Fig. 1

Nestin regulates actin polymerization and smooth muscle contraction without affecting myosin light chain phosphorylation at Ser-19. A Protein expression of human airway smooth muscle (HASM) cells expressing control (Ctrl) or nestin shRNA was evaluated by immunoblot analysis. Nestin shRNA reduces nestin expression in HASM cells. Data are means ± SE (n = 4 batches of culture from 3 donors). B Ctrl and nestin knockdown (KD) HASM cells were treated with 10^–4 M acetylcholine (ACh) for 5 min, or left untreated (UT). F-actin/G-actin ratios in cells were evaluated using the fractionation assay. Data are means ± SE (n = 5 batches of culture from 3 donors). C Myosin light chain (MLC) phosphorylation at Ser-19 in Ctrl cells and nestin KD cells was assessed by immunoblot analysis. Basal and ACh-induced MLC phosphorylation was similar in Ctrl cells and nestin KD cells (NS, not significant). Data are means ± SE (n = 4–5 batches of culture from 3 donors). D Immunoblots showing the effects of nestin siRNA on protein expression in human PLCS. Data are means ± SE (n = 3 slices from 3 donors). E Nestin KD reduces airway constriction of human PLCS. Data are means ± SE (n = 4–6 slices from 3 donors). *p < 0.05; **p < 0.01. Student’s t-test was used for statistical analysis of A, D and E. Two-way ANOVA was used for statistical analysis of B and C