Construction of an In Vitro Blood-Brain Barrier Micro-Organoid Model Using Decellularized Squid Mantle Scaffold Film

Area of Science:

• Biomedical Engineering
• Neuroscience
• Materials Science

Background:

• Existing blood-brain barrier (BBB) models have limitations due to the BBB's complex structure and cell interactions.
• A more authentic in vitro model is needed to accurately study neurological diseases and drug transport.

Purpose of the Study:

• To develop a novel BBB micro-organoid model using decellularized squid mantle scaffold (DSMS) films.
• To create a more reliable in vitro neurovascular unit (NVU) microenvironment for research.
• To validate the model's functional barrier properties and responsiveness to stimuli.

Main Methods:

• Decellularization and defatting of squid mantle scaffolds to create DSMS films.
• Lyophilization of DSMS films to achieve suitable pore size for cell culture.
• Co-culture of endothelial cells (hCMEC/D3) and astrocytes (hACs) on DSMS films to form a BBB-like structure.
• Assessment of barrier function using transendothelial electrical resistance (TEER) and macromolecule permeability.
• Evaluation of tight junctional complex (TJC) protein expression and low-density lipoprotein receptor-related protein 1 (LRP1) levels.

Main Results:

• DSMS films demonstrated good biocompatibility and suitable porosity for NVU cell adhesion and growth.
• The developed BBB micro-organoid model exhibited functional barrier properties with high TEER (~230 Ω/cm²) and restricted macromolecule permeability.
• The model showed expression of key TJCs (VE-cad, ZO-1) and stable expression of LRP1, indicating BBB integrity.
• The model responded to thrombotic conditions, showing its utility in studying BBB pathophysiology.

Conclusions:

• A functionally competent BBB micro-organoid model was successfully constructed using DSMS films and NVU cells.
• The DSMS-based BBB model provides a promising and authentic in vitro platform for neurological disease research.
• This model facilitates further studies on BBB pathological mechanisms and drug permeability evaluation.