Liposomes and Cationic Lipids (Lipofection)
In the existing biological experimental technology, liposome and cationic lipids transfection have become one of the most commonly used methods for gene delivery (introduce foreign genes into cells). In most gene therapy projects, the most important point of transfecting foreign genes with liposomes is to prevent their toxicity from damaging the body. Therefore, such as the ratio of liposomes to plasmids, cell density, length of transfection and serum content in the medium are all important parameters that affect the transfection efficiency. We need to explore suitable transfection conditions through continuous experiments to improve the efficiency and safety of transfection.
Advantages of Lipofection
Because liposomes are naturally occurring substances, they have the unique advantage of being biodegradable, and have been regarded as one of the ideal carriers for gene delivery. They can achieve stable encapsulation of nucleic acid molecules, protect DNA from enzymatic degradation, and promote cellular uptake and endosome escape, thereby mediating efficient gene transfer. Liposomes not only have excellent biocompatibility and low immunogenicity, but also have the ability to deliver large fragments of DNA with well-defined physicochemical composition, and are easy to handle and prepare. In addition, because liposomes have the potential to transfect multiple cell types, they can be used in different gene therapy programs.
Disadvantages of Lipofection
Because cationic liposomes are positively charged, nonspecific interactions with negatively charged cell components (such as serum proteins and enzymes) may occur, leading to decreased cell adhesion, hemolysis, and transfection efficiency. In addition, the preparation of liposomes involves the use of organic reagents such as ether and chloroform, which will cause damage to cells and tissues and need to be removed through additional processes. In general, due to its potential cytotoxicity and low transfection efficiency, cationic liposomes cannot currently achieve the desired gene therapy effect in clinical trials. Therefore, it is very important to develop a new non-toxic cation system with effective gene transfection ability and good safety.
Figure 1. Lipofection. (Carter, 2014)
Optimization of Liposome Transfection Safety
The cytotoxicity of cationic liposomes is mainly due to their cationic properties and linking groups. Optimization is generally performed through two offenses to increase transfection efficiency and reduce cytotoxicity:
- Synthetic modification of positively charged headgroup. For example, heterocycles such as imidazolium pyridinium and protonated polyamine groups are introduced into cationic liposomes to reduce the positive charge of the cation head.
- Modification with linker functional groups improves the properties of liposomes for gene delivery. Cationic liposomes with ether linkages are too stable to be biodegraded. Although they have good transfection efficiency, they lead to higher toxicity. However, ester or amide linkers are more biodegradable and consistently less cytotoxic in cultured cells. Moreover, lipids with these two linkers are more easily degraded in the circulatory system. Carbamate is a widely studied structure that has the ability to keep lipids stable in the circulatory system, and it may break down and release DNA after entering the endosome. As a result, lipids with a urethane linker can quickly degrade into small molecules with much lower toxicity.
In addition, cationic liposomes conjugated with polyethylene glycol (PEG) or other molecules such as ligands and peptides have been greatly improved to achieve smaller particle sizes, controlled structures, more regular morphology, and better stability.
Reference
- Carter, M.; Shieh, J. C. (2015). Guide to research techniques in neuroscience. Academic Press.
