Revisiting Complexation between DNA and Polyethylenimine: Does the Disulfide Linkage Play a Critical Role in Promoting Gene Delivery?

Despite its cytotoxicity, polyethylenimine (PEI) is still used as a golden reference in gene transfection. Long PEI chains are more effective but also more cytotoxic. To solve this problem, an alternative strategy is to link short PEI chains into a longer PEI with disulfide bonds because they are degradable in the cell. However, how PEI promotes gene transfection is still unclear. Also, the chain length of PEI is also increased as disulfide bonds are formed. Therefore, it is important to investigate whether the increase in transfection efficiency is attributable to the disulfide linkage, chain size, or both. To distinguish between such factors, a novel method is developed here tomake longer linear PEI with disulfide bonds (lPEI(s-s)) by linking the mercapto groups of short linear PEI (lPEI(i)). By comparing the physiochemical properties and the transfection efficiencies of short lPEI(i), long lPEI(s-s), and un-degradable long PEI, it is found that introducing disulfide bonds instead of directly using longer PEI chains has less effect on gene transfection, and it is the chain length that plays a key role in promoting gene transfection.