Promoting Autophagic Clearance: Viable Therapeutic Targets in Alzheimer’s Disease

Many neurodegenerative disorders are characterized by the aberrant accumulation of aggregate-prone proteins. Alzheimer’s disease (AD) is associated with the buildup of β-amyloid peptides and tau, which aggregate into extracellular plaques and neurofibrillary tangles, respectively. Multiple studies have linked dysfunctional intracellular degradation mechanisms with AD pathogenesis. One such pathway is the autophagy–lysosomal system, which involves the delivery of large protein aggregates/inclusions and organelles to lysosomes through the formation, trafficking, and degradation of double-membrane structures known as autophagosomes. Converging data suggest that promoting autophagic degradation, either by inducing autophagosome formation or enhancing lysosomal digestion, provides viable therapeutic strategies. In this review, we discuss compounds that can augment autophagic clearance and may ameliorate disease-related pathology in cell and mouse models of AD. Canonical autophagy induction is associated with multiple signaling cascades; on the one hand, the best characterized is mammalian target of rapamycin (mTOR). Accordingly, multiple mTOR-dependent and mTOR-independent drugs that stimulate autophagy have been tested in preclinical models. On the other hand, there is a growing list of drugs that can enhance the later stages of autophagic flux by stabilizing microtubule-mediated trafficking, promoting lysosomal fusion, or bolstering lysosomal enzyme function. Although altering the different stages of autophagy provides many potential targets for AD therapeutic interventions, it is important to consider how autophagy drugs might also disturb the delicate balance between autophagosome formation and lysosomal degradation.

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The online version of this article (doi:10.1007/s13311-014-0320-z) contains supplementary material, which is available to authorized users.     

Mammalian target of rapamycin: A valid therapeutic target through the autophagy pathway for alzheimer's disease?

Autophagy plays a critical role in multiple pathological lesions of Alzheimer's disease (AD), such as the formation of amyloid plaques from amyloid-β (Aβ) production and accumulation via dysregulating amyloid precursor protein turnover and enhancing the activity of β- and/or γ-secretases, intraneuronal neurofibrillary tangles (NFT) because of tau hyperphosphorylation, and neuronal apoptosis. Dysfunction of the autophagy-lysosome system also contributes to Aβ accumulation and the formation of tau oligomers and insoluble aggregates, because induction of autophagy enhances the clearance of both soluble and aggregated forms of Aβ and tau proteins. The mammalian target of rapamycin (mTOR) pathway plays a central role in controlling protein homeostasis and negatively regulates autophagy. Inhibition of mTOR by rapamycin improves cognitive deficits and rescues Aβ pathology and NFTs by increasing autophagy. Several mTOR signaling components may be potential biomarkers of cognitive impairment in the clinical diagnosis of AD. Thus, mTOR-related agents through the control of autophagy-lysosome protein degradation are emerging as an important therapeutic target for AD.     

Neuroprotection of rapamycin in lactacystin-induced neurodegeneration via autophagy enhancement

The ubiquitin-proteasome system (UPS) and the autophagy-lysosomal pathway (ALP) are the two most important cellular mechanisms for protein degradation. To investigate the role of autophagy in reversing neuronal injury, the proteasome inhibitor lactacystin was used to cause UPS dysfunction in differentiated PC12 cells and in C57BL/6 mice and rapamycin was used as an autophagy enhancer. The results showed that rapamycin pre-treatment attenuated lactacystin-induced apoptosis and reduced lactacystin-induced ubiquitinated protein aggregation in differentiated PC12 cells. The observed protection was partially blocked by the autophagy inhibitor 3-methyladenine. Furthermore, post-treatment of mice with rapamycin significantly attenuated lactacystin-induced loss of nigral DA neurons and the reduction of striatal DA levels. The lactacystin-induced high molecular ubiquitinated proteins were also attenuated by rapamycin treatment in vivo. In addition, as a chemical compound, rapamycin caused an increase of bcl2 protein level and blocked the release of cytochrome c from mitochondria to cytosal. We concluded that the neuroprotective effect of rapamycin is partially mediated by autophagy enhancement through enhanced degradation of misfolded proteins and autophagy enhancement may be considered to be a promising strategy to prevent diseases associated with misfolded/aggregated proteins, such as Parkinson's disease.     

The formation of peripheral myelin protein 22 aggregates is hindered by the enhancement of autophagy and expression of cytoplasmic chaperones

The accumulation of misfolded proteins is associated with various neurodegenerative conditions. Peripheral myelin protein 22 (PMP22) is a hereditary neuropathy-linked, short-lived molecule that forms aggresomes when the proteasome is inhibited or the protein is mutated. We previously showed that the removal of pre-existing PMP22 aggregates is assisted by autophagy. Here we examined whether the accumulation of such aggregates could be suppressed by experimental induction of autophagy and/or chaperones. Enhancement of autophagy during proteasome inhibition hinders protein aggregate formation and correlates with a reduction in accumulated proteasome substrates. Conversely, simultaneous inhibition of autophagy and the proteasome augments the formation of aggregates. An increase of heat shock protein levels by geldanamycin treatment or heat shock preconditioning similarly hampers aggresome formation. The beneficial effects of autophagy and chaperones in preventing the accumulation of misfolded PMP22 are additive and provide a potential avenue for therapeutic approaches in hereditary neuropathies linked to PMP22 mutations.     

The Effect of Autophagy in the Process of Adipose-Derived Stromal Cells Differentiation into Astrocytes

Autophagy plays different roles in the growth and development process of different cells. The role of autophagy in the differentiation process of adult adipose-derived stromal cells (ADSCs) into astrocytes is unclear. We researched the role of autophagy in the induction process by adding autophagy agonist rapamycin, which was not added in the control group. Immunocytochemistry showed that the expression of glial fibrillary acidic protein (GFAP) was increased gradually with the extending reaction time and had reached the peak on the 14th day. Typical autophagy ultrastructure, including autophagic bodies and self-macrophage lysosomal, was shown under transmission electron microscopy (TEM) when cells were induced for 14 days. By methyl thiazolyl tetrazolium (MTT) assay, we found that the number of living cells was reduced gradually, and early apoptosis rate was increased by flow cytometry. We observed that the differentiation ratio, the number of living cells, and the positive expression rates of GFAP in the rapamycin group were higher than those in the control group when ADSCs were induced for 48 h and 7 days (P?<?0.01); however, the rates of early apoptosis were lower than those in the control group. The positive rate of microtubule-associated protein light chain 3 (LC3) in the rapamycin group had been up to 88.87 % on the 7th day (P?<?0.01), but not obvious with extending time. After 14 days of induction, the optical density (OD) value of surviving cells was declined, and early apoptosis rate was increased gradually. The results showed that adding autophagy agonist to the inducers may enhance intensity of autophagy, shorten the induction time, and improve the efficiency of differentiation.     

Amyloid-β accumulation caused by chloroquine injections precedes ER stress and autophagosome formation in rat skeletal muscle

Chloroquine, an anti-malaria drug, is known to cause myopathy with rimmed vacuole formation. Although it disrupts the lysosomal degradation of proteins, the precise mechanism underlying muscle fiber degeneration has remained unclear. We investigated the temporal profiles of muscle fiber degeneration in chloroquine-treated rats, paying special attention to endoplasmic reticulum (ER) stress and autophagy. Male Wistar rats were intraperitoneally injected with chloroquine diphosphate at a dosage of 50 mg/kg body weight every day. We examined the localization and levels of proteins related to ER stress and autophagy in soleus muscle by means of immunohistochemistry and Western blotting at 3, 5, and 7 weeks after the beginning of the treatment. At 3 weeks, the levels of LC3-II and amyloid-β (Aβ) were increased. At 5 weeks, an unfolded protein response took place. At 7 weeks, rimmed vacuole formation became obvious. Interestingly, SERCA2, a Ca2+ -pump ATPase located in the endoplasmic/sarcoplasmic reticulum membrane was up-regulated at 5 weeks after treatment, but declined to the control level by 7 weeks. Taken together, these findings suggest that Aβ accumulation (at 3 weeks) caused by the disruption of lysosomal enzymes precedes an unfolded protein response (at 5 weeks). Next, activation of autophagy occurs (at 7 weeks), probably using sarcoplasmic reticulum membrane, the amount of which was increased. Chloroquine-treated rats could be useful for investigating the pathogenesis of diseases related to Aβ accumulation.     

蛋白酶体在信号转导和神经退行性疾病中的作用

哺乳动物细胞内有两条蛋白降解途径：泛素-蛋白酶体通路和自噬-溶酶体通路。蛋白酶体由多种蛋白亚单位组成,在降解细胞短寿命蛋白中有重要作用。近来的研究发现蛋白酶体降解系统在细胞信号转导和细胞功能中有重要作用。蛋白酶体功能障碍或失调可能参与某些神经退行性疾病的致病机制。本文综述蛋白酶体在细胞信号转导中的生物学功能和在神经退行性疾病中的可能作用。     

The emerging role of autophagic-lysosomal dysfunction in Gaucher disease and Parkinson's disease

Gaucher disease(GD),the commonest lysosomal storage disorder,results from the lack or functional deficiency of glucocerebrosidase(GCase) secondary to mutations in the GBA1 gene.There is an established association between GBA1 mutations and Parkinson's disease(PD),and indeed GBA1 mutations are now considered to be the greatest genetic risk factor for PD.Impaired lysosomal-autophagic degradation of cellular proteins,including α-synuclein(α-syn),is implicated in the pathogenesis of PD,and there is increasing evidence for this also in GD and GBA1-PD.Indeed we have recently shown in a Drosophila model lacking neuronal GCase,that there are clear lysosomal-autophagic defects in association with synaptic loss and neurodegeneration.In addition,we demonstrated alterations in mechanistic target of rapamycin complex 1(mTORC1) signaling and functional rescue of the lifespan,locomotor defects and hypersensitivity to oxidative stress on treatment of GCase-deficient flies with the mT OR inhibitor rapamycin.Moreover,a number of other recent studies have shown autophagy-lysosomal system(ALS) dysfunction,with specific defects in both chaperone-mediated autophagy(CMA),as well as macroautophagy,in GD and GBA1-PD model systems.Lastly we discuss the possible therapeutic benefits of inhibiting mT OR using drugs such as rapamycin to reverse the autophagy defects in GD and PD.     

Altered lysosomal positioning affects lysosomal functions in a cellular model of Huntington's disease

Huntington's disease (HD) is a hereditary and devastating neurodegenerative disorder caused by a mutation in the huntingtin protein. Understanding the functions of normal and mutant huntingtin protein is the key to revealing the pathogenesis of HD and developing therapeutic targets. Huntingtin plays an important role in vesicular and organelle trafficking. Lysosomes are dynamic organelles that integrate several degradative pathways and regulate the activity of mammalian target of rapamycin complex 1 (mTORC1). In the present study, we found that the perinuclear accumulation of lysosomes was increased in a cellular model of HD derived from HD knock‐in mice and primary fibroblasts from an HD patient. This perinuclear lysosomal accumulation could be reversed when normal huntingtin was overexpressed in HD cells. When we further investigated the functional significance of the increased perinuclear lysosomal accumulation in HD cells, we demonstrated that basal mTORC1 activity was increased in HD cells. In addition, autophagic influx was also increased in HD cells in response to serum deprivation, which leads to premature fusion of lysosomes with autophagosomes. Taken together, our data suggest that the increased perinuclear accumulation of lysosomes may play an important role in HD pathogenesis by altering lysosomal‐dependent functions.     

Rotenone Inhibits Autophagic Flux Prior to Inducing Cell Death

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Monitoring Autophagy in the Treatment of Protein Aggregate Diseases: Steps Toward Identifying Autophagic Biomarkers

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Dorfin ameliorates phenotypes in a transgenic mouse model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that is characterized by progressive motor neuron degeneration and leads to death within a few years of diagnosis. One of the pathogenic mechanisms of ALS is proposed to be a dysfunction in the protein quality‐control machinery. Dorfin has been identified as a ubiquitin ligase (E3) that recognizes and ubiquitinates mutant SOD1 proteins, thereby accelerating their degradation and reducing their cellular toxicity. We examined the effects of human Dorfin overexpression in G93A mutant SOD1 transgenic mice, a mouse model of familial ALS. In addition to causing a decrease in the amount of mutant SOD1 protein in the spinal cord, Dorfin overexpression ameliorated neurological phenotypes and motor neuron degeneration. Our results indicate that Dorfin overexpression or the activation or induction of E3 may be a therapeutic avenue for mutant SOD1‐associated ALS. © 2009 Wiley‐Liss, Inc.     

Autophagy in the central nervous system: implications for neurodegenerative disorders

The autophagy-lysosomal pathway is a major proteolytic pathway that in mammalian systems mainly comprises of macroautophagy and chaperone-mediated autophagy. The former is relatively non-selective and involves bulk degradation of proteins and organelles, whereas the latter is selective for certain cytosolic proteins. These autophagy pathways are important in development, differentiation, cellular remodeling and survival during nutrient starvation. Autophagy is crucial for neuronal homeostasis and acts as a local housekeeping process, since neurons are post-mitotic cells and require effective protein degradation to prevent accumulation of toxic aggregates. A growing body of evidence now suggests that dysfunction of autophagy causes accumulation of abnormal proteins and/or damaged organelles. Such accumulation has been linked to synaptic dysfunction, cellular stress and neuronal death. Abnormal autophagy may be involved in the pathology of both chronic nervous system disorders, such as proteinopathies (Alzheimer's, Parkinson's, Huntington's disease) and acute brain injuries. Although autophagy is generally beneficial, its aberrant activation may also exert a detrimental role in neurological diseases depending on the environment and the insult, leading to autophagic neuronal death. In this review we summarize the current knowledge regarding the role of autophagy-lysosomal pathway in the central nervous system and discuss the implication of autophagy dysregulation in human neurological diseases and animal models.     

Modulation of autophagy for neuroprotection and functional recovery in traumatic spinal cord injury

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Rapamycin Delays Disease Onset and Prevents PrP Plaque Deposition in a Mouse Model of Gerstmann-Straussler-Scheinker Disease

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