The gut barrier acts as a critical defense mechanism for the host, protecting it from the external environment and tightly regulating the interaction between the commensal microbiota and the intestinal immune system. Increasing lines of evidence demonstrated alterations of the gut microbiota composition with loss of the intestinal barrier integrity and abnormal activation of mucosal immunity in several autoimmune diseases, including multiple sclerosis (MS). These findings suggest that this intricate system plays an important role in influencing autoimmune responses even at sites distant from the intestine, such as the central nervous system (CNS) in MS. In this chapter, we review the current understanding of how alterations in the intestinal barrier integrity together with gut inflammation affect the interplay between the commensal microbiota and the mucosal immune system and how this interaction impacts on CNS autoimmunity.