Bioinformatics analysis reveals different gene expression patterns in the annulus fibrosis and nucleus pulpous during intervertebral disc degeneration

Abstract

Degeneration of the intervertebral disc (IVD), which consists of the annulus fibrosus (AF) and nucleus pulposus (NP), is a multifactorial physiological process associated with lower back pain. Despite decades of research, the knowledge of the underlying molecular mechanisms of IVD degeneration (IDD) has remained limited. The present study aimed to reveal the differential gene expression patterns in AF and NP during the process of IDD and to identify key biomarkers contributing to these differences. The microarray dataset GSE70362 containing 24 AF and 24 NP samples was retrieved from the Gene Expression Omnibus database. Of these, 8 healthy samples were discarded. GeneSpring11.5 software was employed to identify differentially expressed genes (DEGs). Metascape online tools were used to perform enrichment analyses. Finally, the DEGs were mapped with the Search Tool for the Retrieval of Interacting Genes, and a protein-protein interaction (PPI) network was constructed in Cytoscape software. A total of 87 DEGs were identified. Gene ontology enrichment revealed that these DEGs were mainly involved in the inflammatory response, the extracellular matrix and RNA polymerase II transcription factor activity. Pathway enrichment revealed that the DEGs were mainly involved in the transforming growth factor (TGF-β) and estrogen signaling pathways. Matrix metalloproteinase (MMP)1 and interleukin (IL)6 were included in the genes enriched in rheumatoid arthritis, whereas bone morphogenetic protein (BMP)2 and thrombospondin 1 (THBS1) were among the genes enriched in the TGF-β signaling pathway. In the PPI network, IL6 was identified as the central gene. In conclusion, as MMP1 has been demonstrated degrade collagen III at higher rates compared with other types of collagen (which is at a higher quantity in AF than NP), collagen types may be in different distribution patterns, which may contribute to the upregulation of MMP1 in AF. Differences in the expression of BMP2, ESR1 and THBS1 may explain for the pathological differences between AF and NP. IL6 may have a key role in different degeneration processes in AF and NP.

Keywords: differential expression; enrichment analysis; protein-protein interaction network.

Figure 1.

Volcano plot displaying the DEGs. The expression levels of 1,514 genes were significantly different between AF and NP (P<0.05). A total of 87 genes were identified as DEGs (P<0.05, absolute FC>2; red squares). The P-value was adjusted using the Benjamini-Hochberg procedure. FC, fold change; NP, nucleus pulposus; AF, annulus fibrosus; DEG, differentially expressed gene.

Figure 2.

Heatmap displaying hierarchical clustering of the 87 differentially expressed genes in annulus fibrosus and nucleus pulposus (red, upregulated; green, downregulated). The original data were normalized using the z-score to indexes between −3 and 3.

Figure 3.

The top 30 ranked GO terms according to gene count. ‘Qvalue’ is the P-value adjusted using the Benjamini-Hochberg procedure. ‘Log (Qvalue)’ is the log 10 of the q-value. ‘Gene count’ is the number of genes enriched in a GO term. ‘Gene ratio’ is the percentage of total DEGs in the given GO term (only input genes with at least one GO term annotation were included in the calculation). GO, gene ontology; BP, biological process; MF, molecular function; CC, cellular component.

Figure 4.

Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis. hsa, Homo sapiens; TGF, transforming growth factor; TNF, tumor necrosis factor; TRP, transient receptor potential.

Figure 5.

Protein-protein interaction network of differentially expressed genes. All nodes with a combined interaction score of >0.4 are displayed. The blue nodes have a central role with a degree of >5.0.

Figure 6.

Significant modules were screened with the following criteria: Degree centrality cutoff, 2; node score cutoff, 0.2; k-core, 2; max depth, 100.