Adenosine Triphosphate Content In Cells Research Articles

Dual-signal readout sensing of ATP content in single dental pulp stem cells during differentiation via functionalized glass nanopipettes

Adenosine triphosphate (ATP) is regarded as the “energy currency” in living cells, so real-time quantification of content variation of intracellular ATP is highly desired for understanding some important physiological processes. Due to its single-molecule readout ability, nanopipette sensing has emerged as a powerful technique for molecular sensing. In this study, based on the effect of targeting-aptamer binding on ionic current, and fluorescence resonance energy transfer (FRET), we reported a dual-signal readout nanopipette sensing system for monitoring ATP content variation at the subcellular level. In the presence of ATP, the complementary DNA-modified gold nanoparticles (cDNAs-AuNPs) were released from the inner wall of the nanopipette, which leads to sensitive response variations in ionic current rectification and fluorescence intensity. The developed nanopipette sensor was capable of detecting ATP in single cells, and the fluctuation of ATP content in the differentiation of dental pulp stem cells (DPSCs) was further quantified with this method. The study provides a more reliable nanopipette sensing platform due to the introduction of fluorescence readout signals. Significantly, the study of energy fluctuation during cell differentiation from the perspective of energy metabolism is helpful for differentiation regulation and cell therapy.

An electrochemiluminescence microsensor based on DNA-silver nanoclusters amplification for detecting cellular adenosine triphosphate.

Adenosine triphosphate (ATP), as the primary energy source, plays vital roles in many cellular events. Developing an efficient assay is crucial to rapidly evaluate the level of cellular ATP. A portable and integrated electrochemiluminescence (ECL) microsensor array based on a closed bipolar electrode (BPE) was presented. In the BPE unit, the ECL chemicals and oxidation/reduction were separated from the sensing chamber. The ATP aptamer was assembled with single-stranded DNA (ssDNA) in the sensing chamber. ATP capture made the aptamer disassemble from the ssDNA and facilitated DNA-templated silver nanocluster (Ag NC) generation by the target-rolling circle amplification (RCA) reaction. The guanine-rich padlock sequence produced tandem periodic cytosine-rich sequences by the RCA, inducing Ag NC generation in the cytosine-rich region of the produced DNA strands through Ag+ reduction. The in situ Ag NC generation enhanced the circuit conductivity of the BPE and promoted the ECL reaction of [Ru(bpy)2dppz]2+/tripropylamine in the anodic reservoir. On this ECL microsensor, a good linear relationship of ATP was achieved ranging from 30 to 1000 nM. The ATP content in HepG2 cells was selectively and sensitively determined without complex pretreatment. The ATP amount of 25 cells could be successfully detected when a sub-microliter sample was loaded.

Cloning and functional analysis prohibitins protein-coding gene EuPHB1 in Eucommia ulmoides Oliver

As multifunctional proteins, prohibitins(PHBs) participate in many cellular processes and play essential roles in organisms. In this study, using rapid amplification of cDNA end (RACE) technology, EuPHB1 was cloned from Eucommia ulmoides Oliver (E. ulmoides). A subcellular localization assay preliminarily located EuPHB1 in mitochondria. Then EuPHB1 was transformed into tobacco, and phenotype analyses showed that overexpression of EuPHB1 caused leaves to become chlorotic and shrivel. Furthermore, genes related to hormone and auxin signal transduction, auxin binding, and transport, such as ethylene-responsive transcription factor CRF4-like and ABC transporter B family member 11-like, were significantly inhibited in response to EuPHB1 overexpression. Its overexpression disturbs the original signal transduction pathway, thus causing the corresponding phenotypic changes in transgenic tobacco. Indeed, such overexpression caused fading of palisade tissue and an increase in the number of certain mesophyll cells. It also increased adenosine triphosphate (ATP) synthase activity, mitochondrial membrane potential, ATP content, and reactive oxygen species (ROS) levels in cells. Our results suggest that EuPHB1 expression promotes cellular energy metabolism by accelerating the oxidative phosphorylation of the mitochondrial respiratory chain. Elevated levels of EuPHB1 in the mitochondria, which helps supply the extra energy required to support rapid rates of cell division.

Anti-Oxidative and Anti-Diabetic Effects of Electrolyzed Weakly Alkaline Reduced Water on Renal Proximal Tubular Epithelial Cells

Diabetes, a chronic metabolic disorder, affects glucose consumption in the body and leads to severe kidney damage, known as diabetic nephropathy, where oxidative stress (OS) plays a vital role in disease pathogenesis. Electrolyzed weakly alkaline reduced water (EWARW) is known to have anti-oxidative functions in vitro due to its bioactive properties. However, there is an increasing need for adequate research on EWARW-specific therapies to minimize the negative consequences of hyperglycemia-induced OS in the kidney. This study aimed to determine the anti-oxidative and anti-diabetic effects of EWARW at pH 8.5 on human kidney-2 (HK-2) cells. Here, HK-2 cells were treated with 71.68 mM glucose to induce OS and then treated with 10% tap water (TW)_7.7, electrolyzed alkaline reduced water (EARW)_9.5, or EARW_8.5 for 24 h. After treatment, cell viability, OS and apoptotic markers, glucose uptake, adenosine triphosphate (ATP) content, and inflammatory markers were assessed to determine the effect of EARW_8.5. Our results revealed that EARW_8.5 rescued cell viability in comparison to the high glucose (HG) only and TW_7.7 groups. In addition, EARW_8.5 was effective in reducing OS markers (ROS, NO, and CAT), apoptotic markers (cleaved caspase-3), mitogen-activated protein kinase (MAPK) markers (p-JNK and p-p38), diabetic markers (glucose uptake), and inflammatory markers (p-STAT1, p-STAT3, IL-1β, and IL-6). Furthermore, we found EARW_8.5 treatment enhanced GPx and ATP content in HG-induced HK-2 cells compared to the HG-only and TW_7.7 groups. Collectively, EARW_8.5 stabilized antioxidant defenses and modulated inflammation, improving cellular metabolism and rescuing HG-induced harmful effects in HK-2 cells.

Single-Cell Adenosine Triphosphate Content Monitoring during Hyperthermia Cell Death by Using Plasmonic Fluorescent Nanoflare.

Gold nanorods-based plasmonic photothermal therapy (AuNRs-PPTT) is a prospective anticancer approach in which AuNRs absorb near-infrared (NIR) light and convert it into heat, leading to cell death. Investigating the molecular energy metabolism of single cells, especially cancer cells, during the hyperthermia cell death process is therefore of great significance, as it can help us to better understand the photothermal lethal mechanism of cancer cells and design new photothermal probes more rationally. However, during the AuNRs-PPTT process, how the cells respond to heat stimulation and how their energy metabolism changes have rarely been studied. Herein, we selected adenosine triphosphate (ATP) as a target molecule, and by preparing a plasmonic and turn-on type fluorescent nanoprobe, we examined the ATP metabolism difference between cancerous cells and normal cells during the AuNRs-PPTT process. We found that the fluorescence intensity increased ∼60% after 5 min laser irradiation as compared to the initial intensity in single HeLa cells, but only ∼20% increase was observed for single H8 cells; obviously, the increase of ATP content in cancerous cells was notably higher than that in normal cells during the hyperthermia cell death.

Phoenix dactylifera L. Fruits Date Fruit Ameliorate Oxidative Stress in 3-NP Intoxicated PC12 Cells

Introduction: Date palm fruits (DFs) are reported to possess antimutagenic, antiviral, antifungal, and anti-inflammatory properties. Effect of date fruits in the management of Huntington’s is yet to be studied. Methods: The protective effects of DF were measured in terms of superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities and reduced glutathione (rGSH), malondialdehyde (MDA) and nitrate/nitrite (NO2/NO3) content in cells. Cellular adenosine triphosphate (ATP) content was also measured. Cytotoxicity assay revealed that DF has the ability to protected cellular viability against 3-NP intoxication. Results: DFs increased the SOD and GPx activities and rGSH content. On the other hand, DF decreased MDA and NO2/NO3 levels in 3-NP intoxicated cells. Interestingly, DF increased ATP content in pheochromocytoma (PC) cells. Conclusion: DF has the ability to encounter 3-NP intoxication induced biochemical changes and improves cellular ATP contents, hence may be an interestingly candidate for further investigations.

Endometriosis may be associated with mitochondrial dysfunction in cumulus cells from subjects undergoing in vitro fertilization-intracytoplasmic sperm injection, as reflected by decreased adenosine triphosphate production

To determine whether endometriosis is associated with mitochondrial dysfunction in cumulus (granulosa [GC]) cells of subjects undergoing IVF-intracytoplasmic sperm injection (ICSI). Prospective cohort study. An IVF clinic in a tertiary academic care center. Eleven women with endometriosis and 39 controls. None. Cumulus cell adenosine triphosphate (ATP) levels, mitochondrial DNA (mtDNA), and genomic DNA (gDNA) number. Cumulus cell ATP content was 65% lower in subjects with surgically proven endometriosis (median 312.5 attomoles/ng total DNA, interquartile range = 116.0-667.8) compared with controls (median 892.4 attomoles/ng total DNA, interquartile range = 403.0-1,412.2). There was no significant difference in mtDNA:gDNA ratio. There were no significant differences in age, body mass index (BMI), basal serum FSH level, total oocyte number, metaphase II (M2) oocyte number, metaphase I oocyte number, percentage of M2 oocytes, fertilization rate, implantation rate, or pregnancy rate (PR). Multivariate regression analysis showed significant positive correlations between ATP and [1] M2 oocyte number (r = 0.307) and [2] pregnancy (r = 0.332). There were also trends toward positive correlations between ATP and [3] age (r = 0.283), [4] total number of oocytes (r = 0.271), [5] percentage of M2 oocytes (r = 0.249), and [6] implantation rate (r = 0.293). There were no statistically significant correlations between mtDNA:gDNA ratio and any demographic factors or clinical outcomes measured. Surgically confirmed endometriosis may be associated with cumulus cell mitochondrial dysfunction in subjects undergoing IVF-ICSI for infertility, as reflected by decreased ATP production.

Effect and Mechanism of 808 nm Light Pretreatment of Hypoxic Primary Neurons

This study investigated the effect of low intensity 808 nm light pretreatment of hypoxic primary neurons. Cobalt chloride (CoCl2) has been used to induce hypoxic injury in primary mouse cortical neurons. Low intensity 808 nm light was from light-emitting diode (LED). Cells were randomly divided into 4 groups: normal control group, CoCl2-induced group, CoCl2-induced group with 808 nm light irradiation pretreatment, and normal group with 808 nm light irradiation pretreatment. Effect of low intensity 808 nm light on neuronal morphology has been observed by microscope. MTT colorimetric assay has been used to detect the effect of low intensity 808 nm light on neuronal activity. Adenosine triphosphate (ATP) concentration and cytochrome C oxidase (COX) activity has been detected to study the effect of low intensity 808 nm light on neuronal mitochondria function. The results indicated that low intensity 808 nm light pretreatment alone did not affect cell viability, COX activity, and ATP content of neurons and low intensity 808 nm light pretreatment promoted the cell viability, COX activity, and ATP content of neurons with CoCl2exposure; however, low intensity 808 nm light pretreatment did not completely recover COX activity and cellular ATP content of primary neurons with CoCl2exposure to the level of the normal neurons.

Correlation between adenosine triphosphate content and apoptosis in liver of rats treated with alcohol.

Chronic alcohol consumption depresses adenosine triphosphate (ATP) synthesis and induces mitochondrial DNA (Mt-DNA) deletion. ATP content in cells may play a critical role in inducing cell death, apoptosis, or necrosis. However, it is unknown which type of cell death occurs in alcoholic liver disease. In this study, the deletions of hepatic Mt-DNA, hepatic ATP content, and the number of single-stranded DNA (ss-DNA) of hepatocytes in rats treated with ethanol were determined to elucidate the relationship among Mt-DNA deletion, ATP synthesis, and/or hepatic apoptosis. Sixteen male Wistar rats were fed with a liquid diet containing 36% ethanol (E group) or liquid diet without ethanol (C group) for 5 weeks. Hepatic ATP content was measured and the deletions of Mt-DNA encoding complexes I, IV, and V were determined in fresh liver tissue, and ss-DNA was stained in paraffin sections. Fatty change was observed in the E group, but not in the C group. Hepatic ATP content in the E group was 0.44 micromol/g of liver, which was significantly lower than that in the C group (0.84 micromol/g of liver). However, no deletion of Mt-DNA encoding complexes I, IV, and V was detected in either the E or the C group. ss-DNA staining was clearly observed in the nuclei of hepatocytes in both groups. The number of ss-DNA-positive hepatocytes in the E group was 5.6 +/- 1.8/10,000 hepatocytes, which was significantly less than that in the C group: 20.6 +/- 4.8/10,000 hepatocytes. There was a positive correlation between hepatic ATP contents and the number of ss-DNA-positive cells. The results suggest that mitochondrial function, at least in part ATP synthesis, was depressed before the damage of Mt-DNA by chronic ethanol consumption. Chronic ethanol consumption may not be responsible for the apoptosis of hepatocytes.

Fatty acid-induced cytotoxicity: Differences in susceptibility between mdck cells and primary cultures of proximal tubular cells

We have compared the cytotoxicity of exogenously added fatty acid (oleic acid) and that of endogenous free fatty acids released from cell membranes by phospholipase A 2 in primary cultures of mouse proximal tubular (MPT) cells and in Madine-Darby canine kidney (MDCK) cells. Exposure of MPT cell monolayers to oleic acid (125 mmol/L) for 2 hours resulted in severe irreversible injury to 70% ± 4% of MPT cells. In striking contrast, only 8% ± 3% of MDCK cells were killed by the same insult. This striking difference in the response to exogenous oleate by MPT and MDCK cells was associated with modest and comparable reductions in cell adenosine triphosphate (ATP) content in both cell types. Chemical anoxia induced by cyanide plus deoxyglucose (CN-DOG) in the absence of glucose was associated with greater injury in MPT cells (45% ± 6% killed) than in MDCK cells (16% ± 5% cells killed) despite severe and comparable depletion of cell ATP content in both MPT cells (96.0% ± 0.6% reduction) and MDCK cells (96.0% ± 0.5% reduction). The release of endogenous fatty acids by the exposure of cells to exogenous phospholipase A 2 caused mild injury in both cell types that was more severe in MPT cells than in MDCK cells. The combined insult of phospholipase A 2 and chemical anoxia for 2 hours caused substantially greater cell injury in both MPT and MDCK cells than either intervention alone, but the combined insult was still more damaging to MPT cells (73% ± 4% killed) than to MDCK cells (30% ± 4% killed). We conclude that the cell membrane in MDCK cells is intrinsically more resistant to fatty acid-induced injury than the lipid membrane in MPT cells.

Oxygen deprivation and early myocardial contractile failure: A reassessment of the possible role of adenosine triphosphate

The precise mechanism responsible for early contractile failure after the onset of myocardial anoxia or ischemia has attracted speculation and controversy. The simple and attractive hypothesis that adenosine triphosphate (ATP) deficiency is responsible for this failure has often been dismissed on the basis of claims that there is only a small reduction in cell ATP content at a time when contractile activity is severely reduced. The premise of this article is that the changes in cell ATP content and distribution that theoretically should occur after oxygen depletion may not have been adequately considered and that previous measurements of cell ATP content may not have been carried out at the correct time. Using an isolated rat heart preparation and high speed freeze-clamping techniques it has been possible to demonstrate that a substantial decrease in myocardial ATP and creatine phosphate content occurs after the onset of anoxia but before the onset of contractile failure. Thus, during the first 5 seconds of anoxia contractile activity remains constant whereas ATP decreases by 25 percent and creatine phosphate by 50 percent. Thereafter, contractile failure occurs and the rate of utilization of high energy phosphates declines with the cell content at a plateau or possibly increasing. These results are assessed in the light of the dynamic changes in energy metabolism occurring in early anoxia and suggest that ATP depletion in a specific cell compartment may be the primary trigger for early contractile failure.

Utilization of adenosine triphosphate in rat mast cells during histamine release induced by the ionophore A23187.

The role of endogenous adenosine triphosphate (ATP) in histamine release from rat mast cells induced by the ionophore A23187 in vitro has been studied. 2 The amount of histamine released by calcium from rat mast cells primed with the ionophore A23187 was dependent on the ATP content of the mast cells. 3 In aerobic experiments a drastic reduction in mast cell ATP content was found during the time when histamine release induced by A23187 takes place. 4 Anaerobic experiments were performed with metabolic inhibitors (antimycin A, oligomycin, and carbonyl cyanide p-trifluorometroxyphenylnydrazone), which are known to block the energy-dependent calcium uptake by isolated mitochondria. The mast cell ATP content was reduced during A23187-induced histamine release under anaerobic conditions in the presence of glucose. This indicates an increased utilization of ATP during the release process. 5 The observations are consistent with the view that energy requiring processes are involved in ionophore-induced histamine release from rat mast cells although part of the ATP reduction in the aerobic experiments may be due to an uncoupling effect of calcium on the oxidative phosphorylation.

Detection of bacteriuria by luciferase assay of adenosine triphosphate.

A selective method for distinguishing bacterial and nonbacterial adenosine triphosphate (ATP) in clinical bacteriological specimens was studied. The method involved incubation of samples with the detergent Triton X-100 and the ATP-hydrolyzing enzyme apyrase. The incubation selectively destroyed ATP in suspensions of various human cells while not affecting the ATP content in microbial cells. ATP remaining in the sample after incubation was extracted in boiling buffer and assayed by the firefly luciferase assay. Application of the method to 469 clinical urine specimens showed that the ATP level after treatment with Triton/apyrase was correlated to bacterial counts and that the sensitivity of the assay was sufficient for the detection of 10(5) bacteria/ml. The ATP levels per bacterial cell remaining in the urine specimen after treatment with Triton/apyrase were close to values observed in laboratory-grown cultures. The specificity and sensitivity of the luciferase assay for the detection of urinary bacteria and its possible use as a bacteriuria screening method are discussed.