Microcephaly Gene Mcph1 Deficiency Induces p19ARF-Dependent Cell Cycle Arrest and Senescence.

MCPH1 has been identified as the causal gene for primary microcephaly type 1, a neurodevelopmental disorder characterized by reduced brain size and delayed growth. As a multifunction protein, MCPH1 has been reported to repress the expression of TERT and interact with transcriptional regulator E2F1. However, it remains unclear whether MCPH1 regulates brain development through its transcriptional regulation function. This study showed that the knockout of Mcph1 in mice leads to delayed growth as early as the embryo stage E11.5. Transcriptome analysis (RNA-seq) revealed that the deletion of Mcph1 resulted in changes in the expression levels of a limited number of genes. Although the expression of some of E2F1 targets, such as Satb2 and Cdkn1c, was affected, the differentially expressed genes (DEGs) were not significantly enriched as E2F1 target genes. Further investigations showed that primary and immortalized Mcph1 knockout mouse embryonic fibroblasts (MEFs) exhibited cell cycle arrest and cellular senescence phenotype. Interestingly, the upregulation of p19ARF was detected in Mcph1 knockout MEFs, and silencing p19Arf restored the cell cycle and growth arrest to wild-type levels. Our findings suggested it is unlikely that MCPH1 regulates neurodevelopment through E2F1-mediated transcriptional regulation, and p19ARF-dependent cell cycle arrest and cellular senescence may contribute to the developmental abnormalities observed in primary microcephaly.

Ab initio determination of melting and sound velocity of neon up to the deep interior of the Earth.

The thermophysical properties and elemental abundances of the noble gases in terrestrial materials can provide unique insights into the Earth's evolution and mantle dynamics. Here, we perform extensive ab initio molecular dynamics simulations to determine the melting temperature and sound velocity of neon up to 370 GPa and 7500 K to constrain its physical state and storage capacity, together with to reveal its implications for the deep interior of the Earth. It is found that solid neon can exist stably under the lower mantle and inner core conditions, and the abnormal melting of neon is not observed under the entire temperature (T) and pressure (P) region inside the Earth owing to its peculiar electronic structure, which is substantially distinct from other heavier noble gases. An inspection of the reduction for sound velocity along the Earth's geotherm evidences that neon can be used as a light element to account for the low-velocity anomaly and density deficit in the deep Earth. A comparison of the pair distribution functions and mean square displacements of MgSiO3-Ne and Fe-Ne alloys further reveals that MgSiO3 has a larger neon storage capacity than the liquid iron under the deep Earth condition, indicating that the lower mantle may be a natural deep noble gas storage reservoir. Our results provide valuable information for studying the fundamental behavior and phase transition of neon in a higher T-P regime, and further enhance our understanding for the interior structure and evolution processes inside the Earth.

Repopulated microglia after pharmacological depletion decrease dendritic spine density in adult mouse brain.

Microglia are innate immune cells in the brain and show exceptional heterogeneity. They are key players in brain physiological development regulating synaptic plasticity and shaping neuronal networks. In pathological disease states, microglia-induced synaptic pruning mediates synaptic loss and targeting microglia was proposed as a promising therapeutic strategy. However, the effect of microglia depletion and subsequent repopulation on dendritic spine density and neuronal function in the adult brain is largely unknown. In this study, we investigated whether pharmacological microglia depletion affects dendritic spine density after long-term permanent microglia depletion and after short-term microglia depletion with subsequent repopulation. Long-term microglia depletion using colony-stimulating-factor-1 receptor (CSF1-R) inhibitor PLX5622 resulted in increased overall spine density, especially of mushroom spines, and increased excitatory postsynaptic current amplitudes. Short-term PLX5622 treatment with subsequent repopulation of microglia had an opposite effect resulting in activated microglia with increased synaptic phagocytosis and consequently decreased spine density and reduced excitatory neurotransmission, while Barnes maze and elevated plus maze testing was unaffected. Moreover, RNA sequencing data of isolated repopulated microglia showed an activated and proinflammatory phenotype. Long-term microglia depletion might be a promising therapeutic strategy in neurological diseases with pathological microglial activation, synaptic pruning, and synapse loss. However, repopulation after depletion induces activated microglia and results in a decrease of dendritic spines possibly limiting the therapeutic application of microglia depletion. Instead, persistent modulation of pathological microglia activity might be beneficial in controlling synaptic damage.

PARP1 UFMylation ensures the stability of stalled replication forks.

The S-phase checkpoint involving CHK1 is essential for fork stability in response to fork stalling. PARP1 acts as a sensor of replication stress and is required for CHK1 activation. However, it is unclear how the activity of PARP1 is regulated. Here, we found that UFMylation is required for the efficient activation of CHK1 by UFMylating PARP1 at K548 during replication stress. Inactivation of UFL1, the E3 enzyme essential for UFMylation, delayed CHK1 activation and inhibits nascent DNA degradation during replication blockage as seen in PARP1-deficient cells. An in vitro study indicated that PARP1 is UFMylated at K548, which enhances its catalytic activity. Correspondingly, a PARP1 UFMylation-deficient mutant (K548R) and pathogenic mutant (F553L) compromised CHK1 activation, the restart of stalled replication forks following replication blockage, and chromosome stability. Defective PARP1 UFMylation also resulted in excessive nascent DNA degradation at stalled replication forks. Finally, we observed that PARP1 UFMylation-deficient knock-in mice exhibited increased sensitivity to replication stress caused by anticancer treatments. Thus, we demonstrate that PARP1 UFMylation promotes CHK1 activation and replication fork stability during replication stress, thus safeguarding genome integrity.

JADE1 is dispensable for the brain development in mice.

In mammalian brain development, WNT signaling balances proliferation and differentiation of neural progenitor cells, and is essential for the maintenance of regular brain development. JADE1 is a candidate transcription co-factor essential for DNA replication, cell division, and cell cycle regulation. In 293T cells, JADE1 is stabilized by von Hippel-Lindau protein pVHL, promotes the ß-catenin ubiquitination and thus blunts canonical WNT signaling. Furthermore, JADE1 inhibits ß-catenin-induced ectopic axis formation in Xenopus embryos. However, JADE1's role in mammalian brain development remains unknown. Here, we generated a new Jade1 knockout mouse line using CRISPR-Cas9 technology. We found that JADE1 null resulted in decreased survival rate, reduced body weight and brain weight in mice. However, histological analysis revealed a normal brain development. Furthermore, Jade1 null neural progenitor cells proliferated normally in vivo and in vitro. RNA-seq analysis further showed that JADE1 loss did not affect the cerebral cortex gene expression. Our findings indicate that JADE1 is dispensable for developing the cerebral cortex in mice.

Janus 2H-MXTe (M = Zr, Hf; X = S, Se) monolayers with outstanding thermoelectric properties and low lattice thermal conductivities.

Two-dimensional (2D) materials have been one of the most popular objects in the research field of thermoelectric (TE) materials and have attracted substantial attention in recent years. Inspired by the synthesized 2H-MoSSe and numerous theoretical studies, we systematically investigated the electronic, thermal, and TE properties of Janus 2H-MXTe (M = Zr and Hf; X = S and Se) monolayers by using first-principles calculations. The phonon dispersion curves and AIMD simulations confirm the thermodynamic stabilities. Moreover, Janus 2H-MXTe were evaluated as indirect band-gap semiconductors with band gaps ranging from 0.56 to 0.90 eV using the HSE06 + SOC method. To evaluate the TE performance, firstly, we calculated the temperature-dependent carrier relaxation time with acoustic phonon scattering τac, impurity scattering τimp, and polarized scattering τpol. Secondly, the calculation of lattice thermal conductivity (κl) shows that these monolayers possess relatively poor κl with values of 3.4-5.4 W mK-1 at 300 K, which is caused by the low phonon lifetime and group velocity. After computing the electronic transport properties, we found that the n-type doped Janus 2H-MXTe monolayers exhibit a high Seebeck coefficient exceeding 200 µV K-1 at 300 K, resulting in a high TE power factor. Eventually, combining the electrical and thermal conductivities, the optimal dimensionless figure of merit (zT) at 300 K (900 K) can be obtained, which is 0.94 (3.63), 0.51 (2.57), 0.64 (2.72), and 0.50 (1.98) for n-type doping of ZrSeTe, HfSeTe, ZeSTe, and HfSTe monolayers. Particularly, the ZrSeTe monolayer shows the best TE performance with the maximal zT value. These results indicate the excellent application potential of Janus 2H-MXTe (M = Zr and Hf; X = S and Se) monolayers in TE materials.

Two-dimensional Janus pentagonal MSeTe (M = Ni, Pd, Pt): promising water-splitting photocatalysts and optoelectronic materials.

Inspired by the interesting and novel properties exhibited by Janus transition metal dichalcogenides (TMDs) and two-dimensional pentagonal structures, we here investigated the structural stability, mechanical, electronic, photocatalytic, and optical properties for a class of two-dimensional (2D) pentagonal Janus TMDs, namely penta-MSeTe (M = Ni, Pd, Pt) monolayers, by using density functional theory (DFT) combined with Hubbard's correction (U). Our results showed that these monolayers exhibit good structural stability, appropriate band structures for photocatalysts, high visible light absorption, and good photocatalytic applicability. The calculated electronic properties reveal that the penta-MSeTe are semiconductors with a bandgap range of 2.06-2.39 eV, and their band edge positions meet the requirements for water-splitting photocatalysts in various environments (pH = 0-13). We used stress engineering to seek higher solar-to-hydrogen (STH) efficiency in acidic (pH = 0), neutral (pH = 7) and alkaline (pH = 13) environments for penta-MSeTe from 0% to +8% biaxial and uniaxial strains. Our results showed that penta-PdSeTe stretched 8% along the y direction and demonstrates an STH efficiency of up to 29.71% when pH = 0, which breaks the theoretical limit of the conventional photocatalytic model. We also calculated the optical properties and found that they exhibit high absorption (13.11%) in the visible light range and possess a diverse range of hyperbolic regions. Hence, it is anticipated that penta-MSeTe materials hold great promise for applications in photocatalytic water splitting and optoelectronic devices.

The role and advantage of traditional Chinese medicine in the prevention and treatment of COVID-19.

The global coronavirus disease 2019 (COVID-19) pandemic has had a massive impact on global social and economic development and human health. By combining traditional Chinese medicine (TCM) with modern medicine, the Chinese government has protected public health by supporting all phases of COVID-19 prevention and treatment, including community prevention, clinical treatment, control of disease progression, and promotion of recovery. Modern medicine focuses on viruses, while TCM focuses on differential diagnosis of patterns associated with viral infection of the body and recommends the use of TCM decoctions for differential treatment. This differential diagnosis and treatment approach, with its profoundly empirical nature and holistic view, endows TCM with an accessibility advantage and high application value for dealing with COVID-19. Here, we summarize the advantage of and evidence for TCM use in COVID-19 prevention and treatment to draw attention to the scientific value and accessibility advantage of TCM and to promote the use of TCM in response to public health emergencies. Please cite this article as: Huang M, Liu YY, Xiong K, Yang FW, Jin XY, Wang ZQ, Zhang JH, Zhang BL. The role and advantage of traditional Chinese medicine in the prevention and treatment of COVID-19. J Integr Med. 2023; 21(5): 407-412.

Poly(ADP-Ribose) Polymerase-1 Lacking Enzymatic Activity Is Not Compatible with Mouse Development.

Poly(ADP-ribose) polymerase-1 (PARP1) binds DNA lesions to catalyse poly(ADP-ribosyl)ation (PARylation) using NAD+ as a substrate. PARP1 plays multiple roles in cellular activities, including DNA repair, transcription, cell death, and chromatin remodelling. However, whether these functions are governed by the enzymatic activity or scaffolding function of PARP1 remains elusive. In this study, we inactivated in mice the enzymatic activity of PARP1 by truncating its C-terminus that is essential for ART catalysis (PARP1ΔC/ΔC, designated as PARP1-ΔC). The mutation caused embryonic lethality between embryonic day E8.5 and E13.5, in stark contrast to PARP1 complete knockout (PARP1-/-) mice, which are viable. Embryonic stem (ES) cell lines can be derived from PARP1ΔC/ΔC blastocysts, and these mutant ES cells can differentiate into all three germ layers, yet, with a high degree of cystic structures, indicating defects in epithelial cells. Intriguingly, PARP1-ΔC protein is expressed at very low levels compared to its full-length counterpart, suggesting a selective advantage for cell survival. Noticeably, PARP2 is particularly elevated and permanently present at the chromatin in PARP1-ΔC cells, indicating an engagement of PARP2 by non-enzymatic PARP1 protein at the chromatin. Surprisingly, the introduction of PARP1-ΔC mutation in adult mice did not impair their viability; yet, these mutant mice are hypersensitive to alkylating agents, similar to PARP1-/- mutant mice. Our study demonstrates that the catalytically inactive mutant of PARP1 causes the developmental block, plausibly involving PARP2 trapping.

The neurological and non-neurological roles of the primary microcephaly-associated protein ASPM.

Primary microcephaly (MCPH), is a neurological disorder characterized by small brain size that results in numerous developmental problems, including intellectual disability, motor and speech delays, and seizures. Hitherto, over 30 MCPH causing genes (MCPHs) have been identified. Among these MCPHs, MCPH5, which encodes abnormal spindle-like microcephaly-associated protein (ASPM), is the most frequently mutated gene. ASPM regulates mitotic events, cell proliferation, replication stress response, DNA repair, and tumorigenesis. Moreover, using a data mining approach, we have confirmed that high levels of expression of ASPM correlate with poor prognosis in several types of tumors. Here, we summarize the neurological and non-neurological functions of ASPM and provide insight into its implications for the diagnosis and treatment of MCPH and cancer.

Simultaneous Nbs1 and p53 inactivation in neural progenitors triggers high-grade gliomas.

AIMS: Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder caused by hypomorphic mutations of NBS1. NBS1 is a member of the MRE11-RAD50-NBS1 (MRN) complex that binds to DNA double-strand breaks and activates the DNA damage response (DDR). Nbs1 inactivation in neural progenitor cells leads to microcephaly and premature death. Interestingly, p53 homozygous deletion rescues the NBS1-deficient phenotype allowing long-term survival. The objective of this work was to determine whether simultaneous inactivation of Nbs1 and p53 in neural progenitors triggered brain tumorigenesis and if so in which category this tumour could be classified. METHODS: We generated a mouse model with simultaneous genetic inactivation of Nbs1 and p53 in embryonic neural stem cells and analysed the arising tumours with in-depth molecular analyses including immunohistochemistry, array comparative genomic hybridisation (aCGH), whole exome-sequencing and RNA-sequencing. RESULTS: NBS1/P53-deficient mice develop high-grade gliomas (HGG) arising in the olfactory bulbs and in the cortex along the rostral migratory stream. In-depth molecular analyses using immunohistochemistry, aCGH, whole exome-sequencing and RNA-sequencing revealed striking similarities to paediatric human HGG with shared features with radiation-induced gliomas (RIGs). CONCLUSIONS: Our findings show that concomitant inactivation of Nbs1 and p53 in mice promotes HGG with RIG features. This model could be useful for preclinical studies to improve the prognosis of these deadly tumours, but it also highlights the singularity of NBS1 among the other DNA damage response proteins in the aetiology of brain tumours.

Microglia mediate neurocognitive deficits by eliminating C1q-tagged synapses in sepsis-associated encephalopathy.

Sepsis-associated encephalopathy (SAE) is a severe and frequent complication of sepsis causing delirium, coma, and long-term cognitive dysfunction. We identified microglia and C1q complement activation in hippocampal autopsy tissue of patients with sepsis and increased C1q-mediated synaptic pruning in a murine polymicrobial sepsis model. Unbiased transcriptomics of hippocampal tissue and isolated microglia derived from septic mice revealed an involvement of the innate immune system, complement activation, and up-regulation of lysosomal pathways during SAE in parallel to neuronal and synaptic damage. Microglial engulfment of C1q-tagged synapses could be prevented by stereotactic intrahippocampal injection of a specific C1q-blocking antibody. Pharmacologically targeting microglia by PLX5622, a CSF1-R inhibitor, reduced C1q levels and the number of C1q-tagged synapses, protected from neuronal damage and synapse loss, and improved neurocognitive outcome. Thus, we identified complement-dependent synaptic pruning by microglia as a crucial pathomechanism for the development of neuronal defects during SAE.

UPF3A is dispensable for nonsense-mediated mRNA decay in mouse pluripotent and somatic cells.

Nonsense-mediated mRNA decay (NMD) is a highly conserved regulatory mechanism of post-transcriptional gene expression in eukaryotic cells. NMD plays essential roles in mRNA quality and quantity control and thus safeguards multiple biological processes including embryonic stem cell differentiation and organogenesis. UPF3A and UPF3B in vertebrate species, originated from a single UPF3 gene in yeast, are key factors in the NMD machinery. Although UPF3B is a well-recognized weak NMD-promoting factor, whether UPF3A functions in promoting or suppressing NMD is under debate. In this study, we generated a Upf3a conditional knockout mouse strain and established multiple lines of embryonic stem cells and somatic cells without UPF3A. Through extensive analysis on the expressions of 33 NMD targets, we found UPF3A neither represses NMD in mouse embryonic stem cells, somatic cells, nor in major organs including the liver, spleen, and thymus. Our study reinforces that UPF3A is dispensable for NMD when UPF3B is present. Furthermore, UPF3A may weakly and selectively promote NMD in certain murine organs.

TRRAP-mediated acetylation on Sp1 regulates adult neurogenesis.

The adult hippocampal neurogenesis plays a vital role in the function of the central nervous system (CNS), including memory consolidation, cognitive flexibility, emotional function, and social behavior. The deficiency of adult neural stem cells (aNSCs) in maintaining the quiescence and entering cell cycle, self-renewal and differentiation capacity is detrimental to the functional integrity of neurons and cognition of the adult brain. Histone acetyltransferase (HAT) and histone deacetylase (HDAC) have been shown to modulate brain functionality and are important for embryonic neurogenesis via regulation of gene transcription. We showed previously that Trrap, an adapter for several HAT complexes, is required for Sp1 transcriptional control of the microtubule dynamics in neuronal cells. Here, we find that Trrap deletion compromises self-renewal and differentiation of aNSCs in mice and in cultures. We find that the acetylation status of lysine residues K16, K19, K703 and K639 all fail to overcome Trrap-deficiency-incurred instability of Sp1, indicating a scaffold role of Trrap. Interestingly, the deacetylation of Sp1 at K639 and K703 greatly increases Sp1 binding to the promoter of target genes, which antagonizes Trrap binding, and thereby elevates Sp1 activity. However, only deacetylated K639 is refractory to Trrap deficiency and corrects the differentiation defects of Trrap-deleted aNSCs. We demonstrate that the acetylation pattern at K639 by HATs dictates the role of Sp1 in the regulation of adult neurogenesis.

Assessment of Mitochondrial Dysfunctions After Sirtuin Inhibition.

Posttranslational modifications are important for protein functions and cellular signaling pathways. The acetylation of lysine residues is catalyzed by histone acetyltransferases (HATs) and removed by histone deacetylases (HDACs), with the latter being grouped into four phylogenetic classes. The class III of the HDAC family, the sirtuins (SIRTs), contributes to gene expression, genomic stability, cell metabolism, and tumorigenesis. Thus, several specific SIRT inhibitors (SIRTi) have been developed to target cancer cell proliferation. Here we provide an overview of methods to study SIRT-dependent cell metabolism and mitochondrial functionality. The chapter describes metabolic flux analysis using Seahorse analyzers, methods for normalization of Seahorse data, flow cytometry and fluorescence microscopy to determine the mitochondrial membrane potential, mitochondrial content per cell and mitochondrial network structures, and Western blot analysis to measure mitochondrial proteins.

DNA damage response in neurodevelopment and neuromaintenance.

The central nervous system is particularly susceptible to DNA repair deficiency, which renders a variety of neurodevelopmental and neurodegenerative disorders in humans. It is generally believed that DNA damage occurs upon repetitive replication and oxidative stress in highly proliferating neuroprogenitor cells (NPs), or due to high rates of metabolism and active neuronal activity in terminally differentiated neurons. DNA double-stranded breaks (DSBs) and single-stranded breaks (SSBs) constitute the most prevalent forms of DNA damage, which can result in neuronal apoptosis if unrepaired. Despite these notions, there are still gaps in our knowledge regarding the mechanism and specificity of DNA damage and repair in the neural development and the homeostasis of neural tissues. Recent studies have identified recurrent DSBs within neuronal long genes in NPs and 'programmed' SSBs in neuronal activity genes. However, the physiological function of these DNA breakages in the nervous system has not been so far explored. In this review, we summarise the recent advances in the field of DNA damage and DNA repair in neural development and neuropathies.

PARP1: Liaison of Chromatin Remodeling and Transcription.

Poly(ADP-ribosyl)ation (PARylation) is a covalent post-translational modification and plays a key role in the immediate response of cells to stress signals. Poly(ADP-ribose) polymerase 1 (PARP1), the founding member of the PARP superfamily, synthesizes long and branched polymers of ADP-ribose (PAR) onto acceptor proteins, thereby modulating their function and their local surrounding. PARP1 is the most prominent of the PARPs and is responsible for the production of about 90% of PAR in the cell. Therefore, PARP1 and PARylation play a pleotropic role in a wide range of cellular processes, such as DNA repair and genomic stability, cell death, chromatin remodeling, inflammatory response and gene transcription. PARP1 has DNA-binding and catalytic activities that are important for DNA repair, yet also modulate chromatin conformation and gene transcription, which can be independent of DNA damage response. PARP1 and PARylation homeostasis have also been implicated in multiple diseases, including inflammation, stroke, diabetes and cancer. Studies of the molecular action and biological function of PARP1 and PARylation provide a basis for the development of pharmaceutic strategies for clinical applications. This review focuses primarily on the role of PARP1 in the regulation of chromatin remodeling and transcriptional activation.

The Central Domain of MCPH1 Controls Development of the Cerebral Cortex and Gonads in Mice.

MCPH1 is the first gene identified to be responsible for the human autosomal recessive disorder primary microcephaly (MCPH). Mutations in the N-terminal and central domains of MCPH1 are strongly associated with microcephaly in human patients. A recent study showed that the central domain of MCPH1, which is mainly encoded by exon 8, interacts with E3 ligase ßTrCP2 and regulates the G2/M transition of the cell cycle. In order to investigate the biological functions of MCPH1's central domain, we constructed a mouse model that lacked the central domain of MCPH1 by deleting its exon 8 (designated as Mcph1-Δe8). Mcph1-Δe8 mice exhibited a reduced brain size and thinner cortex, likely caused by a compromised self-renewal capacity and premature differentiation of Mcph1-Δe8 neuroprogenitors during corticogenesis. Furthermore, Mcph1-Δe8 mice were sterile because of a loss of germ cells in the testis and ovary. The embryonic fibroblasts of Mcph1-Δe8 mice exhibited premature chromosome condensation (PCC). All of these findings indicate that Mcph1-Δe8 mice are reminiscent of MCPH1 complete knockout mice and Mcph1-ΔBR1 mice. Our study demonstrates that the central domain of MCPH1 represses microcephaly, and is essential for gonad development in mammals.

ASPM promotes ATR-CHK1 activation and stabilizes stalled replication forks in response to replication stress.

ASPM is a protein encoded by primary microcephaly 5 (MCPH5) and is responsible for ensuring spindle position during mitosis and the symmetrical division of neural stem cells. We recently reported that ASPM promotes homologous recombination (HR) repair of DNA double strand breaks. However, its potential role in DNA replication and replication stress response remains elusive. Interestingly, we found that ASPM is dispensable for DNA replication under unperturbed conditions. However, ASPM is enriched at stalled replication forks in a RAD17-dependent manner in response to replication stress and promotes RAD9 and TopBP1 loading onto chromatin, facilitating ATR-CHK1 activation. ASPM depletion results in failed fork restart and nuclease MRE11-mediated nascent DNA degradation at the stalled replication fork. The overall consequence is chromosome instability and the sensitization of cancer cells to replication stressors. These data support a role for ASPM in loading RAD17-RAD9/TopBP1 onto chromatin to activate the ATR-CHK1 checkpoint and ultimately ensure genome stability.

Oxidative Glucose Metabolism Promotes Senescence in Vascular Endothelial Cells.

Vascular aging is based on the development of endothelial dysfunction, which is thought to be promoted by senescent cells accumulating in aged tissues and is possibly affected by their environment via inflammatory mediators and oxidative stress. Senescence appears to be closely interlinked with changes in cell metabolism. Here, we describe an upregulation of both glycolytic and oxidative glucose metabolism in replicative senescent endothelial cells compared to young endothelial cells by employing metabolic profiling and glucose flux measurements and by analyzing the expression of key metabolic enzymes. Senescent cells exhibit higher glycolytic activity and lactate production together with an enhanced expression of lactate dehydrogenase A as well as increases in tricarboxylic acid cycle activity and mitochondrial respiration. The latter is likely due to the reduced expression of pyruvate dehydrogenase kinases (PDHKs) in senescent cells, which may lead to increased activity of the pyruvate dehydrogenase complex. Cellular and mitochondrial ATP production were elevated despite signs of mitochondrial dysfunction, such as an increased production of reactive oxygen species and extended mitochondrial mass. A shift from glycolytic to oxidative glucose metabolism induced by pharmacological inhibition of PDHKs in young endothelial cells resulted in premature senescence, suggesting that alterations in cellular glucose metabolism may act as a driving force for senescence in endothelial cells.