Microarray analysis of differentially expressed genes in L929 mouse fibroblast cells exposed to leptin and hypoxia

Abstract

Leptin and hypoxia are pro-fibrotic factors involved in fibrogenesis, however, the gene expression profiles remain to be fully elucidated. The aim of the present study was to investigate the regulatory roles of leptin and hypoxia on the L929 mouse fibroblast cell line. The cells were assigned to a normoxia, normoxia with leptin, hypoxia, and hypoxia with leptin group. The cDNA expression was detected using an Agilent mRNA array platform. The differentially expressed genes (DEGs) in response to leptin and hypoxia were identified using reverse transcription-quantitative polymerase chain reaction analysis, followed by clustering analysis, Gene Ontology analysis and pathway analysis. As a result, 54, 1,507 and 1,502 DEGs were found in response to leptin, hypoxia and the two combined, respectively, among which 52 (96.30%), 467 (30.99%) and 495 (32.96%) of the DEGs were downregulated. The most significant functional terms in response to leptin were meiosis I for biological process (P=0.0041) and synaptonemal complex for cell component (P=0.0013). Only one significant pathway responded to leptin, which was axon guidance (P=0.029). Flow cytometry confirmed that leptin promoted L929 cell proliferation. The most significant functional terms in response to hypoxia were ion binding for molecular function (P=7.8621E-05), glucose metabolic process for biological process (P=0.0008) and cell projection part for cell component (P=0.003). There were 12 pathways, which significantly responded to hypoxia (P<0.05) and the pathway with the highest significance was the chemokine signaling pathway (P=0.0001), which comprised 28 genes, including C-C motif ligand (CCL)1, C-X-C motif ligand (CXCL)9, CXCL10, son of sevenless homolog 1, AKT serine/threonine kinase 2, Rho-associated protein kinase 1, vav guanine nucleotide exchange factor 1, CCL17, arrestin β1 and C-C motif chemokine receptor 2. In conclusion, the present study showed that leptin and hypoxia altered the profiles of gene expression in L929 cells. These findings not only extend the cell spectrum of leptin on cell proliferation, but also improve current understanding of hypoxia in fibroblast cells.

Hypoxia Alters the Expression of CC Chemokines and CC Chemokine Receptors in a Tumor–A Literature Review

“…CCL17/TARC expression is affected by chronic hypoxia, although the nature of these changes depends on the adopted research model. In L929 mouse fibroblasts, chronic hypoxia reduces CCL17/TARC expression [189], while in human hepatoma cells [193] and lung adenocarcinoma cells [13], chronic hypoxia does not affect CCL17/TARC expression. In MDA-MB-231 breast cancer cells, hypoxia increases CCL17/TARC expression but not in the MCF-7 line [142].…”

“…In a tumor cell, chronic hypoxia does not change the expression of CCL1/I-309 as demonstrated by lung adenocarcinoma cells [13] and hepatocellular carcinoma [143]. Nevertheless, in the L929 mouse fibroblast cell line, chronic hypoxia increases expression [189]. In the tumor, CCL1/I-309 expression occurs, among others, in CAF [236,237].…”

“…In hypoxia, CCR2 expression is reduced in mouse L929 fibroblast cells [ 189 ] and human monocytes [ 79 , 144 ]. It has also been shown that chronic hypoxia disrupts the CCL2/MCP-1-dependent chemotaxis of monocytes [ 190 ], which is associated with a chronic hypoxia-induced increase in the expression of MAPK phosphatase 1 (MKP-1) [ 191 ] which reduces the activation of MAPK cascades and chemotaxis dependent on these signaling pathways [ 191 ].…”

ROCK Inhibition as Potential Target for Treatment of Pulmonary Hypertension

“…Lung fibroblasts isolated from patients with idiopathic pulmonary fibrosis also display increased resistance to apoptosis, a phenotype which reflects the higher expression of B cell lymphoma (Bcl)-2 in these cells by activation of the ROCK/MRTF/SRF pathway [ 104 ]. The exposure of fibroblasts to hypoxia leads to ROCK1 gene upregulation, which further contributes to increased migration, proliferation and phenotypic plasticity in those cells by crosstalk between ROCK1 and HIF-1α [ 105 , 106 ]. Recently, a growth factor-independent pathway for fibroblast activation and differentiation was described, which relies on the activation of the signal transducer and activator of transcription (STAT)-3 by ROCK and may contribute to the exacerbation of tissue fibrosis [ 107 ].…”

Altered expression of differential gene and lncRNA in the lower thoracic spinal cord on different time courses of experimental obstructive jaundice model accompanied with altered peripheral nociception in rats

“…The introduction of high-throughput RNA sequencing (RNA-seq), where thousands of differential genes and lncRNA expression patterns can be studied in parallel [ 21 – 24 ], permits a broad assessment for altered expression of differential gene and lncRNA in the lower thoracic spinal cord on different time courses of obstructive jaundice model. In current study, we performed a comprehensive transcriptome analysis in obstructive jaundice model using RNA-seq, and identified lncRNAs with differential expression.…”