Pharmacology and Therapeutics - Research Publications
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ItemIron overload and impaired iron handling contribute to the dystrophic pathology in models of Duchenne muscular dystrophyAlves, FM ; Kysenius, K ; Caldow, MK ; Hardee, JP ; Chung, JD ; Trieu, J ; Hare, DJ ; Crouch, PJ ; Ayton, S ; Bush, A ; Lynch, GS ; Koopman, R (WILEY, 2022-03-06)BACKGROUND: Oxidative stress is implicated in the pathophysiology of Duchenne muscular dystrophy (DMD, caused by mutations in the dystrophin gene), which is the most common and severe of the muscular dystrophies. To our knowledge, the distribution of iron, an important modulator of oxidative stress, has not been assessed in DMD. We tested the hypotheses that iron accumulation occurs in mouse models of DMD and that modulation of iron through the diet or chelation could modify disease severity. METHODS: We assessed iron distribution and total elemental iron using LA-ICP-MS on skeletal muscle cross-sections of 8-week-old Bl10 control mice and dystrophic mdx mice (with moderate dystrophy) and dystrophin/utrophin-null mice (dko, with severe dystrophy). In addition, mdx mice (4 weeks) were treated with either an iron chelator (deferiprone 150 mg/kg/day) or iron-enriched feed (containing 1% added iron as carbonyl iron). Immunoblotting was used to determine the abundance of iron- and mitochondria-related proteins. (Immuno)histochemical and mRNA assessments of fibrosis and inflammation were also performed. RESULTS: We observed a significant increase in total elemental iron in hindlimb muscles of dko mice (+50%, P < 0.05) and in the diaphragm of mdx mice (+80%, P < 0.05), with both tissues exhibiting severe pathology. Iron dyshomeostasis was further evidenced by an increase in the storage protein ferritin (dko: +39%, P < 0.05) and ferroportin compared with Bl10 control mice (mdx: +152% and dko: +175%, P < 0.05). Despite having features of iron overload, dystrophic muscles had lower protein expression of ALAS-1, the rate-limiting enzyme for haem synthesis (dko -44%, P < 0.05), and the haem-containing protein myoglobin (dko -54%, P < 0.05). Deferiprone treatment tended to decrease muscle iron levels in mdx mice (-30%, P < 0.1), which was associated with lower oxidative stress and fibrosis, but suppressed haem-containing proteins and mitochondrial content. Increasing iron via dietary intervention elevated total muscle iron (+25%, P < 0.05) but did not aggravate the pathology. CONCLUSIONS: Muscles from dystrophic mice have increased iron levels and dysregulated iron-related proteins that are associated with dystrophic pathology. Muscle iron levels were manipulated by iron chelation and iron enriched feed. Iron chelation reduced fibrosis and reactive oxygen species (ROS) but also suppressed haem-containing proteins and mitochondrial activity. Conversely, iron supplementation increased ferritin and haem-containing proteins but did not alter ROS, fibrosis, or mitochondrial activity. Further studies are required to investigate the contribution of impaired ferritin breakdown in the dysregulation of iron homeostasis in DMD.
ItemType-1 Interferons Contribute to the Neuroinflammatory Response and Disease Progression of the MPTP Mouse Model of Parkinson's DiseaseMain, BS ; Zhang, M ; Brody, KM ; Ayton, S ; Frugier, T ; Steer, D ; Finkelstein, D ; Crack, PJ ; Taylor, JM (WILEY, 2016-09-01)Type-1 interferons (IFNs) are pleiotropic cytokines with a critical role in the initiation and regulation of the pro-inflammatory response. However, the contribution of the type-1 IFNs to CNS disorders, specifically chronic neuropathologies such as Parkinson's disease is still unknown. Here, we report increased type-1 IFN signaling in both post mortem human Parkinson's disease samples and in the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) mouse model. In response to MPTP, mice lacking the type-1 IFN receptor (IFNAR1(-/-) ) displayed decreased type-1 IFN signaling, an attenuated pro-inflammatory response and reduced loss of dopaminergic neurons. The neuroprotective potential of targeting the type-1 IFN pathway was confirmed by reduced neuroinflammation and DA cell death in mice treated with a blocking monoclonal IFNAR1 (MAR-1) antibody. The MPTP/MAR-1 treated mice also displayed increased striatal dopamine levels and improved behavioural outcomes compared to their MPTP/IgG controls. These data, implicate for the first time, a deleterious role for the type-1 IFNs as key modulators of the early neuroinflammatory response and therefore the neuronal cell death in Parkinson's disease. GLIA 2016;64:1590-1604.
ItemTreating Alzheimer's disease by targeting ironNikseresht, S ; Bush, A ; Ayton, S (WILEY, 2019-09-01)No disease modifying drugs have been approved for Alzheimer's disease despite recent major investments by industry and governments throughout the world. The burden of Alzheimer's disease is becoming increasingly unsustainable, and given the last decade of clinical trial failures, a renewed understanding of the disease mechanism is called for, and trialling of new therapeutic approaches to slow disease progression is warranted. Here, we review the evidence and rational for targeting brain iron in Alzheimer's disease. Although iron elevation in Alzheimer's disease was reported in the 1950s, renewed interest has been stimulated by the advancement of fluid and imaging biomarkers of brain iron that predict disease progression, and the recent discovery of the iron-dependent cell death pathway termed ferroptosis. We review these emerging clinical and biochemical findings and propose how this pathway may be targeted therapeutically to slow Alzheimer's disease progression. LINKED ARTICLES: This article is part of a themed section on Therapeutics for Dementia and Alzheimer's Disease: New Directions for Precision Medicine. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.18/issuetoc.
ItemIron accumulation in skeletal muscles of old mice is associated with impaired regeneration after ischaemia-reperfusion damageAlves, FM ; Kysenius, K ; Caldow, MK ; Hardee, JP ; Crouch, PJ ; Ayton, S ; Bush, AI ; Lynch, GS ; Koopman, R (WILEY, 2021-03-04)BACKGROUND: Oxidative stress is implicated in the insidious loss of muscle mass and strength that occurs with age. However, few studies have investigated the role of iron, which is elevated during ageing, in age-related muscle wasting and blunted repair after injury. We hypothesized that iron accumulation leads to membrane lipid peroxidation, muscle wasting, increased susceptibility to injury, and impaired muscle regeneration. METHODS: To examine the role of iron in age-related muscle atrophy, we compared the skeletal muscles of 3-month-old with 22- to 24-month-old 129SvEv FVBM mice. We assessed iron distribution and total elemental iron using laser ablation inductively coupled plasma mass spectrometry and Perls' stain on skeletal muscle cross-sections. In addition, old mice underwent ischaemia-reperfusion (IR) injury (90 min ischaemia), and muscle regeneration was assessed 14 days after injury. Immunoblotting was used to determine lipid peroxidation (4HNE) and iron-related proteins. To determine whether muscle iron content can be altered, old mice were treated with deferiprone (DFP) in the drinking water, and we assessed its effects on muscle regeneration after injury. RESULTS: We observed a significant increase in total elemental iron (+43%, P < 0.05) and lipid peroxidation (4HNE: +76%, P < 0.05) in tibialis anterior muscles of old mice. Iron was further increased after injury (adult: +81%, old: +135%, P < 0.05) and associated with increased lipid peroxidation (+41%, P < 0.05). Administration of DFP did not impact iron or measures of lipid peroxidation in skeletal muscle or modulate muscle mass. Increased muscle iron concentration and lipid peroxidation were associated with less efficient regeneration, evident from the smaller fibres in cross-sections of tibialis anterior muscles (-24%, P < 0.05) and an increased percentage of fibres with centralized nuclei (+4124%, P < 0.05) in muscles of old compared with adult mice. Administration of DFP lowered iron after IR injury (PRE: -32%, P < 0.05 and POST: -41%, P < 0.05), but did not translate to structural improvements. CONCLUSIONS: Muscles from old mice have increased iron levels, which are associated with increased lipid peroxidation, increased susceptibility to IR injury, and impaired muscle regeneration. Our results suggest that iron is involved in effective muscle regeneration, highlighting the importance of iron homeostasis in muscle atrophy and muscle repair.
ItemThe hypoxia imaging agent Cu-II(atsm) is neuroprotective and improves motor and cognitive functions in multiple animal models of Parkinson's diseaseHung, LW ; Villemagne, VL ; Cheng, L ; Sherratt, NA ; Ayton, S ; White, AR ; Crouch, PJ ; Lim, S ; Leong, SL ; Wilkins, S ; George, J ; Roberts, BR ; Pham, CLL ; Liu, X ; Chiu, FCK ; Shackleford, DM ; Powell, AK ; Masters, CL ; Bush, AI ; O'Keefe, G ; Culvenor, JG ; Cappai, R ; Cherny, RA ; Donnelly, PS ; Hill, AF ; Finkelstein, DI ; Barnham, KJ (ROCKEFELLER UNIV PRESS, 2012-04-09)Parkinson's disease (PD) is a progressive, chronic disease characterized by dyskinesia, rigidity, instability, and tremors. The disease is defined by the presence of Lewy bodies, which primarily consist of aggregated α-synuclein protein, and is accompanied by the loss of monoaminergic neurons. Current therapeutic strategies only give symptomatic relief of motor impairment and do not address the underlying neurodegeneration. Hence, we have identified Cu(II)(atsm) as a potential therapeutic for PD. Drug administration to four different animal models of PD resulted in improved motor and cognition function, rescued nigral cell loss, and improved dopamine metabolism. In vitro, this compound is able to inhibit the effects of peroxynitrite-driven toxicity, including the formation of nitrated α-synuclein oligomers. Our results show that Cu(II)(atsm) is effective in reversing parkinsonian defects in animal models and has the potential to be a successful treatment of PD.
ItemTau-mediated iron export prevents ferroptotic damage after ischemic strokeTuo, Q-Z ; Lei, P ; Jackman, KA ; Li, X-I ; Xiong, H ; Li, X-L ; Liuyang, Z-Y ; Roisman, L ; Zhang, S-T ; Ayton, S ; Wang, Q ; Crouch, PJ ; Ganio, K ; Wang, X-C ; Pei, L ; Adlard, PA ; Lu, Y-M ; Cappai, R ; Wang, J-Z ; Liu, R ; Bush, AI (NATURE PUBLISHING GROUP, 2017-11-01)Functional failure of tau contributes to age-dependent, iron-mediated neurotoxicity, and as iron accumulates in ischemic stroke tissue, we hypothesized that tau failure may exaggerate ischemia-reperfusion-related toxicity. Indeed, unilateral, transient middle cerebral artery occlusion (MCAO) suppressed hemispheric tau and increased iron levels in young (3-month-old) mice and rats. Wild-type mice were protected by iron-targeted interventions: ceruloplasmin and amyloid precursor protein ectodomain, as well as ferroptosis inhibitors. At this age, tau-knockout mice did not express elevated brain iron and were protected against hemispheric reperfusion injury following MCAO, indicating that tau suppression may prevent ferroptosis. However, the accelerated age-dependent brain iron accumulation that occurs in tau-knockout mice at 12 months of age negated the protective benefit of tau suppression against MCAO-induced focal cerebral ischemia-reperfusion injury. The protective benefit of tau knockout was revived in older mice by iron-targeting interventions. These findings introduce tau-iron interaction as a pleiotropic modulator of ferroptosis and ischemic stroke outcome.
ItemNo Preview AvailableA comparison of ceruloplasmin to biological polyanions in promoting the oxidation of Fe2+ under physiologically relevant conditionsWong, BX ; Ayton, S ; Lam, LQ ; Lei, P ; Adlard, PA ; Bush, AI ; Duce, JA (ELSEVIER SCIENCE BV, 2014-12-01)BACKGROUND: Iron oxidation is thought to be predominantly handled enzymatically in the body, to minimize spontaneous combustion with oxygen and to facilitate cellular iron export by loading transferrin. This process may be impaired in disease, and requires more accurate analytical assays to interrogate enzymatic- and auto-oxidation within a physiologically relevant environment. METHOD: A new triplex ferroxidase activity assay has been developed that overcomes the previous assay limitations of measuring iron oxidation at a physiologically relevant pH and salinity. RESULTS: Revised enzymatic kinetics for ceruloplasmin (Vmax≈35μMFe(3+)/min/μM; Km≈15μM) are provided under physiological conditions, and inhibition by sodium azide (Ki for Ferric Gain 78.3μM, Ki for transferrin loading 8.1×10(4)μM) is quantified. We also used this assay to characterize the non-enzymatic oxidation of iron that proceeded linearly under physiological conditions. CONCLUSIONS AND GENERAL SIGNIFICANCE: These findings indicate that the requirement of an enzyme to oxidize iron may only be necessary under conditions of adverse pH or anionic strength, for example from hypoxia. In a normal physiological environment, Fe(3+) incorporation into transferrin would be sufficiently enabled by the biological polyanions that are prevalent within extracellular fluids.
ItemCeruloplasmin and beta-amyloid precursor protein confer neuroprotection in traumatic brain injury and lower neuronal ironAyton, S ; Zhang, M ; Roberts, BR ; Lam, LQ ; Lind, M ; McLean, C ; Bush, AI ; Frugier, T ; Crack, PJ ; Duce, JA (ELSEVIER SCIENCE INC, 2014-04-01)Traumatic brain injury (TBI) is in part complicated by pro-oxidant iron elevation independent of brain hemorrhage. Ceruloplasmin (CP) and β-amyloid protein precursor (APP) are known neuroprotective proteins that reduce oxidative damage through iron regulation. We surveyed iron, CP, and APP in brain tissue from control and TBI-affected patients who were stratified according to time of death following injury. We observed CP and APP induction after TBI accompanying iron accumulation. Elevated APP and CP expression was also observed in a mouse model of focal cortical contusion injury concomitant with iron elevation. To determine if changes in APP or CP were neuroprotective we employed the same TBI model on APP(-/-) and CP(-/-) mice and found that both exhibited exaggerated infarct volume and iron accumulation postinjury. Evidence supports a regulatory role of both proteins in defence against iron-induced oxidative damage after TBI, which presents as a tractable therapeutic target.