Background Rheumatoid arthritis (RA) is associated with altered miRNA expression profile in peripheral blood, synovial fluid and synovial membrane. There are several reports on the role of miR-223 in the disease pathogenesis but data on its use as biomarkers in the clinical practice are lacking.
Objectives To analyze the expression of miR-223 in peripheral blood (PB) and synovial fluid (SF) from RA patients in regard to its use as biomarker for disease activity and severity.
Methods Total RNA including miRNAs was isolated from the PB of 57 RA patients and 20 healthy controls (HCs) using PAX gene tubes and from paired SF samples from 44 patients and 11 controls by RNeasy Protect Cell Mini kit (according to manufacturer's protocol). Reverse transcription was performed in order to synthesize cDNA by using miScript II RT kit. Expression of miR-223 was analyzed by relative quantitation method (2^-(ΔΔCq)) using quantitative real-time polymerase chain reaction (qPCR). RNU6B gene was used as reference control for normalization. Data were analyzed by SPSS. The results were related to the clinical presentation and laboratory data for disease activity. The latter was assessed using disease activity score 28 (DAS28).
Results MiR-223 was underexpressed in 65.91% of RA SF samples compared to PB samples. We found statistically significant (p<0.05) overexpression of miR-223 in 81.82% of the SF samples and 35.1% of the PB samples from RA patients compared to HCs. Receiver operating characteristic (ROC) curve analysis was constructed in order to evaluate the diagnostic accuracy of miR-223 in SF by using relative expression (RQ) values. Area under the curve (AUC) was 0.841 (95 CI: 0.727-0.956) with 81.4% sensitivity and 72.7% specificity when RQ value is greater than or equal to 2. The result was statistically significant (p=0.001). Levels of miR-223 in SF correlate with laboratory markers for disease activity such as ESR (p<0.01 two tailed) and CRP (p=0.051 two tailed), with the swollen joint count (p=0.029 two tailed), VAS (0.027 two tailed) and DAS28 score (p=0.003 two tailed). The Spearman's correlation coefficients were 0.463, 0.338, 0.300, 0.303 and 0.402 respectively. No correlations between miR-223 levels in blood and clinical pathological characteristics were observed. Independent T test analysis showed that miR-223 expression in blood is associated with the RA diagnosis.
Conclusions MiR-223 is a potential biomarker for diagnosis with high sensitivity and specificity and for evaluating the activity of RA as its expression is altered significantly in synovial fluid and correlates with diseases activity measured by DAS28 score. We did not observed correlation between the expression of miR-223 in blood and the disease activity but we found week correlation with the RA diagnosis.
Disclosure of Interest None declared