Sensitivity of myofibroblasts to H₂O₂‐mediated apoptosis and their antioxidant cell network

Abstract

During wound healing, the transition from granulation to scar tissue shows a decrease in myofibroblast cellularity. Previous results have correlated the disappearance of these cells with the induction of apoptotic cell death by some unknown stimuli. In contrast, hypertrophic scar appearance after wound healing is thought to be linked to a disorder of apoptotic function which induces myofibroblast persistence in granulation tissue. Oxidative stress being an important mediator of apoptosis, we have evaluated the apoptotic response of normal and pathological wound myofibroblasts (WMyo and HMyo respectively) in their interaction with two oxidative stress inducers: hydrogen peroxide, using a high concentration as a single dose, and sodium ascorbate which induced a continuous release of H2O2 at a low concentration. Our results showed that, according to the H2O2 treatment type, HMyo were more sensitive (after ascorbate treatment) or less sensitive (after H2O2 treatment) when compared to WMyo and Fb. We next assessed the presence of several molecules known to be involved in the antioxidant network protecting cells against H2O2 injury and found HMyo to have a higher level of activity of glutathione peroxidase and a lower level of activity of catalase than WMyo. These results can help explain the contradictory responses of myofibroblasts according to the oxidative stress treatment. This is the first study linking refractory oxidative stress mediated cell death to cellular phenotype in hypertrophic myofibroblasts, and indicates a pivotal role for the antioxidant enzyme system in this type of resistance.

Shedding of microparticles by myofibroblasts as mediator of cellular cross‐talk during normal wound healing

“…Cell apoptosis was evaluated using annexin‐V/PI binding (R&D) and TUNEL assay (R&D) as described in Larochelle et al (2004). Caspase 3 activity measurement was performed using the Apo Alert Caspase 3 assay kit (Clontech, Palo Alto, CA).…”

“…The presence of α2‐integrin protein on membrane of cells and MP was analyzed using flow cytometry on freshly trypsinized cells or MP isolated from the same cells using α2‐integrin antibodies directly coupled with FITC as described in Moulin et al (2001). Annexin‐V binding was used to numerate phosphatidylserine‐expressing MP as described in Larochelle et al (2004). VEGF and FGF2 were quantified using ELISA according to the manufacturer protocols (Peprotech, Rocky Hill, NJ).…”

Supplementation with High-Dose Ascorbic Acid Improves Intestinal Anastomotic Healing

“…Oxidative stress may result from either an increase in the levels of superoxide anion (O 2 antioxidant levels [7,8] . The presence of oxidative stress increases antioxidant enzyme activity to protect cells from injury, as demonstrated by the association between levels of antioxidant enzymes and oxidative stress [6] .…”

New and Improved Tissue Engineering Techniques: Production of Exogenous Material-Free Stroma by the Self-Assembly Technique

“…Myofibroblasts, or activated fibroblasts, are contractile, resistant to apoptosis and have an upregulated rate of matrix deposition. They also express different cytokine and chemokine receptors that enable fiber regulation and wound contraction at injury sites [21][22][23][24]. In this sense, activated fibroblasts not only favor wound healing, but can also cause injuries when their activation is uncontrolled, producing a pathological fibrotic response [25,26].…”