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Integrin‐Fibronectin (FN) Mediated Mechanotransduction of Microvascular Smooth Muscle Cells (VSMC) Involves Activation of RhoA and PKC‐α
Author(s) -
Sun Zhe,
Huang Shaoxing,
Meininger Gerald A.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.744.18
Subject(s) - rhoa , mechanotransduction , microbiology and biotechnology , integrin , förster resonance energy transfer , fibronectin , chemistry , protein kinase c , vascular smooth muscle , cell adhesion , transfection , adhesion , biophysics , signal transduction , cell , biology , extracellular matrix , biochemistry , smooth muscle , medicine , fluorescence , physics , organic chemistry , quantum mechanics , gene
Integrin‐mediated mechanotransduction plays an important role in the arteriolar myogenic response. Diverse signaling pathways are believed to be involved in this process. To investigate the activation of RhoA and PKC‐α during VSMC mechanotransduction, we utilized atomic force microscopy (AFM) to mechanically stimulate single VSMC cells. Confocal microscopy was used to visualize RhoA and PKC‐α in live VSMCs isolated from rat skeletal muscle arterioles, 70~100 μm Diameter. VSMCs were transfected with either a FRET sensor of RhoA activation or GFP‐tagged PKC‐α. The tips of the AFM probes were fused with a FN‐coated bead (5 μm Dia). The FN‐coated beads were placed in contact (20 minutes) with the VSMC membrane that leads to the formation of an integrin‐mediated cell adhesion complex. A controlled pulling force was applied through the bead adhesion site using the AFM while simultaneously recording the signal from the RhoA‐FRET sensor or the GFP‐PKC‐α. Following pulling, a significant increase in the FRET signal for the RhoA sensor was detected. Also, the pulling was accompanied by translocation of GFP‐PKC‐α to the cell membrane. These changes began to occur within seconds after force application and were observed throughout the cell. Collectively, these data suggested that RhoA and PKC were activated by pulling force applied through integrin‐FN focal adhesion. (Supported by NIH HL‐062863 to GAM and AHA 0765481Z to ZS)