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1.
Sci Rep ; 14(1): 13625, 2024 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-38871787

RESUMO

Currently, the increasing pollution of the environment by heavy metals is observed, caused both by natural factors and those related to human activity. They pose a significant threat to human health and life. It is therefore important to find an effective way of protecting organisms from their adverse effects. One potential product showing a protective effect is green tea. It has been shown that EGCG, which is found in large amounts in green tea, has strong antioxidant properties and can therefore protect cells from the adverse effects of heavy metals. Therefore, the aim of the study was to investigate the effect of EGCG on cells exposed to Cd. In the study, CHO-K1 cells (Chinese hamster ovary cell line) were treated for 24 h with Cd (5 and 10 µM) and EGCG (0.5 and 1 µM) together or separately. Cell viability, ATP content, total ROS activity, mitochondrial membrane potential and apoptosis potential were determined. The results showed that, in tested concentrations, EGCG enhanced the negative effect of Cd. Further analyses are needed to determine the exact mechanism of action of EGCG due to the small number of publications on the subject and the differences in the results obtained in the research.


Assuntos
Apoptose , Cádmio , Catequina , Sobrevivência Celular , Cricetulus , Potencial da Membrana Mitocondrial , Estresse Oxidativo , Espécies Reativas de Oxigênio , Catequina/análogos & derivados , Catequina/farmacologia , Animais , Células CHO , Apoptose/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Cádmio/toxicidade , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Antioxidantes/farmacologia , Cricetinae , Trifosfato de Adenosina/metabolismo
2.
Exp Oncol ; 46(1): 38-44, 2024 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-38852054

RESUMO

BACKGROUND: Paclitaxel is a highly effective chemotherapeutic agent used to treat breast, ovarian, and other cancers. At the same time, paclitaxel causes peripheral neuropathy as a side effect in 45%-70% of patients. AIM: The aim of the study was to investigate the effect of paclitaxel-induced peripheral neuropathy on the development of pathological changes in the salivary glands of animals and to explore the possibility of correction of the identified changes with vitamin B/ATP complex. MATERIALS AND METHODS: To simulate toxic neuropathy, animals were injected i/p with paclitaxel 2 mg/kg for 4 days. In order to correct the identified changes, rats were injected i/m with vitamin B/ATP complex (1 mg/ kg) for 9 days. In the homogenate of the submandibular salivary glands, α-amylase activity, total proteolytic activity, total antitryptic activity, the content of medium mass molecules, thiobarbituric acid reactive substances (TBARS), oxidatively modified proteins, and catalase activity were determined. RESULTS: A significant increase in the content of oxidatively modified proteins, medium mass molecules, and the content of TBARS and significant decrease in the activity of catalase and amylase were determined in the salivary glands of animals with toxic neuropathy compared to these parameters in intact animals. Administration of vitamin B/ATP complex for 9 days against the background of paclitaxel-induced neuropathy led to normalization of antitryptic activity and amylase activity, a significant decrease in the content of oxidatively modified proteins, medium mass molecules, and TBARS along with a significant increase in catalase activity in the salivary glands of animals compared to the untreated rats with neuropathy. CONCLUSION: Paclitaxel-induced neuropathy caused the development of pathological changes in the salivary glands of rats, which was evidenced by a carbonyl- oxidative stress and impaired protein synthetic function. The correction with vitamin B/ATP complex restored the protein-synthetic function and the proteinase-inhibitor balance, suppressed the oxidative stress and normalized free radical processes in the salivary glands of rats.


Assuntos
Paclitaxel , Doenças do Sistema Nervoso Periférico , Glândulas Salivares , Animais , Paclitaxel/efeitos adversos , Paclitaxel/farmacologia , Glândulas Salivares/efeitos dos fármacos , Glândulas Salivares/patologia , Glândulas Salivares/metabolismo , Ratos , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/patologia , Antineoplásicos Fitogênicos/efeitos adversos , Antineoplásicos Fitogênicos/farmacologia , Ratos Wistar , Estresse Oxidativo/efeitos dos fármacos , Trifosfato de Adenosina/metabolismo , Complexo Vitamínico B/farmacologia , Complexo Vitamínico B/uso terapêutico , Masculino , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Catalase/metabolismo
3.
Structure ; 32(6): 647-649, 2024 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-38848680

RESUMO

In this issue of Structure, Yin et al.1 present the CryoEM structure of the HisRS-like domain of human GCN2 and demonstrate that it is a pseudoenzyme, which binds uncharged tRNA in a different manner than HisRS and does not bind histidine and ATP.


Assuntos
Trifosfato de Adenosina , Humanos , Trifosfato de Adenosina/metabolismo , Trifosfato de Adenosina/química , Microscopia Crioeletrônica , RNA de Transferência/metabolismo , RNA de Transferência/química , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/química , Catálise , Modelos Moleculares , Histidina/química , Histidina/metabolismo
4.
Can Vet J ; 65(6): 553-558, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38827592

RESUMO

Gastrointestinal stromal tumors arising from gastric cardia are uncommon in dogs. A few studies have shown the effectiveness of tyrosine kinase inhibitors in the treatment of canine gastrointestinal stromal tumors, but no standardized protocols are currently available. An 11-year-old spayed female Maltese dog was diagnosed with a gastrointestinal stromal tumor using histopathological and immunohistochemical analyses. An adenosine triphosphate-based tumor chemosensitivity assay revealed that imatinib at lower concentrations had a stronger inhibitory effect than toceranib. Based on the results of the assay, the dog was treated with imatinib after surgery. After 28 mo of therapy, there was no recurrence of the tumor. Key clinical message: Adenosine triphosphate-based tumor chemosensitivity assays may help clinicians to select appropriate postoperative chemotherapeutic drugs for incompletely resected gastrointestinal stromal tumors in dogs.


Gestion réussie à la suite d'une résection incomplète d'une tumeur stromale gastro-intestinale à l'aide de l'imatinib basée sur un test de sensibilité tumorale à base d'adénosine triphosphate chez un chien. Les tumeurs stromales gastro-intestinales résultant du cardia gastrique sont rares chez le chien. Quelques études ont montré l'efficacité des inhibiteurs de la tyrosine kinase dans le traitement des tumeurs stromales gastrointestinales canines, mais aucun protocole standardisé n'est actuellement disponible. Une chienne maltaise stérilisée de 11 ans a reçu un diagnostic de tumeur stromale gastro-intestinale à l'aide d'analyses histopathologiques et immunohistochimiques. Un test de chimiosensibilité tumorale à base d'adénosine triphosphate a révélé que l'imatinib à des concentrations plus faibles avait un effet inhibiteur plus fort que le tocéranib. Sur la base des résultats du test, le chien a été traité avec de l'imatinib après l'opération. Après 28 mois de traitement, il n'y a eu aucune récidive de la tumeur.Message clinique clé :Les tests de chimiosensibilité tumorale à base d'adénosine triphosphate peuvent aider les cliniciens à sélectionner les médicaments chimiothérapeutiques postopératoires appropriés pour les tumeurs stromales gastro-intestinales incomplètement réséquées chez le chien.(Traduit par Dr Serge Messier).


Assuntos
Antineoplásicos , Doenças do Cão , Tumores do Estroma Gastrointestinal , Mesilato de Imatinib , Animais , Tumores do Estroma Gastrointestinal/veterinária , Tumores do Estroma Gastrointestinal/cirurgia , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/patologia , Cães , Mesilato de Imatinib/uso terapêutico , Doenças do Cão/tratamento farmacológico , Doenças do Cão/cirurgia , Feminino , Antineoplásicos/uso terapêutico , Trifosfato de Adenosina/uso terapêutico , Indóis , Pirróis
5.
Biochemistry (Mosc) ; 89(4): 585-600, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38831498

RESUMO

Accurate duplication and separation of long linear genomic DNA molecules is associated with a number of purely mechanical problems. SMC complexes are key components of the cellular machinery that ensures decatenation of sister chromosomes and compaction of genomic DNA during division. Cohesin, one of the essential eukaryotic SMC complexes, has a typical ring structure with intersubunit pore through which DNA molecules can be threaded. Capacity of cohesin for such topological entrapment of DNA is crucial for the phenomenon of post-replicative association of sister chromatids better known as cohesion. Recently, it became apparent that cohesin and other SMC complexes are, in fact, motor proteins with a very peculiar movement pattern leading to formation of DNA loops. This specific process has been called loop extrusion. Extrusion underlies multiple functions of cohesin beyond cohesion, but molecular mechanism of the process remains a mystery. In this review, we summarized the data on molecular architecture of cohesin, effect of ATP hydrolysis cycle on this architecture, and known modes of cohesin-DNA interactions. Many of the seemingly disparate facts presented here will probably be incorporated in a unified mechanistic model of loop extrusion in the not-so-distant future.


Assuntos
Proteínas de Ciclo Celular , Proteínas Cromossômicas não Histona , Coesinas , DNA , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/química , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/química , DNA/metabolismo , DNA/química , Humanos , Animais , Trifosfato de Adenosina/metabolismo , Trifosfato de Adenosina/química , Cromátides/metabolismo , Cromátides/química
6.
Pestic Biochem Physiol ; 202: 105945, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38879302

RESUMO

With the widespread utilization of the sanitizing product benzethonium chloride (BEC) throughout the coronavirus pandemic, concerns have emerged regarding its potential hazards. Nevertheless, the long-term and multigenerational toxic effects of BEC on aquatic organisms remains unexplored. This study investigates acute and chronic toxicity, oxidative stress, mitochondrial membrane potential, ATP concentrations, and gene expression using Daphnia carinata as the model organism. Meanwhile, hierarchical clustering analysis was utilized to investigate phenotypic effects among different treatment groups. The integrated biomarker response index version 2 (IBRv2) was employed to estimate the deviation in toxic effects over two generations. These results indicated that D. carinata in the second generation exhibited higher survival rate and lower levels of oxidative stress than the first generation. However, the higher sublethal effects were found in the second generation as follows, the weakened growth performance, mitochondrial membrane potential depolarization, reduced ATP concentrations, and down-regulated gene expression. The mitochondrial toxicity induced by BEC may account for the distinct toxic effects exhibited in two generations. The findings here can assist with the evaluation of potential risk for BEC on aquatic organisms, and provide new insight into the cross-generational toxicity mechanisms of pollutants in aquatic ecosystems.


Assuntos
Daphnia , Potencial da Membrana Mitocondrial , Estresse Oxidativo , Animais , Daphnia/efeitos dos fármacos , Daphnia/genética , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Trifosfato de Adenosina/metabolismo , Poluentes Químicos da Água/toxicidade
7.
Nat Commun ; 15(1): 5113, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38879529

RESUMO

Factor-dependent termination uses molecular motors to remodel transcription machineries, but the associated mechanisms, especially in eukaryotes, are poorly understood. Here we use single-molecule fluorescence assays to characterize in real time the composition and the catalytic states of Saccharomyces cerevisiae transcription termination complexes remodeled by Sen1 helicase. We confirm that Sen1 takes the RNA transcript as its substrate and translocates along it by hydrolyzing multiple ATPs to form an intermediate with a stalled RNA polymerase II (Pol II) transcription elongation complex (TEC). We show that this intermediate dissociates upon hydrolysis of a single ATP leading to dissociation of Sen1 and RNA, after which Sen1 remains bound to the RNA. We find that Pol II ends up in a variety of states: dissociating from the DNA substrate, which is facilitated by transcription bubble rewinding, being retained to the DNA substrate, or diffusing along the DNA substrate. Our results provide a complete quantitative framework for understanding the mechanism of Sen1-dependent transcription termination in eukaryotes.


Assuntos
Trifosfato de Adenosina , DNA Helicases , RNA Polimerase II , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Imagem Individual de Molécula , Terminação da Transcrição Genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , RNA Polimerase II/metabolismo , Trifosfato de Adenosina/metabolismo , DNA Helicases/metabolismo , DNA Helicases/genética , Imagem Individual de Molécula/métodos , RNA Helicases/metabolismo , RNA Helicases/genética , Transcrição Gênica , RNA Fúngico/metabolismo , RNA Fúngico/genética , DNA Fúngico/metabolismo , DNA Fúngico/genética , Hidrólise
8.
Nat Commun ; 15(1): 5119, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38879572

RESUMO

One open question in the biology of growth factor receptors is how a quantitative input (i.e., ligand concentration) is decoded by the cell to produce specific response(s). Here, we show that an EGFR endocytic mechanism, non-clathrin endocytosis (NCE), which is activated only at high ligand concentrations and targets receptor to degradation, requires a tripartite organelle platform involving the plasma membrane (PM), endoplasmic reticulum (ER) and mitochondria. At these contact sites, EGFR-dependent, ER-generated Ca2+ oscillations are sensed by mitochondria, leading to increased metabolism and ATP production. Locally released ATP is required for cortical actin remodeling and EGFR-NCE vesicle fission. The same biochemical circuitry is also needed for an effector function of EGFR, i.e., collective motility. The multiorganelle signaling platform herein described mediates direct communication between EGFR signaling and mitochondrial metabolism, and is predicted to have a broad impact on cell physiology as it is activated by another growth factor receptor, HGFR/MET.


Assuntos
Trifosfato de Adenosina , Endocitose , Retículo Endoplasmático , Receptores ErbB , Mitocôndrias , Transdução de Sinais , Mitocôndrias/metabolismo , Receptores ErbB/metabolismo , Retículo Endoplasmático/metabolismo , Humanos , Trifosfato de Adenosina/metabolismo , Animais , Membrana Celular/metabolismo , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo
9.
Cell Biochem Funct ; 42(4): e4025, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38845083

RESUMO

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease. Metabolic and mitochondrial dysregulation are critical causal factors in the pathogenesis and progression of RA. Mitochondrial dysfunction include abnormal energy metabolism, and excessive production of reactive oxygen species (ROS). This study aimed to investigate the adenosine triphosphate (ATP), mitochondrial membrane potential (ΔΨm), ROS, and mRNA expression level of ROMO1 (as ROS modulator) and OMA1 (as regulator mitochondrial dynamics) of peripheral blood mononuclear cells (PBMC) in RA patients. The study participants were 50 patients with RA and 50 sex- and age-matched healthy volunteers. PBMC of all participant were isolated by Ficoll-Paque. Alteration in ΔΨm and cellular ROS were measured using flow cytometry, ATP level was also assessed via luminometry, and ROMO1 and OMA1 mRNA expression via qRT-PCR assay. A significant decrease in ATP (p = .005) and ΔΨm (p < .001) was observed in the PBMC of RA compared to control. The ROS levels were significantly higher in the PBMC of RA compared to the control (p < .001). ROMO1 and OMA1 mRNA expression was also significantly increased in RA patients compared to control (p < .001). The decrease in ATP is strongly associated with ROS increasing in PBMC of RA patients, denoting an inverse and negative relationship between ATP and ROS production. Also, a decrease in ΔΨm was observed. It seems that in line with mitochondrial dysfunction in PBMC, increased expression of ROMO1 and OMA1 genes could also be involved in the development of RA.


Assuntos
Artrite Reumatoide , Leucócitos Mononucleares , Mitocôndrias , Espécies Reativas de Oxigênio , Humanos , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Leucócitos Mononucleares/metabolismo , Feminino , Masculino , Espécies Reativas de Oxigênio/metabolismo , Mitocôndrias/metabolismo , Pessoa de Meia-Idade , Biomarcadores/metabolismo , Biomarcadores/sangue , Trifosfato de Adenosina/metabolismo , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Adulto , Potencial da Membrana Mitocondrial , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/genética
10.
Immun Inflamm Dis ; 12(6): e1286, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38860755

RESUMO

OBJECTIVE: This study aimed to link intracellular adenosine triphosphate content in CD4+ T lymphocytes (CD4+ iATP) with sepsis patient mortality, seeking a new predictive biomarker for outcomes and enhanced management. METHODS: 61 sepsis patients admitted to the Intensive Care Unit between October 2021 and November 2022 were enrolled. iATP levels were gauged using whole blood CD4+ T cells stimulated with mitogen PHA-L. Based on CD4+ iATP levels (<132.24 and ≥132.24 ng/mL), patients were categorized into two groups. The primary endpoint was all-cause mortality. To identify factors associated with mortality, both univariate and multivariate Cox proportional hazard analyses were conducted. RESULTS: Of the patients, 40 had high CD4+ iATP levels (≥132.24 ng/mL) and 21 had low levels (<132.24 ng/mL). In a 28-day follow-up, 21 (34.4%) patients perished. Adjusting for confounders like SOFA score, APACHE II score, lactic acid, and albumin, those with low CD4+ iATP had three- to fivefold higher mortality risk compared to high CD4+ iATP patients (61.9% vs. 20.0%; hazard ratio [95% confidence interval], Model 1: 4.515 [1.276-15.974], p = .019, Model 2: 3.512 [1.197-10.306], p = .022). CD4+ iATP correlated positively with white blood cell and neutrophil counts but not with lymphocytes, CD3, and CD4 counts. CONCLUSIONS: Low CD4+ iATP levels were associated with a higher risk of mortality in sepsis patients. Measurement of CD4+ iATP may serve as a useful tool for identifying patients at a higher risk of mortality and could potentially provide a basis for clinical treatment. Further research is warranted to fully elucidate the underlying mechanisms of this association.


Assuntos
Trifosfato de Adenosina , Linfócitos T CD4-Positivos , Sepse , Humanos , Trifosfato de Adenosina/metabolismo , Sepse/mortalidade , Sepse/imunologia , Sepse/sangue , Masculino , Feminino , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Pessoa de Meia-Idade , Estudos Prospectivos , Idoso , Biomarcadores , Prognóstico , Unidades de Terapia Intensiva/estatística & dados numéricos , Adulto
11.
Proc Natl Acad Sci U S A ; 121(25): e2318535121, 2024 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-38865270

RESUMO

The heart beats approximately 100,000 times per day in humans, imposing substantial energetic demands on cardiac muscle. Adenosine triphosphate (ATP) is an essential energy source for normal function of cardiac muscle during each beat, as it powers ion transport, intracellular Ca2+ handling, and actin-myosin cross-bridge cycling. Despite this, the impact of excitation-contraction coupling on the intracellular ATP concentration ([ATP]i) in myocytes is poorly understood. Here, we conducted real-time measurements of [ATP]i in ventricular myocytes using a genetically encoded ATP fluorescent reporter. Our data reveal rapid beat-to-beat variations in [ATP]i. Notably, diastolic [ATP]i was <1 mM, which is eightfold to 10-fold lower than previously estimated. Accordingly, ATP-sensitive K+ (KATP) channels were active at physiological [ATP]i. Cells exhibited two distinct types of ATP fluctuations during an action potential: net increases (Mode 1) or decreases (Mode 2) in [ATP]i. Mode 1 [ATP]i increases necessitated Ca2+ entry and release from the sarcoplasmic reticulum (SR) and were associated with increases in mitochondrial Ca2+. By contrast, decreases in mitochondrial Ca2+ accompanied Mode 2 [ATP]i decreases. Down-regulation of the protein mitofusin 2 reduced the magnitude of [ATP]i fluctuations, indicating that SR-mitochondrial coupling plays a crucial role in the dynamic control of ATP levels. Activation of ß-adrenergic receptors decreased [ATP]i, underscoring the energetic impact of this signaling pathway. Finally, our work suggests that cross-bridge cycling is the largest consumer of ATP in a ventricular myocyte during an action potential. These findings provide insights into the energetic demands of EC coupling and highlight the dynamic nature of ATP concentrations in cardiac muscle.


Assuntos
Trifosfato de Adenosina , Cálcio , Acoplamento Excitação-Contração , Ventrículos do Coração , Miócitos Cardíacos , Miócitos Cardíacos/metabolismo , Trifosfato de Adenosina/metabolismo , Acoplamento Excitação-Contração/fisiologia , Animais , Cálcio/metabolismo , Ventrículos do Coração/metabolismo , Ventrículos do Coração/citologia , Potenciais de Ação/fisiologia , Retículo Sarcoplasmático/metabolismo , Frequência Cardíaca/fisiologia , Humanos , Canais KATP/metabolismo , Contração Miocárdica/fisiologia , Camundongos
12.
Sci Rep ; 14(1): 13550, 2024 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-38866785

RESUMO

Red blood cells (RBCs) exhibit an interesting response to hydrodynamic flow, releasing adenosine triphosphate (ATP). Subsequently, these liberated ATP molecules initiate a crucial interaction with endothelial cells (ECs), thereby setting off a cascade involving the release of calcium ions (Ca 2 + ). Ca 2 + exerts control over a plethora of cellular functions, and acts as a mediator for dilation and contraction of blood vessel walls. This study focuses on the relationship between RBC dynamics and Ca 2 + dynamics, based on numerical simulations under Poiseuille flow within a linear two-dimensional channel. It is found that the concentration of ATP depends upon a variety of factors, including RBC density, channel width, and the vigor of the flow. The results of our investigation reveals several features. Firstly, the peak amplitude of Ca 2 + per EC escalates in direct proportion to the augmentation of RBC concentration. Secondly, increasing the flow strength induces a reduction in the time taken to reach the peak of Ca 2 + concentration, under the condition of a constant channel width. Additionally, when flow strength remains constant, an increase in channel width corresponds to an elevation in calcium peak amplitude, coupled with a decrease in peak time. This implies that Ca 2 + signals should transition from relatively unconstrained channels to more confined pathways within real vascular networks. This notion gains support from our examination of calcium propagation in a linear channel. In this scenario, the localized Ca 2 + release initiates a propagating wave that gradually encompasses the entire channel. Notably, our computed propagation speed agrees with observations.


Assuntos
Trifosfato de Adenosina , Cálcio , Células Endoteliais , Eritrócitos , Eritrócitos/metabolismo , Trifosfato de Adenosina/metabolismo , Cálcio/metabolismo , Células Endoteliais/metabolismo , Humanos , Sinalização do Cálcio
13.
Biophys Chem ; 311: 107270, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38833963

RESUMO

We propose a detailed computational beta cell model that emphasizes the role of anaplerotic metabolism under glucose and glucose-glutamine stimulation. This model goes beyond the traditional focus on mitochondrial oxidative phosphorylation and ATP-sensitive K+ channels, highlighting the predominant generation of ATP from phosphoenolpyruvate in the vicinity of KATP channels. It also underlines the modulatory role of H2O2 as a signaling molecule in the first phase of glucose-stimulated insulin secretion. In the second phase, the model emphasizes the critical role of anaplerotic pathways, activated by glucose stimulation via pyruvate carboxylase and by glutamine via glutamate dehydrogenase. It particularly focuses on the production of NADPH and glutamate as key enhancers of insulin secretion. The predictions of the model are consistent with empirical data, highlighting the complex interplay of metabolic pathways and emphasizing the primary role of glucose and the facilitating role of glutamine in insulin secretion. By delineating these crucial metabolic pathways, the model provides valuable insights into potential therapeutic targets for diabetes.


Assuntos
Glucose , Glutamina , Secreção de Insulina , Insulina , Modelos Biológicos , Glutamina/metabolismo , Glucose/metabolismo , Insulina/metabolismo , Humanos , Células Secretoras de Insulina/metabolismo , Animais , Piruvato Carboxilase/metabolismo , Peróxido de Hidrogênio/metabolismo , Trifosfato de Adenosina/metabolismo
14.
Sci Rep ; 14(1): 13148, 2024 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-38849425

RESUMO

Recent data indicate that extracellular ATP affects wound healing efficacy via P2Y2-dependent signaling pathway. In the current work, we propose double-modified ATP analogue-alpha-thio-beta,gamma-methylene-ATP as a potential therapeutic agent for a skin regeneration. For the better understanding of structure-activity relationship, beside tested ATP analogues, the appropriate single-modified derivatives of target compound, such as alpha-thio-ATP and beta,gamma-methylene-ATP, were also tested in the context of their involvement in the activation of ATP-dependent purinergic signaling pathway via the P2Y2 receptor. The diastereomerically pure alpha-thio-modified-ATP derivatives were obtained using the oxathiaphospholane method as separate SP and RP diastereomers. Both the single- and double- modified ATP analogues were then tested for their impact on the viability and migration of human keratinocytes. The involvement of P2Y2-dependent purinergic signaling was analyzed in silico by molecular docking of the tested compounds to the P2Y2 receptor and experimentally by studying intracellular calcium mobilization in the human keratinocytes HaCaT. The effects obtained for ATP analogues were compared with the results for ATP as a natural P2Y2 agonist. To confirm the contribution of the P2Y2 receptor to the observed effects, the tests were also performed in the presence of the selective P2Y2 antagonist-AR-C118925XX. The ability of the alpha-thio-beta,gamma-methylene-ATP to influence cell migration was analyzed in vitro on the model HaCaT and MDA-MB-231 cells by wound healing assay and transwell migration test as well as in vivo using zebrafish system. The impact on tissue regeneration was estimated based on the regrowth rate of cut zebrafish tails. The in vitro and in vivo studies have shown that the SP-alpha-thio-beta,gamma-methylene-ATP analogue promotes regeneration-related processes, making it a suitable agent for enhance wound healing. Performed studies indicated its impact on the cell migration, induction of epithelial-mesenchymal transition and intracellular calcium mobilization. The enhanced regeneration of cut zebrafish tails confirmed the pro-regenerative activity of this ATP analogue. Based on the performed studies, the SP-alpha-thio-beta,gamma-methylene-ATP is proposed as a potential therapeutic agent for wound healing and skin regeneration treatment.


Assuntos
Trifosfato de Adenosina , Queratinócitos , Cicatrização , Peixe-Zebra , Cicatrização/efeitos dos fármacos , Humanos , Trifosfato de Adenosina/metabolismo , Animais , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Simulação de Acoplamento Molecular , Movimento Celular/efeitos dos fármacos , Receptores Purinérgicos P2Y2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Cálcio/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Relação Estrutura-Atividade
15.
Nat Commun ; 15(1): 5035, 2024 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-38866788

RESUMO

Radio-immunotherapy exploits the immunostimulatory features of ionizing radiation (IR) to enhance antitumor effects and offers emerging opportunities for treating invasive tumor indications such as melanoma. However, insufficient dose deposition and immunosuppressive microenvironment (TME) of solid tumors limit its efficacy. Here we report a programmable sequential therapeutic strategy based on multifunctional fusogenic liposomes (Lip@AUR-ACP-aptPD-L1) to overcome the intrinsic radio-immunotherapeutic resistance of solid tumors. Specifically, fusogenic liposomes are loaded with gold-containing Auranofin (AUR) and inserted with multivariate-gated aptamer assemblies (ACP) and PD-L1 aptamers in the lipid membrane, potentiating melanoma-targeted AUR delivery while transferring ACP onto cell surface through selective membrane fusion. AUR amplifies IR-induced immunogenic death of melanoma cells to release antigens and damage-associated molecular patterns such as adenosine triphosphate (ATP) for triggering adaptive antitumor immunity. AUR-sensitized radiotherapy also upregulates matrix metalloproteinase-2 (MMP-2) expression that combined with released ATP to activate ACP through an "and" logic operation-like process (AND-gate), thus triggering the in-situ release of engineered cytosine-phosphate-guanine aptamer-based immunoadjuvants (eCpG) for stimulating dendritic cell-mediated T cell priming. Furthermore, AUR inhibits tumor-intrinsic vascular endothelial growth factor signaling to suppress infiltration of immunosuppressive cells for fostering an anti-tumorigenic TME. This study offers an approach for solid tumor treatment in the clinics.


Assuntos
Aptâmeros de Nucleotídeos , Imunoterapia , Lipossomos , Melanoma , Microambiente Tumoral , Lipossomos/química , Aptâmeros de Nucleotídeos/química , Animais , Camundongos , Linhagem Celular Tumoral , Imunoterapia/métodos , Melanoma/terapia , Melanoma/imunologia , Humanos , Microambiente Tumoral/efeitos dos fármacos , Metaloproteinase 2 da Matriz/metabolismo , Ouro/química , Camundongos Endogâmicos C57BL , Feminino , Antígeno B7-H1/metabolismo , Antígeno B7-H1/imunologia , Trifosfato de Adenosina/metabolismo
16.
Microbiologyopen ; 13(3): e23, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38867416

RESUMO

The G protein-coupled estrogen receptor, also known as GPER1 or originally GPR30, is found in various tissues, indicating its diverse functions. It is typically present in immune cells, suggesting its role in regulating immune responses to infectious diseases. Our previous studies have shown that G-1, a selective GPER agonist, can limit the pathogenesis mediated by Staphylococcus aureus alpha-hemolysin (Hla). It aids in clearing bacteria in a mouse skin infection model and restricts the surface display of the Hla receptor, ADAM10 (a disintegrin and metalloprotease 10) in HaCaT keratinocytes. In this report, we delve into the modulation of GPER in human immune cells in relation to the NLRP3 inflammasome. We used macrophage-like differentiated THP-1 cells for our study. We found that treating these cells with G-1 reduces ATP release, decreases the activity of the caspase-1 enzyme, and lessens cell death following Hla intoxication. This is likely due to the reduced levels of ADAM10 and NLRP3 proteins, as well as the decreased display of the ADAM10 receptor in the G-1-treated THP-1 cells. Our studies, along with our previous work, suggest the potential therapeutic use of G-1 in reducing Hla susceptibility in humans. This highlights the importance of GPER in immune regulation and its potential as a therapeutic target.


Assuntos
Proteína ADAM10 , Secretases da Proteína Precursora do Amiloide , Toxinas Bacterianas , Proteínas Hemolisinas , Inflamassomos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Receptores de Estrogênio , Receptores Acoplados a Proteínas G , Staphylococcus aureus , Proteína ADAM10/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Humanos , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Hemolisinas/metabolismo , Inflamassomos/metabolismo , Toxinas Bacterianas/metabolismo , Células THP-1 , Receptores de Estrogênio/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Proteínas de Membrana/agonistas , Caspase 1/metabolismo , Trifosfato de Adenosina/metabolismo , Macrófagos/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/microbiologia , Dipeptídeos , Ácidos Hidroxâmicos
17.
PLoS One ; 19(6): e0304062, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38870128

RESUMO

ATP is actively maintained at high concentrations in cancerous tissues, where it promotes a malignant phenotype through P2 receptors. In this study, we first evaluated the effect of extracellular ATP depletion with apyrase in SKOV-3, a cell line derived from metastatic ovarian carcinoma. We observed a decrease in cell migration and an increase in transepithelial electrical resistance and cell markers, suggesting a role in maintaining a mesenchymal phenotype. To identify the P2 receptor that mediated the effects of ATP, we compared the transcript levels of some P2 receptors and found that P2RX7 is three-fold higher in SKOV-3 cells than in a healthy cell line, namely HOSE6-3 (from human ovarian surface epithelium). Through bioinformatic analysis, we identified a higher expression of the P2RX7 transcript in metastatic tissues than in primary tumors; thus, P2X7 seems to be a promising effector for the malignant phenotype. Subsequently, we demonstrated the presence and functionality of the P2X7 receptor in SKOV-3 cells and showed through pharmacological approaches that its activity promotes cell migration and contributes to maintaining a mesenchymal phenotype. P2X7 activation using BzATP increased cell migration and abolished E-cadherin expression. On the other hand, a series of P2X7 receptor antagonists (A438079, BBG and OxATP) decreased cell migration. We used a CRISPR-based knock-out system directed to P2RX7. According to the results of our wound-healing assay, SKOV3-P2X7KO cells lacked receptor-mediated calcium mobilization and decreased migration. Altogether, these data let us propose that P2X7 receptor is a regulator for cancer cell migration and thus a potential drug target.


Assuntos
Trifosfato de Adenosina , Movimento Celular , Neoplasias Ovarianas , Receptores Purinérgicos P2X7 , Humanos , Receptores Purinérgicos P2X7/metabolismo , Receptores Purinérgicos P2X7/genética , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/genética , Feminino , Trifosfato de Adenosina/metabolismo , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica
18.
Mikrochim Acta ; 191(7): 386, 2024 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-38867016

RESUMO

The combination of CRISPR/Cas12a and functional DNA provides the possibility of constructing biosensors for detecting non-nucleic-acid targets. In the current study, the duplex protospacer adjacent motif (PAM) in the activator of CRISPR/Cas12a was used as a molecular switch, and a sensitive adenosine triphosphate (ATP) detection biosensor was constructed using an allosteric probe-conjugated PAM site formation in hybridization chain reaction (HCR) integrated with the CRISPR/Cas12a system (APF-CRISPR). In the absence of ATP, an aptamer-containing probe (AP) is in a stem-loop structure, which blocks the initiation of HCR. In the presence of ATP, the structure of AP is changed upon ATP binding, resulting in the release of the HCR trigger strand and the production of long duplex DNA with many PAM sites. Since the presence of a duplex PAM site is crucial for triggering the cleavage activity of CRISPR/Cas12a, the ATP-dependent formation of the PAM site in HCR products can initiate the FQ-reporter cleavage, allowing ATP quantification by measuring the fluorescent signals. By optimizing the sequence elements and detection conditions, the aptasensor demonstrated superior detection performance. The limit of detection (LOD) of the assay was estimated to be 1.16 nM, where the standard deviation of the blank was calculated based on six repeated measurements. The dynamic range of the detection was 25-750 nM, and the whole workflow of the assay was approximately 60 min. In addition, the reliability and practicability of the aptasensor were validated by comparing it with a commercially available chemiluminescence kit for ATP detection in serum. Due to its high sensitivity, specificity, and reliable performance, the APF-CRISPR holds great potential in bioanalytical studies for ATP detection. In addition, we have provided a proof-of-principle for constructing a CRISPR/Cas12a-based aptasensor, in which the PAM is utilized to regulate Cas12a cleavage activity.


Assuntos
Trifosfato de Adenosina , Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , Sistemas CRISPR-Cas , Trifosfato de Adenosina/química , Trifosfato de Adenosina/análise , Técnicas Biossensoriais/métodos , Aptâmeros de Nucleotídeos/química , Proteínas Associadas a CRISPR/química , Limite de Detecção , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Hibridização de Ácido Nucleico , Endodesoxirribonucleases
19.
Reprod Fertil Dev ; 362024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38870344

RESUMO

In addition to its central role in cellular metabolism, adenosine 5'-triphosphate (ATP) is an important extracellular signalling molecule involved in various physiological processes. In reproduction, extracellular ATP participates in both autocrine and paracrine paths regulating gametogenesis, gamete maturation and fertilisation. This review focusses on how extracellular ATP modulates sperm physiology with emphasis on the mammalian acrosome reaction. The presence of extracellular ATP in the reproductive tract is primarily determined by the ion channels and transporters that influence its movement within the cells comprising the tract. The main targets of extracellular ATP in spermatozoa are its own transporters, particularly species-specific sperm purinergic receptors. We also discuss notable phenotypes from knock-out mouse models and human Mendelian inheritance related to ATP release mechanisms, along with immunological, proteomic, and functional observations regarding sperm purinergic receptors and their involvement in sperm signalling.


Assuntos
Trifosfato de Adenosina , Espermatozoides , Animais , Masculino , Espermatozoides/metabolismo , Espermatozoides/fisiologia , Trifosfato de Adenosina/metabolismo , Humanos , Reação Acrossômica/fisiologia , Receptores Purinérgicos/metabolismo , Transdução de Sinais , Mamíferos/fisiologia , Camundongos
20.
J Ovarian Res ; 17(1): 118, 2024 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-38822408

RESUMO

In women who are getting older, the quantity and quality of their follicles or oocytes and decline. This is characterized by decreased ovarian reserve function (DOR), fewer remaining oocytes, and lower quality oocytes. As more women choose to delay childbirth, the decline in fertility associated with age has become a significant concern for modern women. The decline in oocyte quality is a key indicator of ovarian aging. Many studies suggest that age-related changes in oocyte energy metabolism may impact oocyte quality. Changes in oocyte energy metabolism affect adenosine 5'-triphosphate (ATP) production, but how related products and proteins influence oocyte quality remains largely unknown. This review focuses on oocyte metabolism in age-related ovarian aging and its potential impact on oocyte quality, as well as therapeutic strategies that may partially influence oocyte metabolism. This research aims to enhance our understanding of age-related changes in oocyte energy metabolism, and the identification of biomarkers and treatment methods.


Assuntos
Envelhecimento , Metabolismo Energético , Oócitos , Ovário , Oócitos/metabolismo , Humanos , Feminino , Envelhecimento/metabolismo , Ovário/metabolismo , Animais , Trifosfato de Adenosina/metabolismo
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