Transcriptome, proteome, and metabolome of lens fiber cells from mice exposed to cigarette smoke

We adopted an omics (transcriptome, proteome, and metabolome) approach to characterize the differentially regulated molecules in mouse lens fiber cells exposed to cigarette smoke (CS). Eight pregnant female mice (gestation days 19-20) were placed in a smoking chamber that served as a CS-exposed group. The mothers and newborn pups were exposed to CS for five hours/day, five days/week for 110 days. In parallel, mice were kept in normal cages that served as a control group. The ocular lenses were isolated and fiber cells were separated from the lens epithelium under a microscope. The control and CS-exposed groups were maintained in four biological replicates each consisting of a pool of lens fiber cells from four eyes. The eight biological replicates (four control and four CS-exposed) of fiber cells were used for the next-generation-based transcriptome (RNA-Seq) sequencing, mass-spectrometry-based proteome, and metabolome profiling. RNA-Seq analysis identified the expression (≥1.0 FPKM) of 9,590 and 9,531 genes in control and CS-exposed fiber cells, respectively. The analysis identified 348 differentially expressed genes, which included 186 downregulated and 162 upregulated genes in CS-exposed fiber cells. Proteome profiling revealed a total of 2,424 proteins in control and CS-exposed fiber cells. Metabolome profiling identified a total of 280 metabolites, marked with decreased levels of branched-chain amino acids (BCAAs)-related metabolites in CS-exposed fiber cells. In conclusion, we have established a comprehensive omics profile of CS-exposed lens fiber cells. To the best of our knowledge, this is the first report investigating a comprehensive omics profile of CS-exposed fiber cells. Next-generation (NG)-based whole transcriptome sequencing (RNA-Seq) was performed for lens fiber cells isolated from control (CT-FC) and cigarette smoke (CS-FC) exposed mouse eyes. Total RNA was subjected to RNA-Seq library preparation using NEB Next Ultra RNA Library Prep Kit. Eight RNA-Seq bar-coded pooled libraries were sequenced (2 × 150 bp) on a HiSeq 2500. The raw reads (FASTQ) were processed and analyzed using combinations of algorithms (STAR, HTSeq, and DESeq2). The paired-end reads were aligned to the Mus musculus genome (GRCm38/mm10) using the STAR algorithm (Ver. 2.5). The HTSeq (Ver. 0.6.1) was used for the quantification of gene expression using mapped reads. The expression data were normalized by calculating the fragment per kilobase per million mapped reads (FPKM) values for each gene. Differential gene expression analysis was performed using the DESeq2.