Thermoresponsive block copolymer brush for temperature-modulated hepatocyte separation

Area of Science:

• Biomaterials Science
• Regenerative Medicine
• Cell Biology

Background:

• Hepatic tissue engineering offers a promising avenue for treating liver disease.
• Effective application necessitates cell separation technologies that preserve cell viability and avoid surface modification.
• Current methods often fall short in meeting these critical requirements for clinical translation.

Purpose of the Study:

• To develop novel hepatocyte cell separation materials utilizing thermoresponsive polymers.
• To create a system for selective cell adhesion and detachment based on temperature modulation.
• To assess the material's efficacy in separating hepatocytes from other cell types and maintaining cell activity.

Main Methods:

• Synthesis of a block copolymer brush, poly(N-p-vinylbenzyl-O-β-D-galactopyranosyl-(1→4)-D-gluconamide) (PVLA) and poly(N-isopropylacrylamide) (PNIPAAm) [PVLA-b-PNIPAAm], via atom transfer radical polymerization.
• Evaluation of HepG2 (hepatocyte model), TWNT-1 (stellate cell), and RAW264.7 (macrophage model) cell adhesion and detachment on the copolymer brush at varying temperatures (37 °C and 20 °C).
• Demonstration of temperature-modulated cell separation using a mixture of these cell types and assessment of recovered cell purity and activity.

Main Results:

• The PVLA-b-PNIPAAm brush exhibited temperature-dependent cell adhesion: HepG2 cells adhered at 37 °C and detached at 20 °C.
• TWNT-1 cells did not adhere, while RAW264.7 cells adhered regardless of temperature, enabling selective separation.
• Successful separation of HepG2 cells from a mixed cell population with ~85% purity and maintained normal activity was achieved via temperature modulation. Induced pluripotent stem (iPS) cell-derived hepatocytes were also selectively separated from undifferentiated iPS cells.

Conclusions:

• The developed PVLA-b-PNIPAAm copolymer brush provides an effective method for temperature-modulated hepatic cell separation.
• This technique successfully isolates hepatocytes from contaminating cells without surface modification or compromising cell activity.
• The copolymer brush represents a valuable tool for cell therapy and liver tissue engineering applications.