- The proton sponge effect is a widely accepted mechanism that explains how certain cationic polymers, particularly polyethyleneimine (PEI), enhance gene delivery by promoting endosomal escape. This phenomenon is especially important in non-viral transfection methods, where the cellular uptake of DNA or RNA complexes often results in entrapment within acidic endosomal compartments. Without an effective mechanism for escape, these nucleic acids are typically degraded in lysosomes, leading to inefficient gene expression.
- When PEI–DNA complexes (polyplexes) are taken up by mammalian cells, they are internalized via endocytosis and enclosed within endosomes. These endosomes are progressively acidified by proton pumps (H⁺-ATPases) as part of their normal maturation process. PEI, however, possesses a high density of protonatable amine groups—primary, secondary, and tertiary amines—which can buffer the influx of protons. As the PEI absorbs more and more protons, the endosomal pH does not decrease as expected, effectively “sponging” the protons.
- To maintain electrochemical balance, the accumulation of protons within the endosome is accompanied by an influx of negatively charged ions such as chloride (Cl⁻), along with water. This results in a significant osmotic imbalance, causing the endosome to swell. Eventually, the internal pressure becomes too great for the endosomal membrane to withstand, leading to rupture of the vesicle. As a result, the DNA or RNA cargo is released into the cytoplasm, where it can proceed to the nucleus for expression.
- The proton sponge effect is crucial for the success of PEI-based transfection, as endosomal escape is one of the major bottlenecks in non-viral gene delivery. Without this escape mechanism, most of the internalized genetic material would be degraded. PEI’s ability to combine DNA condensation, cellular uptake, and endosomal release in one molecule makes it a highly efficient transfection reagent, particularly for transient expression systems in mammalian cells.
- Despite its utility, the proton sponge effect is not without controversy. Some researchers argue that direct experimental evidence for osmotic endosomal rupture is limited, and suggest that other mechanisms—such as direct membrane destabilization or fusion—may also play roles. Furthermore, the same properties that enable endosomal escape can also contribute to cytotoxicity, which remains a challenge in the use of PEI for in vivo applications.
