Vitiligo is a common acquired, chronic dermatological disease with loss of functioning melanocytes, affecting skin, hair, mucosa, or all. Non‐segmental vitiligo (NSV) is the most common form of vitiligo Melanocyte destruction under oxidative stress may be promoted by microRNA-25-5p (miR-25-5p), which inhibits keratinocytes' production and release of growth factors stem cell factor (SCF) and basic fibroblast growth factor (bFGF). Microphthalmia-associated transcription factor (MITF) is a primary regulator of the function and survival of melanocytes. 48 NSV sufferers and 48 healthy control participants were enrolled in this study. Peripheral blood samples were taken from each subject to measure the messenger RNA (mRNA) expression levels of MITF and miR-25-5p. This was done by using the real-time reverse transcriptase polymerase chain reaction (RT-PCR). The current study investigated the expression profiles of miR-25-5p and MITF in NSV patients compared to healthy controls. The results indicated that NSV patients exhibited a significant upregulation of miR-25-5p expression compared to the control group. Conversely, MITF expression levels were obviously lower in the NSV cohort. Furthermore, analysis within the NSV patient group revealed a negative correlation between miR-25-5p and MITF expression levels, suggesting an inverse relationship. Additionally, miR-25-5p expression displayed a positive correlation with disease activity as measured by the Vitiligo Disease Activity (VIDA) score. Conversely, MITF expression exhibited a negative correlation with VIDA score. Interestingly, no significant correlation was observed between gene expression and the Vitiligo Area Scoring Index (VASI) score, a marker of disease severity. It was concluded from the results that MITF and miR-25-5p are involved in the pathogenesis of NSV.