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1.
Chem Biol Interact ; 348: 109649, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34516972

RESUMO

Cadmium (Cd) exposure induced lipid metabolic disorder with changes in lipid composition, as well as triglyceride (TG) levels. Liver is the main organ maintaining body TG level and previous studies suggested that Cd exposure might increase TG synthesis but reduce TG uptake in liver. However, the effects of Cd exposure on TG secretion from liver and underlying mechanism are still unclear. In the present study, the data revealed that Cd exposure increased TG levels in the HepG2 cells and the cultured medium by increasing the expression of microsomal triglyceride transfer protein (MTTP), which was abrogated by siRNA knockdown of MTTP. MTTP was synergistically accumulated after Cd exposure or treated with proteasome inhibitor MG132 and lysosome inhibitor chloroquine (CQ), which suggested the Cd increased MTTP protein stability by inhibiting both the proteasome and the lysosomal protein degradation pathways. In addition, our results demonstrated that Cd exposure inhibited the lysosomal acidic degradation pathway through disrupting endoplastic reticulum (ER) Ca2+ homeostasis. Cd-induced MTTP protein and TG levels were significantly reduced by pretreatments of BAPTA/AM chelation of intracellular Ca2+, 2-APB inhibition of ER Ca2+ release channel inositol 1,4,5-trisphosphate receptor (IP3R) and CDN1163 activation of ER Ca2+ reuptake pump sarcoplasmic reticulum Ca2+-ATPase (SERCA). These results suggest that Cd-induced ER Ca2+ release impaired the lysosomal acidity, which associated with MTTP protein accumulation and contributed to increased TG levels.


Assuntos
Cádmio/farmacologia , Proteínas de Transporte/metabolismo , Retículo Endoplasmático/metabolismo , Homeostase/efeitos dos fármacos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Triglicerídeos/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Células Hep G2 , Humanos , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo
2.
Cell Physiol Biochem ; 55(S4): 68-95, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34523304

RESUMO

Protein homeostasis strongly depends on the targeted and selective removal of unneeded or flawed proteins, of protein aggregates, and of damaged or excess organelles by the two main intracellular degradative systems, namely the ubiquitin proteasomal system (UPS) and the autophagosomal lysosomal system. Despite representing completely distinct mechanisms of degradation, which underlie differing regulatory mechanisms, growing evidence suggests that the UPS and autophagy strongly interact especially in situations of overwhelming and impairment, and that both are involved in podocyte proteostasis and in the pathogenesis of podocyte injury. The differential impact of autophagy and the UPS on podocyte biology and on podocyte disease development and progression is not understood. Recent advances in understanding the role of the UPS and autophagy in podocyte biology are reviewed here.


Assuntos
Autofagia , Nefropatias , Podócitos , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina/metabolismo , Animais , Autofagossomos/metabolismo , Autofagossomos/patologia , Humanos , Nefropatias/metabolismo , Nefropatias/patologia , Nefropatias/fisiopatologia , Lisossomos/metabolismo , Lisossomos/patologia , Podócitos/metabolismo , Podócitos/patologia
3.
J Hazard Mater ; 416: 126158, 2021 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-34492938

RESUMO

Graphene oxide (GO) nanomaterials have significant advantages for drug delivery and electrode materials in neural science, however, their exposure risks to the central nervous system (CNS) and toxicity concerns are also increased. The current studies of GO-induced neurotoxicity remain still ambiguous, let alone the mechanism of how complicated GO chemistry affects its biological behavior with neural cells. In this study, we characterized the commercially available GO in detail and investigated its biological adverse effects using cultured SH-SY5Y cells. We found that ultrasonic processing in medium changed the oxidation status and surface reactivity on the planar surface of GO due to its hydration activity, causing lipid peroxidation and cell membrane damage. Subsequently, ROS-disrupted mitochondrial homeostasis, resulting from the activation of NOX2 signaling, was observed following GO internalization. The autophagy-lysosomal network was initiated as a defensive reaction to obliterate oxidative damaged mitochondria and foreign nanomaterials, which was ineffective due to reduced lysosomal degradation capacity. These sequential cellular responses exacerbated mitochondrial stress, leading to apoptotic cell death. These data highlight the importance of the structure-related activity of GO on its biological properties and provide an in-depth understanding of how GO-derived cellular redox signaling induces mitochondrion-related cascades that modulate cell functionality and survival.


Assuntos
Autofagia , Lisossomos , Apoptose , Linhagem Celular Tumoral , Grafite , Homeostase , Lisossomos/metabolismo , Mitocôndrias , Oxirredução , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo
4.
Nat Commun ; 12(1): 5354, 2021 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-34504082

RESUMO

Mitochondrial division is not an autonomous event but involves multiple organelles, including the endoplasmic reticulum (ER) and lysosomes. Whereas the ER drives the constriction of mitochondrial membranes, the role of lysosomes in mitochondrial division is not known. Here, using super-resolution live-cell imaging, we investigate the recruitment of lysosomes to the site of mitochondrial division. We find that the ER recruits lysosomes to the site of division through the interaction of VAMP-associated proteins (VAPs) with the lysosomal lipid transfer protein ORP1L to induce a three-way contact between the ER, lysosome, and the mitochondrion. We also show that ORP1L might transport phosphatidylinositol-4-phosphate (PI(4)P) from lysosomes to mitochondria, as inhibiting its transfer or depleting PI(4)P at the mitochondrial division site impairs fission, demonstrating a direct role for PI(4)P in the division process. Our findings support a model where the ER recruits lysosomes to act in concert at the fission site for the efficient division of mitochondria.


Assuntos
Retículo Endoplasmático/metabolismo , Lisossomos/metabolismo , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Fosfatos de Fosfatidilinositol/metabolismo , Receptores de Esteroides/metabolismo , Transdução de Sinais , Animais , Células COS , Linhagem Celular Tumoral , Chlorocebus aethiops , Células HeLa , Humanos , Microscopia Confocal/métodos , Interferência de RNA , Receptores de Esteroides/genética , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
5.
Biol Aujourdhui ; 215(1-2): 25-43, 2021.
Artigo em Francês | MEDLINE | ID: mdl-34397373

RESUMO

Targeted protein degradation (TPD), discovered twenty years ago through the PROTAC technology, is rapidly developing thanks to the implication of many scientists from industry and academia. PROTAC chimeras are heterobifunctional molecules able to link simultaneously a protein to be degraded and an E3 ubiquitin ligase. This allows the protein ubiquitination and its degradation by 26S proteasome. PROTACs have evolved from small peptide molecules to small non-peptide and orally available molecules. It was shown that PROTACs are capable to degrade proteins considered as "undruggable" i.e. devoid of well-defined pockets and deep grooves possibly occupied by small molecules. Among these "hard to drug" proteins, several can be degraded by PROTACs: scaffold proteins, BAF complex, transcription factors, Ras family proteins. Two PROTACs are clinically tested for breast (ARV471) and prostate (ARV110) cancers. The protein degradation by proteasome is also induced by other types of molecules: molecular glues, hydrophobic tagging (HyT), HaloPROTACs and homo-PROTACs. Other cellular constituents are eligible to induced degradation: RNA-PROTACs for RNA binding proteins and RIBOTACs for degradation of RNA itself (SARS-CoV-2 RNA). TPD has recently moved beyond the proteasome with LYTACs (lysosome targeting chimeras) and MADTACs (macroautophagy degradation targeting chimeras). Several techniques such as screening platforms together with mathematical modeling and computational design are now used to improve the discovery of new efficient PROTACs.


Assuntos
COVID-19/tratamento farmacológico , Desenho de Fármacos , Terapia de Alvo Molecular/métodos , Proteólise , Proteínas Recombinantes de Fusão/farmacologia , SARS-CoV-2/efeitos dos fármacos , Antineoplásicos/química , Antineoplásicos/farmacologia , Autofagia , Catálise , Humanos , Lisossomos/metabolismo , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Complexo de Endopeptidases do Proteassoma/metabolismo , Conformação Proteica , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estabilidade Proteica , Proteólise/efeitos dos fármacos , RNA/efeitos dos fármacos , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/farmacocinética , Relação Estrutura-Atividade , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
6.
Int Rev Cell Mol Biol ; 363: 203-269, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34392930

RESUMO

An increase in intracellular Ca2+ concentration ([Ca2+]i) regulates a plethora of functions in the cardiovascular (CV) system, including contraction in cardiomyocytes and vascular smooth muscle cells (VSMCs), and angiogenesis in vascular endothelial cells and endothelial colony forming cells. The sarco/endoplasmic reticulum (SR/ER) represents the largest endogenous Ca2+ store, which releases Ca2+ through ryanodine receptors (RyRs) and/or inositol-1,4,5-trisphosphate receptors (InsP3Rs) upon extracellular stimulation. The acidic vesicles of the endolysosomal (EL) compartment represent an additional endogenous Ca2+ store, which is targeted by several second messengers, including nicotinic acid adenine dinucleotide phosphate (NAADP) and phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2], and may release intraluminal Ca2+ through multiple Ca2+ permeable channels, including two-pore channels 1 and 2 (TPC1-2) and Transient Receptor Potential Mucolipin 1 (TRPML1). Herein, we discuss the emerging, pathophysiological role of EL Ca2+ signaling in the CV system. We describe the role of cardiac TPCs in ß-adrenoceptor stimulation, arrhythmia, hypertrophy, and ischemia-reperfusion injury. We then illustrate the role of EL Ca2+ signaling in VSMCs, where TPCs promote vasoconstriction and contribute to pulmonary artery hypertension and atherosclerosis, whereas TRPML1 sustains vasodilation and is also involved in atherosclerosis. Subsequently, we describe the mechanisms whereby endothelial TPCs promote vasodilation, contribute to neurovascular coupling in the brain and stimulate angiogenesis and vasculogenesis. Finally, we discuss about the possibility to target TPCs, which are likely to mediate CV cell infection by the Severe Acute Respiratory Disease-Coronavirus-2, with Food and Drug Administration-approved drugs to alleviate the detrimental effects of Coronavirus Disease-19 on the CV system.


Assuntos
COVID-19/complicações , COVID-19/tratamento farmacológico , Sinalização do Cálcio/fisiologia , Doenças Cardiovasculares/etiologia , Doenças Cardiovasculares/metabolismo , Sistema Cardiovascular/metabolismo , Lisossomos/metabolismo , SARS-CoV-2 , ADP-Ribosil Ciclase 1/metabolismo , Animais , Encéfalo/irrigação sanguínea , Encéfalo/metabolismo , COVID-19/metabolismo , Canais de Cálcio/metabolismo , Doenças Cardiovasculares/tratamento farmacológico , Retículo Endoplasmático/metabolismo , Células Endoteliais/metabolismo , Humanos , Modelos Cardiovasculares , Miócitos Cardíacos/metabolismo , NADP/análogos & derivados , NADP/metabolismo , Receptores Adrenérgicos beta/metabolismo , Retículo Sarcoplasmático/metabolismo , Canais de Potencial de Receptor Transitório/metabolismo
7.
Nat Commun ; 12(1): 4739, 2021 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-34362892

RESUMO

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a potent Ca2+-mobilizing second messenger which uniquely mobilizes Ca2+ from acidic endolysosomal organelles. However, the molecular identity of the NAADP receptor remains unknown. Given the necessity of the endolysosomal two-pore channel (TPC1 or TPC2) in NAADP signaling, we performed affinity purification and quantitative proteomic analysis of the interacting proteins of NAADP and TPCs. We identified a Sm-like protein Lsm12 complexed with NAADP, TPC1, and TPC2. Lsm12 directly binds to NAADP via its Lsm domain, colocalizes with TPC2, and mediates the apparent association of NAADP to isolated TPC2 or TPC2-containing membranes. Lsm12 is essential and immediately participates in NAADP-evoked TPC activation and Ca2+ mobilization from acidic stores. These findings reveal a putative RNA-binding protein to function as an NAADP receptor and a TPC regulatory protein and provides a molecular basis for understanding the mechanisms of NAADP signaling.


Assuntos
Cálcio/metabolismo , NADP/análogos & derivados , Porinas/metabolismo , Animais , Canais de Cálcio/metabolismo , Linhagem Celular , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/química , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Endossomos/metabolismo , Fibroblastos , Células HEK293 , Humanos , Lisossomos/metabolismo , Camundongos , Mutação , NADP/genética , NADP/metabolismo , Proteômica
8.
Environ Pollut ; 286: 117295, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34438478

RESUMO

Ambient particulate matter (APM) has been authenticated to exert hazards on human vascular endothelial cells, including abnormal autophagy. However, the potential reasons for autophagosome accumulation are still obscure. Since autophagy is a dynamic process, it is imperative to systemically consider the autophagic induction combined with its degradation to reflect realistic scenarios. Therefore, in the current study, different exposure durations were initially employed for the detection of autophagic marker proteins to assess the dynamic autophagic state preliminarily. Additionally, LC3 turn-over and autophagic flux assays were used to determine the specific cause of LC3II upregulation in EA.hy926 human vascular endothelial cells by a type of standard urban particulate matter, PM SRM1648a. As a result, PM SRM1648a stimulates excess autophagic vacuoles in EA. hy926 cells, in which the underlying causes are probably different at varying incubation endpoints. Intriguingly, LC3II upregulation was due to the intensifying autophagic initiation after 6 h of exposure, whereas as exposure period was extended to 24 h, overloaded autophagic vacuoles were attributed to the defective autophagy. Mechanistically, PM SRM1648a damages EA. hy926 cells by inducing lysosomal disequilibrium and resultant autophagic malfunction which are not directly mediated by oxidative stress. These data indicate that appropriate maintenance of lysosomal function and autophagic flux is probably a protective measure against APM-induced endothelial cell damage.


Assuntos
Células Endoteliais , Material Particulado , Autofagia , Humanos , Lisossomos/metabolismo , Estresse Oxidativo , Material Particulado/metabolismo , Material Particulado/toxicidade
9.
Stem Cell Res Ther ; 12(1): 452, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34380561

RESUMO

Mitophagy is a specific autophagic phenomenon in which damaged or redundant mitochondria are selectively cleared by autophagic lysosomes. A decrease in mitophagy can accelerate the aging process. Mitophagy is related to health and longevity and is the key to protecting stem cells from metabolic stress damage. Mitophagy decreases the metabolic level of stem cells by clearing active mitochondria, so mitophagy is becoming increasingly necessary to maintain the regenerative capacity of old stem cells. Stem cell senescence is the core problem of tissue aging, and tissue aging occurs not only in stem cells but also in transport amplifying cell chambers and the stem cell environment. The loss of the autophagic ability of stem cells can cause the accumulation of mitochondria and the activation of the metabolic state as well as damage the self-renewal ability and regeneration potential of stem cells. However, the claim remains controversial. Mitophagy is an important survival strategy against nutrient deficiency and starvation, and mitochondrial function and integrity may affect the viability, proliferation and differentiation potential, and longevity of normal stem cells. Mitophagy can affect the health and longevity of the human body, so the number of studies in this field has increased, but the mechanism by which mitophagy participates in stem cell development is still not fully understood. This review describes the potential significance of mitophagy in stem cell developmental processes, such as self-renewal, differentiation and aging. Through this work, we discovered the role and mechanism of mitophagy in different types of stem cells, identified novel targets for killing cancer stem cells and curing cancer, and provided new insights for future research in this field.


Assuntos
Mitocôndrias , Mitofagia , Autofagia , Humanos , Lisossomos/metabolismo , Células-Tronco/metabolismo
10.
Nat Commun ; 12(1): 5134, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34446734

RESUMO

The gastrointestinal tract may be a site of origin for α-synuclein pathology in idiopathic Parkinson's disease (PD). Disruption of the autophagy-lysosome pathway (ALP) may contribute to α-synuclein aggregation. Here we examined epigenetic alterations in the ALP in the appendix by deep sequencing DNA methylation at 521 ALP genes. We identified aberrant methylation at 928 cytosines affecting 326 ALP genes in the appendix of individuals with PD and widespread hypermethylation that is also seen in the brain of individuals with PD. In mice, we find that DNA methylation changes at ALP genes induced by chronic gut inflammation are greatly exacerbated by α-synuclein pathology. DNA methylation changes at ALP genes induced by synucleinopathy are associated with the ALP abnormalities observed in the appendix of individuals with PD specifically involving lysosomal genes. Our work identifies epigenetic dysregulation of the ALP which may suggest a potential mechanism for accumulation of α-synuclein pathology in idiopathic PD.


Assuntos
Apêndice/metabolismo , Autofagia , Epigênese Genética , Lisossomos/metabolismo , Doença de Parkinson/metabolismo , Animais , Apêndice/química , Encéfalo/metabolismo , Encéfalo/patologia , Metilação de DNA , Feminino , Humanos , Lisossomos/química , Lisossomos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Doença de Parkinson/genética , Doença de Parkinson/patologia , Agregados Proteicos , alfa-Sinucleína/química , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo
11.
Nat Commun ; 12(1): 5073, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34417467

RESUMO

The contents of numerous membrane lipids change upon ageing. However, it is unknown whether and how any of these changes are causally linked to lifespan regulation. Acyl chains contribute to the functional specificity of membrane lipids. In this study, working with C. elegans, we identified an acyl chain-specific sphingolipid, C22 glucosylceramide, as a longevity metabolite. Germline deficiency, a conserved lifespan-extending paradigm, induces somatic expression of the fatty acid elongase ELO-3, and behenic acid (22:0) generated by ELO-3 is incorporated into glucosylceramide for lifespan regulation. Mechanistically, C22 glucosylceramide is required for the membrane localization of clathrin, a protein that regulates membrane budding. The reduction in C22 glucosylceramide impairs the clathrin-dependent autophagic lysosome reformation, which subsequently leads to TOR activation and longevity suppression. These findings reveal a mechanistic link between membrane lipids and ageing and suggest a model of lifespan regulation by fatty acid-mediated membrane configuration.


Assuntos
Caenorhabditis elegans/fisiologia , Ácidos Graxos não Esterificados/metabolismo , Glicoesfingolipídeos/metabolismo , Homeostase , Longevidade/fisiologia , Lisossomos/metabolismo , Animais , Proteínas de Caenorhabditis elegans/metabolismo , Ceramidas/metabolismo , Colesterol/metabolismo , Clatrina/metabolismo , Mutação em Linhagem Germinativa/genética , Proteínas de Fluorescência Verde/metabolismo , Larva/metabolismo , Modelos Biológicos , Interferência de RNA , Estresse Fisiológico
13.
Int J Mol Sci ; 22(15)2021 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-34360917

RESUMO

Although once perceived as inert structures that merely serve for lipid storage, lipid droplets (LDs) have proven to be the dynamic organelles that hold many cellular functions. The LDs' basic structure of a hydrophobic core consisting of neutral lipids and enclosed in a phospholipid monolayer allows for quick lipid accessibility for intracellular energy and membrane production. Whereas formed at the peripheral and perinuclear endoplasmic reticulum, LDs are degraded either in the cytosol by lipolysis or in the vacuoles/lysosomes by autophagy. Autophagy is a regulated breakdown of dysfunctional, damaged, or surplus cellular components. The selective autophagy of LDs is called lipophagy. Here, we review LDs and their degradation by lipophagy in yeast, which proceeds via the micrometer-scale raft-like lipid domains in the vacuolar membrane. These vacuolar microdomains form during nutrient deprivation and facilitate internalization of LDs via the vacuolar membrane invagination and scission. The resultant intra-vacuolar autophagic bodies with LDs inside are broken down by vacuolar lipases and proteases. This type of lipophagy is called microlipophagy as it resembles microautophagy, the type of autophagy when substrates are sequestered right at the surface of a lytic compartment. Yeast microlipophagy via the raft-like vacuolar microdomains is a great model system to study the role of lipid domains in microautophagic pathways.


Assuntos
Autofagia/fisiologia , Gotículas Lipídicas/metabolismo , Microdomínios da Membrana/metabolismo , Saccharomyces cerevisiae/metabolismo , Vacúolos/metabolismo , Citosol/metabolismo , Retículo Endoplasmático/metabolismo , Homeostase/fisiologia , Lipólise/fisiologia , Lisossomos/metabolismo , Fosfolipídeos/metabolismo
14.
Molecules ; 26(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34361780

RESUMO

Parkinson's disease is characterized by the loss of dopaminergic neurons in substantia nigra pars compacta (SNpc) and the resultant loss of dopamine in the striatum. Various studies have shown that oxidative stress and neuroinflammation plays a major role in PD progression. In addition, the autophagy lysosome pathway (ALP) plays an important role in the degradation of aggregated proteins, abnormal cytoplasmic organelles and proteins for intracellular homeostasis. Dysfunction of ALP results in the accumulation of α-synuclein and the loss of dopaminergic neurons in PD. Thus, modulating ALP is becoming an appealing therapeutic intervention. In our current study, we wanted to evaluate the neuroprotective potency of noscapine in a rotenone-induced PD rat model. Rats were administered rotenone injections (2.5 mg/kg, i.p.,) daily followed by noscapine (10 mg/kg, i.p.,) for four weeks. Noscapine, an iso-qinulinin alkaloid found naturally in the Papaveraceae family, has traditionally been used in the treatment of cancer, stroke and fibrosis. However, the neuroprotective potency of noscapine has not been analyzed. Our study showed that administration of noscapine decreased the upregulation of pro-inflammatory factors, oxidative stress, and α-synuclein expression with a significant increase in antioxidant enzymes. In addition, noscapine prevented rotenone-induced activation of microglia and astrocytes. These neuroprotective mechanisms resulted in a decrease in dopaminergic neuron loss in SNpc and neuronal fibers in the striatum. Further, noscapine administration enhanced the mTOR-mediated p70S6K pathway as well as inhibited apoptosis. In addition to these mechanisms, noscapine prevented a rotenone-mediated increase in lysosomal degradation, resulting in a decrease in α-synuclein aggregation. However, further studies are needed to further develop noscapine as a potential therapeutic candidate for PD treatment.


Assuntos
Autofagia/efeitos dos fármacos , Corpo Estriado/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Noscapina/farmacologia , Doença de Parkinson Secundária/tratamento farmacológico , Doença de Parkinson Secundária/genética , Parte Compacta da Substância Negra/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Astrócitos/patologia , Catalase/genética , Catalase/metabolismo , Corpo Estriado/metabolismo , Corpo Estriado/patologia , Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Estresse Oxidativo/efeitos dos fármacos , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/patologia , Parte Compacta da Substância Negra/metabolismo , Parte Compacta da Substância Negra/patologia , Ratos , Ratos Wistar , Proteínas Quinases S6 Ribossômicas 70-kDa/genética , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Rotenona/toxicidade , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , alfa-Sinucleína/antagonistas & inibidores , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo
15.
Int J Mol Sci ; 22(16)2021 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-34445706

RESUMO

Here, we present the main features of human acid sphingomyelinase (ASM), its biosynthesis, processing and intracellular trafficking, its structure, its broad substrate specificity, and the proposed mode of action at the surface of the phospholipid substrate carrying intraendolysosomal luminal vesicles. In addition, we discuss the complex regulation of its phospholipid cleaving activity by membrane lipids and lipid-binding proteins. The majority of the literature implies that ASM hydrolyses solely sphingomyelin to generate ceramide and ignores its ability to degrade further substrates. Indeed, more than twenty different phospholipids are cleaved by ASM in vitro, including some minor but functionally important phospholipids such as the growth factor ceramide-1-phosphate and the unique lysosomal lysolipid bis(monoacylglycero)phosphate. The inherited ASM deficiency, Niemann-Pick disease type A and B, impairs mainly, but not only, cellular sphingomyelin catabolism, causing a progressive sphingomyelin accumulation, which furthermore triggers a secondary accumulation of lipids (cholesterol, glucosylceramide, GM2) by inhibiting their turnover in late endosomes and lysosomes. However, ASM appears to be involved in a variety of major cellular functions with a regulatory significance for an increasing number of metabolic disorders. The biochemical characteristics of ASM, their potential effect on cellular lipid turnover, as well as a potential impact on physiological processes will be discussed.


Assuntos
Fosfolipídeos/biossíntese , Esfingomielina Fosfodiesterase/biossíntese , Esfingomielina Fosfodiesterase/metabolismo , Transporte Biológico , Ceramidas/metabolismo , Colesterol/metabolismo , Endossomos/metabolismo , Humanos , Lisossomos/metabolismo , Lipídeos de Membrana/metabolismo , Doença de Niemann-Pick Tipo A/metabolismo , Fosfolipídeos/metabolismo , Esfingomielina Fosfodiesterase/fisiologia , Esfingomielinas/metabolismo , Fosfolipases Tipo C/metabolismo , Fosfolipases Tipo C/fisiologia
16.
Int J Mol Sci ; 22(16)2021 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-34445748

RESUMO

In age-related macular degeneration (AMD), hydroquinone (HQ)-induced oxidative damage in retinal pigment epithelium (RPE) is believed to be an early event contributing to dysregulation of inflammatory cytokines and vascular endothelial growth factor (VEGF) homeostasis. However, the roles of antioxidant mechanisms, such as autophagy and the ubiquitin-proteasome system, in modulating HQ-induced oxidative damage in RPE is not well-understood. This study utilized an in-vitro AMD model involving the incubation of human RPE cells (ARPE-19) with HQ. In comparison to hydrogen peroxide (H2O2), HQ induced fewer reactive oxygen species (ROS) but more oxidative damage as characterized by protein carbonyl levels, mitochondrial dysfunction, and the loss of cell viability. HQ blocked the autophagy flux and increased proteasome activity, whereas H2O2 did the opposite. Moreover, the lysosomal membrane-stabilizing protein LAMP2 and cathepsin D levels declined with HQ exposure, suggesting loss of lysosomal membrane integrity and function. Accordingly, HQ induced lysosomal alkalization, thereby compromising the acidic pH needed for optimal lysosomal degradation. Pretreatment with MG132, a proteasome inhibitor and lysosomal stabilizer, upregulated LAMP2 and autophagy and prevented HQ-induced oxidative damage in wildtype RPE cells but not cells transfected with shRNA against ATG5. This study demonstrated that lysosomal dysfunction underlies autophagy defects and oxidative damage induced by HQ in human RPE cells and supports lysosomal stabilization with the proteasome inhibitor MG132 as a potential remedy for oxidative damage in RPE and AMD.


Assuntos
Autofagia , Lisossomos/metabolismo , Degeneração Macular/etiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Catepsina D/metabolismo , Células Cultivadas , Humanos , Hidroquinonas , Leupeptinas , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Degeneração Macular/metabolismo , Mitocôndrias/metabolismo , Epitélio Pigmentado da Retina/citologia
17.
Biomolecules ; 11(7)2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34356645

RESUMO

Microphthalmia-associated transcription factor (MITF) is the principal transcription factor regulating pivotal processes in melanoma cell development, growth, survival, proliferation, differentiation and invasion. In recent years, convincing evidence has been provided attesting key roles of endolysosomal cation channels, specifically TPCs and TRPMLs, in cancer, including breast cancer, glioblastoma, bladder cancer, hepatocellular carcinoma and melanoma. In this review, we provide a gene expression profile of these channels in different types of cancers and decipher their roles, in particular the roles of two-pore channel 2 (TPC2) and TRPML1 in melanocytes and melanoma. We specifically discuss the signaling cascades regulating MITF and the relationship between endolysosomal cation channels, MAPK, canonical Wnt/GSK3 pathways and MITF.


Assuntos
Canais Iônicos/metabolismo , Melanócitos/metabolismo , Melanoma/patologia , Fator de Transcrição Associado à Microftalmia/metabolismo , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Endossomos/metabolismo , Expressão Gênica , Humanos , Canais Iônicos/genética , Lisossomos/metabolismo , Melanócitos/patologia , Melanoma/metabolismo , Fator de Transcrição Associado à Microftalmia/genética , Canais de Potencial de Receptor Transitório/genética , Canais de Potencial de Receptor Transitório/metabolismo
18.
J Cell Sci ; 134(2)2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-34432035

RESUMO

Pharmacological manipulation of lysosome membrane integrity or ionic movements is a key strategy for probing lysosomal involvement in cellular processes. However, we have found an unexpected inhibition of store-operated Ca2+ entry (SOCE) by these agents. Dipeptides [glycyl-L-phenylalanine 2-naphthylamide (GPN) and L-leucyl-L-leucine methyl ester] that are inducers of lysosomal membrane permeabilization (LMP) uncoupled endoplasmic reticulum Ca2+-store depletion from SOCE by interfering with Stim1 oligomerization and/or Stim1 activation of Orai. Similarly, the K+/H+ ionophore, nigericin, that rapidly elevates lysosomal pH, also inhibited SOCE in a Stim1-dependent manner. In contrast, other strategies for manipulating lysosomes (bafilomycin A1, lysosomal re-positioning) had no effect upon SOCE. Finally, the effects of GPN on SOCE and Stim1 was reversed by a dynamin inhibitor, dynasore. Our data show that lysosomal agents not only release Ca2+ from stores but also uncouple this release from the normal recruitment of Ca2+ influx.


Assuntos
Canais de Cálcio , Cálcio , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Sinalização do Cálcio , Retículo Endoplasmático/metabolismo , Lisossomos/metabolismo , Proteína ORAI1/metabolismo , Molécula 1 de Interação Estromal/metabolismo
19.
Molecules ; 26(13)2021 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-34279378

RESUMO

The aim of the study is to evaluate oxidant-antioxidant balance as well as lysosomal and anti-protease activities in ovarian cancer since it has been emphasized that the crucial inducing factor of carcinogenesis may be reactive oxygen/nitrogen species or, more precisely, oxidative stress-induced inflammation. The study involved 15 women with ovarian cancer, aged 59.9 ± 7.8 years, and 9 healthy women aged 56.3 ± 4.3 years (controls). The study material was venous blood collected from fasting subjects. In erythrocytes, the activities of superoxide dismutase, glutathione peroxidase, and catalase, as well as concentrations of conjugated dienes (CDs) and thiobarbituric acid reactive substances (TBARS), were investigated. CD, TBARS, and vitamins A and E plasma concentrations were also determined. Moreover, total antioxidant capacity and concentrations of 4-hydroxynonenal adducts and 8-iso-prostaglandin F2α, as well as activities of acid phosphatase, arylsulfatase, cathepsin D, and α1-antitrypsin, were studied in serum. The vitamin E and 8-iso-prostaglandin F2α concentrations as well as arylsulfatase activity were lower in the women with cancer compared to the controls (p = 0.006, p = 0.03, p = 0.001, respectively). In contrast, cathepsin D activity was lower in the controls (p = 0.04). In the peripheral blood of the women with cancer, oxidant-antioxidant and lysosomal disturbances were observed.


Assuntos
Lisossomos/metabolismo , Recidiva Local de Neoplasia/sangue , Neoplasias Ovarianas/sangue , Estresse Oxidativo , Idoso , Catalase/sangue , Catepsina D/sangue , Dinoprosta/sangue , Feminino , Glutationa Peroxidase/sangue , Humanos , Pessoa de Meia-Idade , Metástase Neoplásica , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Superóxido Dismutase/sangue , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Vitamina A/sangue , Vitamina E/sangue
20.
Int J Mol Sci ; 22(14)2021 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-34299248

RESUMO

Parkinson's disease (PD) is a complex and progressive neurodegenerative disorder with a prevalence of approximately 0.5-1% among those aged 65-70 years. Although most of its clinical manifestations are due to a loss of dopaminergic neurons, the PD etiology is largely unknown. PD is caused by a combination of genetic and environmental factors, and the exact interplay between genes and the environment is still debated. Several biological processes have been implicated in PD, including mitochondrial or lysosomal dysfunctions, alteration in protein clearance, and neuroinflammation, but a common molecular mechanism connecting the different cellular alterations remains incompletely understood. Accumulating evidence underlines a significant role of lipids in the pathological pathways leading to PD. Beside the well-described lipid alteration in idiopathic PD, this review summarizes the several lipid alterations observed in experimental models expressing PD-related genes and suggests a possible scenario in relationship to the molecular mechanisms of neuronal toxicity. PD could be considered a lipid-induced proteinopathy, where alteration in lipid composition or metabolism could induce protein alteration-for instance, alpha-synuclein accumulation-and finally neuronal death.


Assuntos
Metabolismo dos Lipídeos/genética , Lipídeos/fisiologia , Doença de Parkinson/genética , Neurônios Dopaminérgicos/metabolismo , Glucosilceramidase/genética , Glucosilceramidase/metabolismo , Fosfolipases A2 do Grupo VI/genética , Fosfolipases A2 do Grupo VI/metabolismo , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Lisossomos/metabolismo , Mitocôndrias/metabolismo , Degeneração Neural/patologia , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo
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