Decrease In ATP Levels Research Articles

Low phosphorus causes hepatic energy metabolism disorder through Dynamin-related protein 1-mediated mitochondrial fission in fish

BackgroundLow phosphorus (LP) diets perturb hepatic energy metabolism homeostasis in fish. However, the specific mechanisms in LP-induced hepatic energy metabolism disorders remain to be fully elucidated. ObjectivesThis study sought to elucidate the underlying mechanisms of mitochondria involved in LP-induced energy metabolism disorders. MethodsSpotted seabass were fed diets with 0.72% (S-AP, control) or 0.36% (S-LP) available phosphorus for 10 weeks. Drp1 was knocked down or protein kinase A (PKA) was activated using 8Br-cAMP (5 μM, a PKA activator) in spotted seabass hepatocytes under LP medium. Zebrafish were fed Z-LP diets (0.30% available phosphorus) containing Mdivi-1 (5 mg/kg, a Drp1 inhibitor) or 8Br-cAMP (0.5 mg/kg) for 6 weeks. Biochemical and molecular parameters, along with transmission electron microscopy and immunofluorescence, were used to assess hepatic glycolipid metabolism, mitochondrial function and morphology. ResultsSpotted seabass fed S-LP diets showed reduced ATP (0.52-fold) and cyclic adenosine monophosphate (cAMP) (0.52-fold) levels, along with reduced Drp1 (s582) (0.38-fold) and PKA (0.61-fold) phosphorylation levels in the liver compared with those fed S-AP diets (P < 0.05). Drp1 knockdown elevated ATP levels (1.99-fold), decreased mitochondrial DRP1 protein levels (0.45-fold), and increased mitochondrial aspect ratio (1.82-fold) in LP-treated hepatocytes (P < 0.05). Furthermore, 8Br-cAMP-treated hepatocytes exhibited higher PKA phosphorylation (2.85-fold), ATP levels (1.60-fold), and mitochondrial aspect ratio (2.00-fold), along with decreased mitochondrial DRP1 protein levels (0.29-fold) under LP medium (P < 0.05). However, mutating s582 to alanine mimic Drp1 dephosphorylation decreased ATP levels (0.63-fold) and mitochondrial aspect ratio (0.53-fold) in 8Br-cAMP-treated hepatocytes (P < 0.05). In addition, zebrafish were fed Z-LP diets containing Mdivi-1 or 8Br-cAMP had higher ATP levels (3.44-fold or 1.98-fold) than that fed Z-LP diets (P < 0.05). ConclusionsThese findings provide a potential mechanistic elucidation for LP-induced energy metabolism disorders through the cAMP/PKA/Drp1-mediated mitochondrial fission signaling pathway.

Time-restricted feeding mediated modulation of microbiota leads to changes in muscle physiology in Drosophila obesity models.

Recent research has highlighted the essential role of the microbiome in maintaining skeletal muscle physiology. The microbiota influences muscle health by regulating lipid metabolism, protein synthesis, and insulin sensitivity. However, metabolic disturbances such as obesity can lead to dysbiosis, impairing muscle function. Time-restricted feeding (TRF) has been shown to mitigate obesity-related muscle dysfunction, but its effects on restoring healthy microbiomes remain poorly understood. This study utilizes 16S microbiome analysis and bacterial supplementation to investigate the bacterial communities influenced by TRF that may benefit skeletal muscle physiology. In wild-type and obese Drosophila models (axenic models devoid of natural microbial communities), the absence of microbiota influence muscle performance and metabolism differently. Specifically, axenic wild-type Drosophila exhibited reduced muscle performance, higher glucose levels, insulin resistance, ectopic lipid accumulation, and decreased ATP levels. Interestingly, in obese Drosophila (induced by a high-fat diet or predisposed obesity mutant Sk2), the absence of microbiota improved muscle performance, lowered glucose levels, reduced insulin resistance, and increased ATP levels. TRF was found to modulate microbiota composition, notably increasing Acetobacter pasteurianus (AP) and decreasing Staphylococcus aureus (SA) in both obesity models. Supplementation with AP improved muscle performance and reduced glucose and insulin resistance, while SA supplementation had the opposite effect. This study provides novel insights into the complex interactions between TRF, microbiota, and skeletal muscle physiology in different Drosophila models.

Investigating the regulatory mechanism of glucose metabolism by ubiquitin-like protein MNSFβ.

Monoclonal nonspecific suppressor factor β (MNSFβ), a ubiquitously expressed member of the ubiquitin-like protein family, is associated with diverse cell regulatory functions. It has been implicated in glycolysis regulation and cell proliferation enhancement in the macrophage-like cell line Raw264.7. This study aims to show that HIF-1α regulates MNSFβ-mediated metabolic reprogramming. In Raw264.7 cells, MNSFβ siRNA increased the oxygen consumption rate and reactive oxygen species (ROS) production but decreased ATP levels. Cells with MNSFβ knockdown showed a markedly increased ATP reduction rate upon the addition of oligomycin, a mitochondrial ATP synthase inhibitor. In addition, MNSFβ siRNA decreased the expression levels of mRNA and protein of HIF-1α-a regulator of glucose metabolism. Evaluation of the effect of MNSFβ on glucose metabolism in murine peritoneal macrophages revealed no changes in lactate production, glucose consumption, or ROS production. MNSFβ affects both glycolysis and mitochondrial metabolism, suggesting HIF-1α involvement in the MNSFβ-regulated glucose metabolism in Raw264.7 cells.

Urolithin A promotes the degradation of TMSB10 to deformation F-actin in non-small-cell lung cancer

BackgroundLung cancer is one of the most frequently diagnosed cancers and non-small-cell lung cancer (NSCLC) poses major diagnoses. Urolithin A (UA) is a natural compound produced by the gut microbiota through the metabolism of polyphenol ellagitannins (ETs) and ellagic acid (EA), which has been found to inhibit epithelial-mesenchymal transition (EMT) in lung cancer cell lines. However, the mechanism of UA function in NSCLC remains elusive. ProposeThis study aimed to investigate the potential effectiveness of UA in NSCLC therapeutic and uncovering its underlying mechanisms. MethodsEffects of UA treatment, TMSB10 gene knockdown or overexpression on NSCLC cell phenotype were evaluated by availability, transwell assays. The downstream factors and pathways of UA were investigated by proteomics. TMSB10 expression in NSCLC tissues was detected by bioinformatics analysis as well as immunohistochemistry. Confocal imaging, GST pull-down and western blotting investigated the mechanism of UA induced TMSB10 degradation. ResultsIn the present study, we demonstrated that UA shows an inhibitory role in NSCLC cell proliferation, migration, and invasion. This inhibition is attributed to the accelerated degradation of TMSB10, a biomarker among various cancers, via the autophagy-lysosome pathway. Additionally, knocked down of TMSB10 showed a similar phenotype with UA treatment. The reduction of TMSB10 protein level following decreased ATP level inhibits the F-actin formation for cell migration, thereby disrupting the equilibrium between G-actin-TMSB10 and G-actin-ATP interactions in A549 cells. ConclusionOur results reveal that UA is potential for NSCLC therapeutics through reducing the protein level of TMSB10 to deformation the F-actin.

Targeting POLRMT by IMT1 inhibits colorectal cancer cell growth

This study investigates the potential anti-colorectal cancer (CRC) activity of IMT1, a novel specific inhibitor of mitochondrial RNA polymerase (POLRMT). Single-cell RNA sequencing data reveal that POLRMT is overexpressed in CRC cells. Additionally, elevated POLRMT expression was observed in local CRC tissues and cells, while its expression remained relatively low in colon epithelial tissues and cells. IMT1 significantly inhibited colony formation, cell viability, proliferation, cell cycle progression, and migration in both primary and immortalized CRC cells. Furthermore, IMT1 induced apoptosis and cell death in CRC cells. The inhibition of POLRMT by IMT1 disrupted mitochondrial functions in CRC cells, leading to mitochondrial depolarization, oxidative damage, and decreased ATP levels. Using targeted shRNA to silence POLRMT closely mirrored the effects of IMT1, showing robust anti-CRC cell activity. Crucially, the efficacy of IMT1 was diminished in CRC cells with silenced POLRMT. Contrarily, boosting POLRMT expression externally by a lentiviral construct promoted the proliferation and migration of CRC cells. Importantly, treatment with IMT1 or silencing POLRMT in primary colon cancer cells decreased the phosphorylation of Akt1-S6K1, whereas overexpression of POLRMT had the opposite effect. In nude mice, orally administering IMT1 potently restrained primary colon cancer xenograft growth. IMT1 suppressed POLRMT activity, disrupted mitochondrial function, hindered Akt-mTOR activation, and prompted apoptosis within the xenograft tissues. In addition, IMT1 administration suppressed lung metastasis of primary colon cancer cells in nude mice. These combined results highlight the robust anti-CRC activity of IMT1 by specifically targeting POLRMT.

Calcium ion delivery by microbubble-assisted sonoporation stimulates cell death in human gastrointestinal cancer cells

Ultrasound-mediated cell membrane permeabilization – sonoporation, enhances drug delivery directly to tumor sites while reducing systemic side effects. The potential of ultrasound to augment intracellular calcium uptake – a critical regulator of cell death and proliferation – offers innovative alternative to conventional chemotherapy. However, calcium therapeutic applications remain underexplored in sonoporation studies. This research provides a comprehensive analysis of calcium sonoporation (CaSP), which combines ultrasound treatment with calcium ions and SonoVue microbubbles, on gastrointestinal cancer cells LoVo and HPAF-II. Initially, optimal sonoporation parameters were determined: an acoustic wave of 1 MHz frequency with a 50 % duty cycle at intensity of 2 W/cm2. Subsequently, various cellular bioeffects, such as viability, oxidative stress, metabolism, mitochondrial function, proliferation, and cell death, were assessed following CaSP treatment. CaSP significantly impaired cancer cell function by inducing oxidative and metabolic stress, evidenced by increased mitochondrial depolarization, decreased ATP levels, and elevated glucose uptake in a Ca2+ dose-dependent manner, leading to activation of the intrinsic apoptotic pathway. Cellular response to CaSP depended on the TP53 gene's mutational status: colon cancer cells were more susceptible to CaSP-induced apoptosis and G1 phase cell cycle arrest, whereas pancreatic cancer cells showed a higher necrotic response and G2 cell cycle arrest. These promising results encourage future research to optimize sonoporation parameters for clinical use, investigate synergistic effects with existing treatments, and assess long-term safety and efficacy in vivo. Our study highlights CaSP's clinical potential for improved safety and efficacy in cancer therapy, offering significant implications for the pharmaceutical and biomedical fields.

Evaluation of NAD+ precursors for improved metabolism and productivity of antibody-producing CHO cell.

In the previous study, the culture medium was treated with nicotinamide adenine dinucleotide (NAD+) under the hypothesis that NAD+ regeneration is a major factor causing excessive lactate accumulation in Chinese hamster ovary (CHO) cells. The NAD+ treatment improved metabolism by not only reducing the Warburg effect but also enhancing oxidative phosphorylation, leading to enhanced antibody production. Building on this, four NAD+ precursors - nicotinamide mononucleotide (NMN), nicotinic acid (NA), nicotinamide riboside (NR), and nicotinamide (NAM) - were tested to elevate intracellular NAD+ levels more economically. First, the ability of CHO cells to utilize both the salvage and Preiss-Handler pathways for NAD+ biosynthesis was verified, and then the effect of NAD+ precursors on CHO cell cultures was evaluated. These precursors increased intracellular NAD+ levels by up to 70.6% compared to the non-treated group. Culture analysis confirmed that all the precursors induced metabolic changes and that NMN, NA, and NR improved productivity akin to NAD+ treatment, with comparable integral viable cell density. Despite the positive effects such as the increase in the specific productivity and changes in cellular glucose metabolism, none of the precursors surpassed direct NAD+ treatment in antibody titer, presumably due to the reduction in nucleoside availability, as evidenced by the decrease in ATP levels in the NAD+ precursor-treated groups. These results underscore the complexity of cellular metabolism as well as the necessity for further investigation to optimize NAD+ precursor treatment strategies, potentially with the supplementation of nucleoside precursors. Our findings suggest a feasible approach for improving CHO cell culture performances by using NAD+ precursors as medium and feed components for the biopharmaceutical production.

Quiescence Enhances Survival during Viral Infection in Caenorhabditis elegans.

Infection causes reduced activity, anorexia, and sleep, which are components of the phylogenetically conserved but poorly understood sickness behavior. We developed a Caenorhabditis elegans model to study quiescence during chronic infection, using infection with the Orsay virus. The Orsay virus infects intestinal cells yet strongly affects behavior, indicating gut-to-nervous system communication. Infection quiescence has the sleep properties of reduced responsiveness and rapid reversibility. Both the ALA and RIS neurons regulate virus-induced quiescence though ALA plays a more prominent role. Quiescence-defective animals have decreased survival when infected, indicating a benefit of quiescence during chronic infectious disease. The survival benefit of quiescence is not explained by a difference in viral load, indicating that it improves resilience rather than resistance to infection. Orsay infection is associated with a decrease in ATP levels, and this decrease is more severe in quiescence-defective animals. We propose that quiescence preserves energetic resources by reducing energy expenditures and/or by increasing extraction of energy from nutrients. This model presents an opportunity to explore the role of sleep and fatigue in chronic infectious illness.

Rational design of an activatable dual-color fluorogenic probe for revealing the interaction of adenosine-5′-triphosphate and peroxynitrite in pyroptosis associated with acute kidney injury

ATP and ONOO- play unique roles in various biological events and exhibit notable interactions. To date, there is no available chemical tool for investigating the correlation between ATP and ONOO- concentrations in pyroptosis associated with acute kidney injury (AKI). Herein, we designed a novel dual-color near-infrared fluorescent (NIRF) probe P2 for simultaneous visualization of ATP and ONOO- both in vitro and in vivo. Unlike previously reported single-site fluorescent probes, P2 enabled concurrent imaging of ATP and ONOO- in two distinct fluorescence channels, with emission wavelengths centered at 585 and 690 nm, which greatly reduced spectral cross-talk. Employing a HK-2 pyroptosis model, a significant interaction between ATP and ONOO- was unveiled. Elevated ONOO- production was found to correlate with decreased ATP levels; conversely, an increase in ATP levels was associated with rapid ONOO- scavenging. Remarkably, P2 allowed the assessment of cellular hypoxia by monitoring ATP and ONOO- concentrations. The commercial ONOO--scavenger uric acid showcased a protective effect on HK-2 cells via inhibition of the cellular pyroptosis pathway. Furthermore, P2 was successfully employed for imaging of ATP and ONOO- in the AKI mice model. Our findings confirmed that renal ischemia-reperfusion triggered a rise in ONOO- levels, concurrent with a decline in ATP levels. Surprisingly, the cells exhibited a compensatory recovery of ATP levels as the reperfusion time was prolonged. These results suggested the newly devised P2, as a pivotal chemical tool for the simultaneous monitoring of ATP and ONOO-, might open new avenues for disease diagnosis and treatment.

Premeiotic deletion of Eif2s2 causes oocyte arrest at the early diplotene stage and apoptosis in mice.

Eukaryotic translation initiation factor 2 subunit 2 (EIF2S2), a subunit of the heterotrimeric G protein EIF2, is involved in the initiation of translation. Our findings demonstrate that the depletion of Eif2s2 in premeiotic germ cells causes oocyte arrest at the pachytene and early diplotene stages at 1 day postpartum (dpp) and 5 dpp, respectively, and eventually leads to oocyte apoptosis and failure of primordial follicle formation. Further studies reveal that Eif2s2 deletion downregulates homologous recombination-related and mitochondrial fission-related protein levels, and upregulates the integrated stress response-related proteins and mRNA levels. Consistently, Eif2s2 deletion significantly decreases the expression of dictyate genes and compromises mitochondrial function, characterized by elongated shapes, decreased ATP levels and mtDNA copy number, along with an excessive accumulation of reactive oxygen species (ROS) and mitochondrial superoxide. Furthermore, DNA damage response and proapoptotic protein levels increase, while anti-apoptotic protein levels decrease in Eif2s2-deleted mice. An increase in oocytes with positive cleaved-Caspase-3 and TUNEL signals, alongside reduced Lamin B1 intensity, further indicates oocyte apoptosis. Collectively, Eif2s2 deletion in premeiotic germ cells causes oocyte meiotic arrest at the early diplotene stage by impairing homologous recombination, and eventually leads to oocyte apoptosis mainly through the downregulation of mitochondrial fission-related proteins, ROS accumulation and subsequent DNA damage.

Neuronostatin regulates neuronal function and energetic metabolism in Alzheimer's disease in a GPR107-dependent manner

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease, which is characterized by the accumulation and aggregation of amyloid in brain. Neuronostatin (NST) is an endogenous peptide hormone that participates in many fundamental neuronal processes. However, the metabolism and function of NST in neurons of AD mice are not known. In this study, by combining the structural analyses, primary cultures, knockout cells, and various assessments, the behavior, histopathology, brain-wide expression and cellular signaling pathways in the APP/PS1 mice were investigated. It was found that NST directly bound to GPR107, which was primarily expressed in neurons. NST modulated the neuronal survivability and neurite outgrowth induced by Aβ via GPR107 in neurons. Intracerebroventricular (i.c.v.) administration of NST attenuated learning and memory abilities, reduced the synaptic protein levels of hippocampus, but improved amyloid plaques in the cortex and hippocampus of APP/PS1 mice. NST modulated glucose metabolism of hypothalamus-hippocampus-cortex axis in APP/PS1 mice and decreased ATP levels via the regulation of reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in response to Aβ, suppressed energetic metabolism, and mitochondrial function in neurons via GPR107/protein kinase A (PKA) signaling pathway. In summary, our findings suggest that NST regulates neuronal function and brain energetic metabolism in AD mice via the GPR107/PKA signaling pathway, which can be a promising target for the treatment of AD.

Traffic-related ultrafine particles impair mitochondrial functions in human olfactory mucosa cells – Implications for Alzheimer's disease

Constituents of air pollution, the ultrafine particles (UFP) with a diameter of ≤0.1 μm, are considerably related to traffic emissions. Several studies link air pollution to Alzheimer's disease (AD), yet the exact relationship between the two remains poorly understood. Mitochondria are known targets of environmental toxicants, and their dysfunction is associated with neurodegenerative diseases. The olfactory mucosa (OM), located at the rooftop of the nasal cavity, is directly exposed to the environment and in contact with the brain. Mounting evidence suggests that the UFPs can impact the brain directly through the olfactory tract. By using primary human OM cultures established from nasal biopsies of cognitively healthy controls and individuals diagnosed with AD, we aimed to decipher the effects of traffic-related UFPs on mitochondria. The UFP samples were collected from the exhausts of a modern heavy-duty diesel engine (HDE) without aftertreatment systems, run with renewable diesel (A0) and petroleum diesel (A20), and from an engine of a 2019 model diesel passenger car (DI-E6d) equipped with state-of-the-art aftertreatment devices and run with renewable diesel (Euro6). OM cells were exposed to three different UFPs for 24-h and 72-h, after which cellular processes were assessed on the functional and transcriptomic levels. Our results show that UFPs impair mitochondrial functions in primary human OM cells by hampering oxidative phosphorylation (OXPHOS) and redox balance, and the responses of AD cells differ from cognitively healthy controls. RNA-Seq and IPA® revealed inhibition of OXPHOS and mitochondrial dysfunction in response to UFPs A0 and A20. Functional validation confirmed that A0 and A20 impair cellular respiration, decrease ATP levels, and disturb redox balance by altering NAD and glutathione metabolism, leading to increased ROS and oxidative stress. RNA-Seq and functional assessment revealed the presence of AD-related alterations in human OM cells and that different fuels and engine technologies elicit differential effects.

Galactose-1-phosphate inhibits cytochrome c oxidase and causes mitochondrial dysfunction in classic galactosemia

Classic galactosemia is an inborn error of metabolism caused by mutations in the GALT gene resulting in the diminished activity of the galactose-1-phosphate uridyltransferase enzyme. This reduced GALT activity leads to the buildup of the toxic intermediate galactose-1-phosphate and a decrease in ATP levels upon exposure to galactose. In this work, we focused our attention on mitochondrial oxidative phosphorylation in the context of this metabolic disorder. We observed that galactose-1-phosphate accumulation reduced respiratory rates in vivo and changed mitochondrial function and morphology in yeast models of galactosemia. These alterations are harmful to yeast cells since the mitochondrial retrograde response is activated as part of the cellular adaptation to galactose toxicity. In addition, we found that galactose-1-phosphate directly impairs cytochrome c oxidase activity of mitochondrial preparations derived from yeast, rat liver, and human cell lines. These results highlight the evolutionary conservation of this biochemical effect. Finally, we discovered that two compounds - oleic acid and dihydrolipoic acid – that can improve the growth of cell models of mitochondrial diseases, were also able to improve galactose tolerance in this model of galactosemia. These results reveal a new molecular mechanism relevant to the pathophysiology of classic galactosemia - galactose-1-phosphate-dependent mitochondrial dysfunction - and suggest that therapies designed to treat mitochondrial diseases may be repurposed to treat galactosemia.

P-113 Comparison of mitochondrial function between expanded and hatching human blastocysts

Abstract Study question Is there a difference in mitochondrial function between expanded and hatching fresh and vitrified blastocysts? Summary answer Freezing and thawing may lead to mitochondrial dysfunction. What is known already Mitochondria, as crucial cellular organelles, play role in energy production through the electron transport chain, cell motility and organelle movement. They also have a significant impact on embryonic development. Furthermore, during the process of embryonic hatching, the energy generated by these organelles can affect ion pump activity, blastocyst expansion, and collapse. Therefore, factors that stress embryonic mitochondria may contribute to embryogenesis and hatching. Cooling and warming, as stress factors, maybe result in organelle damage, protein denaturation, membrane disruption, and DNA fragmentation. Additionally, they can cause changes in mitochondrial function, leading to decreased ATP levels and increased ROS levels. Study design, size, duration In this study, vitrified and fresh human 8-cell embryos were collected from April 18, 2021, to July 19, 2023. The embryos were cultured until the blastocyst stage, and then four samples were assessed in four groups. These groups included vitrified expanded blastocysts, vitrified hatching blastocysts, fresh expanded blastocysts, and fresh hatching blastocysts. Participants/materials, setting, methods Vitrified embryos were sourced from the embryo bank of the Royan Institute, while fresh embryos were obtained from good-quality 3PN embryos or excess embryos from couples who did not wish to freeze them. A stock solution containing lipophilic cationic JC-1 dye in dimethyl sulfoxide was prepared. Blastocysts were then exposed to a concentration of 1 μg/ml of JC-1 diluted in HamsF10-HSA. After washing with HF-HSA, imaging was performed using a 2-photon confocal microscope. Main results and the role of chance Our results showed that mitochondrial membrane potential in expanded blastocysts of both fresh and vitrified eight-cell embryos was significantly higher than in hatching blastocysts of fresh and vitrified embryos (P &amp;lt; 0.05). However, there was no significant difference in mitochondrial function between the vitrified and fresh groups, neither in expanded nor hatching blastocysts. Limitations, reasons for caution Due to the human origin of the samples, the minimum necessary replicates were considered for each experimental group. Wider implications of the findings In this study, it appears that plump zona breakers can be detected in hatching blastocysts using confocal microscopy. Previous studies, such as the one conducted by Sathananthan, identified these cells using a TEM microscope. Trial registration number -

Mutation of mpv17 results in loss of iridophores due to mitochondrial dysfunction in tilapia

Abstract Mpv17 (mitochondrial inner membrane protein MPV17) deficiency causes severe mitochondrial DNA depletion syndrome in mammals and loss of pigmentation of iridophores and a significant decrease of melanophores in zebrafish. The reasons for this are still unclear. In this study, we established an mpv17 homozygous mutant line in Nile tilapia. The developing mutants are transparent due to the loss of iridophores and aggregation of pigment granules in the melanophores and disappearance of the vertical pigment bars on the side of the fish. Transcriptome analysis using the skin of fish at 30 dpf (days post fertilization) revealed that the genes related to purine (especially pnp4a) and melanin synthesis were significantly downregulated. However, administration of guanine diets failed to rescue the phenotype of the mutants. In addition, no obvious apoptosis signals were observed in the iris of the mutants by TUNEL staining. Significant downregulation of genes related to iridophore differentiation was detected by qPCR. Insufficient ATP, as revealed by ATP assay, α-MSH treatment, and adcy5 mutational analysis, might account for the defects of melanophores in mpv17 mutants. Several tissues displayed less mtDNA and decreased ATP levels. Taken together, these results indicated that mutation of mpv17 led to mitochondrial dTMP deficiency, followed by impaired mtDNA content and mitochondrial function, which in turn, led to loss of iridophores and a transparent body color in tilapia.

Exogenous Oxidative Stress in Human Spermatozoa Induces Opening of the Mitochondrial Permeability Transition Pore: Effect on Mitochondrial Function, Sperm Motility and Induction of Cell Death.

Oxidative stress (OS) and disrupted antioxidant defense mechanisms play a pivotal role in the etiology of male infertility. The alterations in reactive oxygen species (ROS) production and calcium (Ca2+) homeostasis are the main activators for the mitochondrial permeability transition pore (mPTP) opening. The mPTP opening is one of the main mechanisms involved in mitochondrial dysfunction in spermatozoa. This alteration in mitochondrial function adversely affects energy supply, sperm motility, and fertilizing capacity and contributes to the development of male infertility. In human spermatozoa, the mPTP opening has been associated with ionomycin-induced endogenous oxidative stress and peroxynitrite-induced nitrosative stress; however, the effect of exogenous oxidative stress on mPTP opening in sperm has not been evaluated. The aim of this study was to determine the effect of exogenous oxidative stress induced by hydrogen peroxide (H2O2) on mPTP opening, mitochondrial function, motility, and cell death markers in human spermatozoa. Human spermatozoa were incubated with 3 mmol/L of H2O2 for 60 min, and intracellular Ca2+ concentration, mPTP opening, mitochondrial membrane potential (ΔΨm), ATP levels, mitochondrial reactive oxygen species (mROS) production, phosphatidylserine (PS) externalization, DNA fragmentation, viability, and sperm motility were evaluated. H2O2-induced exogenous oxidative stress caused increased intracellular Ca2+, leading to subsequent mPTP opening and alteration of mitochondrial function, characterized by ΔΨm dissipation, decreased ATP levels, increased mROS production, and the subsequent alteration of sperm motility. Furthermore, H2O2-induced opening of mPTP was associated with the expression of apoptotic cell death markers including PS externalization and DNA fragmentation. These results highlight the role of exogenous oxidative stress in causing mitochondrial dysfunction, deterioration of sperm motility, and an increase in apoptotic cell death markers, including PS externalization and DNA fragmentation, through the mPTP opening. This study yielded new knowledge regarding the effects of this type of stress on mitochondrial function and specifically on mPTP opening, factors that can contribute to the development of male infertility, considering that the role of mPTP in mitochondrial dysfunction in human sperm is not completely elucidated. Therefore, these findings are relevant to understanding male infertility and may provide an in vitro model for further research aimed at improving human sperm quality.

UW Supplementation with AP39 Improves Liver Viability Following Static Cold Storage.

Static cold storage of donor livers at 4°C incompletely arrests metabolism, ultimately leading to decreases in ATP levels, oxidative stress, cell death, and organ failure. Hydrogen Sulfide (H2S) is an endogenously produced gas, previously demonstrated to reduce oxidative stress, reduce ATP depletion, and protect from ischemia and reperfusion injury. H2S is difficult to administer due to its rapid release curve, resulting in cellular death at high concentrations. AP39, a mitochondrially targeted, slow-release H2S donor, has been shown to reduce ischemia-reperfusion injury in hearts and kidneys. Thus, we investigated whether the addition of AP39 during 3-day static cold storage can improve liver graft viability. At the end of storage, livers underwent six hours of acellular normothermic machine perfusion, a model of transplantation. During simulated transplantation, livers stored with AP39 showed reduced resistance, reduced cellular damage (ALT and AST), and reduced apoptosis. Additionally, bile production and glucose, as well as energy charge were improved by the addition of AP39. These results indicate that AP39 supplementation improves liver viability during static cold storage.

Distinct epigenetic modulation of differentially expressed genes in the adult mouse brain following prenatal exposure to low-dose bisphenol A.

Bisphenol A (BPA) is a common component in the manufacture of daily plastic consumer goods. Recent studies have suggested that prenatal exposure to BPA can increase the susceptibility of offspring to mental illness, although the underlying mechanisms remain unclear. In this study, we performed transcriptomic and epigenomic profiling in the adult mouse brain following prenatal exposure to low-dose BPA. We observed a sex-specific transcriptional dysregulation in the cortex, with more significant differentially expressed genes was observed in adult cortex from male offspring. Moreover, the upregulated genes primarily influenced neuronal functions, while the downregulated genes were significantly associated with energy metabolism pathways. More evidence supporting impaired mitochondrial function included a decreased ATP level and a reduced number of mitochondria in the cortical neuron of the BPA group. We further investigated the higher-order chromatin regulatory patterns of DEGs by incorporating published Hi-C data. Interestingly, we found that upregulated genes exhibited more distal interactions with multiple enhancers, while downregulated genes displayed relatively short-range interactions among adjacent genes. Our data further revealed decreased H3K9me3 signal on the distal enhancers of upregulated genes, whereas increased DNA methylation and H3K27me3 signals on the promoters of downregulated genes. In summary, our study provides compelling evidence for the potential health risks associated with prenatal exposure to BPA, and uncovers sex-specific transcriptional changes with a complex interplay of multiple epigenetic mechanisms.

Lactate dehydrogenase B as a metabolism-related marker for immunotherapy in head and neck squamous cell carcinoma

BackgroundHead and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies. Lactate dehydrogenase family genes (LDHs) play a critical role in tumor metabolism, but their functions in HNSCC have not been investigated thoroughly. Thus, we aimed to explore the value of LDHs in HNSCC. MethodsThe association between LDHs expression and mutations, methylation, copy number variations (CNVs), alternative splicing (AS) and competing endogenous RNA (ceRNA) was investigated in The Cancer Genome Atlas (TCGA). The expression level of LDHs in OSCC tissues and adjacent normal tissues was verified by qPCR. Algorithms, such as ssGSEA, ESTIMATE, xCell and TIDE were utilized to analyze the characteristics of immune infiltration. Pathway alternations were enriched by GO, GSEA and KEGG analysis. The Mantel test was employed to elucidate the correlation between metabolism and the tumor microenvironment (TME). Subsequently, MTT and colony formation assays were utilized to assess the impact of LDHB knockdown on cellular proliferation. Additionally, ATP and lactate assays were performed to examine metabolic alterations. Co-culture experiments further investigated the effect of LDHB knockdown on T cell differentiation. ResultsLDHs were completely analyzed in multiple databases, among which LDHB was differentially expressed in HNSCC and significantly associated with prognosis. Low LDHB expression had better clinicopathological characteristics. Downregulated LDHB expression was associated with enhanced immune cell infiltration and could influence tumor metabolism. Despite having worse cytotoxic T lymphocyte dysfunction, the LDHBlow group was predicted to respond more favorably to immune checkpoint inhibitors (ICIs) therapy. Moreover, the correlation between metabolism and TME was depicted. In vitro, LDHB knockdown resulted in inhibited cell proliferation, increased lactate levels and decreased ATP levels, while promoted the Th1 differentiation of T cells. ConclusionsOur study provided a comprehensive analysis of the LDHs and illustrated low LDHB expression could inhibit tumor cell proliferation and ATP production by influencing metabolism, with improved immune cell infiltration and better response to immunotherapy.

Impaired Long-Chain Fatty Acid Metabolism in Carnitine Palmitoyltransferase II Knockout Mouse Leads to Chronic Pancreatitis

Background: To support high rates of protein synthesis, pancreatic acinar cells must generate large quantities of ATP. Mitochondrial dysfunction and impaired energy metabolism play a significant role in the development of pancreatitis. Patients with elevated plasma triglycerides are at increased risk for development of severe acute pancreatitis. Carnitine palmitoyltransferase II (CPT2) is required for importing and thus metabolizing long-chain fatty acids (LCFAs) in mitochondria. To inhibit LCFA metabolism, we utilized the CPT2acinarKO mouse to probe the consequences of impaired LCFA import on pancreatic function. Hypothesis: Loss of CPT2 causes an accumulation of long-chain acylcarnitines and disruption of the tissue acylcarnitine/free carnitine ratio which is thought to alter cellular function. Given acinar cells’ high metabolic requirement, impaired LCFA utilization would reduce energy production, secretory function, and alter pancreatic lipid composition. Methods: We generated mice lacking CPT2 specifically in acinar cells (CPT2acinarKO) by crossing tamoxifen-inducible, Elastase-Cre mice with CPT2 floxed mice. Mice were fed a high fat (60% kcal fat), low fat (10% kcal fat), or chow (14% kcal fat) diet for 12 weeks. Analysis of fat accumulation, edema, and fibrosis was performed with H&amp;E and picrosirius staining. Mass Spec. lipid profiling was performed in pancreas and serum. Acinar cells were isolated from wildtype, CPT2 floxed, and CPT2acinarKO mice to assess secretory function, ATP levels, and necrosis via lactate dehydrogenase (LDH) release. Results: Histologic examination revealed CPT2acinarKO mice on all three diets displayed pancreatic fat accumulation. Strikingly, CPT2acinarKO mice fed a chow or low fat diet developed chronic pancreatitis with a 25% loss in pancreatic weight, immune cell infiltration, edema, and fibrosis. As anticipated, long-chain acylcarnitines were significantly elevated in CPT2acinarKO pancreas. Assessment of secretion in 2 weeks post tamoxifen cells revealed significantly reduced maximal (10 pM) and supramaximal (1 nM) cholecystokinin (CCK) stimulated amylase release as a percent of total cellular amylase. Acini isolated at 4 weeks post tamoxifen revealed significantly increased basal secretion and reduced maximal and supramaximal CCK stimulated amylase release. When normalized to cellular DNA, secretion was similar between CPT2acinarKO and CPT2 floxed acinar cells indicating CPT2acinarKO cells had reduced amylase levels. Wildtype acinar cells treated with unsaturated and saturated free fatty acid concentrations as low as 100 μM for 3 hours caused significant necrosis. CPT2acinarKO pancreas had significantly decreased ATP levels indicative of mitochondrial dysfunction. Conclusion: Loss of CPT2 in the pancreatic acinar cells impairs LCFA import, reduces ATP levels, disrupts the secretory pathway, promoting chronic pancreatitis. In contrast to other CPT2 knockout models, pancreatitis occurs independent of high fat feeding in CPT2acinarKO mice. Further investigation is needed to identify metabolic alterations leading to acinar damage and mitochondrial dysfunction following the loss of CPT2. UW-Madison Comprehensive Diabetes Center Pilot Grant. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.