Drosophila: a "model" model system to study neurodegeneration

The fruit fly, Drosophila melanogaster, is a powerful model genetic organism that has been used since the turn of the previous century in the study of complex biological problems. In the last decade, numerous researchers have focused their attention on understanding neurodegenerative diseases by utilizing this model system. Numerous Drosophila mutants have been isolated that profoundly affect neural viability and integrity of the nervous system with age. Additionally, many transgenic strains have been developed as models of human disease conditions. We review the existing Drosophila neurodegenerative mutants and transgenic disease models, and discuss the role of the fruit fly in therapeutic development for neurodegenerative diseases.     

Illuminating the role of caspases during Drosophila oogenesis

Cell death is a prominent feature of animal germline development. In Drosophila, the death of 15 nurse cells is linked to the development of each oocyte. In addition, females respond to poor environmental conditions by inducing egg chamber death prior to yolk uptake by the oocyte. To study these two forms of cell death, we analyzed caspase activity in the germline by expressing a transgene encoding a caspase cleavage site flanked by cyan fluorescent protein and yellow fluorescent protein. When expressed in ovaries undergoing starvation-induced apoptosis, this construct was an accurate reporter of caspase activity. However, dying nurse cells at the end of normal oogenesis showed no evidence of cytoplasmic caspase activity. Furthermore, although expression of the caspase inhibitors p35 or Drosophila inhibitor of apoptosis protein 1 blocked starvation-induced death, it did not affect normal nurse cell death or overall oogenesis in well-fed females. Our data suggest that caspases play no role in developmentally programmed nurse cell death.     

Stem cells and neurodegenerative diseases

Neurodegenerative diseases are characterized by the neurodegenerative changes or apoptosis of neurons involved in networks, which are important to specific physiological functions. With the de-velopment of old-aging society, the incidence of neurodegenerative diseases is on the increase. How-ever, it is difficult to diagnose for most of neurodegenerative diseases. At present, there are too few effective therapies. Advances in stem cell biology have raised the hope and possibility for the therapy of neurodegenerative diseases. Recently, stem cells have been widely attempted to treat neurodegen-erative diseases of animal model. Here we review the progress and prospects of various stem cells, including embryonic stem cells, mesenchymal stem cell and neural stem cells and so on, for the treatments of neurodegenerative diseases, such as Parkinson’s disease, Alzheimer’s disease, Hunt-ington’s disease and Amyotrophic lateral sclerosis/Lou Gehrig’s disease.     

The accumulation of neurotoxic proteins, induced by proteasome inhibition, is reverted by trehalose, an enhancer of autophagy, in human neuroblastoma cells

Neurodegenerative diseases like Parkinson's disease, Alzheimer's disease, Huntington's disease and others are due to accumulation of abnormal proteins which fold improperly and impair neuronal function. Accumulation of these proteins could be achieved by several mechanisms including mutation, overproduction or impairment of its degradation. Inhibition of the normal protein degradation is produced by blockade of the ubiquitin proteasome system. We have shown that epoxomicin, a proteasome inhibitor, increases the levels of proteins involved in neurodegenerative disorders such as α-synuclein and hyper phosphorylated tau in NB69 human neuroblastoma cells and that such increase correlates with an enhanced rate of cell death. We then investigated whether the stimulation of autophagy, an alternative mechanism for elimination of abnormal proteins, by treatment with trehalose, counteracts the effects of proteasomal blockade. Trehalose, a disaccharide present in many non-mammalian species, known to enhance autophagy, protects cells against various environmental stresses. Treatment with trehalose produced a dose and time-dependent increase in the number of autophagosomes and markers of autophagy in NB69 cells. Trehalose did not change the number of total neither the number of dividing cells in the culture but it completely prevented the necrosis of NB69 induced by epoxomicin. In addition, the treatment with trehalose reverted the accumulation, induced by epoxomicin, of polyubiquitinated proteins, total and phosphorylated tau, p-GSK-3, and α-synuclein, as well as the α-synuclein intracellular aggregates. The effects of trehalose were not mediated through activation of free radical scavenging compounds, like GSH, or mitochondrial proteins, like DJ1, but trehalose reduced the activation of ERK and chaperone HSP-70 induced by epoxomicin. Inhibition of ERK phosphorylation prevented the epoxomicin-induced cell death. Inhibition of autophagy reverted the neuroprotective effects of trehalose in epoxomicin-induced cell death. These results suggest that trehalose is a powerful modifier of abnormal protein accumulation in neurodegenerative diseases.     

利用黑腹果蝇（Drosophila melanogaster）研究微量金属元素代谢

微量金属参与了生物体许多化学反应过程,同时也可作为蛋白质的辅基或辅因子起作用,对机体生长发育以及正常生物功能的维持具有重要作用;微量金属元素的代谢失衡与生物体许多疾病密切相关,如威尔森氏病、门克斯病、铁色素沉积、肠变性皮炎以及一些神经退行性疾病。黑腹果蝇（Drosophila melanogaster）是遗传背景清楚、生活周期短、操作方便的模式生物,利用果蝇研究金属离子代谢以及金属离子代谢与疾病的联系具有独特的优势,近年来,随着果蝇基因组测序的完成以及许多转基因果蝇株的建立,果蝇也越来越多的用于金属离子代谢的研究。介绍了近年来果蝇在金属离子代谢研究领域的进展,以及其与神经退行性疾病关系研究上的一些应用。     

Atypical antiinflammatory activation of microglia induced by apoptotic neurons

In the central nervous system (CNS), apoptosis plays an important role during development and is a primary pathogenic mechanism in several adult neurodegenerative diseases. A main feature of apoptotic cell death is the efficient and fast removal of dying cells by macrophages and nonprofessional phagocytes, without eliciting inflammation in the surrounding tissue. Apoptotic cells undergo several membrane changes, including the externalization of so-called "eat me" signals whose cognate receptors are present on professional phagocytes. Among these signals, the aminophospholipid phosphatidylserine (PS) appears to have a crucial and unique role in preventing the classical pro-inflammatory activation of macrophages, thus ensuring the silent and safe removal of apoptotic cells. Although extensively studied in the peripheral organs, the process of recognition and removal of apoptotic cells in the brain has only recently begun to be unraveled. Here, we summarize the evidence suggesting that upon interaction with PS-expressing apoptotic neurons, microglia may no longer promote the inflammatory cascade, but rather facilitate the elimination of damaged neurons through antiinflammatory and neuroprotective functions. We propose that the anti-inflammatory microglial phenotype induced through the activation of the specific PS receptor (PtdSerR), expressed by resting and activated microglial cells, could be relevant to the final outcome of neurodegenerative diseases, in which apoptosis seems to play a crucial role.     

生命科学与人类疾病研究的重要模型——果蝇

黑腹果蝇(Drosophilamelanogaster)是生物学研究中最重要的模式生物之一,它在遗传的染色体理论建立中起到非常重要的作用。由于果蝇自身独特的优势,20世纪70年代以来,它又在发育生物学、神经科学、人类疾病研究等领域得到广泛应用,作出许多新的重要贡献。果蝇在神经退行性疾病研究中是非常有用的模型。可以预期,随着研究手段的丰富及科学的发展,果蝇将作为一种理想的模式生物在生物医学中发挥更大的作用。     

Extracellular proteases and neuronal cell death.

Neuronal cell death occurs during development of the central nervous system as well as in pathological situations such as acute injury and progressive degenerative diseases. For instance, granule cells in the developing cerebellum and neuronal precursor cells in the cortex undergo programmed cell death, or apoptosis. There is currently strong debate conceming the mechanism of death in many degenerative events such as ischemia, blunt head trauma, excitotoxicity and neurodegenerative diseases, i.e. Alzheimer's disease. Neurons can die a necrotic death when the initial insult is too great; apoptosis requires "planning." For example, the cell death seen in the core of an ischemic infarct is necrotic, while in the surrounding penumbra region the death is probably apoptotic. Regardless of the degenerative pathway, damaged or dead neurons are a hallmark of many diseases including Alzheimer's, Parkinson's, glaucoma, ischemia and multiple sclerosis. Molecules such as cytokines, chemokines, reactive nitrogen/oxygen species, and proteases play an important role in promoting and/or mediating neurodegeneration. Proteases have been implicated in both physiological and pathological events, suggesting their intervention in key points when things go awry. In this review we will summarize recent findings linking extracellular proteases with neuronal cell death in both human diseases and their animal models.     

Induction of neuronal death by microglial AGE-albumin: implications for Alzheimer's disease

Advanced glycation end products (AGEs) have long been considered as potent molecules promoting neuronal cell death and contributing to neurodegenerative disorders such as Alzheimer's disease (AD). In this study, we demonstrate that AGE-albumin, the most abundant AGE product in human AD brains, is synthesized in activated microglial cells and secreted into the extracellular space. The rate of AGE-albumin synthesis in human microglial cells is markedly increased by amyloid-β exposure and oxidative stress. Exogenous AGE-albumin upregulates the receptor protein for AGE (RAGE) and augments calcium influx, leading to apoptosis of human primary neurons. In animal experiments, soluble RAGE (sRAGE), pyridoxamine or ALT-711 prevented Aβ-induced neuronal death in rat brains. Collectively, these results provide evidence for a new mechanism by which microglial cells promote death of neuronal cells through synthesis and secretion of AGE-albumin, thereby likely contributing to neurodegenerative diseases such as AD.     

Understanding the importance of autophagy in human diseases using Drosophila

Autophagy is a lysosome-dependent intracellular degradation pathway that has been implicated in the pathogenesis of various human diseases, either positively or negatively impacting disease outcomes depending on the specific context. The majority of medical conditions including cancer, neurodegenerative diseases, infections and immune system disorders and inflammatory bowel disease could probably benefit from therapeutic modulation of the autophagy machinery. Drosophila represents an excellent model animal to study disease mechanisms thanks to its sophisticated genetic toolkit, and the conservation of human disease genes and autophagic processes. Here, we provide an overview of the various autophagy pathways observed both in flies and human cells(macroautophagy, microautophagy and chaperone-mediated autophagy), and discuss Drosophila models of the above-mentioned diseases where fly research has already helped to understand how defects in autophagy genes and pathways contribute to the relevant pathomechanisms.     

Drosophila models of human neurodegenerative disease

Drosophila has provided a powerful genetic system in which to elucidate fundamental cellular pathways in the context of a developing and functioning nervous system. Recently, Drosophila has been applied toward elucidating mechanisms of human neurodegenerative disease, including Alzheimer's, Parkinson's and Huntington's diseases. Drosophila allows study of the normal function of disease proteins, as well as study of effects of familial mutations upon targeted expression of human mutant forms in the fly. These studies have revealed new insight into the normal functions of such disease proteins, as well as provided models in Drosophila that will allow genetic approaches to be applied toward elucidating ways to prevent or delay toxic effects of such disease proteins. These, and studies to come that follow from the recently completed sequence of the Drosophila genome, underscore the contributions that Drosophila as a model genetic system stands to contribute toward the understanding of human neurodegenerative disease.     

Tackling neurodegenerative diseases: animal models of Alzheimer’s disease and Parkinson’s disease

Our ageing society is confronted with a dramatic increase in incidence of age-related neurodegenerative diseases; biomedical research leading to novel therapeutic strategies is crucial to address this problem. Animal models of neurodegenerative conditions are invaluable in improving our understanding of the molecular basis of pathology, potentially revealing novel targets for intervention. Here, we review transgenic animal models of Alzheimer’s and Parkinson’s disease reported in mice, zebrafish, Caenorhabditis elegans and Drosophila melanogaster. This information will enable researchers to compare different animal models targeting disease-associated molecules by genomic engineering and to facilitate the development of novel animal models for any particular study, depending on the ultimate research goals.     

Spatio-temporal pattern of programmed cell death in the developing Drosophila optic lobe

A large number of cells die via programmed cell death during the normal development of the Drosophila optic lobe. In this study, we report the precise spatial and temporal pattern of cell death in this organ. Cell death in the developing optic lobe occurs in two distinct phases. The first phase extends from the start of metamorphosis to the mid-pupal stage. During this phase, a large number of cells die in the optic lobe as a whole, with a peak of cell death at an early pupal stage in the lamina and medulla cortices and the region of the T2/T3/C neurons, and a smaller number of dead cells observed in the lobula plate cortex. The second phase extends from the mid-pupal stage to eclosion. Throughout this period, a small number of dying cells can be observed, with a small peak at a late pupal stage. Most of the dying cells are neurons. During the first phase, dying cells are distributed in specific patterns in cortices. The lamina cortex contains two distinct clusters of dying cells; the medulla cortex, four clusters; the lobula plate cortex, one cluster; and the region of the T2/T3/C neurons, one cluster. Many of the clusters maintain their distinct positions in the optic lobe but others extend the region they cover during development. The presence of distinct clusters of dying cells at different phases suggests that distinct mechanisms control cell death during different stages of optic lobe development in Drosophila.     

CD8+ T cells are increased in the subventricular zone with physiological and pathological aging

Age‐related cognitive decline and neurodegenerative diseases are associated with less functional neurogenic niches. It has been recently shown that aged subventricular zone (SVZ) suffers an infiltration of T cells, which affects neural stem cell activity in mice. Whether this occurs in human neurogenic niches or to which extent T‐cell infiltration is also taking place in neurodegenerative diseases remains unknown. In this work, we studied the presence of T cells in both human neurogenic niches in young and old individuals. There was a significant increase in the number of CD3⁺ and CD8⁺ T cells in the SVZ of elderly individuals, which was not detected in the dentate gyrus. Moreover, we also found CD3⁺ and CD8⁺ T cells in the SVZ of individuals with neurodegenerative diseases. However, T‐cell count was similar when compared non‐neuropathological elderly with disease diagnosed patients. Our study reveals the infiltration of T cells in old human brains, particularly in the SVZ under non‐pathological conditions and also in neurodegenerative contexts.     

Flexibility exists in the region of [A6-A11, A7-B7] disulfide bonds during insulin precursor folding

Programmed cell death or apoptosis is the regulatory mechanism for removing unneeded cells during animal development and in tissue homeostasis. Perturbation of the cell death mechanisms leads to various disorders, including neurodegenerative diseases, immunodeficiency diseases, and tumors. c-Jun N-terminal kinase (JNK) has crucial roles in the regulation of cell death in response to many stimuli. Since JNK is highly conserved from yeast to mammals, genetic studies using model animals are helpful in understanding the principal cell death mechanisms regulated by JNK. For example, loss-of-function studies using the targeted disruption of murine genes have established the genetic framework of the mechanisms of the cell death induced by UV radiation. Also, in Drosophila, many cell death-related genes have been identified by genetics. Genetic studies of JNK-dependent cell death mechanisms should shed light on the regulation of both physiological and pathological cell death.     

Tau phosphorylation: physiological and pathological consequences

The microtubule-associated protein tau, abundant in neurons, has gained notoriety due to the fact that it is deposited in cells as fibrillar lesions in numerous neurodegenerative diseases, and most notably Alzheimer's disease. Regulation of microtubule dynamics is the most well-recognized function of tau, but it is becoming increasingly evident that tau plays additional roles in the cell. The functions of tau are regulated by site-specific phosphorylation events, which if dysregulated, as they are in the disease state, result in tau dysfunction and mislocalization, which is potentially followed by tau polymerization, neuronal dysfunction and death. Given the increasing evidence that a disruption in the normal phosphorylation state of tau plays a key role in the pathogenic events that occur in Alzheimer's disease and other neurodegenerative conditions, it is of crucial importance that the protein kinases and phosphatases that regulate tau phosphorylation in vivo as well as the signaling cascades that regulate them be identified. This review focuses on recent literature pertaining to the regulation of tau phosphorylation and function in cell culture and animal model systems, and the role that a dysregulation of tau phosphorylation may play in the neuronal dysfunction and death that occur in neurodegenerative diseases that have tau pathology.     

Modeling human mitochondrial diseases in flies

Human mitochondrial diseases are associated with a wide range of clinical symptoms, and those that result from mutations in mitochondrial DNA affect at least 1 in 8500 individuals. The development of animal models that reproduce the variety of symptoms associated with this group of complex human disorders is a major focus of current research. Drosophila represents an attractive model, in large part because of its short life cycle, the availability of a number of powerful techniques to alter gene structure and regulation, and the presence of orthologs of many human disease genes. We describe here Drosophila models of mitochondrial DNA depletion, deafness, encephalopathy, Freidreich's ataxia, and diseases due to mitochondrial DNA mutations. We also describe several genetic approaches for gene manipulation in flies, including the recently developed method of targeted mutagenesis by recombinational knock-in.     

Connecting cell-cycle activation to neurodegeneration in Drosophila

Studies in cell-culture systems and in postmortem tissue from human disease have suggested a connection between cell-cycle activation and neurodegeneration. The fruit fly Drosophila melanogaster has recently emerged as a powerful model system in which to model neurodegenerative diseases. Here we review work in the fly that has begun to address some of the important questions regarding the relationship between cell-cycle activation and neurodegeneration in vivo, including recent data implicating cell-cycle activation as a downstream effector of tau-induced neurodegeneration. We suggest how powerful research tools in Drosophila might be utilized to approach fundamental questions that remain.     

Modelling neurodegenerative diseases in Drosophila: a fruitful approach?

Human neurodegenerative diseases are characterized by the progressive loss of specific neuronal populations, resulting in substantial disability and early death. The identification of causative single-gene mutations in families with inherited neurodegenerative disorders has facilitated the modelling of these diseases in experimental organisms, including the fruitfly Drosophila melanogaster. Many neurodegenerative diseases have now been successfully modelled in Drosophila, and genetic analysis is under way in each of these models. Using fruitfly genetics to define the molecular pathways that underlie the neurodegenerative process is likely to improve substantially our understanding of the pathogenesis of the human diseases, and to provide new therapeutic targets.