The tRNA-derived small RNAs (tsRNAs) expression profiles in human aortic vascular smooth muscle cells (HAVSMCs)

Purpose: Next-generation sequencing (NGS) is the main way to mine ncRNA. The purpose of this study is to screen out tRNA-derived small RNAs (tsRNAs) that are differentially expressed in proliferative and quiescent HASMCs Methods: tsRNAs profiles of proliferative and quiescent HASMCs were generated by deep sequencing, in triplicate, using Illumina NextSeq 500. The sequence reads that passed Illumina chastity filter were used for the following analysis. Trimmed reads (with 5ʹ, 3ʹ-adaptor bases removed) were aligned to mature-tRNA and pre-tRNA reference sequences. The alignment statistical analysis was applied to retain the valid sequences for subsequent tsRNAs expression profiles and differential expression analysis. Results: Using an optimized data analysis workflow, we mapped about 8 million sequence reads per sample to mature-tRNA and pre-tRNA sequences from GtRNAdb and identified 3,891 tsRNAs in the proliferative and quiescent HASMCs. According to the standardized TPM, 887 up-regulated and 951 down-regulated tsRNAs were found in the proliferative HASMCs (fold change > 2, P < 0.05). HASMCs were cultured using Smooth Muscle Cell Medium containing 2% fetal bovine serum, 1% 100 × Smooth Muscle Cell Growth Supplement and 1% 100 × Penicillin/Streptomycin Solution at 37°C in a humidified chamber with 5% CO2 and 95% O2. Quiescent HASMCs were induced by FBS starvation for 24 hours.