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Investigative Ophthalmology & Visual... Apr 2024To undertake the first ultrastructural characterization of human retinal pigment epithelial (RPE) differentiation from fetal development to adolescence.
PURPOSE
To undertake the first ultrastructural characterization of human retinal pigment epithelial (RPE) differentiation from fetal development to adolescence.
METHODS
Ten fetal eyes and three eyes aged six, nine, and 17 years were examined in the temporal retina adjacent to the optic nerve head by transmission electron microscopy. The area, number, and distribution of RPE organelles were quantified and interpreted within the context of adjacent photoreceptors, Bruch's membrane, and choriocapillaris maturation.
RESULTS
Between eight to 12 weeks' gestation (WG), pseudostratified columnar epithelia with apical tight junctions differentiate to a simple cuboidal epithelium with random distribution of melanosomes and mitochondria. Between 12 to 26 WG, cells enlarge and show long apical microvilli and apicolateral junctional complexes. Coinciding with eye opening at 26 WG, melanosomes migrate apically whereas mitochondria distribute to perinuclear regions, with the first appearance of phagosomes, complex granules, and basolateral extracellular space (BES) formation. Significantly, autophagy and heterophagy, as evidenced by organelle recycling, and the gold standard of ultrastructural evidence for autophagy of double-membrane autophagosomes and mitophagosomes were evident from 32 WG, followed by basal infoldings of RPE cell membrane at 36 WG. Lipofuscin formation and deposition into the BES evident at six years increased at 17 years.
CONCLUSIONS
We provide compelling ultrastructural evidence that heterophagy and autophagy begins in the third trimester of human fetal development and that deposition of cellular byproducts into the extracellular space of RPE takes place via exocytosis. Transplanted RPE cells must also demonstrate the capacity to subserve autophagic and heterophagic functions for effective disease mitigation.
Topics: Humans; Retinal Pigment Epithelium; Adolescent; Autophagy; Microscopy, Electron, Transmission; Child; Lipofuscin; Exocytosis; Extracellular Space; Gestational Age; Female; Male; Fetal Development; Mitochondria; Cell Differentiation
PubMed: 38648041
DOI: 10.1167/iovs.65.4.32 -
Cell Communication and Signaling : CCS Apr 2024p66Shc, as a redox enzyme, regulates reactive oxygen species (ROS) production in mitochondria and autophagy. However, the mechanisms by which p66Shc affects...
BACKGROUND
p66Shc, as a redox enzyme, regulates reactive oxygen species (ROS) production in mitochondria and autophagy. However, the mechanisms by which p66Shc affects autophagosome formation are not fully understood.
METHODS
p66Shc expression and its location in the trophoblast cells were detected in vivo and in vitro. Small hairpin RNAs or CRISPR/Cas9, RNA sequencing, and confocal laser scanning microscope were used to clarify p66Shc's role in regulating autophagic flux and STING activation. In addition, p66Shc affects mitochondrial-associated endoplasmic reticulum membranes (MAMs) formation were observed by transmission electron microscopy (TEM). Mitochondrial function was evaluated by detected cytoplastic mitochondrial DNA (mtDNA) and mitochondrial membrane potential (MMP).
RESULTS
High glucose induces the expression and mitochondrial translocation of p66Shc, which promotes MAMs formation and stimulates PINK1-PRKN-mediated mitophagy. Moreover, mitochondrial localized p66Shc reduces MMP and triggers cytosolic mtDNA release, thus activates cGAS/STING signaling and ultimately leads to enhanced autophagy and cellular senescence. Specially, we found p66Shc is required for the interaction between STING and LC3II, as well as between STING and ATG5, thereby regulates cGAS/STING-mediated autophagy. We also identified hundreds of genes associated several biological processes including aging are co-regulated by p66Shc and ATG5, deletion either of which results in diminished cellular senescence.
CONCLUSION
p66Shc is not only implicated in the initiation of autophagy by promoting MAMs formation, but also helps stabilizing active autophagic flux by activating cGAS/STING pathway in trophoblast.
Topics: Src Homology 2 Domain-Containing, Transforming Protein 1; Autophagosomes; Extravillous Trophoblasts; Autophagy; DNA, Mitochondrial; Trophoblasts; Glucose; Nucleotidyltransferases
PubMed: 38643181
DOI: 10.1186/s12964-024-01621-x -
Journal of Molecular Biology Apr 2024Proteins are known to perform an astonishing array of functions thanks to their ability to cooperate and modulate each other's properties. Inside cells, proteins can... (Review)
Review
Proteins are known to perform an astonishing array of functions thanks to their ability to cooperate and modulate each other's properties. Inside cells, proteins can assemble into large multi-subunit complexes to carry out complex cellular functions. The correct assembly and maintenance of the functional state of macromolecular protein complexes is crucial for human health. Failure to do so leads to loss of function and potential accumulation of harmful materials, which is associated with a variety of human diseases such as neurodegeneration and cancer. Autophagy engulfs cytosolic material in autophagosomes, and therefore is best suited to eliminate intact macromolecular complexes without disassembling them, which could interfere with de novo assembly. In this review, we discuss the role of autophagy in the selective degradation of macromolecular complexes. We highlight the current state of knowledge for different macromolecular complexes and their selective autophagic degradation. We emphasize the gaps in our understanding of what it takes for these large macromolecular complexes to be degraded and point to future work that may shed light on the regulation of the selective degradation of macromolecular complexes by autophagy.
PubMed: 38636617
DOI: 10.1016/j.jmb.2024.168574 -
Heliyon Apr 2024Alzheimer's disease is the most common form of dementia and is characterized by cognitive impairment. The disruption of autophagosome-lysosome function has been linked...
Alzheimer's disease is the most common form of dementia and is characterized by cognitive impairment. The disruption of autophagosome-lysosome function has been linked to the pathogenesis of Alzheimer's disease. Tris (1,3-dichloro-2-propyl) phosphate (TDCIPP) is a widely used organophosphorus flame retardant that has the potential to cause neuronal damage. We found that TDCIPP significantly increased the expression of β-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1), presenilin-1 (PS1) and Aβ42. Proteomic studies with TMT labeling revealed changes in the profiles of N2a-APPswe cells after exposure to TDCIPP. Proteomic and bioinformatics analyses revealed that lysosomal proteins were dysregulated in N2a-APPswe cells after treatment with TDCIPP. The LC3, P62, CTSD, and LAMP1 levels were increased after TDCIPP exposure, and dysregulated protein expression was validated by Western blotting. The exposure to TDCIPP led to the accumulation of autophagosomes, and this phenomenon was enhanced in the presence of chloroquine (CQ). Our results revealed for the first time that TDCIPP could be a potential environmental risk factor for AD development. The inhibition of autophagosome-lysosome fusion may have a significant impact on the generation of Aβ1-42 in response to TDCIPP.
PubMed: 38628727
DOI: 10.1016/j.heliyon.2024.e26832 -
Heliyon Apr 2024Leptin resistance represents a primary pathological manifestation in obesity. Investigating potential treatments and associated mechanisms to restore leptin sensitivity...
PURPOSE
Leptin resistance represents a primary pathological manifestation in obesity. Investigating potential treatments and associated mechanisms to restore leptin sensitivity is crucial for effective obesity management. This study aimed to explore the therapeutic potential of acupoints catgut embedding (ACE) in addressing obesity and its associated leptin resistance.
METHODS
A simple obesity model was established by subjecting C57 male mice to a high-fat diet (HFD) for 12 weeks, followed by ACE treatment administered to half of the obese mice for a duration of 4 weeks. The levels of leptin and its receptor-lepRb, were assessed using enzyme-linked immunosorbent assay (ELISA) and Western blot analysis, respectively. Autophagy progression markers were evaluated through quantitative polymerase chain reaction (qPCR) and Western blot analysis. Also, the liver autophagosomes were photographed using electron microscopy. The role of autophagy in regulating leptin resistance was elucidated using an autophagy suppression model.
RESULTS
Comparative analyses demonstrated that ACE treatment resulted in a significant reduction in body weight and blood lipid levels compared to the HFD group. Furthermore, serum leptin levels decreased, while liver lepRb expression increased following ACE treatment. The mRNA and protein expression levels of autophagy in liver were adjusted by ACE treatment. Interestingly, the beneficial effects of ACE were attenuated upon the administration of an autophagy inhibitor. Additionally, ACE treatment led to the activation of the AMPK-mTOR signaling pathway, a crucial regulator of autophagy.
CONCLUSION
These findings suggest that ACE therapy holds promise for recovering leptin resistance by enhancing autophagy progression, mediated via the AMPK-mTOR signaling pathway in liver.
PubMed: 38623207
DOI: 10.1016/j.heliyon.2024.e29094 -
Cell Discovery Apr 2024Macroautophagy is a process that cells engulf cytosolic materials by autophagosomes and deliver them to lysosomes for degradation. The biogenesis of autophagosomes...
Macroautophagy is a process that cells engulf cytosolic materials by autophagosomes and deliver them to lysosomes for degradation. The biogenesis of autophagosomes requires ATG2 as a lipid transfer protein to transport lipids from existing membranes to phagophores. It is generally believed that endoplasmic reticulum is the main source for lipid supply of the forming autophagosomes; whether ATG2 can transfer lipids from other organelles to phagophores remains elusive. In this study, we identified a new ATG2A-binding protein, ANKFY1. Depletion of this endosome-localized protein led to the impaired autophagosome growth and the reduced autophagy flux, which largely phenocopied ATG2A/B depletion. A pool of ANKFY1 co-localized with ATG2A between endosomes and phagophores and depletion of UVRAG, ANKFY1 or ATG2A/B led to reduction of PI3P distribution on phagophores. Purified recombinant ANKFY1 bound to PI3P on membrane through its FYVE domain and enhanced ATG2A-mediated lipid transfer between PI3P-containing liposomes. Therefore, we propose that ANKFY1 recruits ATG2A to PI3P-enriched endosomes and promotes ATG2A-mediated lipid transfer from endosomes to phagophores. This finding implicates a new lipid source for ATG2A-mediated phagophore expansion, where endosomes donate PI3P and other lipids to phagophores via lipid transfer.
PubMed: 38622126
DOI: 10.1038/s41421-024-00659-y -
Journal of Dental Sciences Apr 2024Autophagy is an intracellular degradative process occurring under stressful conditions. Nitric oxide (NO), a free radical, regulates autophagy and apoptosis in several...
BACKGROUND/PURPOSE
Autophagy is an intracellular degradative process occurring under stressful conditions. Nitric oxide (NO), a free radical, regulates autophagy and apoptosis in several cancers. However, the effect of head and neck squamous cell carcinoma (HNSCC) cell adaptation to high nitric oxide (HNO) on autophagy remains unknown. We investigated the autophagy and apoptotic changes in the HNO-adapted HNSCC cell lines.
MATERIALS AND METHODS
Isogenic primary HNSCC (HN18/HN30) and metastatic (HN17/HN31) cell lines were evaluated. The cells were induced with 1, 2, 3 and 4 mM DEA-NONOate, an NO donor, for 72 h and assessed for cell viability by MTT assay. "HNO-adapted cells" were defined when the cell viability in the treatment group was <10%. The surviving cells were re-treated with HNO to confirm their adaptation. HNO-adapted cells were quantified for apoptosis using flow cytometry. Autophagic structures (autophagosomes) and proteins (LC3A/B and LC3B-II) were investigated using transmission electron and confocal microscopy, respectively.
RESULTS
HNO-adapted concentration for HN18, HN17, HN30 and HN31 cells was 3, 2, 4 and 4 mM, respectively. The HNO-adapted HN18 cells demonstrated a significantly increased apoptotic percentage, whereas no significant apoptotic change was detected in the HNO-adapted HN17, HN30 and HN31 cells compared with the parent cells. Autophagosomes were widely observed across the HNO-adapted cells. Moreover, LC3A/B and LC3B-II proteins were increased in all HNO-adapted cells.
CONCLUSION
Our results demonstrate that apoptosis and/or autophagy are increased during HNO adaptation in HNSCC cell lines.
PubMed: 38618052
DOI: 10.1016/j.jds.2023.08.023 -
World Journal of Gastroenterology Mar 2024Autophagy is a cellular catabolic process characterized by the formation of double-membrane autophagosomes. Transmission electron microscopy is the most rigorous method... (Review)
Review
Autophagy is a cellular catabolic process characterized by the formation of double-membrane autophagosomes. Transmission electron microscopy is the most rigorous method to clearly visualize autophagic engulfment and degradation. A large number of studies have shown that autophagy is closely related to the digestion, secretion, and regeneration of gastrointestinal (GI) cells. However, the role of autophagy in GI diseases remains controversial. This article focuses on the morphological and biochemical characteristics of autophagy in GI diseases, in order to provide new ideas for their diagnosis and treatment.
Topics: Humans; Gastrointestinal Diseases; Autophagy; Microscopy, Electron, Transmission
PubMed: 38617452
DOI: 10.3748/wjg.v30.i11.1524 -
International Journal of Molecular... Apr 2024The Golgi apparatus, long recognized for its roles in protein processing and vesicular trafficking, has recently been identified as a crucial contributor to innate... (Review)
Review
The Golgi apparatus, long recognized for its roles in protein processing and vesicular trafficking, has recently been identified as a crucial contributor to innate immune signaling pathways. This review discusses our expanding understanding of the Golgi apparatus's involvement in initiating and activating these pathways. It highlights the significance of membrane connections between the Golgi and other organelles, such as the endoplasmic reticulum, mitochondria, endosomes, and autophagosomes. These connections are vital for the efficient transmission of innate immune signals and the activation of effector responses. Furthermore, the article delves into the Golgi apparatus's roles in key immune pathways, including the inflammasome-mediated activation of caspase-1, the -STING pathway, and TLR/RLR signaling. Overall, this review aims to provide insights into the multifunctional nature of the Golgi apparatus and its impact on innate immunity.
Topics: Golgi Apparatus; Immunity, Innate; Inflammasomes; Autophagosomes; Caspase 1
PubMed: 38612929
DOI: 10.3390/ijms25074120 -
International Journal of Molecular... Apr 2024In the last two decades, alpha-synuclein (alpha-syn) assumed a prominent role as a major component and seeding structure of Lewy bodies (LBs). This concept is driving... (Review)
Review
In the last two decades, alpha-synuclein (alpha-syn) assumed a prominent role as a major component and seeding structure of Lewy bodies (LBs). This concept is driving ongoing research on the pathophysiology of Parkinson's disease (PD). In line with this, alpha-syn is considered to be the guilty protein in the disease process, and it may be targeted through precision medicine to modify disease progression. Therefore, designing specific tools to block the aggregation and spreading of alpha-syn represents a major effort in the development of disease-modifying therapies in PD. The present article analyzes concrete evidence about the significance of alpha-syn within LBs. In this effort, some dogmas are challenged. This concerns the question of whether alpha-syn is more abundant compared with other proteins within LBs. Again, the occurrence of alpha-syn compared with non-protein constituents is scrutinized. Finally, the prominent role of alpha-syn in seeding LBs as the guilty structure causing PD is questioned. These revisited concepts may be helpful in the process of validating which proteins, organelles, and pathways are likely to be involved in the damage to meso-striatal dopamine neurons and other brain regions involved in PD.
Topics: Humans; alpha-Synuclein; Lewy Bodies; Parkinson Disease; Corpus Striatum; Disease Progression
PubMed: 38612739
DOI: 10.3390/ijms25073929