Circulating miR-221/222 and Serum IL-23 in Treatment-Naïve Multiple Sclerosis: A Case-Control Study

Background and Objectives: Circulating microRNAs (miRNAs) are emerging as accessible biomarkers for multiple sclerosis (MS). In experimental models, miR-221 and miR-222 have been linked to immune and Th17-related pathways, while interleukin-23 (IL-23) is a cytokine driving autoimmune inflammation. This observational study evaluated the expression of circulating miR-221/222 and serum IL-23 concentrations in treatment-naive adult patients with MS. Materials and Methods: This prospective, cross-sectional case-control study included 43 untreated adult patients with MS (36 with relapsing-remitting MS and 7 with primary progressive MS) and 37 healthy controls. Demographic features, Expanded Disability Status Scale (EDSS) scores, magnetic resonance imaging involvement, initial symptoms, serum IL-23 concentration, and miR-221/222 expression were recorded. Total RNA, including small RNAs, was isolated; complementary DNA was synthesized by reverse transcription from RNA templates; and reverse transcription-quantitative real-time PCR (RT-qPCR) was performed using RNU6-2 as the endogenous small RNA reference. Individual ΔΔCt values were specified as the primary inferential scale, while 2^-ΔΔCt fold-change was retained only for descriptive reporting. Results: The relative expression (2^-ΔΔCt) of miR-221 and miR-222 was higher in patients than in controls. Within the patient group, miR-221 and miR-222 relative expression did not differ by age, sex, EDSS, time since MS diagnosis, MRI involvement area, or initial symptoms. No statistically significant difference in IL-23 levels was observed between the patient and control groups. Conclusions: The presence of unaltered IL-23 levels alongside significantly elevated miR-221 and miR-222 expressions in treatment-naive MS patients during the remission phase suggests their potential utility as biomarkers for immune regulation. Given the cross-sectional design of this study, no causal or regulatory relationship between miR-221/222 and IL-23 can be inferred.