Collagen fibrillar structures in vocal fold scarring and repair using stem cell therapy: a detailed histological, immunohistochemical and atomic force microscopy study

Abstract

Summary Regenerative medicine opens new opportunities in the repair of cicatricial lesions of the vocal folds. Here, we present a thorough morphological study, with the focus on the collagen structures in the mucosa of the vocal folds, dedicated to the effects of stem cells on the vocal folds repair after cicatricial lesions. We used a conventional experimental model of a mature scar of the rabbit vocal folds, which was surgically excised with a simultaneous implantation of autologous bone marrow‐derived mesenchymal stem cells (MSC) into the defect. The restoration of the vocal folds was studied 3 months postimplantation of stem cells and 6 months after the first surgery. The collagen structure assessment included histology, immunohistochemistry and atomic force microscopy (AFM) studies. According to the data of optical microscopy and AFM, as well as to immunohistochemical analysis, MSC implantation into the vocal fold defect leads not only to the general reduction of scarring, normal ratio of collagens type I and type III, but also to a more complete restoration of architecture and ultrastructure of collagen fibres in the mucosa, as compared to the control. The collagen structures in the scar tissue in the vocal folds with implanted MSC are more similar to those in the normal mucosa of the vocal folds than to those of the untreated scars. AFM has proven to be an instrumental technique in the assessment of the ultrastructure restoration in such studies. Lay Description Regenerative medicine opens new opportunities in the repair of the vocal fold scars. Because collagen is a main component in the vocal fold mucosa responsible for the scar formation and repair, we focus on the collagen structures in the mucosa of the vocal folds, using a thorough morphological study based on histology and atomic force microscopy (AFM). Atomic force microscopy is a scanning microscopic technique which allows revealing the internal structure of a tissue with a resolution up to nanometres. We used a conventional experimental model of a mature scar of the rabbit vocal folds, surgically excised and treated with a mesenchymal stem cells transplant. Our morphological study, primarily AFM, explicitly shows that the collagen structures in the scarred vocal folds almost completely restore after the stem cell treatment. Thus, the modern microscopic methods, and especially AFM are instrumental tools for monitoring the repair of the vocal folds scars.

Autologous fibroblasts for vocal scars and age‐related atrophy: A randomized clinical trial

“…1,28,29 Cell-based therapy includes transplantation of pluripotent stem cells, mesenchymal stromal cells, or mature fibroblasts. Recent literature reviews [30][31][32] identified more than 30 in vitro and in vivo studies on cell therapy for vocal fold scarring alone. Translating these promising concepts to human trials has been much slower, as significant assurances of feasibility and safety are prerequisites before human administration.…”

Stem cell therapy for vocal fold regeneration after scarring: a review of experimental approaches

“…The most widespread stains applied in the histology analysis are hematoxylin–eosin and Masson trichrome for the general tissue structure, picrosirius red for revealing collagen, orcein and van Gieson stain to visualize elastic fibers, and alcian blue to identify glycosaminoglycans [ 25 – 27 , 38 , 41 , 46 , 47 , 50 , 53 , 54 , 60 – 62 ]. When applying immunohistochemistry staining, antibodies to collagen type I and fibronectin are mainly used [ 27 , 38 , 41 , 57 ]; in some studies, collagen type III, elastin, hyaluronic acid and decorin were also visualized [ 39 , 62 ]. Application of these approaches reflects the known data on the structure of the native and scarred VF mucosa [ 4 , 13 , 20 , 65 ].…”

Hyaluronan Hydrogels for the Local Delivery of Mesenchymal Stromal Cells to the Injured Vocal Fold

“…Supporting studies have investigated in detail ultrastructural changes in fibrous components of the VF ECM post-MSC injection, demonstrating improved collagen structure (thinner fibrils and fibers) and organization, as visualized via atomic force microscopy (190). Effects of MSC delivery have also been reported on scarring with relation to endogenous stromal fibroblasts.…”