Intracellular Adenosine Triphosphate Content Research Articles

Sodium butyrate activates the KATP channels to regulate the mechanism of Parkinson's disease microglia model inflammation.

Parkinson's disease (PD) is a common neurodegenerative disorder. Microglia-mediated neuroinflammation has emerged as an involving mechanism at the initiation and development of PD. Activation of adenosine triphosphate (ATP)-sensitive potassium (KATP ) channels can protect dopaminergic neurons from damage. Sodium butyrate (NaB) shows anti-inflammatory and neuroprotective effects in some animal models of brain injury and regulates the KATP channels in islet β cells. In this study, we aimed to verify the anti-inflammatory effect of NaB on PD and further explored potential molecular mechanisms. We established an in vitro PDmodel in BV2 cells using 1-methyl-4-phenylpyridinium (MPP+ ). The effects of MPP+ and NaB on BV2 cell viability were detected by cell counting kit-8 assays. The morphology of BV2 cells with or without MPP+ treatment was imaged via an optical microscope. The expression of Iba-1 was examined by the immunofluorescence staining. The intracellular ATP content was estimated through the colorimetric method, and Griess assay was conducted to measure the nitric oxide production. The expression levels of pro-inflammatory cytokines and KATP channel subunits were evaluated by reverse transcription-quantitativepolymerase chain reaction and western blot analysis. NaB (5 mM) activated the KATP channels through elevating Kir6.1 and Kir6.1 expression in MPP+ -challenged BV2 cells. Both NaB and pinacidil (a KATP opener) suppressed the MPP+ -induced activation of BV2 cells and reduced the production of nitrite and pro-inflammatory cytokines in MPP+ -challenged BV2 cells. NaB treatment alleviates the MPP+ -induced inflammatory responses in microglia via activation of KATP channels.

Exploring the Role of Surface and Mitochondrial ATP-Sensitive Potassium Channels in Cancer: From Cellular Functions to Therapeutic Potentials.

ATP-sensitive potassium (KATP) channels are found in plasma membranes and mitochondria. These channels are a type of ion channel that is regulated by the intracellular concentration of adenosine triphosphate (ATP) and other nucleotides. In cell membranes, they play a crucial role in linking metabolic activity to electrical activity, especially in tissues like the heart and pancreas. In mitochondria, KATP channels are involved in protecting cells against ischemic damage and regulating mitochondrial function. This review delves into the role of KATP channels in cancer biology, underscoring their critical function. Notably responsive to changes in cellular metabolism, KATP channels link metabolic states to electrical activity, a feature that becomes particularly significant in cancer cells. These cells, characterized by uncontrolled growth, necessitate unique metabolic and signaling pathways, differing fundamentally from normal cells. Our review explores the intricate roles of KATP channels in influencing the metabolic and ionic balance within cancerous cells, detailing their structural and operational mechanisms. We highlight the channels' impact on cancer cell survival, proliferation, and the potential of KATP channels as therapeutic targets in oncology. This includes the challenges in targeting these channels due to their widespread presence in various tissues and the need for personalized treatment strategies. By integrating molecular biology, physiology, and pharmacology perspectives, the review aims to enhance the understanding of cancer as a complex metabolic disease and to open new research and treatment avenues by focusing on KATP channels. This comprehensive approach provides valuable insights into the potential of KATP channels in developing innovative cancer treatments.

NH2-MIL-101(Fe)-mediated photo-Fenton reaction enhanced simultaneous removal of nitrogen and refractory organics in anammox process through interfacial electron transfer

In this study, H2O2 (0.1 ‰) and NH2-MIL-101(Fe)-driven (150 mg/L) photo-Fenton-coupled anammox were proposed to simultaneously improve the removal efficiency of nitrogen and humic acid. Long-term experiments showed that the total nitrogen removal efficiency was increased by the photo-Fenton reaction to 91.9 ± 1.5 % by altering the bioavailability of refractory organics. Correspondingly, the total organic carbon removal efficiency was significantly increased. Microbial community analyses indicated that Candidatus_Brocadia maintained high activity during photo-Fenton reaction and was the most abundant genus in the reactor. Dissimilatory nitrate reduction to ammonium process and denitrification process were enhanced, resulting in reduced NO3–-N production. The establishment of electron transfer between microorganisms and NH2-MIL-101 (Fe) improved the charge separation efficiency of the quantum dots and increased the intracellular adenosine triphosphate content of anammox bacteria. These results indicated that photo-Fenton-anammox process promoted the removal of nitrogen and refractory organics in one reactor which had good economic value and application prospects.

Adenosine Triphosphate (ATP) and Protein Aggregation in Age-Related Vision-Threatening Ocular Diseases.

Protein aggregation is the etiopathogenesis of the three most profound vision-threatening eye diseases: age-related cataract, presbyopia, and age-related macular degeneration. This perspective organizes known information on ATP and protein aggregation with a fundamental unrecognized function of ATP. With recognition that maintenance of protein solubility is related to the high intracellular concentration of ATP in cells, tissues, and organs, we hypothesize that (1) ATP serves a critical molecular function for organismal homeostasis of proteins and (2) the hydrotropic feature of ATP prevents pathological protein aggregation while assisting in the maintenance of protein solubility and cellular, tissue, and organismal function. As such, the metabolite ATP plays an extraordinarily important role in the prevention of protein aggregation in the leading causes of vision loss or blindness worldwide.

Endocytic internalization mechanism of bioactive antibacterial nanoparticles by fibroblasts

Studying the uptake mechanism of photosensitizers is an important step in developing an ideal photosensitizer for use in photodynamic therapy (PDT). Understanding the uptake mechanism can help design novel photosensitizers that are selectively accumulated in the target tissue, with improved pharmacokinetics, and are dosed optimally to maximize the efficacy of the treatment. In our previous studies we synthesized and characterized the use of chitosan nanoparticles functionalized with rose bengal (CSRBnp) as a photosensitizer against dental biofilm. The aim of this study is to analyze the internalization mechanism and cellular proinflammatory activities of CSRBnps on fibroblasts. Fibroblasts (NIH 3T3) were incubated with chlorpromazine (5 µg/ml), nystatin (5 µg/ml), wortmannin (100 ng/ml) and at 4 °C for 30 min followed by CSRBnp (0.3 mg/ml). Cell viability (MTS assay), intracellular adenosine triphosphate content (Luminescence assay), cytokine expression (TNF-α) using ELISA and nitric oxide (NO) production by Griess reaction system were conducted at different time intervals (30 min, 1, 4, and 12 h). The internalization of CSRBnps was analyzed using live cell imaging confocal microscope with excitation wavelengths of 405 and 568 to detect nuclei (Hoechst 33,342) and CSRBnps respectively. CSRBnps and inhibitors at the applied concentrations were not cytotoxic. ATP content in chlorpromazine and without inhibitors groups were significantly lower than the control group at 12 h. All inhibitors showed significantly lower CSRBnps uptake compared to the control group at 30 min, 1 h, and 4 h time. Wortmannin resulted in the most significant inhibition of CSRBnps uptake as compared to chlorpromazine and nystatin (P < 0.05). TNF-α expression and NO production were not significant during the entire CSRBnps uptake. The results showed macropinocytosis was a dominant CSRBnps uptake mechanism by fibroblasts in the early stages and non-specific uptake pathways were activated after prolonged incubation time. CSRBnps uptake by fibroblasts was energy dependent and did not cause any proinflammatory response.

Morphofunctional state of platelets in their concentrates depending on storage time

Background.Evaluation of the functional state of platelets in the composition of concentrates is necessary to improve the methods of their production, optimize storage conditions and terms, increase therapeutic efficacy, and reduce the risk of transfusion complications.&#x0D; Aim.Comprehensive study of the morphofunctional state of platelets during the storage of their concentrates.&#x0D; Material and methods.202samples of human platelet apheresis concentrates were studied for storage periods of up to 17days at a temperature of 2224C. The functional state of platelets was assessed using flow cytometry for the expression of P-selectin, active IIb3 integrin, and phosphatidylserines before and after biochemical stimulation. In addition, the mitochondrial potential of platelets, the concentration of intracellular adenosine triphosphate, contractile properties, induced aggregation, and morphological characteristics were studied according to scanning electron microscopy. Statistical analysis was performed by analysis of variance (ANOVA or KruskalWallis).&#x0D; Results.In non-activated platelets during storage, the spontaneous expression of P-selectin increased by an average of 7times, and phosphatidylserines by 3times. The induced expression of P-selectin, integrin IIb3 and phosphatidylserines was the highest on the day of receiving the concentrate and gradually decreased during storage to 1/2of the initial level. Platelet aggregation activity in response to collagen stimulation progressively decreased by an average of 100times, in response to the TRAP peptide by 1.5times. In contrast to the expression of activation markers, the ability of platelets to compress plasma clots in most samples changed slightly, within a few percent; in single samples, the contractile function dropped to zero by 47days of storage, which was combined with platelet hyperactivation and depletion. The mitochondrial potential of platelets and the content of adenosine triphosphate were relatively constant. Morphologically, discoid platelets predominated, however, starting from the 1st day of storage, spindle-shaped platelets, and from the 3rd day spherical forms and microaggregates of platelets accumulated.&#x0D; Conclusion.Platelets in the composition of concentrates initially have a high functional potential, which gradually decreases due to progressive spontaneous activation with a simultaneous decrease in reactivity and structural changes.

Role of Coptis chinensis in antibiotic susceptibility of carbapenem-resistant Klebsiella pneumoniae.

The incidence of carbapenem-resistant Klebsiella pneumoniae (CRKP) has rapidly increased. This study aimed to assess the effect of Coptis chinensis and its compounds on the minimal inhibitory concentrations (MICs) of eight antibiotics against CRKP. Cell cultures were used to investigate the effects of C. chinensis and its compounds on the MICs of eight antibiotics against CRKP. The MICs for antibiotics alone and antibiotics with C. chinensis or compounds were measured and compared. Furthermore, the effects of C. chinensis on cell membrane injury and intracellular adenosine triphosphate (ATP) CRKP concentration were also measured. The Mann-Whitney rank-sum test was used to analyze the differences between means. C. chinensis exhibits a notable MIC bacteriostatic effect at 5mg/mL on CRKP. A significant MIC reduction against CRKP exists when C. chinensis was added to colistin and colistin-containing two-antibiotic combinations. Moreover, C. chinensis could damage cell membrane integrity and decrease intracellular ATP concentration in CRKP. Thus, C. chinensis exhibits antimicrobial activity superiority with colistin against CRKP. Furthermore, the effects of identified compounds in C. chinensis on the MICs of colistin, four-to eight-, two-to four-, and one-to two-fold reductions were found in ferulic acid, magnoflorine, and jatrorrhizine hydrochloride, respectively. Among these compounds, ferulic acid destroys membrane integrity and decreases intracellular ATP concentration. C. chinensis and ferulic acid can potentiate the antimicrobial activity of colistin and may represent a promising component of combination therapy against CRKP infections in a clinical setting.

Strategies for variable regulation of methanogenesis efficiency and velocity.

Combinations of various strategic approaches to the suppression of methanogenesis and the formation of biogas with a simultaneous decrease in the ratio of methane in its composition were investigated. Introduction of methanogenesis suppressors such as redox derivatives of humic acids, potassium persulfate (K2S2O8), possessing oxidizing and electron acceptor properties, enzyme hexahistidine-containing organophosphorus hydrolase with high lactonase activity and polypeptide antimicrobial agent bacitracin into the media with anaerobic consortia were studied. The effect of these substances was directed at various participants of the natural methanogenic consortium, as well as on the biochemical processes carried out by them. The use of K2S2O8 together with bacitracin provided maximum and almost complete suppression of CH4 production. The measured concentration of intracellular adenosine triphosphate has shown that viability of cells in the consortium remained almost the same, whereas their metabolic activity decreased. Various combinations of the above-mentioned suppressors provided different degrees of methanogenesis suppression, but redox agents played a key role in all the cases studied. Based on the accumulated data, combining suppressors in different concentrations can be used to manage the methanogenesis (efficiency and velocity of its decrease) in media with anaerobic consortia. KEY POINTS: • Various strategies for suppression of the methanogenesis were combined. • The enzyme His6-OPH was firstly used for quorum quenching in methanogenic consortium. • Velocity of methanogenesis decrease can be managed by combinations of suppressors.

Connecting single-cell ATP dynamics to overflow metabolism, cell growth, and the cell cycle in Escherichia coli

Adenosine triphosphate (ATP) is an abundant and essential metabolite that cells consume and regenerate in large amounts to support growth. Although numerous studies have inferred the intracellular concentration of ATP in bacterial cultures, what happens in individual bacterial cells under stable growth conditions is less clear. Here, we use the QUEEN-2m biosensor to quantify ATP dynamics in single Escherichia coli cells in relation to their growth rate, metabolism, cell cycle, and cell lineage. We find that ATP dynamics are more complex than expected from population studies and are associated with growth-rate variability. Under stable nutrient-rich condition, cells can display large fluctuations in ATP level that are partially coordinated with the cell cycle. Abrogation of aerobic acetate fermentation (overflow metabolism) through genetic deletion considerably reduces both the amplitude of ATP level fluctuations and the cell-cycle trend. Similarly, growth in media in which acetate fermentation is lower or absent results in the reduction of ATP level fluctuation and cell-cycle trend. This suggests that overflow metabolism exhibits temporal dynamics, which contributes to fluctuating ATP levels during growth. Remarkably, at the single-cell level, growth rate negatively correlates with the amplitude of ATP fluctuation for each tested condition, linking ATP dynamics to growth-rate heterogeneity in clonal populations. Our work highlights the importance of single-cell analysis in studying metabolism and its implication to phenotypic diversity and cell growth.

Modeling ATP-mediated endothelial cell elongation on line patterns

Endothelial cell (EC) migration is crucial for a wide range of processes including vascular wound healing, tumor angiogenesis, and the development of viable endovascular implants. We have previously demonstrated that ECs cultured on 15-μm wide adhesive line patterns exhibit three distinct migration phenotypes: (a) “running” cells that are polarized and migrate continuously and persistently on the adhesive lines with possible spontaneous directional changes, (b) “undecided” cells that are highly elongated and exhibit periodic changes in the direction of their polarization while maintaining minimal net migration, and (c) “tumbling-like” cells that migrate persistently for a certain amount of time but then stop and round up for a few hours before spreading again and resuming migration. Importantly, the three migration patterns are associated with distinct profiles of cell length. Because of the impact of adenosine triphosphate (ATP) on cytoskeletal organization and cell polarization, we hypothesize that the observed differences in EC length among the three different migration phenotypes are driven by differences in intracellular ATP levels. In the present work, we develop a mathematical model that incorporates the interactions between cell length, cytoskeletal (F-actin) organization, and intracellular ATP concentration. An optimization procedure is used to obtain the model parameter values that best fit the experimental data on EC lengths. The results indicate that a minimalist model based on differences in intracellular ATP levels is capable of capturing the different cell length profiles observed experimentally.

Dextran-coated Gd-based ultrasmall nanoparticles as phosphatase-like nanozyme to increase ethanol yield via reduction of yeast intracellular ATP level

Nanozymes—functional materials that possess intrinsic enzyme-like characteristics—have gained tremendous attention in recent years owing to their unique advantages; however, further research is required to understand their scope in biological applications. In this study, dextran-coated nanogadolinia (DCNG) was synthesised, and its phosphatase mimetic activity was demonstrated. Specifically, the dephosphorylation of adenosine triphosphate (ATP), an important biomolecule, by DCNG was investigated. The results showed that DCNG could selectively catalyse the hydrolysis of the terminal high-energy phosphate bonds of ATP under physiological conditions. Furthermore, the biocompatible DCNG, with remarkable phosphatase mimicking activity, decreased the intracellular ATP content by dephosphorylation and increased ethanol yield during glucose fermentation by S. cerevisiae. These results indicate potential alternatives for improving ethanol yields and exploring novel biological applications of nanozymes.

Transcriptomics analyses and biochemical characterization of Aspergillus flavus spores exposed to 1-nonanol.

The exploitation of plant volatile organic compounds as biofumigants to control postharvest decaying of agro-products has received considerable research attention. Our previous study reported that 1-nonanol, the main constituent of cereal volatiles, can inhibit Aspergillus flavus growth and has the potential as a biofumigant to control the fungal spoilage of cereal grains. However, the antifungal mechanism of 1-nonanol against A. flavus is still unclear at the molecular level. In this study, the minimum inhibitory concentration and minimum fungicidal concentration of 1-nonanol against A. flavus spores were 2 and 4 μL/mL, respectively. Scanning electron microscopy revealed that the 1-nonanol can distort the morphology of A. flavus spore. Annexin V-FITC/PI double staining showed that 1-nonanol induced phosphatidylserine eversion and increased membrane permeability of A. flavus spores. Transcriptional profile analysis showed that 1-nonanol treatment mainly affected the expression of genes related to membrane damage, oxidative phosphorylation, blockage of DNA replication, and autophagy in A. flavus spores. Flow cytometry analysis showed that 1-nonanol treatment caused hyperpolarization of mitochondrial membrane potential and accumulation of reactive oxygen species in A. flavus spores. 4',6-diamidino-2-phenylindole staining showed that treatment with 1-nonanol destroyed the DNA. Biochemical analysis results confirmed that 1-nonanol exerted destructive effects on A. flavus spores by decreasing intracellular adenosine triphosphate content, reducing mitochondrial ATPase activity, accumulating hydrogen peroxide and superoxide anions, and increasing catalase and superoxide dismutase enzyme activities. This study provides new insights into the antifungal mechanisms of 1-nonanol against A. flavus. KEY POINTS: • 1-Nonanol treatment resulted in abnormal morphology of A. flavus spores. • 1-Nonanol affects the expression of key growth-related genes of A. flavus. • The apoptosis of A. favus spores were induced after exposed to 1-nonanol.

Kaempferol increases intracellular ATP content in C2C12 myotubes under hypoxic conditions by suppressing the HIF-1α stabilization and/or by enhancing the mitochondrial complex IV activity

Kaempferol (KMP) has numerous important biological functions, and we recently showed that it remarkably increased intracellular adenosine triphosphate (ATP) content in C2C12 myotubes under hypoxic conditions. Since intracellular ATP is generated by aerobic energy metabolism or anaerobic glycolysis, hypoxia inducible factor-1α (HIF-1α) has been shown to be associated with metabolic remodeling and causes metabolic shift from aerobic energy metabolism to anaerobic glycolysis in response to hypoxic conditions. Here, we investigate the effects of KMP under hypoxic conditions on the stabilization of HIF-1α in C2C12 myotubes and its underlying molecular mechanisms. Constitutive HIF-1α protein expression was observed in C2C12 myotubes, and its expression under hypoxic conditions was remarkably suppressed by KMP by reducing its stability; thus, resulting in an increase in ATP content. Furthermore, KMP strikingly increased the ubiquitination of HIF-1α and promoted its degradation via the ubiquitin proteasome system. Inhibition of HIF-1α by KMP resulted in the abrogation of the expression of glycolytic enzymes such as lactate dehydrogenase A and pyruvate dehydrogenase kinase isozyme 1. In addition, the metabolome profiling showed that KMP promoted oxidative energy production, while the mitochondrial complex activity assay indicated that KMP increased the activity of mitochondrial complex IV. Finally, we showed that KMP inhibited HIF-1α expression and increased intracellular ATP content in the soleus muscle of rats. Taken together, these results suggest that KMP increases intracellular ATP content under hypoxic conditions by suppressing the HIF-1α stabilization and/or by enhancing the mitochondrial complex IV activity in muscle.

Antibacterial Effect of Oregano Essential Oil against Vibrio vulnificus and Its Mechanism.

Oregano essential oil (OEO) is an effective natural antibacterial agent, but its antibacterial activity against Vibrio vulnificus has not been widely studied. The aim of this study was to investigate the inhibitory effect and germicidal activity of OEO on V. vulnificus and its possible inhibition mechanism. The minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of OEO against four V. vulnificus strains (ATCC 27562 and three isolates from seafoods) were from 0.06 to 0.15 μL/mL. Compared with untreated bacteria, OEO reduced the concentration of intracellular adenosine triphosphate (ATP), hyperpolarized the cell membrane, increased the level of reactive oxygen species (ROS), and increased the concentration of intracellular malondialdehyde (MDA), but there was no obvious DNA damage at the OEO test concentration. It was indicated that OEO inactivated V. vulnificus by generating ROS which caused lipid peroxidation of cell membranes, thereby reducing the permeability and integrity of cell membranes and causing morphological changes to cells, but there was no obvious damage to DNA. In addition, OEO could effectively kill V. vulnificus in oysters at 25 °C, and the number of bacteria decreased by 48.2% after 0.09% OEO treatment for 10 h. The good inhibitory effect and bactericidal activity of OEO showed in this study, and the economy and security of OEO make it possible to apply OEO to control V. vulnificus contamination in oysters and other seafoods.

[Metabolic system of hematopoietic stem cells and its age-related changes revealed by real-time ATP quantification in living cells].

Hematopoietic stem cells' (HSCs) metabolic dynamics regulate cell fate and differentiation. To maintain the stemness of HSCs, it is necessary to maintain an appropriate balance of adenosine triphosphate (ATP) production by the glycolytic system and mitochondria. In conventional intracellular ATP measurement, snapshot analysis is performed after the lysis of a large number of cells, resulting in the loss of single-cell level and spatiotemporal information. To overcome this technical limitation, we recently developed a methodology for measuring ATP concentration in living HSCs using knock-in mice of an intracellular ATP concentration biosensor called GO-ATeam2, which employs the principle of Förster resonance energy transfer. The current understanding of metabolism in HSCs, the metabolic dependence of HSCs revealed by the GO-ATeam2 system, and the prospects for novel metabolic measurement techniques in the future are reviewed in this paper.

Alterations in mitochondrial function and energy metabolism-related properties in thyroid cancer stem cells.

Increasing evidence indicates that cancer stem cells (CSCs) are initiators of the occurrence, development, and recurrence of malignant tumors. Mitochondria are important organelles in eukaryotic cells, not only responsible for converting part of energy released during nutrients oxidation into the energy-yielding molecule adenosine triphosphate (ATP) to fuel the activities of cell, but also play essential roles in processes such as cell apoptosis and cellular proliferation. The mitochondrial-related abnormalities have also been considered to have an important role in the origin and development of tumors. This study aimed at testing the abnormalities in mitochondrial function and energy/metabolism-related phenotypes in thyroid cancer stem cells (TCSCs). TCSCs were isolated and identified from MDA-T32 thyroid carcinoma cell line. The mitochondrial mass and mitochondrial arrangement, amount of mitochondrial DNA (mtDNA), mitochondrial membrane potential (MMP), oxygen/glucose consumption, and intracellular concentrations of reactive oxygen species (ROS) and ATP levels were examined. Perinuclear mitochondrial distribution, low amount of mtDNA and oxygen/glucose consumption, high MMP, and low intracellular ROS and ATP concentrations were observed in TCSCs. Alterations in mitochondrial function and cellular energy metabolism may be used as novel indicators of thyroid cancer.

Antibacterial Activity and Mechanism of Coenzyme Q0 Against Escherichia coli.

Coenzyme Q0 (CoQ0) is a natural compound found in Antrodia cinnamomea, which has a variety of biological activities. Here, the antibacterial activity and possible antibacterial mechanism of CoQ0 against Escherichia coli were investigated. The antibacterial effect was evaluated by determining minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values, and by assessing bacterial survival and the effect on the growth of E. coli after CoQ0 treatment in Luria-Bertani (LB) broth. To reveal the antibacterial mechanism of CoQ0, changes in intracellular adenosine triphosphate (ATP) concentration, membrane potential, and bacterial protein content, as well as effects on cell morphology and membrane integrity, were investigated. Both the MICs and MBCs of CoQ0 against E. coli were 0.1 mg/mL. After treatment of E. coli (6.5 log colony-forming units/mL) with 0.1 mg/mL of CoQ0 in LB broth for 3 h, the number of viable cells dropped below the detection limit. In addition, CoQ0 treatment resulted in the reduction in intracellular ATP concentration, cell membrane hyperpolarization, decreased bacterial protein concentrations, and damage to cell membrane integrity and cellular morphology. These results indicated that CoQ0 has effective antibacterial activity against E. coli, suggesting potential applications in food industry safety.

Effect of Oleanolic Acid on Apoptosis and Autophagy of SMMC-7721 Hepatoma Cells.

BackgroundLiver cancer is a common cancer with high morbidity and mortality. Due to the large toxic side effects of chemotherapeutic drugs and the overexpression of multidrug resistance genes in liver cancer, no effective chemotherapeutic drug has yet been found. Therefore, the search for a highly effective, low-toxic, and safe natural anticancer therapy is a hot issue.Material/MethodsSMMC-7721 cells (a hepatocellular carcinoma cell line) were treated with different concentrations of oleanolic acid (OA) plus autophagy inhibitor 3-methyladenine (3-MA) (3-MA+OA) or chloroquine (CQ) plus OA (CQ+OA). We used MTT and Hoechst 33258 staining methods to determine the proliferation and apoptotic effect of OA on cells. Flow cytometry was used to detect apoptosis. Mitochondrial function was assessed by measuring mitochondrial membrane potential and adenosine triphosphate (ATP) concentration. To evaluate the ability of OA on apoptosis and autophagy mechanisms on SMMC 7721 cells, the related protein expression for apoptosis, autophagy, and the autophagic pathway were detected and analyzed by western blot.ResultsOA can inhibit and induce apoptosis of SMMC-7721 in a dose-dependent manner. Compared with the control group, OA significantly reduced the intracellular mitochondrial membrane potential, and the intracellular ATP concentration was also significantly reduced. Moreover, OA reduced the expression of p-Akt and p-mTOR. The expression of p62 was decreased, and LC3-II and Beclin-1 protein expression levels increased. After inhibiting autophagy with 3-MA or CQ, compared with OA alone, cell mitochondrial membrane potential and ATP concentration were significantly reduced, cell p62 expression was reduced, and LC3-II expression was increased, apoptosis-related protein Bax protein was increased, and Bcl-2 protein was decreased, which suggested that 3-MA or CQ treatment increased OA-induced apoptosis of SMMC-7721 cells. This suggested that OA activated autophagy of SMMC-7721 cells in a protective autophagic manner.ConclusionsThe study findings suggest that OA combined with autophagy inhibitor 3-MA can better exert its anticancer effect.

Optimization potential of anaerobic biocatalytic processes using intracellular ATP concentration as the main criterion for decision making

The effective use of such an analytical indicator as the concentration of intracellular adenosine triphosphate (ATP) in the process of anaerobic transformation of sulfones was shown. The bioluminescent method for determining ATP evaluates to quickly evaluate the state of biocatalysts in such processes, to select the conditions for effective conducted of biotechnological processes and the functioning of cells, as well as media compositions used in the conversion of sulfur-containing compounds.

Into the heart of blood flow

Signal Transduction The flow of blood within the heart is exactly matched to its metabolic needs and, throughout life, never runs out of energy. How is this possible? Zhao et al. identified a local and instantaneous feedback control system that couples metabolism in cardiac myocytes to the regulation of blood flow. Several such mechanisms exist, but these authors characterize one that depends on adenosine triphosphate (ATP)–sensitive K+ (KATP) channels. Studies of mouse ventricular myocytes showed that as active cells depleted intracellular concentrations of ATP, KATP channels were activated. This caused an efflux of K+, thus depolarizing the cells. Electrical coupling of the myocytes to neighboring endothelial cells and contractile pericytes and smooth muscle cells allowed a compensatory increase in blood flow. Proc. Natl. Acad. Sci. U.S.A. 117 , 7461 (2020).