Photo-Degradable Protein-Polymer Hybrid Shells for Caging Living Cells.

There is growing demand for the precise remote control of cellular functions in various fields. Herein, a method for caging mammalian cells by coating with photodegradable protein-polymer hybrid shells to photo-control their functions without genetic engineering is reported. A layer-by-layer assembly of photocleavable synthetic materials through biotin-streptavidin (SA) binding was employed for cell coating. The cell surfaces were first biotinylated with photocleavable biotinylated poly(ethylene glycol)(PEG)-lipid and then coated by repeatedly layering SA and micelles of the PEG-lipid and photocleavable biotinylated four-arm PEG. The cell extension and adhesion were suppressed with the shells and then triggered with the degradation of the shells by light exposure. Macrophage phagocytosis was also stopped by caging with the shells and restarted by light-guided uncaging. This study provides the first proof of principle that cellular functions can be remotely controlled by steric hinderance of cell surfaces with photodegradable materials.

Intracellular controlled release prolongs the time period of siRNA-based gene suppression.

RNA interference (RNAi) is a gene silencing process by inhibiting a target messenger RNA (mRNA) in the sequence-specific manner in the cell cytoplasm. Small interfering RNA (siRNA) cleaves the target mRNA. However, siRNA is not generally internalized into cells in the native state. The objective of this study is to prepare cationized gelatin nanospheres (cGNS) incorporating small interfering RNA (siRNA) and to prolong the time period of gene expression suppression. The cGNS with different degradabilities were prepared to evaluate the effect on the suppression of gene expression. There was no difference in the apparent size and zeta potential of cGNS among the amounts of glutaraldehyde (GA) added for crosslinking. The degradation of cGNS tended to become slowly with an increase of GA amounts used in preparation. After MC3T3-E1 cells were incubated with cGNS incorporating siRNA, the gene expression of cells was evaluated by real-time polymerase chain reaction (PCR). The time period of gene suppression increased with an increased amount of siRNA incorporated in cGNS. Moreover, the significant gene suppression was extended over 4 days. It is concluded that the intracellular controlled release with the cGNS enabled siRNA to prolong the time period of gene expression suppression.

Protein cell-surface display through in situ enzymatic modification of proteins with a poly(Ethylene glycol)-lipid.

Cell-surface display of functional proteins is a powerful and useful tool for regulating and reinforcing cellular functions. Direct incorporation of site-specifically lipidated proteins from the extracellular medium is more rapid, easily controllable and reliable in displaying active proteins than expression through gene transfer. However, undesirable amphiphilic reagents such as organic co-solvents and detergents were required for suppressing aggregation of ordinary lipidated proteins in solution. We report here sortase A-catalyzed modification of proteins with a poly(ethylene glycol)(PEG)-lipid in situ on the surface of living cells. Proteins fused with a recognition tag were site-specifically ligated with the PEG-lipid which was preliminary incorporated into cell membranes. Accordingly, target proteins were successfully displayed on living cells without aggregation under an amphiphilic reagent-free condition. Furthermore, to demonstrate the availability of the present method, Fc domains of immunoglobulin G were displayed on cancer cells, and the phagocytosis of cancer cells with dendritic cells were enhanced through the Fc-Fc receptor interaction. Thus, the present facile chemoenzymatic method for protein display can be utilized for modulating cell-cell interactions in cell and tissue engineering fields.