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1.
Clin Interv Aging ; 14: 1277-1283, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371933

RESUMO

Age-related macular degeneration (AMD) is the leading cause of irreversible blindness among the elderly. Considering the relatively limited effect of therapy on early AMD, it is important to focus on the pathogenesis of AMD, especially early AMD. Ageing is one of the strongest risk factors for AMD, and analysis of the impact of ageing on AMD development is valuable. Among all the ageing hallmarks, increased DNA damage accumulation is regarded as the beginning of cellular senescence and is related to abnormal expression of inflammatory cytokines, which is called the senescence-associated secretory phenotype (SASP). The exact pathway for DNA damage that triggers senescence-associated hallmarks is poorly understood. Recently, mounting evidence has shown that the cGAS/STING pathway is an important DNA sensor related to proinflammatory factor secretion and is associated with another hallmark of ageing, SASP. Thus, we hypothesized that the cGAS/STING pathway is a vital signalling pathway for early AMD development and that inhibition of STING might be a potential therapeutic strategy for AMD cases.


Assuntos
Dano ao DNA/fisiologia , Inflamação/metabolismo , Inflamação/fisiopatologia , Degeneração Macular/metabolismo , Degeneração Macular/fisiopatologia , Idoso , Senescência Celular/genética , Humanos , Degeneração Macular/patologia , Nucleotidiltransferases/metabolismo , Transdução de Sinais
2.
Cell Mol Life Sci ; 76(22): 4511-4524, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31338556

RESUMO

The nucleolus is a sub-nuclear body known primarily for its role in ribosome biogenesis. Increased number and/or size of nucleoli have historically been used by pathologists as a prognostic indicator of cancerous lesions. This increase in nucleolar number and/or size is classically attributed to the increased need for protein synthesis in cancer cells. However, evidences suggest that the nucleolus plays critical roles in many cellular functions in both normal cell biology and disease pathologies, including cancer. As new functions of the nucleolus are elucidated, there is mounting evidence to support the role of the nucleolus in regulating additional cellular functions, particularly response to cellular stressors, maintenance of genome stability, and DNA damage repair, as well as the regulation of gene expression and biogenesis of several ribonucleoproteins. This review highlights the central role of the nucleolus in carcinogenesis and cancer progression and discusses how cancer cells may become "addicted" to nucleolar functions.


Assuntos
Nucléolo Celular/fisiologia , Neoplasias/patologia , Animais , Carcinogênese/patologia , Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Progressão da Doença , Instabilidade Genômica/fisiologia , Humanos
3.
Nat Rev Cancer ; 19(6): 326-338, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31053804

RESUMO

The cyclin-dependent kinase (CDK)-RB-E2F axis forms the core transcriptional machinery driving cell cycle progression, dictating the timing and fidelity of genome replication and ensuring genetic material is accurately passed through each cell division cycle. The ultimate effectors of this axis are members of a family of eight distinct E2F genes encoding transcriptional activators and repressors. E2F transcriptional activity is tightly regulated throughout the cell cycle via transcriptional and translational regulation, post-translational modifications, protein degradation, binding to cofactors and subcellular localization. Alterations in one or more key components of this axis (CDKs, cyclins, CDK inhibitors and the RB family of proteins) occur in virtually all cancers and result in heightened oncogenic E2F activity, leading to uncontrolled proliferation. In this Review, we discuss the activities of E2F proteins with an emphasis on the newest atypical E2F family members, the specific and redundant functions of E2F proteins, how misexpression of E2F transcriptional targets promotes cancer and both current and developing therapeutic strategies being used to target this oncogenic pathway.


Assuntos
Fatores de Transcrição E2F/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Animais , Apoptose/fisiologia , Proliferação de Células , Instabilidade Cromossômica , Dano ao DNA/fisiologia , Fatores de Transcrição E2F/genética , Humanos , Terapia de Alvo Molecular/métodos , Família Multigênica , Neoplasias/genética , Processamento de Proteína Pós-Traducional , Estabilidade Proteica
4.
Nat Protoc ; 14(5): 1489-1508, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30962605

RESUMO

Non-coding RNA (ncRNA) molecules have been shown to play a variety of cellular roles; however, the contributions of different types of RNA to specific phenomena are often hard to dissect. To study the role of RNA in the assembly of DNA damage response (DDR) foci, we developed the RNase A treatment and reconstitution (RATaR) method, in which cells are mildly permeabilized, incubated with recombinant RNase A and subsequently reconstituted with different RNA species, under conditions of RNase A inactivation and inhibition of endogenous transcription. The block of transcription right after RNase A removal represents a key innovation of RATaR, preventing potential contributions of endogenously neo-synthesized transcripts to the phenotypes studied. A critical aspect of this technique is the balance between sufficient permeabilization of membranes to allow enzyme/RNA access into the cell nucleus and cell viability. Here, we present our protocol for RNA-dependent DDR foci disassembly and reassembly using fluorescent DDR RNAs (DDRNAs) in NIH2/4 cells, an engineered NIH3T3-derived cell line. The use of sequence-specific, fluorescent RNA molecules permits the concomitant determination of their subcellular localization and biological functions. We also outline adaptations of RATaR when implemented in different cell lines exposed to various genotoxic treatments, such as γ-radiation, restriction enzymes and telomere deprotection. In all these cases, the entire procedure can be completed within 2 h without the need for special equipment or uncommon skills. We believe this technique will prove useful for investigating the contribution of RNA to a variety of relevant cellular processes.


Assuntos
Dano ao DNA , Reparo do DNA , RNA não Traduzido , Ribonuclease Pancreático/metabolismo , Animais , Dano ao DNA/genética , Dano ao DNA/fisiologia , Reparo do DNA/genética , Reparo do DNA/fisiologia , Técnicas Genéticas , Células HeLa , Humanos , Camundongos , Células NIH 3T3 , RNA/análise , RNA/genética , RNA/metabolismo , RNA não Traduzido/genética , RNA não Traduzido/fisiologia
5.
Life Sci ; 222: 228-234, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30858123

RESUMO

AIMS: In comparison with a low linear energy transfer (LET) radiation, a high-LET radiation induces more complex DNA damage. This study wonders whether radiation-induced bystander effect (RIBE) is dependent of LET. MATERIALS AND METHODS: Chinese hamster ovary CHO-9 cells and its subline EM-C11 cells (SSB repair deficient) and XR-C1 cells (DSB repair deficient) were irradiated by γ-rays, α-particles, or carbon ions with different LETs of 13, 30 and 70 keV/µm. Cell proliferation, cell death, DNA damage, cell cycle distribution and some protein expressions were measured with the cell counting kit-8 (CCK-8), colony formation, micronuclei (MN), flow cytometry and western blot, respectively. KEY FINDINGS: A series of cell responses were induced by these radiations in a LET-dependent manner, including proliferation inhibition, cell death, MN induction, G2/M phase arrest and the expression of γH2AX protein. These cell injuries were also depended on DNA repair capacity, and XR-C1 cells were the most sensitive to each radiation. Furthermore, when the cells were treated with the conditioned medium (CM) collected from irradiated CHO-9 cells, the MN induction and cell death response in the bystander cells of EM-C11 or XR-C1 increased along with LET of irradiation, and the bystander damage was easier to be induced in EM-C11 and XR-C1 cells than that in CHO-9 cells. SIGNIFICANCE: Both cellular DNA repair capacity and the LET value of radiation could deeply influence damage extents of not only the irradiated cells but also the bystander cells.


Assuntos
Efeito Espectador/efeitos da radiação , Dano ao DNA/efeitos da radiação , Transferência Linear de Energia/efeitos da radiação , Animais , Efeito Espectador/fisiologia , Células CHO , Cricetinae , Cricetulus , Dano ao DNA/fisiologia , Transferência Linear de Energia/fisiologia
6.
Fertil Steril ; 111(4): 699-707.e1, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30826116

RESUMO

OBJECTIVE: To analyze the effect of single- and double-stranded sperm DNA fragmentation (ssSDF and dsSDF) on human embryo kinetics monitored under a time-lapse system. DESIGN: Observational, double blind, prospective cohort study. SETTING: University spin-off and private center. PATIENT(S): One hundred ninety-six embryos from 43 infertile couples were included prospectively. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): SsSDF and dsSDF were analyzed in the same semen sample used for intracytoplasmic sperm injection. Embryo kinetics was then monitored using time-lapse technology, and the timing of each embryo division was obtained. RESULT(S): When comparing embryos obtained from semen samples with low dsSDF and high dsSDF, splitting data using a statistically significant delay in high dsSDF was observed in second polar body extrusion, T4, T8, morula, and starting blastocyst and embryo implantation rates were impaired. Embryo kinetics and implantation rates are not significantly affected when high values of ssSDF are present. Different patterns of delay in embryo kinetics were observed for these different types of DNA damage: dsSDF caused a delay along all stages of embryo development; however, its major effect was observed at the second polar body extrusion and morula stages, coinciding with embryo DNA damage checkpoint activation as described before; ssSDF had its major effect at the pronucleus stage, but embryo kinetics was then restored at all following stages. The results show that dsSDF could be the main type of DNA damage that affects embryo development in intracytoplasmic sperm injection cycles, probably due to motility-based sperm selection in this assisted reproduction procedure. CONCLUSION(S): Double-stranded sperm DNA damage caused a delay in embryo development and impaired implantation, while single-stranded DNA damage did not significantly affect embryo kinetics and implantation.


Assuntos
Dano ao DNA/fisiologia , DNA/genética , Implantação do Embrião/genética , Desenvolvimento Embrionário/genética , Infertilidade/genética , Espermatozoides/metabolismo , Adulto , Método Duplo-Cego , Feminino , Fertilização In Vitro , Humanos , Infertilidade/terapia , Masculino , Gravidez , Taxa de Gravidez , Estudos Prospectivos , Injeções de Esperma Intracitoplásmicas , Imagem com Lapso de Tempo
7.
Environ Sci Pollut Res Int ; 26(13): 13366-13380, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30903470

RESUMO

Eight nuclear abnormalities of genotoxicity and cytotoxicity were studied in peripheral blood erythrocytes of herring (Clupea harengus membras), flounder (Platichthys flesus), and Atlantic cod (Gadus morhua) sampled (2010-2017) from the Polish and the Lithuanian Exclusive Economic Zones (EEZ) in the Baltic Sea. At all study stations, total genotoxicity (∑Gentox) was found to be higher than total cytotoxicity (∑Cytox). A significant time-related decrease in genotoxicity was detected in the Lithuanian EEZ (2015-2017), while in the Polish EEZ (2014-2016), the opposite tendency was revealed. The highest ∑Gentox and ∑Cytox values recorded in fish sampled at the study stations located relatively close to each other clearly indicate an increased environmental genotoxicity and cytotoxicity pressure for fish in these areas. Exceptionally high and high-level genotoxicity risks to herring followed by those to flounder and cod were determined at a higher percentage of the stations studied.


Assuntos
Núcleo Celular/química , Dano ao DNA/genética , Peixes/sangue , Animais , Países Bálticos , Dano ao DNA/fisiologia , Linguado , Gadus morhua , Oceanos e Mares , Polônia
8.
BMC Biol ; 17(1): 23, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30866939

RESUMO

BACKGROUND: Female mammals have a limited reproductive lifespan determined by the size of the primordial follicle pool established perinatally. Over two thirds of fetal oocytes are abolished via programmed cell death during early folliculogenesis. However, the underlying mechanisms governing fetal oocyte attrition remain largely elusive. RESULTS: Here, we demonstrate that glycogen synthase kinase-3 beta (GSK-3ß) is indispensable for fetal oocyte maintenance during meiotic prophase I in mice. In vitro inhibition of GSK-3ß activity or in vivo conditional deletion of Gsk-3ß in the germline led to a dramatic loss of fetal oocytes via apoptosis, which subsequently resulted in a reduced capacity of the primordial follicle pool. Inhibition of GSK-3ß also impeded meiotic progression in fetal oocytes and led to a deficiency in DNA double-strand break (DSB) repair associated with premature upregulation of Tap63, the major genome guardian of the female germline, following GSK-3ß inhibition in fetal ovaries. Mechanistically, we demonstrated that aberrant nuclear translocation of ß-catenin was responsible for the abnormal expression of TAp63 and global fetal oocyte attrition following GSK-3ß inhibition. CONCLUSIONS: In summary, GSK-3ß was essential for sustaining fetal oocyte survival and folliculogenesis via fine-tuning the cytoplasmic-nuclear translocation of ß-catenin, which in turn modulates timely TAp63 expression during meiotic prophase I in mice. Our study provides a perspective on the physiological regulatory role of DNA damage checkpoint signaling in fetal oocyte guardianship and female fertility.


Assuntos
Glicogênio Sintase Quinase 3 beta/metabolismo , Oócitos/metabolismo , Fosfoproteínas/metabolismo , Transdução de Sinais/fisiologia , Transativadores/metabolismo , Animais , Apoptose/fisiologia , Dano ao DNA/fisiologia , Feminino , Glicogênio Sintase Quinase 3 beta/genética , Prófase Meiótica I/fisiologia , Camundongos , Fosfoproteínas/genética , Transativadores/genética , Regulação para Cima , beta Catenina/metabolismo
9.
Nucleic Acids Res ; 47(6): 2766-2777, 2019 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-30773596

RESUMO

Structural variations (SVs) in the human genome originate from different mechanisms related to DNA repair, replication errors, and retrotransposition. Our analyses of 26 927 SVs from the 1000 Genomes Project revealed differential distributions and consequences of SVs of different origin, e.g. deletions from non-allelic homologous recombination (NAHR) are more prone to disrupt chromatin organization while processed pseudogenes can create accessible chromatin. Spontaneous double stranded breaks (DSBs) are the best predictor of enrichment of NAHR deletions in open chromatin. This evidence, along with strong physical interaction of NAHR breakpoints belonging to the same deletion suggests that majority of NAHR deletions are non-meiotic i.e. originate from errors during homology directed repair (HDR) of spontaneous DSBs. In turn, the origin of the spontaneous DSBs is associated with transcription factor binding in accessible chromatin revealing the vulnerability of functional, open chromatin. The chromatin itself is enriched with repeats, particularly fixed Alu elements that provide the homology required to maintain stability via HDR. Through co-localization of fixed Alus and NAHR deletions in open chromatin we hypothesize that old Alu expansion had a stabilizing role on the human genome.


Assuntos
Cromatina/química , Genoma Humano , Variação Estrutural do Genoma/genética , Característica Quantitativa Herdável , Cromatina/metabolismo , Mapeamento Cromossômico , Biologia Computacional , Quebras de DNA de Cadeia Dupla , Dano ao DNA/fisiologia , Reparo do DNA , Recombinação Homóloga , Humanos , Reparo de DNA por Recombinação
10.
Nucleic Acids Res ; 47(6): 2757-2765, 2019 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-30773597

RESUMO

Molecular insight into electronic rearrangements and structural trajectories arising from oxidative damages to DNA backbone is of crucial importance in understanding the effect of ionizing radiation, developing DNA biosensors and designing effective DNA cleaving molecules. Employing a Density Functional Theory based multi-scale Quantum-Mechanical-Molecular-Mechanical (QM/MM) simulation and a suitable partitioning of the Hamiltonian on solvated nucleotide, and single-, and double-stranded DNA, we mimic hydrogen transfer reactions from the backbone by OH radicals and report structural trajectories arising from on-the-fly electronic charge- and spin-density redistribution in these three different structural topologies of DNA. Trajectories reveal that H4' abstraction can disrupt the deoxyribose moiety through the formation of C4'=O4' ketone and a π-bond with base at C1'-N9 in a nucleotide versus only partial ketone formation in single- and double-stranded DNA, where the orientation of the base is topologically restrained. However, H5' abstraction can lead DNA cleavage at 5' end through the formation of C5'=O5' ketone and breakage of P-O5' bond. Results demonstrate that structural damages from oxidative reactions are restrained by base stacking and base-pair hydrogen bonding. The methodology can be suitably used to study targeted DNA and RNA damages from radicals and radiomimetic drugs to design DNA cleaving molecules for chemotherapy.


Assuntos
Dano ao DNA/fisiologia , DNA/química , DNA/metabolismo , Simulação de Dinâmica Molecular , Estresse Oxidativo/genética , Teoria Quântica , Biologia Computacional/métodos , Teoria da Densidade Funcional , Radicais Livres/química , Radicais Livres/metabolismo , Hidrogênio/química , Modelos Moleculares , Estrutura Molecular , Oxirredução , Estresse Oxidativo/fisiologia
11.
DNA Cell Biol ; 38(3): 272-280, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30657337

RESUMO

Helicobacter pylori is a major cause of gastric cancer. This study was aimed to explore the characteristic of DNA damage induced by H. pylori infection in gastric cancer AGS cells. After infection with H. pylori, the reactive oxygen species (ROS) levels in AGS cells were significantly higher than those in the uninfected cells. Cells with longer comet tails were detected after infection with H. pylori. The number of apurinic/apyrimidinic endonuclease 1- and phosphorylated H2AX-positive cells was significantly increased compared with the number of negative control cells. The expression of pChk1 and pChk2 was significantly upregulated by H. pylori infection. Cell growth was inhibited after H. pylori infection. All these results were dose dependent. The cell alterations were more significant upon infection with H. pylori at a multiplicity of infection (MOI) of 100:1 than at an MOI of 50:1. H. pylori infection can induce DNA single-strand breaks, DNA double-strand breaks, and cell cycle checkpoint activation after ROS generation in the gastric cancer cell line AGS, which is a potential driver for gastric cancer.


Assuntos
Dano ao DNA/fisiologia , Helicobacter pylori/patogenicidade , Neoplasias Gástricas/microbiologia , Proteínas de Bactérias/genética , Linhagem Celular Tumoral , DNA/metabolismo , Quebras de DNA , Dano ao DNA/genética , Células Epiteliais/metabolismo , Infecções por Helicobacter/genética , Infecções por Helicobacter/metabolismo , Helicobacter pylori/metabolismo , Humanos , Estresse Oxidativo/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Fatores de Risco , Neoplasias Gástricas/genética
12.
Int J Mol Sci ; 20(3)2019 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-30678294

RESUMO

Chemotherapeutic dosing, is largely based on the tolerance levels of toxicity today. Molecular imaging strategies can be leveraged to quantify DNA cytotoxicity and thereby serve as a theranostic tool to improve the efficacy of treatments. Methoxyamine-modified cyanine-7 (Cy7MX) is a molecular probe which binds to apurinic/apyrimidinic (AP)-sites, inhibiting DNA-repair mechanisms implicated by cytotoxic chemotherapies. Herein, we loaded (Cy7MX) onto polyethylene glycol-coated gold nanoparticles (AuNP) to selectively and stably deliver the molecular probe intravenously to tumors. We optimized the properties of Cy7MX-loaded AuNPs using optical spectroscopy and tested the delivery mechanism and binding affinity using the DLD1 colon cancer cell line in vitro. A 10:1 ratio of Cy7MX-AuNPs demonstrated a strong AP site-specific binding and the cumulative release profile demonstrated 97% release within 12 min from a polar to a nonpolar environment. We further demonstrated targeted delivery using imaging and biodistribution studies in vivo in an xenografted mouse model. This work lays a foundation for the development of real-time molecular imaging techniques that are poised to yield quantitative measures of the efficacy and temporal profile of cytotoxic chemotherapies.


Assuntos
Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Ouro/química , Nanopartículas Metálicas/química , Nanomedicina Teranóstica/métodos , Linhagem Celular Tumoral , Humanos , Estrutura Molecular
13.
Nat Commun ; 10(1): 241, 2019 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-30651562

RESUMO

Cell survival after oxidative DNA damage requires signaling, repair and transcriptional events often enabled by nucleosome displacement, exchange or removal by chromatin remodeling enzymes. Here, we show that Chromodomain Helicase DNA-binding protein 6 (CHD6), distinct to other CHD enzymes, is stabilized during oxidative stress via reduced degradation. CHD6 relocates rapidly to DNA damage in a manner dependent upon oxidative lesions and a conserved N-terminal poly(ADP-ribose)-dependent recruitment motif, with later retention requiring the double chromodomain and central core. CHD6 ablation increases reactive oxygen species persistence and impairs anti-oxidant transcriptional responses, leading to elevated DNA breakage and poly(ADP-ribose) induction that cannot be rescued by catalytic or double chromodomain mutants. Despite no overt epigenetic or DNA repair abnormalities, CHD6 loss leads to impaired cell survival after chronic oxidative stress, abnormal chromatin relaxation, amplified DNA damage signaling and checkpoint hypersensitivity. We suggest that CHD6 is a key regulator of the oxidative DNA damage response.


Assuntos
Montagem e Desmontagem da Cromatina/fisiologia , Dano ao DNA/fisiologia , DNA Helicases/metabolismo , Reparo do DNA/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/fisiologia , Células A549 , Sobrevivência Celular/fisiologia , Dano ao DNA/efeitos da radiação , DNA Helicases/genética , Pontos de Checagem da Fase G2 do Ciclo Celular/fisiologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Células HEK293 , Humanos , Microscopia Intravital , Lasers/efeitos adversos , Proteínas do Tecido Nervoso/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo
14.
Cell ; 176(1-2): 127-143.e24, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30633903

RESUMO

DNA damage provokes mutations and cancer and results from external carcinogens or endogenous cellular processes. However, the intrinsic instigators of endogenous DNA damage are poorly understood. Here, we identify proteins that promote endogenous DNA damage when overproduced: the DNA "damage-up" proteins (DDPs). We discover a large network of DDPs in Escherichia coli and deconvolute them into six function clusters, demonstrating DDP mechanisms in three: reactive oxygen increase by transmembrane transporters, chromosome loss by replisome binding, and replication stalling by transcription factors. Their 284 human homologs are over-represented among known cancer drivers, and their RNAs in tumors predict heavy mutagenesis and a poor prognosis. Half of the tested human homologs promote DNA damage and mutation when overproduced in human cells, with DNA damage-elevating mechanisms like those in E. coli. Our work identifies networks of DDPs that provoke endogenous DNA damage and may reveal DNA damage-associated functions of many human known and newly implicated cancer-promoting proteins.


Assuntos
Dano ao DNA/genética , Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Proteínas de Bactérias/metabolismo , Instabilidade Cromossômica/fisiologia , Replicação do DNA/fisiologia , Proteínas de Ligação a DNA/metabolismo , Escherichia coli/metabolismo , Instabilidade Genômica , Humanos , Proteínas de Membrana Transportadoras/fisiologia , Mutagênese , Mutação , Fatores de Transcrição/metabolismo
15.
Arch Ital Urol Androl ; 90(4): 254-259, 2019 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-30655635

RESUMO

OBJECTIVE: the aim of our study was to put forward insights to treat any possible correlation among sperm quality, sperm DNA damage and male age as they may have fertility implications for men who choose to delay fatherhood. MATERIALS AND METHODS: Our study is a non-interventional retrospective analysis of 3124 semen samples from patients that were investigated for the conventional semen parameters. Tunel test assay was set up for the evaluation of the sperm DNA fragmentation index (DFI). We applied the Kappa index to compare both the 1999 and the 2010 World Health Organization (WHO) reference criteria to evaluate the competence of such semen parameters categorization during the standard routine of our laboratory. RESULTS: With regards to our findings, it is possible to underline a significant relationship between aging and semen volume (p = 0.001), motility (p = 0.009), semen viscosity (p < 0.003) and sperm DNA damage (p < 0.009). We found a trend when focusing on the semen concentration (p = 0.05). The analysis of sperm morphology did not show any influence with advancing age (p = 0.606). When comparing both the 1999 and the 2010 WHO scales we found no accordance in the appraisal of sperm morphology but a very good one in the evaluation of the other parameters. CONCLUSIONS: Conventional semen analysis represents the opportunity to draw up a proxy insight on the male fertility status even if semen quality can only indirectly assess the probability of pregnancy. Several studies have verified a decay in the male reproductive system, sperm quality and fertility with advancing age although the reported results are not yet conclusive. Our results substantially agree with those findings outlined in the literature. Moreover we find that the discrepancy between the two WHO reference scales would eventually lead to an improper diagnosis of infertility.


Assuntos
Dano ao DNA/fisiologia , Contagem de Espermatozoides , Motilidade Espermática/fisiologia , Espermatozoides/fisiologia , Adulto , Idoso , Humanos , Infertilidade Masculina/etiologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sêmen/fisiologia , Análise do Sêmen , Adulto Jovem
16.
Biomed Pharmacother ; 111: 331-337, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30590321

RESUMO

In Brazilian folk medicine, copaiba oleoresin is widely known for its therapeutic activity, especially its wound healing and anti-inflammatory actions. Considering the relationship between inflammatory processes and carcinogenesis, this paper reports on the Copaifera reticulata Ducke oleoresin (CRO) chemopreventive potential in the colon carcinogenesis model in rats. To understand the mechanisms involved in this effect, the anti-inflammatory activity of CRO and its major chemical constituent, the diterpene ent-polyalthic acid (PA), were evaluated on the production of nitric oxide (NO) and prostaglandin E2 (PGE2) in mouse macrophages. For the chemoprevention assessment, the effect of CRO administered by gavage was investigated on DNA damage, pre-neoplastic lesions and mitotic frequencies induced by the 1,2-dimethylhydrazine (DMH; intraperitoneal injection) carcinogen by comet, aberrant crypt focus (ACF) and long-term assays, respectively. CRO reduced DNA damage (average 31.5%) and pre-neoplastic lesions (average 64.5%) induced by DMH, which revealed that CRO has antigenotoxic and anticarcinogenic effects. In the long-term assay, treatment with CRO significantly decreased mitoses in the tumor tissue, which suggested that CRO influenced carcinogenesis progression. PA reduced NO levels induced by lipopolysaccharides in macrophages. However, this diterpene showed no effect on PGE2. Taken together, our results suggest that PA exerts anti-inflammatory action via the NO pathway. The CRO chemopreventive effect may be partly due to the anti-inflammatory property of its major chemical constituent, PA. Our findings indicate that CRO is a promising agent to suppress colon carcinogenesis.


Assuntos
Carcinogênese/efeitos dos fármacos , Neoplasias do Colo/prevenção & controle , Fabaceae , Extratos Vegetais/farmacologia , Animais , Carcinogênese/metabolismo , Carcinogênese/patologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Quimioprevenção/métodos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/fisiologia , Relação Dose-Resposta a Droga , Masculino , Camundongos , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/uso terapêutico , Ratos , Ratos Wistar
17.
BMB Rep ; 52(1): 13-23, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30545443

RESUMO

Aging is accompanied by a time-dependent progressive deterioration of multiple factors of the cellular system. The past several decades have witnessed major leaps in our understanding of the biological mechanisms of aging using dietary, genetic, pharmacological, and physical interventions. Metabolic processes, including nutrient sensing pathways and mitochondrial function, have emerged as prominent regulators of aging. Mitochondria have been considered to play a key role largely due to their production of reactive oxygen species (ROS), resulting in DNA damage that accumulates over time and ultimately causes cellular failure. This theory, known as the mitochondrial free radical theory of aging (MFRTA), was favored by the aging field, but increasing inconsistent evidence has led to criticism and rejection of this idea. However, MFRTA should not be hastily rejected in its entirety because we now understand that ROS is not simply an undesired toxic metabolic byproduct, but also an important signaling molecule that is vital to cellular fitness. Notably, mitochondrial function, a term traditionally referred to bioenergetics and apoptosis, has since expanded considerably. It encompasses numerous other key biological processes, including the following: (i) complex metabolic processes, (ii) intracellular and endocrine signaling/communication, and (iii) immunity/inflammation. Here, we will discuss shortcomings of previous concepts regarding mitochondria in aging and their emerging roles based on recent advances. We will also discuss how the mitochondrial genome integrates with major theories on the evolution of aging. [BMB Reports 2019; 52(1): 13-23].


Assuntos
Envelhecimento/fisiologia , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Envelhecimento/genética , Animais , Apoptose , Dano ao DNA/fisiologia , DNA Mitocondrial/genética , Radicais Livres , Humanos , Estresse Oxidativo/fisiologia , Espécies Reativas de Oxigênio
18.
Oncol Rep ; 41(1): 552-558, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30365131

RESUMO

Breast cancer metastasis suppressor 1 (BRMS1) is a tumor metastasis suppressor implicated in multiple steps during the metastatic cascade. Many proteins interacting with BRMS1 have been identified to unravel the intracellular signaling mechanisms. In the present study, we report that FANCI is a novel interacting protein of BRMS1 as determined by co­immunoprecipitation assay. The linker region between two coiled­coil motifs of BRMS1 is required for BRMS1­FANCI interaction. FANCI is an essential protein in the Fanconi anemia (FA) pathway responsible for the repair of DNA interstrand crosslinks (ICLs). We demonstrated that knockdown or knockout of BRMS1 significantly diminished the monoubiquitination of FANCI and FANCD2 in response to DNA ICL damage. BRMS1­deficient cells exhibited suppressed FANCD2 foci formation and hypersensitivity to ICLs. Moreover, rescue assays by utilizing different BRMS1 constructs suggested that BRMS1­FANCI interaction is necessary for the regulatory role of BRMS1 in the FA pathway. Overall, our findings characterize BRMS1 as a novel regulatory protein functioning in the DNA repair pathway via protein interaction.


Assuntos
Dano ao DNA/fisiologia , DNA/metabolismo , Proteínas de Grupos de Complementação da Anemia de Fanconi/metabolismo , Proteínas Repressoras/metabolismo , Linhagem Celular , Reparo do DNA/fisiologia , Anemia de Fanconi/metabolismo , Células HEK293 , Humanos , Transdução de Sinais/fisiologia , Ubiquitinação/fisiologia
19.
Cell ; 176(1-2): 144-153.e13, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30554877

RESUMO

Abasic sites are one of the most common DNA lesions. All known abasic site repair mechanisms operate only when the damage is in double-stranded DNA. Here, we report the discovery of 5-hydroxymethylcytosine (5hmC) binding, ESC-specific (HMCES) as a sensor of abasic sites in single-stranded DNA. HMCES acts at replication forks, binds PCNA and single-stranded DNA, and generates a DNA-protein crosslink to shield abasic sites from error-prone processing. This unusual HMCES DNA-protein crosslink intermediate is resolved by proteasome-mediated degradation. Acting as a suicide enzyme, HMCES prevents translesion DNA synthesis and the action of endonucleases that would otherwise generate mutations and double-strand breaks. HMCES is evolutionarily conserved in all domains of life, and its biochemical properties are shared with its E. coli ortholog. Thus, HMCES is an ancient DNA lesion recognition protein that preserves genome integrity by promoting error-free repair of abasic sites in single-stranded DNA.


Assuntos
5-Metilcitosina/análogos & derivados , Reparo do DNA/fisiologia , DNA de Cadeia Simples/fisiologia , 5-Metilcitosina/metabolismo , Ácido Apurínico/metabolismo , DNA/metabolismo , Dano ao DNA/fisiologia , Replicação do DNA/fisiologia , DNA de Cadeia Simples/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Endonucleases , Escherichia coli/metabolismo , Polinucleotídeos/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo
20.
Proc Natl Acad Sci U S A ; 115(52): E12453-E12462, 2018 12 26.
Artigo em Inglês | MEDLINE | ID: mdl-30541889

RESUMO

To combat DNA damage, organisms mount a DNA damage response (DDR) that results in cell cycle regulation, DNA repair and, in severe cases, cell death. Underscoring the importance of gene regulation in this response, studies in Arabidopsis have demonstrated that all of the aforementioned processes rely on SUPPRESSOR OF GAMMA RESPONSE 1 (SOG1), a NAC family transcription factor (TF) that has been functionally equated to the mammalian tumor suppressor, p53. However, the expression networks connecting SOG1 to these processes remain largely unknown and, although the DDR spans from minutes to hours, most transcriptomic data correspond to single time-point snapshots. Here, we generated transcriptional models of the DDR from GAMMA (γ)-irradiated wild-type and sog1 seedlings during a 24-hour time course using DREM, the Dynamic Regulatory Events Miner, revealing 11 coexpressed gene groups with distinct biological functions and cis-regulatory features. Within these networks, additional chromatin immunoprecipitation and transcriptomic experiments revealed that SOG1 is the major activator, directly targeting the most strongly up-regulated genes, including TFs, repair factors, and early cell cycle regulators, while three MYB3R TFs are the major repressors, specifically targeting the most strongly down-regulated genes, which mainly correspond to G2/M cell cycle-regulated genes. Together these models reveal the temporal dynamics of the transcriptional events triggered by γ-irradiation and connects these events to TFs and biological processes over a time scale commensurate with key processes coordinated in response to DNA damage, greatly expanding our understanding of the DDR.


Assuntos
Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Reparo do DNA/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Pontos de Checagem do Ciclo Celular , Dano ao DNA/fisiologia , Reparo do DNA/genética , Regulação da Expressão Gênica de Plantas/genética , Mutação/genética , Transativadores/metabolismo , Ativação Transcricional , Transcriptoma/genética
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