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1.
Adv Biol (Weinh) ; : e2400083, 2024 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-38717792

RESUMO

The regulation of complex energy metabolism is intricately linked to cellular energy demands. Caloric restriction (CR) plays a pivotal role in modulating the expression of genes associated with key metabolic pathways, including glycolysis, the tricarboxylic acid (TCA) cycle, and the glyoxylate cycle. In this study, the chronological lifespan (CLS) of 35 viable single-gene deletion mutants under both non-restricted and CR conditions, focusing on genes related to these metabolic pathways is evaluated. CR is found to increase CLS predominantly in mutants associated with the glycolysis and TCA cycle. However, this beneficial effect of CR is not observed in mutants of the glyoxylate cycle, particularly those lacking genes for critical enzymes like isocitrate lyase 1 (icl1Δ) and malate synthase 1 (mls1Δ). This analysis revealed an increase in isocitrate lyase activity, a key enzyme of the glyoxylate cycle, under CR, unlike the activity of isocitrate dehydrogenase, which remains unchanged and is specific to the TCA cycle. Interestingly, rapamycin, a compound known for extending lifespan, does not increase the activity of the glyoxylate cycle enzyme. This suggests that CR affects lifespan through a distinct metabolic mechanism.

2.
Nano Lett ; 24(6): 1882-1890, 2024 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-38198287

RESUMO

Understanding the spatial organization of membrane proteins is crucial for unraveling key principles in cell biology. The reaction-diffusion model is commonly used to understand biochemical patterning; however, applying reaction-diffusion models to subcellular phenomena is challenging because of the difficulty in measuring protein diffusivity and interaction kinetics in the living cell. In this work, we investigated the self-organization of the plasmalemma vesicle-associated protein (PLVAP), which creates regular arrangements of fenestrated ultrastructures, using single-molecule tracking. We demonstrated that the spatial organization of the ultrastructures is associated with a decrease in the association rate by actin destabilization. We also constructed a reaction-diffusion model that accurately generates a hexagonal array with the same 130 nm spacing as the actual scale and informs the stoichiometry of the ultrastructure, which can be discerned only through electron microscopy. Through this study, we integrated single-molecule experiments and reaction-diffusion modeling to surpass the limitations of static imaging tools and proposed emergent properties of the PLVAP ultrastructure.


Assuntos
Proteínas de Transporte , Proteínas de Membrana , Proteínas de Membrana/metabolismo , Difusão , Modelos Biológicos
3.
Nat Commun ; 14(1): 7619, 2023 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-37993434

RESUMO

The biological process of aging is thought to result in part from accumulation of senescent cells in organs. However, the present study identified a subset of fibroblasts and smooth muscle cells which are the major constituents of organ stroma neither proliferative nor senescent in tissues of the elderly, which we termed "mid-old status" cells. Upregulation of pro-inflammatory genes (IL1B and SAA1) and downregulation of anti-inflammatory genes (SLIT2 and CXCL12) were detected in mid-old cells. In the stroma, SAA1 promotes development of the inflammatory microenvironment via upregulation of MMP9, which decreases the stability of epithelial cells present on the basement membrane, decreasing epithelial cell function. Remarkably, the microenvironmental change and the functional decline of mid-old cells could be reversed by a young cell-originated protein, SLIT2. Our data identify functional reversion of mid-old cells as a potential method to prevent or ameliorate aspects of aging-related tissue dysfunction.


Assuntos
Envelhecimento , Senescência Celular , Humanos , Idoso , Senescência Celular/genética , Envelhecimento/genética , Células Epiteliais/fisiologia , Fibroblastos , Miócitos de Músculo Liso
4.
Pharmacol Ther ; 252: 108560, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37952903

RESUMO

Biological responsiveness refers to the capacity of living organisms to adapt to changes in both their internal and external environments through physiological and behavioral mechanisms. One of the prominent aspects of aging is the decline in this responsiveness, which can lead to a deterioration in the processes required for maintenance, survival, and growth. The vital link between physiological responsiveness and the essential life processes lies within the signaling systems. To devise effective strategies for controlling the aging process, a comprehensive reevaluation of this connecting loop is imperative. This review aims to explore the impact of aging on signaling systems responsible for responsiveness and introduce a novel perspective on intervening in the aging process by restoring the compromised responsiveness. These innovative mechanistic approaches for modulating altered responsiveness hold the potential to illuminate the development of action plans aimed at controlling the aging process and treating age-related disorders.


Assuntos
Envelhecimento , Transdução de Sinais , Humanos , Envelhecimento/fisiologia
5.
Cell Rep ; 42(10): 113183, 2023 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-37777962

RESUMO

Recent developments in genome sequencing have expanded the knowledge of genetic factors associated with late-onset Alzheimer's disease (AD). Among them, genetic variant ε4 of the APOE gene (APOE4) confers the greatest disease risk. Dysregulated glucose metabolism is an early pathological feature of AD. Using isogenic ApoE3 and ApoE4 astrocytes derived from human induced pluripotent stem cells, we find that ApoE4 increases glycolytic activity but impairs mitochondrial respiration in astrocytes. Ultrastructural and autophagy flux analyses show that ApoE4-induced cholesterol accumulation impairs lysosome-dependent removal of damaged mitochondria. Acute treatment with cholesterol-depleting agents restores autophagic activity, mitochondrial dynamics, and associated proteomes, and extended treatment rescues mitochondrial respiration in ApoE4 astrocytes. Taken together, our study provides a direct link between ApoE4-induced lysosomal cholesterol accumulation and abnormal oxidative phosphorylation.


Assuntos
Doença de Alzheimer , Células-Tronco Pluripotentes Induzidas , Humanos , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Astrócitos/metabolismo , Fosforilação Oxidativa , Células Cultivadas , Células-Tronco Pluripotentes Induzidas/metabolismo , Apolipoproteína E3/metabolismo , Colesterol/metabolismo , Doença de Alzheimer/metabolismo , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo
6.
Heliyon ; 9(9): e20201, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37809635

RESUMO

Impaired cutaneous wound healing is a major complication in patients with diabetes mellitus (DM), leading to increased amputation and mortality rates in affected patients. Adipose-derived stem cells (ASCs) are widely used seed cells for promoted tissue regeneration to improve wound closure under diabetic conditions. However, ASCs-based therapies remain limited due to difficulties in maintaining cell quality during transplantation. To overcome this problem, extracellular matrix mimetic biomaterials have been developed for use in biomedical engineering field, including tissue engineering and regenerative medicine. Herein, a biosynthesized arginine-glycine-aspartate amino acid residues (RGD motif, known as a cell adhesion motif)-containing elastin-like polypeptides (REPs) improved the efficacy of ASCs in enhancing wound closure and skin elasticity in diabetic wounds by promoting the expression of angiogenic growth factors. Therefore, REPs can be used as potential supplements to stem cell-based therapeutic approach to accelerate diabetic wound repair.

7.
Anal Chem ; 95(43): 15924-15932, 2023 10 31.
Artigo em Inglês | MEDLINE | ID: mdl-37774148

RESUMO

In live cells, the plasma membrane is composed of lipid domains separated by hundreds of nanometers in dynamic equilibrium. Lipid phase separation regulates the trafficking and spatiotemporal organization of membrane molecules that promote signal transduction. However, visualizing domains with adequate spatiotemporal accuracy remains challenging because of their subdiffraction limit size and highly dynamic properties. Here, we present a single lipid-molecular motion analysis pipeline (lipid-MAP) for analyzing the phase heterogeneity of lipid membranes by detecting the instantaneous velocity change of a single lipid molecule using the excellent optical properties of nanoparticles, high spatial localization accuracy of single-molecule localization microscopy, and separation capability of the diffusion state of the hidden Markov model algorithm. Using lipid-MAP, individual lipid molecules were found to be in dynamic equilibrium between two statistically distinguishable phases, leading to the formation of small (∼170 nm), viscous (2.5× more viscous than surrounding areas), and transient domains in live cells. Moreover, our findings provide an understanding of how membrane compositional changes, i.e., cholesterol and phospholipids, affect domain formation. This imaging method can contribute to an improved understanding of spatiotemporal-controlled membrane dynamics at the molecular level.


Assuntos
Fosfolipídeos , Transdução de Sinais , Membrana Celular/metabolismo , Fosfolipídeos/metabolismo , Membranas , Difusão , Bicamadas Lipídicas/metabolismo
8.
Tissue Eng Regen Med ; 20(7): 1133-1143, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37610706

RESUMO

BACKGROUND: Cryopreservation is a crucial method for long-term storage and stable allocation of human pluripotent stem cells (hPSCs), which are increasingly being used in various applications. However, preserving hPSCs in cryogenic conditions is challenging due to reduced recovery rates. METHODS: To address this issue, the Arginine-Glycine-Aspartate (RGD) motif was incorporated into a recombinant elastin-like peptide (REP). Human embryonic stem cells (hESCs) were treated with REP containing RGD motif (RGD-REP) during suspension and cryopreservation, and the survival rate was analyzed. The underlying mechanisms were also investigated. RESULTS: The addition of RGD-REP to the cryopreservation solution improved cell survival and pluripotency marker expression. The improvement was confirmed to be due to the activation of the FAK-AKT cascade by RGD-REP binding to hESC surface interin protein, and consequent inhibition of FoxO3a. The inactivation of FoxO3a reduced the expression of apoptosis-related genes, such as BIM, leading to increased survival of PSCs in a suspension state. CONCLUSION: RGD-REP, as a ligand for integrin protein, improves the survival and maintenance of hPSCs during cryopreservation by activating survival signals via the RGD motif. These results have potential implications for improving the efficiency of stem cell usage in both research and therapeutic applications.


Assuntos
Células-Tronco Embrionárias Humanas , Células-Tronco Pluripotentes , Humanos , Células-Tronco Embrionárias Humanas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Elastina/metabolismo , Criopreservação/métodos , Transdução de Sinais , Oligopeptídeos/farmacologia
9.
J Korean Med Sci ; 38(33): e258, 2023 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-37605497

RESUMO

BACKGROUND: This study aimed to identify the specific T cell co-stimulatory and co-inhibitory factors that play prognostic roles in patients with glioblastoma. Additionally, the unique histone H3 modification enzymes that regulate the expression levels of these specific co-stimulatory and co-inhibitory factors were investigated. METHODS: The medical records of 84 patients newly diagnosed with glioblastoma at our institution from January 2006 to December 2020 were retrospectively reviewed. Immunohistochemical (IHC) staining for T cell co-stimulatory factors (CD27, CD28, CD137, OX40, and ICOS), T cell co-inhibitory factors (CTLA4, PD1, PD-L1, TIM3, and CD200R), and histone H3 lysine modification enzymes (MLL4, RIZ, EZH1, NSD2, KDM5c, JMJD1a, UTX, and JMJD5) was performed on archived paraffin-embedded tissues obtained by biopsy or resection. Quantitative real time-polymerase chain reaction (qRT-PCR) was performed for specific factors, which demonstrated causal relationships, in order to validate the findings of the IHC examinations. RESULTS: The mean follow-up duration was 27.5 months (range, 4.1-43.5 months). During this period, 76 patients (90.5%) died, and the mean OS was 19.4 months (95% confidence interval, 16.3-20.9 months). Linear positive correlations were observed between the expression levels of CD28 and JMJD1a (R2 linear = 0.982) and those of CD137 and UTX (R2 linear = 1.528). Alternatively, significant negative correlations were observed between the expression levels of CTLA4 and RIZ (R2 linear = -1.746) and those of PD-L1 and EZH1 (R2 linear = -2.118); these relationships were confirmed by qRT-PCR. In the multivariate analysis, increased expression levels of CD28 (P = 0.042), and CD137 (P = 0.009), and decreased expression levels of CTLA4 (P = 0.003), PD-L1 (P = 0.020), and EZH1 (P = 0.040) were significantly associated with longer survival. CONCLUSION: These findings suggest that the expression of certain T cell co-stimulatory factors, such as CD28 and CD 137, and co-inhibitory factors, such as CTLA4 and PD-L1 are associated with prognosis of glioblastoma patients.


Assuntos
Glioblastoma , Histonas , Humanos , Antígeno CTLA-4/genética , Antígeno B7-H1 , Lisina , Prognóstico , Antígenos CD28 , Glioblastoma/diagnóstico , Glioblastoma/genética , Epigênese Genética , Estudos Retrospectivos , Linfócitos T
10.
J Invest Dermatol ; 143(11): 2295-2310.e17, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37211200

RESUMO

An effective healing response is critical to healthy aging. In particular, energy homeostasis has become increasingly recognized as a factor in effective skin regeneration. ANT2 is a mediator of adenosine triphosphate import into mitochondria for energy homeostasis. Although energy homeostasis and mitochondrial integrity are critical for wound healing, the role played by ANT2 in the repair process had not been elucidated to date. In our study, we found that ANT2 expression decreased in aged skin and cellular senescence. Interestingly, overexpression of ANT2 in aged mouse skin accelerated the healing of full-thickness cutaneous wounds. In addition, upregulation of ANT2 in replicative senescent human diploid dermal fibroblasts induced their proliferation and migration, which are critical processes in wound healing. Regarding energy homeostasis, ANT2 overexpression increased the adenosine triphosphate production rate by activating glycolysis and induced mitophagy. Notably, ANT2-mediated upregulation of HSPA6 in aged human diploid dermal fibroblasts downregulated proinflammatory genes that mediate cellular senescence and mitochondrial damage. This study shows a previously uncharacterized physiological role of ANT2 in skin wound healing by regulating cell proliferation, energy homeostasis, and inflammation. Thus, our study links energy metabolism to skin homeostasis and reports, to the best of our knowledge, a previously unreported genetic factor that enhances wound healing in an aging model.

11.
Sensors (Basel) ; 23(6)2023 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-36992066

RESUMO

Lightning strikes can cause significant damage to critical infrastructure and pose a serious threat to public safety. To ensure the safety of facilities and investigate the causes of lightning accidents, we propose a cost-effective design method for a lightning current measuring instrument that uses a Rogowski coil and dual signal conditioning circuits to detect a wide range of lightning currents, ranging from hundreds of A to hundreds of kA. To implement the proposed lightning current measuring instrument, we design signal conditioning circuits and software capable of detecting and analyzing lightning currents from ±500 A to ±100 kA. By employing dual signal conditioning circuits, it offers the advantage of detecting a wide range of lightning currents compared to existing lightning current measuring instruments. The proposed instrument has the following features: First, the peak current, polarity, T1 (front time), T2 (time to half value), and Q (amount of energy of the lightning current) can be analyzed and measured with a fast sampling time of 380 ns. Second, it can distinguish whether a lightning current is induced or direct. Third, a built-in SD card is provided to save the detected lightning data. Finally, it provides Ethernet communication capability for remote monitoring. The performance of the proposed instrument is evaluated and validated by applying induced and direct lightning using a lightning current generator.

12.
Elife ; 112022 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-36214665

RESUMO

Lack of oxygen (hypoxia and anoxia) is detrimental to cell function and survival and underlies many disease conditions. Hence, metazoans have evolved mechanisms to adapt to low oxygen. One such mechanism, metabolic suppression, decreases the cellular demand for oxygen by downregulating ATP-demanding processes. However, the molecular mechanisms underlying this adaptation are poorly understood. Here, we report on the role of ndrg1a in hypoxia adaptation of the anoxia-tolerant zebrafish embryo. ndrg1a is expressed in the kidney and ionocytes, cell types that use large amounts of ATP to maintain ion homeostasis. ndrg1a mutants are viable and develop normally when raised under normal oxygen. However, their survival and kidney function is reduced relative to WT embryos following exposure to prolonged anoxia. We further demonstrate that Ndrg1a binds to the energy-demanding sodium-potassium ATPase (NKA) pump under anoxia and is required for its degradation, which may preserve ATP in the kidney and ionocytes and contribute to energy homeostasis. Lastly, we show that sodium azide treatment, which increases lactate levels under normoxia, is sufficient to trigger NKA degradation in an Ndrg1a-dependent manner. These findings support a model whereby Ndrg1a is essential for hypoxia adaptation and functions downstream of lactate signaling to induce NKA degradation, a process known to conserve cellular energy.


Assuntos
Hipóxia , Peixe-Zebra , Trifosfato de Adenosina/metabolismo , Animais , Hipóxia/genética , Lactatos , Oxigênio/metabolismo , Potássio/metabolismo , Sódio/metabolismo , Azida Sódica/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Peixe-Zebra/metabolismo
13.
Cell Death Dis ; 13(9): 760, 2022 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-36055997

RESUMO

Selective removal of senescent cells, or senolytic therapy, has been proposed to be a potent strategy for overcoming age-related diseases and even for reversing aging. We found that nintedanib, a tyrosine kinase inhibitor, selectively induced the death of primary human dermal fibroblasts undergoing RS. Similar to ABT263, a well-known senolytic agent, nintedanib triggered intrinsic apoptosis in senescent cells. Additionally, at the concentration producing the senolytic effect, nintedanib arrested the cell cycle of nonsenescent cells in the G1 phase without inducing cytotoxicity. Interestingly, the mechanism by which nintedanib activated caspase-9 in the intrinsic apoptotic pathway differed from that of ABT263 apoptosis induction; specifically, nintedanib did not decrease the levels of Bcl-2 family proteins in senescent cells. Moreover, nintedanib suppressed the activation of the JAK2/STAT3 pathway, which caused the drug-induced death of senescent cells. STAT3 knockdown in senescent cells induced caspase activation. Moreover, nintedanib reduced the number of senescence-associated ß-galactosidase-positive senescent cells in parallel with a reduction in STAT3 phosphorylation and ameliorated collagen deposition in a mouse model of bleomycin-induced lung fibrosis. Consistently, nintedanib exhibited a senolytic effect through bleomycin-induced senescence of human pulmonary fibroblasts. Overall, we found that nintedanib can be used as a new senolytic agent and that inhibiting STAT3 may be an approach for inducing the selective death of senescent cells. Our findings pave the way for expanding the senolytic toolkit for use in various aging statuses and age-related diseases.


Assuntos
Indóis , Senoterapia , Animais , Bleomicina/farmacologia , Senescência Celular , Fibroblastos/metabolismo , Humanos , Indóis/metabolismo , Indóis/farmacologia , Camundongos , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
14.
Commun Biol ; 5(1): 702, 2022 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-35835838

RESUMO

The multifaceted nature of senescent cell cycle arrest necessitates the targeting of multiple factors arresting or promoting the cell cycle. We report that co-inhibition of ATM and ROCK by KU-60019 and Y-27632, respectively, synergistically increases the proliferation of human diploid fibroblasts undergoing replicative senescence through activation of the transcription factors E2F1 and FOXM1. Time-course transcriptome analysis identified FOXM1 and E2F1 as crucial factors promoting proliferation. Co-inhibition of the kinases ATM and ROCK first promotes the G2/M transition via FOXM1 activation, leading to accumulation of cells undergoing the G1/S transition via E2F1 activation. The combination of both inhibitors increased this effect more significantly than either inhibitor alone, suggesting synergism. Our results demonstrate a FOXM1- and E2F1-mediated molecular pathway enhancing cell cycle progression in cells with proliferative potential under replicative senescence conditions, and treatment with the inhibitors can be tested for senomorphic effect in vivo.


Assuntos
Senescência Celular , Fator de Transcrição E2F1 , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células , Fator de Transcrição E2F1/genética , Fator de Transcrição E2F1/metabolismo , Fator de Transcrição E2F1/farmacologia , Proteína Forkhead Box M1/genética , Proteína Forkhead Box M1/metabolismo , Proteína Forkhead Box M1/farmacologia , Humanos
15.
Sci Rep ; 12(1): 7310, 2022 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-35508617

RESUMO

NR2E3 is an orphan nuclear receptor whose loss-of-function causes abnormal retinal photoreceptor development and degeneration. However, despite that many nuclear receptors are regulated by binding of small molecule ligands, biological small molecule ligands regulating NR2E3 have not been identified. Identification of an endogenous NR2E3 ligand might reveal a previously unrecognized component contributing to retinal development and maintenance. Here we report that biliverdin, a conserved green pigment from heme catabolism, regulates NR2E3 and is necessary for zebrafish retinal photoreceptor development. Biliverdin from retinal extracts specifically bound to NR2E3's ligand-binding domain and induced NR2E3-dependent reporter gene expression. Inhibition of biliverdin synthesis decreased photoreceptor cell populations in zebrafish larvae, and this phenotype was alleviated by exogenously supplied biliverdin. Thus, biliverdin is an endogenous small molecule ligand for NR2E3 and a component necessary for the proper development of photoreceptor cells. This result suggests a possible role of heme metabolism in the regulation of retinal photoreceptor cell development.


Assuntos
Degeneração Retiniana , Peixe-Zebra , Animais , Biliverdina , Heme , Ligantes , Receptores Nucleares Órfãos/genética , Células Fotorreceptoras/metabolismo , Células Fotorreceptoras de Vertebrados/metabolismo , Receptores Citoplasmáticos e Nucleares , Degeneração Retiniana/metabolismo , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética
16.
Biochemistry ; 61(7): 575-582, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35285625

RESUMO

Human phosphoribosylaminoimidazole carboxylase phosphoribosylaminoimdiazole succinocarboxamide synthetase (PAICS) is a dual activity enzyme catalyzing two consecutive reactions in de novo purine nucleotide synthesis. Crystallographic structures of recombinant human PAICS suggested the channeling of 4-carboxy-5-aminoimidazole-1-ribose-5'-phosphate (CAIR) between two active sites of PAICS, while a prior work of an avian PAICS suggested otherwise. Here, we present time-course mass spectrometric data supporting the channeling of CAIR between domains of recombinant human PAICS. Time-course mass spectral analysis showed that CAIR added to the bulk solution (CAIRbulk) is decarboxylated and re-carboxylated before the accumulation of succinyl-5-aminoimidazole-4-carboxamide-1-ribose-5'-phosphate (SAICAR). An experiment with 13C-bicarbonate showed that SAICAR production was proportional to re-carboxylated CAIR instead of total CAIR or CAIRbulk. This result indicates that the SAICAR synthase domain selectively uses enzyme-made CAIR over CAIRbulk, which is consistent with the channeling model. This channeling between PAICS domains may be a part of a larger channeling process in de novo purine nucleotide synthesis.


Assuntos
Bicarbonatos , Carboxiliases , Peptídeo Sintases , Carboxiliases/química , Domínio Catalítico , Humanos , Enzimas Multifuncionais/química , Peptídeo Sintases/química
17.
Cell Chem Biol ; 29(3): 476-489.e6, 2022 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-34529934

RESUMO

Topoisomerase II (topo II) is essential for disentangling newly replicated chromosomes. DNA unlinking involves the physical passage of one duplex through another and depends on the transient formation of double-stranded DNA breaks, a step exploited by frontline chemotherapeutics to kill cancer cells. Although anti-topo II drugs are efficacious, they also elicit cytotoxic side effects in normal cells; insights into how topo II is regulated in different cellular contexts is essential to improve their targeted use. Using chemical fractionation and mass spectrometry, we have discovered that topo II is subject to metabolic control through the TCA cycle. We show that TCA metabolites stimulate topo II activity in vitro and that levels of TCA flux modulate cellular sensitivity to anti-topo II drugs in vivo. Our work reveals an unanticipated connection between the control of DNA topology and cellular metabolism, a finding with ramifications for the clinical use of anti-topo II therapies.


Assuntos
Antineoplásicos , Inibidores da Topoisomerase II , Antineoplásicos/farmacologia , DNA/metabolismo , DNA Topoisomerases Tipo II/metabolismo , Inibidores da Topoisomerase II/farmacologia
18.
Cancer Res Treat ; 54(3): 690-708, 2022 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34793663

RESUMO

PURPOSE: This study aimed to investigate the methylation status of major histone modification sites in primary central nervous system lymphoma (PCNSL) samples and examine their prognostic roles in patients with PCNSL. MATERIALS AND METHODS: Between 2007 and 2020, 87 patients were histopathologically diagnosed with PCNSL. We performed immunohistochemical staining of the formalin-fixed paraffin-embedded samples of PCNSL for major histone modification sites, such as H3K4, H3K9, H3K27, H3K14, and H3K36. After detection of meaningful methylation sites, we examined histone modification enzymes that induce methylation or demethylation at each site using immunohistochemical staining. The meaningful immunoreactivity was validated by western blotting using fresh tissue of PCNSL. RESULTS: More frequent recurrences were found in hypomethylation of H3K4me3 (p=0.004) and hypermethylation of H3K27me2 (p<0.001) and H3K27me3 (p=0.002). These factors were also statistically related to short PFS and overall survival in the univariate and multivariate analyses. Next, histone modification enzymes inducing the demethylation of H3K4 (lysine-specific demethylase-1/2 and Jumonji AT-rich interactive domain [JARID] 1A-D]) and methylation of H3K27 (enhancer of zeste homolog [EZH]-1/2) were immu- nohistochemically stained. Among them, the immunoreactivity of JARID1A inversely associated with the methylation status of H3K4me3 (R2=-1.431), and immunoreactivity of EZH2 was directly associated with the methylation status of H3K27me2 (R2=0.667) and H3K27me3 (R2=0.604). These results were validated by western blotting in fresh PCNSL samples. CONCLUSION: Our study suggests that hypomethylation of H3K4me3 and hypermethylation of H3K27me2 and H3K27me3 could be associated with poor outcomes in patients with PCNSL and that these relationships are modified by JARID1A and EZH2.


Assuntos
Histonas , Linfoma , Biomarcadores , Sistema Nervoso Central/metabolismo , Metilação de DNA , Epigênese Genética , Histonas/genética , Histonas/metabolismo , Humanos , Linfoma/diagnóstico , Linfoma/genética , Lisina/metabolismo , Prognóstico
19.
FASEB J ; 35(10): e21894, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34460995

RESUMO

Neuromyelitis optica (NMO) is an autoimmune disease that primarily targets astrocytes. Autoantibodies (NMO-IgG) against the water channel protein, aquaporin 4 (AQP4), are a serologic marker in NMO patients, and they are known to be responsible for the pathophysiology of the disease. In the brain, AQP4 is mainly expressed in astrocytes, especially at the end-feet, where they form the blood-brain barrier. Following the interaction between NMO-IgG and AQP4 in astrocytes, rapid AQP4 endocytosis initiates pathogenesis. However, the cellular and molecular mechanisms of astrocyte destruction by autoantibodies remain largely elusive. We established an in vitro human astrocyte model system using induced pluripotent stem cells (iPSCs) technology in combination with NMO patient-derived serum and IgG to elucidate the cellular and functional changes caused by NMO-IgG. Herein, we observed that NMO-IgG induces structural alterations in mitochondria and their association with the endoplasmic reticulum (ER) and lysosomes at the ultrastructural level, which potentially leads to impaired mitochondrial functions and dynamics. Indeed, human astrocytes display impaired mitochondrial bioenergetics and autophagy activity in the presence of NMO-IgG. We further demonstrated NMO-IgG-driven ER membrane deformation into a multilamellar structure in human astrocytes. Together, we show that NMO-IgG rearranges cellular organelles and alter their functions and that our in vitro system using human iPSCs offers previously unavailable experimental opportunities to study the pathophysiological mechanisms of NMO in human astrocytes or conduct large-scale screening for potential therapeutic compounds targeting astrocytic abnormalities in patients with NMO.


Assuntos
Astrócitos/imunologia , Autoanticorpos/imunologia , Retículo Endoplasmático/imunologia , Imunoglobulina G/imunologia , Células-Tronco Pluripotentes Induzidas/imunologia , Mitocôndrias/imunologia , Neuromielite Óptica/imunologia , Aquaporina 4/imunologia , Humanos
20.
Exp Mol Med ; 53(6): 1092-1108, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-34188179

RESUMO

Senescent cells exhibit a reduced response to intrinsic and extrinsic stimuli. This diminished reaction may be explained by the disrupted transmission of nuclear signals. However, this hypothesis requires more evidence before it can be accepted as a mechanism of cellular senescence. A proteomic analysis of the cytoplasmic and nuclear fractions obtained from young and senescent cells revealed disruption of nucleocytoplasmic trafficking (NCT) as an essential feature of replicative senescence (RS) at the global level. Blocking NCT either chemically or genetically induced the acquisition of an RS-like senescence phenotype, named nuclear barrier-induced senescence (NBIS). A transcriptome analysis revealed that, among various types of cellular senescence, NBIS exhibited a gene expression pattern most similar to that of RS. Core proteomic and transcriptomic patterns common to both RS and NBIS included upregulation of the endocytosis-lysosome network and downregulation of NCT in senescent cells, patterns also observed in an aging yeast model. These results imply coordinated aging-dependent reduction in the transmission of extrinsic signals to the nucleus and in the nucleus-to-cytoplasm supply of proteins/RNAs. We further showed that the aging-associated decrease in Sp1 transcription factor expression was critical for the downregulation of NCT. Our results suggest that NBIS is a modality of cellular senescence that may represent the nature of physiological aging in eukaryotes.


Assuntos
Senescência Celular , Proteômica , Núcleo Celular/metabolismo , Senescência Celular/genética , Regulação para Baixo
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