Mechanisms of bradykinin-induced contraction in human fetal lung fibroblasts.
Bradykinin (BK) induces fibroblast contraction but the structural changes and intracellular mechanisms involved have not been completely explored. We stimulated HFL-1 fibroblasts with BK to assess: 1) fibroblast contractility; 2) the role of α-smooth muscle actin (SMA) in contraction by small interfering RNA (siRNA); 3) α-SMA protein expression; 4) α-SMA and F-actin structure; 5) intracellular calcium concentration ([Ca(2+)](i)); and 6) phosphorylated myosin light-chain (pMLC) and MLC kinase (MLCK) expression. BK triggered concentration- and time-dependent fibroblast gel contraction in conjunction with α-SMA over expression, but not in α-SMA-siRNA-treated cells. BK also increased α-SMA(+) and F-actin(+) cell number and stress fibre polymerisation (detectable at 5-60 min). These BK-induced changes were associated with an increase in [Ca(2+)](i), which peaked within 15 s, and activation of pMLC, which was detectable at 5-60 min. No MLCK content modification was observed. The different manifestations of the BK-induced fibroblast activation were downregulated at different levels (25-100%) by HOE140, a specific BK B2 receptor (B2R) antagonist and by the Ca(2+) chelator, EGTA. Thus, BK-induced fibroblast contraction, associated with differentiation into α-SMA(+) myofibroblasts, is mediated through the activation of the B2R and involves the Ca(2+)/calmodulin pMLC-dependent pathway.
Petecchia
L
L
Pulmonary Diseases Unit, G Gaslini Institute, 16147 Genoa, Italy.
Sabatini
F
F
Usai
C
C
Carnevali
S
S
Ognibene
M
M
Vanni
C
C
Eva
A
A
Fabbri
L M
LM
Rossi
G A
GA
Ricciardolo
F L M
FL
D000199:Actins / Q000378:metabolism; D001920:Bradykinin / Q000494:pharmacology*; D002454:Cell Differentiation; D003094:Collagen / Q000737:chemistry; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008168:Lung / Q000187:drug effects* / Q000196:embryology*; D008856:Microscopy, Fluorescence / Q000379:methods; D009130:Muscle, Smooth / Q000166:cytology; D009218:Myosins / Q000737:chemistry; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000378:metabolism; D013997:Time Factors; D014665:Vasodilator Agents / Q000494:pharmacology*
D000199:Actins; D034741:RNA, Small Interfering; D014665:Vasodilator Agents; D003094:Collagen; D009218:Myosins; D001920:Bradykinin