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Alterations in gene expression in lacrimal and salivary glands of male NOD mice due to LTBR-Ig treatment relative to control antibody

ABSTRACT: NOD mice were injected once a week with LTBR-Ig to block the LTBR-pathway, or with control monoclonal antibody MOPC from age 8 to 16 weeks old. Extraorbital lacrimal glands or submaxillary glands were dissected and total mRNA prepared. Each sample was either the combined lacrimals (2) from each mouse or individual salivary glands. There were 4 mice in each treatment group. Total mRNA was isolated and the quality was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). Reverse transcription to prepare cDNA was performed using Invitrogen M-MLV system. The purpose was to determine changes in gene expression in glands due to blockade of the LTBR-pathway.

ORGANISM(S): Mus musculus

PROVIDER: GSE32681 | GEO | 2011/10/07

SECONDARY ACCESSION(S): PRJNA146907

REPOSITORIES: GEO

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Project description:One µg of total RNA from either an individual rat or from a pooled sample was amplified and labeled with a fluorescent dye (either Cy3 or Cy5) using the Low RNA Input Linear Amplification Labeling kit (Agilent Technologies, Palo Alto, CA) following the manufacturer’s protocol. The amount and quality of the resulting fluorescently labeled cRNA was assessed using a Nanodrop ND-100 spectrophometer and an Agilent Bioanalyzer. Equal amounts of Cy3- or Cy5-labeled cRNA were hybridized to the Agilent Rat Oligo Microarray (Agilent Technologies, Inc., Palo Alto, CA) for 17 hrs, prior to washing and scanning. Data was extracted from the resulting images using Agilent’s Feature Extraction Software (Agilent Technologies, Inc., Palo Alto, CA). For day/night comparisons, two types of complementing hybridizations were performed: hepatic RNA from individual day rats was hybridized against a pool made up of RNA from 12 hour offset night rats, and RNA from individual night rats was hybridized against a pool made up of RNA from 12-hour offset day rats. Specifically, hepatic RNA samples from three individual day rats collected ten hours after light on (CT10) were hybridized against a pooled RNA sample composed of equal aliquots of RNA from the livers of six night rats collected ten hrs after lights off (CT22); and RNA samples from three individual night rats collected at CT22 were hybridized against a pooled RNA sample composed of equal aliquots of RNA from the livers of six day rats collected at CT10. This was replicated in a second study for a total of 24 hybridizations of 12 hour offset samples. For time of day comparisons, equal aliquots of RNA from the livers of six rats collected at each of four different times of the circadian day (CT4, CT10, C16 and CT22) were pooled. The two replicate studies thus resulted in 8 pools (two replicate pools/time point times four time points); each pool was hybridized against a Universal Rat Reference RNA Standard (Stratagene, La Jolla, CA). Keywords: time-course

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Alterations in gene expression in lacrimal and salivary glands of male NOD mice due to LTBR-Ig treatment relative to control antibody

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