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ChIP assay

Yiwen Ouyang, Jingyi Li, Doris M Haverstick, James P Landers
We recently defined a magnetic bead-based assay that exploited an agglutination-like response for DNA and applied it to DNA-containing cell enumeration using inexpensive benchtop hardware [ J. Am. Chem. Soc. 2012 , 134 ( 12 ), 5689 - 96 ]. Although cost-efficient, the open-well format assay required numerous manual steps, and the magnetic field actuation scheme was not readily adaptable for integration. Here, we demonstrate a low-cost (<$2 in-lab), higher-throughput "pinwheel assay" platform that relies on a combination of a disposable rotation-driven microdisc (RDM), and a simple bidirectional rotating magnetic field (bi-RMF)...
October 25, 2016: Analytical Chemistry
Kyung Suk Lee, Lucy E Lee, Erel Levine
Small animals such as the roundworm C. elegans are excellent models for studying bacterial infection and host response, as well as for genetic and chemical screens. A key methodology is the killing assay, in which the number of surviving animals is tracked as a function of the time post infection. This is a labor-intensive procedure, prone to human error and subjective choices, and often involves undesired perturbation to the animals and their environment. In addition, the survival of animals is just one aspect of a multi-dimensional complex biological process...
October 24, 2016: Scientific Reports
Qiang Sun, Tao Zhou, Jun Niu, Peng Wu, Yanan Guo, Yanfei Yang
The aim of this study was to screen the biomarkers of diabetic limb arterial stenosis. Fasting blood samples of 40 patients with diabetic limb arterial stenosis (experimental group), 40 diabetes patients (diabetic control group), and 40 healthy individuals (healthy control group) were collected. Protein-chip assay analysis and ELISA were used to detect tumor necrosis factor α, interleukin-6, endothelin-1, calcitonin gene-related peptide and high-sensitivity C-reactive protein in the three groups. Protein-chip array analysis and ELISA found consistent results that endothelin-1, tumor necrosis factor α, interleukin-6 and high-sensitivity C-reactive protein in the experimental group were significantly up-regulated while the expression of calcitonin gene-related peptide was down-regulated compared with the healthy control group (P < 0...
October 21, 2016: Vascular
Saeed Rafizadeh-Tafti, Mohammad Hossein Haqiqatkhah, Mehrdad Saviz, Mohsen Janmaleki, Reza Faraji Dana, Somayeh Zanganeh, Mohammad Abdolahad
A highly sensitive impedimetric bio-chip based on vertically aligned multiwall carbon nanotubes (VAMWCNTs), was applied in direct interaction with lung cancer cells. Our tool provided both inducing and monitoring the bioelectrical changes in the cells initiated by electromagnetic (EM) wave stimulation. EM wave of 940MHz frequency with different intensities was used. Here, wave ablation might accumulate electrical charge on the tips of nanotubes penetrated into cell's membrane. The charge might induce ionic exchanges into the cell and cause alterations in electrical states of the membrane...
January 1, 2017: Materials Science & Engineering. C, Materials for Biological Applications
Hui Dong, Hao Sun, Jianping Zheng
With the development of large-scale biologic databases, precision medicine is becoming a frontier in biomedical research. As a main focus of precision medicine study, cancer has been widely accepted as a disease born out of inherited genetic variations or accumulating genomic damage. At the single-cell level, microfluidics or lab-on-a-chip technology for cancer study is an emerging tool for improving risk assessment, diagnostic categories and therapeutic strategies. This work presents a multi-layer microchip for single-cell gene expression profiling...
December 1, 2016: Talanta
Yuxin Shu, Yan Lu, Xiaojuan Pang, Wei Zheng, Yahong Huang, Jiahong Li, Jianguo Ji, Can Zhang, Pingping Shen
Peroxisome proliferator-activating receptor γ (PPARγ), a transcription factor, is involved in many important biological processes, including cell terminal differentiation, survival and apoptosis. However, the role of PPARγ, which regulates tumour promoter and oncogene expression, is not well understood in hepatocellular carcinoma (HCC). In the present study, based on evidence from clinical samples that phosphorylation of PPARγ at Ser84 is up-regulated in human liver tumours, we confirmed that phosphorylation of PPARγ was also significantly increased in an HCC mouse model and was increased by Mitogen-activated protein kinase (MEK)/ Extracellular-signal-regulated kinases (ERK) kinase...
October 19, 2016: Oncotarget
Qian-Qian Luo, Zhong-Ming Qian, Yu-Fu Zhou, Meng-Wan Zhang, Dang Wang, Li Zhu, Ya Ke
The inconsistent of responses of IRP1 and HIF-1 alpha to hypoxia and the similar tendencies in the changes of IRP1 and pCREB contents led us to hypothesize that pCREB might be involved in the regulation of IRP1 under hypoxia. Here, we investigated the role of pCREB in IRP1 expression in HepG2 cells under hypoxia using quantitative PCR, western blot, immunofluorescence, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). We demonstrated that 1) Hypoxia increased pCREB levels inside of the nucleus; 2) Putative CREs were found in the IRP1 gene; 3) Nuclear extracts of HepG2 cells treated with hypoxia could bind to CRE1 and CRE3, and 100-fold competitor of putative CREs could abolish the binding activity to varying degrees; 4) pCREB was found in the CRE1 and CRE3 DNA-protein complexes of EMSA; 5) CRE1 and CRE3 binding activity of IRP1 depended on CREB activation but not on HIF-1; 6) Increased IRP1 expression under hypoxia could be prevented by LY294002; 7) ChIP assays demonstrated that pCREB binds to IRP1 promoter; and 8) HIF-1 and/or HIF-2 siRNA had no effect on the expression of pCREB and IRP1 proteins in cells treated with hypoxia for 8 hours...
2016: International Journal of Biological Sciences
Chengxing Zhu, Jiong Yu, Qiaoling Pan, Jinfeng Yang, Guangshu Hao, Yingjie Wang, Lanjuan Li, Hongcui Cao
Human placenta-derived mesenchymal stem cells (hPMSCs) reside in a physiologically low-oxygen microenvironment. Hypoxia influences a variety of stem cell cellular activities, frequently involving hypoxia-inducible factor-2 alpha (HIF-2α). This research showed that hPMSCs cultured in hypoxic conditions (5% O2) exhibited a more naïve morphology and had a higher proliferative capability and higher HIF-2α expression than hPMSCs cultured in normoxic conditions (21% O2). Similar to the hypoxic cultures, hPMSCs over-expressing HIF-2α showed higher proliferative potential and higher expression of CCND1 (CyclinD1), MYC (c-Myc), POU5F1 (Oct4) and the components of the MAPK/ERK pathway...
October 21, 2016: Scientific Reports
Takayuki Koga, Pei-Li Yao, Maryam Goudarzi, Iain A Murray, Gayathri Balandaram, Frank J Gonzalez, Gary H Perdew, Albert J Fornace, Jeffrey M Peters
Alcoholic liver disease is a pathological condition caused by over-consumption of alcohol. Due to the high morbidity and mortality associated with this disease, there remains a need to elucidate the molecular mechanisms underlying its etiology and to develop new treatments. Since peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) modulates ethanol-induced hepatic effects, the present study examined alterations in gene expression that may contribute to this disease. Chronic ethanol treatment causes increased hepatic CYP2B10 expression in Pparβ/δ+/+ mice, but not in Pparβ/δ-/- mice...
October 20, 2016: Journal of Biological Chemistry
Chuanguang Yao, Hongxin Song, Ying Wan, Kefeng Ma, Chenyu Zheng, Hongda Cui, Peng Xin, Xubo Ji, Sheng-Yuan Deng
A porphyrin-based electro-photodynamic imaging system was fabricated for monitoring the concentration of oxygen. Distinct from the electrochemiluminescent (ECL) inability of numerous organic species in aqueous solutions, a strong and stable red irradiation at 634 nm could be stimulated electrochemically on zinc(II) meso-tetra(4-carboxyphenyl) porphine (ZnTCPP)/tetraoctylammonium bromide (TOAB) in the physiological condition. In terms of in situ electron paramagnetic resonance and ECL spectroscopies, the nature of ECL was thoroughly investigated, being exactly the chemiluminescence from singlet oxygen (1O2) produced during the successive electro-reduction of ZnTCPP...
October 20, 2016: ACS Applied Materials & Interfaces
David Olagnier, Cindy Chiang, John Hiscott
The dynamics of chromatin structure contribute to the regulation of gene transcription and in part, the changes in chromatin structure associated with gene activation/repression are a function of the state of histone acetylation. Histone deacetylases (HDACs) deacetylate histone tails leading to a more compact structure of chromatin that in turn represses gene transcription. Given the rapid activation and/or repression of gene networks following microbial infection, the role of HDACs in the epigenetic regulation of genes involved in the innate and adaptive immune responses has become an area of extensive research...
2017: Methods in Molecular Biology
Wen-Jin Ding, Min Zhou, Mei-Mei Chen, Chun-Ying Qu
PURPOSE: The homeobox B8 (HOXB8) functions as a sequence-specific transcription factor that is involved in development. Increased expression of this gene is associated with a wide variety of tumor; however, its function in gastric cancer has not been clarified. In the present study, the expression of HOXB8 in gastric cancer tissues and influence of HOXB8 on gastric cancer cellular were evaluated. METHODS: The expression levels of HOXB8 mRNA in human gastric cancer tissues were analyzed through quantitative RT-PCR...
October 19, 2016: Journal of Cancer Research and Clinical Oncology
Amel Dudakovic, Emily T Camilleri, Scott M Riester, Christopher R Paradise, Martina Gluscevic, Thomas M O'Toole, Roman Thaler, Jared M Evans, Huihuang Yan, Malayannan Subramaniam, John R Hawse, Gary S Stein, Martin A Montecino, Meghan E McGee-Lawrence, Jennifer J Westendorf, Andre J van Wijnen
Perturbations in skeletal development and bone degeneration may result in reduced bone mass and quality leading to greater fracture risk. Bone loss is mitigated by bone protective therapies, but there is a clinical need for new bone-anabolic agents. Previous work has demonstrated that enhancer of zeste homolog 2 (Ezh2), a histone 3 lysine 27 (H3K27) methyltransferase, suppressed differentiation of osteogenic progenitors. Here, we investigated if inhibition of Ezh2 can be leveraged for bone stimulatory applications...
October 10, 2016: Journal of Biological Chemistry
Xiang Du, Mi-Die Xu, Yiqin Wang, Wei-Wei Weng, Ping Wei, Peng Qi, Qiongyan Zhang, Cong Tan, Shujuan Ni, Lei Dong, Yusi Yang, Wanrun Lin, Qinghua Xu, Dan Huang, Zhaohui Huang, Yuqing Ma, Wei Zhang, Weiqi Sheng
PURPOSE: The long noncoding RNA (lncRNA) PVT1 is an important epigenetic regulator with a critical role in human tumors. Here, we aimed to investigate the clinical application and the potential molecular mechanisms of PVT1 in gastric cancer (GC) tumorigenesis and progression. EXPERIMENTAL DESIGN: The expression level of PVT1 was determined by RT-qPCR analysis in 190 pairs of GC tissues and adjacent normal gastric mucosa tissues (ANTs). The biological functions of PVT1 were assessed by in vitro and in vivo functional experiments...
October 18, 2016: Clinical Cancer Research: An Official Journal of the American Association for Cancer Research
Dan Li, Jie Hong, Weibiao Cao
We have shown that NOX5-S may mediate the acid-induced decrease in cell apoptosis. However, mechanisms of NOX5-S-dependent decrease in cell apoptosis are not fully understood. In this study we found that silencer of death domain (SODD) was significantly increased in esophageal adenocarcinoma (EA) tissues, EA cell lines FLO and OE33, and a dysplastic cell line CP-B. Strong SODD immunostaining was significantly higher in low-grade dysplasia (66.7%), high-grade dysplasia (81.2%) and EA (71.2%) than in Barrett's mucosa (10...
October 18, 2016: Journal of Pharmacology and Experimental Therapeutics
Zheng Zhang, Bo Jin, Yaqiong Jin, Shengquan Huang, Xiaohua Niu, Zebin Mao, Dianqi Xin
Androgens (AR) play an important role in initiation and progression of prostate cancer. It has been shown that AR exert their effects mainly through the androgen-activated AR which binds to androgen response elements (AREs) in the regulatory regions of target genes to regulate the transcription of androgen-responsive genes, thus, identification of AR downstream target gene is critical to understand androgen function in prostate cancer. In this study, our results showed that androgen treatment of LNCaP cells induced PTTG1 expression, which was blocked by the androgen receptor antagonist, Casodex...
October 15, 2016: Experimental Cell Research
Xiong Liao, Xiao Guo, Qi Wang, Yantao Wang, Di Zhao, Liping Yao, Shuang Wang, Guojie Liu, Tianhong Li
Dehydration-responsive element binding factors (DREBs) play important roles in plant growth, development, and stress signaling pathways in model plants. However, little is known about the function of DREBs in apple (Malus × domestica), a widely cultivated crop that is frequently threatened by drought. We isolated a DREB gene from Malus sieversii (Ledeb.) Roem., MsDREB6.2, and investigated its functions using overexpression analysis and chimeric repressor gene-silencing technology (CRES-T). We identified possible target genes of the protein encoded by MsDREB6...
October 18, 2016: Plant Journal: for Cell and Molecular Biology
Juan Wang, Hiroyuki Kobori, Yuki Shibayama, Daisuke Nakano, Hirofumi Hitomi, Akira Nishiyama
OBJECTIVE: Previous studies have shown that angiotensinogen (AGT) synthesis is enhanced by high glucose (HG) in rat renal proximal tubular cells. We aimed to investigate the glucose-responsive elements within human AGT (hAGT) promoter in human immortalized proximal tubular HK-2 cells. DESIGN AND METHOD: HK-2 cells were treated with normal glucose (NG: 5.5 mmol/L) and high glucose (HG: 15 mmol/L). Human AGT promoter region was cloned from -4358 to +122. Consecutive 5'-end deletion mutant constructs and different site-direct mutagenesis products were transfected into HK-2, followed by promoter activity measurement by dual luciferase assay...
September 2016: Journal of Hypertension
Deepesh Pandey, Daijiro Tori, Yohei Nomura, Dan Berkowitz, Lewis Romer
OBJECTIVE: Kruppel Like Factor 15 (KLF15) has recently been shown to be critical for activation of proinflammatory processes in vascular smooth muscle and atherogenesis. Although KLF15 is abundantly expressed in vascular endothelium there is a significant lack of knowledge regarding the role of KLF15 in the regulation of vascular endothelial function. Here we tested the hypothesis that KLF15 is a critical regulator of Arg2 transcription in hypoxia exposed human pulmonary microvascular endothelial cells (HPMEC) and that it plays critical role in pathogenesis of pulmonary hypertension (PHTN)...
September 2016: Journal of Hypertension
M Gokulnath, R Swetha, G Thejaswini, P Shilpa, N Selvamurugan
Transforming growth factor-beta1 (TGF-β1) plays a significant role in breast cancer mediated bone metastasis, and it stimulated expression of matrix metalloproteinase-13 (MMP-13; an invasive and metastasis gene) via activating transcription factor-3 (ATF-3) in human breast cancer cells (MDA-MB231). We further dissected the role of ATF-3 and its interacting proteins (activator protein-1; AP-1) for TGF-β1-stimulation of MMP-13 expression in these cells. Chromatin immunoprecipitation (ChIP) experiment identified the TGF-β1-stimulation of ATF-3 interaction at the AP-1 site of the MMP-13 promoter in a sustained and prolonged manner in MDA-MB231 cells...
October 15, 2016: International Journal of Biological Macromolecules
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