Asthma and lower airway diseaseMeasures of gene expression in sputum cells can identify TH2-high and TH2-low subtypes of asthma
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Subjects and clinical samples
We studied induced sputum samples stored in the Airway Tissue Bank at the University of California, San Francisco (UCSF). The biological samples were collected in multiple research studies at UCSF from 2009-2013 in which the characterization studies and biospecimen collections had all been collected according to standardized and uniform protocols (NCT00917787, NCT01073410, NCT00595153, and NCT01508078). Asthmatic patients had a prior physician's diagnosis of asthma and airway
RNA quality in cell pellets from induced sputum
Storage buffer and DNA extraction before RNA extraction had marked effects on the RIN values of sputum cell RNA (Fig 1, A). Specifically, 32% of the 52 sputum cell pellets stored in PBS had RIN values of greater than 5, 46% of the 60 sputum cell pellets stored in RNA protection buffer (RLT/BME or the Qiagen RNAprotect Saliva Reagent) had RIN values of greater than 5, and 92% of 26 sputum cell pellets stored in RLT/BME or RNA Saliva Reagent and processed with a DNA elimination column before RNA
Discussion
We describe optimized methods for extracting high-quality RNA from cells in induced sputum, and we show that measures of gene expression in sputum cells can identify TH2-high and TH2-low subtypes of asthma. Our data suggest that measures of gene expression in sputum cells could be applied in molecular phenotyping studies of asthma, including studies to investigate the unknown molecular abnormalities occurring in patients with TH2-low disease.
In initial studies we set out to examine the quality
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Supported in part by a research grant from Genentech, and National Institutes of Health grants P01HL107202 and T32HL007185.
Disclosure of potential conflict of interest: M. C. Peters has received grants from the National Institutes of Health (NIH; T325T32HL007185). N. R. Bhakta has received grants from the NIH (F321F32HL110720 and NIH T325T32HL007185). P. G. Woodruff has received grants from Genentech; has consultant arrangements with Genentech, MedImmune, Astra Zeneca, Boehringer Ingelheim, Merck, and Kalobios; has grants/grants pending with Genentech and Pfizer; and has a patent application for asthma diagnostics. J. V. Fahy has received research grants from the NIH; has received consulting fees or honoraria from Merck, Regeneron, Boehringer Ingelheim, Pathway Therapeutics, Cytokinetics, Amgen, and the University of Calgary; has received support for travel to meetings for study or other purposes from Boehringer Ingelheim to the Transatlantic Airway Conference; has received fees for participation in review activities, such as data monitoring boards, statistical analysis, end point committees, and the like from the NIH; and has patents planned, pending, or issued for a patent describing biomarkers of TH2-high asthma. The rest of the authors declare that they have no relevant conflicts of interest.