Inmovilización de nanoparticulas magnéticas en membranas celulares mediante cadherinas para estudios de hipertermia magnética

Castro Hinojosa, Christian
Fratila, Raluca Maria (dir.) ; Moros Caballero, María (dir.)

Universidad de Zaragoza, CIEN, 2019
Departamento de Química Orgánica, Área de Química Orgánica

Máster Universitario en Biotecnología Cuantitativa

Resumen: Magnetic nanoparticles (MNPs) have the ability to generate heat when exposed to an alternating magnetic field. In biomedicine this property can be exploited to study the effects of localized heat generation on cell membranes. For this purpose, it is essential to target the MNPs to specific cell membrane receptors and achieve a stable and long-lasting immobilization of the MNPs on the cell membrane. Several molecules can be used as vectors; however, a stable conjugation is often difficult to achieve. Herein we explore the use of E-cadherin membrane protein as a potential targeting tool to generate a stable immobilization of MNPs on the cell membrane, based on homophilic E-cadherin – E-cadherin interactions. MNPs (12 nm diameter) were functionalized with polyethylene glycol (PEG) chains (MW: 750 or 5000 Da) and a derivative of nitrilotriacetic acid (NTA-Cu2+) in a single step using EDC chemistry; then, E-cadherin protein fragments (wild type: WT or mutant: W2A) were conjugated on the surface of the nanoparticles in an oriented manner through their histidine tail (His-tag), generating a protocol that could be easily adapted to any protein containing a His-tag in its structure. The cell-nanoparticle interactions were evaluated in Madin-Darby canine kidney (MDCK) and mouse fibroblast BALB/c3T3 cells, expressing and lacking E-cadherins respectively. The results obtained showed that the MNPs coated with a larger PEG decreased the extent of non-specific interaction with cells. On the other hand, when the MNPs were immobilized by the W2A mutant protein fragments, their interaction with the MDCK cells was lower in comparison with the one of the MNPs functionalized with WT fragments. Furthermore, in the case of BALB/c3T3 cells, lacking E-cadherins on their membranes, the interaction with the different types of MNPs did not show clear differences. These results suggested that the MNPs interact with MDCK cells through homophilic interactions; furthermore, the mutation present in the protein fragments W2A generates changes in the binding of nanomaterials to the cell membrane, which could be used in future studies to improve the specificity and stability of the MNPs immobilization on cell membrane.

Tipo de Trabajo Académico: Trabajo Fin de Master