BRC-D-13-00319R2
Suppl. Fig. 1: Cardiac APN expression is up-regulated in acute CVB3 myocarditis.
(a) mRNA expression of APN in hearts of WT mice was determined by qRT-PCR on day 3 p.i. (n = 8 animals per group). (b) Cardiac myocytes were infected in vitro with CVB3 (MOI =
1 PFU) and mRNA expression of APN was determined by qRT-PCR at 6 h p.i. (n = 6). (c)
Serum concentrations of APN in WT mice, APN-KO mice and reconstituted APN-KO mice on day 3 p.i., i.e. 10 days after APN-expressing vector (Ad-APN) injection was determined by
ELISA (n = 8 animals per group).
Suppl. Fig. 2: APN-KO mice exhibit reduced virus load, inflammation and fibrosis in subacute CVB3 myocarditis.
(a) CVB3 virus load in hearts of WT and APN-KO mice on day 7 p.i. was determined by qRT-PCR (n = 15 animals per group). (b) mRNA expression of
IFNß, IFN 
, TNF

, IL1ß, IL-6 and IL-12(p40) in hearts of WT and APN-KO mice on day 7 p.i. was determined by qRT-PCR (n = 15 animals per group). (c) mRNA expression of Col1a1 and Col3a1 in hearts of WT and APN-KO mice on day 7 p.i. was determined by qRT-PCR
(n = 10 animals per group).
Suppl. Fig. 3: APN-KO mice exhibit similar CAR expression, virus load and T cell infiltration in acute CVB3 myocarditis.
(a) mRNA expression of CAR in hearts of WT and
APN-KO mice on day 3 post CVB3 infection was determined by qRT-PCR (n = 8 animals per group). (b) CVB3 virus load in hearts of WT and APN-KO mice on day 3 p.i. was determined by plaque assay (left panel) and qRT-PCR (right panel) (n = 8 animals per group). (c) mRNA expression of CD3z in hearts of WT and APN-KO mice on day 3 p.i. was determined by qRT-
PCR (n = 8 animals per group).
Suppl. Fig. 4: APN has no influence on TLR-mediated up-regulation of CD14 expression in cardiac myocytes.
Cardiac myocytes were incubated with APN (10 µg/mL) or vehicle (Albumin 10 µg/mL) for 18 h before stimulation with TLR ligands (a) P(I:C)
(50 µg/mL) and R-848 (5 µg/mL) or (b) LPS (1 µg/mL) for 6 h. mRNA expression of CD14 was determined by qRT-PCR (n = 8).
Suppl. Fig. 5: APN-KO mice exhibit increased expression of pro-inflammatory cytokines and chemokines in acute CVB3 myocarditis. mRNA expression of TNF

,
IL-12(p40), CCL5 and CXCL10 in hearts of WT and APN-KO mice on day 3 p.i. was determined by qRT-PCR (n = 8 animals per group).