RESUMEN
7-Deaza-2'-deoxyisoguanosine forms stable inverse Watson-Crick base pairs with 5-methyl-2'-deoxyisocytidine and purine-purine base pairs with 2'-deoxyguanosine or 5-aza-7-deaza-2'-deoxyguanosine. Both base pairs expand the genetic coding system. The manuscript reports on the functionalization of these base pairs with halogen atoms and clickable side chains introduced at 7-position of the 7-deazapurine base. Oligonucleotides containing the functionalized base pairs were prepared by solid-phase synthesis. To this end, a series of phosphoramidites were synthesized and clickable side chains with short and long linkers were incorporated in oligonucleotides. Fluorescent pyrene conjugates were obtained by postmodification. Functionalization of DNA with a single inverse Watson-Crick base pair by halogens or clickable residues has only a minor impact on duplex stability. Pyrene click adducts increase (long linker) or decrease (short linker) the double helix stability. Stable hybrid duplexes were constructed containing three consecutive purine-purine pairs of 7-functionalized 7-deaza-2'-deoxyisoguanine with guanine or 5-aza-7-deazaguanine in the center and Watson-Crick pairs at both ends. The incorporation of a hybrid base pair tract of 7-deaza-2'-deoxyisoguanosine/5-aza-7-deaza-2'-deoxyguanosine pairs stabilizes the double helix strongly. Fluorescence intensity of pyrene short linker adducts increased when the 7-deazapurine base was positioned opposite to 5-methylisocytosine (inverse base pair) compared to purine-purine base pairs with guanine or 5-aza-7-deazaguanine in opposite positions. For long liker adducts, the situation is more complex. Circular dichroism (CD) spectra of purine DNA differ to those of Watson-Crick double helices and are indicative for the new DNA constructs. The impact of 7-deaza-2'-deoxyisoguanine base pair functionalization is studied for the first time and all experimental details are reported to prepare DNA functionalized at the 7-deazaisoguanine site. The influence of single and multiple incorporations on DNA structure and stability is shown. Clickable residues introduced at the 7-position of the 7-deazaisoguanine base provide handles for Huisgen-Sharpless-Meldal click cycloadditions without harming the stability of purine-pyrimidine and purine-purine base pairs. Other chemistries might be used for bioconjugation. Our investigation paves the way for the functionalization of a new DNA related recognition system expanding the common Watson-Crick regime.
Asunto(s)
Emparejamiento Base , ADN , Purinas , Purinas/química , ADN/química , Guanosina/química , Guanosina/análogos & derivados , Pirenos/química , Oligonucleótidos/química , Desoxiguanosina/química , Desoxiguanosina/análogos & derivadosRESUMEN
Papillary thyroid carcinoma (PTC) is a common endocrine cancer with a good prognosis. Radioactive iodine is thought to be useful for individuals who have had a total or almost total thyroidectomy, but its effects are still controversial. The effects of radioactive iodine-131 (I-131) treatment on oxidative and chromosomal damage in PTC patients were examined in this study, which was carried out with 16 patients newly diagnosed with PTC and 20 healthy control subjects with similar age and gender. Blood samples were taken from patients with PTC at five sampling times (before total thyroidectomy, after total thyroidectomy, and seven days, six months, and one year after treatment) and from control subjects. The cytokinesis block micronucleus cytome (CBMN-cyt) assay parameters in peripheral blood lymphocytes of patients with PTC and controls were evaluated and plasma 8-hydroxydeoxyguanosine (8-OHdG) levels were measured. Furthermore, genome instability and oxidative DNA damage in peripheral blood lymphocytes and plasma of patients with PTC were evaluated before total thyroidectomy (n=16), after total thyroidectomy (before I-131 treatment) (n=16), seven days (n=10), six months (n=5), and one year after treatment (n=5). The numbers of CBMN-cyt assay parameters (micronucleus; MN and nucleoplasmic bridges; NPB) and 8-OHdG levels in patients with PTC were determined to be significantly higher than in those of the control subjects and these values significantly decreased after total thyroidectomy (before I-131 treatment). While the number of MN, apoptotic, and necrotic cells increased after I-131 treatment, it significantly decreased after six months and one year after treatment. The results achieved in this study suggest that I-131 treatment may pose a threat to cells and that radioactive iodine therapy should be avoided (if possible) for patients with PTC after total thyroidectomy.
Asunto(s)
Daño del ADN , Radioisótopos de Yodo , Estrés Oxidativo , Cáncer Papilar Tiroideo , Neoplasias de la Tiroides , Tiroidectomía , Humanos , Radioisótopos de Yodo/uso terapéutico , Radioisótopos de Yodo/efectos adversos , Neoplasias de la Tiroides/sangre , Neoplasias de la Tiroides/radioterapia , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/genética , Femenino , Masculino , Adulto , Persona de Mediana Edad , Cáncer Papilar Tiroideo/sangre , Cáncer Papilar Tiroideo/radioterapia , Estrés Oxidativo/efectos de los fármacos , Pruebas de Micronúcleos , Carcinoma Papilar/sangre , Carcinoma Papilar/patología , Carcinoma Papilar/radioterapia , Carcinoma/radioterapia , Carcinoma/sangre , Carcinoma/genética , Linfocitos/efectos de la radiación , Linfocitos/efectos de los fármacos , 8-Hidroxi-2'-Desoxicoguanosina/sangre , Desoxiguanosina/análogos & derivados , Desoxiguanosina/sangre , Estudios de Casos y Controles , Inestabilidad GenómicaRESUMEN
Tandem lesions, which are defined by two or more contiguously damaged nucleotides, are a hallmark of ionizing radiation. Recently, tandem lesions containing 5-formyl-2'-deoxyuridine (5-fdU) flanked by a 5'-8-OxodGuo or Fapyâ¢dG were discovered, and they are more mutagenic in human cells than the isolated lesions. In the current study, we examined replication of these tandem lesions in Escherichia coli. Bypass efficiency of both tandem lesions was reduced by 30-40% compared to the isolated lesions. Mutation frequencies (MFs) of isolated 8-OxodGuo and Fapyâ¢dG were low, and no mutants were isolated from replication of a 5-fdU construct. The types of mutations from 8-OxodGuo were targeted G â T transversion, whereas Fapyâ¢dG predominantly gave G â T and G deletion. 5'-8-OxodGuo-5-fdU also gave exclusively G â T mutation, which was 3-fold and 11-fold greater, without and with SOS induction, respectively, compared to that of an isolated 8-OxodGuo. In mutY/mutM cells, the MF of 8-OxodGuo and 5'-8-OxodGuo-5-fdU increased 13-fold and 7-fold, respectively. The MF of 5'-8-OxodGuo-5-fdU increased 2-fold and 3-fold in Pol II- and Pol IV-deficient cells, respectively, suggesting that these polymerases carry out largely error-free bypass. The MF of 5'- Fapyâ¢dG-5-fdU was similar without (13 ± 1%) and with (16 ± 2%) SOS induction. Unlike the complex mutation spectrum reported earlier in human cells for 5'- Fapyâ¢dG-5-fdU, with G â T as the major type of errors, in E. coli, the mutations were predominantly from deletion of 5-fdU. We postulate that removal of adenine-incorporated opposite 8-OxodGuo by Fpg and MutY repair proteins is partially impaired in the tandem 5'-8-OxodGuo-5-fdU, resulting in an increase in the G â T mutations, whereas a slippage mechanism may be operating in the 5'- Fapyâ¢dG-5-fdU mutagenesis. This study showed that not only are these tandem lesions more mutagenic than the isolated lesions but they may also exhibit different types of mutations in different organisms.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Escherichia coli , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , 8-Hidroxi-2'-Desoxicoguanosina/metabolismo , Desoxiuridina/análogos & derivados , Desoxiuridina/química , Desoxiuridina/farmacología , Mutágenos/toxicidad , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Mutación , Mutagénesis , Daño del ADNRESUMEN
8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) base is the predominant form of commonly observed DNA oxidative damage. DNA impairment profoundly impacts gene expression and serves as a pivotal factor in stimulating neurodegenerative disorders, cancer, and aging. Therefore, precise quantification of 8-oxoG has clinical significance in the investigation of DNA damage detection methodologies. However, at present, the existing approaches for 8-oxoG detection pose challenges in terms of convenience, expediency, affordability, and heightened sensitivity. We employed the sandwich enzyme-linked immunosorbent assay (ELISA) technique, a highly efficient and swift colorimetric method, to detect variations in 8-oxo-dG content in MCF-7 cell samples stimulated with different concentrations of hydrogen peroxide (H2O2). We determined the concentration of H2O2 that induced oxidative damage in MCF-7 cells by detecting its IC50 value in MCF-7 cells. Subsequently, we treated MCF-7 cells with 0, 0.25, and 0.75 mM H2O2 for 12 h and extracted 8-oxo-dG from the cells. Finally, the samples were subjected to ELISA. Following a series of steps, including plate spreading, washing, incubation, color development, termination of the reaction, and data collection, we successfully detected changes in the 8-oxo-dG content in MCF-7 cells induced by H2O2. Through such endeavors, we aim to establish a method to evaluate the degree of DNA oxidative damage within cell samples and, in doing so, advance the development of more expedient and convenient approaches for DNA damage detection. This endeavor is poised to make a meaningful contribution to the exploration of associative analyses between DNA oxidative damage and various domains, including clinical research on diseases and the detection of toxic substances.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Daño del ADN , Ensayo de Inmunoadsorción Enzimática , Peróxido de Hidrógeno , Estrés Oxidativo , Humanos , Daño del ADN/efectos de los fármacos , Células MCF-7 , Ensayo de Inmunoadsorción Enzimática/métodos , Peróxido de Hidrógeno/farmacología , Estrés Oxidativo/efectos de los fármacos , Desoxiguanosina/análogos & derivados , Desoxiguanosina/análisisRESUMEN
The genome-the source of life and platform of evolution-is continuously exposed to harmful factors, both extra- and intra-cellular. Their activity causes different types of DNA damage, with approximately 80 different types of lesions having been identified so far. In this paper, the influence of a clustered DNA damage site containing imidazolone (Iz) or oxazolone (Oz) and 7,8-dihydro-8-oxo-2'-deoxyguanosine (OXOdG) on the charge transfer through the double helix as well as their electronic properties were investigated. To this end, the structures of oligo-Iz, d[A1Iz2A3OXOG4A5]*d[T5C4T3C2T1], and oligo-Oz, d[A1Oz2A3OXOG4A5]*d[T5C4T3C2T1], were optimized at the M06-2X/6-D95**//M06-2X/sto-3G level of theory in the aqueous phase using the ONIOM methodology; all the discussed energies were obtained at the M06-2X/6-31++G** level of theory. The non-equilibrated and equilibrated solvent-solute interactions were taken into consideration. The following results were found: (A) In all the discussed cases, OXOdG showed a higher predisposition to radical cation formation, and B) the excess electron migration toward Iz and Oz was preferred. However, in the case of oligo-Oz, the electron transfer from Oz2 to complementary C4 was noted during vertical to adiabatic anion relaxation, while for oligo-Iz, it was settled exclusively on the Iz2 moiety. The above was reflected in the charge transfer rate constant, vertical/adiabatic ionization potential, and electron affinity energy values, as well as the charge and spin distribution. It can be postulated that imidazolone moiety formation within the CDL ds-oligo structure and its conversion to oxazolone can significantly influence the charge migration process, depending on the C2 carbon hybridization sp2 or sp3. The above can confuse the single DNA damage recognition and removal processes, cause an increase in mutagenesis, and harm the effectiveness of anticancer therapy.
Asunto(s)
Daño del ADN , Imidazoles , Imidazoles/química , Oxazolona/química , 8-Hidroxi-2'-Desoxicoguanosina/química , ADN/química , Modelos Moleculares , Desoxiguanosina/química , Desoxiguanosina/análogos & derivados , TermodinámicaRESUMEN
Formamidopyrimidine (Fapyâ¢dG) is a major lesion arising from oxidation of dG that is produced from a common chemical precursor of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-OxodGuo). In human cells, replication of single-stranded shuttle vectors containing Fapyâ¢dG is more mutagenic than 8-OxodGuo. Here, we present the first data regarding promoter dependent RNA polymerase II bypass of Fapyâ¢dG. 8-OxodGuo bypass was examined side-by-side. Experiments were carried out using double-stranded shuttle vectors in HeLa cell nuclear lysates and in HEK 293T cells. The lesions do not significantly block transcriptional bypass efficiency. Less than 2% adenosine incorporation occurred in cells when the lesions were base paired with dC. Inhibiting base excision repair in HEK 293T cells significantly increased adenosine incorporation, particularly from Fapyâ¢dG:dC bypass which yielded â¼25% adenosine incorporation. No effect was detected upon transcriptional bypass of either lesion in nucleotide excision repair deficient cells. Transcriptional mutagenesis was significantly higher when shuttle vectors containing dA opposite one of the lesions were employed. For Fapyâ¢dG:dA bypass, adenosine incorporation was greater than 85%; whereas 8-OxodGuo:dA yielded >20% point mutations. The combination of more frequent replication mistakes and greater error-prone Pol II bypass suggest that Fapyâ¢dG is more mutagenic than 8-OxodGuo.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Daño del ADN , Desoxiguanosina , Regiones Promotoras Genéticas , ARN Polimerasa II , Humanos , ARN Polimerasa II/metabolismo , ARN Polimerasa II/genética , Células HEK293 , 8-Hidroxi-2'-Desoxicoguanosina/metabolismo , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Células HeLa , Reparación del ADN , Transcripción Genética , Pirimidinas , Dímeros de Pirimidina/metabolismo , Dímeros de Pirimidina/genéticaRESUMEN
DNA is constantly damaged by various external and internal factors. In particular, oxidative damage occurs in a steady state, and 8-oxo-2'-deoxyguanosine (oxodG) is known as the main oxidative damage. OxodG is a strong genotoxic nucleoside and is thought to be involved in the pathogenesis of cancer and neurological diseases. However, a breakthrough method to detect the position of oxodG in DNA has not yet been developed. Therefore, we attempted to develop a novel method to detect oxodG in DNA using artificial nucleosides. Recently, we have succeeded in the recognition of oxodG in DNA by a single nucleotide elongation reaction using nucleoside derivatives based on a purine skeleton with a 1,3-diazaphenoxazine unit. In this study, we developed a new nucleoside derivative with a pyrimidine skeleton in order to further improve the recognition ability and enzymatic reaction efficiency. We, therefore, designed and synthesized 2'-deoxycytidine-1,3-diazaphenoxazine (Cdap) and its triphosphate derivatives. The results showed that it was incorporated into the primer strand relative to the dG template because of its cytidine skeleton, but it was more effective at the complementary position of the oxodG template. These results indicate that the new nucleoside derivative can be considered as one of the new candidates for the detection of oxodG in DNA.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , ADN , Desoxicitidina , Oxazinas , ADN/química , Desoxicitidina/análogos & derivados , Desoxicitidina/química , Oxazinas/química , Desoxiguanosina/química , Desoxiguanosina/análogos & derivados , Daño del ADN , Nucleótidos/química , PolifosfatosRESUMEN
Hairdressers are constantly occupationally exposed to many chemicals have the potential to cause allergies and carcinogenic effects, act as skin and eye irritants and induce oxidative stress and DNA damage. This study aimed to evaluate occupation-induced genotoxicity based on the presence of micronucleus (MN) and other nuclear anomalies in urothelial cells and measure oxidative DNA damage based on the 8-hydroxy-2'-deoxyguanosine level in the urine of Turkish hairdressers. Originality of this study comes from that there was no study on MN and other nuclear anomalies frequencies and oxidative DNA damage in urine samples of hairdressers in the literature. The mean±standard deviation frequency () of micronucleated (MNed) cells was higher in the hairdresser group (n=56) (4.81±7.87, p<0.001) than in the control group (n=56) (0.93±1.85). Nuclear buds were not observed in either group. While the frequency of basal cells was higher in the control group (446.6±106.21) than in the hairdresser group (367.78±101.51, p<0.001), the frequency of binuclear, karyolytic, pycnotic and karyorrhectic cells were higher in the hairdresser group (0.41±0.80, p<0.001; 438.02±118.27, p<0.001; 0.43±0.76, p<0.001; and 47.27±28.40, p<0.001) than in the control group (0.04±0.27, 358.57±95.71, 0.05±0.23 and 24.41±14.50). Condensed chromatins were observed only in the hairdresser group. Specific gravity adjusted 8-hydroxy-2'-deoxyguanosine level was statistically lower in the hairdresser group (908.21±403.25â¯ng/mL-SG) compared to the control group (1003.09±327.09â¯ng/mL-SG) (p=0.024). No significant correlation was found between the 8-hydroxy-2'-deoxyguanosine level and the frequency MN. The amount of formaldehyde released during Brazilian keratin treatment was higher than the American Conference of Governmental Industrial Hygienists -Threshold Limit Value (ACGIH-TLV; 0.1â¯ppm). Similarly, the amount of ethyl acetate released in three salons was above the recommended limit (400â¯ppm). These findings suggest that hairdressers have an increased risk of genotoxicity and cytotoxicity owing to occupational exposure, regardless of age, working hours, smoking and alcohol consumption.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Daño del ADN , Desoxiguanosina , Micronúcleos con Defecto Cromosómico , Pruebas de Micronúcleos , Exposición Profesional , Urotelio , Humanos , 8-Hidroxi-2'-Desoxicoguanosina/orina , Exposición Profesional/efectos adversos , Adulto , Turquía , Urotelio/efectos de los fármacos , Urotelio/patología , Urotelio/metabolismo , Urotelio/citología , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Masculino , Micronúcleos con Defecto Cromosómico/inducido químicamente , Daño del ADN/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Persona de Mediana Edad , Femenino , Adulto Joven , Estudios de Casos y Controles , Núcleo Celular/efectos de los fármacosRESUMEN
Previous evidence suggests elevated levels of oxidatively-induced DNA damage, particularly 8-hydroxy-2'-deoxyguanosine (8-OH-dG), and abnormalities in the repair of 8-OH-dG by the base excision repair (BER) in bipolar disorder (BD). However, the genetic disposition of these abnormalities remains unknown. In this study, we aimed to investigate the levels of oxidatively-induced DNA damage and BER mechanisms in individuals with BD and their siblings, as compared to healthy controls (HCs). 46 individuals with BD, 41 siblings of individuals with BD, and 51 HCs were included in the study. Liquid chromatography-tandem mass spectrometry was employed to evaluate the levels of 8-OH-dG in urine, which were then normalized based on urine creatinine levels. The real-time-polymerase chain reaction was used to measure the expression levels of 8-oxoguanine DNA glycosylase 1 (OGG1), apurinic/apyrimidinic endonuclease 1 (APE1), poly ADP-ribose polymerase 1 (PARP1), and DNA polymerase beta (POLß). The levels of 8-OH-dG were found to be elevated in both individuals with BD and their siblings when compared to the HCs. The OGG1 and APE1 expressions were downregulated, while POLß expressions were upregulated in both the patient and sibling groups compared to the HCs. Age, smoking status, and the number of depressive episodes had an impact on APE1 expression levels in the patient group while body mass index, smoking status, and past psychiatric history had an impact on 8-OH-dG levels in siblings. Both individuals with BD and unaffected siblings presented similar abnormalities regarding oxidatively-induced DNA damage and BER, suggesting a link between abnormalities in DNA damage/BER mechanisms and familial susceptibility to BD. Our findings suggest that targeting the oxidatively-induced DNA damage and BER pathway could offer promising therapeutic strategies for reducing the risk of age-related diseases and comorbidities in individuals with a genetic predisposition to BD.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Trastorno Bipolar , Daño del ADN , ADN Glicosilasas , Reparación del ADN , Estrés Oxidativo , Hermanos , Humanos , Trastorno Bipolar/genética , Trastorno Bipolar/metabolismo , Femenino , Masculino , Adulto , ADN Glicosilasas/genética , Estrés Oxidativo/genética , Persona de Mediana Edad , ADN Polimerasa beta/genética , ADN Polimerasa beta/metabolismo , ADN-(Sitio Apurínico o Apirimidínico) Liasa/genética , Estudios de Casos y Controles , Adulto Joven , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Reparación por EscisiónRESUMEN
The major product of DNA-methylating agents, N7-methyl-2'-deoxyguanosine (MdG), is a persistent lesion in vivo, but it is not believed to have a large direct physiological impact. However, MdG reacts with histone proteins to form reversible DNA-protein cross-links (DPCMdG), a family of DNA lesions that can significantly threaten cell survival. In this paper, we developed a tandem mass spectrometry method for quantifying the amounts of MdG and DPCMdG in nuclear DNA by taking advantage of their chemical lability and the concurrent release of N7-methylguanine. Using this method, we determined that DPCMdG is formed in less than 1% yield based upon the levels of MdG in methyl methanesulfonate (MMS)-treated HeLa cells. Despite its low chemical yield, DPCMdG contributes to MMS cytotoxicity. Consequently, cells that lack efficient DPC repair by the DPC protease SPRTN are hypersensitive to MMS. This investigation shows that the downstream chemical and biochemical effects of initially formed DNA damage can have significant biological consequences. With respect to MdG formation, the initial DNA lesion is only the beginning.
Asunto(s)
ADN , Desoxiguanosina , Metilmetanosulfonato , Humanos , Células HeLa , ADN/metabolismo , ADN/química , ADN/efectos de los fármacos , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Desoxiguanosina/química , Metilmetanosulfonato/química , Metilmetanosulfonato/farmacología , Espectrometría de Masas en Tándem , Supervivencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Reactivos de Enlaces Cruzados/química , Proteínas de Unión al ADNRESUMEN
INTRODUCTION: Peritoneal dialysis (PD) is an effective treatment modality for advanced kidney failure, offering patients a significant degree of independence. However, the long-term use of PD is limited due to the degeneration of the peritoneal membrane, resulting in reduced dialysis adequacy. Evaluating the peritoneal membrane condition in patients with advanced kidney failure who are undergoing PD is challenging with existing methods. Therefore, this study aimed to investigate the correlation between 8-hydroxy-2'-deoxyguanosine (8OHDG) levels in the peritoneal solution of patients undergoing PD and various factors, such as peritoneal equilibration test (PET), dialysis adequacy (Kt/V), underlying diseases, serum ferritin, and albumin levels. 8OHDG is a sensitive marker of oxidative stress caused by DNA damage. METHODS: A total of 56 patients were included in this cross-sectional study. Five milliliters of PD fluid were collected from the patients, and 8-OHdG levels were measured using ELISA method. Then, they were compared with PET, Kt/V, albumin, and ferritin markers in the patients' files, and the results were analyzed by statistical tests. RESULTS: The study examined the correlation between 8OHDG and other markers. It was found that this index had significant associations with PET and underlying HTN (P < .05), whereas no significant associations were identified with the other markers. CONCLUSION: The results of the present study demonstrate that the level of 8OHDG, as one of the oxidative stress markers, could be used to evaluate the function of the peritoneum in patients undergoing PD. DOI: 10.52547/ijkd.7654.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Estrés Oxidativo , Diálisis Peritoneal , Femenino , Humanos , Masculino , 8-Hidroxi-2'-Desoxicoguanosina/análisis , Biomarcadores/sangre , Biomarcadores/metabolismo , Estudios Transversales , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Desoxiguanosina/sangre , Ferritinas/sangre , Ferritinas/análisis , Fallo Renal Crónico/terapia , Fallo Renal Crónico/sangre , Diálisis Peritoneal/efectos adversos , Peritoneo/química , Peritoneo/metabolismo , Peritoneo/patología , Albúmina Sérica/análisis , Albúmina Sérica/metabolismoRESUMEN
This study was designed to investigate the potential neuronal damage mechanism of the okadaic acid (OA) in the brain tissues of zebrafish embryos by evaluating in terms of immunofluorescence of Nf KB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG signaling pathways. We also evaluated body malformations. For this purpose, zebrafish embryos were exposed to 0.5 µg/ml, 1 µg/ml and 2.5 µg/ml of OA for 5 days. After application, FITC/GFP labeled protein-specific antibodies were used in immunofluorescence assay for NfKB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG respectively. The results indicated that OA caused immunofluorescence positivity of NfKB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG in a dose-dependent manner in the brain tissues of zebrafish embryos. Pericardial edema (PE), nutrient sac edema (YSE) and body malformations, tail malformation, short tail and head malformation (BM) were detected in zebrafish embryos. These results suggest that OA induces neuronal damage by affecting the modulation of DNA damage, apoptotic, and inflammatory activities in the brain tissues of zebrafish embryos. The increase in signaling pathways shows that OA can cause damage in the structure and function of brain nerve cells. Our results provide a new basis for the comprehensive assessment of the neural damage of OA and will offer enable us to better understand molecular the mechanisms underlying the pathophysiology of OA toxicity.
Asunto(s)
Encéfalo , FN-kappa B , Ácido Ocadaico , Transducción de Señal , Receptor Toll-Like 4 , Pez Cebra , Animales , Pez Cebra/inmunología , Encéfalo/efectos de los fármacos , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Transducción de Señal/efectos de los fármacos , Ácido Ocadaico/toxicidad , FN-kappa B/metabolismo , FN-kappa B/inmunología , 8-Hidroxi-2'-Desoxicoguanosina , Caspasa 3/metabolismo , Caspasa 3/genética , Larva/efectos de los fármacos , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-fos/genética , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismoRESUMEN
INTRODUCTION: Benzotriazoles and benzothiazoles (BTs) are high-production volume chemicals as well as widely distributed emerging pollutants with potential health risk. However, information about human exposure to BTs and associated health outcomes is limited. OBJECTIVE: We aimed to characterise exposure to BTs among Czech men, including possible occupational exposure among firefighters, its predictors, and its associations with liver function, serum lipids and oxidative stress. METHODS: 165 participants (including 110 firefighters) provided urine and blood samples that were used to quantify the urinary levels of 8 BTs (high-performance liquid chromatography-tandem mass spectrometry), and 4 liver enzymes, cholesterol, low-density lipoprotein, and 8-hydroxy-2'-deoxyguanosine. Linear regression was used to assess associations with population characteristics and biomarkers of liver function, serum lipids and oxidative stress. Regression models were adjusted for potential confounding variables and false discovery rate procedure was applied to account for multiplicity. RESULTS: The BTs ranged from undetected up to 46.8 ng/mL. 2-hydroxy-benzothiazole was the most predominant compound (detection frequency 83%; median 1.95 ng/mL). 1-methyl-benzotriazole (1M-BTR) was measured in human samples for the first time, with a detection frequency 77% and median 1.75 ng/mL. Professional firefighters had lower urinary 1M-BTR compared to non-firefighters. Urinary 1M-BTR was associated with levels of γ-glutamyl transferase (ß = - 17.54%; 95% CI: - 26.127, - 7.962). CONCLUSION: This is the first study to investigate BT exposure in Central Europe, including potentially exposed firefighters. The findings showed a high prevalence of BTs in the study population, the relevance of 1M-BTR as a new biomarker of exposure, and an urgent need for further research into associated adverse health outcomes.
Asunto(s)
Benzotiazoles , Biomarcadores , Exposición Profesional , Estrés Oxidativo , Triazoles , Humanos , Masculino , Estrés Oxidativo/efectos de los fármacos , Exposición Profesional/análisis , Biomarcadores/sangre , Biomarcadores/orina , Adulto , Persona de Mediana Edad , República Checa , Bomberos , Hígado/efectos de los fármacos , Lípidos/sangre , 8-Hidroxi-2'-Desoxicoguanosina/orina , 8-Hidroxi-2'-Desoxicoguanosina/sangre , Colesterol/sangre , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Desoxiguanosina/sangreRESUMEN
Importance: All-cause mortality and the risk for age-related medical disease is increased in individuals with psychiatric illness, but the underlying biological mechanisms are not known. Oxidative stress on nucleic acids (DNA and RNA; NA-OXS) is a molecular driver of aging and a potential pathophysiological mechanism in a range of age-related disorders. Objective: To study the levels of markers of NA-OXS in a large cohort of community-dwelling individuals with and without psychiatric illness and to evaluate their association with prospective all-cause mortality. Design, Setting, and Participants: This cohort study used a combined cohort of participants from 2 population-based health studies: the Danish General Suburban Population Study (January 2010 to October 2013) and nondiabetic control participants from the Vejle Diabetes Biobank study (March 2007 to May 2010). Individual history of psychiatric illness was characterized using register data on psychiatric diagnoses and use of psychotropic drugs before baseline examination. Urinary markers of systemic RNA (8-oxo-7,8-dihydroguanosine [8-oxoGuo]) and DNA (8-oxo-7,8-dihydro-2'-deoxyguanosine [8-oxodG]) damage from oxidation were measured by ultraperformance liquid chromatography-tandem mass spectrometry. Cox proportional hazard regression models were applied for survival analyses, using register-based all-cause mortality updated to May 2023. The follow-up time was up to 16.0 years. Exposures: History of psychiatric illness. Main Outcomes and Measures: Mortality risk according to psychiatric illness status and 8-oxoGuo or 8-oxodG excretion level. Results: A total of 7728 individuals were included (3983 [51.5%] female; mean [SD] age, 58.6 [11.9] years), 3095 of whom (40.0%) had a history of psychiatric illness. Mean (SD) baseline 8-oxoGuo was statistically significantly higher in individuals with psychiatric illness than in those without (2.4 [1.2] nmol/mmol vs 2.2 [0.9] nmol/mmol; P < .001), whereas 8-oxodG was not. All-cause mortality was higher in the psychiatric illness group vs the no psychiatric illness group (hazard ratio [HR], 1.44; 95% CI, 1.27-1.64; P < .001) and increased sequentially with each increasing tertile of 8-oxoGuo excretion in both groups to an almost doubled risk in the psychiatric illness/high 8-oxoGuo group compared to the no psychiatric illness/low 8-oxoGuo reference group (HR, 1.99; 95% CI, 1.58-2.52; P < .001). These results persisted after adjustment for a range of potential confounders and in a sensitivity analysis stratified for sex. Conclusions and Relevance: This study establishes systemic oxidative stress-induced damage to RNA as a potential mechanism in the accelerated aging observed in psychiatric disorders and urinary 8-oxoGuo as a potentially useful marker of mortality risk in individuals with psychiatric illness.
Asunto(s)
8-Hidroxi-2'-Desoxicoguanosina , Daño del ADN , Guanosina , Guanosina/análogos & derivados , Trastornos Mentales , Estrés Oxidativo , ARN , Humanos , Estrés Oxidativo/fisiología , Femenino , Masculino , Trastornos Mentales/epidemiología , Persona de Mediana Edad , 8-Hidroxi-2'-Desoxicoguanosina/orina , Guanosina/orina , Anciano , ARN/genética , Dinamarca/epidemiología , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Estudios de Cohortes , Adulto , Biomarcadores , Estudios Prospectivos , MortalidadRESUMEN
Oxidative DNA lesions cause significant detrimental effects on a living species. Two major DNA lesions resulting from dG oxidation, 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-OxodGuo) and formamidopyrimidine (Fapy·dG), are produced from a common chemical intermediate. Fapy·dG is formed in comparable yields under oxygen-deficient conditions. Replicative bypass of Fapy·dG in human cells is more mutagenic than that of 8-OxodGuo. Despite the biological importance of transcriptional mutagenesis, there are no reports of the effects of Fapy·dG on RNA polymerase II (Pol II) activity. Here we perform comprehensive kinetic studies to investigate the impact of Fapy·dG on three key transcriptional fidelity checkpoint steps by Pol II: insertion, extension, and proofreading steps. The ratios of error-free versus error-prone incorporation opposite Fapy·dG are significantly reduced in comparison with undamaged dG. Similarly, Fapy·dG:A mispair is extended with comparable efficiency as that of the error-free, Fapy·dG:C base pair. The α- and ß-configurational isomers of Fapy·dG have distinct effects on Pol II insertion and extension. Pol II can preferentially cleave error-prone products by proofreading. To further understand the structural basis of transcription processing of Fapy·dG, five different structures were solved, including Fapy·dG template-loading state (apo), error-free cytidine triphosphate (CTP) binding state (prechemistry), error-prone ATP binding state (prechemistry), error-free Fapy·dG:C product state (postchemistry), and error-prone Fapy·dG:A product state (postchemistry), revealing distinctive nucleotide binding and product states. Taken together, our study provides a comprehensive mechanistic framework for better understanding how Fapy·dG lesions impact transcription and subsequent pathological consequences.
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Daño del ADN , Pirimidinas , ARN Polimerasa II , Humanos , ARN Polimerasa II/metabolismo , 8-Hidroxi-2'-Desoxicoguanosina , Cinética , Mutagénesis , DesoxiguanosinaRESUMEN
Excess oxidative stress generated in the body causes various types of cellular damage, including DNA damage. Certain trace minerals act as antioxidants by functioning as cofactors for antioxidant enzymes. This study was conducted to evaluate the serum and hair concentrations of major antioxidant trace minerals (zinc, manganese, selenium, and chromium) and to determine the association between the oxidative stress marker urinary 8-hydroxy-2'-deoxyguanosine (8-OHdG) and serum or hair antioxidant trace mineral concentrations, according to the general characteristics of healthy adults. Study participants were selected after screening, and 108 participants aged 19-69 years were finally included. Serum and hair trace mineral concentrations were analyzed using inductively coupled plasma mass spectrometry, and urine 8-OHdG levels were quantified using an ELISA kit. Results showed that urinary 8-OHdG levels were significantly higher in exercisers than in those who did not exercise. Correlation analysis revealed that urinary 8-OHdG was negatively correlated with hair zinc in participants over 60 years of age and with poor health status, and positively correlated with hair chromium in participants with irregular dietary habits. In conclusion, these results suggest that urinary 8-OHdG is particularly correlated with hair zinc and chromium levels. Additional large-scale epidemiological studies are needed to generally confirm these findings.
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Selenio , Oligoelementos , Adulto , Humanos , Persona de Mediana Edad , Anciano , Antioxidantes/metabolismo , Oligoelementos/análisis , Estudios Transversales , Estrés Oxidativo , Selenio/metabolismo , Zinc/metabolismo , 8-Hidroxi-2'-Desoxicoguanosina/metabolismo , Cromo/metabolismo , Cabello/química , Desoxiguanosina/metabolismoRESUMEN
In recent years, research has shown that oxidative stress plays a significant role in chronic inflammatory conditions. The alteration of the oxidant/antioxidant balance leads to the appearance of free radicals, important molecules involved in both diabetes mellitus and periodontal disease. Diabetes is considered to be one of the major risk factors of periodontal disease and the inflammation characterizing this condition is associated with oxidative stress, implicitly resulting in oxidative damage to DNA. 8-Hydroxydeoxyguanosine (8-OHdG) is the most common stable product of oxidative DNA damage caused by reactive oxygen species, and its levels have been reported to increase in body fluids and tissues during inflammatory conditions. 8-OHdG emerges as a pivotal biomarker for assessing oxidative DNA damage, demonstrating its relevance across diverse health conditions, including neurodegenerative disorders, cancers, inflammatory conditions, and periodontal disease. Continued research in this field is crucial for developing more precise treatments and understanding the detailed link between oxidative stress and the progression of periodontitis. The use of the 8-OHdG biomarker in assessing and managing chronic periodontitis is an area of increased interest in dental research, with the potential to provide crucial information for diagnosis and treatment.
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Periodontitis Crónica , Diabetes Mellitus , Humanos , 8-Hidroxi-2'-Desoxicoguanosina , Desoxiguanosina , Saliva/metabolismo , Biomarcadores/metabolismo , Estrés Oxidativo , Diabetes Mellitus/diagnóstico , Daño del ADNRESUMEN
Unspliced HIV-1 RNAs function as messenger RNAs for Gag or Gag-Pol polyproteins and progeny genomes packaged into virus particles. Recently, it has been reported that fate of the RNAs might be primarily determined, depending on transcriptional initiation sites among three consecutive deoxyguanosine residues (GGG tract) downstream of TATA-box in the 5' long terminal repeat (LTR). Although HIV-1 RNA transcription starts mostly from the first deoxyguanosine of the GGG tract and often from the second or third deoxyguanosine, RNAs beginning with one guanosine (G1-form RNAs), whose transcription initiates from the third deoxyguanosine, were predominant in HIV-1 particles. Despite selective packaging of G1-form RNAs into virus particles, its biological impact during viral replication remains to be determined. In this study, we revealed that G1-form RNAs are primarily selected as a template for provirus DNA rather than other RNAs. In competitions between HIV-1 and lentiviral vector transcripts in virus-producing cells, approximately 80% of infectious particles were found to generate provirus using HIV-1 transcripts, while lentiviral vector transcripts were conversely selected when we used HIV-1 mutants in which the third deoxyguanosine in the GGG tract was replaced with deoxythymidine or deoxycytidine (GGT or GGC mutants, respectively). In the other analyses of proviral sequences after infection with an HIV-1 mutant in which the GGG tract in 3' LTR was replaced with TTT, most proviral sequences of the GGG-tract region in 5' LTR were found to be TTG, which is reasonably generated using the G1-form transcripts. Our results indicate that the G1-form RNAs serve as a dominant genome to establish provirus DNA.IMPORTANCESince the promoter for transcribing HIV-1 RNA is unique, all viral elements including genomic RNA and viral proteins have to be generated by the unique transcripts through ingenious mechanisms including RNA splicing and frameshifting during protein translation. Previous studies suggested a new mechanism for diversification of HIV-1 RNA functions by heterogeneous transcriptional initiation site usage; HIV-1 RNAs whose transcription initiates from a certain nucleotide were predominant in virus particles. In this study, we established two methods to analyze heterogenous transcriptional initiation site usage by HIV-1 during viral infection and showed that RNAs beginning with one guanosine (G1-form RNAs), whose transcription initiates from the third deoxyguanosine of the GGG tract in 5' LTR, were primarily selected as viral genome in infectious particles and thus are used as a template to generate provirus for continuous replication. This study provides insights into the mechanism for diversification of unspliced RNA functions and requisites of lentivirus infectivity.
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VIH-1 , Provirus , Desoxiguanosina/genética , Guanosina/genética , Duplicado del Terminal Largo de VIH/genética , VIH-1/fisiología , Provirus/genética , ARN Viral/genética , Secuencias Repetidas TerminalesRESUMEN
There are few effective treatments for small cell lung cancer (SCLC) underscoring the need for innovative therapeutic approaches. This study focuses on exploiting telomerase, a critical SCLC dependency as a therapeutic target. A prominent characteristic of SCLC is their reliance on telomerase activity, a key enzyme essential for their continuous proliferation. Here we utilize a nucleoside analog, 6-Thio-2'-deoxyguanosine (6TdG) currently in phase II clinical trials, that is preferentially incorporated by telomerase into telomeres leading to telomere dysfunction. Using preclinical mouse and human derived models we find low intermittent doses of 6TdG inhibit tumor growth and reduce metastatic burden. Anti-tumor efficacy correlates with a reduction in a subpopulation of cancer initiating like cells (CICs) identified by their expression of L1CAM/CD133 and highest telomerase activity. 6TdG treatment also leads to activation of innate and adaptive anti-tumor responses. Mechanistically, 6TdG depletes CICs and induces type-I interferon signaling leading to tumor immune visibility by activating tumor cell STING signaling. We also observe increased sensitivity to irradiation after 6TdG treatment in both syngeneic and humanized SCLC xenograft models both of which are dependent on the presence of host immune cells. This study underscores the immune-enhancing and metastasis-reducing effects of 6TdG, employing a range of complementary in vitro and in vivo SCLC preclinical models providing a potential therapeutic approach to SCLC.
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Desoxiguanosina/análogos & derivados , Neoplasias Pulmonares , Carcinoma Pulmonar de Células Pequeñas , Telomerasa , Tionucleósidos , Humanos , Animales , Ratones , Carcinoma Pulmonar de Células Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Sistemas de Liberación de Medicamentos , TelómeroRESUMEN
One of the most distinguished features in biological effects of heavy ions would be the decrease of oxygen effect in the high-LET region. This feature has been referred to as the radiobiological basis for the control of hypoxic fraction in cancer radiotherapy. However, mechanisms to explain this phenomenon have not been fully understood. One of the explanations was given by the oxygen in the track hypothesis, which proposes that oxygen is produced along ion tracks even in the hypoxic irradiation condition. In the present study, we designed an experimental approach to support this hypothesis by using 8-hydroxy-2'-deoxyguanosine (8-OHdG) as DNA damage requiring oxygen to produce. The LET dependence of 8-OHdG under hypoxic condition revealed that with increasing LET 8-OHdG yield seems to increase, despite that the yield of OH radical, which is also required for the production of 8-OHdG, decreases in the high-LET region. This result is consistent with the explanation that the local generation of oxygen along ion tracks contributes to the increase of 8-OHdG yield.