The utilisation of carbon nanotubes (CNTs) in the biomedical field is being realised. Short multi-walled CNTs (MWNTs) has the better toxicological profile, thus is preferable over its single-walled counterpart. Functionalisation of MWNTs (producing fMWNTs) confer modifiable functional groups with which therapeutic molecules can be attached; employment of fMWNTs as drug delivery nanovectors could be evaluated. Plenty of potentially beneficial neurological therapeutics have been hampered by poor penetration of the blood-brain barrier (BBB). Therefore, the objective was to demonstrate cellular internalisation of fMWNTs into cells of the BBB.
Pristine MWNTs were functionalised to produce MWNT:PF127, MWNT-COOH and MWNT-NH3+. All MWNT preparations were dispersed in water before used in experiments and were characterised using transmission electron microscopy (TEM), Fourier-transform infrared (FTIR) spectroscopy and thermogravimetric analysis (TGA). Three types of cells were used for the internalisation experiments: human brain microvascular endothelial cells (hBEC-5i, ATCC CRL-3245), human pericytes (hPeric) and human astrocytes (hAstro). Internalisation of MWNTs were assessed using flow cytometry analysis (FCA) of HBEC-5i monolayers and confocal microscopy (CFM) of fluorescently-labelled heterocellular spheroidal model of the BBB. Measures to indicate internalisation are increased side-scatter (FCA) and reduction in fluorescence intensity fraction (CFM).
All fMWNT preparations were easily dispersible in water. Compared to MWNT:PF127, both MWNT-COOH and MWNT-NH3+ were shorter. FTIR confirmed the presence of the functional groups on MWNT-COOH and MWNT-NH3+. TGA showed that whilst pristine MWNTs were thermally stable, MWNT-COOH and MWNT-NH3+ lost 33% and 80% of their weight, respectively, after heated to 1000°C. The quantitative Kaiser test performed on MWNT-NH3+ showed amine group loading of 674 µmol/g of MWNT. HBEC-5i monolayer exposed to MWNT:PF127 for 24 and 48 hours did not show any dose-dependent difference in side scatter nor AUC. However, exposure of the same cell line to MWNT-COOH and MWNT-NH3+ showed a dose-dependent increase in cellular side scatter. Exposure of the heterocellular spheroidal model of the BBB to the fMWNTs resulted in reduced fluorescence intensity fraction in both hBECs and hPerics.
fMWNTs were shown to be safely internalised into cells of the BBB, with MWNT-NH3+ possessing the higher potential as a drug-delivery nanovector due to its modifiable functional group.
