Figure 9.

Impact of the NBD peptide on cellular responses to TLR2 signaling. A through F, Cellular responses to Pam3CSK4, a specific synthetic ligand for TLR2, under treatment of the NBD peptide or the control peptide were assessed after 48‐hour stimulation in cardiomyocytes (A and D) and after 24‐hour stimulation in cardiac fibroblasts (B and E) and vascular endothelial cells (C and F). A, Cell surface areas of cardiomyocytes. Cell surface areas of 25 cardiomyocytes were measured in specimens with anti‐sarcomeric α‐actinin staining in each group. Scale bars=20 μm. B and C, Proliferation of fibroblasts (B) and vascular endothelial cells (C). Cell proliferation was assessed by means of dimethylthiazol‐carboxymethoxyphenyl‐sulfophenyl‐tetrazolium assay and was expressed as the percentage of the absorbance to the well with cells treated by control and the control peptide. Data were obtained from 7 independent experiments in fibroblasts and 6 independent experiments in vascular endothelial cells. D through F, Expression levels of IL‐1β (Il1β), IL‐6 (Il6), and TNF‐α (Tnfα) mRNAs were measured in cardiomyocytes (D), cardiac fibroblasts (E), and vascular endothelial cells (F). Three samples were obtained from cells cultured in 6‐well plates in each group. Expression levels of each gene were normalized to those of GAPDH (Gapdh). *P<0.05 vs control; ^†P<0.05 vs the control peptide. IL indicates interleukin; GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; NBD, NEMO‐binding domain; NF, nuclear factor; TLR, Toll‐like receptor; TNF, tumor necrosis factor.