B Cell-Related Circulating MicroRNAs With the Potential Value of Biomarkers in the Differential Diagnosis, and Distinguishment Between the Disease Activity and Lupus Nephritis for Systemic Lupus Erythematosus

Abstract

Our understanding of circulating microRNAs (miRNAs) related to systemic lupus erythematosus (SLE) remains very limited. In this study, we screened SLE-specific miRNAs in plasma from 42 B cell-related miRNAs by using miRNA PCR Array. The selected miRNAs were first confirmed in plasma samples from 50 SLE patients, 16 rheumatoid arthritis (RA) patients, and 20 healthy donors using qRT-PCR. We then investigated the relationship between expressions of the selected miRNAs and SLE clinical indicators. As a result, 14 miRNAs (miR-103, miR-150, miR-20a, miR-223, miR-27a, miR-15b, miR-16, miR-181a, miR-19b, miR-22, miR-23a, miR-25, miR-92a, and miR-93) were significantly decreased in the plasma of SLE patients compared with healthy controls (P < 0.05) and could act as the diagnostic signature to distinguish SLE patients from healthy donors. Six miRNAs (miR-92a, miR-27a, miR-19b, miR-23a, miR-223, and miR-16) expressed in plasma were significantly lower in SLE patients than in RA patients (P < 0.05), revealing the potentially diagnostic signature to distinguish SLE patients from RA patients. Furthermore, the downregulated expression of miR-19b, miR-25, miR-93, and miR-15b was associated with SLE disease activity (P < 0.05) while miR-15b and miR-22 expressions were significantly lower in SLE patients with low estimate glomerular filtration rate (eGFR < 60 ml/min/1.73 m²) (P < 0.05). The diagnostic potential of miR-15b for SLE disease activity and lupus nephritis (LN) with low eGFR was validated on an independent validation set with 69 SLE patients and a cross-validation set with 80 SLE patients. In summary, the signature of circulating miRNAs will provide novel biomarkers for the diagnosis of SLE and evaluation of disease activity and LN.

Keywords: biomarker; circulating microRNA; diagnosis; miR-15b; systemic lupus erythematosus.

Figure 1

A flowchart of the study design.

Figure 2

15 microRNA (miRNA) differentially expressed in the plasma of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA) patients, and healthy control (HC). Expressions of the selected miRNAs in the plasma obtained from patients with SLE (n = 50), RA (n = 16), and HC (n = 20) were determined by qRT-PCR. The expression levels of miRNAs were normalized to cel-miR-39.

Figure 3

Receiver operating characteristic (ROC) curve of 14 microRNAs (miRNAs) for systemic lupus erythematosus (SLE) diagnosis from healthy control. The areas under curve (AUC) for miRNAs are shown in Figure 3 and Table 4, respectively. The diagnostic sensitivity for SLE difference from healthy people was 0.61~0.97 and the specificity was 0.61~1.

Figure 4

Receiver operating characteristic (ROC) curve of six microRNAs (miRNAs) for systemic lupus erythematosus (SLE) diagnosis from rheumatoid arthritis patients. The areas under curve (AUC) for miRNAs are shown in Figure 4 and Table 5, respectively. The diagnostic sensitivity for SLE difference from healthy people was 0.59~0.87 and the specificity was 0.71~0.94.

Figure 5

Expressions of four plasma microRNAs (miRNAs) in patients with active systemic lupus erythematosus (SLE). (A) Four plasma miRNAs expressed differentially in the active SLE and stable SLE patients. Expressions of selected miRNAs in the plasma obtained from patients with SLE (n = 50) were determined by RT-qPCR. The expression levels of miRNAs were normalized to cel-miR-39. (B) Receiver operating characteristic curve analysis of four plasma miRNAs expressed in the active SLE and stable SLE patients.

Figure 6

Receiver operating characteristic curve analysis of miR-15b and miR-22 microRNAs (miRNAs) expression in patients with lupus nephritis.

Figure 7

Expression of miR-15b in double validation group. (A) Plasma miR-15b expression of 69 samples from First Affiliated Hospital of Wenzhou Medical University. Plasma miR-15b was significantly different in active systemic lupus erythematosus (SLE) patients. (B) Receiver operating characteristic (ROC) curves with corresponding AUC for miR-15b in discriminating patients (n = 69) with active SLE from stable SLE. (C) Comparison of plasma miR-15b expression level in SLE patients (n = 69) with lupus nephritis (LN). (D) MiR-15b expressions of 80 serum samples from Ruijin Hospital of Shanghai Jiao Tong University. MiR-15b was significantly different in active SLE patients. (E) ROC curves with corresponding AUC for miR-15b in discriminating patients (n = 80) with active SLE from stable SLE. (F) Comparison of plasma miR-15b expression level in SLE patients (n = 69) with LN.