This study shows the potential of a thermally induced human serum albumin (HSA) hydrogel to serve as a drug depot for sustained release of a highly cytotoxic modified paullone ligand bearing a TEMPO free radical (HL). The binding of HL to HSA was studied by electron paramagnetic resonance (EPR) spectroscopy and imaging. The EPR protocol was also implemented for the study of matrix degradation, and ligand diffusion rate, in two additional spin-labeled hydrogels, containing 5-doxylstearate and 3-carbamoyl-proxyl. The results showed that the hydrogel is an efficient HL reservoir as it retained 60% of the ligand during 11 days of dialysis in physiological saline. Furthermore, upon incubation with Colo 205 human colon adenocarcinoma cells for 3 days, the HL/HSA hydrogel did not exhibit cytotoxic activity, demonstrating that it is also an efficient ligand depot in the presence of living cells. It was observed that the percentage of HL release is independent of its initial concentration in the hydrogel, suggesting that HSA possesses a specific binding site for the ligand, most likely Sudlow site 2, as predicted by molecular docking. The intrinsic property of albumin to bind and transport various substances, including hydrophobic drugs, may be fine-tuned by appropriate physical/chemical hydrogel preparation procedures, providing optimal drug delivery.
Background:
Alzheimer’s disease (AD) is the most common neurodegenerative disorder characterized by cognitive decline and total brain atrophy. Despite the substantial scientific effort, the pathological mechanisms underlying neurodegeneration in AD are currently unknown. In
most studies, amyloid β peptide has been considered the key pathological change in AD. However,
numerous Aβ-targeting treatments have failed in clinical trials. This implies the need to shift the re-
search focus from Aβ to other pathological features of the disease.
Objective:
The aim of this study was to examine the interplay between mitochondrial dysfunction,
oxidative stress and blood-brain barrier (BBB) disruption in AD pathology, using a novel approach
that involves the application of electron paramagnetic resonance (EPR) spectroscopy.
Method:
In vivo and ex vivo EPR spectroscopy using two spin probes (aminoxyl radicals) exhibit-
ing different cell-membrane and BBB permeability were employed to assess BBB integrity and
brain tissue redox status in the 5xFAD mouse model of AD. In vivo spin probe reduction decay
was analyzed using a two-compartment pharmacokinetic model. Furthermore, 15 K EPR spectros-
copy was employed to investigate the brain metal content.
Results:
This study has revealed an altered brain redox state, BBB breakdown, as well as ROS-me-
diated damage to mitochondrial iron-sulfur clusters, and up-regulation of MnSOD in the 5xFAD
model.
Conclusion:
The EPR spin probes were shown to be excellent in vivo reporters of the 5xFAD neu-
ronal tissue redox state, as well as the BBB integrity, indicating the importance of in vivo EPR spec-
troscopy application in preclinical studies of neurodegenerative diseases.
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