Genetic ablation of the p66(Shc) adaptor protein reverses cognitive deficits and improves mitochondrial function in an APP transgenic mouse model of Alzheimer's disease

The mammalian ShcA adaptor protein p66(Shc) is a key regulator of mitochondrial reactive oxygen species (ROS) production and has previously been shown to mediate amyloid β (Aβ)-peptide-induced cytotoxicity in vitro. Moreover, p66(Shc) is involved in mammalian longevity and lifespan determination as revealed in the p66(Shc) knockout mice, which are characterized by a 30% prolonged lifespan, lower ROS levels and protection from age-related impairment of physical and cognitive performance. In this study, we hypothesized a role for p66(Shc) in Aβ-induced toxicity in vivo and investigated the effects of genetic p66(Shc) deletion in the PSAPP transgenic mice, an established Alzheimer's disease mouse model of β-amyloidosis. p66(Shc)-ablated PSAPP mice were characterized by an improved survival and a complete rescue of Aβ-induced cognitive deficits at the age of 15 months. Importantly, these beneficial effects on survival and cognitive performance were independent of Aβ levels and amyloid plaque deposition, but were associated with improved brain mitochondrial respiration, a reversal of mitochondrial complex I dysfunction, restored adenosine triphosphate production and reduced ROS levels. The results of this study support a role for p66(Shc) in Aβ-related mitochondrial dysfunction and oxidative damage in vivo, and suggest that p66(Shc) ablation may be a promising novel therapeutic strategy against Aβ-induced toxicity and cognitive impairment.Molecular Psychiatry advance online publication, 19 July 2016; doi:10.1038/mp.2016.112

Deleterious role of endothelial lectin-like oxidized low-density lipoprotein receptor-1 in ischaemia/reperfusion cerebral injury

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is implicated in cardiovascular disease by modulating apoptosis and oxidative stress. We hypothesized that LOX-1 may be involved in pathophysiology of stroke by mediating ischaemia/reperfusion (I/R)-dependent cell death. Transient middle cerebral artery occlusion (tMCAO) was performed in wild-type (WT) mice, endothelial-specific LOX-1 transgenic mice (eLOX-1TG) and WT animals treated with LOX-1 silencing RNA (siRNA). In WT mice exposed to tMCAO, LOX-1 expression and function were increased in the MCA. Compared to WT animals, eLOX-1TG mice displayed increased stroke volumes and worsened outcome after I/R. Conversely, LOX-1-silencing decreased both stroke volume and neurological impairment. Similarly, in HBMVECs, hypoxia/reoxygenation increased LOX-1 expression, while LOX-1 overexpressing cells showed increased death following hypoxia reoxygenation. Increased caspase-3 activation was observed following LOX-1 overexpression both in vivo and in vitro, thus representing a likely mediator. Finally, monocytes from ischaemic stroke patients exhibited increased LOX-1 expression which also correlated with disease severity. Our data unequivocally demonstrate a key role for LOX-1 in determining outcome following I/R brain damage. Our findings could be corroborated in human brain endothelial cells and monocytes from patients, underscoring their translational relevance and suggesting siRNA-mediated LOX-1 knockdown as a novel therapeutic strategy for stroke patients

Detecting aquatic and terrestrial biodiversity in a tropical estuary using environmental DNA

Estuaries are characterized by a tidal regime and are strongly influenced by hydrodynamics and host diverse and highly dynamic habitats, from fresh, brackish, or saltwater to terrestrial, whose biodiversity is especially difficult to monitor. Here, we investigated the potential of environmental DNA (eDNA) metabarcoding, with three primer sets targeting different regions of the mitochondrial DNA 12S ribosomal RNA gene, to detect vertebrate diversity in the estuary of the Don Diego River in Colombia. With eDNA, we detected not only aquatic organisms, including fishes, amphibians, and reptiles, but also a large diversity of terrestrial, arboreal, and flying vertebrates, including mammals and birds, living in the estuary surroundings. Further, the eDNA signal remained relatively localized along the watercourse. A transect from the deep outer section of the estuary, across the river mouth toward the inner section of the river, showed marked taxonomic turnover from typical marine to freshwater fishes, while eDNA of terrestrial and arboreal species was mainly found in the inner section of the estuary. Our results indicate that eDNA enables the detection of a large diversity of vertebrates and could become an important tool for biodiversity monitoring in estuaries, where water integrates information across the ecosystem. Abstract in Spanish is available with online material

Post-ischaemic silencing of p66Shc reduces ischaemia/reperfusion brain injury and its expression correlates to clinical outcome in stroke

Aim: Constitutive genetic deletion of the adaptor protein p66Shc was shown to protect from ischaemia/reperfusion injury. Here, we aimed at understanding the molecular mechanisms underlying this effect in stroke and studied p66Shc gene regulation in human ischaemic stroke. Methods and Results: Ischaemia/reperfusion brain injury was induced by performing a transient middle cerebral artery occlusion surgery on wild-type mice. After the ischaemic episode and upon reperfusion, small interfering RNA targeting p66Shc was injected intravenously. We observed that post-ischaemic p66Shc knockdown preserved blood-brain barrier integrity that resulted in improved stroke outcome, as identified by smaller lesion volumes, decreased neurological deficits, and increased survival. Experiments on primary human brain microvascular endothelial cells demonstrated that silencing of the adaptor protein p66Shc preserves claudin-5 protein levels during hypoxia/reoxygenation by reducing nicotinamide adenine dinucleotide phosphate oxidase activity and reactive oxygen species production. Further, we found that in peripheral blood monocytes of acute ischaemic stroke patients p66Shc gene expression is transiently increased and that this increase correlates with short-term neurological outcome. Conclusion: Post-ischaemic silencing of p66Shc upon reperfusion improves stroke outcome in mice while the expression of p66Shc gene correlates with short-term outcome in patients with ischaemic stroke

Endothelial LOX-1 activation differentially regulates arterial thrombus formation depending on oxLDL levels: role of the Oct-1/SIRT1 and ERK1/2 pathways

Aims The lectin-like oxLDL receptor-1 (LOX-1) promotes endothelial uptake of oxidized low-density lipoprotein (oxLDL) and plays an important role in atherosclerosis and acute coronary syndromes (ACS). However, its role in arterial thrombus formation remains unknown. We investigated whether LOX-1 plays a role in arterial thrombus formation in vivo at different levels of oxLDL using endothelial-specific LOX-1 transgenic mice (LOX-1TG) and a photochemical injury thrombosis model of the carotid artery. Methods and results In mice fed a normal chow diet, time to arterial occlusion was unexpectedly prolonged in LOX-1TG as compared to WT. In line with this, tissue factor (TF) expression and activity in carotid arteries of LOX-1TG mice were reduced by half. This effect was mediated by activation of octamer transcription factor 1 (Oct-1) leading to upregulation of the mammalian deacetylase silent information regulator-two 1 (SIRT1) via binding to its promoter and subsequent inhibition of NF-κB signaling. In contrast, intravenous injection of oxLDL as well as high cholesterol diet for 6 weeks led to a switch from the Oct-1/SIRT1 signal transduction pathway to the ERK1/2 pathway and in turn to an enhanced thrombotic response with shortened occlusion time. Conclusions Thus, LOX-1 differentially regulates thrombus formation in vivo depending on the degree of activation by oxLDL. At low oxLDL levels LOX-1 activates the protective Oct-1/SIRT1 pathway, while at higher levels of the lipoprotein switches to the thrombogenic ERK1/2 pathway. These findings may be important for arterial thrombus formation in ACS and suggest that SIRT1 may represent a novel therapeutic target in this context