Optimization of amniotic membrane (AM) denuding for tissue engineering

Area of Science:

• Biomaterials Science
• Tissue Engineering
• Regenerative Medicine

Background:

• Amniotic membrane (AM) is a popular scaffold for ex vivo-expanded cells in tissue engineering, especially for ocular surface reconstruction.
• Current AM preparation methods are inconsistent and their impact on the basement membrane (BM) is unclear.
• Standardized AM processing is crucial for reliable tissue-engineered constructs.

Purpose of the Study:

• To compare the effects of common denuding procedures on AM basement membrane (BM) integrity.
• To introduce and validate a novel, standardized thermolysin-based method for AM preparation.
• To assess the structural and molecular preservation of the BM after denudation.

Main Methods:

• Amniotic membranes (AMs) were processed using ethylenediaminetetraacetic acid (EDTA), dispase, and a novel thermolysin-based procedure.
• Scanning and transmission electron microscopy were employed to evaluate BM structure.
• Immunohistochemistry was used to assess the molecular composition of key BM components.

Main Results:

• EDTA- and dispase-based methods led to significant disaggregation and destruction of the AM basement membrane (BM).
• The novel thermolysin procedure effectively removed the epithelium.
• Thermolysin treatment preserved the structural and molecular integrity of the AM basement membrane (BM).

Conclusions:

• Existing AM denuding methods are variable and compromise BM integrity, leading to non-standardized membranes.
• The novel thermolysin-based technique offers an effective and standardized approach to AM preparation.
• This standardized method has the potential to enhance the generation of tissue-engineered constructs utilizing AM.