Lignin Nanotubes As Vehicles for Gene Delivery into Human Cells

Abstract

Lignin nanotubes (LNTs) synthesized from the aromatic plant cell wall polymer lignin in a sacrificial alumina membrane template have as useful features their flexibility, ease of functionalization due to the availability of many functional groups, label-free detection by autofluorescence, and customizable optical properties. In this report we show that the physicochemical properties of LNTs can be varied over a wide range to match requirements for specific applications by using lignin with different subunit composition, a function of plant species and genotype, and by choosing the lignin isolation method (thioglycolic acid, phosphoric acid, sulfuric acid (Klason), sodium hydroxide lignin), which influences the size and reactivity of the lignin fragments. Cytotoxicity studies with human HeLa cells showed that concentrations of up to 90 mg/mL are tolerated, which is a 10-fold higher concentration than observed for single- or multiwalled carbon nanotubes (CNTs). Confocal microscopy imaging revealed that all LNT formulations enter HeLa cells without auxiliary agents and that LNTs made from NaOH-lignin penetrate the cell nucleus. We further show that DNA can adsorb to LNTs. Consequently, exposure of HeLa cells to LNTs coated with DNA encoding the green fluorescent protein (GFP) leads to transfection and expression of GFP. The highest transfection efficiency was obtained with LNTs made from NaOH-lignin due to a combination of high DNA binding capacity and DNA delivery directly into the nucleus. These combined features of LNTs make LNTs attractive as smart delivery vehicles of DNA without the cytotoxicity associated with CNTs or the immunogenicity of viral vectors.

Recent Advances in Nanomaterials for Gene Delivery—A Review

“…Caicedo et al [122] developed a method to synthesize lignin-based nanotubes (LNTs) and nanowires using an alumina membrane as a sacrificial template. After showing the ability of DNA to adsorb onto LNTs, Ten et al [123] demonstrated the potential of LNTs to be used as a gene delivery vector. The morphology and properties of the LNTs were dependent on the plant species from which the lignin originated as well as the lignin isolation procedure [123].…”

“…After showing the ability of DNA to adsorb onto LNTs, Ten et al [123] demonstrated the potential of LNTs to be used as a gene delivery vector. The morphology and properties of the LNTs were dependent on the plant species from which the lignin originated as well as the lignin isolation procedure [123]. For example, LNTs synthesized with lignin that had been isolated from loblolly pine ( Pinus taeda L) wood with thioglycolic acid localized to the cytosol of HeLa cells, whereas LNTs synthesized from pine wood lignin isolated using NaOH penetrated the nuclei of the HeLa cells.…”

Organ Distribution and Bone Tropism of Cellulose Nanocrystals in Living Mice

“…36,40 This limited uptake could be likely due to the negative ζ-potential or the elongated shape: both factors may influence the interaction with cell membrane and the endosomal vesicle formation. 45,47 After 48 h of incubation, CNCs progressively penetrated in cells, with a concentration-dependent manner ( Figure 3A, middle column; Figure 3B). The merge between the fluorescent signals and the differential interference contrast (Nomarsky), which enabled us to visualize the shape of the cells in a 3D-like manner, showed spotted red circles exclusively confined to the cytoplasm.…”

Preparation and structural characterization of lignin micro/nano-particles with ionic liquid treatment by self-assembly

“…And from the DLS, the large size particles may be due to the agglomeration of lignin particles after centrifugation, drying and dispersed in the water ( Figure 3). Interests in using lignin micro/nano-particles for developing biodegradable materials are growing for many potential applications [16][17][18]. Many studies on lignin micro/nano-particles production used alkali lignin that was dissolved in organic solvents, such as ethylene glycol [18], acetone, tetrahydrofuran (THF) etc.…”