Identification of long non-coding RNAs GAS5, linc0597 and lnc-DC in plasma as novel biomarkers for systemic lupus erythematosus

Abstract

Despite increasing evidence that long non-coding RNAs (lncRNAs) widely take part in human diseases, the role of lncRNAs in systemic lupus erythematosus (SLE) is largely unknown. In this study, we performed a two-stage study to explore the plasma levels of five lncRNAs (GAS5, linc0949, linc0597, HOTAIRM1 and lnc-DC) and their potential as SLE biomarkers. Compared with healthy controls, plasma levels of GAS5 and lnc-DC were significantly decreased (P < 0.001 and P = 0.002, respectively) while linc0597 were overexpressed in SLE patients (P < 0.001). When SLE patients were divided into SLE without nephritis and lupus nephritis (LN), the levels of lnc-DC were significantly higher in LN compared with SLE without nephritis (P = 0.018), but no significant difference in levels of GAS5 and linc0597 were found between LN and SLE without nephritis; plasma linc0949 level showed no significant difference in all comparisons. Further evaluation on potential biomarkers showed that GAS5, linc0597 and lnc-DC may specifically identify patients with SLE, the combination of GAS5 and linc0597 provided better diagnostic accuracy; lnc-DC may discriminate LN from SLE without nephritis. In summary, GAS5, linc0597 and lnc-DC in plasma could be potential biomarkers for SLE.

Keywords: biomarker; diagnosis; long non-coding RNA; systemic lupus erythematosus.

Figure 1. Validation of candidate lncRNAs (GAS5, linc0949, linc0597 and lnc-DC) identified in the first stage screening in plasma of SLE and healthy subjects

Each symbol represents an individual subjects; horizontal lines indicate median values. The expression levels of the four candidate lncRNAs in 163 SLE patients, 80 healthy controls were analyzed by qRT-PCR and normalized by GAPDH. (A) Decreased expression of GAS5 in total SLE patients vs healthy controls. (B) Decreased expression of GAS5 in both SLE patients with and without nephritis vs healthy controls, but no significant difference in GAS5 between SLE patients with and without nephritis. (C) No significant difference in linc0949 between total SLE patients and healthy controls. (D) No significant difference in linc0949 among healthy controls, SLE patients with and without nephritis. (E) Increased expression of linc0597 in total SLE patients vs healthy controls. (F) Increased expression of linc0597 in both SLE patients with and without nephritis vs healthy controls, but no significant difference in linc0597 between SLE patients with and without nephritis. (G) Decreased expression of lnc-DC in total SLE patients vs healthy controls. (H) Decreased expression of lnc-DC in SLE patients without nephritis vs healthy controls and in SLE patients without nephritis vs SLE patients with nephritis, but no significant difference in lnc-DC in SLE patients with nephritis vs healthy controls. SLE: systemic lupus erythematosus; HC: healthy controls; NS: not significant; ^*** P< 0.001, ^* P< 0.01, * P< 0.05.

Figure 2. Receiver operating characteristic (ROC) curve analysis of lncRNAs for the discriminative ability of SLE patients vs healthy controls and SLE with nephritis vs SLE without nephritis

(A) lnc-DC alone, linc0597 alone, GAS5 alone and GAS5 combined with linc0597 for the discriminative ability of SLE patients vs healthy controls in the validation set. (B) linc0597 alone, GAS5 alone and GAS5 combined with linc0597 for the discriminative ability of SLE patients vs healthy controls in the combination set. (C) lnc-DC alone for the discriminative ability of SLE with nephritis vs SLE without nephritis in the combination set. (D) lnc-DC combined with GAS5 for the discriminative ability of SLE with nephritis vs SLE without nephritis in the combination set.

Figure 3. Relative expression of lncRNAs in the validation set of patients with SLE and disease controls

Each symbol represents an individual subjects; horizontal lines indicate median values. (A) GAS5, (B) linc0597, (C) lnc-DC. SLE: systemic lupus erythematosus; RA: rheumatoid arthritis; SS: Sjögren's syndrome; NS: not significant; ^*** P< 0.001, * P< 0.05.

Figure 4. Receiver operating characteristic (ROC) curve analysis of GAS5 combined with linc0597 for the risk-score in

(A) SLE patients in the validation set vs all controls (healthy controls in the validation set, RA and SS), (B) SLE patients in the validation set vs RA and SS, (C) combination set.