Synthetic Thermo-Responsive Terpolymers as Tunable Scaffolds for Cell Culture Applications

Abstract

The use of tailored synthetic hydrogels for in vitro tissue culture and biomanufacturing provides the advantage of mimicking the cell microenvironment without issues of batch-to-batch variability. To that end, this work focused on the design, characterization, and preliminary evaluation of thermo-responsive, transparent synthetic terpolymers based on N-isopropylacrylamide, vinylphenylboronic acid, and polyethylene glycol for cell manufacturing and in vitro culture applications. Polymer physical properties were characterized by FT-IR, 1H-NMR, DLS, rheology, and thermal-gravimetric analysis. Tested combinations provided polymers with a lower critical solution temperature (LCST) between 30 and 45 °C. Terpolymer elastic/shear modulus varied between 0.3 and 19.1 kPa at 37 °C. Cellular characterization indicated low cell cytotoxicity on NIH-3T3. Experiments with the ovarian cancer model SKOV-3 and Jurkat T cells showed the terpolymers’ capacity for cell encapsulation without interfering with staining or imaging protocols. In addition, cell growth and high levels of pluripotency demonstrated the capability of terpolymer to culture iPSCs. Characterization results confirmed a promising use of terpolymers as a tunable scaffold for cell culture applications.

Microglia isolation from aging mice for cell culture: A beginner’s guide

“…Primary cell cultures are essential for certain research and have a number of applications. Researchers use cell cultures for various cell types, e.g., cardiac cells ( Avazzadeh et al, 2022 ), nerve cells ( Wei et al, 2022 ), hepatocytes ( Miyoshi et al, 2022 ) etc., and applications such as understanding cell morphology ( Giulian and Baker, 1986 ), functionality ( Stansley et al, 2012 ), stimulated cytokine production ( Bokarewa et al, 2021 ), scaffold research ( Lizana-Vasquez et al, 2022 ), and model systems ( Law et al, 2021 ). These aspects can be studied and visualized more reliably by cell culture techniques than by protein and RNA expression analyzes.…”

In vitro encapsulation and expansion of T and CAR-T cells using 3D synthetic thermo-responsive matrices

“…This hydrogel was characterized by its capability of easily attaching and detaching cells, supporting cell proliferation, transparency, and 3D encapsulation of cells including Jurkat T cells. 35 Two terpolymer combinations composed of 2% 4-VPBA, 4% PEGMMA400, and 94% NiPAAm (henceforth named 2:4:94 P400), and 4% 4-VPBA, 12% PEGMMA400, and 84% NiPAAm (henceforth named 4:12:84 P400) were specially selected for this study. The combination of these monomer molar ratios yielded terpolymers that possessed thermal and mechanical properties suitable for T cell encapsulation.…”

“…As detailed elsewhere, the thermo-responsive terpolymer samples were synthesized, characterized, washed, and sterilized for cell culture applications as described elsewhere. 35 Terpolymers are a polymerization of N -isopropylacrylamide (NiPAAm), 4-vinylphenylboronic acid (4-VPBA), and polyethylene glycol monomethyl ether monomethacrylate (PEGMMA) of 400 g mol −1 monomers.…”

“…Cells were encapsulated in 96 well plates using the sandwich methodology previously described by Lizana-Vasquez et al 35 Briefly, a polymer solution 15 wt% was prepared by mixing the appropriate amount of polymer and solubilized using culture media. This solution was further sterilized by filtration.…”

“…Furthermore, this smart and non-cytotoxic hydrogel was proven to have several advantageous features for Jurkat T cell encapsulation and culture. 35 In this study, two thermo-responsive terpolymers with different mechanical properties (0.5 ± 0.2 kPa 17 ± 2.5 kPa) were investigated. The selected stiffness values were based on mimicking some biologically relevant stiffnesses T cells face, while traveling to secondary lymphoid organs and peripheral tissues.…”