Am J Clin Exp Immunol 2012;1(2):70-89

Original Article
An integrated nano-scale approach to profile miRNAs in limited clinical
samples

Grégory Seumois, Pandurangan Vijayanand, Christopher J Eisley, Nada Omran, Lukas Kalinke, Mal North, Asha P
Ganesan, Laura J Simpson, Nathan Hunkapiller, Felix Moltzahn, Prescott G Woodruff, John V Fahy, David J Erle, Ratko
Djukanovic, Robert Blelloch, K Mark Ansel

Sandler Asthma Basic Research Center, University of California San Francisco, San Francisco, CA, USA; Division
of Infection, Inflammation and Immunity, University of Southampton, School of Medicine, Southampton NIHR Respiratory
Biomedical Research Unit, Sir Henry Wellcome Laboratories, Southampton General Hospital, Southampton,
UK; Pulmonary and Critical Care Division, Department of Medicine, University of California San Francisco;
Lung Biology Center, University of California San Francisco; Center for Reproductive Sciences, Department of
Obstetrics, Gynecology & Reproductive Sciences, University of California San Francisco; The Eli and Edythe Broad
Center of Regeneration Medicine and Stem Cell Research, Center for Reproductive Sciences, and Department of
Urology, University of California San Francisco; Department of Microbiology & Immunology, University of California
San Francisco; Current address: Division of Cell Signaling and Gene Expression, La Jolla Institute for Allergy
and Immunology, San Diego, USA. These authors contributed equally to this work.

Received September 19, 2012; Accepted September 26, 2012; Epub September 27, 2012; Published November 30, 2012

Abstract: Profiling miRNA expression in cells that directly contribute to human disease pathogenesis is likely to aid
the discovery of novel drug targets and biomarkers. However, tissue heterogeneity and the limited amount of human
diseased tissue available for research purposes present fundamental difficulties that often constrain the scope and
potential of such studies. We established a flow cytometry-based method for isolating pure populations of pathogenic
T cells from bronchial biopsy samples of asthma patients, and optimized a high-throughput nano-scale qRT-PCR
method capable of accurately measuring 96 miRNAs in as little as 100 cells. Comparison of circulating and airway
T cells from healthy and asthmatic subjects revealed asthma-associated and tissue-specific miRNA expression
patterns. These results establish the feasibility and utility of investigating miRNA expression in small populations of
cells involved in asthma pathogenesis, and set a precedent for application of our nano-scale approach in other
human diseases. The microarray data from this study (Figure 7) has been submitted to the NCBI Gene Expression
Omnibus (GEO; http://ncbi.nlm.nih.gov/geo) under accession no. GSE31030. (AJCEI1209003)

Keywords: microRNA (miRNA), asthma, helper T cell, microfluidic, qPCR arrays


Address all correspondence to:
Dr. K Mark Ansel
513 Parnassus Avenue, UCSF Box 0414
San Francisco, CA 94143-0414, USA.
Tel: +1-415-476-5368; Fax:
+1-415-502-4995
E-mail: mark.ansel@ucsf.edu
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