Arthritis & Rheumatism, Volume 63,
November 2011 Abstract Supplement
Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Chicago, Illinois November 4-9, 2011.
Deep Sequencing of Small RNAs Reveals Novel MicroRNAs in Minor Salivary Glands of Sjgren's Syndrome Patients.
Tandon1, Mayank, Gallo1, Alessia, Illei2, Gabor G., Alevizos2, Ilias
Sjögren's Syndrome (SS) is a complex autoimmune disease of the salivary gland with an unknown etiology, so a thorough characterization of the transcriptome would facilitate our understanding of the disease. We use ultra deep sequencing of small RNAs from Sjögren's patients and healthy volunteers, primarily to identify and discover novel miRNA sequences that may play a role in the disease.
Total RNA was isolated from minor salivary glands of healthy volunteers (n=3) and patients with either high (n=3) or low (n=3) salivary flow. RNA from individuals was pooled and sequenced on the SOLiD 4 platform using the small RNA protocol from Applied Biosystems. Prediction of mature miRNAs from the sequenced reads was done using miRanalyzer and expression was validated using custom Taqman qPCR assays. Additional validation was performed on several cell lines, including human salivary gland (HSG) cells, Jurkat cells, and primary culture cells from patient salivary glands and fibroblasts.
The number of total reads generated, as well as the reads mapped during each step of the analysis, was greatly varied between samples. The Low Flow sample generated more reads (~26 million) than the Healthy Volunteer sample (~15 million). However, a smaller proportion of those reads was matched to the human genome in Low Flow patients, 3.6% compared to 27.8% in Healthy Volunteers, suggesting an elevated level of exogenous sequences in patients. We also validated the presence of six previously unidentified miRNA sequences in patient samples and in several cell lines, and one shows promise as a biomarker candidate. This novel miRNA is elevated 4.6-fold in patients with high salivary flow and 12.5-fold in patients with low salivary flow when compared to healthy volunteers.
Sequencing small RNAs in the salivary gland is largely unprecedented, but here we show the feasibility of discovering novel miRNAs and disease biomarkers by sequencing the transcriptome of pooled patient samples.
To cite this abstract, please use the following information:
Tandon, Mayank, Gallo, Alessia, Illei, Gabor G., Alevizos, Ilias; Deep Sequencing of Small RNAs Reveals Novel MicroRNAs in Minor Salivary Glands of Sjgren's Syndrome Patients. [abstract]. Arthritis Rheum 2011;63 Suppl 10 :498