Experimental autoimmune encephalomyelitis in mice

C57BL/6J mice were purchased from Harlan (Borchen, Germany) and housed at the in-house animal care facility of the University of Erlangen under standardized conditions. EAE induction was done as described before^{9 (link)}. Briefly, male mice were immunized with 200 μg MOG (35–55) (Charite, Berlin, Germany) in an equal amount of complete Freund’s adjuvant and received 200ng pertussis toxin (List Biochemicals, Campbell, CA) intraperitonally (i.p.) on days 0 and 2 post induction (p.i). The clinical evaluation was performed on a daily bases by a 5 point scale ranging from 0, no clinical sign; 1, limp tail; 2, limp tail, impaired righting reflex, and paresis of one limb; 3, hind limb paralysis; 4, hind limb and forelimb paralysis; 5, moribund. Mice received normal chow and tap water ad libitum (control group) or sodium-rich chow containing 4% NaCl (SSNIFF, Soest, Germany) and tap water containing 1% NaCl ad libitum (high-salt group). Inhibition of p38/MAPK in vivo was done as described before^{36 (link)}. In brief, mice were maintained on a control or high-salt diet and either received 1mg/kg/d SB202190 (TOCRIS) i.p. or vehicle from day −3 p.i. of EAE. Brain leukocytes were isolated by percoll gradient centrifugation on day 17 post EAE induction, stimulated by PMA/ionomycin and analysed by flow cytometry for IL-17A and CD4 expression. Mx-Cre⁺/p38α^fl/fl mice^{37 (link)} maintained on C57BL/6 background were a kind gift of Dr. Jean-Pierre David. Mice were injected with 13mg/kg/body-weight Polyinosinic-polycydidylic acid (poly(I:C), Sigma-Aldrich) on days 0, 2, 6 and were sacrificed on day 8 for isolation of splenocytes. Blood pressure analysis was performed by the tail cuff method as described previously^{9 (link)}. All animal experimentation was performed in accordance to the German animal protection law.