The Biological Implications of Sialic Acid Binding Nanoparticles in T cells
Typ
Examensarbete för masterexamen
Master's Thesis
Master's Thesis
Program
Biomedical engineering (MPBME), MSc
Publicerad
2023
Författare
de Carvalho, Viktoria
Modellbyggare
Tidskriftstitel
ISSN
Volymtitel
Utgivare
Sammanfattning
The immune system is heavily regulated and affected by changes in the composition
of cell surface molecules, such as glycans. One sugar usually located at the end of
glycans is sialic acid (SA). Due to its negative charge and location, it is involved in
many immune processes, such as immune cell migration and maturation, but many
functions of SA are still unknown. The position and charge provide us with an easy
target for engineered particles with SA affinity. Seeing that immune cells are sialylated,
we can use this to target the immune system and possibly modulate it. This
project has focused on polymeric SA affine nanoparticles (NPs) and T cells. The
project primarily worked with the Jurkat E6-1 cell line of CD4+ T cells. Peripheral
blood mononuclear cells (PBMCs) purified from buffy coat were also used. Cytotoxicity
of NPs was investigated with the Alamar Blue assay. Activation markers
and NP-binding to cells were investigated with flow cytometry and the cytokine
profile was evaluated with enzyme-linked immunosorbent assay (ELISA). Confocal
microscopy was used to qualitatively evaluate NP binding specificity. The results
show that the NPs are not cytotoxic to T cells in tested concentrations ranging up
to 500 μg/mL. Flow cytometry revealed that the NPs bind less to activated Jurkat
cells and CD4+ primary cells, but more to activated CD8+ primary cells, compared
to the respective unactivated cells. Confocal microscopy confirmed that NPs bind
to unactivated cells to a higher extent. The NPs decreased the cellular expression
of CD62L, which could be due to some activation asserted by the NPs, or steric
hindering of the anti-CD62L antibody. ELISA showed that the IL-10 concentration
in supernatant from PBMCs was lower for cells exposed to NPs, but further
research is needed to confirm whether the NPs have a significant effect on the IL-10
profile of PBMCs. The IL-10 profile of Jurkats was not changed between particle
concentrations. Finally, flow cytometry showed that the level of activation markers
CD69 and CD25 were not affected by the particle concentrations, nor was the
IL-8 concentration after evaluation with ELISA, leading to the conclusion that the
activation level in T cells is not affected by the SA affine polymeric NPs. Thus,
the NPs prove promising for future use in immunomodulation, with diagnostic or
T cell activation-based targeting possibilities, and as a tool for understanding SA
expression.
Beskrivning
Ämne/nyckelord
T cells , polymeric nanoparticles , sialic acid , Jurkat , PBMC , glycans , CD69 , CD62L