Circ_0005918 Sponges miR-622 to Aggravate Intervertebral Disc Degeneration

Abstract

Intervertebral discdegeneration (IDD) is the most common cause of lower back pain, but the exact molecular mechanism of IDD is still unknown. Recently, studies have shown that circular RNAs (circRNAs) regulate diverse biological procedures such as cell metastasis, growth, metabolism, migration, apoptosis, and invasion. We demonstrated that IL-1β and TNF-α induced circ_0005918 expression in the NP cell, and circ_0005918 was overexpressed in the IDD group compared with the control group. Moreover, the upregulated expression of circ_0005918 was associated with disc degeneration degree. The elevated expression of circ_0005918 promoted cell growth and ECM degradation, and it induced secretion of inflammatory cytokines including IL-1β, IL-6, and TNF-α. Moreover, we found that circ_0005918 sponged miR-622 in the NP cell. In addition, the exposure to IL-1β and TNF-α suppressed the expression of miR-622, which was downregulated in the IDD group compared with the control group. Furthermore, the downregulated expression of miR-622 was associated with disc degeneration degree. The expression level of miR-622 was negatively associated with circ_0005918 expression in the IDD group. In conclusion, circ_0005918 regulated cell growth, ECM degradation, and secretion of inflammatory cytokines by regulating miR-622 expression. These data suggested that circ_0005918 played important roles in the development of IDD via sponging miR-622.

Keywords: cancers; circ_0005918; circular RNAs; intervertebral disc degeneration; miR-622.

FIGURE 1

Exposure to IL-1β and TNF-α induced circ_0005918 expression. (A) Expression of circ_0005918 was measured by qRT-PCR assay. (B) IL-1β induced circ_0005918 expression in a time-dependent manner. (C) The level of circ_0005918 was detected via qRT-PCR assay. (D) TNF-α induced circ_0005918 expression in a time-dependent manner.

FIGURE 2

Upregulated circ_0005918 levels were associated with IDD. (A) The circ_0005918 level was upregulated 5 NP cells from IDD compared to control. (B) Circ_0005918 was overexpressed in the IDD group compared to the control group. (C) The upregulated expression of circ_0005918 was associated with disc degeneration levels. *p < 0.05 and ***p < 0.001.

FIGURE 3

Circ_0005918 induced cell growth and ECM degradation. (A) The expression of circ_0005918 was measured by qRT-PCR assay. (B) The elevated expression of circ_0005918 promoted cell growth in the NP cell. (C) The overexpression of circ_0005918 promoted MMP-9 expressions in the NP cell. (D) The ectopic expression of circ_0005918 induced MMP-13 expression in the NP cell. (E) The overexpression of circ_0005918 suppressed collagen II expression in the NP cell. *p < 0.05, **p < 0.01, ***p < 0.001.

FIGURE 4

Circ_0005918 induced inflammatory cytokines secretion. (A) The ectopic expression of circ_0005918 promoted IL-1β expression. (B) The level of IL-6 in the cell supernatants was measured by ELISA. (C) The level of TNF-α in the cell supernatants was measured by ELISA. ***p < 0.001.

FIGURE 5

Circ_0005918 regulated miR-622 expression in NP cell. (A) By using Circular RNA Interactome software, it shows that circ_0005918 may sponge miR-622 expression. (B) The expression of miR-622 was measured by qRT-PCR assay. (C) The luciferase reporter indicated the ectopic expression of miR-622 suppressed the luciferase reporter value of circ_0005918 wild reporter vector but did not change the value of the mutant reporter. (D) The overexpression of circ_0005918 inhibited miR-622 expression. **p < 0.01, ***p < 0.001.

FIGURE 6

Exposure to IL-1β and TNF-α suppressed miR-622 expression. (A) The expression of miR-622 was measured by qRT-PCR assay. (B) IL-1β suppressed miR-622 expression in a time-dependent manner. (C) The level of miR-622 was detected via qRT-PCR assay. (D) TNF-α inhibited miR-622 expression in a time-dependent manner.

FIGURE 7

Downregulated miR-622 levels were associated with IDD. (A) miR-622 level was downregulated 5 NP cells from IDD compared to control. (B) miR-622 was lower in the IDD group than the control group. (C) The downregulated expression of miR-622 was associated with disc degeneration levels. (D) The level of miR-622 was negatively associated with circ_0005918 expression in the IDD group. **p < 0.01, ***p < 0.001.

FIGURE 8

Circ_0005918 regulated the cell growth and ECM degradation by regulating miR-622. (A) Cell proliferation was detected by CCK-8 assay. (B) The ectopic expression of miR-622 suppressed MMP-9 expression in the circ_0005918-overexpressing NP cell. (C) The level of MMP-13 was measured by qRT-PCR analysis. (D) The level of collagen II was measured by qRT-PCR analysis. *p < 0.05, **p < 0.01.

FIGURE 9

Circ_0005918 regulated the inflammatory cytokines secretion by regulating miR-622. (A) Elevated expression of circ_0005918 induced IL-1β expression, while miR-622 mimic suppressed this function in the NP cell. (B) The level of IL-6 in the cell supernatants was measured by ELISA. (C) The level of TNF-α in the cell supernatants was measured by ELISA. ***p < 0.001.