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Background: Human adipose tissue-derived mesenchymal stem cells (ASCs) have potential utility as modulators of the regeneration of tissue that is inflamed or scarred secondary to injuries such as burns or trauma. However, the effect of ASCs on one particular type of scarring, keloidal disease, remains unknown. The absence of an optimal model for investigation has hindered the development of an effective therapy using ASCs for keloids.Objective: To investigate the influence of ASCs on angiogenesis, extracellular matrix deposition, and inflammatory cell influx in keloids.Methods: We analyzed the proliferation, migration, and apoptosis of human keloid-derived fibroblasts treated with a starvation-induced, conditioned medium from ASCs (ASCs-CM). This was achieved by Brdu proliferation assay, a validated co-culture migration assay, and flow cytometry, respectively. To assess the change in phenotype to a pro-fibrotic state, fibroblasts were analyzed by real-time PCR and contraction assay. A keloid implantation animal model was used to assess the paracrine effect of ASCs histochemically and immunohistochemically on scar morphology, collagen deposition, inflammatory cell composition, and blood vessel density. In tandem, an antibody-based array was used to identify protein concentration in the presence of ASCs-CM at time point 0, 24, and 48h.Results: ASCs-CM inhibited the proliferation and collagen synthesis of human keloid-derived fibroblasts. ASCs-CM was associated with reduced inflammation and fibrosis in the keloid implantation model. Thirty-four cytokines were differentially regulated by ASCs-CM at 24h. These included molecules associated with apoptosis, matrix metalloproteases, and their inhibitors. The same molecules were present at relatively higher concentrations at the 48h timepoint.Conclusion: These results suggest that ASCs are associated with the inhibition of fibrosis in keloids by a paracrine effect. This phenomenon may have utility as a therapeutic approach in the clinical environment. [ABSTRACT FROM AUTHOR]

Systemic Sclerosis (SSc) is an autoimmune disease characterized by vascular and immune system dysfunction, along with tissue fibrosis. Our previous study found GRB2 was downregulated by salvianolic acid B, a small molecule drug that attenuated skin fibrosis of SSc. Here we aim to investigate the role of GRB2 in SSc. The microarray data of SSc skin biopsies in Caucasians were obtained from the Gene Expression Omnibus (GEO) database. The expression of GRB2 was further detected in Chinese SSc and healthy controls. Bleomycin (BLM)-induced skin fibrosis mice were used to explore how GRB2 downregulation affected fibrosis. The apoptosis of EA.hy926 endothelial cells was induced by H 2 O 2 and apoptosis ratio was measured by flow cytometric. Transcriptome and phosphoproteomic analyses were performed to explore the regulated pathway. The expression of GRB2 was significantly enhanced in SSc patient skin, 1.51-fold in Caucasians and 1.40-fold in Chinese. Double immunofluorescence staining showed the endothelial cells of SSc patient's skin highly expressed GRB2. The in vivo study revealed that GRB2 knockdown alleviated skin fibrosis and apoptosis of endothelial cells in BLM mouse skin. The in vitro study showed that GRB2 downregulation inhibited the apoptosis of EA.hy926 and protected them from H 2 O 2 -induced hyperpermeability. Moreover, transcriptome and phosphoproteomic analysis suggested the focal adhesion pathway was enriched in GRB2 siRNA transfected endothelial cells. Our results demonstrated GRB2 highly expressed in endothelial cells of SSc skin, and inhibiting GRB2 could effectively attenuate BLM-induced skin fibrosis and endothelial cell apoptosis. GRB2 is expected to be a new therapeutic target for SSc. • Enhanced GRB2 in SSc patient skin, 1.51-fold in Caucasians and 1.40-fold in Chinese. • The endothelial cells of SSc patient skin highly expressed GRB2. • Inhibiting GRB2 attenuated BLM-induced skin fibrosis and endothelial cell apoptosis. • Inhibiting GRB2 protected EA.hy926 from apoptosis and H 2 O 2 -induced hyperpermeability. • Focal adhesion pathway was regulated by GRB2 to influence endothelial cell apoptosis. [ABSTRACT FROM AUTHOR]