Construction of glucosamine/DNA composite nanomaterials and its effect on function of Raw264.7 cells

Abstract

Objective To construct a new type of glucosamine/DNA composite nanostructure (NTGlcN) assembled without magnesium, verify whether or not glucosamine can mediate the assembly of DNA nanotubes (NT) and assess its effect on the function of Raw264.7 cells. Methods Utilizing the gradient annealing method with 3 DNA single strands Y1, Y2, and Y3, glucosamine (GlcN) was employed to mediate the assembly of DNA NT, resulting in the formation of glucosamine/DNA composite nanostructures. Atomic force microscopy (AFM) was used to observe the surface structure of the nanomaterial and dynamic light scattering (DLS) was used to measure its size. RAW264.7 cells were used in cell experiments. The cytotoxicity of GlcN and NTGlcN was assessed using CCK-8 assay. Flow cytometry and laser confocal microscopy were employed to investigate the cellular uptake efficiency of the nanostructures. The effects of NTGlcN and NTMg (Mg2+-assembled of DNA NT) on the expression levels of inflammatory cytokines (IL-1β, IL-6) in macrophages induced by lipopolysaccharides (LPS) were evaluated using RT-qPCR. Results GlcN successfully mediated the synthesis of NTGlcN, which exhibited good stability. AFM characterization results revealed that NTGlcN formed tubular particles that were uniformly distributed on the surface of mica. DLS measurements indicated that the diameter of NTGlcN was approximately 15.26±3.86 nm. Cell experiments demonstrated that, compared to NTMg, macrophages exhibited a higher cellular uptake efficiency for NTGlcN, with a higher cell survival rate following treatment with NTGlcN (P < 0.05). After NTGlcN treatment, the expression of inflammatory cytokines in LPS-induced macrophages was reduced (P < 0.05). Conclusion The glucosamine/DNA composite nanostructures have been successfully developed, possessing excellent stability, biocompatibility and cell uptake efficiency. NTGlcN is capable of reducing the cytotoxicity of GlcN and can suppress cellular inflammatory responses by decreasing the expression of inflammatory cytokines in RAW264.7 cells.

Keywords

• dna nanostructures
• glucosamine
• macrophages