroki Chikumi, Miyako Takata, Kosuke Yamaguchi, Haruhiko Makino, Tsuyoshi Kitaura, Masaki Nakamoto, Akira Yamasaki, Tadashi Igishi, Naoto Burioka, Eiji Shimizu
BACKGROUND: Clarithromycin is a macrolide antibiotic that possesses anti-inflammatory and immunomodulatory properties. Although recent data suggests that macrolide antibiotics enhance Pseudomonas aeruginosa clearance from the lung, involving natural killer (NK) T cells in this process by activating the NKG2D-NKG2D ligand system, the precise underlying mechanism is still unclear. In this study, we examined the effect of clarithromycin on a potent NKG2D ligand, UL16-binding protein 2 (ULBP2), in the lung and its shedding mechanism...
March 2015: Yonago Acta Medica
Jing Ren, Yunzhong Nie, Mingming Lv, Sunan Shen, Ruijing Tang, Yujun Xu, Yayi Hou, Shuli Zhao, Tingting Wang
Estrogen is involved in promoting lung cancer cell division and metastasis. MICA and MICB function as ligands for NKG2D, an important immunoreceptor expressed on natural killer (NK) cells. However, whether estrogen regulates MICA/B expression and affects tumor immune escape remains unknown. In this study, we measured the mRNA levels of MICA, MICB and ADAM17in non-small cell lung cancer (NSCLC) cell lines treated with estrogen. Surface expression of MICA/B on LTEP-a2 and A549 was detected using flow cytometry...
November 2015: Cellular & Molecular Immunology
Huaqun Zhu, Xiaolin Sun, Lei Zhu, Fanlei Hu, Lianjie Shi, Zhanguo Li, Yin Su
OBJECTIVE: To investigate the expression and clinical significance of trans-membrane MerTK (mMer) on circulating CD14+ monocytes/macrophages and soluble MerTK (sMer) levels in plasma in systemic lupus erythematosus (SLE). METHOD: 108 SLE patients and 42 healthy controls were recruited in this study. The expression of mMer on the surfaces of CD14+ monocytes/macrophages was evaluated by flow cytometry (FCM). The sMer levels were measured by ELISA. Real-time quantitative PCR was applied to evaluate the mRNA levels of MerTK and ADAM17...
2014: Journal of Immunology Research
Christin Dewitz, Katja Möller-Hackbarth, Olga Schweigert, Karina Reiss, Athena Chalaris, Jürgen Scheller, Stefan Rose-John
T-cell immunoglobulin and mucin domain (TIM)-2 is expressed on activated B cells. Here, we provide evidence that murine TIM-2 is a target of ADAM10-mediated ectodomain shedding, resulting in the generation of a soluble form of TIM-2. We identified ADAM10 but not ADAM17 as the major sheddase of TIM-2, as shown by pharmacological ADAM10 inhibition and with ADAM10-deficient and ADAM17-deficient murine embryonic fibroblasts. Ionomycin-induced or 2'(3')-O-(4-benzoylbenzoyl) ATP triethylammonium salt-induced shedding of TIM-2 was abrogated by deletion of 10 juxtamembrane amino acids from the stalk region but not by deletion of two further N-terminally located blocks of 10 amino acids, indicating a membrane-proximal cleavage site...
January 2014: FEBS Journal
Sonia Y Velásquez, Luis F García, Gerhard Opelz, Cristiam M Alvarez, Caner Süsal
BACKGROUND: Membrane CD30 is an important costimulatory molecule for activated T lymphocytes, and serum level of soluble CD30 (sCD30) is considered a marker for predicting outcome in kidney transplantation. METHODS: We investigated the kinetics of CD30 expression on CD4 and CD8 T-cell populations and the source of sCD30 during alloimmune responses in vitro. The effect of neutralizing antibodies against interferon (IFN)-γ and other cytokines on sCD30 release and the involvement of metalloproteinases ADAM10 and ADAM17/TACE that are responsible for sCD30 shedding were also assessed...
July 27, 2013: Transplantation
Jia-yan Ling, Lin Shen, Qing Liu, Sha Xue, Wei Ma, Hui Wu, Zi-xi Li, Rui Zhu
Glycoprotein (GP) Ibα ectodomain shedding has important implications for thrombosis and hemostasis. A disintegrin and metalloproteinase 17 (ADAM17) was identified to play an essential role in agonist induced GPIbα shedding. The relationship of GPIbα shedding and ADAM17 in the acute stage of atherosclerotic ischemic stroke (AIS) patients has not been thoroughly studied. A total of 306 patients and 230 controls matched for age, sex, race, history of hypertension and diabetes mellitus were enrolled in the study...
June 2013: Journal of Huazhong University of Science and Technology. Medical Sciences
Michelle Dang, Nicole Armbruster, Miles A Miller, Efrain Cermeno, Monika Hartmann, George W Bell, David E Root, Douglas A Lauffenburger, Harvey F Lodish, Andreas Herrlich
Ectodomain cleavage of cell-surface proteins by A disintegrin and metalloproteinases (ADAMs) is highly regulated, and its dysregulation has been linked to many diseases. ADAM10 and ADAM17 cleave most disease-relevant substrates. Broad-spectrum metalloprotease inhibitors have failed clinically, and targeting the cleavage of a specific substrate has remained impossible. It is therefore necessary to identify signaling intermediates that determine substrate specificity of cleavage. We show here that phorbol ester or angiotensin II-induced proteolytic release of EGF family members may not require a significant increase in ADAM17 protease activity...
June 11, 2013: Proceedings of the National Academy of Sciences of the United States of America
Xiang-Jun Wang, Chang-Wei Feng, Min Li
A disintegrin and metalloproteinase-17 (ADAM17) is a member of the metalloproteinase superfamily and involved in the cleavage of ectodomain of many transmembrane proteins. ADAM17 is overexpressed in a variety of human tumors, which is associated with tumor development and progression. In the present study, we sought to investigate the expression and function of ADAM17 in hypoxia-treated hepatocellular carcinoma (HCC) cells. Western blot analysis was used to measure the expression of ADAM17 in HCC cell lines (Hep3B and HepG2 cells)...
August 2013: Molecular and Cellular Biochemistry
Guranda Chitadze, Marcus Lettau, Jaydeep Bhat, Daniela Wesch, Alexander Steinle, Daniel Fürst, Joannis Mytilineos, Holger Kalthoff, Ottmar Janssen, Hans-Heinrich Oberg, Dieter Kabelitz
The interaction of the MHC class I-related chain molecules A and B (MICA and MICB) with the corresponding natural killer group 2, member D (NKG2D) receptor triggers cytotoxic effector activity of natural killer cells and certain T-cell subsets and provides a costimulatory signal for cytokine production. Thus, the presence of MICA/B on transformed cells contributes to tumor immunosurveillance. Consequently, the proteolytic cleavage of MICA/B is regarded as an important immune escape mechanism of various cancer cells...
October 1, 2013: International Journal of Cancer. Journal International du Cancer
Alessandra F Perna, Immacolata Sepe, Diana Lanza, Rosanna Capasso, Silvia Zappavigna, Giovambattista Capasso, Michele Caraglia, Diego Ingrosso
H2S is the third endogenous gaseous mediator, after nitric oxide and carbon monoxide, possessing pleiotropic effects, including cytoprotection and anti-inflammatory action. We analyzed, in an in vitro model entailing monocyte adhesion to an endothelial monolayer, the changes induced by H2S on various potential targets, including cytokines, chemokines, and proteases, playing a crucial role in inflammation and cell adhesion. Results show that H2S prevents the increase in monocyte adhesion induced by tumor necrosis factor-α (TNF-α)...
July 2013: Journal of Cellular Biochemistry
Yue Wang, Jianming Wu, Robert Newton, Nooshin S Bahaie, Chunmei Long, Bruce Walcheck
CD16b (FcγRIIIb) is exclusively expressed by human neutrophils and binds IgG in immune complexes. Cell surface CD16b undergoes efficient ectodomain shedding upon neutrophil activation and apoptosis. Indeed, soluble CD16b is present at high levels in the plasma of healthy individuals, which appears to be maintained by the daily turnover of apoptotic neutrophils. At this time, the principal protease responsible for CD16b shedding is not known. We show that CD16b plasma levels were significantly decreased in patients administered a selective inhibitor targeting the metalloproteases ADAM10 and ADAM17...
March 2013: Biochimica et Biophysica Acta
Grazia Carbotti, Anna Maria Orengo, Delia Mezzanzanica, Marina Bagnoli, Antonella Brizzolara, Laura Emionite, Andrea Puppo, Maria Grazia Centurioni, Milena Bruzzone, Paola Marroni, Armando Rossello, Silvana Canevari, Silvano Ferrini, Marina Fabbi
Activated leukocyte cell adhesion molecule (ALCAM) is involved in cell-cell interactions in cancer. Shedding of its ectodomain by the metalloprotease ADAM17/TACE generates a soluble form (sALCAM). Here, we show that serum sALCAM levels were significantly higher in epithelial ovarian cancer (EOC) (p < 0.005) than in controls. The performance of sALCAM as classifier, tested by receiver operating characteristic curve, resulted in an area under the curve (AUC) of 0.8067. Serum sALCAM levels showed direct correlation with Carbohydrate Antigen-125 (CA125/MUC16)...
June 1, 2013: International Journal of Cancer. Journal International du Cancer
Feng Lin, Ping Lin, Xin Liu, Dong Li, Zi-Jun Liu, Hai-Feng Zou, Ying Jiang, Xue-Fei Zhao, Jin-Liang Feng, Xiao-Guang Yu
OBJECTIVE: To study the effect of siRNA targeting ADAM17 (ADAM17-siRNA) on the proliferation of prostate cancer PC-3 cells. METHODS: After transfecting PC-3 cells with ADAM17-siRNA 1 and ADAM17-siRNA 2, we detected the expressions of ADAM17 mRNA and protein by RT- PCR and Western blotting, respectively. We measured the changes in the proliferation and DNA synthesis of PC-3 cells by MTT and bromodeoxyuridine (BrdU) incorporation assay, examined the cell cycle profile by flow cytometry, and determined the expressions of the genes associated with PC-3 cell proliferation by Western blotting...
August 2012: Zhonghua Nan Ke Xue, National Journal of Andrology
Keisuke Nagao, Tetsuro Kobayashi, Manabu Ohyama, Haruhiko Akiyama, Keisuke Horiuchi, Masayuki Amagai
Hair follicles (HFs) are equipped with stem cell niches that allow regeneration. Tumor necrosis factor-α converting enzyme (TACE), also known as A disintegrin and metalloproteinase 17, is a proteolytic enzyme that regulates a variety of cell surface molecules including TNF-α, via ectodomain shedding. We found TACE expression on mouse HFs and conditionally depleted it in cells that expressed sex-determining region Y-related high-mobility-group box 9 (SOX9) transcription factor, an HF stem cell transcription factor (Tace(flox/flox) -Sox9-Cre, hereafter, "Tace/Sox9")...
August 2012: Stem Cells
Xiaojin Li, Liliana Pérez, Huizhou Fan
AIM: To determine if the cytotail of the principal sheddase tumor necrosis factor-α converting enzyme (TACE; ADAM17) controls protein ectodomain shedding. METHODS: Site-directed mutagenesis was performed to derive TACE variants. The resulting TACE expression plasmids with amino acid substitutions in the extracellular, cysteine-rich disintegrin domain (CRD) and/or deleted cytotail, along with an expression vector for the enhanced green fluorescence protein were transfected into shedding-defective M1 mutants stably expressing transmembrane L-selectin or transforming growth factor (TGF)-α...
November 26, 2011: World Journal of Biological Chemistry
Ahmad Trad, Nina Hedemann, Mohammad Shomali, Verena Pawlak, Joachim Grötzinger, Inken Lorenzen
ADAM17 (a disintegrin and metalloproteinase-containing protein 17) is a membrane-bound metalloproteinase, implicates in many physiological processes, including cell migration and proliferation. Of particular note, most of the studies so far are restricted on the analysis of ADAM17 mRNA levels. In this study we generated, utilizing hybridoma technology, three monoclonal antibodies (mAbs) (A 300, A 309 and A 318) against the extracellular domain of human ADAM17 to enable quantification of protein expression. The specificity of these mAbs against ADAM17 was tested by enzyme-linked immunoadsorbent assay (ELISA), flourescence-activated cell sorting (FACS) and western blotting...
August 31, 2011: Journal of Immunological Methods
Steeve Kwan Tat, Jean-Pierre Pelletier, François Mineau, Judith Caron, Johanne Martel-Pelletier
INTRODUCTION: In osteoarthritis (OA) the progression of cartilage degeneration has been associated with remodeling of the subchondral bone. Human OA subchondral bone osteoblasts were shown to have an abnormal phenotype and altered metabolism leading to an abnormal resorptive process. Bone resorption is suggested to occur, at least in part, through the increased levels of two proteolytic enzymes, MMP-2 and MMP-9, and RANKL, which are mainly produced by osteoblasts. In this study, we investigated in human OA subchondral bone osteoblasts the modulatory effect of strontium ranelate on the above key factors...
September 2011: Bone
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