Rac1 Expression by Fibroblasts Is Required for Tissue Repair in Vivo

Abstract

Tissue repair requires that fibroblasts migrate into the wound to produce and remodel extracellular matrix, a process that requires adhesion. Failure to suppress the tissue repair program results in fibrotic disorders that are characterized by excessive adhesive signaling. The role of specific components of adhesive signaling in fibrogenic responses is unclear, but may involve small GTPases such as Rac1. To address the functions of Rac1 in fibroblasts, we generated mice containing a fibroblast-specific deletion of Rac1. These mice show delayed cutaneous wound closure, including reduced collagen production and myofibroblast formation. In cultured Rac1-deficient fibroblasts , adhesion , spreading , and migration were significantly inhibited. Rac1-deficient fibroblasts possessed impaired myofibroblast formation and function as visualized by reduced ␣-smooth muscle actin expression as well as matrix contraction. Both in vivo and in vitro, Rac1-deficient fibroblasts showed a reduced generation of reactive oxygen species; in vitro, hydrogen peroxide alleviated the phenotype of Rac1-deficient fibroblasts. Thus, Rac1 is an essential signaling integrator that is required for normal wound healing and dermal homeostasis. (Am J

Defective granulation tissue formation in mice with specific ablation of integrin-linked kinase in fibroblasts – role of TGFβ1 levels and RhoA activity

“…In vivo analysis of ILK function in fibroblasts was achieved by breeding Ilk fl/fl mice with a strain that carries a tamoxifen-inducible Cre recombinase under the control of a minimal a2(I) procollagen promoter linked to a fibroblast-specific enhancer (Denton et al, 2009;Liu et al, 2009;Sonnylal et al, 2007;Zheng et al, 2002). Efficient deletion of ILK was seen in fibroblasts of Col1-ILK mice, but not in keratinocytes or endothelial cells.…”

“…These mice were crossed to mice in which both Ilk alleles were mutated by the insertion of loxP sites flanking exon 2 (homozygous Ilk fl/fl ) ; Cre neg were used as controls. Cre activity was induced in all animals including controls at 12 days of postnatal life by intraperitoneal administration of tamoxifen for five consecutive days as previously described (Zheng et al, 2002;Liu et al, 2009;Sonnylal et al, 2007). Induction of Cre activity was examined in parallel experiments, in which the collagen Cre line was mated to reporter mice harboring a lacZ gene encoding bgalactosidase that was interrupted by a loxP-flanked stop cassette (Soriano, 1999).…”

Expression of integrin β1 by fibroblasts is required for tissue repair in vivo

“…Hydroxyproline assay was performed essentially as described previously Liu et al, 2009a;Reddy and Enwemeka, 1996). Wound tissues were homogenized in saline, hydrolyzed with 2N NaOH for 30 minutes at 120°C, followed by the determination of hydroxyproline by modification of the Neumann and Logan's reaction using Chloramine T and Ehrlich's reagent using a hydroxyproline standard curve and measuring at 550 nm.…”

“…As an example, fibroblasts lacking focal adhesion kinase (FAK) spread poorly, cannot migrate, and cannot properly respond to TGF (Ilic et al, 1995;Liu et al, 2007). Recently, we have shown that fibroblast expression of Rac1, a small GTPase that is expressed ubiquitously and is recruited to FAs by paxillin (Burridge and Wennerberg, 2004;Ishibe et al, 2004), is required for fibrogenesis and cutaneous tissue repair in vivo Liu et al, 2009a) and for induction of myofibroblast formation by endothelin-1 (Shi-wen et al, 2004). Moreover, we have shown that activation of cell adhesion is sufficient to cause increased expression of mRNAs encoding fibrogenic proteins .…”

Rac1 is deactivated at integrin activation sites via an IQGAP1/filamin-A/RacGAP1 pathway

“…Integrin-mediated adhesion modulates the activity of the small GTPase Rac1 (Del Pozo and Schwartz, 2007;Morgan et al, 2007;Price et al, 1998), and therefore dynamically regulates cell migration by promoting cytoskeletal re-organisation and membrane protrusion (Ridley, 2003;Ridley, 2006). Disruption of Rac1 signalling results in major migration defects, such as loss of cell directionality on fibrillar matrices Pankov et al, 2005) and suppressed wound healing in vivo (Liu et al, 2009;Tscharntke et al, 2007). Productive cell migration requires coordinated activation and deactivation of Rac1, and accordingly a range of guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs) have been reported to be involved in integrin-dependent Rac1 regulation (Katoh and Negishi, 2003;Nishiya et al, 2005).…”