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Supplement 1 A B C * ** D E Supplement 1. CpG ODN induced CCN1 in time and dose dependent but TLR9 independent manner. (A) CCN1 was secretion by time dependent.

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Presentation on theme: "Supplement 1 A B C * ** D E Supplement 1. CpG ODN induced CCN1 in time and dose dependent but TLR9 independent manner. (A) CCN1 was secretion by time dependent."— Presentation transcript:

1 Supplement 1 A B C * ** D E Supplement 1. CpG ODN induced CCN1 in time and dose dependent but TLR9 independent manner. (A) CCN1 was secretion by time dependent manner. (B) CCN1 was secreted by ODN dose dependent manner. (C) Each CpG ODN (1 μM) was treated for 4h to Beas2B cells and CCN1 was assayed by ELISA. (D) TLR9 siRNA was transfected to Beas2B cells for 2 days and siRNA efficiency was confirmed by WB. (E) NF-κB signaling inhibitor, Bay , was 1h pre-treated to Beas2B cells and ODN treated for 4h. CCN1 was measured by ELSIA

2 Supplement 2 A B Figure 2. Anti-inflammatory effects of CCN1 in LPS stimulating alveolar macrophages. (A) Level of IL-6 and TNF-α after stimulating LPS with CCN1 in alveolar macrophages (MH-s). *p < .05, compared to LPS (B) Reversed IL-6 and TNF-α secretion level by neutralizing CCN1 antibody in LPS+CCN1 stimulating alveolar macrophages (MH-s). *p < .05, compared to LPS+CCN1

3 Supplement 3 B A * * Supplement 2
(A) CCN1 was treated to A549 cells for 4h and IL-10 was measured by ELISA (B) CCN1 (1 μg/ml) was treated to Beas2B cells for 6h and mRNA expression of integrin αV and β6 was measured by real-time PCR and then normalized by HPRT

4 Supplement 4 A B * * Supplement 3 Cav-1 and CCN1 secretion
(A) Level of CCN1 secretion by Cav-1 inhibitor, Daidzen, with ODN stimulation, Daidzein was pre-treated for 3h. (B) Cav-1 over-expression plasmid was transfected into Beas2B cells for 2 days and CCN1 was measured by ELISA

5 Supplement 5 A B C β-actin BiP siRNA - + β-actin JNK siRNA - + D E F
Supplement 4. Expression of BiP and CHOP was independent on Src activation (A) Src siRNA was transfected to Beas2B cells and ODN (1 μM) was treated. BiP, CHOP, Src and β-actin were detected by WB (B) PP2 was 1h pre-treated and then ODN treated. BiP, CHOP, Src (Y527), Src and β-actin were detected by WB (C) Each BiP and JNK siRNA were transfected to Beas2B cells and siRNA efficiency was confirmed by WB (D) JNK inhibitor was 1h pre-treated and then ODN treated to Beas2B cells. JNK, pJNK, Cav-1, pCav-1 and β-actin were detected by WB (E) PP2 was 1h pre-treated and ODN treated to time points. Src (Y527) regulated pJNK expression (F) Cav-1 siRNA was transfected and ODN treated. Cav-1 siRNA was independent of pJNK expression.

6 Neutrophil infiltration
Supplement 6 A B CCN1 CCN1 SP IGFbp vWF αV β6 Src Cav-1 PKC CpG ODN (5~30 min) CCN1 IGFbp vWF 4 IL-10 ER stress Cav-1 1 2 Src BiP 5 3 P(Y527) JNK CpG ODN (45~60 min) Supplement 6. Schemata (A) CCN1 vWF MIP-2, TNF-α IGFbp 7 Cav-1 Neutrophil infiltration Src p P(Y14) 6 JNK 8 ALI CCN1 secretion

7 Supplement 6 C 1 4 5 3 6 7 2 K.pneumoniae αVβ6 DNA/CpG ODN IL-10 CCN1
TNF-α


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