"Mucosal immunization is a field of ever-increasing significance due to its ease of use, and nanospheres seem to have a very interesting effect that we tried to explain using high-density DNA microarrays," Carl Borrebaeck of Lund University told nanotechweb.org. "The power of genome-wide analysis is tremendous and has not been applied at all to explain the biological effects of nanospheres used in, for example, mucosal immunization."
Borrebaeck and colleagues attached tetanus toxoid antigen to the surface of polystyrene nanospheres with a diameter of about 460 nm. They introduced the tetanus-carrying nanospheres to immature human monocyte derived dendritic cells in vitro.
Dendritic cells are present throughout the body, but occur in greater numbers in areas that are particularly exposed to antigens, for example the skin and mucous membranes. An encounter with an antigen causes an immune response, in which the dendritic cells mature, migrate to secondary lymphoid tissue and activate T cells.
Fluorescence microscopy using nanospheres attached to fluorescent dye molecules showed that the dendritic cells internalized the nanospheres by phagocytosis. The nanospheres did not appear to be toxic to the cells over a three-day test period.
The tetanus-carrying nanospheres caused maturation of the dendritic cells, which expressed molecular maturation markers. The cells released HLA-DR and CD86 - markers that are both involved in the induction of an immune response.
"We have clearly shown that nanospheres as carriers of antigen directly affect the sentinels of the immune response, i.e. the dendritic cells," said Borrebaeck. "[This] also points to the fact that differently constructed nanospheres could have different immunological effects, thus pointing to a very interesting development within nanobiotechnology."
The researchers used high-density microarrays to do a global transcriptional analysis of the nanosphere-treated dendritic cells. The treatment caused 100-175 genes to up-regulate more than two-fold. Treatment with nanospheres that weren't attached to tetanus toxoid also caused an increase in up-regulation, but for fewer genes. And treatment with tetanus toxoid alone only caused a marginal response. The results for down-regulation of genes were even more marked.
According to the researchers, their findings demonstrate that antigen immobilized onto the surface of a nanosphere has a significantly more pronounced effect on the gene regulation in dendritic cells than antigen alone.
"By custom-designing nanospheres using different polymers, including biodegradable [ones], it might be possible to educate the immune system and obtain especially desired effects such as increased CTL [cytotoxic T lymphocyte] response or induction of certain Ig [immunoglobulin] classes," said Borrebaeck. "This has not been shown but there are indications from our collaboration that this might be achievable."
The researchers reported their work in Nano Letters.