Lysosome-associated membrane glycoprotein 1 precursor (LAMP-1) (Lysosome-associated membrane protein 1) (CD107 antigen-like family member A) (CD107a antigen)
Dietary restriction (DR) is the strategy ameliorating the morbidity of various pathologies, including age-associated diseases. Acute kidney injury (AKI) remains a problem for the elderly with DR being a promising approach for diminishing its consequences. We evaluated the possible nephroprotective potential of short-term DR in young and old rats. DR in young rats resulted in pronounced beneficial effects normalizing lipid metabolism (triglycerides concentration, adiponectin level) activating autophagic-lysosomal system evaluated by LC3II/LC3I ratio, LAMP1, p62/SQSTM1 levels, and LysoTracker Green staining. DR had a remarkable recovering effect on mitochondrial structure and functions including regaining of mitochondrial membrane potential, the elevation of SIRT-3, PGC-1α, levels and partial restoration of ultrastructure. The beneficial effects of DR resulted in the mitigation of oxidative stress including a decrease in levels of protein carbonylation and lipid peroxidation. Aging led to decreased activity of autophagy, elevated oxidative stress and impaired kidney regenerative capacity. Eventually, in old rats, even 8-week DR was not able to ameliorate AKI, but it caused some rejuvenating effects including elevation of mitochondrial membrane potential and Bcl-X levels, as well as lowered severity of the oxidative stress. Thus, the age-associated decline of protective signaling demands extended DR to achieve nephroprotective potential in old animals.
- caloric restriction
- kidney injury
Impaired cargo trafficking and the aggregation of intracellular macromolecules are key features of neurodegeneration, and a hallmark of aged as well as diseased retinal pigment epithelial (RPE) cells in the eye. Here, photoreceptor outer segments (POS), which are internalized daily by RPE cells, were modified by UV-irradiation to create oxidatively modified POS (OxPOS). Oxidative modification was quantified by a protein carbonyl content assay. Human ARPE-19 cells were synchronously pulsed with POS or OxPOS to study whether oxidatively modified cargos can recapitulate features of RPE pathology associated with blinding diseases. Confocal immunofluorescence microscopy analysis showed that OxPOS was trafficked to LAMP1, LAMP2 lysosomes and to LC3b autophagy vacuoles. Whilst POS were eventually degraded, OxPOS cargos were sequestered in late compartments. Co-localization of OxPOS was also associated with swollen autolysosomes. Ultrastructural analysis revealed the presence of electron-dense OxPOS aggregates in RPE cells, which appeared to be largely resistant to degradation. Measurement of cellular autofluorescence, using parameters used to assess fundus autofluorescence (FAF) in age-related macular disease (AMD) patients, revealed that OxPOS contributed significantly to a key feature of aged and diseased RPE. This in vitro cell model therefore represents a versatile tool to study disease pathways linked with RPE damage and sight-loss.
- oxidized POS
To date, microglia subsets in the healthy CNS have not been identified. Utilizing autofluorescence (AF) as a discriminating parameter, we identified two novel microglia subsets in both mice and non-human primates, termed autofluorescence-positive (AF ) and negative (AF ). While their proportion remained constant throughout most adult life, the AF signal linearly and specifically increased in AF microglia with age and correlated with a commensurate increase in size and complexity of lysosomal storage bodies, as detected by transmission electron microscopy and LAMP1 levels. Post-depletion repopulation kinetics revealed AF cells as likely precursors of AF microglia. At the molecular level, the proteome of AF microglia showed overrepresentation of endolysosomal, autophagic, catabolic, and mTOR-related proteins. Mimicking the effect of advanced aging, genetic disruption of lysosomal function accelerated the accumulation of storage bodies in AF cells and led to impaired microglia physiology and cell death, suggestive of a mechanistic convergence between aging and lysosomal storage disorders.
- lysosomal storage disorder
- rhesus macaque
Studies have found that autophagy could promote the clearance of Aβ. To promote and maintain the occurrence of autophagy in Alzheimer's Disease (AD) might be a potential way to reduce neuronal loss and improve the learning and memory of AD. To investigate the possible mechanisms of Yishen Huazhuo Decoction (YHD) against AD model. Forty 7-month-old male SAMP8 mice were randomly divided into model (P8) group and YHD group, 20 in each group, with 20 SAMR1 mice as control (R1) group. All mice were intragastrically administered for 4 weeks, YHD at the dosage of 6.24g/kg for YHD group, and distilled water for P8 group and R1 group. Morris Water Maze (MWM) test, Nissl's staining, TEM, TUNEL staining, immunofluorescence double staining, and western blot analysis were applied to learning and memory, structure and ultrastructure of neurons, autophagosome, apoptosis index, Aβ, LAMP1, and autophagy related proteins. The escape latency time of YHD group was significantly shorter on the 4th and 5th day during MWM test than those in P8 group (P=0.011, 0.008<0.05), and the number of crossing platform in YHD group increased significantly (P=0.02<0.05). Nissl's staining showed that the number of neurons in YHD group increased significantly (P<0.0001). TEM showed in YHD group that the nucleus of neurons was slightly irregular, with slightly reduced organelles, partially fused and blurred cristae and membrane of mitochondria. The apoptosis index of YHD group showed a decreasing trend, without statistically significant difference (P=0.093>0.05), while Caspase3 expression in YHD group was significantly lower (P=0.044<0.05). YHD could promote the clearance of Aβ1-42 protein, improve the expression of Beclin-1 and p-Bcl2 proteins, reduce mTOR and p62 proteins. YHD could induce autophagy initiation, increase the formation of autophagosomes and autolysosome, promote the degradation of autophagy substrates, thereby regulating autophagy, and promoting the clearance of Aβ1-42 to improve memory impairment in SAMP8 mice.
- Alzheimer`s disease
- Nissl’s staining
- Yishen Huazhuo Decoction
- senescence accelerated mouse prone 8
The senescence-accelerated mouse (SAM) prone 8 (SAMP8) has been demonstrated for muscular aging research including sarcopenia, but its underlying mechanisms remain scarce. Physiological indices and histology of skeletal muscle were analyzed in SAMP8 mice at different ages. SAMP8 mice exhibited typical features of sarcopenia at 40 weeks of age and were more time-efficient than that at 88 weeks of age in bothSAM resistant 1 (SAMR1) and C57BL/6 mice. Increase in FoxO3a-mediated transcription of Atrogin-1 and MuRF1 and decrease in phosphorylated mTOR/P70 were observed at week 40 in SAMP8 mice. High oxidative stress was observed from week 24 and persisted to week 40 in SAMP8 mice evidenced by overexpression of protein carbonyl groups and reduced activities of CAT, SOD, and GPx. Downregulation of genes involved in mitochondrial biogenesis (PGC-1α, Nrf-1, Tfam, Ndufs8, and Cox5b) and in mitochondrial dynamics fission (Mfn2 and Opa1) from week 24 indicated dysregulation of mitochondrial quality control in SAMP8 mice. Impaired autophagic flux was observed in SAMP8 mice evidenced by elevated Atg13 and LC3-II accompanied with the accumulation of P62 and LAMP1. Increases in inflammatory factors (IL-6 and MCP-1), adipokines (leptin and resistin), and myostatin in serum at week 32 and decline in Pax7 satellite cell resided next to muscle fibers at week 24 implied that muscle microenvironment contributed to the progression of sarcopenia in SAMP8 mice. Our data suggest that early alterations of mitochondrial quality control and autophagic flux worsen muscle microenvironment prior to the onset of sarcopenia.
- Autophagic flux
- Mitochondria quality/dynamics
- Senescence accelerated mouse prone 8
Therapy-induced senescence in cancer cells is an irreversible antiproliferative state, which inhibits tumor growth and is therefore a potent anti-neoplastic mechanism. In this study, low doses of Abrus agglutinin (AGG)-induced senescence through autophagy in prostate carcinoma cells (PC3) and inhibited proliferation. The inhibition of autophagy with 3-methyl adenine reversed AGG-induced senescence, thus confirming that AGG-triggered senescence required autophagy. AGG treatment also led to lipophagy-mediated accumulation of free fatty acids (FFAs), with a concomitant decrease in the number of lipid droplets. Lalistat, a lysosomal acid lipase inhibitor, abrogated AGG-induced lipophagy and senescence in PC3 cells, indicating that lipophagy is essential for AGG-induced senescence. The accumulation of FFAs increased reactive oxygen species generation, a known facilitator of senescence, which was also reduced in the presence of lalistat. Furthermore, AGG upregulated silent mating type information regulator 2 homolog 1 (SIRT1), while the presence of sirtinol reduced autophagy flux and the senescent phenotype in the AGG-treated cells. Mechanistically, AGG-induced cytoplasmic SIRT1 deacetylated a Lys residue on the cytoplasmic domain of lysosome-associated membrane protein 1 (LAMP1), an autolysosomal protein, resulting in lipophagy and senescence. Taken together, our findings demonstrate a novel SIRT1/LAMP1/lipophagy axis mediating AGG-induced senescence in prostate cancer cells.
- Abrus agglutinin
- free fatty acid
- reactive oxygen species
Bone marrow-derived mesenchymal stem cells (BMSCs) are gradually getting attention because of its multi-directional differentiation potential, hematopoietic support, and promotion of stem cell implantation. However, cultured BMSCs in vitro possess a very limited proliferation potential, and the presence of stem cell aging has substantially restricted the effect together with the efficiency in clinical treatment. Recently, increasing attention has been paid to the connection between cellular aging and lysosomal acidification as new reports indicated that vacuolar H -ATPase (v-ATPase) activity was altered and lysosomal pH was dysregulated in the process of cellular aging. Therefore, promoting lysosomal acidification might contribute to inhibition of cell senescence. Our previous studies showed that a novel small molecule, 3-butyl-1-chloro imidazo [1, 5-a] pyridine-7-carboxylic acid (SGJ), could selectively and sensitively respond to acidic pH with fast response (within 3 min), but whether SGJ can promote lysosomal acidification and inhibit senescence in BMSCs is unknown. Rat BMSCs were cultured based on our system that had been already documented. BMSCs were treated with SGJ and/or Bafilomycin-A1 (Baf-A1). The co-localization between SGJ and lysosomes was assessed by a confocal microscope. Acridine orange (AO) staining and the Lysosensor™ Green DND-189 reagents were used for indicating changes in lysosomal concentration of H . Changes of senescence were detected by immunoblotting of p21 and senescence-associated beta-galactosidase (SA-β-gal) staining as well as immunofluorescence assay of senescence-associated heterochromatin foci (SAHF). Changes of autophagy were detected by immunoblotting of MAP1LC3 (LC3B) and SQSTM1 (p62). Cell proliferation was determined by flow cytometry. Cell viability was calculated by sulforhodamine B assay (SRB). The V0 proton channel of v-ATPase was knocked down by transfecting with its small interfering RNA (si-ATP6V0C). Our work showed that SGJ can promote lysosomal acidification and inhibit senescence in BMSCs. Firstly, SGJ and lysosomes were well co-located in senescent BMSCs with the co-localization coefficient of 0.94. Secondly, SGJ increased the concentration of H and the protein expression of lysosome-associated membrane protein 1 (LAMP1) and lysosome-associated membrane protein 2 (LAMP2). Thirdly, SGJ suppressed the expression of p21 in the senescent BMSCs and reduced SA-β-gal positive cells. Fourthly, SGJ promoted senescent BMSCs' proliferation and protein level of LC3B but reduced the p62/SQSTM1 protein level. Furthermore, experimental group pretreated with 20 μM SGJ showed a stronger red fluorescent intensity, thinner cell morphology, less SA-β-gal positive cell, and less p21 protein level as well as higher cell viability in the presence of Baf-A1. Notably, ATP6V0C knockdown decreased the activity of v-ATPase and SGJ increased the concentration of H . Our work showed that SGJ could inhibit senescence in BMSCs and protect lysosomes by promoting expression of LAMP1 and LAMP2. Meanwhile, SGJ could promote autophagy. Furthermore, our study also suggested that SGJ was a new Baf-A1 antagonist because SGJ could target and occupy the V0 proton channel of v-ATPase.
- Cell Proliferation
- Cellular Senescence
- Cyclin-Dependent Kinase Inhibitor p21
- Hydrogen-Ion Concentration
- Lysosomal-Associated Membrane Protein 1
- Lysosomal-Associated Membrane Protein 2
- Mesenchymal Stem Cells
- Models, Biological
- Molecular Probes
- Rats, Wistar
- Vacuolar Proton-Translocating ATPases
- Bafilomycin-A1 antagonist
- Bone marrow-derived mesenchymal stem cells
To determine the impact of the loss of syndecan 1 (SDC1) on intraepithelial corneal nerves (ICNs) during homeostasis, aging, and in response to 1.5-mm trephine and debridement injury. Whole-mount corneas are used to quantify ICN density and thickness over time after birth and in response to injury in SDC1-null and wild-type (WT) mice. High-resolution three-dimensional imaging is used to visualize intraepithelial nerve terminals (INTs), axon fragments, and lysosomes in corneal epithelial cells using antibodies against growth associated protein 43 (GAP43), βIII tubulin, and LAMP1. Quantitative PCR was performed to quantify expression of SDC1, SDC2, SDC3, and SDC4 in corneal epithelial mRNA. Phagocytosis was assessed by quantifying internalization of fluorescently labeled 1-μm latex beads. Intraepithelial corneal nerves innervate the corneas of SDC1-null mice more slowly. At 8 weeks, ICN density is less but thickness is greater. Apically projecting intraepithelial nerve terminals and lysosome-associated membrane glycoprotein 1 (LAMP1) are also reduced in unwounded SDC1-null corneas. Quantitative PCR and immunofluorescence studies show that SDC3 expression and localization are increased in SDC1-null ICNs. Wild-type and SDC1-null corneas lose ICN density and thickness as they age. Recovery of axon density and thickness after trephine but not debridement wounds is slower in SDC1-null corneas compared with WT. Experiments assessing phagocytosis show reduced bead internalization by SDC1-null epithelial cells. Syndecan-1 deficiency alters ICN morphology and homeostasis during aging, reduces epithelial phagocytosis, and impairs reinnervation after trephine but not debridement injury. These data provide insight into the mechanisms used by sensory nerves to reinnervate after injury.
- Corneal Injuries
- Disease Models, Animal
- Epithelium, Corneal
- Mice, Inbred BALB C
- Nerve Fibers
Retinal degeneration and visual impairment are the first signs of juvenile neuronal ceroid lipofuscinosis caused by CLN3 mutations, followed by inevitable progression to blindness. We investigated retinal degeneration in Cln3(Δex1-6) null mice, revealing classic 'fingerprint' lysosomal storage in the retinal pigment epithelium (RPE), replicating the human disease. The lysosomes contain mitochondrial F0-ATP synthase subunit c along with undigested membranes, indicating a reduced degradative capacity. Mature autophagosomes and basal phagolysosomes, the terminal degradative compartments of autophagy and phagocytosis, are also increased in Cln3(Δex1) (-6) RPE, reflecting disruption to these key pathways that underpin the daily phagocytic turnover of photoreceptor outer segments (POS) required for maintenance of vision. The accumulated autophagosomes have post-lysosome fusion morphology, with undigested internal contents visible, while accumulated phagosomes are frequently docked to cathepsin D-positive lysosomes, without mixing of phagosomal and lysosomal contents. This suggests lysosome-processing defects affect both autophagy and phagocytosis, supported by evidence that phagosomes induced in Cln3(Δex1) (-) (6)-derived mouse embryonic fibroblasts have visibly disorganized membranes, unprocessed internal vesicles and membrane contents, in addition to reduced LAMP1 membrane recruitment. We propose that defective lysosomes in Cln3(Δex1) (-) (6) RPE have a reduced degradative capacity that impairs the final steps of the intimately connected autophagic and phagocytic pathways that are responsible for degradation of POS. A build-up of degradative organellar by-products and decreased recycling of cellular materials is likely to disrupt processes vital to maintenance of vision by the RPE.
- Disease Models, Animal
- Membrane Fusion
- Membrane Glycoproteins
- Mice, Inbred C57BL
- Mice, Knockout
- Mitochondrial Proton-Translocating ATPases
- Molecular Chaperones
- Neuronal Ceroid-Lipofuscinoses
- Retinal Pigment Epithelium