ARPE-19 Cell Uptake of Small and Ultrasmall Superparamagnetic Iron Oxide

作者：Grottone Gustavo Teixeira^*; Loureiro Renata Ruoco; Covre Joyce; Rodrigues Eduardo Buchele; Pereira Gomes Jose Alvaro

来源：Current Eye Research, 2014, 39(4): 403-410.

DOI：10.3109/02713683.2013.845228

• Cell culture techniques
• tissue therapy
• nanoparticles
• retina
• translational medical research

Purpose: To investigate the cytotoxicity, cellular intake and magnetic field interaction of three superparamagnetic iron oxide (SPIO) and one ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles on ARPE-19 cells.
Methods: Two FDA-approved SPIO nanoparticles (Endorem and Lumirem), one commercial SPIO(FluidMag-L) and one FDA-approved USPIO (Feraheme) were tested. Nanoparticle suspensions were diluted and prepared in high- (1000 Fe-ug/ml) and low- (100 Fe-ug/ml) dose suspensions. ARPE-19 cells were incubated in four 24-well plates and the medium changed every other day until cells attained 80% confluence. Nanoparticle cytotoxicity was evaluated using the XTT cytotoxicity assay. Cellular attraction was tested after digestion of the cells in collagenase A (1 mg/ml) overnight. A 3500 Gauss neodymium magnet was used to attract cells to the well walls. ARPE-19 cell ultrastructure was evaluated by transmission electron microscopy (TEM) to determine the specific locations of nanoparticles within the cell membranes.
Results: Cytotoxicity assessment by the XTT assay revealed that ARPE-19 cells that were exposed to either concentration of Endorem, FLuidMag-L, Feraheme non-conjugated with protamine and heparin or Lumirem demonstrated no statistically significant toxicity. Cells exposed to Feraheme when conjugated with protamine and heparin presented severe toxicity in both concentrations. When a magnetic field was applied, all nanoparticle-containing samples, except Feraheme non-conjugated form, were promptly attracted. TEM and prussian blue staining examination revealed that Feraheme alone was not initially capable of cellular uptake. This issue was solved by conjugating Feraheme with protamine and heparin (although cytotoxicity was found on those samples). Endorem, FLuidMag-L, Feraheme conjugated form were found within the cytoplasm of ARPE-19 cells.
Conclusions: Ferahame when conjugated with protamine and heparin was cytotoxic at the higher and lower doses, as revealed by the XTT assay. Endorem and FluidMag-L were not toxic at the studied concentrations. Feraheme non-conjugated solutions and Lumirem solutions provided were harmless but were not internalized by ARPE-19 cells. All the studied nanoparticles were attracted to the magnetic field except Feraheme in the non-conjugated form.

• 出版日期2014-4