A new electrochemical assay method for gene expression using hela cells with a secreted alkaline phosphatase (SEAP) reporter system

2012 ◽  
Vol 109 (8) ◽  
pp. 2163-2167 ◽  
Author(s):  
Mustafa Şen ◽  
Kosuke Ino ◽  
Hitoshi Shiku ◽  
Tomokazu Matsue
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Hela Cells ◽  
Assay Method ◽  
Reporter System ◽  
Electrochemical Assay

scholarly journals Quantification of Gene Expression with a Secreted Alkaline Phosphatase Reporter System

BioTechniques ◽  
1997 ◽  
Vol 23 (6) ◽  
pp. 1110-1114 ◽  
Author(s):  
Te-Tuan Yang ◽  
Parisa Sinai ◽  
Paul A. Kitts ◽  
Steven R. Kain
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Reporter System

scholarly journals Cell-Based Electrochemical Assay for Endotoxin Using a Secreted Alkaline Phosphatase Reporter System

2008 ◽  
Vol 76 (8) ◽  
pp. 525-528 ◽  
Author(s):  
Kumi Y. INOUE ◽  
Tomoyuki YASUKAWA ◽  
Hitoshi SHIKU ◽  
Tomokazu MATSUE
Keyword(s):  
Alkaline Phosphatase ◽  
Reporter System ◽  
Electrochemical Assay

scholarly journals Connectivity Map-based discovery of parbendazole reveals targetable human osteogenic pathway

2015 ◽  
Vol 112 (41) ◽  
pp. 12711-12716 ◽  
Author(s):  
Andrea M. Brum ◽  
Jeroen van de Peppel ◽  
Cindy S. van der Leije ◽  
Marijke Schreuders-Koedam ◽  
Marco Eijken ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Osteoblast Differentiation ◽  
Target Genes ◽  
Focal Adhesions ◽  
Bone Fragility ◽  
Bone Morphogenetic Protein 2 ◽  
Marker Genes ◽  
Connectivity Map ◽  
Morphogenetic Protein

Osteoporosis is a common skeletal disorder characterized by low bone mass leading to increased bone fragility and fracture susceptibility. In this study, we have identified pathways that stimulate differentiation of bone forming osteoblasts from human mesenchymal stromal cells (hMSCs). Gene expression profiling was performed in hMSCs differentiated toward osteoblasts (at 6 h). Significantly regulated genes were analyzed in silico, and the Connectivity Map (CMap) was used to identify candidate bone stimulatory compounds. The signature of parbendazole matches the expression changes observed for osteogenic hMSCs. Parbendazole stimulates osteoblast differentiation as indicated by increased alkaline phosphatase activity, mineralization, and up-regulation of bone marker genes (alkaline phosphatase/ALPL, osteopontin/SPP1, and bone sialoprotein II/IBSP) in a subset of the hMSC population resistant to the apoptotic effects of parbendazole. These osteogenic effects are independent of glucocorticoids because parbendazole does not up-regulate glucocorticoid receptor (GR) target genes and is not inhibited by the GR antagonist mifepristone. Parbendazole causes profound cytoskeletal changes including degradation of microtubules and increased focal adhesions. Stabilization of microtubules by pretreatment with Taxol inhibits osteoblast differentiation. Parbendazole up-regulates bone morphogenetic protein 2 (BMP-2) gene expression and activity. Cotreatment with the BMP-2 antagonist DMH1 limits, but does not block, parbendazole-induced mineralization. Using the CMap we have identified a previously unidentified lineage-specific, bone anabolic compound, parbendazole, which induces osteogenic differentiation through a combination of cytoskeletal changes and increased BMP-2 activity.

scholarly journals pTRA – A reporter system for monitoring the intracellular dynamics of gene expression

PLoS ONE ◽  
2018 ◽  
Vol 13 (5) ◽  
pp. e0197420 ◽  
Author(s):  
Sabine G. Wagner ◽  
Martin Ziegler ◽  
Hannes Löwe ◽  
Andreas Kremling ◽  
Katharina Pflüger-Grau
Keyword(s):  
Gene Expression ◽  
Reporter System

The influence of titanium surfaces treated by alkalis on macrophage and osteoblast-like cell adhesion and gene expression in vitro

RSC Advances ◽  
2015 ◽  
Vol 5 (99) ◽  
pp. 81378-81387 ◽  
Author(s):  
Ting Ma ◽  
Xi-Yuan Ge ◽  
Sheng-Nan Jia ◽  
Xi Jiang ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Cell Adhesion ◽  
Phosphatase Activity ◽  
Alkaline Phosphatase Activity ◽  
Related Gene ◽  
Titanium Surfaces

The effect of alkali-treated titanium surfaces on inflammation-related gene expression of macrophages and alkaline phosphatase activity of osteoblast-like cells.

ATF is important to late S phase-dependent regulation of DNA topoisomerase IIα gene expression in HeLa cells

2002 ◽  
Vol 184 (1) ◽  
pp. 81-88 ◽  
Author(s):  
Mee-Young Son ◽  
Tae-Jeong Kim ◽  
Kwang-In Kweon ◽  
Jong-Il Park ◽  
Chung Park ◽  
...  
Keyword(s):  
Gene Expression ◽  
Hela Cells ◽  
S Phase ◽  
Topoisomerase Iiα ◽  
Dna Topoisomerase ◽  
Dna Topoisomerase Iiα

Simulated microgravity using the Random Positioning Machine inhibits differentiation and alters gene expression profiles of 2T3 preosteoblasts

2005 ◽  
Vol 288 (6) ◽  
pp. C1211-C1221 ◽  
Author(s):  
Steven J. Pardo ◽  
Mamta J. Patel ◽  
Michelle C. Sykes ◽  
Manu O. Platt ◽  
Nolan L. Boyd ◽  
...  
Keyword(s):  
Gene Expression ◽  
Alkaline Phosphatase ◽  
Bone Loss ◽  
Simulated Microgravity ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Underlying Mechanism ◽  
Bone Biology ◽  
Random Positioning Machine

Exposure to microgravity causes bone loss in humans, and the underlying mechanism is thought to be at least partially due to a decrease in bone formation by osteoblasts. In the present study, we examined the hypothesis that microgravity changes osteoblast gene expression profiles, resulting in bone loss. For this study, we developed an in vitro system that simulates microgravity using the Random Positioning Machine (RPM) to study the effects of microgravity on 2T3 preosteoblast cells grown in gas-permeable culture disks. Exposure of 2T3 cells to simulated microgravity using the RPM for up to 9 days significantly inhibited alkaline phosphatase activity, recapitulating a bone loss response that occurs in real microgravity conditions without altering cell proliferation and shape. Next, we performed DNA microarray analysis to determine the gene expression profile of 2T3 cells exposed to 3 days of simulated microgravity. Among 10,000 genes examined using the microarray, 88 were downregulated and 52 were upregulated significantly more than twofold using simulated microgravity compared with the static 1-g condition. We then verified the microarray data for some of the genes relevant in bone biology using real-time PCR assays and immunoblotting. We confirmed that microgravity downregulated levels of alkaline phosphatase, runt-related transcription factor 2, osteomodulin, and parathyroid hormone receptor 1 mRNA; upregulated cathepsin K mRNA; and did not significantly affect bone morphogenic protein 4 and cystatin C protein levels. The identification of gravisensitive genes provides useful insight that may lead to further hypotheses regarding their roles in not only microgravity-induced bone loss but also the general patient population with similar pathological conditions, such as osteoporosis.

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