Cell Culture on a Thermo-Responsive Polymer Surface

Abstract

We have used a thermo-responsive polymer, poly-N-isopropyl acrylamide (PNI-PAAm), as a substratum for the culture of human dermal fibroblasts by conjugating it with collagen. The cells attached well, spread, and grew on the substratum, indicating that the polymer has no toxicity towards the cells. PNIPAAm is insoluble in water over the lower critical solution temperature (LCST; about 32 degrees C) and reversibly solubilized below the LCST. Taking advantage of this conversion, monolayered fibroblasts cultured on the substratum containing the PNIPAAm over the LCST, were completely detachable from the substratum by simply lowering the temperature below the LCST, without the use of conventional detaching agents such as trypsin and EDTA. The detached cell sheet gradually aggregated and finally formed a multicellular spheroid. This polymer may provide a convenient and potentially useful technology for cell culture.

Tunable Stimuli-Responsive Polymers for Cell Sheet Engineering

“…The use of thermoresponsive substrate to detach the confluent cell sheets without the use of conventional enzymatic treatments was first reported by Takezawa et al in 1990 (Takezawa et al, 1990). In this study they have used PIPAAm as a substratum by conjugating it with collagen for the culture of human dermal fibroblasts.…”

Scaffolding in tissue engineering: general approaches and tissue-specific considerations

“…Such approach can be repeated to laminate multiple single cell layers to form thicker matrix. This technology is pioneered by Japanese group [84,85,110] and has been systemically applied to a number of applications such as cornea [82] in clinical trials and myocardium [106] in preclinical trials. The same approach has been modified and improved to produce patterned substrates for thicker tissue fabrication and for injectable applications [57].…”

Biology on a Chip: Microfabrication for Studying the Behavior of Cultured Cells

“…The first reported switch was thermally actuated, using the thermo-responsive polymer poly N-isopropyl acrylamide (PNIPAAm), which is hydrophobic above the lower critical solubility temperature (LCST, ∼32 °C) and very hydrophilic below the LCST. Takezawa et al 59 conjugated PNIPAAm to collagen as a substrate for culturing of human dermal fibroblasts and showed that cells could spread and grew normally on this surface above the LCST but could be detached by lowering the temperature below the LCST without the use of conventional (damaging) enzymes such as trypsin. Without compromised functions found in cultured cells damaged by trypsinization, cells recovered by this method maintained substrate adhesivity, growth, and secretion activities similar to those of primary cultured cells.…”