title: MicroRNA Profiling in Aqueous Humor of Individual Human Eyes by Next-Generation Sequencing.
abstract: OBJECTIVE
  Extracellular microRNAs (miRNAs) in aqueous humor were suggested to have a role in transcellular signaling and may serve as disease biomarkers. The authors adopted next-generation sequencing (NGS) techniques to further characterize the miRNA profile in single samples of 60 to 80 μL human aqueous humor.
  
  
  METHODS
  Samples were obtained at the outset of cataract surgery in nine independent, otherwise healthy eyes. Four samples were used to extract RNA and generate sequencing libraries, followed by an adapter-driven amplification step, electrophoretic size selection, sequencing, and data analysis. Five samples were used for quantitative PCR (qPCR) validation of NGS results. Published NGS data on circulating miRNAs in blood were analyzed in comparison.
  
  
  RESULTS
  One hundred fifty-eight miRNAs were consistently detected by NGS in all four samples; an additional 59 miRNAs were present in at least three samples. The aqueous humor miRNA profile shows some overlap with published NGS-derived inventories of circulating miRNAs in blood plasma with high prevalence of human miR-451a, -21, and -16. In contrast to blood, miR-184, -4448, -30a, -29a, -29c, -19a, -30d, -205, -24, -22, and -3074 were detected among the 20 most prevalent miRNAs in aqueous humor. Relative expression patterns of miR-451a, -202, and -144 suggested by NGS were confirmed by qPCR.
  
  
  CONCLUSIONS
  Our data illustrate the feasibility of miRNA analysis by NGS in small individual aqueous humor samples. Intraocular cells as well as blood plasma contribute to the extracellular aqueous humor miRNome. The data suggest possible roles of miRNA in intraocular cell adhesion and signaling by TGF-β and Wnt, which are important in intraocular pressure regulation and glaucoma.
authors.affiliation: Eye Center Freiburg University Medical Center, Freiburg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Wecker
authors.affiliation: GenXPro GmbH, Frankfurt Innovation Center Biotechnology, Frankfurt, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Hoffmeier
authors.affiliation: GenXPro GmbH, Frankfurt Innovation Center Biotechnology, Frankfurt, Germany.
authors.forename: Anne
authors.initials: A
authors.lastname: Plötner
authors.affiliation: Bioinformatics Group, Department of Computer Science, University of Freiburg, Germany 4Centre for Biological Systems Analysis (ZBSA), University of Freiburg, Germany.
authors.forename: Björn Andreas
authors.initials: BA
authors.lastname: Grüning
authors.affiliation: GenXPro GmbH, Frankfurt Innovation Center Biotechnology, Frankfurt, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Horres
authors.affiliation: Bioinformatics Group, Department of Computer Science, University of Freiburg, Germany 4Centre for Biological Systems Analysis (ZBSA), University of Freiburg, Germany 5Centre for Biological Signalling Studies (BIOSS), University of Freiburg, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Backofen
authors.affiliation: Eye Center Freiburg University Medical Center, Freiburg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Reinhard
authors.affiliation: Eye Center Freiburg University Medical Center, Freiburg, Germany.
authors.forename: Günther
authors.initials: G
authors.lastname: Schlunck
mesh_terms: D000368:Aged; D001082:Aqueous Humor / Q000378:metabolism; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods; D005786:Gene Expression Regulation; D005901:Glaucoma / Q000235:genetics* / Q000378:metabolism; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics*; D008875:Middle Aged; D012313:RNA / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D012016:Reference Values
chemical_list: C505362:MIRN16 microRNA, human; C524940:MIRN21 microRNA, human; C531201:MIRN451 microRNA, human; D035683:MicroRNAs; D012313:RNA