Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 29
Filtrar
1.
Mol Cell Neurosci ; 131: 103972, 2024 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-39349151

RESUMO

Gut microbe-derived short-chain fatty acids (SCFAs) are known to have a profound impact on various brain functions, including cognition, mood, and overall neurological health. However, their role, if any, in protecting against hypoxic injury and ischemic stroke has not been extensively studied. In this study, we investigated the effects of two major SCFAs abundant in the gut, propionate (P) and butyrate (B), on hypoxia-reperfusion injury using a neuronal cell line and a zebrafish model. Neuro 2a (N2a) cells treated with P and B exhibited reduced levels of mitochondrial and cytosolic reactive oxygen species (ROS), diminished loss of mitochondrial membrane potential, suppressed caspase activation, and lower rates of cell death when exposed to CoCl2, a chemical commonly used to simulate hypoxia. Furthermore, adult zebrafish fed SCFA-supplemented feeds showed less susceptibility to hypoxic conditions compared to the control group, as indicated by multiple behavioral measures. Histological analysis of 2,3,5-Triphenyltetrazolium chloride (TTC) stained brain sections revealed less damage in the SCFA-fed group. We also found that Fatty Acid Binding Protein 7 (FABP7), also known as Brain Lipid Binding Protein (BLBP), a neuroprotective fatty acid binding protein, was upregulated in the brains of the SCFA-fed group. Additionally, when FABP7 was overexpressed in N2a cells, it protected the cells from injury caused by CoCl2 treatment. Overall, our data provide evidence for a neuroprotective role of P and B against hypoxic brain injury and suggest the potential of dietary supplementation with SCFAs to mitigate stroke-induced brain damage.

2.
Biochem Biophys Res Commun ; 664: 20-26, 2023 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-37130457

RESUMO

The cellular implications of the interaction between Pannexin-1 (Panx1) channel and P2X7 receptor (P2X7R) have not been fully elucidated. Evidence suggests that ATP, released through Panx1, activates P2X7R, which in turn promotes further activation of Panx1. In a previous study, we reported that the C-terminus of Panx1 (Panx1-CT) attenuates P2X7R-mediated Ca2+ influx and cell death. One of the distinctive features of P2X7R is the gradual increase in current with repetitive stimulation. In the current study, we report an effect of Panx1-CT (amino acid residues 350 to 426) on P2X7R current, which differs from the effect of full-length Panx1. Panx1-CT inhibited P2X7R current, which persisted in all consecutive agonist applications. However, full-length Panx1 reduced P2X7R current at initial stimulations, followed by gradual augmentation. When P2X7R was activated for an extended period, cells expressing Panx1-CT exhibited less mitochondrial depolarization, reactive oxygen species (ROS) generation, Caspase 3 activation and cell death, whereas cells overexpressing full-length Panx1 showed the opposite effect. Taken together, these findings suggest that Panx1 can either attenuate or augment P2X7R-mediated cellular processes depending on the degree of P2X7R activation.


Assuntos
Conexinas , Receptores Purinérgicos P2X7 , Receptores Purinérgicos P2X7/genética , Receptores Purinérgicos P2X7/metabolismo , Conexinas/genética , Conexinas/metabolismo , Transdução de Sinais , Morte Celular , Trifosfato de Adenosina/metabolismo
3.
Biochem Biophys Res Commun ; 548: 143-147, 2021 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-33640607

RESUMO

Pannexin 1 (Panx1) has been implicated in a plethora of physiological and pathophysiological processes. It is one of the major ATP release channels in many cell types. Extracellular ATP, activates purinergic P2X and P2Y receptors, triggering several signaling cascades. A disease-associated mutation, Arg-217-His (R217H) in the 3rd transmembrane domain of Panx1 attenuates channel functions through an unknown mechanism. Since carboxyl terminus (CT) gates the channel, we hypothesized that R217 interacts with the CT, and this interaction is required for optimum channel activities. R217H mutation though reduced the currents in the full-length channel, did not affect CT-truncated Panx1-Δ386. Also, compared to the wild-type, Panx1-R217H expressing cells showed lesser cell death when activated through P2X7 receptor. However, cell death in Panx1-R217H-Δ386 and Panx1-Δ386 expressing cells were similar. The mutation is ineffective unless the channel has an intact CT. Based on our results we propose that R217H mutation perturbs the conformational flexibility of CT, leading to channel dysfunction.


Assuntos
Conexinas/química , Conexinas/genética , Mutação/genética , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Sequência de Aminoácidos , Morte Celular , Conexinas/metabolismo , Regulação da Expressão Gênica , Células HEK293 , Humanos , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Receptores Purinérgicos P2X7/genética , Receptores Purinérgicos P2X7/metabolismo
4.
Biochem Biophys Res Commun ; 527(1): 71-75, 2020 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-32446393

RESUMO

Acid-sensing ion channels (ASICs) have been implicated in many physiological and patho-physiological processes like synaptic plasticity, inflammation, pain perception, stroke-induced brain damage and, drug-seeking behaviour. Although ASICs have been shown to be modulated by gasotransmitters like nitric oxide (NO), their regulation by hydrogen sulfide (H2S) is not known. Here, we present strong evidence that H2S potentiates ASICs-mediated currents. Low pH-induced current in Chinese hamster ovary (CHO) cells, expressing homomeric either ASIC1a, ASIC2a or ASIC3, increased significantly by an H2S donor NaHS. The effect was reversed by washing the cells with NaHS-free external solution of pH 7.4. MTSES, a membrane impermeable cysteine thiol-modifier failed to abrogate the effect of NaHS on ASIC1a, suggesting that the target cysteine residues are not in the extracellular region of the channel. The effect of NaHS is not mediated through NO, as the basal NO level in cells did not change following NaHS application. This previously unknown mechanism of ASICs-modulation by H2S adds a new dimension to the ASICs in health and disease.


Assuntos
Canais Iônicos Sensíveis a Ácido/metabolismo , Sulfeto de Hidrogênio/farmacologia , Animais , Células CHO , Cricetulus , Concentração de Íons de Hidrogênio , Óxido Nítrico/metabolismo , Técnicas de Patch-Clamp , Agonistas de Canais de Sódio/farmacologia
5.
J Cell Sci ; 130(22): 3818-3828, 2017 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-28972132

RESUMO

Leucine-rich repeat-containing 8 (LRRC8) proteins have been proposed to evolutionarily originate from the combination of the channel protein pannexin, and a leucine-rich repeat (LRR) domain. Five paralogs of LRRC8, namely LRRC8A, LRRC8B, LRRC8C, LRRC8D and LRRC8E have been reported. LRRC8A has been shown to be instrumental in cell swelling. Here, we identify LRRC8B as a key player in the cellular Ca2+ signaling network. Overexpression of human LRRC8B in HEK293 cells reduced the Ca2+ level in the endoplasmic reticulum (ER). LRRC8B-overexpressing cells exhibited a lesser release of Ca2+ from the ER in response to ATP, carbachol and intracellular administration of inositol (1,4,5)-trisphosphate (IP3). LRRC8B-knockdown cells showed a slower depletion of the ER Ca2+ stores when sarco-endoplasmic reticulum Ca2+-ATPase was blocked with thapsigargin (TG), while overexpression of LRRC8B had the opposite effect. LRRC8B-overexpressing cells exhibited a higher level of store-operated Ca2+ entry following store-depletion by TG. Collectively, LRRC8B participates in intracellular Ca2+ homeostasis by acting as a leak channel in the ER. This study gives a fundamental understanding of the role of a novel protein in the elemental cellular process of ER Ca2+ leak and expands the known roles for LRRC8 proteins.This article has an associated First Person interview with the first author of the paper.


Assuntos
Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/metabolismo , Trifosfato de Adenosina/fisiologia , Sinalização do Cálcio , Células HEK293 , Humanos , Transporte Proteico
6.
Planta Med ; 85(17): 1316-1325, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31618777

RESUMO

Alzheimer's disease is a neurodegenerative disease that leads to irreversible neuronal damage. Senile plaques, composed of amyloid beta peptide, is the principal abnormal characteristic of the disease. Among the factors involved, the secretase enzymes, namely, α secretase, beta-site amyloid precursor protein-cleaving enzyme, ß secretase, and γ secretase, hold consequential importance. Beta-site amyloid precursor protein-cleaving enzyme 1 is considered to be the rate-limiting factor in the production of amyloid beta peptide. Research supporting the concept of inhibition of beta-site amyloid precursor protein-cleaving enzyme activity as one of the effective therapeutic targets in the mitigation of Alzheimer's disease is well accepted. The identification of natural compounds, such as ß-amyloid precursor protein-selective beta-site amyloid precursor protein-cleaving enzyme inhibitors, and the idea of compartmentalisation of the beta-site amyloid precursor protein-cleaving enzyme 1 action have caused a dire need to closely examine the natural compounds and their effectiveness in the disease mitigation. Many natural compounds have been reported to effectively modulate beta-site amyloid precursor protein-cleaving enzyme 1. At lower doses, compounds like 2,2',4'-trihydroxychalcone acid, quercetin, and myricetin have been shown to effectively reduce beta-site amyloid precursor protein-cleaving enzyme 1 activity. The currently used five drugs that are marketed and used for the management of Alzheimer's disease have an increased risk of toxicity and restricted therapeutic efficiency, hence, the search for new anti-Alzheimer's disease drugs is of primary concern. A variety of natural compounds having pure pharmacological moieties showing multitargeting activity and others exhibiting specific beta-site amyloid precursor protein-cleaving enzyme 1 inhibition as discussed below have superior biosafety. Many of these compounds, which are isolated from medicinal herbs and marine flora, have been long used for the treatment of various ailments since ancient times in the Chinese and Ayurvedic medical systems. The aim of this article is to review the available data on the selected natural compounds, giving emphasis to the inhibition of beta-site amyloid precursor protein-cleaving enzyme 1 activity as a mode of Alzheimer's disease treatment.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Produtos Biológicos/uso terapêutico , Alcaloides/uso terapêutico , Flavonoides/uso terapêutico , Humanos , Fenóis/uso terapêutico , Fitoterapia
7.
Biochemistry ; 55(49): 6832-6847, 2016 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-27951646

RESUMO

Since its discovery, neuroglobin (Ngb), a neuron-specific oxygen binding hemoglobin, distinct from the classical myoglobin and blood hemoglobin, has attracted attention as an endogenous neuroprotectant. Recent reports suggest that Ngb protects neurons from brain stroke, ischemic stress-induced degeneration, and other brain disorders. Proteins with a specific role in neuroprotection are often associated with neurodegeneration, as well, depending on the cellular environment or specific cellular triggers that tilt the balance one way or the other. This investigation explored the potential role of Ngb in amyloid fibril-related neuronal disorder. Ngb was capable of amyloid formation in vitro at neutral pH and ambient temperature, in both apo and holo forms, albeit at a slower rate in the holo form, unlike other hemoglobins that exhibit such behavior exclusively in the apo states. Elevated temperature enhanced the rate of fibril formation significantly. The B-helix, which is known to play a major role in Ngb ligand binding kinetics, was found to be amyloidogenic with the Phe28B10 amino acid side chain as the key inducer of fibrillation. The Ngb amyloid fibril was also significantly cytotoxic to neuroblastoma cell lines, compared to those obtained from reference hemoglobins. The Ngb fibril probably promoted toxicity by inducing channel formation in the cell membrane, as investigated here using synthetic lipid bilayer membranes and the propidium iodide uptake assay. These findings imply that Ngb plays a role in neurodegenerative disorders in vivo, for which there seems to be indirect evidence by association. Ngb thus presents a novel prospect for understanding amyloid-related brain disorders beyond the limited set of proteins currently investigated for such diseases.


Assuntos
Amiloide/química , Encéfalo/metabolismo , Globinas/química , Hemoglobinas/química , Proteínas do Tecido Nervoso/química , Fenilalanina/química , Linhagem Celular Tumoral , Dicroísmo Circular , Globinas/genética , Humanos , Concentração de Íons de Hidrogênio , Microscopia Eletrônica de Transmissão , Mutagênese Sítio-Dirigida , Proteínas do Tecido Nervoso/genética , Neuroglobina , Temperatura
8.
Biochim Biophys Acta ; 1848(1 Pt A): 151-8, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25459681

RESUMO

Voltage dependent anion channel (VDAC) of mitochondria plays a crucial role in apoptosis. Human VDAC-1, reconstituted in planar lipid bilayer showed reduced conductance when treated with curcumin. Curcumin interacts with residues in the α helical N-terminus of VDAC and in the channel wall, as revealed by molecular docking, followed by mutational analysis. N-terminus mimicking peptide showed conformational changes in circular dichroism, upon curcumin treatment. We propose that the interaction of curcumin with amino acids in N-terminus and in channel wall fixes the α helix in closed conformation. This restricts its movement which is required for the opening of the channel.


Assuntos
Curcumina/farmacologia , Mitocôndrias/efeitos dos fármacos , Proteínas Mitocondriais/química , Canal de Ânion 1 Dependente de Voltagem/química , Animais , Sítios de Ligação/genética , Western Blotting , Bovinos , Dicroísmo Circular , Curcumina/química , Curcumina/metabolismo , Relação Dose-Resposta a Droga , Humanos , Ativação do Canal Iônico/efeitos dos fármacos , Ativação do Canal Iônico/genética , Ativação do Canal Iônico/fisiologia , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Simulação de Acoplamento Molecular , Estrutura Molecular , Mutação , Ligação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Espectrofotometria , Canal de Ânion 1 Dependente de Voltagem/genética , Canal de Ânion 1 Dependente de Voltagem/metabolismo
9.
Biochem Biophys Res Commun ; 477(3): 490-4, 2016 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-27318085

RESUMO

Dietary trans fatty acids (TFAs) are known to increase the risk of cardiovascular diseases by altering plasma lipid profile and activating various inflammatory signaling pathways. Here we show that elaidic acid (EA), the most abundant TFA in diet, alters the electrophysiological properties of voltage-dependent anion channel (VDAC) of mitochondria. Purified bovine brain VDAC, when incorporated in the planar lipid bilayer (PLB) composed of 1,2-diphytanoyl-sn-glycero-3 phosphatidyl choline (DPhPC) and EA in a 9 to 1 ratio (wt/wt), exhibited complete closing events at different voltages. The closing events were observed at even -10 mV, a voltage at which VDAC usually remains fully open all the time. Additionally, the voltage sensitivity of VDAC was lost in presence of EA; the channel conductance did not decrease with increasing voltages. In identical experimental conditions, membrane containing oleic acid (OA), the cis isomer of EA did not produce any such effect. We propose that EA possibly exerts its adverse effect by modulating VDAC.


Assuntos
Mitocôndrias/fisiologia , Ácido Oleico/farmacologia , Canais de Ânion Dependentes de Voltagem/efeitos dos fármacos , Animais , Mitocôndrias/efeitos dos fármacos , Ácidos Oleicos , Canais de Ânion Dependentes de Voltagem/fisiologia
10.
Angiogenesis ; 18(2): 175-89, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25510468

RESUMO

Angiogenesis, the formation of new blood vessels from pre-existing vessels, is a complex process that warrants cell migration, proliferation, tip cell formation, ring formation, and finally tube formation. Angiogenesis is initiated by a single leader endothelial cell called "tip cell," followed by vessel elongation by "stalk cells." Tip cells are characterized by their long filopodial extensions and expression of vascular endothelial growth factor receptor-2 and endocan. Although nitric oxide (NO) is an important modulator of angiogenesis, its role in angiogenic sprouting and specifically in tip cell formation is poorly understood. The present study tested the role of endothelial nitric oxide synthase (eNOS)/NO/cyclic GMP (cGMP) signaling in tip cell formation. In primary endothelial cell culture, about 40% of the tip cells showed characteristic sub-cellular localization of eNOS toward the anterior progressive end of the tip cells, and eNOS became phosphorylated at serine 1177. Loss of eNOS suppressed tip cell formation. Live cell NO imaging demonstrated approximately 35% more NO in tip cells compared with stalk cells. Tip cells showed increased level of cGMP relative to stalk cells. Further, the dissection of NO downstream signaling using pharmacological inhibitors and inducers indicates that NO uses the sGC/cGMP pathway in tip cells to lead angiogenesis. Taken together, the present study confirms that eNOS/NO/cGMP signaling defines the direction of tip cell migration and thereby initiates new blood vessel formation.


Assuntos
Óxido Nítrico/fisiologia , Animais , Bovinos , Linhagem Celular Transformada , Galinhas , GMP Cíclico/metabolismo , Humanos , Neovascularização Fisiológica/fisiologia , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Fosforilação , RNA Interferente Pequeno/genética , Regulação para Cima
12.
Nitric Oxide ; 47: 77-84, 2015 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-25917852

RESUMO

Nitric oxide (NO), a major gaseous signaling molecule, modulates several ion channels and receptors. Here we show that NO attenuates pannexin 1 (Panx1) mediated currents in HEK-293 cells. NO exerts its effect by activating a cGMP-protein kinase G (PKG) dependent pathway. NO donors, sodium nitroprusside (SNP), S-nitroso-N-acetylpenicillamine (SNAP) and S-nitrosoglutathione (GSNO), reduced Panx1 currents by 25-41%. 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), an inhibitor of soluble guanylyl cyclase (sGC), blocked the inhibition completely, whereas sGC activator YC-1 mimicked the effect of NO, suggesting the involvement of a cGMP dependent pathway. Supporting this, NO had no effect in the presence of the PKG inhibitor, KT5823. Further, immuno-precipitated Panx1 was recognized by an anti-phosphoserine antibody in Western blot. Phosphorylation was enhanced significantly when cells were treated with SNP. The target for phosphorylation is possibly Ser 206 of Panx1, as its mutation to Ala completely abolished the NO mediated inhibition.


Assuntos
Conexinas/antagonistas & inibidores , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Óxido Nítrico/farmacologia , Conexinas/metabolismo , Células HEK293 , Humanos , Proteínas do Tecido Nervoso/metabolismo , Óxido Nítrico/metabolismo
13.
Cell Biochem Funct ; 30(2): 89-100, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22392438

RESUMO

Hemichannels, which are one half of the gap junction channels, have independent physiological roles. Although hemichannels consisting of connexins are more widely documented, hemichannels of pannexins, proteins homologous to invertebrate gap junction proteins also have been studied. There are at least 21 different connexin and three pannexin isotypes. This variety in isotypes results in tissue-specific hemichannels, which have been implicated in varied events ranging from development, cell survival, to cell death. Hemichannel function varies with its spatio-temporal opening, thus demanding a refined degree of regulation. This review discusses the activity of hemichannels and the molecules released in different physiological states and their impact on tissue functioning.


Assuntos
Conexinas/metabolismo , Junções Comunicantes/metabolismo , Crescimento e Desenvolvimento , Animais , Morte Celular , Humanos
14.
Life Sci ; 291: 120268, 2022 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-34973275

RESUMO

AIMS: High-mobility group (HMG) proteins are oncogenic in different cancers, including cervical cancer; silencing their individual expression using sh-RNAs, siRNAs, and miRNAs has had anti-tumorigenic effects, but the consequences of their collective downregulation are not known. Since multiple gene targeting is generally very effective in cancer therapy, the present study highlighted the consequences of silencing the expression of HMGA1, A2, B1, and B3 using sh-RNAs or miR-142-3p (that can potentially target HMGA1, A2, B1, and B3) in cervical cancer cell lines. MAIN METHODS: 3' UTR luciferase reporter assays were performed to validate HMGA1, A2, B1, and B3 as targets of miR-142-3p in human cervical cancer cells. Annexin V/PI dual staining and flow cytometry analyses were used to detect apoptotic cells. miR-142-3p-mediated regulation of cell death, colony formation, migration, and invasion was investigated in human cervical cancer cells together with in vivo metastasis in zebrafish. KEY FINDINGS: Concurrent knockdown of HMGA1, A2, B1, and B3 through their corresponding sh-RNAs inhibited cell viability and colony formation but induced apoptosis, and these effects were relatively reduced upon their individual knockdown. miR-142-3p targeted HMGA1, A2, B1, and B3 by binding to their 3'UTRs and induced apoptosis but inhibited proliferation, migration, and invasion of human cervical cancer cells. In addition, miR-142-3p expression decreased phospho-p65 and EMT-related proteins in cervical cancer cells and their in vivo metastatic potential upon implantation in zebrafish. SIGNIFICANCE: These findings suggest that miR-142-3p acts as a tumor-suppressive miRNA by targeting HMGA1, A2, B1, and B3 and may serve as a potential therapeutic agent in human cervical cancer.


Assuntos
MicroRNAs/genética , Neoplasias do Colo do Útero/metabolismo , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Feminino , Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Proteína HMGA1a/genética , Proteína HMGA1a/metabolismo , Proteína HMGA2/genética , Proteína HMGA2/metabolismo , Proteína HMGB1/genética , Proteína HMGB1/metabolismo , Proteína HMGB3/genética , Proteína HMGB3/metabolismo , Células HeLa , Humanos , MicroRNAs/metabolismo , Modelos Animais , Invasividade Neoplásica/genética , Oncogenes , Neoplasias do Colo do Útero/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-Zebra
15.
Life Sci ; 304: 120727, 2022 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-35753437

RESUMO

AIMS: High mobility group box (HMGB) family proteins, HMGB1, HMGB2, HMGB3, and HMGB4 are oncogenic. The oncogenic nature of HMGB1 is characterized by its association with autophagy, ROS, and MMP. Since HMGB3 is its paralog, we hypothesized that it might also modulate autophagy, ROS, and MMP. Hence, we targeted HMGB3 using its shRNA or miR-142-3p and assessed the changes in autophagy, ROS, MMP, and tumorigenic properties of human breast cancer cells. MAIN METHODS: Cell viability was assessed by resazurin staining and annexin-V/PI dual staining was used for confirming apoptosis. Colony formation, transwell migration, invasion and luciferase reporter (for miRNA-target validation) assays were also performed. ROS and MMP were detected using DHE and MitoTracker dyes, respectively. A zebrafish xenograft model was used to assess the role of miR-142-3p on in vivo metastatic potential of breast cancer cells. KEY FINDINGS: Breast cancer tissues from Indian patients and TCGA samples exhibit overexpression of HMGB3. miR-142-3p binds to 3' UTR of HMGB3, leading to its downregulation that subsequently inhibits colony formation and induces apoptosis involving increased ROS accumulation and decreased MMP, phospho-mTOR and STAT3. Our findings show that HMGB3 is directly involved in the miR-142-3p-mediated disruption of autophagy and induction of apoptotic cell death via modulation of LC3, cleaved PARP and Bcl-xL. In addition, miR-142-3p inhibited migration, invasion and metastatic potential of breast cancer cells. SIGNIFICANCE: Our findings highlighted the role of HMGB3, for the first time, in the modulation of autophagy and apoptosis in human breast cancer cells, and these results have therapeutic implications.


Assuntos
Neoplasias da Mama , Proteína HMGB1 , Proteína HMGB3 , MicroRNAs , Regiões 3' não Traduzidas , Animais , Apoptose/genética , Autofagia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/genética , Feminino , Proteína HMGB1/genética , Proteína HMGB3/genética , Proteína HMGB3/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio , Peixe-Zebra/genética
16.
Front Mol Biosci ; 9: 1050768, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36387285

RESUMO

Many neurodegenerative diseases, such as Alzheimer's disease (AD) and frontotemporal dementia with Parkinsonism linked to chromosome 17, are characterized by tau pathology. Numerous motor proteins, many of which are involved in synaptic transmission, mediate transport in neurons. Dysfunction in motor protein-mediated neuronal transport mechanisms occurs in several neurodegenerative disorders but remains understudied in AD. Kinesins are the most important molecular motor proteins required for microtubule-dependent transport in neurons, and kinesin-1 is crucial for neuronal transport among all kinesins. Although kinesin-1 is required for normal neuronal functions, the dysfunction of these motor domains leading to neurodegenerative diseases is not fully understood. Here, we reported that the kinesin-I heavy chain (KIF5B), a key molecular motor protein, is involved in tau homeostasis in AD cells and animal models. We found that the levels of KIF5B in P301S tau mice are high. We also found that the knockdown and knockout (KO) of KIFf5B significantly decreased the tau stability, and overexpression of KIF5B in KIF5B-KO cells significantly increased the expression of phosphorylated and total tau levels. This suggested that KIF5B might prevent tau accumulation. By conducting experiments on P301S tau mice, we showed that partially reducing KIF5B levels can reduce hyperphosphorylation of the human tau protein, formation of insoluble aggregates, and memory impairment. Collectively, our results suggested that decreasing KIF5B levels is sufficient to prevent and/or slow down abnormal tau behavior of AD and other tauopathies.

17.
Cell Calcium ; 99: 102458, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34479067

RESUMO

Extracellular ATP works as an autocrine and/or paracrine signaling molecule by activating plasma membrane-localized purinergic receptors. Stimulation of purinergic P2X7 receptor (P2X7R) increases cytosolic Ca2+ ([Ca2+]c), which in turn activates Pannexin 1 (Panx1) channel. In earlier studies, Panx1 and P2X7R have been shown to interact physically. Also, both the channels have been implicated in similar pathophysiological processes. In this study, we investigated the effect of Panx1 on P2X7R-mediated Ca2+influx. Panx1 attenuated P2X7R-mediated [Ca2+]c rise in CHO-K1 and HEK-293 cells. [Ca2+]c rise was higher in Panx1 knockdown astrocytes. The inhibitory effect was unaffected in the presence of Panx1 blocker, carbenoxolone. The region between 350th and 386th amino acid residues in the carboxyl terminus (CT) of Panx1 was found to be crucial for inhibiting P2X7R. Like full-length Panx1, the CT (350th to 426th amino acids) alone was able to attenuate the [Ca2+]c rise. Further, CT prevented cell death caused by P2X7R overactivation. Based on our results, we propose a novel pro-survival role of Panx1 exerted by modulating P2X7R.


Assuntos
Conexinas , Receptores Purinérgicos P2X7 , Trifosfato de Adenosina , Morte Celular , Conexinas/genética , Conexinas/metabolismo , Células HEK293 , Humanos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo
18.
Cancer Lett ; 522: 211-224, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34571083

RESUMO

Breast cancer cells evade cell death by overexpressing SLC7A11, which functions by transporting cystine into cells in exchange for intracellular glutamate facilitating glutathione synthesis and reducing reactive oxygen species (ROS)-mediated stress. Using an in silico approach, we predicted an miRNA (miR-5096) that can target and downregulate SLC7A11. We demonstrated SLC7A11 as a target of miR-5096 by 3'UTR luciferase assay and further validated it by identifying reduced mRNA and protein levels of SLC7A11 upon miR-5096 overexpression. miR-5096-induced ferroptotic cell death in human breast cancer cells was confirmed by concurrently increased ROS, OH-, lipid ROS, and iron accumulation levels and decreased GSH and mitochondrial membrane potential (MitoTracker™ Orange) with mitochondrial shrinkage and partial cristae loss (observed by TEM). miR-5096 inhibited colony formation, transwell migration, and breast cancer cell invasion, whereas antimiR-5096 promoted these tumorigenic properties. Ectopic expression of SLC7A11 partly reversed miR-5096-mediated effects on cell survival, ROS, lipid peroxides, iron accumulation, GSH, hydroxyl radicals, mitochondrial membrane potential, and colony formation. miR-5096 modulated the expression of epithelial-mesenchymal transition markers in vitro and inhibited the metastatic potential of MDA-MB-231 cells in a tumor xenograft model of zebrafish larvae. Our results demonstrate that miR-5096 is a tumor-suppressive miRNA in breast cancer cells, and this paper discusses its therapeutic implications.


Assuntos
Sistema y+ de Transporte de Aminoácidos/genética , Neoplasias da Mama/genética , Carcinogênese/genética , MicroRNAs/genética , Animais , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Ferroptose/genética , Regulação Neoplásica da Expressão Gênica/genética , Glutationa/metabolismo , Xenoenxertos , Humanos , Peroxidação de Lipídeos/genética , Potencial da Membrana Mitocondrial , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Peixe-Zebra
19.
Nitric Oxide ; 22(3): 213-9, 2010 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-20045740

RESUMO

Acid sensing ion channels (ASICs) are widely expressed in central and peripheral nervous system. They are involved in a variety of physiological and pathophysiological processes: synaptic transmission, learning and memory, pain perception, ischemia, etc. During ischemia, metabolic acidosis causes the drop of extracellular pH (pHe) which in turn activates ASICs. Activation of calcium permeable ASIC1a has been implicated in neuronal death. ASICs are modulated by several redox reagents, divalent cations and nitric oxide (NO). Although NO potentiates ASIC mediated currents, the physiological significance of such modulation has not been studied in detail. We have evaluated the role of endogenous NO in cell death at different pH, mediated by the activation of ASICs. At pH 6.1, death rates of ASIC1 expressing Neuro2A (N2A) cells are significantly higher in comparison to the cells that do not express ASICs. Amiloride, a blocker of ASICs protects the cell from acid-injury. Sodium nitroprusside, a potent NO donor not only increases the ASIC mediated currents but also increases cell death at low pH. L-Arg, the precursor of NO also potentiates ASICs in a pH dependent manner. L-Arg-induced NO production and potentiation of ASICs were observed at pHs 7.4, 7.2, 7.0 and 6.8. Lowering the pH below 6.8 did not result in significant production of NO or potentiation of ASICs upon L-Arg stimulation. Our results suggest that potentiation of ASICs by NO and subsequent cell death in vivo depends on the severity of acidosis. During mild and moderate acidosis, NO promotes cell death by potentiating ASICs, whereas this potentiation subsides in severe acidosis due to inhibition of NO synthase.


Assuntos
Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Óxido Nítrico/metabolismo , Canais de Sódio/metabolismo , Canais Iônicos Sensíveis a Ácido , Ácidos/farmacologia , Animais , Células CHO , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cricetinae , Cricetulus , Células HeLa , Humanos , Concentração de Íons de Hidrogênio , Neurônios/efeitos dos fármacos , Óxido Nítrico/análise , Doadores de Óxido Nítrico/farmacologia , Nitroprussiato/farmacologia , Ratos , Ratos Wistar
20.
Colloids Surf B Biointerfaces ; 176: 360-370, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30658284

RESUMO

Two-dimensional (2D) tungsten disulfide (WS2) quantum dots offer numerous promising applications in materials and optoelectronic sciences. Additionally, the catalytic and photoluminescence properties of ultra-small WS2 nanoparticles are of potential interest in biomedical sciences. Addressing the use of WS2 in the context of infection, the present study describes the conjugation of two potent antimicrobial peptides with WS2 quantum dots, as well as the application of the resulting conjugates in antimicrobial therapy and bioimaging. In doing so, we determined the three-dimensional solution structure of the quantum dot-conjugated antimicrobial peptide by a series of high-resolution nuclear magnetic resonance (NMR) techniques, correlating this to the disruption of both model lipid and bacterial membranes, and to several key biological performances, including antimicrobial and anti-biofilm effects, as well as cell toxicity. The results demonstrate that particle conjugation enhances the antimicrobial and anti-biofilm potency of these peptides, effects inferred to be due to multi-dendate interactions for the conjugated peptides. As such, our study provides information on the mode-of-action of such conjugates, laying the foundation for their potential use in treatment and monitoring of infections.


Assuntos
Anti-Infecciosos/farmacologia , Diagnóstico por Imagem , Dissulfetos/química , Peptídeos/química , Pontos Quânticos/química , Tungstênio/química , Sequência de Aminoácidos , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Candida albicans/ultraestrutura , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/ultraestrutura
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA