Investigation Title Transcription profiling of human pulp and odontoblasts reponse to TGF-beta1 Comment[Submitted Name] Effects of TGF-β1 on expression profile of human pulp and odontoblasts Experimental Design unknown_experiment_design_type transcription profiling by array Experimental Design Term Source REF EFO Comment[ArrayExpressReleaseDate] 2008-06-16 Comment[SecondaryAccession] GSE8730 Comment[AEMIAMESCORE] 4 Comment[ArrayExpressAccession] E-GEOD-8730 Comment[MAGETAB TimeStamp_Version] 2011-01-23 03:11:54 Last Changed Rev: 14857 Experimental Factor Name Experimental Factor Type Experimental Factor Term Source REF Person Last Name Pääkkönen Person First Name Virve Person Mid Initials Person Email virve.paakkonen@oulu.fi Person Phone Person Fax Person Address Institute of Dentistry, University of Oulu, Aapistie 3, Oulu, 90014, Finland Person Affiliation University of Oulu Person Roles submitter Person Roles Term Source REF Quality Control Type Quality Control Term Source REF Replicate Type Replicate Term Source REF Normalization Type Normalization Term Source REF Date of Experiment Public Release Date 2008-06-16 PubMed ID 17889552 Publication DOI 17889552 Publication Author List Virve Pääkkönen, Jussi Vuoristo, Tuula Salo, Leo Tjäderhane Publication Title Effects of TGF-beta1 on interleukin profile of human dental pulp and odontoblasts. Publication Status journal_article Publication Status Term Source REF The MGED Ontology Experiment Description Transforming growth factor beta 1 (TGF-β1) is the most extensively studied growth factor in dentin-pulp complex, with pleiotropic effects on pulp response and healing. Our main objective was to analyze the expression profile of pulp tissue and odontoblasts, and the effects of TGF-β1 on these profiles in cultured human pulp and odontoblasts with a specific interest in the anti- and pro-inflammatory cytokines. Experiment Overall Design: Pulp tissues and odontoblasts were cultured for different time periods, and microarray was performed to both cultured and native samples to detect the effects of TGF-β1. Expression of various interleukins (IL) were confirmed by RT-PCR, and in +/- TGF-β1 treated pulps also by antibody array. Protocol Name P-G8730-14 P-G8730-9 P-G8730-15 P-G8730-5 P-G8730-10 P-G8730-1 P-G8730-13 P-G8730-7 P-G8730-6 P-G8730-12 P-G8730-4 P-G8730-11 P-G8730-3 P-G8730-8 P-G8730-2 Affymetrix:Protocol:Hybridization-Unknown P-AFFY-6 Affymetrix:Protocol:ExpressionStat Protocol Type specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action specified_biomaterial_action nucleic_acid_extraction labeling labeling hybridization feature_extraction bioassay_data_transformation Protocol Description Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 24h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 5h without TGF-beta. Freshly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 48h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The crowns were embedded into agarose gel pulp chambers facing upwards, and the pulp chambers were filled with OPTIMEM. Odontoblasts were cultured for 24h without TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 24h without TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The crowns were embedded into agarose gel pulp chambers facing upwards, and the pulp chambers were filled with OPTIMEM. Odontoblasts were cultured for 1h without TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 5h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 1h without TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The crowns were embedded into agarose gel pulp chambers facing upwards, and the pulp chambers were filled with OPTIMEM. Odontoblasts were cultured for 24h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 1h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The crowns were embedded into agarose gel pulp chambers facing upwards, and the pulp chambers were filled with OPTIMEM. Odontoblasts were cultured for 1h with TGF-beta. Fresly extracted human third molars were cut 2-3mm apically from cement-enamel junction and pulp tissues were removed. The pulps were cultured on 24-well culture plates in OPTIMEM-medium for 48h without TGF-beta. Total RNA was isolated using Trizol-protocol and further purified by Qiagen RNeasy Mini Kit Double-stranded DNA was synthesized by means of the Superscript Choice System (Gibco BRL Life Technologies, Rockville, MD, USA) and T7-(dT)24 primer. The DNA was purified using GeneChip Sample Cleanup Module (Qiagen, Valencia, CA, USA). In vitro transcription was performed to produce biotin labeled cRNA using a BioArray HighYield RNA Transcription Labeling Kit (Enzo Diagnostics, Farmingdale, NY, USA) according to the manufacturer’s instructions. Biotinylated cRNA was cleaned with an GeneChip Sample Cleanup Module (Qiagen), fragmented to 35 to 200 nt, and hybridized to Affymetrix HGU133A . After being washed, the array was stained with streptavidin-phycoerythrin (Molecular Probes, Eugene, OR, USA). Staining signal was amplified by biotinylated anti-streptavidin (Vector Laboratories, Burlingame, CA, USA) and second staining with streptavidin-phycoerythrin. Double-stranded DNA was synthesized by means of the Superscript Choice System (Gibco BRL Life Technologies, Rockville, MD, USA) and T7-(dT)24 primer. The DNA was purified using GeneChip Sample Cleanup Module (Qiagen, Valencia, CA, USA). In vitro transcription was performed to produce biotin labeled cRNA using a BioArray HighYield RNA Transcription Labeling Kit (Enzo Diagnostics, Farmingdale, NY, USA) according to the manufacturer’s instructions. Biotinylated cRNA was cleaned with an GeneChip Sample Cleanup Module (Qiagen), fragmented to 35 to 200 nt, and hybridized to Affymetrix HGU133plus 2.0 (odontoblast samples). After being washed, the array was stained with streptavidin-phycoerythrin (Molecular Probes, Eugene, OR, USA). Staining signal was amplified by biotinylated anti-streptavidin (Vector Laboratories, Burlingame, CA, USA) and second staining with streptavidin-phycoerythrin. Title: Affymetrix Generic Hybridization. Description: Title: Affymetrix CEL analysis. Description: Title: Affymetrix CHP Analysis (ExpressionStat). Description: Protocol Parameters Protocol Hardware Protocol Software MicroArraySuite 5.0 MicroArraySuite 5.0 MicroArraySuite 5.0 Protocol Contact Protocol Term Source REF mo SDRF File E-GEOD-8730.sdrf.txt Term Source Name mo The MGED Ontology ArrayExpress EFO The MGED Ontology mo Term Source File http://mged.sourceforge.net/ontologies/MGEDontology.php http://mged.sourceforge.net/ontologies/MGEDontology.php http://www.ebi.ac.uk/arrayexpress http://www.ebi.ac.uk/efo/ http://mged.sourceforge.net/ontologies/MGEDontology.php http://mged.sourceforge.net/ontologies/MGEDontology.php Term Source Version