Fabrication of Myomucosal Flap Using Tissue‐engineered Bioartificial Mucosa Constructed With Oral Keratinocytes Cultured on Amniotic Membrane

Abstract

The purpose of this study was to fabricate bioartificial mucosa using cultured oral keratinocytes (OKCs) on an amniotic membrane (AM), and to evaluate the possibility of developing a prelaminated myomucosal flap using the fabricated bioartificial mucosa and local muscle flap. Buccal mucosa was harvested from male New Zealand rabbits (n = 40, 2.5-3.0 kg) and primary cultivation was performed. The cultured OKCs were seeded on the AM and a submerged culture was performed. Prelamination of the bioartificial mucosa was performed on the latissimus dorsi (LD) muscle of rabbits. Survival rate, layer of OKCs, and Cinamon's score (CS) based on macroscopic and microscopic examinations were evaluated 7, 10, 14, and 21 days after prelamination (n = 10 per day). The OKCs cultured on AM showed multiple layers (3.85 +/- 1.32) and cells were tightly adhered with desmosomes. Basal layer cells adhered to the AM with hemidesmosomes. In addition, the AM played an excellent role as a substrate for the OKCs and simplified handling during prelamination. A myomucosal flap with OKCs cultured on AM was fabricated within 2 weeks (CS: 11.05 +/- 2.63). The basement component of laminin was observed 2 weeks after prelamination and showed enough strength to adhere to the underlying fascia. A myomucosal flap was successfully developed using prelamination of bioartificial mucosa on the LD muscle between 10 and 14 days.

Sequential keratinocytic differentiation and maturation in a three‐dimensional model of human artificial oral mucosa

“…In this work, we have performed a sequential study of epithelial layer development of a human AOM model by evaluating key basement membrane proteins and intercellular junctions. Most of the works focused on the study of AOM have used partial and non‐autologous oral mucosa models to identify epithelial patterns such as DG3, laminin and others . On the one hand, our results confirmed the epithelial phenotype of the bioengineered epithelial layer as determined by the positive expression of pancytokeratin in all AOM samples as we previously demonstrated .…”

In vivo behavior of complete human oral mucosa equivalents: characterization in athymic mice

“…In our view, maturation in vitro prolongs culture time at early stages. In this regard, in the first week of development, our findings are consistent with those reported by other authors (12–14), whose CAOMEs were ripened using an air–liquid interface for 7 d. Fourteen days after grafting, we observed a progressive increase in the number of layers; the most notable feature was the remarkable organization the epithelium achieved, with a basal layer and some suprabasal layers not yet arranged. The epithelium began to mature in the second week as long as the basal layer proliferation was preserved.…”

“…However, Ahn et al. (14) could not find this positivity at 14 d of grafting in their rabbit model. In our work, we observed discontinuity throughout the first week of grafting, but in the second week the immunostaining was virtually continuous, and at the third week, completely continuous.…”

“…Of the major components that make up the basement membrane, laminin 5 is the one which appears earliest (17), and it seems to stabilize in vitro after 14 d of air-liquid interface (20). However, Ahn et al (14) could not find this positivity at 14 d of grafting in their rabbit model. In our work, we observed discontinuity throughout the first week of grafting, Fig.…”

Überexpression von β1-Integrin in oralen Keratinozyten korreliert mit deren hohen Proliferationskapazität

“…Replacing oral mucosa is difficult to achieve and allowing oral mucosal wounds to close by secondary inten-* Contributed equally tion often causes scarring and contracture. During past decades skin has been substituted for mucosa in the form of skin grafts [2], axial pattern flaps [3], musculocutaneous flaps [4,5] or free-tissue transfer [6,7]. However, problems with these treatments include unpredictable graft survival, contracture, hair growth, excessive bulk of flap tissue and difficult dental prosthetic reconstruction [8], and therefore their applications are limited.…”