*4.3. MicroRNA Expression Profiling in CD14*+ *Monocytes by Next-Generation Sequencing (NGS)*

RNA of CD14+ monocytes was extracted using Direct-zol™ RNA Kits (Zymo Research, Irvine, CA, USA) according to the manufacturer's protocol and measured quantitatively using Agilent Bioanalyzer 2100 O (Agilent, Santa Clara, CA, USA). Only samples with RNA integrity number (RIN) ≥ 8.0 were then used for the TruSeq Small RNA Preparation protocol (Illumina, San Diego, CA, USA). The prepared amplicons were sequenced with a V3 150-cycle sequencing reagent on the MiSeq system (Illumina, San Diego, CA, USA) to generate 51-nt single-end reads. The generated NGS data were first analyzed with miRSeq [29], a toolkit for sequencing quality evaluation and miRNA quantification (in transcripts per million, TPM).
