Adenosine Triphosphate Levels Research Articles

Aflatoxin B1 induces ROS-dependent mitophagy by modulating the PINK1/Parkin pathway in HepG2 cells.

Aflatoxin B1 (AFB1) is extremely harmful to both humans and animals. Mitophagy is a selective process of self-elimination and has an important role in controlling mitochondrial quality. The present study aimed to investigate the effect of reactive oxygen species (ROS) accumulation on AFB1-induced mitophagy in HepG2 cells to provide a new perspective from which to design novel therapeutic strategies to treat AFB1 poisoning. ROS release was induced in HepG2 cells with AFB1 (10μmol/L). Cell autophagy activity, mitochondrial membrane potential (MMP), adenosine triphosphate (ATP) levels, Parkin translocation and both the transcription and expression of mitophagy-related proteins were measured when N-acetyl-L-cysteine (NAC) partially decreased the ROS level, while the knockdown of nuclear factor erythroid 2-related factor 2 (Nrf2) resulted in a large accumulation of ROS. The results reveal that NAC pretreatment ameliorated the decline in both the MMP and the ATP levels while also activating phosphoglycerate mutase 5 (PGAM5)-PTEN-induced kinase 1 (PINK1)/Parkin, while the Nrf2 knockdown group exhibited the opposite trend. These results suggest that AFB1-induced mitophagy in HepG2 cells depends on ROS, and proper ROS activates mitophagy to play a protective role.

Vibration Emissions Reduce Boar Sperm Quality via Disrupting Its Metabolism

Artificial insemination (AI) with liquid-preserved semen has recently become common in pig breeding. The semen doses are produced in a centralized manner at the boar stud and then subsequently distributed and transported to sow farms. However, vibration emissions during transportation by logistic vehicles may adversely affect the quality of boar sperm. Therefore, this study aimed to explore the impact of vibration-induced emissions on sperm quality and function under simulated transportation conditions. Each time, ejaculates from all 15 boars were collected and then pooled together to minimize individual variations, and the sample was split using an extender for dilution. Different rotational speeds (0 rpm, 80 rpm, 140 rpm, 200 rpm) were utilized to simulate varying intensities of vibration exposure using an orbital shaker, considering different transportation times (0 h, 3 h, and 6 h). Subsequently, evaluations were conducted regarding sperm motility, plasma membrane integrity, acrosome integrity, mitochondrial function, adenosine triphosphate (ATP) levels, mitochondrial reactive oxygen species (ROS) levels, pH, glycolytic pathway enzyme activities, and capacitation following exposure to vibration emissions. Both vibration time and intensity impact sperm motility, plasma membrane integrity, and acrosomal integrity. Vibration exposure significantly reduced sperm ATP levels, mitochondrial membrane potential, and the levels of mitochondria-encoded proteins (MT-ND1, MT-ND6) (p < 0.05). After vibration emission treatment, the pH value and mitochondrial ROS levels significantly increased (p < 0.05). Inhibition of sperm glycolysis was observed, with reduced activities of hexokinase (HK), pyruvate kinase (PK), and lactate dehydrogenase (LDH), along with decreased lactate levels (p < 0.05). Additionally, sperm tyrosine phosphorylation levels were significantly reduced by vibration emissions compared to the control group (p < 0.05). After the vibration emission treatment, the number of sperm bound to each square millimeter of oviduct explants decreased significantly compared to the control group (p < 0.05). Similarly, compared to the control group, using semen subjected to vibration stress for AI results in significantly reduced pregnancy rates, total born litter size, live-born litter size, and healthy born litter size (p < 0.05).

Lactococcus lactis subsp. cremoris C60 Upregulates Macrophage Function by Modifying Metabolic Preference in Enhanced Anti-Tumor Immunity

Lactococcus lactis subsp. cremoris C60 is a probiotic strain of lactic acid bacteria (LAB) which induces various immune modifications in myeloid lineage cells. These modifications subsequently regulate T cell function, resulting in enhanced immunity both locally and systemically. Here, we report that C60 suppresses tumor growth by enhancing macrophage function via metabolic alterations, thereby increasing adenosine triphosphate (ATP) production in a murine melanoma model. Intragastric (i.g.) administration of C60 significantly reduced tumor volume compared to saline administration in mice. The anti-tumor function of intratumor (IT) macrophage was upregulated in mice administered with C60, as evidenced by an increased inflammatory phenotype (M1) rather than an anti-inflammatory/reparative (M2) phenotype, along with enhanced antigen-presenting ability, resulting in increased tumor antigen-specific CD8+ T cells. Through this functional modification, we identified that C60 establishes a glycolysis-dominant metabolism, rather than fatty acid oxidation (FAO), in IT macrophages, leading to increased intracellular ATP levels. To address the question of why orally supplemented C60 exhibits functions in distal places, we found a possibility that bacterial cell wall components, which could be distributed throughout the body from the gut, may induce stimulatory signals in peripheral macrophages via Toll-like receptors (TLRs) signaling activation. Thus, C60 strengthens macrophage anti-tumor immunity by promoting a predominant metabolic shift towards glycolysis upon TLR-mediated stimulation, thereby increasing substantial energy production.

IATPSnFR2: A high-dynamic-range fluorescent sensor for monitoring intracellular ATP

We developed a significantly improved genetically encoded quantitative adenosine triphosphate (ATP) sensor to provide real-time dynamics of ATP levels in subcellular compartments. iATPSnFR2 is a variant of iATPSnFR1, a previously developed sensor that has circularly permuted superfolder green fluorescent protein (GFP) inserted between the ATP-binding helices of the ε-subunit of a bacterial F0-F1 ATPase. Optimizing the linkers joining the two domains resulted in a ~fivefold to sixfold improvement in the dynamic range compared to the previous-generation sensor, with excellent discrimination against other analytes, and affinity variants varying from 4 µM to 500 µM. A chimeric version of this sensor fused to either the HaloTag protein or a suitable spectrally separated fluorescent protein provides an optional ratiometric readout allowing comparisons of ATP across cellular regions. Subcellular targeting the sensor to nerve terminals reveals previously uncharacterized single-synapse metabolic signatures, while targeting to the mitochondrial matrix allowed direct quantitative probing of oxidative phosphorylation dynamics.

RUNX3 alleviates mitochondrial dysfunction and tubular damage by inhibiting TLR4/NF-κB signalling pathway in diabetic kidney disease.

The impaired function of tubular mitochondria is critical in diabetic kidney disease (DKD) progression. RUNX3 is down-regulated in DKD models. We intend to explore the effects of RUNX3 on mitochondrial dysfunction and renal tubule injury in DKD and related mechanisms. The development of diabetes models involved injecting mice with streptozotocin while treating HK-2 cells with high glucose (HG). By using immunohistochemical techniques, the renal localizations of RUNX3 were identified. Levels of adenosine triphosphate (ATP), mitochondrial membrane potential, and biochemical index were detected by appropriate kits. Reactive oxygen species (ROS) generation was assessed with dihydroethidium and MitoSOX Red staining. Apoptosis was assessed by flow cytometry and TUNEL. RUNX3 ubiquitination was measured. RUNX3 was mainly present in renal tubules. Overexpressing RUNX3 increased Mfn2, Mfn1, ATP levels, and mitochondrial membrane potential, reduced Drp1 and ROS levels and cell apoptosis, as well as Cyt-C release into the cytoplasm. RUNX3 overexpression displayed a reduction in urinary albumin to creatinine ratio, Hemoglobin A1c, serum creatinine, and blood urea nitrogen. Overexpressing TLR4 attenuated the inhibitory effect of RUNX3 overexpression on mitochondrial dysfunction and cell apoptosis. HG promoted RUNX3 ubiquitination and SMURF2 expression. RUNX3 knockdown cancelled the inhibitory effect of SMURF2 on mitochondrial dysfunction and cell apoptosis. SMURF2 interference inhibits RUNX3 ubiquitination and TLR4/NF-κB signalling pathway, thereby alleviating renal tubule injury.

Rosmarinic acid ameliorated oxidative stress, neuronal injuries, and mitochondrial dysfunctions mediated by polyglutamine and ɑ-synuclein in Caenorhabditis elegans models.

Numerous natural antioxidants have been developed into agents for neurodegenerative diseases (NDs) treatment. Rosmarinic acid (RA), an excellent antioxidant, exhibits neuroprotective activity, but its anti-NDs efficacy remains puzzling. Here, Caenorhabditis elegans models were employed to systematically reveal RA-mediated mechanisms in delaying NDs from diverse facets, including oxidative stress, the homeostasis of neural and protein, and mitochondrial disorders. Firstly, RA significantly inhibited reactive oxygen species accumulation, reduced peroxide malonaldehyde production, and strengthened the antioxidant defense system via increasing superoxide dismutase activity. Besides, RA reduced neuronal loss and ameliorated polyglutamine and ɑ-synuclein-mediated dyskinesia in NDs models. Further, in combination with the data and molecular docking results, RA may bind specifically to Huntington protein and ɑ-synuclein to prevent toxic protein aggregation and thus enhance proteostasis. Finally, RA ameliorated mitochondrial dysfunction including increasing adenosine triphosphate and mitochondrial membrane potential levels and rescuing mitochondrial membrane proteins' expressions and mitochondrial structural abnormalities via regulating mitochondrial dynamics genes and improving the mitochondrial kinetic homeostasis. Thus, this study systematically revealed the RA-mediated neuroprotective mechanism and promoted RA as a promising nutritional intervention strategy to prevent NDs.

Antifibrotic effect of the P2X7 receptor antagonist A740003 against acute myocardial infarction-induced fibrotic remodelling

Post-acute myocardial infarction (AMI) fibrosis is a pathophysiologic process characterised by activation of the profibrotic mediator, transforming growth factor-β (TGF-β). AMI is associated with a substantial increase in the levels of extracellular adenosine triphosphate (eATP), which acts on the purinergic P2X7-receptor (P2X7-R) and triggers an inflammatory response that contributes to myocardial fibrotic remodelling. P2X7-R has been implicated in several cardiovascular diseases; however, its role in the regulation of cardiac fibrosis remains unclear. Therefore, the current study aimed to determine the effect of the P2X7-R antagonist, A740003, on post-AMI fibrosis, via the profibrotic TGF-β1/Smad signalling pathway, and elucidate whether its effect is mediated via the modulation of GSK-3β. AMI was induced by surgical ligation of the left anterior descending coronary artery, Thereafter, animals were divided into groups: sham control, MI-untreated, MI-vehicle, and MI-A740003 (50 mg/kg/day) and treated for seven days accordingly. The heart weight/body weight ratio of untreated-ligated rats significantly increased by 15.1%, creatine kinase‐MB (CK‐MB) significantly increased by 40 %, troponin‐I levels significantly increased by 25.4 %, and lactate dehydrogenase significantly increased by 47.2 %, indicating myocardial damage confirmed by morphological changes and massive cardiac fibrosis. The protein expression of cardiac fibronectin, TGF-β1, and p-Smad2 were also upregulated by 143 %, 40 %, and 8 %, respectively, indicating cardiac fibrosis. The treatment of ligated rats with A740003 led to improvement in all the above-mentioned parameters. Overall, A740003 exhibits potential cardio-protective effects on post-AMI fibrotic remodelling in the animal model of AMI through P2X7-R blockade, possibly by downregulating the profibrotic TGF-β1/Smad signalling pathway and restoring GSK-3β phosphorylation. Altogether, treatment with A740003 could serve as a new cardioprotective strategy to attenuate post-AMI fibrotic remodelling.

Ginseng total saponin improves red blood cell oxidative stress injury by regulating tyrosine phosphorylation and glycolysis in red blood cells

BackgroundOxidative stress is the main cause of many diseases, but because of its complex pathogenic factors, there is no clear method for treating it. Ginseng total saponin (GTS) an important active ingredients in Panax ginseng C.A. Mey (PG) and has potential therapeutic ability for oxidative stress due to various causes. However, the molecular mechanism of GTS in the treating oxidative stress damage in red blood cells (RBCs) is still unclear. PurposeThis study aimed to examine the protective effect of GTS on RBCs under oxidative stress damage and to determine its potential mechanism. MethodsThe oxidative stress models of rat RBCs induced by hydrogen peroxide (H2O2) and exhaustive swimming in vivo and in vitro was used. We determined the cell morphology, oxygen carrying capacity, apoptosis, antioxidant capacity, and energy metabolism of RBCs. The effect of tyrosine phosphorylation (pTyr) of Band 3 protein on RBCs glycolysis was also examined. ResultsGTS reduced the hemolysis of RBCs induced by H2O2 at the lowest concentration. Moreover, GTS effectively improved the morphology, enhanced the oxygen carrying capacity, and increased antioxidant enzyme activity, adenosine triphosphate (ATP) levels, and adenosine triphosphatase (ATPase) activity in RBCs. GTS also promoted the expression of membrane proteins in RBCs, inhibited pTyr of Band 3 protein, and further improved glycolysis, restoring the morphological structure and physiological function of RBCs. ConclusionsThis study shows, that GTS can protect RBCs from oxidative stress damage by improving RBCs morphology and physiological function. Changes in pTyr expression and its related pTyr regulatory enzymes before and after GTS treatment suggest that Band 3 protein is the main target of GTS in the treating endogenous and exogenous oxidative stress. Moreover, GTS can enhance the glycolytic ability of RBCs by inhibiting pTyr of Band 3 protein, thereby restoring the function of RBCs.

Cypate-sensitized upconversion nanoprobes for intracellular and in-vivo ATP ratiometric detection

As energy currency of life, adenosine triphosphate (ATP) is highly dynamic in living organisms. It is crucial yet challenging to probe intracellular or in-vivo ATP precisely without destruction of the living biological systems. Herein, we have designed novel ATP-responsive ratiometric nanoprobes by employing upconversion nanoparticles (UCNPs) with cypate as antennas. Nuclear magnetic resonance spectroscopy was used to analyze the interfacial interactions among lanthanide nanocrystal, cypate and adenosine phosphates, disclosing the origin of detection specificity. ATP induced interruption of energy transfer pathway in cypate-sensitized UCNPs was further investigated by femtosecond transient absorption and time-resolved photoluminescence spectroscopy. The nanoprobes with dual near-infrared (NIR)-excited ratiometric UCL were successfully applied for quantification of ATP level in living cells under vegetatively growing, starving, and glycolysis-inhibiting states, respectively. Furthermore, the increase of available energy by feeding for zebrafish larvae was confirmed through monitoring the in-vivo ATP variation with the nanoprobes. Such nanoprobes showed great potential in ATP-associated biological research and clinical applications. These findings may gain deep insights into the surface science and photophysics in dye-lanthanide nanocomposites, and open new avenues for the design and bioapplications of lanthanide nanoprobes.

Biocompatible aggregation-induced emission active polyphosphate-manganese nanosheets with glutamine synthetase-like activity in excitotoxic nerve cells

Glutamine synthetase (GS) is vital in maintaining ammonia and glutamate (Glu) homeostasis in living organisms. However, the natural enzyme relies on adenosine triphosphate (ATP) to activate Glu, resulting in impaired GS function during ATP-deficient neurotoxic events. To date, no reports demonstrate using artificial nanostructures to mimic GS function. In this study, we synthesize aggregation-induced emission active polyP-Mn nanosheets (STPE-PMNSs) based on end-labeled polyphosphate (polyP), exhibiting remarkable GS-like activity independent of ATP presence. Further investigation reveals polyP in STPE-PMNSs serves as phosphate source to activate Glu at low ATP levels. This self-feeding mechanism offers a significant advantage in regulating Glu homeostasis at reduced ATP levels in nerve cells during excitotoxic conditions. STPE-PMNSs can effectively promote the conversion of Glu to glutamine (Gln) in excitatory neurotoxic human neuroblastoma cells (SH-SY5Y) and alleviate Glu-induced neurotoxicity. Additionally, the fluorescence signal of nanosheets enables precise monitoring of the subcellular distribution of STPE-PMNSs. More importantly, the intracellular fluorescence signal is enhanced in a conversion-responsive manner, allowing real-time tracking of reaction progression. This study presents a self-sustaining strategy to address GS functional impairment caused by ATP deficiency in nerve cells during neurotoxic events. Furthermore, it offers a fresh perspective on the potential biological applications of polyP-based nanostructures.

Potential mitigating impact of a dipeptidyl peptidase-IV inhibitor, vildagliptin, on oxazolone-induced ulcerative colitis: Targeting the role of PI3K/AKT/mTOR and AMPK/Nrf2 signaling pathways

Growing evidence suggests that phosphoinositide 3-kinase (PI3K) and adenosine monophosphate-activated protein kinase (AMPK) signaling cascades are critical in ulcerative colitis (UC) pathophysiology by influencing gut mucosal inflammation. Recently, the coloprotective properties of dipeptidyl peptidase-IV (DPP-IV) inhibitors have emerged. Thus, this study assessed for the first time the potential mitigating impact of a DPP-IV inhibitor, vildagliptin (Vilda), on oxazolone (OXZ)-induced colitis in rats, targeting the role of PI3K/AKT/mTOR and AMPK/Nrf2 pathways. Thirty-two adult Albino rats were divided into four groups: control, Vilda (10 mg/kg/day orally), OXZ (300 µL of 5 % OXZ in 50 % aqueous ethanol solution introduced once into the colon via catheter), and Vilda+OXZ. Inflammatory cytokines (interleukin 13, tumor necrosis factor-α, interleukin 10), oxidative/endoplasmic reticulum stress markers (myeloperoxidase, reduced glutathione, catalase, CHOP), mitochondrial reactive oxygen species, adenosine triphosphate levels, and mitochondrial transmembrane potential were estimated. p-AMPK, p-AKT, beclin-1, and SQSTM1 levels were immunoassayed. Nrf2, PI3K, and mTOR expression levels were quantified using the real-time polymerase chain reaction. Furthermore, p-NF-ĸBp65 and LC3II immunoreactivity were evaluated. Vilda administration effectively ameliorated OXZ-induced colitis, as evidenced by the reduced Disease Activity Index, macroscopic colon damage score, colon weight/length ratio, ulcer index, and histopathological and electron microscopic changes in the colon tissues. Vilda treatment also counteracted OXZ-triggered inflammation, oxidative/endoplasmic reticulum stress, mitochondrial dysfunction, and enhanced autophagy in the colon. Vilda substantially suppressed PI3K/AKT/mTOR and activated the AMPK/Nrf2 pathway. Vilda has potent coloprotective and anti-ulcerogenic properties, primarily attributed to its antiinflammatory, antioxidant, and modulatory impact on mitochondrial dysfunction and autophagy activity. These effects were mostly mediated by suppressing PI3K/AKT/mTOR and activating AMPK/Nrf2 signaling cascades, suggesting a potential role of Vilda in UC therapy.

The ring rules the chain - inositol pyrophosphates and the regulation of inorganic polyphosphate.

The maintenance of phosphate homeostasis serves as a foundation for energy metabolism and signal transduction processes in all living organisms. Inositol pyrophosphates (PP-InsPs), composed of an inositol ring decorated with monophosphate and diphosphate moieties, and inorganic polyphosphate (polyP), chains of orthophosphate residues linked by phosphoanhydride bonds, are energy-rich biomolecules that play critical roles in phosphate homeostasis. There is a complex interplay between these two phosphate-rich molecules, and they share an interdependent relationship with cellular adenosine triphosphate (ATP) and inorganic phosphate (Pi). In eukaryotes, the enzymes involved in PP-InsP synthesis show some degree of conservation across species, whereas distinct enzymology exists for polyP synthesis among different organisms. In fact, the mechanism of polyP synthesis in metazoans, including mammals, is still unclear. Early studies on PP-InsP and polyP synthesis were conducted in the slime mould Dictyostelium discoideum, but it is in the budding yeast Saccharomyces cerevisiae that a clear understanding of the interplay between polyP, PP-InsPs, and Pi homeostasis has now been established. Recent research has shed more light on the influence of PP-InsPs on polyP in mammals, and the regulation of both these molecules by cellular ATP and Pi levels. In this review we will discuss the cross-talk between PP-InsPs, polyP, ATP, and Pi in the context of budding yeast, slime mould, and mammals. We will also highlight the similarities and differences in the relationship between these phosphate-rich biomolecules among this group of organisms.

Effect of transcutaneous electrical acupoint stimulation on postoperative urinary function in elderly patients undergoing total hip arthroplasty.

To observe the effect of transcutaneous electrical acupoint stimulation (TEAS) on postoperative urinary function in elderly patients undergoing total hip arthroplasty (THA). One hundred and eighty elderly patients undergoing unilateral THA without indwelling urinary catheters were randomly assigned to a TEAS group (90 cases, 3 cases dropped out, 4 cases were eliminated) and a sham TEAS group (90 cases, 1 case dropped out, 4 cases were eliminated). Both groups received fascia iliac block and subarachnoid block anesthesia under ultrasound guidance. The patients in the TEAS group were treated with TEAS at Zhongji (CV 3), Guanyuan (CV 4), and bilateral Huiyang (BL 35), Ciliao (BL 32) 30 minutes before anesthesia initiation, with dissperse-dense wave, frequency of 2 Hz/100 Hz, until 30 minutes after surgery. The patients in the sham TEAS group underwent the same procedure with the device applied at the same acupoints but without electrical stimulation. The incidence of postoperative urinary retention (POUR), time to first void, voiding threshold, urinary adenosine triphosphate (ATP) level, postoperative abnormal voiding status (bladder residual volume, re-catheterization rate, nocturia occurrence), and postoperative incidence of urinary tract infection (UTI) and prosthetic joint infection (PJI) were observed in both groups. The incidence of POUR in the TEAS group was lower than that in the sham TEAS group (P<0.05); the time to first void in the TEAS group was shorter than that in the sham TEAS group (P<0.05); the voiding threshold in the TEAS group was lower than that in the sham TEAS group (P<0.05); the urinary ATP level in the TEAS group was higher than that in the sham TEAS group (P<0.05); the bladder residual volume in the TEAS group was lower than that in the sham TEAS group (P<0.05); the nocturia occurrence in the TEAS group was lower than that in the sham TEAS group (P<0.05). However, there was no statistically significant difference in re-catheterization rate, incidence of UTI, and incidence of PJI between the two groups (P>0.05). TEAS could effectively reduce the occurrence of postoperative urinary retention and improve the postoperative urinary function in elderly patients undergoing THA, which might be related with increasing the urinary ATP level.

Early manifestation of hypophosphatemic rickets in goslings: a potential role of insufficient muscular adenosine triphosphate in motility impairment of early P-deficient geese

We aimed to determine the onset time of hypophosphatemic rickets and investigate the mechanism of motility impairment through adenosine triphosphate (ATP) production in goslings. Two hundred and sixteen 1-day-old male Jiangnan white geese were randomly divided into 3 groups, with 6 replicates and 12 geese per replicate. Birds were fed on 3 diets: a control diet (nonphytic phosphorus, NPP, 0.38%), a P-deficient diet (PD; NPP, 0.08%), and a high P diet (HP; NPP, 0.80%) for 14 d. Subsequently, all birds were shifted to the control diet for an additional 14 d. The cumulative incidence of lameness increased significantly (P < 0.01) starting on d 4, reaching over 80% on d 7 and 100% on d 12 in the PD group. Drinking and eating frequency decreased from d 4 and d 5, respectively, in the PD group compared to the other groups (most P < 0.01). The PD group exhibited shorter and narrower beaks, higher (worse) curvature scores of the beak and costochondral junctions, swelling caput costae, and dirtier feathers since d 4, in contrast to the control and HP groups (most P < 0.01). The HP had bigger (P < 0.05) beak and sternum sizes than the control groups on d 4 to 11. Leg muscle ATP levels were lower (P < 0.01 or 0.05) on d 4 to 11; in contrast, adenosine diphosphate (d 7-11) was higher in PD compared to the control (P < 0.05). Leg muscle ATP level had positive linear (R2 > 0.40) correlations (r > 0.60) with eating and drinking frequencies on d 7 and 11 (P < 0.01). Bone stiffness, feather cleanliness, and ATP levels recovered (P > 0.05) to the control level, whereas bone size did not recover (P < 0.05) in PD and HP after eating the control diet for 2 wk. The onset time of hypophosphatemic rickets was around 4 d in goslings, and insufficient leg muscle ATP was related to the impaired motility observed in early P-deficient geese.

Context-dependent modification of PFKFB3 in hematopoietic stem cells promotes anaerobic glycolysis and ensures stress hematopoiesis.

Metabolic pathways are plastic and rapidly change in response to stress or perturbation. Current metabolic profiling techniques require lysis of many cells, complicating the tracking of metabolic changes over time after stress in rare cells such as hematopoietic stem cells (HSCs). Here, we aimed to identify the key metabolic enzymes that define differences in glycolytic metabolism between steady-state and stress conditions in murine HSCs and elucidate their regulatory mechanisms. Through quantitative 13C metabolic flux analysis of glucose metabolism using high-sensitivity glucose tracing and mathematical modeling, we found that HSCs activate the glycolytic rate-limiting enzyme phosphofructokinase (PFK) during proliferation and oxidative phosphorylation (OXPHOS) inhibition. Real-time measurement of ATP levels in single HSCs demonstrated that proliferative stress or OXPHOS inhibition led to accelerated glycolysis via increased activity of PFKFB3, the enzyme regulating an allosteric PFK activator, within seconds to meet ATP requirements. Furthermore, varying stresses differentially activated PFKFB3 via PRMT1-dependent methylation during proliferative stress and via AMPK-dependent phosphorylation during OXPHOS inhibition. Overexpression of Pfkfb3 induced HSC proliferation and promoted differentiated cell production, whereas inhibition or loss of Pfkfb3 suppressed them. This study reveals the flexible and multilayered regulation of HSC glycolytic metabolism to sustain hematopoiesis under stress and provides techniques to better understand the physiological metabolism of rare hematopoietic cells.

Yiqigubiao pill treatment regulates Sirtuin 5 expression and mitochondrial function in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a heterogeneous group of pathophysiological bases of airway inflammation and its anti-inflammatory response. Aberrant mitochondrial signaling and mitochondrial dysfunction underlie the pathomechanisms leading to COPD. This study aims to investigate the effects of the Yiqigubiao (YQGB) pill, a traditional Chinese medicine (TCM), on Sirtuin 5 (SIRT5) and mitochondrial function in patients with COPD. Thirty-four patients with COPD were randomized into oral YQGB or placebo groups concurrent with a 24-week routine treatment. The pulmonary function was assessed by examining the levels of forced expiratory volume in one second (FEV1)/forced vital capacity (FVC), FEV1, and FVC. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot were used to detect SIRT5 expression in mitochondria isolated from peripheral blood. Flow cytometry was used to detect changes in mitochondrial membrane potential and reactive oxygen species (ROS) in peripheral blood lymphocytes. Human bronchial epithelial (HBE) cells stimulated by cigarette smoke extract (CSE) were treated with YQGB. After SIRT5 was knocked down in cells, the changes in mitochondrial membrane potential, levels of adenosine triphosphate (ATP), and ROS were detected. YQGB treatment significantly improved lung function in patients with COPD. The expression of SIRT5 and the mitochondrial membrane potential significantly increased and ROS decreased in patients with COPD after YQGB treatment. The CSE decreased cell proliferation and SIRT5 expression, which was alleviated after YQGB treatment. Furthermore, SIRT5 was knocked down in CSE-stimulated HBE cells, and its expression was elevated upon YQGB treatment. The knockdown of SIRT5 significantly altered the CSE-stimulation-induced dysregulation of mitochondrial membrane potential, ATP levels, and ROS. This was also restored after YQGB treatment. YQGB treatment can elevate SIRT5 expression, restore mitochondrial function in COPD, and exert protective effects.

Bisphenol S impairs mitochondrial function by targeting Myo19/oxidative phosphorylation pathway contributing to axonal and dendritic injury

Exposure to bisphenol S (BPS) is known to adversely affect neuronal development. As pivotal components of neuronal polarization, axons and dendrites are indispensable structures within neurons, crucial for the maintenance of nervous system function. Here, we investigated the impact of BPS exposure on axonal and dendritic development both in vivo and in vitro. Our results revealed that exposure to BPS during pregnancy and lactation led to a reduction in the complexity, density, and length of axons and dendrites in the prefrontal cortex (PFC) of offspring. Employing RNA sequencing technology to elucidate the underlying mechanisms of axonal and dendritic damage induced by BPS, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis highlighted a significant alteration in the oxidative phosphorylation (OXPHOS) pathway, essential for mitochondrial function. Subsequent experiments demonstrate BPS-induced impairment in mitochondrial function, including damaged morphology, decreased adenosine triphosphate (ATP) and superoxide dismutase (SOD) levels, and increased reactive oxygen species and malondialdehyde (MDA). These alterations coincided with the downregulated expression of OXPHOS pathway-related genes (ATP6V1B1, ATP5K, NDUFC1, NDUFC2, NDUFA3, COX6B1) and Myosin 19 (Myo19). Notably, Myo19 overexpression restored the BPS-induced mitochondrial dysfunction by alleviating the inhibition of OXPHOS pathway. Consequently, this amelioration was associated with a reduction in BPS-induced axonal and dendritic injury observed in cultured neurons of the PFC.

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.

Comparative Bioactivities of Chemically Modified Fucoidan and λ-Carrageenan toward Cells Encapsulated in Covalently Cross-Linked Hydrogels.

The sulfated marine polysaccharides, fucoidan and λ-carrageenan, are known to possess anti-inflammatory, immunomodulatory, and cellular protective properties. Although they hold considerable promise for tissue engineering constructs, their covalent cross-linking in hydrogels and comparative bioactivities to cells are absent from the literature. Thus, fucoidan and λ-carrageenan were modified with methacrylate groups and were covalently cross-linked with the synthetic polymer poly(vinyl alcohol)-methacrylate (PVA-MA) to form 20 wt % biosynthetic hydrogels. Identical degrees of methacrylation were confirmed by 1H NMR, and covalent conjugation was determined by using a colorimetric 1,9-dimethyl-methylene blue (DMMB) assay. Pancreatic beta cells were encapsulated in the hydrogels, followed by culturing in the 3D environment for a prolonged period of 32 days and evaluation of the cellular functionality by live/dead, adenosine 5'-triphosphate (ATP) level, and insulin secretion. The results confirmed that fucoidan and λ-carrageenan exhibited ∼12% methacrylate substitution, which generated hydrogels with stable conjugation of the polysaccharides with PVA-MA. The cells encapsulated in the PVA-fucoidan hydrogels demonstrated consistently high ATP levels over the culture period. Furthermore, only cells in the PVA-fucoidan hydrogels retained glucose responsiveness, demonstrating comparatively higher insulin secretion in response to glucose. In contrast, cells in the PVA-λ-carrageenan and the PVA control hydrogels lost all glucose responsiveness. The present work confirms the superior effects of chemically modified fucoidan over λ-carrageenan on pancreatic beta cell survival and function in covalently cross-linked hydrogels, thereby illustrating the importance of differential polysaccharide structural features on their biological effects.

Dihydromyricetin regulates RIPK3-CaMKII to prevent necroptosis in high glucose-stimulated cardiomyocytes

BackgroundDiabetic cardiomyopathy is one common cardiovascular complication without effective treatments. Dihydromyricetin (DHY), a natural dihydroflavonol compound extracted from Ampelopsis grossedentata, possesses versatile pharmacologically important effects. In our current research, we planned to evaluate the impact and probable DHY mechanisms in high glucose (HG)-induced cardiomyocytes. MethodsPrimary cardiomyocytes were pretreated with different concentrations of DHY (0, 20, 40, 80, 160, and 320 μM) for various time (0, 1, 2, 4, 12, and 24 h). They were then stimulated for 48 h with 5.5 mmol/L normal glucose (NG) and 33.3 mmol/L high glucose (HG). Cell viability, adenosine-triphosphate (ATP) levels, and lactate dehydrogenase (LDH) release of cardiomyocytes were detected. JC-1 staining was employed to measure the mitochondrial membrane potential. MitoSOX staining and dihydroethidium (DHE) staining were applied to evaluate the oxidative stress levels. TDT mediated dUTP nick end labeling (TUNEL) was used to measure apoptotic levels. Expressions of calcium/calmodulin-dependent protein kinase II (CaMKII), phospholamban (PLB), optic atrophy 1 (OPA1), dynamin-related protein 1 (DRP1), caspase 3, mixed kinase lineage domain like protein (MLKL), receptor interacting protein kinase 3 (RIPK3), and receptor interacting protein kinase 1 (RIPK1) were detected by immunofluorescence and/or Western blot. ResultsDHY improved cell viability, enhanced ATP level, and decreased LDH content in HG-stimulated cardiomyocytes, suggesting DHY attenuating cell injury. DHY reduced number of TUNEL positive cells, inhibited RIPK3 and cleaved-caspase 3 expression, implying DHY alleviated necroptosis in HG-stimulated cardiomyocytes. DHY diminished JC-1 monomers, DHE and MitoSOX fluorescence intensity as well as DRP1 expression but increased JC-1 aggregates intensity and OPA1 expression, indicating that DHY attenuated oxidative stress in HG-stimulated cardiomyocytes. DHY also attenuated CaMKII activity by suppressed PLB phosphorylation and inhibited CaMKII oxidation in HG-stimulated cardiomyocytes. ConclusionsHG-induced cardiomyocytes injury was alleviated wherein DHY attenuated necroptosis, repressed ROS production, and inhibited CaMKII oxidation, suggesting that DHY may serve as potential agent to prevent and treat diabetic cardiomyopathy.