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Nature Aug 2023Alveolar epithelial type 1 (AT1) cells are necessary to transfer oxygen and carbon dioxide between the blood and air. Alveolar epithelial type 2 (AT2) cells serve as a...
Alveolar epithelial type 1 (AT1) cells are necessary to transfer oxygen and carbon dioxide between the blood and air. Alveolar epithelial type 2 (AT2) cells serve as a partially committed stem cell population, producing AT1 cells during postnatal alveolar development and repair after influenza A and SARS-CoV-2 pneumonia. Little is known about the metabolic regulation of the fate of lung epithelial cells. Here we report that deleting the mitochondrial electron transport chain complex I subunit Ndufs2 in lung epithelial cells during mouse gestation led to death during postnatal alveolar development. Affected mice displayed hypertrophic cells with AT2 and AT1 cell features, known as transitional cells. Mammalian mitochondrial complex I, comprising 45 subunits, regenerates NAD and pumps protons. Conditional expression of yeast NADH dehydrogenase (NDI1) protein that regenerates NAD without proton pumping was sufficient to correct abnormal alveolar development and avert lethality. Single-cell RNA sequencing revealed enrichment of integrated stress response (ISR) genes in transitional cells. Administering an ISR inhibitor or NAD precursor reduced ISR gene signatures in epithelial cells and partially rescued lethality in the absence of mitochondrial complex I function. Notably, lung epithelial-specific loss of mitochondrial electron transport chain complex II subunit Sdhd, which maintains NAD regeneration, did not trigger high ISR activation or lethality. These findings highlight an unanticipated requirement for mitochondrial complex I-dependent NAD regeneration in directing cell fate during postnatal alveolar development by preventing pathological ISR induction.
Topics: Animals; Mice; Alveolar Epithelial Cells; Cell Differentiation; Cell Lineage; Lung; Mitochondria; NAD; NADH Dehydrogenase; Protons; RNA-Seq; Saccharomyces cerevisiae; Single-Cell Gene Expression Analysis; Stress, Physiological
PubMed: 37558881
DOI: 10.1038/s41586-023-06423-8 -
Frontiers in Immunology 2023Dysregulated inflammation is associated with many skeletal diseases and disorders, such as osteolysis, non-union of fractures, osteonecrosis, osteoarthritis and...
BACKGROUND
Dysregulated inflammation is associated with many skeletal diseases and disorders, such as osteolysis, non-union of fractures, osteonecrosis, osteoarthritis and orthopaedic infections. We previously showed that continuous infusion of lipopolysaccharide (LPS) contaminated polyethylene particles (cPE) caused prolonged inflammation and impaired bone formation. However, the metabolic and bioenergetic processes associated with inflammation of bone are unknown. Mitochondria are highly dynamic organelles that modulate cell metabolism and orchestrate the inflammatory responses that involve both resident and recruited cells. Glycolytic reprogramming, the shift from oxidative phosphorylation (OXPHOS) to glycolysis causes inappropriate cell activation and function, resulting in dysfunctional cellular metabolism. We hypothesized that impaired immunoregulation and bone regeneration from inflammatory states are associated with glycolytic reprogramming and mitochondrial dysfunction in macrophages (Mφ) and mesenchymal stromal cells (MSCs).
METHODS
We used the Seahorse XF96 analyzer and real-time qPCR to study the bioenergetics of Mφ and MSCs exposed to cPE. To understand the oxygen consumption rate (OCR), we used Seahorse XF Cell Mito Stress Test Kit with Seahorse XF96 analyzer. Similarly, Seahorse XF Glycolytic Rate Assay Kit was used to detect the extracellular acidification rate (ECAR) and Seahorse XF Real-Time ATP Rate Assay kit was used to detect the real-time ATP production rates from OXPHOS and glycolysis. Real-time qPCR was performed to analyze the gene expression of key enzymes in glycolysis and mitochondrial biogenesis. We further detected the gene expression of proinflammatory cytokines in Mφ and genes related to cell differentiation in MSC during the challenge of cPE.
RESULTS
Our results demonstrated that the oxidative phosphorylation of Mφ exposed to cPE was significantly decreased when compared with the control group. We found reduced basal, maximal and ATP-production coupled respiration rates, and decreased proton leak in Mφ during challenge with cPE. Meanwhile, Mφ showed increased basal glycolysis and proton efflux rates (PER) when exposed to cPE. The percentage (%) of PER from glycolysis was higher in Mφ exposed to cPE, indicating that the contribution of the glycolytic pathway to total extracellular acidification was elevated during the challenge of cPE. In line with the results of OCR and ECAR, we found Mφ during cPE challenge showed higher glycolytic ATP (glycoATP) production rates and lower mitochondrial ATP (mitoATP) production rates which is mainly from OXPHOS. Interestingly, MSCs showed enhanced glycolysis during challenge with cPE, but no significant changes in oxygen consumption rates (OCR). In accordance, seahorse assay of real-time ATP revealed glycoATP rates were elevated while mitoATP rates showed no significant differences in MSC during challenge with cPE. Furthermore, Mφ and MSCs exposed to cPE showed upregulated gene expression levels of glycolytic regulators and Mφ exposed to cPE expressed higher levels of pro-inflammatory cytokines.
CONCLUSION
This study demonstrated the dysfunctional bioenergetic activity of bone marrow-derived Mφ and MSCs exposed to cPE, which could impair the immunoregulatory properties of cells in the bone niche. The underlying molecular defect related to disordered mitochondrial function could represent a potential therapeutic target during the resolution of inflammation.
Topics: Humans; Protons; Glycolysis; Inflammation; Mesenchymal Stem Cells; Macrophages; Cytokines; Adenosine Triphosphate
PubMed: 37675119
DOI: 10.3389/fimmu.2023.1199751 -
Pflugers Archiv : European Journal of... Apr 2024Low pH in the gut is associated with severe inflammation, fibrosis, and colorectal cancer (CRC) and is a hallmark of active inflammatory bowel disease (IBD).... (Review)
Review
Low pH in the gut is associated with severe inflammation, fibrosis, and colorectal cancer (CRC) and is a hallmark of active inflammatory bowel disease (IBD). Subsequently, pH-sensing mechanisms are of interest for the understanding of IBD pathophysiology. Tissue hypoxia and acidosis-two contributing factors to disease pathophysiology-are linked to IBD, and understanding their interplay is highly relevant for the development of new therapeutic options. One member of the proton-sensing G protein-coupled receptor (GPCR) family, GPR65 (T-cell death-associated gene 8, TDAG8), was identified as a susceptibility gene for IBD in a large genome-wide association study. In response to acidic extracellular pH, GPR65 induces an anti-inflammatory response, whereas the two other proton-sensing receptors, GPR4 and GPR68 (ovarian cancer G protein-coupled receptor 1, OGR1), mediate pro-inflammatory responses. Here, we review the current knowledge on the role of these proton-sensing receptors in IBD and IBD-associated fibrosis and cancer, as well as colitis-associated cancer (CAC). We also describe emerging small molecule modulators of these receptors as therapeutic opportunities for the treatment of IBD.
Topics: Humans; Protons; Genome-Wide Association Study; Colitis; Receptors, G-Protein-Coupled; Inflammatory Bowel Diseases; Hydrogen-Ion Concentration; Fibrosis
PubMed: 38514581
DOI: 10.1007/s00424-024-02943-y -
GeroScience Aug 2023Age-associated diseases are becoming progressively more prevalent, reflecting the increased lifespan of the world's population. However, the fundamental mechanisms of... (Review)
Review
Age-associated diseases are becoming progressively more prevalent, reflecting the increased lifespan of the world's population. However, the fundamental mechanisms of physiologic aging are poorly understood, and in particular, the molecular pathways that mediate cardiac aging and its associated dysfunction are unclear. Here, we focus on certain ion flux abnormalities of the mitochondria that may contribute to cardiac aging and age-related heart failure. Using oxidative phosphorylation, mitochondria pump protons from the matrix to the intermembrane space to generate a proton gradient across the inner membrane. The protons are returned to the matrix by the ATPase complex within the membrane to generate ATP. However, a portion of protons leak back to the matrix and do not drive ATP production, and this event is called proton leak or uncoupling. Accumulating evidence suggests that mitochondrial proton leak is increased in the cardiac myocytes of aged hearts. In this mini-review, we discuss the measurement methods and major sites of mitochondrial proton leak with an emphasis on the adenine nucleotide transporter 1 (ANT1), and explore the possibility of inhibiting augmented mitochondrial proton leak as a therapeutic intervention to mitigate cardiac aging.
Topics: Protons; Ion Channels; Mitochondrial Proteins; Mitochondria; Adenosine Triphosphate
PubMed: 36856945
DOI: 10.1007/s11357-023-00757-x -
Nature Structural & Molecular Biology Jul 2023Proton transport is indispensable for cell life. It is believed that molecular mechanisms of proton movement through different types of proton-conducting molecules have...
Proton transport is indispensable for cell life. It is believed that molecular mechanisms of proton movement through different types of proton-conducting molecules have general universal features. However, elucidation of such mechanisms is a challenge. It requires true-atomic-resolution structures of all key proton-conducting states. Here we present a comprehensive function-structure study of a light-driven bacterial inward proton pump, xenorhodopsin, from Bacillus coahuilensis in all major proton-conducting states. The structures reveal that proton translocation is based on proton wires regulated by internal gates. The wires serve as both selectivity filters and translocation pathways for protons. The cumulative results suggest a general concept of proton translocation. We demonstrate the use of serial time-resolved crystallography at a synchrotron source with sub-millisecond resolution for rhodopsin studies, opening the door for principally new applications. The results might also be of interest for optogenetics since xenorhodopsins are the only alternative tools to fire neurons.
Topics: Protons; Proton Pumps; Ion Transport
PubMed: 37386213
DOI: 10.1038/s41594-023-01020-9 -
Acta Neurologica Taiwanica Dec 2023Myalgia (also called muscle pain or muscle ache) is a symptom associated with many diseases, including fibromyalgia, neurodegenerative diseases, degenerative spine...
Myalgia (also called muscle pain or muscle ache) is a symptom associated with many diseases, including fibromyalgia, neurodegenerative diseases, degenerative spine diseases, etc. Myalgia is a major medical problem affecting 60~85% of the population (lifetime prevalence). However, our understanding of chronic myalgia is still limited and effective treatment for intractable myalgia like fibromyalgia is still lacking. Although multifactorial, one known source of muscle pain is tissue acidosis. Experimental muscle pain can be induced by the intramuscular infusion of a buffered acidic solution in humans. As well, animal studies have revealed that acidic infusion activates chemosensitive nociceptors via the proton-sensing ion channels and receptors. Intriguingly, acid signaling in muscle afferents is promiscuous and could be either pro-nociceptive or antinociceptive, so we have coined the term sngception to describe the somatosensory function of acid sensation. Recent single-cell RNAseq studies have shown proton-sensing ion channels and receptors are expressed in all subpopulations of the somatosensory neurons, including nociceptors and non-nociceptive mechanoreceptors. Here, we address how the acid signaling is integrated in muscle afferents and why muscle pain can be chronic and intractable in mouse models of fibromyalgia. Besides acidosis, we have recently found oxidative stress can be another factor to activate proton-sensing ion channels and thus trigger fibromyalgia-like pain in mice. Together, understanding how the acid signaling works in muscle afferents will provide novel therapeutic strategies for myalgia.
Topics: Humans; Mice; Animals; Myalgia; Fibromyalgia; Protons; Ion Channels; Acidosis
PubMed: 37967833
DOI: No ID Found -
JAMA Network Open Aug 2023Radiotherapy plays an important role in the treatment of esophageal cancer. Proton therapy has unique physical properties and higher relative biological effectiveness.... (Meta-Analysis)
Meta-Analysis
IMPORTANCE
Radiotherapy plays an important role in the treatment of esophageal cancer. Proton therapy has unique physical properties and higher relative biological effectiveness. However, whether proton therapy has greater benefit than photon therapy is still unclear.
OBJECTIVE
To evaluate whether proton was associated with better efficacy and safety outcomes, including dosimetric, prognosis, and toxic effects outcomes, compared with photon therapy and to evaluate the efficacy and safety of proton therapy singly.
DATA SOURCES
A systematic search of PubMed, Embase, the Cochrane Library, Web of Science, SinoMed, and China National Knowledge Infrastructure databases was conducted for articles published through November 25, 2021, and updated to March 25, 2023.
STUDY SELECTION
For the comparison of proton and photon therapy, studies including dosimetric, prognosis, and associated toxic effects outcomes were included. The separate evaluation of proton therapy evaluated the same metrics.
DATA EXTRACTION AND SYNTHESIS
Data on study design, individual characteristics, and outcomes were extracted. If I2 was greater than 50%, the random-effects model was selected. This meta-analysis is reported following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) reporting guideline.
MAIN OUTCOMES AND MEASURES
The main outcomes were organs at risk (OARs) dosimetric outcomes, prognosis (overall survival [OS], progression-free survival [PFS], and objective response rate [ORR]), and radiation-related toxic effects.
RESULTS
A total of 45 studies were included in the meta-analysis. For dosimetric analysis, proton therapy was associated with significantly reduced OARs dose. Meta-analysis showed that photon therapy was associated with poor OS (hazard ratio [HR], 1.31; 95% CI, 1.07-1.61; I2 = 11%), but no difference in PFS was observed. Subgroup analysis showed worse OS (HR, 1.42; 95% CI, 1.14-1.78; I2 = 34%) and PFS (HR, 1.48; 95% CI, 1.06-2.08; I2 = 7%) in the radical therapy group with photon therapy. The pathological complete response rate was similar between groups. Proton therapy was associated with significantly decreased grade 2 or higher radiation pneumonitis and pericardial effusion, and grade 4 or higher lymphocytopenia. Single-rate analysis of proton therapy found 89% OS and 65% PFS at 1 year, 71% OS and 56% PFS at 2 years, 63% OS and 48% PFS at 3 years, and 56% OS and 42% PFS at 5 years. The incidence of grade 2 or higher radiation esophagitis was 50%, grade 2 or higher radiation pneumonitis was 2%, grade 2 or higher pleural effusion was 4%, grade 2 or higher pericardial effusion was 3%, grade 3 or higher radiation esophagitis was 8%, and grade 4 or higher lymphocytopenia was 17%.
CONCLUSIONS AND RELEVANCE
In this meta-analysis, proton therapy was associated with reduced OARs doses and toxic effects and improved prognosis compared with photon therapy for esophageal cancer, but caution is warranted. In the future, these findings should be further validated in randomized clinical trials.
Topics: Humans; Proton Therapy; Protons; Pericardial Effusion; Radiation Pneumonitis; Esophageal Neoplasms
PubMed: 37581887
DOI: 10.1001/jamanetworkopen.2023.28136 -
Inorganic Chemistry Sep 2023Nitrogenase is a fascinating enzyme in biology that reduces dinitrogen from air to ammonia through stepwise reduction and protonation. Despite it being studied in detail...
Nitrogenase is a fascinating enzyme in biology that reduces dinitrogen from air to ammonia through stepwise reduction and protonation. Despite it being studied in detail by experimental and computational groups, there are still many unknown factors in the catalytic cycle of nitrogenase, especially related to the addition of protons and electrons and their order. A recent biomimetic study characterized a potential dinitrogen-bridged diiron cluster as a synthetic model of nitrogenase. Using strong acid and reductants, the dinitrogen was converted into ammonia molecules, but details of the mechanism remains unknown. In particular, it was unclear from the experimental studies whether the proton and electron transfer steps are sequential or alternating. Moreover, the work failed to establish what the function of the diiron core is and whether it split into mononuclear iron fragments during the reaction. To understand the structure and reactivity of the biomimetic dinitrogen-bridged diiron complex [(PFeH)(μ-N)] with triphenylphosphine ligands, we performed a density functional theory study. Our computational methods were validated against experimental crystal structure coordinates, Mössbauer parameters, and vibrational frequencies and show excellent agreement. Subsequently, we investigated the alternating and consecutive addition of electrons and protons to the system. The calculations identify a number of possible reaction channels, namely, same-site protonation, alternating protonation, and complex dissociation into mononuclear iron centers. The calculations show that the overall mechanism is not a pure sequential set of electron and proton transfers but a mixture of alternating and consecutive steps. In particular, the first reaction steps will start with double proton transfer followed by an electron transfer, while thereafter, there is another proton transfer and a second electron transfer to give a complex whereby ammonia can split off with a low energetic barrier. The second channel starts with alternating protonation of the two nitrogen atoms, whereafter the initial double proton transfer, electrons and protons are added sequentially to form a hydrazine-bound complex. The latter split off ammonia spontaneously after further protonation. The various reaction channels are analyzed with valence bond and orbital diagrams. We anticipate the nitrogenase enzyme to operate with mixed alternating and consecutive protonation and electron transfer steps.
Topics: Ammonia; Protons; Iron; Nitrogen; Nitrogenase
PubMed: 37650683
DOI: 10.1021/acs.inorgchem.3c02089 -
Annual Review of Physiology Feb 2024A handful of biological proton-selective ion channels exist. Some open at positive or negative membrane potentials, others open at low or high pH, and some are light... (Review)
Review
A handful of biological proton-selective ion channels exist. Some open at positive or negative membrane potentials, others open at low or high pH, and some are light activated. This review focuses on common features that result from the unique properties of protons. Proton conduction through water or proteins differs qualitatively from that of all other ions. Extraordinary proton selectivity is needed to ensure that protons permeate and other ions do not. Proton selectivity arises from a proton pathway comprising a hydrogen-bonded chain that typically includes at least one titratable amino acid side chain. The enormously diverse functions of proton channels in disparate regions of the phylogenetic tree can be summarized by considering the chemical and electrical consequences of proton flux across membranes. This review discusses examples of cells in which proton efflux serves to increase pH, decrease pH, control the membrane potential, generate action potentials, or compensate transmembrane movement of electrical charge.
Topics: Humans; Protons; Ion Channel Gating; Hydrogen-Ion Concentration; Phylogeny; Ion Channels
PubMed: 37931166
DOI: 10.1146/annurev-physiol-042222-023242 -
ELife Mar 2024The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation...
The vesicular monoamine transporter 2 (VMAT2) is a proton-dependent antiporter responsible for loading monoamine neurotransmitters into synaptic vesicles. Dysregulation of VMAT2 can lead to several neuropsychiatric disorders including Parkinson's disease and schizophrenia. Furthermore, drugs such as amphetamine and MDMA are known to act on VMAT2, exemplifying its role in the mechanisms of actions for drugs of abuse. Despite VMAT2's importance, there remains a critical lack of mechanistic understanding, largely driven by a lack of structural information. Here, we report a 3.1 Å resolution cryo-electron microscopy (cryo-EM) structure of VMAT2 complexed with tetrabenazine (TBZ), a non-competitive inhibitor used in the treatment of Huntington's chorea. We find TBZ interacts with residues in a central binding site, locking VMAT2 in an occluded conformation and providing a mechanistic basis for non-competitive inhibition. We further identify residues critical for cytosolic and lumenal gating, including a cluster of hydrophobic residues which are involved in a lumenal gating strategy. Our structure also highlights three distinct polar networks that may determine VMAT2 conformational dynamics and play a role in proton transduction. The structure elucidates mechanisms of VMAT2 inhibition and transport, providing insights into VMAT2 architecture, function, and the design of small-molecule therapeutics.
Topics: Humans; Tetrabenazine; Vesicular Monoamine Transport Proteins; Protons; Cryoelectron Microscopy; Huntington Disease
PubMed: 38517752
DOI: 10.7554/eLife.91973