Comment[ArrayExpressAccession]	E-GEOD-41825
MAGE-TAB Version	1.1											
Public Release Date	2015-10-24
Investigation Title	Bruton's Tyrosine Kinase Regulates TLR9 but not TLR7 Signaling in Human pDCs											
Comment[Submitted Name]	Bruton's Tyrosine Kinase Regulates TLR9 but not TLR7 Signaling in Human pDCs
Experiment Description	Plasmacytoid dendritic cells (pDCs) can be activated by the endosomal TLRs, and contribute to the pathogenesis of systemic lupus erythematosus (SLE) by producing type I IFNs. Thus, blocking TLR-mediated pDC activation may represent a useful approach for the treatment of SLE. In an attempt to identify a therapeutic target for blocking TLR signaling in pDCs, we investigated the contribution of Bruton's tyrosine kinase (Btk) to the activation of pDCs by TLR7 and TLR9 stimulation by using a selective Btk inhibitor RN486. Stimulation of TLR7 and 9 with their respective agonist, namely, gardiquimod and type A CpG ODN2216, resulted in the activation of human pDCs, as demonstrated by the expression of activation markers (CD69, CD40, and CD86), elevated production of IFN-alpha and other inflammatory cytokines, as well as up-regulation of numerous genes including IFN-alpha-inducible genes and activation of interferon regulatory factor 7 (IRF7) and NF-kB. RN486 inhibited all of these events induced by TLR9, but not TLR7 stimulation, with a nanomolar potency for inhibiting type A CpG ODN2216-mediated production of cytokines (e.g., IC50=386 nM for inhibiting IFN-alpha). Our data reveal Btk as an important regulatory enzyme in the TLR9 pathway, and a potential therapeutic target for SLE and other TLR-driven diseases. pDCs from healthy donors (n=4) were treated with gardiquimod (TLR7 agonist) or ODN 2216 (TLR9 agonist) with or without BTK inhibitor for 3 hours.											
Term Source Name	ArrayExpress	EFO
Term Source File	http://www.ebi.ac.uk/arrayexpress/	http://www.ebi.ac.uk/efo/efo.owl										
Person Last Name	Lau	Wang	Lau	Jung	Ravindran	Hang	Carvajal	Majmudar	Allard	Bitter	Narula	Peterson
Person First Name	Kai	Jingming	Kai	Jimmy	Palanikumar	Julie	Valerie	Rupal	John	Hans	Satwant	Larry
Person Mid Initials			Y									
Person Email	geo@ncbi.nlm.nih.gov											
Person Affiliation	Hoffmann-La Roche											
Person Address	Hoffmann-La Roche, 340 Kingsland St, Nutley, USA											
Person Roles	submitter											
Protocol Name	P-GSE41825-1	P-GSE41825-3	P-GSE41825-4	P-GSE41825-2	P-GSE41825-5							
Protocol Description	Data was normalized in R/Bioconductor using the robust multi-array average (RMA) expression measure. Probesets were annotated using the latest annotation available in Bioconductor and curated using an internal Roche probe inspector tool, which updates gene annotation per probeset based on the latest human genome draft release. As an additional quality measure, probesets were also curated using the tool for sequence specificity; probesets mapping to non-unique locations in the genome were flagged and ignored in subsequent analysis. A total of 18,935 out of 54,675 probesets passed the QC and curation processes, corresponding to 10,260 unique genes. ID_REF =  VALUE = Log2 RMA signal intensity	Total RNA (100ng) was transcribed and labeled into amplified RNA (aRNA) using the 3'IVT Express Kit (Affymetrix Inc., Santa Clara, CA) according to the manufacturer's instructions on a GCAS (GeneChip Array Station).	Fragmented and labeled aRNA was hybridized to the Affymetrix U133 Plus 2.0 chips according to the manufacturer's instructions. Wash and stain protocol FS450_0004.	Total RNA was extracted using the mirVana RNA Isolation Kit (Ambion, Inc., Austin, TX, USA) according to the manufacturer's protocol. RNA was quantified using a ND-1000 spectrophotometer (NanoDrop Technology®, Cambridge, UK), and RNA integrity was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). The 260/280 ratios were between 1.7 and 2.1, and the RINs (RNA Integrity Numbers) were greater than 6 for all samples.	GeneChips were scanned using the GC 30007G scanner.							
Protocol Type	normalization data transformation protocol	labelling protocol	hybridization protocol	nucleic acid extraction protocol	array scanning protocol							
Experimental Factor Name	treatment	subject										
Experimental Factor Type	treatment	subject										
Comment[SecondaryAccession]	GSE41825											
Comment[GEOReleaseDate]	2015-10-24											
Comment[ArrayExpressSubmissionDate]	2012-10-24											
Comment[GEOLastUpdateDate]	2015-10-26											
Comment[AEExperimentType]	transcription profiling by array											
SDRF File	E-GEOD-41825.sdrf.txt