Next Generation Sequencing for miRNA Detection on the Exhaled Breath Condensate: A Pilot Study

Cherchi, Roberto;Cusano, Roberto;Orrù, Sandro

;Ferrari, Paolo A;Massidda, Matteo;Fotia, Giorgio;De Matteis, Sara;Cocco, Pierluigi

2023-01-01

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Abstract

INTRODUCTION: Exhaled breath condensate (EBC) sampling has been suggested as a less-invasive and cost-effective method to detect biological macromolecules, including miRNA. To explore the feasibility of its use as a biomarker of early effects of asbestos exposure, we conducted a preliminary test on male volunteers by comparing the miRNA profile in the EBC and the plasma using 2 different sequencing platforms.METHODS: Six male volunteers, all retired and unexposed to dust or fumes, participated in the test. RNA was extracted from 200 mu L EBC samples and same-size plasma samples. Sample aliquots were processed in 2 laboratories using 2 different sequencing platforms: a MiSeq Illumina((R)) platform and a more performing HiSeq Illumina((R)) platform.RESULTS: The HiSeq3000((R)) sequencing platform identified twice as many unique molecular indexes (UMI)-validated miRNA as the MiSeq((R)) platform. The Spearman's correlation coefficient between EBC counts and plasma counts was significant in 5/6 subjects with either platform (MiSeq((R)) = 0.128-0.508, P = .026-<.001; HiSeq((R)) = 0.156-0.412, P = .001-<.001). The intraclass correlation coefficient confirmed the consistency of the miRNA profile over the 6 participants with both biospecimens. Exploring the agreement between the EBC and plasma samples with Bland-Altman plots showed that using the HiSeq3000((R)) platform substantially improved the EBC miRNA detection rate.CONCLUSION: Our preliminary study confirms that, when using the HiSeq((R)) sequencing platform, EBC sampling is a suitable, non-invasive method to detect the miRNA profile in healthy subjects.