Abstract Description

Cryptococcus (C.) neoformans is a critical priority pathogen that causes the often-deadly inflammatory condition of the brain. The latter is complicated by treatment challenges associated with controlling cryptococcal growth in the brain. This is, in part, because the gold-standard drug amphotericin B poorly penetrates the neuroprotective blood-brain barrier (BBB). Our group has previously shown that aspirin displays anti-Cryptococcus activity; however, it is reported to have poor BBB penetration. To this end, we encapsulated it in D-α-tocopheryl polyethene glycol succinate (TPGS), confirmed its colloidal dispersion using Fourier transform infrared spectroscopy (FTIR), and lastly characterised its physicochemical properties using a Zeta particle analyser. This was followed by biological studies aimed at assessing if the prepared aspirin-loaded TPGS micelles could: 1) disrupt the integrity of the artificial BBB made of immortalised mouse b.End5 endothelial cells that were prepared in a transwell plate, 2) cross this artificial BBB, and 3) inhibit the growth of cryptococcal cells overlayed in the basolateral chamber of the plate. Furthermore, Galleria mellonella larvae were used to measure the efficacy of aspirin-loaded TPGS micelles in clearing cells in the brain tissue. Here, melanisation was visualised and used as an indicator of fungal clearance. For the biological studies, aspirin (not dispersed in TPGS) was used as a control for referencing purposes. FTIR analyses confirmed aspirin encapsulation, and these micelles were determined to have an average size of 10.97 nm, polydispersity index of 0.175 and a 3.668 mV zeta potential. The prepared micelles were determined to be not detrimental to the BBB, in part due to their near-zero charge (3.668 mV). The latter was further supported by the obtained TEER measurements and sodium fluorescein penetration readings. Importantly, the micelles could penetrate the BBB, due to their determined small size (10.97 nm), and inhibited significantly more cryptococcal cells in the basolateral chamber than aspirin (p < 0.05). Consistent with these in vitro findings, the Galleria mellonella studies revealed that the brain tissue of larvae infected with cells treated with micelles was less melanised compared to that treated with non-reformulated aspirin or non-treated cells. Taken together, these findings suggest that aspirin could be delivered across the brain endothelium to control the possible growth of cryptococcal cells in the brain parenchyma.