Mechanotransduction-Mediated Expansion of Rabbit Vocal Fold Epithelial Cells via ROCK Inhibition and Stromal...

Area of Science:

• Regenerative Medicine
• Cell Biology
• Biotechnology

Background:

• Therapeutic options for vocal fold (VF) disorders are restricted by a lack of available VF-derived epithelial cells (VFEs).
• Existing in vitro methods yield limited VFE expansion due to growth arrest, necessitating the use of less suitable cell sources.
• This scarcity hinders the development of regenerative therapies for vocal fold repair.

Purpose of the Study:

• To establish an optimized cell culture strategy for the long-term expansion of rabbit VFEs (rbVFEs).
• To investigate the mechanisms underlying VFE self-renewal and proliferation in vitro.
• To provide a consistent supply of rbVFEs for advancing vocal fold regenerative research.

Main Methods:

• Cultured rabbit VFEs (rbVFEs) using a combination of Rho kinase inhibitor (ROCKi), epidermal growth factor (EGF), and STO cell conditioned media (STO-CM).
• Assessed rbVFE proliferation, senescence, and self-renewal capacity under different culture conditions.
• Analyzed cellular mechanisms, including cytoskeletal remodeling, YAP localization, focal adhesion, and cellular tension, in self-renewing versus non-self-renewing rbVFEs.

Main Results:

• ROCKi alone supported only short-term rbVFE proliferation, leading to senescence.
• The addition of EGF or STO-CM to ROCKi-containing media enabled sustained rbVFE proliferation.
• Self-renewing rbVFEs showed distinct mechanobiological profiles (cytoplasmic YAP, reduced adhesion, lower tension) compared to senescent cells.

Conclusions:

• An optimized culture strategy using ROCKi, EGF, and STO-CM effectively promotes long-term expansion of rbVFEs.
• This method overcomes previous limitations in VFE culture, providing a robust cell supply.
• The findings offer a promising avenue for developing regenerative therapies for vocal fold disorders.