Fig. 1

Identification of lncRNAs in human HSC myofibroblasts. a ACTA2, LOX, and LOXL2 mRNA levels were quantified in HSC myofibroblasts cultured on plastic (blue) and compared with mRNA levels in HSC myofibroblasts that were cultured in Matrigel for 3 days to induce reversion to an inactive phenotype (green). Samples were normalized using GAPDH, and expression of each gene in HSC myofibroblasts was set to 1. Error bars represent standard deviation. This experiment is representative of three biological replicates. b Culture in Matrigel leads to accumulation of fat droplets characteristic of HSCs that have reverted to an inactive phenotype. Microscopy images are shown for HSC myofibroblasts cultured without (No Matrigel) and with Matrigel for 3 days. Light microscopy images (left) show that the cells grown in Matrigel become rounded and clump together. Bodipy staining for neutral fat (green fluorescence, right) shows accumulation of lipid droplets in HSCs cultured in Matrigel. Images are shown at 10× magnification. c COL1A1 mRNA levels were quantified in HSC myofibroblasts cultured in 0.2 % bovine serum albumin and after treatment with TGF-β for 16 h. d Pipeline for identification of lncRNAs (see the “ Methods ” for details). miRNA microRNA. e Examples of lncRNAs identified in HSC myofibroblasts. lncRNA-054349 is a novel lncRNA discovered in HSCs (top) and EPHA5-AS1 is an lncRNA expressed in HSCs that is expressed antisense to EPHA5 (blue). For each lncRNA track, H3K4me3 enrichment (log likelihood ratio (LogLR) value normalized by subtracting the background) marks the site of transcription initiation (black, top). RNA-seq reads supporting lncRNA transcripts are shown in red (RNA-seq sense). Antisense transcripts are shown in blue. The genomic structure for each gene is shown below the RNA-seq tracks. lncRNAs are shown in red and protein-coding genes are shown in blue. Boxes indicate exons, lines indicate introns, and arrows indicate the start and direction of transcription. For EPHA5-AS1, the annotated lncRNA and protein-coding gene are shown below the RNA-seq tracks. The multiple isoforms of EPHA5-AS1 that are identified in HSC myofibroblasts (labeled as HSC lncRNA) are shown at the bottom. Novel lncRNAs identified in this study are named with the prefix “lncRNA” followed by the locus number assigned during assembly