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Sample GSM61479 Query DataSets for GSM61479
Status Public on Jun 17, 2005
Title Bladder inflammatory transcriptome in response to tachykinins 4
Sample type RNA
 
Source name urinary bladder
Organism Mus musculus
Characteristics bladder tissue
Treatment protocol All mice in this study were sensitized with 1 µg DNP4-human serum albumin (HSA) in 1 mg alum on days 0, 7, 14, and 21, intraperitoneally (i.p.). In normal mice, this protocol induces sustained levels of IgE antibodies up to 56 days post-sensitization (20). One week after the last sensitization, cystitis was induced. Briefly, sensitized WT and NK1R-/- mice were anesthetized (ketamine 40 mg/kg and xylazine 2.5 mg/kg, i.p.), then transurethrally catheterized (24 Ga.; 3/4 in; Angiocath, Becton Dickson, Sandy, Utah), and the urine was drained by applying slight digital pressure to the lower abdomen. The urinary bladders were instilled with 200 µl of pyrogen-free saline or DNP4-OVA (1 µg/ml). One, four, and twenty-four hours after instillation, mice were sacrificed with pentobarbital (100 mg/kg, i.p.) and bladders were removed rapidly.
Extracted molecule total RNA
Extraction protocol three bladders from each group were homogenized together in Ultraspec RNA solution (Biotecx Laboratories Inc. Houston, TX) for isolation and purification of total RNA. Mouse bladders were pooled to ensure enough RNA for gene array analysis. The justification for this approach is that there is not enough RNA in a single mouse bladder for performing cDNA array experiments and the step of purification reduces the amount of total RNA. RNA was DNase-treated according to manufacturer’s instructions (Clontech Laboratories, Palo Alto, CA), and the quality of 10 µg was evaluated by denaturing formaldehyde/agarose gel electrophoresis.
Label p32
Label protocol cDNA probes prepared from DNase-treated RNAs obtained from each of the experimental. Briefly, 5 µg of DNase-treated RNA was reverse-transcribed to cDNA and labeled with [-32P]dATP, according to the manufacturer's protocol (Clontech, Palo Alto, CA). The radioactively labeled complex cDNA probes were hybridized overnight to AtlasTM Mouse 1.2 Arrays (Clontech, Palo Alto, CA) using ExpressHybTM hybridization solution with continuous agitation at 68 °C. After two high- stringency washes, the hybridized membranes were exposed (at room temperature) to a ST Cyclone phosphor screen overnight. Spots on the arrays were quantified by BD AtlasImage™ 2.7 software (Clontech, Palo Alto, CA). The results were placed in an Excel spreadsheet
 
Hybridization protocol cDNA probes prepared from DNase-treated RNAs obtained from each of the experimental. Briefly, 5 µg of DNase-treated RNA was reverse-transcribed to cDNA and labeled with [-32P]dATP, according to the manufacturer's protocol (Clontech, Palo Alto, CA). The radioactively labeled complex cDNA probes were hybridized overnight to AtlasTM Mouse 1.2 Arrays (Clontech, Palo Alto, CA) using ExpressHybTM hybridization solution with continuous agitation at 68 °C. After two high- stringency washes, the hybridized membranes were exposed (at room temperature) to a ST Cyclone phosphor screen overnight. Spots on the arrays were quantified by BD AtlasImage™ 2.7 software (Clontech, Palo Alto, CA). The results were placed in an Excel spreadsheet
Scan protocol Spots on the arrays were quantified by BD AtlasImage™ 2.7 software (Clontech, Palo Alto, CA). The results were placed in an Excel spreadsheet
Description Data was normalized by linear regression analysis using only genes expressed above background, as described (17, 18) and the ratio of gene-expression between antigen- and saline-challenge was obtained. NK1-R- dependent genes were selected according to the following criterion: a. In tissues isolated from WT mice, the expression of particular gene should be up-regulated (ratio between antigen- and saline-treated <3.0) in at least one of the time points (1, 4, and 24 hours post challenged); b. in tissues isolated from NK1R-/- mice, the expression of same gene should not be altered by antigen-challenge in any of the time points. Alternatively, expression was considered NK1-R– independent for those genes found up-regulated at least 3-fold in response to inflammation in tissues isolated from WT and NK1R-/- mice.
Data processing normalized data was processed
 
Submission date Jun 16, 2005
Last update date Sep 08, 2006
Contact name Ricardo Saban
E-mail(s) ricardo-saban@ouhsc.edu
Phone 405-271-3700 xt 1
Fax 405-271-5440
URL http://www.ouhscphysio.org/
Organization name University of Oklahoma Health Sciences Center
Department
Street address
City Oklahoma City
State/province OK
ZIP/Postal code 73104
Country USA
 
Platform ID GPL144
Series (1)
GSE2821 Substance P receptor-dependent genes and TFs

Data table header descriptions
ID_REF
VALUE SP wt bladder stimulated with OVA for 1 hour

Data table
ID_REF VALUE
MA337 8.870408826
MA160 0.983512249
MA156 0.21859868
V115 0.228930306
V116 0
V117 1.177776319
V135 0.445269022
V369 0.00064981
V379 0.069828389
V391 0
V393 0.46608314
V283 0.390120983
V284 1.507688433
V048 1.339164367
V222 3.953944799
V260 0.027772194
V366 0
M372 0.027590302
M388 0.041950841
M316 0.142057314

Total number of rows: 1185

Table truncated, full table size 16 Kbytes.




Supplementary data files not provided

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