046 – THE ROLE OF GAP JUNCTION-DEPENDENT CYTOKINES IN THE EAE MODEL OF MULTIPLE SCLEROSIS
Filippos Stavropoulos – Elena Georgiou, Irene Sargiannidou, Kleopas A. Kleopa
The Cyprus Institute of Neurology and Genetics
Connexin 47 (Cx47), and to a lesser extent Cx32, are assembled into Gap Junctions (GJs) and couple oligodendrocytes mainly to astrocytes via heterotypic GJs and to other oligodendrocytes via homotypic GJs. In the demyelinating and inflammatory disease Multiple Sclerosis (MS) these oligodendrocyte connexins appear disrupted. Recently published studies in our lab utilizing mice with Experimental Autoimmune Encephalomyelitis (EAE), a mouse model of MS, demonstrates that the absence of Cx32 or Cx47 affects the severity and progression of the disease and alters the profile of several CNS-inflammation-related cytokines. From these 64 cytokines tested in wild type (WT) EAE, Cx32-knockout (KO) EAE, and Cx47-KO EAE mice, we selected the most relevant and altered molecules, GM-CSF, G-CSF, CCL2, and VCAM-1 for further testing in the same mouse genotypes. Their expression was analyzed at 7, 12, and 24 days post injection (dpi) using fluorescent microscopy, real-time PCR, and Western blot analysis. This analysis showed that Cx47-KO EAE mice had the greatest alteration in cytokine levels at almost all disease time points. Moreover, while Vcam-1 mRNA levels were correlated with its protein levels at 7dpi, Ccl2 mRNA and protein levels were inversely correlated, denoting a possible post-transcriptional regulation of Ccl2. Further experiments will shed light on the relationship between connexins and neuroinflammation in the context of MS/ EAE and will provide a better understanding of the inflammatory process in patients with Cx47 and Cx32 mutations.