- 作者: Qifeng Jiang; Renjian Huang; Shaoxi Cai; Chih-Lueh A Wang
- 作者服務機構: Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing, China
- 中文摘要: --
- 英文摘要:
Background:
Migration of vascular smooth muscle cells (SMCs) from the media to intima
constitutes a critical step in the development of proliferative vascular diseases. To
elucidate the regulatory mechanism of vacular SMC motility, the roles of caldesmon
(CaD) and its phosphorylation were investigated.
Methods:
We have performed Transwell migration assays, immunofluorescence microscopy,
traction microscopy and cell rounding assays using A7r5 cells transfected with EGFP
(control), EGFP-wtCaD or phosphomimetic CaD mutants, including EGFP-A1A2
(the two PAK sites Ser452 and Ser482 converted to Ala), EGFP-A3A4 (the two Erk
sites Ser497 and Ser527 converted to Ala), EGFP-A1234 (both PAK- and Erk-sites
converted to Ala) and EGFP-D1234 (both PAK- and Erk-sites converted to Asp).
Results :
We found that cells transfected with wtCaD, A1A2 or A3A4 mutants of CaD
migrated at a rate approximately 50% more slowly than those EGFP-transfected cells.
The migration activity for A1234 cells was only about 13% of control cells. Thus it
seems both MAPK and PAK contribute to the motility of A7r5 cells and the effects
are comparable and additive. The A1234 mutant also gave rise to highest strain
energy and lowest rate of cell rounding. The migratory and contractile properties of
these cells are consistent with stabilized actin cytoskeletal structures. Indeed, the
A1234 mutant cells exhibited most robust stress fibers, whereas cells transfected with
wtCaD or A3A4 (and A1A2) had moderately reinforced actin cytoskeleton. The
control cells (transfected with EGFP alone) exhibited actin cytoskeleton that was
similar to that in untransfected cells, and also migrated at about the same speed as the
untransfected cells.
Conclusions : These results suggest that both the expression level and the level of MAPK- and/or
PAK-mediated phosphorylation of CaD play key roles in regulating the cell motility by modulating the actin cytoskeleton stability in dedifferentiated vascular SMCs such
as A7r5. - 中文關鍵字: --
- 英文關鍵字: --