Wainwright, Luke; (2018) Mechanisms of Coenzyme Q10 Blood-Brain Barrier Transport. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Coenzyme Q10 (CoQ10) deficiencies are unique among mitochondrial respiratory chain (MRC) disorders in that they are potentially treatable. While there is clear evidence, both clinically and biochemically, for the improvement of peripheral abnormalities associated with CoQ10 deficiency following CoQ10 supplementation, neurological symptoms are only partially ameliorated. The reasons for the refractory nature of the neurological sequelae associated with a CoQ10 deficiency are as yet unknown and may be a consequence of irreversible damage prior to CoQ10 supplementation, the retention of CoQ10 in the blood-brain barrier (BBB) itself, or simply reflect poor transport of CoQ10 across the BBB. This thesis presents the first isolated investigations into the mechanisms that govern bi-directional BBB transport of CoQ10 and its synthetic analogue, idebenone, using normal and pathophysiological cell models relevant to disorders of CoQ10 biosynthesis. The mouse BBB endothelial cell line bEnd.3 and porcine primary brain endothelial cells (PBECs) co-cultured with primary astrocytes were used to assess transcytosis from ‘blood-to-brain’ or ‘brain-to-blood’, revealing that although CoQ10 can traverse the BBB, CoQ10 is being effluxed back to the blood, which could explain the refractory nature of CoQ10 therapy, whereas, idebenone appeared to cross the BBB passively. Using inhibitors of known transport systems for lipoproteins, the circulatory bio-carriers of CoQ10 in vivo, three systems mediating the BBB transport of lipoprotein-bound CoQ10 were identified. Inhibitors of the scavenger receptor class B type 1 (SR-B1), BLT-1, and the receptor for advanced glycation end products (RAGE), FPS-ZM1, reduced uptake of lipoprotein-bound CoQ10 towards the brain, implicating RAGE and SR-B1 as modes for CoQ10 brain uptake.. In the reverse direction, the low-density lipoprotein receptor-related protein-1 (LRP-1) inhibitor, RAP, reduced efflux of lipoprotein-bound CoQ10 towards the blood, implicating LRP-1 as a major impediment to brain entry of CoQ10. This study is the first to generate a BBB endothelial cell model of CoQ10 deficiency, using para-aminobenzoic acid (pABA) to pharmacologically induce a depletion of cellular CoQ10 status, resulting in a global reduction of MRC enzyme activities. The CoQ10 deficient BBB models were leakier to large permeability markers, with poor BBB tight-junction formation, and altered CoQ10 transport dynamics in favour of an increased net efflux towards the blood, suggesting BBB pathophysiology is key to the neurological presentation and refractory nature of CoQ10 supplementation in symptomatic patients. In addition, the effects of vitamin E, a common clinical co-therapy in the ‘mito-cocktail’, and simvastatin were assessed. Interestingly, vitamin E co-administration reduced net efflux of CoQ10 from the brain. It is unknown why this occurs, but oxidative effects on the BBB transporters and/or carrier-lipoproteins may be factors to consider. In-line with its deleterious effect on CoQ10 biosynthesis, simvastatin therapy appeared to disrupt BBB integrity, increasing the paracellular leak of the BBB. This would be detrimental to normal brain homeostasis, particularly given the BBBs major role in limiting brain entry of the small molecule plasma excitotoxins, calcium, and glutamate. Throughout this study CoQ10 was quantified using a novel and rapid mass spectrometric method (ESI+ LC-MS/MS), which could potentially enable detection of CoQ10 in the CSF of patients presenting with neurological symptoms, perhaps providing a new analytical tool for the diagnosis of CoQ10 deficiencies in clinical laboratories. In conclusion, this thesis has demonstrated for the first time the pathophysiological consequences of a CoQ10 deficiency on the BBB. It has highlighted the impact of a deficit in CoQ10 status on CoQ10 delivery to the brain parenchyma and has elucidated some of the mechanisms by which CoQ10 is transported across the BBB, which are ultimately dictated by lipoprotein interactions. Additionally, this thesis outlines the potential dangers of statin therapy in patients with an underlying or established MRC dysfunction. Overall, this thesis provides insights into the limitations of CoQ10 supplementation as a therapy for neurological disorders associated with MRC dysfunction and indicates that further work will be required to improve the delivery of exogenous CoQ10 across the BBB, alongside a need for further investigations into the composition of the widely administered ‘mito-cocktail’.

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