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Silk fibroin reactive inks for 3D printing crypt-like structures.

Danielle L Heichel^1,2, Julia A Tumbic^1,2, Marisa E Boch³

¹Polymer Program, Institute of Materials Science, University of Connecticut, Storrs, CT, United States of America.

Biomedical Materials (Bristol, England)

|September 14, 2020

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Researchers developed a novel silk fibroin ink for room-temperature 3D printing of stable protein hydrogels. This bioink allows complex shapes with excellent structural integrity for advanced cell studies.

Area of Science:

Biomaterials Engineering
Tissue Engineering
3D Bioprinting

Background:

Existing methods for 3D printing protein hydrogels often require elevated temperatures, specific additives, or sacrificial materials, limiting their applicability.
There is a need for robust, printable protein-based hydrogels that maintain structural integrity at ambient temperatures for complex tissue engineering applications.

Purpose of the Study:

To develop a reactive silk fibroin ink formulation for extrusion 3D printing of protein hydrogels at room temperature.
To achieve stable, geometrically accurate 3D printed constructs without synthetic additives or templates.
To evaluate the suitability of these hydrogels for long-term in vitro cell culture and studies.

Main Methods:

Purification of silk fibroin from Bombyx mori silkworm.

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Synthesis of reactive inks via enzyme-catalyzed dityrosine bond formation.

Rheological characterization and extrusion 3D printing studies with varying peroxide concentrations.

Assessment of shape retention, cell attachment, proliferation (Caco-2), and encapsulated cell viability (intestinal myofibroblasts).

Main Results:

Tailoring peroxide concentration enabled extrusion of silk fibroin into stable filaments and hydrogel constructs with excellent shape fidelity.
3D printed silk hydrogels demonstrated remarkable shape retention over nine weeks in culture medium without dimensional changes.
Caco-2 cell attachment, proliferation, and tight junction formation were unaffected by construct geometry, and encapsulated myofibroblasts survived printing and proliferated.

Conclusions:

A novel reactive silk fibroin ink formulation facilitates extrusion 3D printing of protein hydrogels at room temperature.
The developed bioink produces stable, geometrically precise constructs suitable for long-term in vitro studies.
This approach enables the creation of complex 3D cellularized constructs with potential applications in regenerative medicine and drug screening.