Adenosine Triphosphate Activity Research Articles

Patient tissue-derived FGFR4-variant and wild-type colorectal cancer organoid development and anticancer drug sensitivity testing

ObjectivesFGFR4-variant and wild-type colorectal cancer (CRC) organoids were developed to investigate the effects of FGFR4-targeted drugs, including FGFR4-IN and erdafitinib, on CRC and their possible molecular mechanism. MethodsClinical CRC tissues were collected, seven CRC organoids were developed, and whole exome sequencing (WES) was performed. CRC organoids were cultured and organoid drug sensitivity studies were conducted. Finally, an FGFR4-variant (no wild-type) CRC patient-derived orthotopic xenograft mouse model was developed. Western blot measured ERK/AKT/STAT3 pathway-related protein levels. ResultsWES results revealed the presence of FGFR4-variants in 5 of the 7 CRC organoids. The structural organization and integrity of organoids were significantly altered under the influence of targeted drugs (FGFR4-IN-1 and erdafitinib). The effects of FGFR4 targeted drugs were not selective for FGFR4 genotypes. FGFR4-IN-1 and erdafitinib significantly reduced the growth, diameter, and Adenosine Triphosphate (ATP) activity of organoids. Furthermore, chemotherapeutic drugs, including 5-fluorouracil and cisplatin, inhibited FGFR4-variant and wild-type CRC organoid activity. Moreover, the tumor volume of mice was significantly reduced at week 6, and p-ERK1/2, p-AKT, and p-STAT3 levels were down-regulated following FGFR4-IN-1 and erdafitinib treatment. ConclusionsFGFR4-targeted and chemotherapeutic drugs inhibited the activity of FGFR4-variant and wild-type CRC organoids, and targeted drugs were more effective than chemotherapeutic drugs at the same concentration. Additionally, FGFR4 inhibitors hindered tumorigenesis in FGFR4-variant CRC organoids through ERK1/2, AKT, and STAT3 pathways. However, no wild-type control was tested in this experiment, which need further confirmation in the next study.

A deep learning-based approach for fully automated segmentation and quantitative analysis of muscle fibers in pig skeletal muscle

Muscle fiber properties exert a significant influence on pork quality, with cross-sectional area (CSA) being a crucial parameter closely associated with various meat quality indicators, such as shear force. Effectively identifying and segmenting muscle fibers in a robust manner constitutes a vital initial step in determining CSA. This step is highly intricate and time-consuming, necessitating an accurate and automated analytical approach. One limitation of existing methods is their tendency to perform well on high signal-to-noise ratio images of intact, healthy muscle fibers but their lack of validation on more complex image datasets featuring significant morphological changes, such as the presence of ice crystals. In this study, we undertake the fully automatic segmentation of muscle fiber microscopic images stained with myosin adenosine triphosphate (mATPase) activity using a deep learning architecture known as SOLOv2. Our objective is to efficiently derive accurate measurements of muscle fiber size and distribution. Tests conducted on actual images demonstrate that our method adeptly handles the intricate task of muscle fiber segmentation, yielding quantitative results amenable to statistical analysis and displaying reliability comparable to manual analysis.

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.

Abstract A044: P2Y2 signaling disruption reduces ATP-dependent calcium elevation and actin localization to promote breast tumor cell dissemination

Abstract The tumor microenvironment and wound healing after injury both contain extremely high concentrations of the extracellular signaling molecule, adenosine triphosphate (ATP) compared to normal tissue. P2Y2 receptor, an ATP-activated purinergic receptor, is typically associated with pulmonary, endothelial, and neural cell signaling. Here we report the importance of ATP-dependent P2Y2 signaling in breast epithelial cells and how it is altered in metastatic breast cancer. In response to ATP activation, P2Y2 receptor signaling causes an increase of intracellular calcium (Ca2+) in non-tumorigenic breast epithelial cells, while their tumorigenic and metastatic counterparts have significantly reduced Ca2+ responses. The non-tumorigenic cells respond to increased Ca2+ with actin polymerization and localization to cell edges, while the metastatic cells remained unaffected. The increase in intracellular Ca2+ after ATP stimulation was blunted using a P2Y2 antagonist, which also prevented actin mobilization and caused cell dissemination from spheroids in non-tumorigenic breast epithelial cells. Furthermore, the lack of Ca2+ concentration changes and actin mobilization in the metastatic breast cancer cells could be due to reduced P2Y2 expression, which correlates with poorer overall survival in breast cancer patients. This study elucidates rapid changes that occur after elevated intracellular Ca2+ in breast epithelial cells and how metastatic cancer cells have adapted to evade this cellular response. Citation Format: Makenzy Mull, Stephen Pratt, Keyata Thompson, David Annis, Julia Ju, Megan Stemberger, Liron Boyman, Michele Vitolo, W. Jonathan Lederer, Stuart Martin. P2Y2 signaling disruption reduces ATP-dependent calcium elevation and actin localization to promote breast tumor cell dissemination [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A044.

Glia-derived adenosine in the ventral hippocampus drives pain-related anxiodepression in a mouse model resembling trigeminal neuralgia

Glial activation and dysregulation of adenosine triphosphate (ATP)/adenosine are involved in the neuropathology of several neuropsychiatric illnesses. The ventral hippocampus (vHPC) has attracted considerable attention in relation to its role in emotional regulation. However, it is not yet clear how vHPC glia and their derived adenosine regulate the anxiodepressive-like consequences of chronic pain. Here, we report that chronic cheek pain elevates vHPC extracellular ATP/adenosine in a mouse model resembling trigeminal neuralgia (rTN), which mediates pain-related anxiodepression, through a mechanism that involves synergistic effects of astrocytes and microglia. We found that rTN resulted in robust activation of astrocytes and microglia in the CA1 area of the vHPC (vCA1). Genetic or pharmacological inhibition of astrocytes and connexin 43, a hemichannel mainly distributed in astrocytes, completely attenuated rTN-induced extracellular ATP/adenosine elevation and anxiodepressive-like behaviors. Moreover, inhibiting microglia and CD39, an enzyme primarily expressed in microglia that degrades ATP into adenosine, significantly suppressed the increase in extracellular adenosine and anxiodepressive-like behaviors. Blockade of the adenosine A2A receptor (A2AR) alleviated rTN-induced anxiodepressive-like behaviors. Furthermore, interleukin (IL)-17A, a pro-inflammatory cytokine probably released by activated microglia, markedly increased intracellular calcium in vCA1 astrocytes and triggered ATP/adenosine release. The astrocytic metabolic inhibitor fluorocitrate and the CD39 inhibitor ARL 67156, attenuated IL-17A-induced increases in extracellular ATP and adenosine, respectively. In addition, astrocytes, microglia, CD39, and A2AR inhibitors all reversed rTN-induced hyperexcitability of pyramidal neurons in the vCA1. Taken together, these findings suggest that activation of astrocytes and microglia in the vCA1 increases extracellular adenosine, which leads to pain-related anxiodepression via A2AR activation. Approaches targeting astrocytes, microglia, and adenosine signaling may serve as novel therapies for pain-related anxiety and depression.

Systemic Delivery of Paclitaxel by Find-Me Nanoparticles Activates Antitumor Immunity and Eliminates Tumors.

Local delivery of immune-activating agents has shown promise in overcoming an immunosuppressive tumor microenvironment (TME) and stimulating antitumor immune responses in tumors. However, systemic therapy is ultimately needed to treat tumors that are not readily locatable or accessible. To enable systemic delivery of immune-activating agents, we employ poly(lactic-co-glycolide) (PLGA) nanoparticles (NPs) with a track record in systemic application. The surface of PLGA NPs is decorated with adenosine triphosphate (ATP), a damage-associated molecular pattern to recruit antigen-presenting cells (APCs). The ATP-conjugated PLGA NPs (NPpD-ATP) are loaded with paclitaxel (PTX), a chemotherapeutic agent inducing immunogenic cell death to generate tumor antigens in situ. We show that the NPpD-ATP retains ATP activity in hostile TME and provides a stable "find-me" signal to recruit APCs. Therefore, the PTX-loaded NPpD-ATP helps populate antitumor immune cells in TME and attenuate the growth of CT26 and B16F10 tumors better than a mixture of PTX-loaded NPpD and ATP. Combined with anti-PD-1 antibody, PTX-loaded NPpD-ATP achieves complete regression of CT26 tumors followed by antitumor immune memory. This study demonstrates the feasibility of systemic immunotherapy using a PLGA NP formulation that delivers ICD-inducing chemotherapy and an immunostimulatory signal.

Adjustable luminescence copper nanoclusters nanoswitch based on competitive coordination of samarium ions for cascade detection of adenosine triphosphate and acid phosphatase activity.

Benefit from the strong coordination property, lanthanide metal ions have been used as competitive reagents to modulate the fluorescence changes of the system. However, lanthanide metal ions as inducers for aggregation-induced emission enhancement in nanosystems is rare. Herein, we report a "turn on-off-on" fluorescent switch for cascade detection of acid phosphatase (ACP) and adenosine triphosphate (ATP) based on the competitive coordination of samarium ions (Sm3+). Novel copper nanoclusters (CuNCs) with long wavelength emission (614nm) stabilized by glutathione (GSH) and glycylglycine (Gly-Gly) have been confirmed to have AIE property. With the continuous aggregation of GSH/Gly-Gly CuNCs under the induction of Sm3+, the fluorescence of the system increased to achieve the "turn-on" process. The coordinated behaviour between Sm3+ and GSH/Gly-Gly CuNCs is discussed. Due to the strong metal coordination ability of ATP, the Sm3+ coordinated with the GSH/Gly-Gly CuNCs is competed out, resulting in the fluorescence "turn-off" process of the system. As the substrate of enzymatic hydrolysis of ACP, with the continuous hydrolysis of ATP by ACP, Sm3+ coordinates with GSH/Gly-Gly CuNCs again, which leads to the AIE effect and realize the fluorescence "turn-on" process of the system. This strategy results inATP linear range of 0.508 ~ 120.0μM with a detection limit of 0.508μM (S/N = 3) and ACP linear range of 0.011 ~ 30.0 U·L-1 with a detection limit of 0.011 U·L-1 (S/N = 3). Application tobiologic samples wassuccessful.

The Pyruvate Kinase (PK) Activator AG-946 Improves PK Properties and Red Blood Cell (RBC) Characteristics upon Ex Vivo treatment of RBCs from Patients with Myelodysplastic Syndromes

Background: Myelodysplastic syndromes (MDS) consist of a heterogenous group of clonal myeloid malignancies, characterized by dysplasia, ineffective hematopoiesis and cytopenias. MDS patients frequently suffer from anemia, directly affecting quality of life. The limited therapeutic options often do not result in the desired clinical improvement. Interestingly, in MDS the activity of the red blood cell (RBC) enzyme pyruvate kinase (PK), a key regulatory enzyme of glycolysis, may be decreased. The activation of PK via small molecules is hypothesized to be beneficial in a wide range of anemias. Currently, a phase 2a, open-label, proof of concept trial is running on the use of the PK activator AG-946 in low-risk MDS patients (NCT05490446). In light of these developments, we investigated the effect of ex vivo treatment of MDS RBCs with AG-946. Objectives: To evaluate RBC PK and cellular properties of patients with MDS, and to determine the effect of ex vivo treatment with the PK activator AG-946. Methods: Eleven low-risk non-transfusion dependent MDS patients and six healthy controls (HCs) were studied. Baseline PK activity and PK thermostability were measured in RBC lysates of purified RBCs, under V max conditions (phosphoenolpyruvate (PEP) 5mM final concentration). Hexokinase (HK) activity was included to relate PK activity to mean RBC age. Ex vivo treatment was performed by incubating MDS RBCs for 16 hours at 37 °C in presence or absence of the PK activator AG-946 (10μM or 5μM) using dimethylsulfoxide (DMSO) as a blank. Effect was assessed by measuring PK activity (using a final concentration of PEP 0.5mM) and levels of adenosine triphosphate (ATP) (LC-MS/MS). Functional RBC analysis was performed by osmotic gradient ektacytometry (Lorrca MaxSis). To investigate the effect on PK thermostability, RBC lysates were incubated with AG-946 (2μM or 1μM) or DMSO, after which residual PK activity was measured (PEP 0.5mM). The effect of AG-946 on erythroid development was studied using peripheral blood mononuclear cells, cultured in MethoCult TM H4434 medium for 14 days in the presence or absence of AG-946 (10μM or 625nM, DMSO as blank). Differences between conditions were determined using Unpaired T-test, ANOVA with Dunnett's test or Kruskal-Wallis with Dunn's test via GraphPad Prism. Results: Mean PK/HK ratio was significantly lower in RBCs from MDS patients compared to HCs (6.1 (SD 1.8) versus 10.5 (SD 1.5), p<0.001). Upon ex vivo treatment, PK activity significantly increased (independent of dosage) when compared to DMSO (10μM AG-946, mean increase 30% ( p<0.001); 5μM, 28% ( p<0.0001)) (Figure 1A). MDS RBCs displayed decreased PK thermostability at baseline under V max conditions (HCs 79% residual activity versus 68% in MDS, p<0.01). This was restored by ex vivo treatment with AG-946 (DMSO, 33% residual activity, 2μM AG-946, 85% ( p<0.0001); 1μM, 87% ( p<0.0001)). The increase in PK activity was accompanied by significantly increased ATP levels (N=10, DMSO, mean ATP 14.9 μg/mL RBC; 10μM AG-946, 21.0 μg/mL RBC ( p<0.01); 5μM, 21.1 μg/mL RBC ( p<0.01)) (Figure 1B). Functionally, RBC properties improved upon ex vivo treatment with AG-946 as reflected by the modest yet significant increase in O hyper, indicating improved hydration status (10μM AG-946, mean increase 2.3% ( p<0.001); 5μM, 2.9% ( p<0.0001)). To date, culture assays were performed in 8 patients; 3/8 showed an increase in the number of burst forming units-erythroid (BFU-Es) upon treatment with AG-946 when compared to DMSO. The percentual increases differed per patient: increase of 25% with 10μM AG946 and 150% with 625nM, increase of 40% (10μM) and 39% (625nM), and for the third an increase of 22% (10μM) and 39% (625nM). Conclusion: Our findings show that RBCs from MDS patients have decreased PK activity and thermostability. We demonstrate that ex vivo treatment with AG-946 increases PK activity and ATP levels and restores PK thermostability. Moreover, ex vivo treatment with AG-946 improves RBC hydration, suggesting that improved energy status directly improves RBC functional properties. The preliminary results of the culture assay could indicate that in certain patients, dyserythropoiesis may be ameliorated upon PK activation. To better understand the effects on erythropoiesis additional experiments are required. In conclusion, our data might support a rationale for the use of PK activators as a novel therapeutic option for MDS.

Mechanism of DaiTongXiao in the treatment of gouty arthritis through the NLRP3 signaling pathway

Ethnopharmacological relevanceDaiTongXiao (DTX) is a traditional Chinese Dai folk formulation utilized for gouty arthritis treatment, with substantial evidence supporting its anti-inflammatory properties. The NLRP3 inflammasome disorder is tightly linked to the development of many inflammatory diseases. Aim of the studyTo elucidate the therapeutic efficacy of DTX in gouty arthritis and reveal its potential underlying mechanism. Materials and methodsThe primary active constituents in DTX were determined through ultraviolet spectrophotometry and gas chromatography. Rats underwent induction with monosodium urate (MSU), followed by treatment of J774A.1 cells with adenosine triphosphate (ATP) activation and lipopolysaccharide (LPS) induction and the subsequent culture in Dulbecco’s modified Eagle’s medium. The degree of foot joint swelling in rats was assessed, and ankle joints were evaluated through H&E staining. Enzyme-linked immunosorbent assay was performed to measure the levels of interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor (TNF)-α in both serum and cells. Reverse transcription–polymerase chain reaction (RT-PCR) was performed to determine the relative mRNA expression levels of NLRP3, ASC, Caspase-1, and NF-κB in J774A.1 macrophages. The expression of NLRP3, ASC, Caspase-1, and NF-κB was examined by western blotting. ResultsDTX could alleviate MSU-induced joint swelling in rats, as evidenced by a reduction in joint inflammation. Moreover, DTX effectively enhanced the survival rate of J774A.1 cells following LPS induction and ATP activation. Furthermore, DTX significantly reduced IL-1β, IL-6, IL-8, and TNF-α levels in both cell culture medium and rat serum. RT-PCR results revealed that DTX notably downregulated the mRNA expression levels of NLRP3, ASC, Caspase-1, and NF-κB in J774A.1 cells. Additionally, DTX downregulated NLRP3, ASC, NF-κB, and Caspase-1 expression in the joint tissue. ConclusionsDTX exerts a significant anti–gouty arthritis effect, with its mechanism being tightly linked to the NLRP3 inflammatory signaling pathway. This pathway may be modulated by inhibiting IL-1β differentiation and maturation by downregulating NLRP3, ASC, Caspase-1, and NF-κB protein expression. This, in turn, leads to a reduction in the release of IL-6, IL-8, and TNF-α, ultimately impeding gouty arthritis progression.

Mitochondrial transplantation and transfer: The promising method for diseases.

Mitochondria are organelles that serve as the powerhouses for cellular bioenergetics in eukaryotic cells. It is responsible for mitochondrial adenosine triphosphate (ATP) generation, cell signaling and activity, calcium balance, cell survival, proliferation, apoptosis, and autophagy. Mitochondrial transplantation is a promising disease therapy that involves the recovery of mitochondrial dysfunction using isolated functioning mitochondria. The objective of the present article is to provide current knowledge on natural mitochondrial transfer processes, in vitro and in vivo applications of mitochondrial transplantation, clinical trials, and challenges associated with mitochondrial transplantation.

Effect of ibuprofen (IBU) on the sulfur-based and calcined pyrite-based autotrophic denitrification (SCPAD) systems with two filling modes: Performance and toxic response mechanism

To investigate the effect of ibuprofen (IBU) on the sulfur-based and calcined pyrite-based autotrophic denitrification (SCPAD) systems, two individual reactors with the layered filling (L-SCPAD) and mixed filling (M-SCPAD) systems were established via sulfur and calcined pyrite. Effluent NO3−-N concentration of the L-SCPAD and M-SCPAD systems was first increased to 6.44, 0.93 mg/L under 0.5 mg/L IBU exposure and gradually decreased to 1.66 mg/L, 0 mg/L under 4.0 mg/L IBU exposure, indicating that NO3−-N removal performance of the M-SCPAD system was better than that of the L-SCPAD system. The variation of extracellular polymeric substances (EPS) characteristics demonstrated that more EPS was secreted in the M-SCPAD system compared to the L-SCPAD system, which contributed to forming a more stable biofilm structure and protecting microorganisms against the toxicity of IBU in the M-SCPAD system. Moreover, the increased electron transfer impedance and decreased cytochrome c implied that IBU inhibited the electron transfer efficiency of the L-SCPAD and M-SCPAD systems. The decreased adenosine triphosphate (ATP) and electron transfer system activity (ETSA) content showed that IBU inhibited metabolic activity, but the M-SCPAD system exhibited higher metabolic activity compared to the L-SCPAD system. In addition, the analysis of the bacterial community indicated a more stable abundance of nitrogen removal function bacteria (Bacillus) in the M-SCPAD system compared to the L-SCPAD system, which was conducive to maintaining a stable denitrification performance. The toxic response mechanism based on the biogeobattery effect was proposed in the SCPAD systems under IBU exposure. This study provided an important reference for the long-term toxic effect of IBU on the SCPAD systems.

301 Measuring ATPase activity of adenosine triphosphate–binding cassette subfamily C member 4, lamprey CFTR, and human CFTR using antimony-phosphomolybdate assay

301 Measuring ATPase activity of adenosine triphosphate–binding cassette subfamily C member 4, lamprey CFTR, and human CFTR using antimony-phosphomolybdate assay

Photocatalytic titanium dioxide reduces postharvest decay of nectarine fruit packaged in different materials through modulating central carbon and energy metabolisms

The capacity of titanium dioxide (TiO2) photocatalysis photocatalytic reactor to prevent and control pathogen infection in nectarine fruit packed in laminated nylon/LDPE, low density polyethylene and microperforated LDPE films was evaluated. Results showed that TiO2 combined with microperforated LDPE packaging (TPL) exhibited superior inhibition of microbial growth, reducing total viable counts by 4.18 log CFU g−1 and yeast and mold counts by 3.20 log CFU g−1, compared to microperforated LDPE packaging alone. TiO2 photocatalysis primed the defense systems in nectarine fruit packed in microperforated LDPE, improving the activity of defense-related enzymes. Metabolomics analysis indicated that l-aspartate, oxaloacetate, and succinic acid involved in central carbon metabolism including the glycolysis and tricarboxylic acid cycle pathways, were significantly upregulated by TPL. TiO2 increased the activity of energy metabolism-related enzymes, adenosine triphosphate, adenosine diphosphate, and energy charge levels to provide adequate energy, thus reducing fruit decay.

Translocation of telomerase reverse transcriptase coincided with ATP release in postnatal cochlear supporting cells.

The spontaneous bursts of electrical activity in the developing auditory system are derived from the periodic release of adenosine triphosphate (ATP) by supporting cells in the Kölliker's organ. However, the mechanisms responsible for initiating spontaneous ATP release have not been determined. Our previous study revealed that telomerase reverse transcriptase (TERT) is expressed in the basilar membrane during the first postnatal week. Its role in cochlear development remains unclear. In this study, we investigated the expression and role of TERT in postnatal cochlea supporting cells. Our results revealed that in postnatal cochlear Kölliker's organ supporting cells, TERT shifts from the nucleus into the cytoplasm over time. We found that the TERT translocation tendency in postnatal cochlear supporting cells in vitro coincided with that observed in vivo. Further analysis showed that TERT in the cytoplasm was mainly located in mitochondria in the absence of oxidative stress or apoptosis, suggesting that TERT in mitochondria plays roles other than antioxidant or anti-apoptotic functions. We observed increased ATP synthesis, release and activation of purine signaling systems in supporting cells during the first 10 postnatal days. The phenomenon that TERT translocation coincided with changes in ATP synthesis, release and activation of the purine signaling system in postnatal cochlear supporting cells suggested that TERT may be involved in regulating ATP release and activation of the purine signaling system. Our study provides a new research direction for exploring the spontaneous electrical activity of the cochlea during the early postnatal period.

ATP and ssDNA aptamer-mediated peroxidase-like activity of rGO@PDA@CeO2 nanozyme: Exosomal proteins profiling and detection at physiological pH for colorimetric sensor

Exosomes are extracellular vesicles that inherit molecular markers from their parent cells for the transfer and delivery of membrane and cytosolic molecules between cells. Exosome detection and protein profiling provide noninvasive access to disease diagnosis and treatment. The nanozyme designed in this paper contains polydopamine-functionalization of graphene oxide (rGO@PDA) units that can adsorb ssDNA aptamers and CeO2 metal oxide nanosheet units with peroxidase activity. As auxiliary regulator, adenosine triphosphate (ATP) enhances peroxidase activity of rGO@PDA@CeO2 under a wide range of pH (4–7.4), which breaks through the limitation that traditional nanozymes can only exhibit peroxidase activity under acidic conditions. The ssDNA aptamers, which are recognition probes of exosome, can be adsorbed on the surface of rGO@PDA by aromatic stacking, competing with ATP molecules. Thus, the ssDNA aptamer acts as the switch for colorimetric sensor to flexibly regulate the peroxidase activity of the rGO@PDA@CeO2 under physiological conditions, and realizes sensitive detection of exosomes. This assay achieves a detection linear range of 0.46 × 107–108 particle/mL and a limit down to 3.81 × 105 particles/mL. In addition, this aptasensor sensitively profiles six exosomal protein across five cell types and clinical samples, suggesting their potential in clinic diagnosis.

Mechanism of mycobacterial ATP synthase inhibition by squaramides and second generation diarylquinolines.

Mycobacteria, such as Mycobacterium tuberculosis, depend on the activity of adenosine triphosphate (ATP) synthase for growth. The diarylquinoline bedaquiline (BDQ), a mycobacterial ATP synthase inhibitor, is an important medication for treatment of drug-resistant tuberculosis but suffers from off-target effects and is susceptible to resistance mutations. Consequently, both new and improved mycobacterial ATP synthase inhibitors are needed. We used electron cryomicroscopy and biochemical assays to study the interaction of Mycobacterium smegmatis ATP synthase with the second generation diarylquinoline TBAJ-876 and the squaramide inhibitor SQ31f. The aryl groups of TBAJ-876 improve binding compared with BDQ, while SQ31f, which blocks ATP synthesis ~10 times more potently than ATP hydrolysis, binds a previously unknown site in the enzyme's proton-conducting channel. Remarkably, BDQ, TBAJ-876, and SQ31f all induce similar conformational changes in ATP synthase, suggesting that the resulting conformation is particularly suited for drug binding. Further, high concentrations of the diarylquinolines uncouple the transmembrane proton motive force while for SQ31f they do not, which may explain why high concentrations of diarylquinolines, but not SQ31f, have been reported to kill mycobacteria.

Age-related changes in energy metabolism in peripheral mononuclear blood cells (PBMCs) and the brains of cognitively healthy seniors

Mitochondrial dysfunction is a hallmark of cellular senescence and many age-related neurodegenerative diseases. We therefore investigated the relationship between mitochondrial function in peripheral blood cells and cerebral energy metabolites in young and older sex-matched, physically and mentally healthy volunteers. Cross-sectional observational study involving 65 young (26.0 ± 0.49 years) and 65 older (71.7 ± 0.71 years) women and men recruited. Cognitive health was evaluated using established psychometric methods (MMSE, CERAD). Blood samples were collected and analyzed, and fresh peripheral blood mononuclear cells (PBMCs) were isolated. Mitochondrial respiratory complex activity was measured using a Clarke electrode. Adenosine triphosphate (ATP) and citrate synthase activity (CS) were determined by bioluminescence and photometrically. N-aspartyl-aspartate (tNAA), ATP, creatine (Cr), and phosphocreatine (PCr) were quantified in brains using 1H- and 31P-magnetic resonance spectroscopic imaging (MRSI). Levels of insulin-like growth factor 1 (IGF-1) were determined using a radio-immune assay (RIA). Complex IV activity (CIV) (− 15%) and ATP levels (− 11%) were reduced in PBMCs isolated from older participants. Serum levels of IGF-1 were significantly reduced (− 34%) in older participants. Genes involved in mitochondrial activity, antioxidant mechanisms, and autophagy were unaffected by age. tNAA levels were reduced (− 5%), Cr (+ 11%), and PCr (+ 14%) levels were increased, and ATP levels were unchanged in the brains of older participants. Markers of energy metabolism in blood cells did not significantly correlate with energy metabolites in the brain. Age-related bioenergetic changes were detected in peripheral blood cells and the brains of healthy older people. However, mitochondrial function in peripheral blood cells does not reflect energy related metabolites in the brain. While ATP levels in PBMCs may be be a valid marker for age-related mitochondrial dysfunction in humans, cerebral ATP remained constant.

Bioinspired Structure Regulation of Apyrase-Like Nanozyme with Intracellular-Generated H2O2 for Tumor Catalytic Therapy

Adenosine triphosphate (ATP) is the major resource of energy supply in tumor activity. Therefore, improving ATP consumption efficiencies is a promising approach for cancer therapy. Herein, inspired by the H2O2-involved structure regulation effect during the catalysis of natural protein enzymes, we developed an artificial H2O2-driven ATP catalysis-promoting system, the Ce-based metal-organic framework (Ce-MOF), for catalytic cancer therapy. In the presence of H2O2, the hydrolysis ATP activity of Ce-MOF(H2O2) was enhanced by around 1.6 times. Taking advantage of the endogenous H2O2 in cancerous cells, catalytic hydrolysis for intracellular ATP of the Ce-MOF achieves the inhibition of cancerous cell growth, which involves damaged mitochondrial function and autophagy-associated cell death. Furthermore, in vivo studies suggest that the Ce-MOF has a good tumor inhibition effect. The artificial H2O2-driven ATP catalysis-promoting system not only demonstrates high catalytic ATP consumption efficiencies for cancer therapy but also highlights a bioinspired strategy to expedite nanozyme research in both design and applied sciences.

Neuroprotective effect of ginsenoside Re on drosophila model of Parkinson's disease

This study aims to explore the neuroprotective mechanism of ginsenoside Re(GS-Re) on drosophila model of Parkinson's disease(PD) induced by rotenone(Rot). To be specific, Rot was used to induce PD in drosophilas. Then the drosophilas were grouped and respectively treated(GS-Re: 0.1, 0.4, 1.6 mmol·L~(-1); L-dopa: 80 μmol·L~(-1)). Life span and crawling ability of drosophilas were determined. The brain antioxidant activity [content of catalase(CAT), malondialdehyde(MDA), reactive oxygen species(ROS), superoxide dismutase(SOD)], dopamine(DA) content, and mitochondrial function [content of adenosine triphosphate(ATP), NADH:ubiquinone oxidoreductase subunit B8(NDUFB8) Ⅰ activity, succinate dehydrogenase complex, subunit B(SDHB) Ⅱ activity] were detected by enzyme-linked immunosorbent assay(ELISA). The number of DA neurons in the brains of drosophilas was measured with the immunofluorescence method. The levels of NDUFB8 Ⅰ, SDHB Ⅱ, cytochrome C(Cyt C), nuclear factor-E2-related factor 2(Nrf2), heme oxygenase-1(HO-1), B-cell lymphoma/leukemia 2(Bcl-2)/Bcl-2-assaciated X protein(Bax), and cleaved caspase-3/caspase-3 in the brain were detected by Western blot. The results showed that model group [475 μmol·L~(-1) Rot(IC_(50))] demonstrated significantly low survival rate, obvious dyskinesia, small number of neurons and low DA content in the brain, high ROS level and MDA content, low content of SOD and CAT, significantly low ATP content, NDUFB8 Ⅰ activity, and SDHB Ⅱ activity, significantly low expression of NDUFB8 Ⅰ, SDHB Ⅱ, and Bcl-2/Bax, large amount of Cyt C released from mitochondria to cytoplasm, low nuclear transfer of Nrf2, and significantly high expression of cleaved caspase-3/caspase-3 compared with the control group. GS-Re(0.1, 0.4, and 1.6 mmol·L~(-1)) significantly improved the survival rate of PD drosophilas, alleviated the dyskinesia, increased DA content, reduced the loss of DA neurons, ROS level, and MDA content in brain, improved content of SOD and CAT and antioxidant activity in brain, maintained mitochondrial homeostasis(significantly increased ATP content and activity of NDUFB8 Ⅰ and SDHB Ⅱ, significantly up-regulated expression of NDUFB8 Ⅰ, SDHB Ⅱ, and Bcl-2/Bax), significantly reduced the expression of Cyt C, increased the nuclear transfer of Nrf2, and down-regulated the expression of cleaved caspase-3/caspase-3. In conclusion, GS-Re can significantly relieve the Rot-induced cerebral neurotoxicity in drosophilas. The mechanism may be that GS-Re activates Keap1-Nrf2-ARE signaling pathway by maintaining mitochondrial homeostasis, improves antioxidant capacity of brain neurons, then inhibits mitochondria-mediated caspase-3 signaling pathway, and the apoptosis of neuronal cells, thereby exerting the neuroprotective effect.

Antibacterial activity and mechanism of sanguinarine against Staphylococcus aureus by interfering with the permeability of the cell wall and membrane and inducing bacterial ROS production.

Staphylococcus aureus (SA) is representative of gram-positive bacteria. Sanguinarine chloride hydrate (SGCH) is the hydrochloride form of sanguinarine (SG), one of the main extracts of Macleaya cordata (M. cordata). There are few reports on its antibacterial mechanism against SA. Therefore, in this study, we investigated the in vitro antibacterial activity and mechanism of SGCH against SA. The inhibitory zone, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) were measured, and the bactericidal activity curve was plotted. In addition, the micromorphology, alkaline phosphatase (AKP) activity, Na+K+, Ca2+Mg2+-adenosine triphosphate (ATP) activity, intracellular reactive oxygen species (ROS), and fluorescein diacetate (FDA) were observed and detected. The results showed that the inhibitory zone of SGCH against SA was judged as medium-sensitive; the MIC and MBC were 128 and 256 μg/mL, respectively; in the bactericidal activity curve, SGCH with 8 × MIC could completely kill SA within 24 h. SGCH was able to interfere with the integrity and permeability of the SA cell wall and membrane, as confirmed by the scanning electron microscopy (SEM) images, the increase in extracellular AKP and Na+ K+, Ca2+ Mg2+-ATP activities as well as the fluorescein diacetate (FDA) staining experiment results. Moreover, a high concentration of SGCH could induce SA to produce large amounts of ROS. In summary, these findings revealed that SGCH has a preferable antibacterial effect on SA, providing an experimental and theoretical basis for using SG as an antibiotic substitute in animal husbandry and for the clinical control and treatment of diseases caused by SA.