Objective: Fas is a positive regulator of Th17 cells differentiation in experimental autoimmune encephalomyelitis (EAE). However, its upstream regulators are still not fully determined. This study was designed to explore the upstream regulators of Fas in regulating Th17 cells differentiation in EAE., Methods: The mouse model of EAE was established by myelin oligodendrocyte glycoprotein injection. Th17 cells differentiation was induced by IL-23, IL-6 and TGF-β., Results: Down-regulated Hsp70 and miR-374c and up-regulated Fas were observed in the spleen and brain of EAE mice. Hsp70 overexpression evidently reduced Fas protein level, but not mRNA level. The luciferase reporter assay indicated that miR-374c targets Fas. Overexpression of miR-374c down-regulated the mRNA and protein level of Fas. The concentration of IL-17A in CD4+ T-cells was reduced by miR-374c or Hsp70 overexpression, and Fas overexpression altered this trend. Hsp70 did not regulate the expression of miR-374c, and likewise, miR-374c did not regulate the expression of Hsp70. Further results suggested that Hsp70 and miR-374c regulated Fas expression through different ways to affect Th17 cells differentiation in EAE., Conclusions: This study suggested that down-regulated miR-374c and Hsp70 promote Th17 cell differentiation by inducing Fas expression in EAE., Competing Interests: Declaration of competing interest All authors declare that they have no conflicts of interest to this work., (Copyright © 2019 Elsevier B.V. All rights reserved.)