According to manufacturers' instructions, total RNA was isolated using TRIzol (Invitrogen) and purified with RNeasy mini kit (QIAGEN). RNA quality and quantity were measured on a NanoDrop spectrophotometer (ND-1000, Nanodrop Technologies) and listed in Supplementary Table 1 . After quality control, the miRCURY™ Hy3™/Hy5™ Power labeling kit (Exiqon, Vedbaek, Denmark) was used according to the manufacturer's guideline for microRNA labeling. After hybridized Hy3™-labeled samples on a miRCURYTM LNA Array (v.18.0) (Exiqon) and washed slides several times with a Wash buffer kit (Exiqon), the slides were scanned on an Axon GenePix 4000B microarray scanner (Axon Instruments, Foster City, CA). Scanned images were then imported into the GenePix Pro 6.0 software (Axon) for grid alignment and data extraction. Replicated microRNAs were averaged, and probes with intensities ≥30 in all samples were selected for normalization. Detailed instructions were as previously described (Liu et al., 2016 (link)). After normalization, significantly differentially expressed microRNAs between the two groups were identified using Fold change and P-value cutoffs of 2 and 0.05, respectively.
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