Stem Cell Expansion and Fate Decision on Liquid Substrates Are Regulated by Self-Assembled Nanosheets

Abstract

The culture of adherent cells is overwhelmingly relying on the use of solid substrates to support cell adhesion. Indeed, it is typically thought that relatively strong bulk mechanical properties (bulk moduli in the range of kPa to GPa) are essential to promote cell adhesion and, in turn, regulate cell expansion and fate decision. In this report, we show that adherent stem cells such as mesenchymal stem cells and primary keratinocytes can be cultured at the surface of liquid substrates and that this phenomenon is mediated by the assembly of polymer nanosheets at the liquid-liquid interface. We use interfacial rheology to quantify this assembly and demonstrate the strong mechanical properties of such nanosheets. Importantly, we show that cell adhesion to such quasi-2D materials is mediated by the classical integrin/acto-myosin machinery, despite the absence of bulk mechanical properties of the underlying liquid substrate. Finally, we show that stem cell proliferation and fate decision are also regulated by the mechanical properties of these self-assembled protein nanosheets. Liquid substrates offer attractive features for the culture of adherent cells and stem cells, and the development of novel stem cell technologies, such as liquid-liquid systems, are particularly well-adapted to automated parallel processing and scale up.

Engineering the cellular mechanical microenvironment – from bulk mechanics to the nanoscale

“…In contrast, HPKs on soft nanosheets present a lower degree of cell spreading, decreased FAs and disorganised actin fibres. Reproduced with permission from Kong et al (2018b) where it was published under a Creative Commons Attribution license (CC BY 4.0).…”

“…oil). These nanosheets can be assembled from bovine serum albumin (BSA) (Kong et al, 2018a) or poly-L-lysine (PLL) (Kong et al, 2018b), and their interfacial shear stress moduli can be tuned by modifying the concentration of pro-surfactant and the pH during self-assembly, respectively. Moreover, PLL polymer nanosheets can be coated with fibronectin, which enables the adhesion and culture of adherent stem cells, such as MSCs and human primary keratinocytes (HPKs) (Fig.…”

“…Moreover, PLL polymer nanosheets can be coated with fibronectin, which enables the adhesion and culture of adherent stem cells, such as MSCs and human primary keratinocytes (HPKs) (Fig. 4D) (Kong et al, 2018b).…”

Materials Nanoarchitectonics as Cell Regulators

“…With addition of reactive surfactant pentafluorobenzoyl chloride, the mechanics of protein layer is enhanced, which can promote cell adhesion and proliferation. The same research team reported expansion and fate decision of stem cells with self‐assembled nanosheets . The used polymer nanosheets was made from cationic poly(L‐lysine) and reactive surfactant pentafluorobenzoyl chloride, to which extracellular matrix protein can be adsorbed.…”

“…The same research team reported expansion and fate decision of stem cells with self-assembled nanosheets. [29] The used polymer nanosheets was made from cationic poly(L-lysine) and reactive surfactant pentafluorobenzoyl chloride, to which extracellular matrix protein can be adsorbed. With strong mechanical properties, cell spreading and proliferation can be controlled.…”