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After a 24 h incubation period, the CM was collected and freshly used for cell treatment or stored at -80 ºC until used for ELISA measurements
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However, recent studies suggest that methotrexate also has anti-inflammatory, immune-modulating, and anti-proliferative effects by regulating IL-6 synthesis and reactive oxygen species (ROS) production, showing significant efficacy in treating immune-related diseases such as vitiligo, psoriasis, and rheumatoid arthritis (Speeckaert and Geel, 2023)
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