The isolation, cultivation and identification of laryngeal mucosal mesenchymal stem cells

Abstract

To investigate the presence of mesenchymal stem cells (MSC) in laryngeal mucosa, and to seek for the method to isolate, cultivate and identify these cells. Normal laryngeal mucosa was obtained from patients with laryngeal carcinoma during surgery, and the generating mesenchymal cells was obtained by digestive method. The cell growth curve was evaluated by 3-(4.5-methylthiazol-2yl)-2.5-diphenyltetrazolium bromide (MTT) assay. Colony forming cell assay was used to screen different morphologic colonies and evaluate clone formation ability. Flow cytometry was performed for the expression of the cells of the 4th passage surface marker profiles. Multiple differentiation potentials were confirmed by adipogenic, osteogenic and neural lineages induction. MTT assay and colony forming cell assay showed that laryngeal mucosa MSC had a relatively rapid proliferation capacity and a relatively high clone formation capacity (clone formation rate 7.1%). Flow cytometry analysis revealed that the laryngeal mucosa MSC were positive for CD29 (16.9%), CD44 (97.4%), CD90 (89.5%), CD105 (85.9%), CD146 (2.5%) and stro-1 (20.4%), but negative for CD34 (1.2%) and CD45 (0.8%). Laryngeal mucosa MSC undergone adipogenic, osteogenic and neural lineages induction were positive for oil red staining, alizarin red staining and S100 staining respectively, which suggested that laryngeal mucosa MSC could differentiate into adipogenic, osteogenic and neural lineages. This study demonstrated that MSC with rapid proliferative capacity and multiple differentiation potential could be obtained from lamina propria of laryngeal mucosa.