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1.
Talanta ; 236: 122884, 2022 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-34635264

RESUMO

This work describes (Z)-N-((Z)-2-(1,3,3-trimethylindolin-2ylidene)ethylidene)quinoline-8-amine (LYSO-QF), a high-performing and biocompatible dye comprised of quinoline and Fisher aldehyde moieties linked via an imine vinyl backbone with lysosome targeting ability that can be used to quantitatively detect the mercury ion (Hg2+) in biosystems and the natural environment. This is achieved by forming three different tetrameric, trimeric and dimeric complexes between Hg2+ and LYSO-QF with the limit of detection (LOD) of 11 nm. The complexes formed were analyzed with the aid of time-dependent density functional theory (TD-DFT) calculations. The concentration dependence of the Hg2+ complex fluorescence emission changes from grey-green to jade green and then to red as the different types of complex are formed. The favorable sensor properties of the LYSO-QF probe are demonstrated by monitoring different Hg2+ concentrations in buffer solutions, HeLa cells, zebrafish model samples and several different types of water sample. Experiments with Whatman paper strips demonstrate that the cost-effective LYSO-QF also has considerable potential for use in on-site Hg2+ detection with the naked eye.


Assuntos
Complexos de Coordenação , Mercúrio , Quinolinas , Aldeídos , Animais , Células HeLa , Humanos , Lisossomos , Peixe-Zebra
2.
J Cell Sci ; 135(5)2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33912962

RESUMO

Membrane contact sites (MCSs) between the endoplasmic reticulum (ER) and late endosomes/lysosomes (LE/lys) are emerging as critical hubs for diverse cellular events, and changes in their extents are linked to severe neurological diseases. While recent studies show that the synaptotagmin-like mitochondrial-lipid-binding (SMP) domain-containing protein PDZD8 may mediate the formation of ER-LE/lys MCSs, the cellular functions of PDZD8 remain largely elusive. Here, we attempt to investigate the lipid transfer activities of PDZD8 and the extent to which its cellular functions depend on its lipid transfer activities. In accordance with recent studies, we demonstrate that PDZD8 is a protrudin (ZFYVE27)-interacting protein and that PDZD8 acts as a tether at ER-LE/lys MCSs. Furthermore, we discover that the SMP domain of PDZD8 binds glycerophospholipids and ceramides both in vivo and in vitro, and that the SMP domain can transport lipids between membranes in vitro. Functionally, PDZD8 is required for LE/lys positioning and neurite outgrowth, which is dependent on the lipid transfer activity of the SMP domain.


Assuntos
Retículo Endoplasmático , Endossomos , Lipídeos , Lisossomos , Crescimento Neuronal
3.
J Nutr Sci Vitaminol (Tokyo) ; 67(4): 243-248, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34470999

RESUMO

Daily fat and sugar intake has increased in Japan, while total energy intake has decreased. However, the number of type 2 diabetes mellitus patients has increased, and this often causes renal injury characterized by autophagic vacuoles. Although many studies with comparisons of high fat or sugar versus a normal macronutrient balanced diet have been reported, there are few studies that equalized calorie intake and body weights. In the current study, AIN93M diets (CONT group) with matching energy content with lard derived high saturated fat (LARD group), soybean oil derived unsaturated fat (SOY OIL group) and sucrose (SUCROSE group) were provided to compare their effects on renal morphology in streptozotocin-injected CD-1 mice without causing obesity. The number of renal tubular vacuoles was higher in SUCROSE and slightly higher in LARD compared with CONT mice, and was higher in LARD and SUCROSE compared with SOY OIL mice. Most of those vacuoles were LAMP1-positive, a marker of lysosomal autophagy. These results suggest that despite identical energy contents, diets with high sucrose or saturated fat compared to unsaturated fat may aggravate lysosomal renal injury in a non-obese, streptozotocin-induced model of diabetes mellitus.


Assuntos
Diabetes Mellitus Tipo 2 , Sacarose , Animais , Dieta , Gorduras na Dieta , Humanos , Rim , Lisossomos , Camundongos , Estreptozocina , Sacarose/efeitos adversos
4.
Talanta ; 235: 122659, 2021 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-34517575

RESUMO

Sensitive detection of ß-galactosidase (ß-gal) is of great significance for early diagnosis of ovarian cancer. Fluorescent probes for detecting ß-gal have received great interest due to the non-invasiveness, excellent sensitivity, high temporal, and superior spatial resolution. However, most reported fluorescent sensors for ß-gal suffer from aggregation caused quenching effect when accumulated, and cannot discriminate ß-gal from other species, especially, Escherichia coliß-gal. Herein, we report the first aggregation-induced emission (AIE)-active fluorescent probe HBTTPAG, which achieves species-selective detection of ß-gal. Probe HBTTPAG can discriminate Aspergillus oryzae ß-gal from Escherichia coliß-gal, with high sensitivity (detection limit of 3.7 × 10-3 UmL-1), superior selectivity and low cytotoxicity. Furthermore, HBTTPAG is utilized to visualize endogenous ß-gal in lysosomes of SKOV-3 cells, as well as to detect ß-gal activity in ovarian cancer tissues. Notably, owing to the AIE-active, HBTTPAG realizes long-term (12 h) tracking ß-gal in ovarian cancer cells. This work provides a promising method for species-selective detection of ß-gal in preclinical.


Assuntos
Imagem Óptica , Neoplasias Ovarianas , Feminino , Corantes Fluorescentes , Humanos , Lisossomos , beta-Galactosidase
5.
J Biomed Nanotechnol ; 17(8): 1647-1653, 2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-34544541

RESUMO

MicroRNA (miRNA) has emerged as an important gene-regulator that shows great potential in gene therapy because of its unique roles in gene-regulation. However, the knowledge on their function and transportation in vivo is still lacking, and there are limited obvious evidences to define intracellular transportation of miRNA. In this study, the dynamics of exogenous miR-21 transfected into HeLa cells was traced by live-cell microscopy. Their transportation at key time points was recorded and dynamic properties were analyzed by single particle tracking (SPT) and mean square displacement (MSD) calculation. Results showed that the exogenous miRNAs bounded to cells quickly and went through lysosome into cytosol, where they were subsequently recruited into p-body. They finally were degraded, otherwise went back to cytosol in some way. Long time observation and analysis of motion mode showed that the miRNAs were confined in a small region and their motion modes were flexible in different intracellular microenvironment after entering the cells.


Assuntos
MicroRNAs , Regulação da Expressão Gênica , Células HeLa , Humanos , Lisossomos , MicroRNAs/genética , Microscopia
6.
Langmuir ; 37(36): 10818-10826, 2021 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-34470217

RESUMO

Direct visualization of the dynamic events in lysosomes during drug-mediated programmed cell death (apoptosis) is a great challenge. This is due to the lack of resolving power of a conventional microscope and also the unavailability of a suitable multimodal probe that simultaneously can carry the drug with high loading capacity and ensure its specific internalization into lysosomes. In this work, using super-resolution microscopy, we observed the lysosomal expansion during apoptosis that was treated with epigallocatechin gallate (EGCG) conjugated to bovine serum albumin (BSA). Albumin protein is known to internalize into lysosomes via endocytosis, thus helping in the specific delivery of EGCG to the lysosomal compartment. The conjugation of EGCG to BSA not only helped in increasing the killing efficiency of cancer cells but it also reduces the side effects and produces minimal reactive oxygen species. The decrease in local viscosity helped in lysosomal expansion during apoptosis.


Assuntos
Catequina , Microscopia , Apoptose , Catequina/análogos & derivados , Lisossomos , Espécies Reativas de Oxigênio
7.
Orphanet J Rare Dis ; 16(1): 379, 2021 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-34496908

RESUMO

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) is the causative agent of the current COVID-19 pandemic. Lysosomal storage disorders (LSD) comprise of 70 inherited inborn errors of metabolism. Affected individuals suffer from multi-systemic involvement with variable severity and rate of disease progression between different diseases. Some of the LSDs have established treatments, whether parenteral or oral therapies. The full impact of the COVID-19 pandemic together with the lockdown on the wellbeing and medical management of patients with rare diseases, such as LSDs, is widely unknown. Herein, we describe the effects of the COVID-19 pandemic and its associated mandatory home lockdown on patients with LSDs in Israel. RESULTS: We present a prospective multi-center questionnaire study including 48 LSD patients from four medical centers in Israel. The study objective was to assess the impact of the COVID-19 pandemic restrictions on individuals with LSDs in Israel, as reported by their caregivers. Secondary objectives were to assess the morbidity from SARS CoV-2 in LSD patients and the impact of changes in mood and behavior on compliance to treatment and to assess the relationship between changes in mood to changes in cognition and behavior. Thirty one of 38 patients (82%) who received any kind of regular treatment did not miss treatments. Among patients receiving enzyme replacement therapy (ERT) in the in-hospital setting, 5 patients (20%) experienced treatment disruptions. Four patients had tested positive for SARS-Cov-2 virus infection by PCR. Seven out of the 48 patients (14%) described mood changes with cognitive and motor deterioration during the home quarantine. CONCLUSIONS: We observed high rates of treatment adherence and low morbidity through the COVID-19 pandemic in patients with LSDs in Israel. LSDs patients can be a model for patients with complex chronic diseases requiring routine treatments and surveillance during a pandemic or other disruption of daily routine.


Assuntos
COVID-19 , Pandemias , Controle de Doenças Transmissíveis , Humanos , Israel/epidemiologia , Lisossomos , Estudos Prospectivos , SARS-CoV-2
8.
Anal Chem ; 93(37): 12639-12647, 2021 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-34491716

RESUMO

Lysosomal membrane permeabilization (LMP) engaged in multiple human diseases is accompanied by relocation of cytosolic galectin into LMP+ lysosomes. We herein reported a galectin trafficking-targeted method to image LMP using two kinds of glyco-dendrimers, a sialic acid-terminated dendrimer labeled with pH-inert rhodamine and a lactose-terminated dendrimer labeled with fluorescein that becomes green-emissive in pH-elevated lysosomes. Albeit both accumulated in physiological lysosomes, the former is released from LMP+ lysosomes while the latter binds to galectin accumulated in LMP+ lysosomes and thus trapped in LMP+ lysosomes. Accordingly, LMP+ lysosomes exhibit loss of red fluorescence and turn-on green fluorescence due to loss of lysosomal acidity. This red-to-green color switch enables discernment of LMP+ lysosomes from physiological lysosomes and pH-elevated lysosomes and can be further utilized to detect LMP in distinct cell death pathways. These results suggest the utility of galectin trafficking pathway-integrated synthetic probes for detection of LMP, a key factor for diseased cells.


Assuntos
Galectinas , Lisossomos , Morte Celular , Citosol , Humanos , Membranas Intracelulares
9.
Anal Chem ; 93(38): 13038-13044, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34519497

RESUMO

Autophagy plays a critical role in many vitally important physiological and pathological processes, such as the removal of damaged and aged organelles and redundant proteins. Although autophagy is mainly a protective process for cells, it can also cause cell death. In this study, we employed in situ and ex situ surface-enhanced Raman scattering (SERS) spectroscopies to obtain chemical information of lysosomes of HepG2 cells. Results reveal that the SERS profiles of the isolated lysosomes are different from the in situ spectra, indicating that lysosomes lie in different microenvironments in these two cases. We further investigated the molecular changes of isolated lysosomes according to the autophagy induced by starvation via ex situ SERS. During autophagy, the conformation of proteins and the structures of lipids have been affected, and autophagy-related molecular evidence is given for the first time in the living lysosomes. We expect that this study will provide a reference for understanding the cell autophagy mechanism.


Assuntos
Lisossomos , Análise Espectral Raman , Autofagia , Células Hep G2 , Humanos , Organelas
10.
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
11.
Nan Fang Yi Ke Da Xue Xue Bao ; 41(8): 1207-1213, 2021 Aug 20.
Artigo em Chinês | MEDLINE | ID: mdl-34549712

RESUMO

OBJECTIVE: To study the effect of lysosomal membrane protein Sidt2 deletion on autophagy in human hepatocytes. METHODS: Crispr-Cas9 technology was used to construct a human hepatocyte (HL7702) model of Sidt2 knockout (Sidt2-/-), and the expression levels of the key autophagy proteins LC3II/I, P62 and autophagy-related proteins Atg5, Atg7, and Atg12 were detected.The co-localization of LC3B and P62 in the cells were analyzed with immunofluorescence assay to assess the identification and storage of P62 cargo proteins by the autophagosomes and the degradation of the autophagolysosomes.The co-localization of LC3B and LAMP1 was also determined with immunofluorescence assay to detect the fusion of the autophagosomes with the lysosomes, and LysoTracker was used to trace the acidic lysosomes. RESULTS: We successfully constructed a HL7702 cell model of Sidt2+/+ and Sidt2-/-, and compared with Sidt2+/+ cells, the Sidt2-/- cell model showed significantly increased expressions of LC3-II/I and P62 (P < 0.01).Immunofluorescence assay showed a significant increase of LC3B and P62 expressions (P < 0.001) and obviously lowered expressions of Atg5, Atg7, and Atg12 in Sidt2-/- cells (P < 0.05).The co-localization of LC3B and P62 and that of LC3B and LAMP1 were both reduced and the number of acidic lysosomes was significantly lowered in Sidt2-/- cells (P < 0.05). CONCLUSION: Sidt2 gene deletion disturbs the recognition and sequestration of P62 cargo protein by autophagosomes in human hepatocytes.At the same time, the decreased number of acidic lysosomes and the dysfunction of autophagosome and lysosome fusion cause the block of the autophagy-lysosome pathway, leading eventually to LC3B and P62 accumulation and impaired autophagy in the hepatocytes.


Assuntos
Autofagia , Hepatócitos , Proteínas de Transporte de Nucleotídeos , Autofagossomos , Células Cultivadas , Técnicas de Inativação de Genes , Humanos , Glicoproteínas de Membrana Associadas ao Lisossomo/genética , Lisossomos
12.
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
13.
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
14.
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
15.
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
16.
Free Radic Biol Med ; 174: 225-235, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34407426

RESUMO

Oxidative stress has been implicated in the aging process and the progression of many neurodegenerative disorders. We previously reported that a novel oxindole compound, GIF-0726-r, effectively prevents endogenous oxidative stress, such as oxytosis/ferroptosis, an iron-dependent form of non-apoptotic cell death, in mouse hippocampal cells. In this study, using two hundred compounds that were developed based on the structure-activity relationship of GIF-0726-r, we screened for the most potent compounds that prevent glutamate- and erastin-induced oxytosis and ferroptosis. Using submicromolar concentrations, we identified nine neuroprotective compounds that have N,N-dimethylaniline as a common structure but no longer contain an oxindole ring. The most potent derivatives, GIF-2114 and GIF-2197-r (the racemate of GIF-2115 and GIF-2196), did not affect glutathione levels, had no antioxidant activity in vitro, or ability to activate the Nrf2 pathway, but prevented oxytosis/ferroptosis via reducing reactive oxygen production and decreasing ferrous ions. Furthermore, we developed fluorescent probes of GIF-2114 and GIF-2197-r to image their distribution in live cells and found that they preferentially accumulated in late endosomes/lysosomes, which play a central role in iron metabolism. These results suggest that GIF-2114 and GIF-2197-r protect hippocampal cells from oxytosis/ferroptosis by targeting late endosomes and lysosomes, as well as decreasing ferrous ions.


Assuntos
Ferroptose , Fármacos Neuroprotetores , Compostos de Anilina , Animais , Endossomos , Lisossomos , Camundongos , Fármacos Neuroprotetores/farmacologia
17.
Int J Mol Sci ; 22(15)2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34360836

RESUMO

Lysyl oxidase-like 3 (LOXL3), belonging to the lysyl oxidase family, is responsible for the crosslinking in collagen or elastin. The cellular localization of LOXL3 is in the extracellular space by reason of its canonical function. In tumors, the presence of LOXL3 has been associated with genomic stability, cell proliferation, and metastasis. In silico analysis has shown that glioblastoma was among tumors with the highest LOXL3 expression levels. LOXL3 silencing of U87MG cells by siRNA led to the spreading of the tumor cell surface, and the transcriptome analysis of these cells revealed an upregulation of genes coding for extracellular matrix, cell adhesion, and cytoskeleton components, convergent to an increase in cell adhesion and a decrease in cell invasion observed in functional assays. Significant correlations of LOXL3 expression with genes coding for tubulins were observed in the mesenchymal subtype in the TCGA RNA-seq dataset of glioblastoma (GBM). Conversely, genes involved in endocytosis and lysosome formation, along with MAPK-binding proteins related to focal adhesion turnover, were downregulated, which may corroborate the observed decrease in cell viability and increase in the rate of cell death. Invasiveness is a major determinant of the recurrence and poor outcome of GBM patients, and downregulation of LOXL3 may contribute to halting the tumor cell invasion.


Assuntos
Aminoácido Oxirredutases/metabolismo , Adesão Celular , Regulação Neoplásica da Expressão Gênica , Glioblastoma/enzimologia , Aminoácido Oxirredutases/genética , Aminoácido Oxirredutases/fisiologia , Linhagem Celular Tumoral , Proliferação de Células , Simulação por Computador , Citoesqueleto/metabolismo , Endocitose , Matriz Extracelular/metabolismo , Perfilação da Expressão Gênica , Glioblastoma/genética , Glioblastoma/patologia , Glioblastoma/fisiopatologia , Humanos , Lisossomos/fisiologia , Invasividade Neoplásica
18.
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
19.
Biomolecules ; 11(7)2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-34356615

RESUMO

The gut epithelial barrier provides the first line of defense protecting the internal milieu from the environment. To circumvent the exposure to constant challenges such as pathogenic infections and commensal bacteria, epithelial and immune cells at the gut barrier require rapid and efficient means to dynamically sense and respond to stimuli. Numerous studies have highlighted the importance of proteolysis in maintaining homeostasis and adapting to the dynamic changes of the conditions in the gut environment. Primarily, proteolytic activities that are involved in immune regulation and inflammation have been examined in the context of the lysosome and inflammasome activation. Yet, the key to cellular and tissue proteostasis is the ubiquitin-proteasome system, which tightly regulates fundamental aspects of inflammatory signaling and protein quality control to provide rapid responses and protect from the accumulation of proteotoxic damage. In this review, we discuss proteasome-dependent regulation of the gut and highlight the pathophysiological consequences of the disarray of proteasomal control in the gut, in the context of aberrant inflammatory disorders and tumorigenesis.


Assuntos
Mucosa Intestinal , Complexo de Endopeptidases do Proteassoma , Proteólise , Transdução de Sinais/imunologia , Animais , Ativação Enzimática/imunologia , Humanos , Inflamação/enzimologia , Mucosa Intestinal/enzimologia , Mucosa Intestinal/imunologia , Lisossomos/enzimologia , Lisossomos/imunologia , Complexo de Endopeptidases do Proteassoma/imunologia , Complexo de Endopeptidases do Proteassoma/metabolismo
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