miR-129 Promotes the Proliferation of Alzheimer’s Neuronal Cells by Binding the 3′ Untranslated Regions (3′ UTR) of Amyloid Precursor Protein (APP)

Alzheimer’s disease (AD) is a neurodegenerative disease with memory loss and cognitive impairment. Short non-coding RNAs (miRNAs) are potential biomarkers and therapeutic targets for AD. This study aims to investigate miR-129’s role in AD. miR-129 and amyloid precursor protein (APP) expression was measured by Q-PCR, and LC3, p62, ATG5, Bcl-2, p-Tau and Caspase3 protein was detected by Western blot. Hydrogenase kits and DCFH-DA detected cell apoptosis, cytotoxicity and ROS generation. The interaction between APP and miR-129 was assessed by luciferase report experiment. HE staining and TUNEL assay evaluated hippocampal neuron damage. In AD patient serum, AD transgenic (TG) mouse brain tissue, and AB1-42-treated SH-SY5Y cells, miR-129 was downregulated but autophagy was increased. Overexpression of miR-129 reduced cell damage induced by AB1-42, and miR-129 can directly regulate APP expression by binding APP 3′-UTR. miR-129 inhibitors reversed the protective effect of shAPP on AB1-42-induced cell damage. In addition, miR-129 overexpression reduced neuronal damage through inhibiting autophagy in vivo. APP expression in AD patient and AD cell model was significantly increased compared to controls. Aβ-42 treatment caused up-regulation of APP expression, while APP knockdown inhibited neurons through autophagy. In conclusion, miR-129 overexpression can regulate autophagy by targeting APP5, thereby reducing neuronal damage in AD. These findings provide a new perspective for treating AD.

The orphan drug dichloroacetate reduces amyloid beta-peptide production whilst promoting non-amyloidogenic proteolysis of the amyloid precursor protein

The amyloid cascade hypothesis proposes that excessive accumulation of amyloid beta-peptides is the initiating event in Alzheimer’s disease. These neurotoxic peptides are generated from the amyloid precursor protein via sequential cleavage by β- and γ-secretases in the ’amyloidogenic’ proteolytic pathway. Alternatively, the amyloid precursor protein can be processed via the ’non-amyloidogenic’ pathway which, through the action of the α-secretase a disintegrin and metalloproteinase (ADAM) 10, both precludes amyloid beta-peptide formation and has the additional benefit of generating a neuroprotective soluble amyloid precursor protein fragment, sAPPα. In the current study, we investigated whether the orphan drug, dichloroacetate, could alter amyloid precursor protein proteolysis. In SH-SY5Y neuroblastoma cells, dichloroacetate enhanced sAPPα generation whilst inhibiting β–secretase processing of endogenous amyloid precursor protein and the subsequent generation of amyloid beta-peptides. Over-expression of the amyloid precursor protein partly ablated the effect of dichloroacetate on amyloidogenic and non-amyloidogenic processing whilst over-expression of the β-secretase only ablated the effect on amyloidogenic processing. Similar enhancement of ADAM-mediated amyloid precursor protein processing by dichloroacetate was observed in unrelated cell lines and the effect was not exclusive to the amyloid precursor protein as an ADAM substrate, as indicated by dichloroacetate-enhanced proteolysis of the Notch ligand, Jagged1. Despite altering proteolysis of the amyloid precursor protein, dichloroacetate did not significantly affect the expression/activity of α-, β- or γ-secretases. In conclusion, dichloroacetate can inhibit amyloidogenic and promote non-amyloidogenic proteolysis of the amyloid precursor protein. Given the small size and blood-brain-barrier permeability of the drug, further research into its mechanism of action with respect to APP proteolysis may lead to the development of therapies for slowing the progression of Alzheimer’s disease.

Accumulation of APP-CTF induces mitophagy dysfunction in the iNSCs model of Alzheimer’s disease

Mitochondrial dysfunction is associated with familial Alzheimer’s disease (fAD), and the accumulation of damaged mitochondria has been reported as an initial symptom that further contributes to disease progression. In the amyloidogenic pathway, the amyloid precursor protein (APP) is cleaved by β-secretase to generate a C-terminal fragment, which is then cleaved by γ-secretase to produce amyloid-beta (Aβ). The accumulation of Aβ and its detrimental effect on mitochondrial function are well known, yet the amyloid precursor protein-derived C-terminal fragments (APP-CTFs) contributing to this pathology have rarely been reported. We demonstrated the effects of APP-CTFs-related pathology using induced neural stem cells (iNSCs) from AD patient-derived fibroblasts. APP-CTFs accumulation was demonstrated to mainly occur within mitochondrial domains and to be both a cause and a consequence of mitochondrial dysfunction. APP-CTFs accumulation also resulted in mitophagy failure, as validated by increased LC3-II and p62 and inconsistent PTEN-induced kinase 1 (PINK1)/E3 ubiquitin ligase (Parkin) recruitment to mitochondria and failed fusion of mitochondria and lysosomes. The accumulation of APP-CTFs and the causality of impaired mitophagy function were also verified in AD patient-iNSCs. Furthermore, we confirmed this pathological loop in presenilin knockout iNSCs (PSEN KO-iNSCs) because APP-CTFs accumulation is due to γ-secretase blockage and similarly occurs in presenilin-deficient cells. In the present work, we report that the contribution of APP-CTFs accumulation is associated with mitochondrial dysfunction and mitophagy failure in AD patient-iNSCs as well as PSEN KO-iNSCs.

Effect of soluble amyloid precursor protein-alpha on adult hippocampal neurogenesis in a mouse model of Alzheimer’s disease

Soluble amyloid precursor protein-alpha (sAPPα) is a regulator of neuronal and memory mechanisms, while also having neurogenic and neuroprotective effects in the brain. As adult hippocampal neurogenesis is impaired in Alzheimer’s disease, we tested the hypothesis that sAPPα delivery would rescue adult hippocampal neurogenesis in an APP/PS1 mouse model of Alzheimer’s disease. An adeno-associated virus-9 (AAV9) encoding murine sAPPα was injected into the hippocampus of 8-month-old wild-type and APP/PS1 mice, and later two different thymidine analogues (XdU) were systemically injected to label adult-born cells at different time points after viral transduction. The proliferation of adult-born cells, cell survival after eight weeks, and cell differentiation into either neurons or astrocytes was studied. Proliferation was impaired in APP/PS1 mice but was restored to wild-type levels by viral expression of sAPPα. In contrast, sAPPα overexpression failed to rescue the survival of XdU+-labelled cells that was impaired in APP/PS1 mice, although it did cause a significant increase in the area density of astrocytes in the granule cell layer across both genotypes. Finally, viral expression of sAPPα reduced amyloid-beta plaque load in APP/PS1 mice in the dentate gyrus and somatosensory cortex. These data add further evidence that increased levels of sAPPα could be therapeutic for the cognitive decline in AD, in part through restoration of the proliferation of neural progenitor cells in adults.

Amyloid Precursor Protein A713T Mutation in Calabrian Patients with Alzheimer’s Disease: A Population Genomics Approach to Estimate Inheritance from a Common Ancestor

Mutation A713T in the amyloid precursor protein (APP) has been linked to cases of Alzheimer’s disease (AD), cerebral amyloid angiopathy (CAA) and cerebrovascular disease. Despite its rarity, it has been observed in several families from the same geographical area, in the Calabria region in Southern Italy. Genotyping of 720,000 genome-wide SNPs with the HumanOmniExpress BeadChip was performed for six patients that were representative of apparently unrelated Calabrian families, as well as a Belgian subject of Italian descent (all with the same A713T mutation and disease). Their genomic structure and genetic relationships were analyzed. Demographic reconstruction and coalescent theory were applied to estimate the time of the most recent common ancestor (tMRCA) among patients. Results show that all A713T carriers fell into the genetic variability of Southern Italy and were not more closely related to each other than to any other healthy Calabrian individual. However, five out of seven patients shared a 1.7 Mbp-long DNA segment centered on the A713T mutation, making it possible to estimate a tMRCA for its common origin in the Calabrian region dating back over 1000 years. The analysis of affected individuals with methodologies based on human population genomics thus provides informative insights in support of clinical observations and biomedical research.

Control of β-Site Amyloid Precursor Protein-Cleaving Enzyme-1 Expression by Protein Kinase C-λ/ι and Nuclear Factor κ-B

Βackground: β-Amyloid precursor protein-cleaving enzyme-1 (BACE1) initiates the production of Aβ-peptides that form Aβ-plaque in Alzheimer’s disease. Methods: Reportedly, acute insulin treatment in normal mice, and hyperinsulinemia in high-fat-fed (HFF) obese/diabetic mice, increase BACE1 activity and levels of Aβ-peptides and phospho- -thr-231-tau in the brain; moreover, these effects are blocked by PKC-λ/ι inhibitors. However, as chemical inhibitors may affect unsuspected targets, we presently used knockout methodology to further examine PKC-λ/ι requirements. We found that total-body heterozygous PKC-λ knockout reduced acute stimulatory effects of insulin and chronic effects of hyperinsulinemia in HFF/obese/diabetic mice, on brain PKC-λ activity and production of Aβ1-40/42 and phospho-thr-231-tau. This protection in HFF mice may reflect that hepatic PKC-λ haploinsufficiency prevents the development of glucose intolerance and hyperinsulinemia. Results: On the other hand, heterozygous knockout of PKC-λ markedly reduced brain levels of BACE1 protein and mRNA, and this may reflect diminished activation of nuclear factor kappa-B (NFκB), which is activated by PKC-λ and increases BACE1 and proinflammatory cytokine transcription. Accordingly, whereas intravenous administration of aPKC inhibitor diminished aPKC activity and BACE1 levels by 50% in the brain and 90% in the liver, nasally-administered inhibitor reduced aPKC activity and BACE1 mRNA and protein levels by 50-70% in the brain while sparing the liver. Additionally, 24-hour insulin treatment in cultured human-derived neurons increased NFκB activity and BACE1 levels, and these effects were blocked by various PKC-λ/ι inhibitors. Conclusion: PKC-λ/ι controls NFκB activity and BACE1 expression; PKC-λ/ι inhibitors may be used nasally to target brain PKC-λ/ι or systemically to block both liver and brain PKC-λ/ι, to regulate NFκB-dependent BACE1 and proinflammatory cytokine expression.

Molecular Mechanisms of Cardiac Amyloidosis

Cardiac involvement has a profound effect on the prognosis of patients with systemic amyloidosis. Therapeutic methods for suppressing the production of causative proteins have been developed for ATTR amyloidosis and AL amyloidosis, which show cardiac involvement, and the prognosis has been improved. However, a method for removing deposited amyloid has not been established. Methods for reducing cytotoxicity caused by amyloid deposition and amyloid precursor protein to protect cardiovascular cells are also needed. In this review, we outline the molecular mechanisms and treatments of cardiac amyloidosis.

White Matter Beta-Amyloid Precursor Protein Immunoreactivity in Autopsied Subjects With and Without COVID-19

The coronavirus SARS-CoV-2 causes COVID-19, a predominantly respiratory disease that has been reported to be associated with numerous neurological signs, symptoms and syndromes. More than 20 published studies have used RT-PCR methods to determine viral SARS-CoV-2 genomic presence in postmortem brain tissue and the overall impression is that viral brain invasion is relatively uncommon and occurs in low copy numbers, supporting indirect mechanisms as the cause of most neurological phenomena. Hypoxic-ischemic brain injury and stroke are one such possible indirect mechanism, as acute ischemia or stroke concurrence with COVID-19 has been reported as being 0.5% to 20%. Immunohistochemical stains for beta-amyloid precursor protein (APP) have been suggested to be a signature change of hypoxic leukoencephalopathy or COVID-19 brain disease, although prior reports have not had a non-COVID-19 control group. We therefore compared the prevalence and intensity of white matter APP staining in the brains of subjects dying with and without COVID-19. Clinical and neuropathological results, including semi-quantitative assessment of the density of white matter APP staining, were compared between 20 COVID-19 cases and 20 pre-COVID-19 autopsy cases, including 10 cases with autopsy-proven non-COVID-19 pneumonia and 10 cases without pneumonia. Positive APP white matter staining in at least one of the two brain regions (precentral gyrus and cingulate gyrus) studied was not significantly more common in COVID-19 vs controls (14/20 vs 12/20). Comparing density scores from both brain regions combined, the mean scores for COVID-19 cases were higher than those for controls of both types together but not significantly different when restricting to controls with pneumonia. Among control cases, cases with pneumonia had significantly higher scores. The presence or absence of a major neuropathologically-defined neurodegenerative disorder did not significantly affect the APP scores. The major finding is that while APP white matter staining cannot be regarded as a specific marker of COVID-19, as it does not occur with significantly greater probability in in COVID-19 brains as compared to non-COVID-19 brains, it is possible that white matter APP staining, representing acute or subacute axonal damage, may be a common occurrence in the perimortem period, and that it may be more intense in subjects dying with pneumonia, regardless of cause.