Adenosine Triphosphate Content Research Articles

Involvement of oxidative stress-AMPK-Cx43-NLRP3 pathway in extracellular matrix remodeling of gastric smooth muscle cells in rats with diabetic gastroparesis.

This study aimed to investigate the changes in oxidative stress, adenosine monophosphate-activated protein kinase (AMPK), connexin43 (Cx43), nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) expression, and extracellular matrix (ECM) in the gastric smooth muscle tissues of rats with diabetic gastroparesis (DGP) and high glucose-cultured gastric smooth muscle cells, determine the existence of oxidative stress-AMPK-Cx43-NLRP3 pathway under high glucose condition, and the involvement of this pathway in ECM remodeling in DGP rats. The results showed that with increasing duration of diabetes, oxidation stress levels gradually increased, the AMPK activity decreased first and then increased, NLRP3, CX43 expression, and membrane/cytoplasmratio of Cx43 expression were increased in the gastric smooth muscle tissues of diabetic rats. Changes in ECM of gastric smooth muscle cells wereobserved in DGP rats. The DGP group showed higher collagen type I content, increased expression of Caspase-1, transforming growth factor-beta 3 (TGF-β3), and matrix metalloproteinase-2 (MMP-2), decreased tissue inhibitor of metalloproteinase-1 (TIMP-1) expression, and higher interleukin-1betacontent when compared with the control group. For gastric smooth muscle cells cultured under higher glucose, the MMP-2 and TGF-β3 expression wasdecreased, TGF-β1 and TIMP-1 expression was increased, the interleukin-1betacontent was decreased in cells after inhibition of NLRP3 expression; the NLRP3 and Caspase-1expression was decreased, and adenosine triphosphate content was lower after inhibition of Cx43; the expression of NLRP3, Caspase-1, P2X7, and the membrane/cytoplasm ratio of CX43 expression was decreased in cells after inhibition of AMPK and oxidative stress, the phospho-AMPK expression was also decreased after suppressing oxidative stress. Our findings suggest that high glucose induced the activation of the AMPK-Cx43-NLRP3 pathway through oxidative stress, and this pathway was involved in the ECM remodeling of gastric smooth muscles in DGP rats by regulating the biological functions of TGF-β3, TGF-β1, MMP-2, and TIMP-1.

Nanopore single molecule ATP detection based on the dual effects of specific capturing and signal amplification

Adenosine triphosphate (ATP) plays a crucial role in energy metabolism. Some serious diseases such as Alzheimer’s disease are related to abnormal ATP concentrations. Fast, sensitive, and specific ATP detection is important and challenging in disease diagnosis and biomedical research. In present study, conical nanopore was designed and employed to detect ATP at single molecule level. For overcoming the inherent defects such as non-selectivity and low sensitivity of nanopore in sensing small target molecule, two collaborative improvements of specific identification and signal amplification were proposed. We first used split aptamer capture probes (H1 and S1) to anchor ATP molecule, and then used auxiliary probes (H2 and H3) to form a long-nicked duplex ATP-DNA concatemer by hybridization chain reaction (HCR). Referring ATP-DNA concatemer rather than native ATP to nanopore sensor, characteristic current signals with high signal-to-noise ratio were obtained. By such indirect way, specific and sensitive ATP detection was realized with a linear range from 20 pM to 5 nM and a detection limit of 3.9 pM (S/N = 3). Additionally, the sensor exhibited many merits such as simplify, anti-interference, and especially allowing ATP detection in real sample. So, we believe that the nanopore sensor has good application prospects in screening and management of ATP-related diseases.

A Vacancy-Engineering Ferroelectric Nanomedicine for Cuproptosis/Apoptosis Co-Activated Immunotherapy.

Low efficacy of immunotherapy due to the poor immunogenicity of most tumors and their insufficient infiltration by immune cells highlights the importance of inducing immunogenic cell death and activating immune system for achieving better treatment outcomes. Herein, ferroelectric Bi2CuO4 nanoparticles with rich copper vacancies (named BCO-VCu) are rationally designed and engineered for ferroelectricity-enhanced apoptosis, cuproptosis, and the subsequently evoked immunotherapy. In this structure, the suppressed recombination of the electron-hole pairs by the vacancies and the band bending by the ferroelectric polarization lead to high catalytic activity, triggering reactive oxygen species bursts and inducing apoptosis. The cell fragments produced by apoptosis serve as antigens to activate T cells. Moreover, due to the generated charge by the ferroelectric catalysis, this nanomedicine can act as "a smart switch" to open the cell membrane, promote nanomaterial endocytosis, and shut down the Cu+ outflow pathway to evoke cuproptosis, and thus a strong immune response is triggered by the reduced content of adenosine triphosphate. Ribonucleic acid transcription tests reveal the pathways related to immune response activation. Thus, this study firstly demonstrates a feasible strategy for enhancing the efficacy of immunotherapy using single ferroelectric semiconductor-induced apoptosis and cuproptosis.

Transcriptomics coupled with proteomics reveals osimertinib-induced myocardial mitochondrial dysfunction

Osimertinib, an irreversible epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) used for cancer treatment, can cause significant cardiac toxicity. However, the specific mechanism of osimertinib-induced cardiotoxicity is not fully understood. In this study, we administered osimertinib to mice and neonatal rat ventricular myocytes (NRVMs). We observed significant structural and functional damage to the hearts of these mice, along with a marked increase in cardiac injury biomarkers and accompanying ultrastructural damage to mitochondria. We integrated 4D label-free protein quantification and RNA-Seq methods to analyze the sequencing data of NRVMs under osimertinib treatment (0 and 2.5 μM). Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis evidenced that differentially expressed genes (DEGs)and differentially expressed proteins (DEPs) were distinctly enriched for oxidative phosphorylation (OXPHOs). Simultaneously, osimertinib primarily affected the contents of adenosine triphosphate (ATP). Further investigations revealed that osimertinib disrupts the functions of the ATP synthase (complex V), leading to a reduction in ATP production. Taken together, our data demonstrated that osimertinib causes mitochondrial dysfunction, which in turn leads to the onset of cardiac toxicity.

AMP, ADP, and ATP Concentrations Differentially Affected by Meat Processing, Manufacturing, and Nonmeat Ingredients

Given its presence in a wide spectrum of soils relevant to food process hygiene, the biological metabolite adenosine triphosphate (ATP) is used as a target for surface hygiene assessments in food processing facilities. Yet, ample evidence demonstrates that ATP is depleted into adenosine di- (ADP) and monophosphate (AMP) homologs resulting in a loss of sensitivity for ATP-based hygiene assays. Yet, there are few studies that denote the degree of these shifts under routine processing conditions such as those encountered during various meat processing steps that may likely alter redox potential and adenosine profiles (e.g., tissue/cellular disruption, application of reducing additives, fermentation, or thermal treatment steps). In this study, meat samples were collected from homogenized beef tissue treated with nonmeat ingredients (sodium chloride, sodium nitrite, sodium erythorbate, natural smoke condensate, and sodium acid pyrophosphate) during manufacture at predetermined steps, and from retail meat products purchased from local markets. Concentrations of ATP, ADP, AMP, and AXP (sum concentration of all homologs) in a lab setting and in situ meat processing venues were determined and compared. Greater differences in AXP were seen during manufacture, where ADP generally comprised ∼90% as a mole fraction of AXP across all treatments, with the exception of the final cook step where AMP predominated. ATP concentrations averaged 2 log values lower than ADP and AMP. Adenosine profiles in retail samples followed similar trends with minimal ATP concentrations with ADP predominant in uncooked samples and AMP predominant in cooked samples. Resultingly, meat processing steps during product manufacture will alter AXP-reliant test sensitivities which should be considered when such technologies are utilized for hygiene verification in meat processing.

Oral supplementation with Scabiosa artropurperea L. aqueous extract improves the invivo sexual behaviour, sperm quality and androgen level in rams.

The effects of an aqueous extract of Scabiosa atropurpurea L. (AES) on the reproduction potential of Queue Fine de l'Ouest rams were evaluated over 9 weeks. Eighteen mature (4-6 years old) rams (52.8 ± 2.6 kg) were divided into three groups. The control (C) group was fed oat hay adlibitum with 700 g of concentrate and the other two groups were fed the same diet supplemented with AES at 1 and 2 mg/kg body weight (AES1 and AES2, respectively). Ram sperm was collected with an artificial vagina (2 × 2 days/week) to evaluate sperm production and quality, antioxidant activity, the adenosine triphosphate (ATP) and calcium concentrations. Sexual behaviour and plasma testosterone concentrations were also investigated. The administration of AES improved sexual behaviour (the duration of contact and the number of lateral approaches). The addition of AES also improved individual spermatozoa motility (C: 71.7% ± 6.3%; AES1: 78.3% ± 4.9%; AES2: 83.8% ± 4.4%), the sperm concentration (C: 5.6 ± 0.36; AES1: 6.4 ± 0.81; AES2: 6.7 ± 0.52 × 109 spermatozoa/mL), the ATP ratio (C: 1 ± 0.08; AES1: 2.1 ± 0.08; AES2: 3.3 ± 0.08) and the calcium concentration (C: 5.6 ± 0.24; AES1: 7.7 ± 0.21; AES2: 8.1 ± 0.24 mmol/L). AES treatment decreased the percentage of abnormal sperm (C: 18.5% ± 1.2%; AES1: 16.2% ± 1.1%; AES2: 14.8% ± 0.94%) and DNA damage (C: 62%; AES1: 27%; AES2: 33%) and was associated with elevated seminal fluid antioxidant activity (C: 22 ± 0.27; AES1: 27.1 ± 1.08 and AES2: 27.5 ± 0.36 mmol Trolox equivalents/L) and plasma testosterone (C: 8.3 ± 0.7; AES1: 11.7 ± 0.4; AES2: 15 ± 0.7 ng/L). In conclusion, our study suggests that S. atropurpurea may be potentially useful to enhance libido and sperm production and quality in ram.

AKR1B10 accelerates glycolysis through binding HK2 to promote the malignant progression of oral squamous cell carcinoma

BackgroundOral squamous cell carcinoma (OSCC) remains a rampant oral cavity neoplasm with high degree of aggressiveness. Aldo–keto reductase 1B10 (AKR1B10) that is an oxidoreductase dependent on nicotinamide adenine dinucleotide phosphate (NADPH) has been introduced to possess prognostic potential in OSCC. The present work was focused on specifying the involvement of AKR1B10 in the process of OSCC and its latent functional mechanism.MethodsAKR1B10 expression in OSCC tissues and cells were detected by RT-qPCR and Western blot analysis. CCK-8 method, EdU staining, wound healing and transwell assays respectively assayed cell viability, proliferation, migration and invasion. Immunofluorescence staining and Western blot evaluated epithelial mesenchymal transition (EMT). Adenosine triphosphate (ATP) contents, glucose consumption and extracellular acidification rate (ECAR) were measured by relevant commercially available kits and Seahorse XF96 Glycolysis Analyzer, severally. The expressions of proteins associated with metastasis and glycolysis were examined with Western blot. Co-IP assay confirmed the binding between AKR1B10 and hexokinase 2 (HK2).ResultsIt was observed that AKR1B10 expression was increased in OSCC tissues and cells. After AKR1B10 was knocked down, the proliferation, migration, invasion and EMT of OSCC cells were all hampered. Additionally, AKR1B10 silencing suppressed glycolysis and bound to HK2 in OSCC cells. Up-regulation of HK2 partially abolished the hampered glycolysis, proliferation, migration, invasion and EMT of AKR1B10-silenced OSCC cells.ConclusionTo sum up, AKR1B10 could bind to HK2 to accelerate glycolysis, thereby facilitating the proliferation, migration, invasion and EMT of OSCC cells.

Effect of Elamipretide on the Vitrification of Mouse Ovarian Tissue by Freezing.

The importance of ovarian cortical cryopreservation in fertility preservation is receiving increasing attention from reproductive specialists, and mitochondrial dysfunction is an important cause of reduced ovarian tissue cryopreservation. Elamipretide (SS-31) is a novel mitochondria-targeted antioxidant. However, whether it has a protective effect on mouse ovarian tissue cryopreservation remains to be studied. In this study, we examined follicular morphology and viability, mitochondrial function and oxidative stress levels, apoptosis, and culture in vitro after vitrification cryoresuscitation operation by treating ovarian tissues with SS-31 in cryoprotectant resuscitation solution. At the end of the experiment, the addition of 100 μmol/L SS-31 significantly improved follicle quality and oocyte maturation rate in vitro (p < 0.05) and significantly reduced apoptosis (p < 0.05) and oxidative stress levels (superoxide dismutase, catalase, malondialdehyde, p < 0.05). Meanwhile, mitochondrial respiratory chain complex enzyme activity, mtDNA copy number (p < 0.05), and adenosine triphosphate (p < 0.05) content were significantly increased in the 100 μmol/L SS-31-treated group. In addition, the mRNA expression levels of mitochondrial energy metabolism- and biosynthesis-related genes (STRT1, PGC-1a, PPAR-a, TFAM, p < 0.05) were markedly upregulated (p < 0.05) in the 100 μmol/L SS-31 group. In conclusion, SS-31 improved the cryopreservation of ovarian tissues, and 100 μmol/L SS-31 was found to be the most effective.

Cellular adenylate energy charge and adenine nucleotides in brain tissue during hypoglycemia in newly born BALB/c mice pups

Aim: Hypoglycemia occurs in the neonatal period but the exact pathophysiology of the resulting brain injury at the cellular level is not well known. Therefore, a neonatal murine model was developed with insulin-induced hypoglycemia, to analyze the in-vitro effects of hypoglycemia on brain nucleotides and adenylate energy charge (AEC) throughout the first ten days of life. Methods: Newly born BALB/c pups between one and ten days of age were used. In each age group, six pups were subjected to insulin-induced hypoglycemia and six others served as controls. In both groups, immediately after euthanasia, brain tissues were collected. The in-vitro effects of hypoglycemia on brain nucleotides [adenosine monophosphate (AMP), adenosine diphosphate (ADP), and adenosine triphosphate (ATP)] were analyzed using liquid chromatography with tandem mass spectrometry (LC-MS/MS) as well on AEC. Results: In the controls, the cellular AEC steadily decreased with age by at least 50% over the 10-day study period (P &lt; 0.05) except in the parietal tissue (P = 0.30) where it remained stable throughout that period. The most marked decrease was observed in the occipital tissue (P &lt; 0.001). In the hypoglycemic mice, AEC in both the parietal and occipital tissues decreased significantly more than in the controls, more rapidly and pronounced between day 2 and 5 in the occipital tissue, reaching very low levels from day 5 onward. Except in the occipital tissue, none of the adenine nucleotides on its own, including ATP, reflected the cellular AEC. Conclusions: Over the first ten days of life, hypoglycemia progressively depleted cellular AEC in the brain, unlike cellular ATP concentration which did not appropriately reflect cellular energy.

Enhancement of skin rejuvenation and hair growth through novel near-infrared light emitting diode (nNIR) lighting: in vitro and in vivo study

The study aimed to explore the impact of a novel near-infrared LED (nNIR) with an extended spectrum on skin enhancement and hair growth. Various LED sources, including White and nNIRs, were compared across multiple parameters: cytotoxicity, adenosine triphosphate (ATP) synthesis, reactive oxygen species (ROS) reduction, skin thickness, collagen synthesis, collagenase expression, and hair follicle growth. Experiments were conducted on human skin cells and animal models. Cytotoxicity, ATP synthesis, and ROS reduction were evaluated in human skin cells exposed to nNIRs and Whites. LED irradiation effects were also studied on a UV-induced photoaging mouse model, analyzing skin thickness, collagen synthesis, and collagenase expression. Hair growth promotion was examined as well. Results revealed both White and nNIR were non-cytotoxic to human skin cells. nNIR enhanced ATP and collagen synthesis while reducing ROS levels, outperforming the commonly used 2chip LEDs. In the UV-induced photoaging mouse model, nNIR irradiation led to reduced skin thickness, increased collagen synthesis, and lowered collagenase expression. Additionally, nNIR irradiation stimulated hair growth, augmented skin thickness, and increased hair follicle count. In conclusion, the study highlighted positive effects of White and nNIR irradiation on skin and hair growth. However, nNIR exhibited superior outcomes compared to White. Its advancements in ATP content, collagen synthesis, collagenase inhibition, and hair growth promotion imply increased ATP synthesis activity. These findings underscore nNIR therapy’s potential as an innovative and effective approach for enhancing skin and promoting hair growth.

Influence of trimetazidine on myocardial injury in mice with diabetic cardiomyopathy

IntroductionThe prevalence of diabetes mellitus is increasing year by year globally, and diabetic cardiomyopathy (DCM), as the most common complication of type 2 diabetes mellitus, seriously affects the prognosis of patients. Trimetazidine (TMZ), as a drug affecting myocardial energy metabolism, mainly reduces the oxidation rate of β-oxidation by inhibiting 3-ketoacyl-CoA thiolase (3-KAT), a key enzyme in β-oxidation of free fatty acid (FFA), so that the energy metabolism substrate of cardiomyocytes preferentially selects glucose rather than fatty acids, increases the content of intracellular adenosine triphosphate (ATP), enhances the contractile function of cardiomyocytes, and improves the state of cellular ischemia and hypoxia. Previous studies have shown that TMZ is closely related to the activation and induction of apoptosis of the MAPK pathway and AMPK pathway, and plays a role in the treatment of diabetic cardiomyopathy, but the specific mechanism is still unclear. ObjectiveThis study aims to investigate the impact of TMZ on myocardial damage in mice exhibiting diabetic cardiomyopathy (DCM), and to furnish a laboratory foundation for the clinical treatment of diabetic cardiomyopathy. MethodMale db/db mice (6 weeks old, n = 21) and male wild-type (wt) (6 weeks old, n = 20) mice were selected for the study. The wt mice were randomly assigned to the wt group (n = 10) and wt + TMZ group (n = 10), while the remaining db/db mice were randomly allocated to the db/db group (n = 11) and db/db + TMZ group (n = 10). Following 8 weeks of feeding, the wt + TMZ group and db/db + TMZ group received TMZ via gavage, whereas the remaining groups were administered physiological saline. Periodic measurements of blood glucose, blood lipids, and myocardial enzymes were conducted in mice, with samples obtained after the 12th week for subsequent biochemical analysis, myocardial pathology assessment, immunohistochemistry, western blot analysis, and TUNEL staining (TdT-mediated dUTP Nick-End Labeling). ResultGLU, TC, TG, LDL-C, and CK-MB levels were significantly higher in db/db mice compared to wt mice (GLU: M ± SD wt 5.94 ± 0.37, db/db 17.63 ± 0.89, p < 0.05, ES = 0.991; TC: M ± SD wt 3.01 ± 0.32, db/db 6.97 ± 0.36, p < 0.05, ES = 0.972; TG: M ± SD wt 0.58 ± 0.2, db/db 1.75 ± 0.14, p < 0.05, ES = 0.920; LDL-C: M ± SD wt 1.59 ± 0.12, db/db 3.87 ± 0.14, p < 0.05, ES = 0.989; CK-MB: M ± SD wt 0.12 ± 0.01, db/db 0.31 ± 0.04, p < 0.05, ES = 0.928). HDL-C levels were significantly lower in db/db mice (M ± SD wt 1.89 ± 0.08, db/db 0.64 ± 0.09, p < 0.05, ES = 0.963). Histopathological analysis confirmed myocardial damage in db/db mice. Treatment with TMZ reduced GLU, TC, TG, LDL-C, and CK-MB levels (p < 0.05, ES > 0.9) and increased HDL-C levels compared to untreated db/db mice. Additionally, TMZ treatment significantly decreased myocardial cell apoptosis (p < 0.05, ES = 0.980). These results demonstrate the efficacy of TMZ in reversing myocardial injury in DCM mice. ConclusionTMZ can mitigate myocardial damage in db/db mice by downregulating the expression of caspase-12, a protein associated with the endoplasmic reticulum stress (ERS) cell apoptosis pathway, consequently diminishing cell apoptosis. This underscores the protective efficacy of TMZ against myocardial damage in mice afflicted with DCM.

Development of a rhodamine-based fluorescent probe for ATP detection for potential applications in meat freshness assessment

Adenosine triphosphate (ATP) plays a vital role in physiological processes and is an essential indicator of microbial content in food. Herein, a new sensitive, rapid and water-soluble probe for ATP detection was developed. Rhodamine B and pentaethylenehexamine were employed to design and synthesise the probe rhodamine-pentaethylenehexamine (RP) for selective ATP detection. The synthesised probe RP was characterized using Fourier transform infrared, NMR and dynamic light scattering size distributions. Upon the addition of ATP, the probe exhibited a distinct change in fluorescence intensity, with fluorescence emission at 580 nm. A linear relationship was observed between fluorescence intensity and ATP concentrations at 0–50 μmol/L, with a limit of detection of 10.97 × 10−9 mol/L. The results of the zeta potential and molecular dynamics simulation demonstrated that the detection mechanism of the probe RP is associated with the electrostatic adsorption interaction between the multi-positively charged sites of RP and the negatively charged triphosphate structure of ATP. Our study provides new insights into improving charge site identification in small molecule detection. Furthermore, the successful detection of ATP on meat surfaces indicates that RP has the potential to assess meat freshness.

Astragaloside IV combined with ligustrazine ameliorates abnormal mitochondrial dynamics via Drp1 SUMO/deSUMOylation in cerebral ischemia-reperfusion injury.

Astragaloside IV (AST IV) and ligustrazine (Lig), the main ingredients of Astragali Radix and Chuanxiong Rhizoma respectively, have demonstrated significant benefits in treatment of cerebral ischemia -reperfusion injury (CIRI); however, the mechanisms underlying its benificial effects remain unclear. SUMO-1ylation and deSUMO-2/3ylation of dynamin-related protein 1 (Drp1) results in mitochondrial homeostasis imbalance following CIRI, which subsequently aggravates cell damage. This study investigates the mechanisms by which AST IV combined with Lig protects against CIRI, focusing on the involvement of SUMOylation in mitochondrial dynamics. Rats were administrated AST IV and Lig for 7 days, and middle cerebral artery occlusion was established to mimic CIRI. Neural function, cerebral infarction volume, cerebral blood flow, cognitive function, cortical pathological lesions, and mitochondrial morphology were measured. SH-SY5Y cells were subjected to oxygen-glucose deprivation/reoxygenation (OGD/R) injury. Mitochondrial membrane potential and lactic dehydrogenase (LDH), reactive oxygen species (ROS), and adenosine triphosphate (ATP) levels were assessed with commercial kits. Moreover, co-immunoprecipitation (Co-IP) was used to detect the binding of SUMO1 and SUMO2/3 to Drp1. The protein expressions of Drp1, Fis1, MFF, OPA1, Mfn1, Mfn2, SUMO1, SUMO2/3, SENP1, SENP2, SENP3, SENP5, and SENP6 were measured using western blot. In rats with CIRI, AST IV and Lig improved neurological and cognitive functions, restored CBF, reduced brain infarct volume, and alleviated cortical neuron and mitochondrial damage. Moreover, in SH-SY5Y cells, the combination of AST IV and Lig enhanced cellular viability, decreased release of LDH and ROS, increased ATP content, and improved mitochondrial membrane potential. Furthermore, AST IV combined with Lig reduced the binding of Drp1 with SUMO1, increased the binding of Drp1 with SUMO2/3, suppressed the expressions of Drp1, Fis1, MFF, and SENP3, and increased the expressions of OPA1, Mfn1, Mfn2, SENP1, SENP2, and SENP5. SUMO1 overexpression promoted mitochondrial fission and inhibited mitochondrial fusion, whereas SUMO2/3 overexpression suppressed mitochondrial fission. AST IV combined with Lig could reverse the effects of SUMO1 overexpression while enhancing those of SUMO2/3 overexpression. This study posits that the combination of AST IV and Lig has the potential to reduce the SUMO-1ylation of Drp1, augment the SUMO-2/3ylation of Drp1, and thereby exert a protective effect against CIRI.

Mitochondrial Transplantation Alleviates Doxorubicin-Induced Toxicity in Rat Renal Cells

Background: Doxorubicin (DOX) is used in the treatment of various cancers and has good effectiveness. However, its therapeutic use is limited due to its effects on various organs and healthy cells. Doxorubicin can affect the kidneys and cause toxicity. Evidence shows that DOX induces nephrotoxicity through oxidative stress. Objectives: In this research, we examined the effect of mitochondrial transplantation on improving mitochondrial and cellular toxicity caused by DOX on renal proximal tubular cells (RPTCs). Methods: The research measured 7 toxicity parameters, including cell lysis, reactive oxygen species (ROS) formation, mitochondrial membrane potential (MMP) decline, GSH and GSSG content, lipid peroxidation (LPO), adenosine triphosphate (ATP) content, and Caspase-3 activity (the final mediator of apoptosis). Active fresh mitochondria were prepared from Wistar rat kidney. Results: The findings indicated that DOX caused cytotoxicity in RPTCs. Additionally, DOX induced oxidative stress by increasing the level of reactive oxygen species, reducing glutathione content, and elevating lipid peroxidation. Moreover, it led to damage to the mitochondrial membrane, increased caspase-3 activity, and decreased ATP content. Mitochondrial transplantation, as a new therapeutic approach, reduced oxidative stress, mitochondrial membrane damage, and apoptosis caused by DOX in RPTCs. Furthermore, this therapeutic approach increased the ATP content in RPTCs. Conclusions: Our study suggests that this therapeutic approach could be helpful in the treatment of drug-induced nephrotoxicity.

Kaempferol Improves Exercise Performance by Regulating Glucose Uptake, Mitochondrial Biogenesis, and Protein Synthesis via PI3K/AKT and MAPK Signaling Pathways.

Kaempferol is a natural flavonoid with reported bioactivities found in many fruits, vegetables, and medicinal herbs. However, its effects on exercise performance and muscle metabolism remain inconclusive. The present study investigated kaempferol's effects on improving exercise performance and potential mechanisms in vivo and in vitro. The grip strength, exhaustive running time, and distance of mice were increased in the high-dose kaempferol group (p < 0.01). Also, kaempferol reduced fatigue-related biochemical markers and increased the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) related to antioxidant capacity. Kaempferol also increased the glycogen and adenosine triphosphate (ATP) content in the liver and skeletal muscle, as well as glucose in the blood. In vitro, kaempferol promoted glucose uptake, protein synthesis, and mitochondrial function and decreased oxidative stress in both 2D and 3D C2C12 myotube cultures. Moreover, kaempferol activated the PI3K/AKT and MAPK signaling pathways in the C2C12 cells. It also upregulated the key targets of glucose uptake, mitochondrial function, and protein synthesis. These findings suggest that kaempferol improves exercise performance and alleviates physical fatigue by increasing glucose uptake, mitochondrial biogenesis, and protein synthesis and by decreasing ROS. Kaempferol's molecular mechanism may be related to the regulation of the PI3K/AKT and MAPK signaling pathways.

2,4,6-trinitrotoluene causes mitochondrial toxicity in Caenorhabditis elegans by affecting electron transport

As a typical energetic compound widely used in military activities, 2,4,6-trinitrotoluene (TNT) has attracted great attention in recent years due to its heavy pollution and wide distribution in and around the training facilities, firing ranges, and demolition sites. However, the subcellular targets and the underlying toxic mechanism of TNT remain largely unknown. In this study, we explored the toxic effects of TNT biological reduction on the mitochondrial function and homeostasis in Caenorhabditis elegans (C. elegans). With short-term exposure of L4 larvae, 10–1000 ng/mL TNT reduced mitochondrial membrane potential and adenosine triphosphate (ATP) content, which was associated with decreased expression of specific mitochondrial complex involving gas-1 and mev-1 genes. Using fluorescence-labeled transgenic nematodes, we found that fluorescence expression of sod-3 (muls84) and gst-4 (dvls19) was increased, suggesting that TNT disrupted the mitochondrial antioxidant defense system. Furthermore, 10 ng/mL TNT exposure increased the expression of the autophagy-related gene pink-1 and activated mitochondrial unfolded protein response (mt UPR), which was indicated by the increased expression of mitochondrial stress activated transcription factor atfs-1, ubiquitin-like protein ubl-5, and homeobox protein dve-1. Our findings demonstrated that TNT biological reduction caused mitochondrial dysfunction and the development of mt UPR protective stress responses, and provided a basis for determining the potential risks of energetic compounds to living organisms.

Dual-Channel/Localization Single-Molecule Fluorescence Probe for Monitoring ATP and HOCl in Early Diagnosis and Therapy of Rheumatoid Arthritis.

Rheumatoid arthritis (RA), a common chronic inflammatory illness, is still incurable, reducing the sufferers' quality of life significantly. Adenosine 5'-triphosphate (ATP) and hypochlorous acid (HOCl) are key indicators in RA, but their precise mechanisms in RA pathophysiology are unknown. As a result, in order to detect ATP and HOCl simultaneously, we created two new dual-channel/localization single-molecule fluorescence probes, RhTNMB and RhFNMB. Furthermore, RhFNMB outperformed RhTNMB in terms of detection performance. ATP and HOCl produce independent fluorescence responses in the light red channel (λex = 520 nm, λem = 586 nm) and deep red channel (λex = 620 nm, λem = 688 nm), respectively, without spectral crosstalk. It should be noted that the probe RhFNMB successfully imaged ATP in mitochondria and HOCl in cells. Surprisingly, the probe RhFNMB demonstrated remarkable detection ability in the diagnosis and treatment of Pseudomonas aeruginosa-induced abdominal inflammation in mice. We continued to apply the probe RhFNMB to track ATP and HOCl in RA and discovered that ATP and HOCl concentrations were considerably greater in RA joints than in normal joints. We also confirmed the therapeutic effect of methotrexate on RA. This study is the first to achieve dual-channel imaging of ATP and HOCl, which is of great value for the early diagnosis and therapy of RA.

Abstract 5416: Purinergic P2Y6 receptor antagonists as potential anti tumor agents

Abstract Introduction: Cancer places a tremendous strain on society in both developed and economically less developed nations. It remains the most recurring and deadliest disease despite rapid advances in treatment modalities. The major contributors are the prevalence of known risk factors, urbanization, population growth and ageing. Cancer resurgence depends on the organ in which the tumor first appears, immune response, patient’s characteristics, disease stage and tumor microenvironment (TME). Among the several inflammatory molecules found within the TMEs, the levels of adenosine triphosphate (ATP) has been found to be high. In healthy tissue, the concentration of ATP in the extracellular space (eATP) is negligible and does not accumulate significantly due to the activity of ectonucleotidases. However, tissue injury, cellular stress and chemotherapy mediated cell death can increase the levels of ATP in the TME. The eATP exerts its effect mainly through purinergic P2 receptors expressed by most malignant cells. We have previously shown that eATP promoted migration and invasion of cancer cells in vitro through increased expression of the enzyme cyclooxygenase 2 (COX-2) (Sharma et al., 2021). We showed that eATP mediated COX-2 expression is through P2 receptors (Akter et al., 2021). Accordingly, we hypothesized that blocking the P2 receptors would abolish the COX-2-mediated migration of tumor cells and tested the same in an animal model. Methodology: Tumors were generated in 8-15-week-old male C57BL/6 mice (20-25 g weight) through subcutaneous injection of syngeneic EL4 lymphoma cells in the left flank of the mice. When tumors were palpable around day 10, saline or P2 receptor, antagonists were directly injected into the center of the tumor and mice were divided into three groups: saline (vehicle control), suramin (broad-spectrum P2 receptor antagonist), and MRS 2578 (P2Y6 receptor-specific antagonist). Drugs were chosen based on prior in vitro research and a vernier caliper was used to assess the growth of the tumor every day. Behavioral tests were performed on day 8 (prior to P2 receptor injections) and days 11, 14 and 17 post-tumor injection. Mice were continuously monitored for their health and were sacrificed on day 18 with deep CO2 anesthesia and cervical dislocation. Results: We found that suramin-injected tumors had a substantial reduction in tumor volume when compared to saline-injected ones. Intra-tumoral injections of P2 receptor antagonists such as MRS-2578 also resulted in similar outcomes as suramin. In tumors treated with P2 receptor antagonists, the expression of COX-2, cell cycle proteins and epithelial-to-mesenchymal (EMT) markers, which aid in metastasis, were significantly downregulated, indicating a suppressive effect. Therefore, we propose that P2 receptor-dependent anti-inflammatory drugs (PBAIDs) could be considered a potential alternative therapeutic option for preventing cancer recurrence. Citation Format: Khagendra Ghimeray, Shilpa Sharma, Ravi Shankar Akundi. Purinergic P2Y6 receptor antagonists as potential anti tumor agents [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5416.

An ATP-free packaging of T4 DNA

Packaging of viral DNA into a capsid with liquid crystalline density is a crucial step in viral reproduction. DNA packaging with an adenosine triphosphate (ATP)-fueled molecular motor is an established viral DNA packaging mechanism system. On the other hand, DNA is compacted by the conformational change attributable to multivalent cations exclusively at valences higher than three. The conformational change of DNA was not considered as a mechanism of DNA packaging of viruses. Here, T4 DNA, ejected from a capsid in the ambient concentration of phosphate corresponding to the intra-cell concentration, is packaged into the capsid when the phosphate concentration decreases to the extra-cell concentration in coexistence with divalent cations, that is, Ca2+ and Mg2+. The compaction and packaging processes coincide with the conformational change of DNA. Divalent cations can compact T4 DNA when the counter anion is phosphate. The DNA-packaged and re-generated virions showed equivalent infective ability with the original populations. Fluorescent microscopy distinguished the conformational changes of DNA between compact forms and coil forms. Packaged or unpackaged DNA was confirmed enzymatically. Plaque-forming unit (Pfu) was used as the measure of infectious ability of virions. The concentration of ATP was measured by the luminometric method. The packaging process was proceeded in picomolar or lower concentration of ATP. This series of procedures may constitute a new ATP-free or low-ATP approach to packaging viral DNA with DNA conformational change underpinning the process. The results from this study may disclose an undiscovered facet of T4 life cycle.

Impact of Zero-Valent Iron Nanoparticles and Ampicillin on Adenosine Triphosphate and Lactate Metabolism in the Cyanobacterium Fremyella diplosiphon.

In cyanobacteria, the interplay of ATP and lactate dynamics underpins cellular energetics; their pronounced shifts in response to zero-valent iron (nZVI) nanoparticles and ampicillin highlight the nuanced metabolic adaptations to environmental challenges. In this study, we investigated the impact of nZVIs and ampicillin on Fremyella diplosiphon cellular energetics as determined by adenosine triphosphate (ATP) content, intracellular and extracellular lactate levels, and their impact on cell morphology as visualized by transmission electron microscopy. While a significant increase in ATP concentration was observed in 0.8 mg/L ampicillin-treated cells compared to the untreated control, a significant decline was noted in cells treated with 3.2 mg/L nZVIs. ATP levels in the combination regimen of 0.8 mg/L ampicillin and 3.2 mg/L nZVIs were significantly elevated (p < 0.05) compared to the 3.2 mg/L nZVI treatment. Intracellular and extracellular lactate levels were significantly higher in 0.8 mg/L ampicillin, 3.2 mg/L nZVIs, and the combination regimen compared to the untreated control; however, extracellular lactate levels were the highest in cells treated with 3.2 mg/L nZVIs. Visualization of morphological changes indicated increased thylakoid membrane stacks and inter-thylakoidal distances in 3.2 mg/L nZVI-treated cells. Our findings demonstrate a complex interplay of nanoparticle and antibiotic-induced responses, highlighting the differential impact of these stressors on F. diplosiphon metabolism and cellular integrity.