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1.
Invest Ophthalmol Vis Sci ; 65(6): 7, 2024 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-38833258

RESUMO

Purpose: The purpose of this study was to analyze the extent of DNA breaks in primary uveal melanoma (UM) with regard to radiotherapy dose delivery (single-dose versus fractionated) and monosomy 3 status. Methods: A total of 54 patients with UM were included. Stereotactic radiotherapy (SRT) was performed in 23 patients, with 8 undergoing single-dose SRT (sdSRT) treatment and 15 receiving fractionated SRT (fSRT). DNA breaks in the enucleated or endoresected tumors were visualized by a TUNEL assay and quantified by measuring the TUNEL-positive area. Protein expression was analyzed by immunohistochemistry. Co-detection of chromosome 3 with proteins was performed by immuno-fluorescent in situ hybridization. Results: The amount of DNA breaks in the total irradiated group was increased by 2.7-fold (P < 0.001) compared to non-irradiated tissue. Tumors treated with fSRT were affected more severely, showing 2.1-fold more DNA damage (P = 0.007) compared to the cases after single (high) dose irradiation (sdSRT). Monosomy 3 tumors showed less DNA breaks compared to disomy 3 samples (P = 0.004). The presence of metastases after radiotherapy correlated with monosomy 3 and less DNA breaks compared to patients with non-metastatic cancer in the combined group with fSRT and sdSRT (P < 0.05). Conclusions: Fractionated irradiation led to more DNA damage than single-dose treatment in primary UM. As tumors with monosomy 3 showed less DNA breaks than those with disomy 3, this may indicate that they are less radiosensitive, which may influence the efficacy of irradiation.


Assuntos
Cromossomos Humanos Par 3 , Dano ao DNA , Melanoma , Neoplasias Uveais , Humanos , Neoplasias Uveais/radioterapia , Neoplasias Uveais/genética , Melanoma/radioterapia , Melanoma/genética , Feminino , Cromossomos Humanos Par 3/genética , Masculino , Pessoa de Meia-Idade , Idoso , Adulto , Idoso de 80 Anos ou mais , Hibridização in Situ Fluorescente , Marcação In Situ das Extremidades Cortadas , Dosagem Radioterapêutica , Imuno-Histoquímica , Radiocirurgia/efeitos adversos , Radiocirurgia/métodos , Relação Dose-Resposta à Radiação
2.
Front Endocrinol (Lausanne) ; 15: 1393111, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38846492

RESUMO

Non-obstructive azoospermia (NOA) is a disease characterized by spermatogenesis failure and comprises phenotypes such as hypospermatogenesis, mature arrest, and Sertoli cell-only syndrome. Studies have shown that FA cross-linked anemia (FA) pathway is closely related to the occurrence of NOA. There are FA gene mutations in male NOA patients, which cause significant damage to male germ cells. The FA pathway is activated in the presence of DNA interstrand cross-links; the key step in activating this pathway is the mono-ubiquitination of the FANCD2-FANCI complex, and the activation of the FA pathway can repair DNA damage such as DNA double-strand breaks. Therefore, we believe that the FA pathway affects germ cells during DNA damage repair, resulting in minimal or even disappearance of mature sperm in males. This review summarizes the regulatory mechanisms of FA-related genes in male azoospermia, with the aim of providing a theoretical reference for clinical research and exploration of related genes.


Assuntos
Azoospermia , Anemia de Fanconi , Transdução de Sinais , Humanos , Azoospermia/genética , Azoospermia/metabolismo , Azoospermia/patologia , Masculino , Anemia de Fanconi/metabolismo , Anemia de Fanconi/genética , Anemia de Fanconi/patologia , Reparo do DNA , Animais , Proteínas de Grupos de Complementação da Anemia de Fanconi/metabolismo , Proteínas de Grupos de Complementação da Anemia de Fanconi/genética , Dano ao DNA , Espermatogênese
3.
J Cancer Res Clin Oncol ; 150(6): 299, 2024 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-38850382

RESUMO

BACKGROUND: Microvesicles are membraned particles produced by different types of cells recently investigated for anticancer purposes. The current study aimed to investigate the effects of human bone marrow mesenchymal stem cell-derived microvesicles (BMSC-MVs) on the multiple myeloma cell line U266. BMSC-MVs were isolated from BMSCs via ultracentrifugation and characterized using transmission electron microscopy (TEM) and dynamic light scattering (DLS). U266 cells were treated with 15, 30, 60, and 120 µg/mL BMSC-MVs for three and seven days and the effects of treatment in terms of viability, cytotoxicity, and DNA damage were investigated via the MTT assay, lactate dehydrogenase (LDH) assay, and 8­hydroxy-2'-deoxyguanosine (8­OHdG) measurement, respectively. Moreover, the apoptosis rate of the U266 cells treated with 60 µg/mL BMSC-MVs was also assessed seven days following treatment via flow cytometry. Ultimately, the expression level of BCL2, BAX, and CCND1 by the U266 cells was examined seven days following treatment with 60 µg/mL BMSC-MVs using qRT-PCR. RESULTS: BMSC-MVs had an average size of ~ 410 nm. According to the MTT and LDH assays, BMSC-MV treatment reduced the U266 cell viability and mediated cytotoxic effects against them, respectively. Moreover, elevated 8­OHdG levels following BMSC-MV treatment demonstrated a dose-dependent increase of DNA damage in the treated cells. BMSC-MV-treated U266 cells also exhibited an increased apoptosis rate after seven days of treatment. The expression level of BCL2 and CCND1 decreased in the treated cells whereas the BAX expression demonstrated an incremental pattern. CONCLUSIONS: Our findings accentuate the therapeutic benefit of BMSC-MVs against the multiple myeloma cell line U266 and demonstrate how microvesicles could be of therapeutic advantage. Future in vivo studies could further corroborate these findings.


Assuntos
Apoptose , Micropartículas Derivadas de Células , Células-Tronco Mesenquimais , Mieloma Múltiplo , Humanos , Mieloma Múltiplo/patologia , Mieloma Múltiplo/metabolismo , Células-Tronco Mesenquimais/metabolismo , Linhagem Celular Tumoral , Micropartículas Derivadas de Células/metabolismo , Sobrevivência Celular , Dano ao DNA
4.
Mol Biol Rep ; 51(1): 725, 2024 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-38851636

RESUMO

Exposure to UV affects the development and growth of a wide range of organisms. Nowadays, researchers are focusing on the impact of UV radiation and its underlying molecular mechanisms, as well as devising strategies to mitigate its harmful effects. Different forms of UV radiation, their typical exposure effects, the impact of UV on DNA integrity, and the deterioration of genetic material are discussed in this review; furthermore, we also review the effects of UV radiation that affect the biological functions of the organisms. Subsequently, we address the processes that aid organisms in navigating the damage in genetic material, neuroinflammation, and neurodegeneration brought on by UV-mediated double-strand breaks. To emphasize the molecular pathways, we conclude the review by going over the animal model studies that highlight the genes and proteins that are impacted by UV radiation.


Assuntos
Quebras de DNA de Cadeia Dupla , Doenças Neurodegenerativas , Doenças Neuroinflamatórias , Raios Ultravioleta , Raios Ultravioleta/efeitos adversos , Doenças Neurodegenerativas/etiologia , Doenças Neurodegenerativas/genética , Animais , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Humanos , Doenças Neuroinflamatórias/etiologia , Reparo do DNA/genética , Dano ao DNA/efeitos da radiação
5.
Mol Brain ; 17(1): 32, 2024 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-38840222

RESUMO

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that affects the motor neuron. One aspect of the neuropathology involved in ALS includes increased genomic damage and impaired DNA repair capability. The TAR-DNA binding protein 43 (TDP43) has been associated with both sporadic and familial forms of ALS, and is typically observed as cytosolic mislocalization of protein aggregates, termed TDP43 proteinopathy. TDP43 is a ubiquitous RNA/DNA binding protein with functional implications in a wide range of disease processes, including the repair of DNA double-strand breaks (DSBs). While TDP43 is widely known to regulate RNA metabolism, our lab has reported it also functions directly at the protein level to facilitate DNA repair. Here, we show that the TDP43 protein interacts with DNA mismatch repair (MMR) proteins MLH1 and MSH6 in a DNA damage-inducible manner. We utilized differentiated SH-SY5Y neuronal cultures to identify this inducible relationship using complementary approaches of proximity ligation assay (PLA) and co-immunoprecipitation (CoIP) assay. We observed that signals of TDP43 interaction with MLH1 and MSH6 increased significantly following a 2 h treatment of 10 µM methylmethanesulfonate (MMS), a DNA alkylating agent used to induce MMR repair. Likewise, we observed this effect was abolished in cell lines treated with siRNA directed against TDP43. Finally, we demonstrated these protein interactions were significantly increased in lumbar spinal cord samples of ALS-affected patients compared to age-matched controls. These results will inform our future studies to understand the mechanisms and consequences of this TDP43-MMR interaction in the context of ALS-affected neurons.


Assuntos
Dano ao DNA , Proteínas de Ligação a DNA , Proteína 1 Homóloga a MutL , Ligação Proteica , Humanos , Proteínas de Ligação a DNA/metabolismo , Proteína 1 Homóloga a MutL/metabolismo , Ligação Proteica/efeitos dos fármacos , Linhagem Celular Tumoral , Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Neurônios/metabolismo , Pessoa de Meia-Idade , Masculino
6.
J Exp Clin Cancer Res ; 43(1): 159, 2024 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-38840237

RESUMO

BACKGROUND: Renal cell carcinoma (RCC) was historically considered to be less responsive to radiation therapy (RT) compared to other cancer indications. However, advancements in precision high-dose radiation delivery through single-fraction and multi-fraction stereotactic ablative radiotherapy (SABR) have led to better outcomes and reduced treatment-related toxicities, sparking renewed interest in using RT to treat RCC. Moreover, numerous studies have revealed that certain therapeutic agents including chemotherapies can increase the sensitivity of tumors to RT, leading to a growing interest in combining these treatments. Here, we developed a rational combination of two radiosensitizers in a tumor-targeted liposomal formulation for augmenting RT in RCC. The objective of this study is to assess the efficacy of a tumor-targeted liposomal formulation combining the mTOR inhibitor everolimus (E) with the survivin inhibitor YM155 (Y) in enhancing the sensitivity of RCC tumors to radiation. EXPERIMENTAL DESIGN: We slightly modified our previously published tumor-targeted liposomal formulation to develop a rational combination of E and Y in a single liposomal formulation (EY-L) and assessed its efficacy in RCC cell lines in vitro and in RCC tumors in vivo. We further investigated how well EY-L sensitizes RCC cell lines and tumors toward radiation and explored the underlying mechanism of radiosensitization. RESULTS: EY-L outperformed the corresponding single drug-loaded formulations E-L and Y-L in terms of containing primary tumor growth and improving survival in an immunocompetent syngeneic mouse model of RCC. EY-L also exhibited significantly higher sensitization of RCC cells towards radiation in vitro than E-L and Y-L. Additionally, EY-L sensitized RCC tumors towards radiation therapy in xenograft and murine RCC models. EY-L mediated induction of mitotic catastrophe via downregulation of multiple cell cycle checkpoints and DNA damage repair pathways could be responsible for the augmentation of radiation therapy. CONCLUSION: Taken together, our study demonstrated the efficacy of a strategic combination therapy in sensitizing RCC to radiation therapy via inhibition of DNA damage repair and a substantial increase in mitotic catastrophe. This combination therapy may find its use in the augmentation of radiation therapy during the treatment of RCC patients.


Assuntos
Carcinoma de Células Renais , Reparo do DNA , Neoplasias Renais , Survivina , Serina-Treonina Quinases TOR , Ensaios Antitumorais Modelo de Xenoenxerto , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/radioterapia , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/metabolismo , Animais , Survivina/metabolismo , Humanos , Camundongos , Linhagem Celular Tumoral , Neoplasias Renais/patologia , Neoplasias Renais/radioterapia , Neoplasias Renais/tratamento farmacológico , Reparo do DNA/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo , Mitose/efeitos dos fármacos , Mitose/efeitos da radiação , Imidazóis/farmacologia , Dano ao DNA , Everolimo/farmacologia , Naftoquinonas/farmacologia , Naftoquinonas/uso terapêutico , Radiossensibilizantes/farmacologia , Radiossensibilizantes/uso terapêutico , Lipossomos/farmacologia , Inibidores de MTOR/farmacologia , Inibidores de MTOR/uso terapêutico
7.
Life Sci Alliance ; 7(8)2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38843934

RESUMO

RNA-binding proteins are frequently deregulated in cancer and emerge as effectors of the DNA damage response (DDR). The non-POU domain-containing octamer-binding protein NONO/p54nrb is a multifunctional RNA-binding protein that not only modulates the production and processing of mRNA, but also promotes the repair of DNA double-strand breaks (DSBs). Here, we investigate the impact of Nono deletion in the murine KP (KRas G12D , Trp53 -/- ) cell-based lung cancer model. We show that the deletion of Nono impairs the response to DNA damage induced by the topoisomerase II inhibitor etoposide or the radiomimetic drug bleomycin. Nono-deficient KP (KPN) cells display hyperactivation of DSB signalling and high levels of DSBs. The defects in the DDR are accompanied by reduced RNA polymerase II promoter occupancy, impaired nascent RNA synthesis, and attenuated induction of the DDR factor growth arrest and DNA damage-inducible beta (Gadd45b). Our data characterise Gadd45b as a putative Nono-dependent effector of the DDR and suggest that Nono mediates a genome-protective crosstalk of the DDR with the RNA metabolism via induction of Gadd45b.


Assuntos
Dano ao DNA , Reparo do DNA , Proteínas de Ligação a RNA , Animais , Camundongos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Quebras de DNA de Cadeia Dupla , Antígenos de Diferenciação/metabolismo , Antígenos de Diferenciação/genética , Bleomicina/farmacologia , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Etoposídeo/farmacologia , Transdução de Sinais , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/genética , Linhagem Celular Tumoral , RNA Polimerase II/metabolismo , Humanos , Proteínas GADD45
8.
Sci Rep ; 14(1): 13079, 2024 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-38844507

RESUMO

As patient exposure to ionizing radiation from medical imaging and its risks are continuing issues, this study aimed to evaluate DNA damage and repair markers after myocardial perfusion single-photon emission computed tomography (MPS). Thirty-two patients undergoing Tc-99m sestamibi MPS were studied. Peripheral blood was collected before radiotracer injection at rest and 60-90 min after injection. The comet assay (single-cell gel electrophoresis) was performed with peripheral blood cells to detect DNA strand breaks. Three descriptors were evaluated: the percentage of DNA in the comet tail, tail length, and tail moment (the product of DNA tail percentage and tail length). Quantitative PCR (qPCR) was performed to evaluate the expression of five genes related to signaling pathways in response to DNA damage and repair (ATM, ATR, BRCA1, CDKN1A, and XPC). Mann-Whitney's test was employed for statistical analysis; p < 0.05 was considered significant. Mean Tc-99m sestamibi dose was 15.1 mCi. After radiotracer injection, comparing post-exposure to pre-exposure samples of each of the 32 patients, no statistically significant differences of the DNA percentage in the tail, tail length or tail moment were found. qPCR revealed increased expression of BRCA1 and XPC, without any significant difference regarding the other genes. No significant increase in DNA strand breaks was detected after a single radiotracer injection for MPS. There was activation of only two repair genes, which may indicate that, in the current patient sample, the effects of ionizing radiation on the DNA were not large enough to trigger intense repair responses, suggesting the absence of significant DNA damage.


Assuntos
Dano ao DNA , Reparo do DNA , Tomografia Computadorizada de Emissão de Fóton Único , Humanos , Feminino , Masculino , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Reparo do DNA/genética , Pessoa de Meia-Idade , Idoso , Tecnécio Tc 99m Sestamibi , Imagem de Perfusão do Miocárdio/métodos , Proteína BRCA1/genética , Ensaio Cometa
9.
Sci Rep ; 14(1): 13015, 2024 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-38844752

RESUMO

Calcium hydroxide (Ca(OH)2NPs), calcium titanate (CaTiO3NPs) and yttrium oxide (Y2O3NPs) nanoparticles are prevalent in many industries, including food and medicine, but their small size raises concerns about potential cellular damage and genotoxic effects. However, there are very limited studies available on their genotoxic effects. Hence, this was done to investigate the effects of multiple administration of Ca(OH)2NPs, CaTiO3NPs or/and Y2O3NPs on genomic DNA stability, mitochondrial membrane potential integrity and inflammation induction in mouse brain tissues. Mice were orally administered Ca(OH)2NPs, CaTiO3NPs or/and Y2O3NPs at a dose level of 50 mg/kg b.w three times a week for 2 weeks. Genomic DNA integrity was studied using Comet assay and the level of reactive oxygen species (ROS) within brain cells was analyzed using 2,7 dichlorofluorescein diacetate dye. The expression level of Presenilin-1, tumor necrosis factor-alpha (TNF-α) and Interleukin-6 (IL-6) genes and the integrity of the mitochondrial membrane potential were also detected. Oral administration of Ca(OH)2NPs caused the highest damage to genomic DNA and mitochondrial membrane potential, less genomic DNA and mitochondrial damage was induced by CaTiO3NPs administration while administration of Y2O3NPs did not cause any remarkable change in the integrity of genomic DNA and mitochondrial membrane potential. Highest ROS generation and upregulation of presenilin-1, TNF-α and IL-6 genes were also observed within the brain cells of mice administrated Ca(OH)2NPs but Y2O3NPs administration almost caused no changes in ROS generation and genes expression compared to the negative control. Administration of CaTiO3NPs alone slightly increased ROS generation and the expression level of TNF-α and IL-6 genes. Moreover, no remarkable changes in the integrity of genomic DNA and mitochondrial DNA potential, ROS level and the expression level of presenilin-1, TNF-α and IL-6 genes were noticed after simultaneous coadministration of Y2O3NPs with Ca(OH)2NPs and CaTiO3NPs. Coadministration of Y2O3NPs with Ca(OH)2NPs and CaTiO3NPs mitigated Ca(OH)2NPs and CaTiO3NPs induced ROS generation, genomic DNA damage and inflammation along with restoring the integrity of mitochondrial membrane potential through Y2O3NPs scavenging free radicals ability. Therefore, further studies are recommended to study the possibility of using Y2O3NPs to alleviate Ca(OH)2NPs and CaTiO3NPs induced genotoxic effects.


Assuntos
Hidróxido de Cálcio , Dano ao DNA , Inflamação , Potencial da Membrana Mitocondrial , Nanopartículas , Espécies Reativas de Oxigênio , Titânio , Ítrio , Animais , Espécies Reativas de Oxigênio/metabolismo , Camundongos , Dano ao DNA/efeitos dos fármacos , Hidróxido de Cálcio/farmacologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Titânio/química , Titânio/toxicidade , Inflamação/metabolismo , Inflamação/patologia , Ítrio/química , Nanopartículas/química , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Masculino , Encéfalo/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , DNA Mitocondrial/metabolismo
11.
Nat Commun ; 15(1): 4696, 2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38824133

RESUMO

Age-related microangiopathy, also known as small vessel disease (SVD), causes damage to the brain, retina, liver, and kidney. Based on the DNA damage theory of aging, we reasoned that genomic instability may underlie an SVD caused by dominant C-terminal variants in TREX1, the most abundant 3'-5' DNA exonuclease in mammals. C-terminal TREX1 variants cause an adult-onset SVD known as retinal vasculopathy with cerebral leukoencephalopathy (RVCL or RVCL-S). In RVCL, an aberrant, C-terminally truncated TREX1 mislocalizes to the nucleus due to deletion of its ER-anchoring domain. Since RVCL pathology mimics that of radiation injury, we reasoned that nuclear TREX1 would cause DNA damage. Here, we show that RVCL-associated TREX1 variants trigger DNA damage in humans, mice, and Drosophila, and that cells expressing RVCL mutant TREX1 are more vulnerable to DNA damage induced by chemotherapy and cytokines that up-regulate TREX1, leading to depletion of TREX1-high cells in RVCL mice. RVCL-associated TREX1 mutants inhibit homology-directed repair (HDR), causing DNA deletions and vulnerablility to PARP inhibitors. In women with RVCL, we observe early-onset breast cancer, similar to patients with BRCA1/2 variants. Our results provide a mechanistic basis linking aberrant TREX1 activity to the DNA damage theory of aging, premature senescence, and microvascular disease.


Assuntos
Dano ao DNA , Exodesoxirribonucleases , Fosfoproteínas , Animais , Exodesoxirribonucleases/genética , Exodesoxirribonucleases/metabolismo , Humanos , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Camundongos , Reparo de DNA por Recombinação , Fenótipo , Mutação , Drosophila/genética , Envelhecimento/genética , Envelhecimento/metabolismo , Feminino , Drosophila melanogaster/genética , Masculino , Doenças Retinianas , Doenças Vasculares , Doenças Desmielinizantes Hereditárias do Sistema Nervoso Central
12.
Sci Adv ; 10(23): eadm9589, 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38838142

RESUMO

DNA replication stress (RS) is a widespread phenomenon in carcinogenesis, causing genomic instability and extensive chromatin alterations. DNA damage leads to activation of innate immune signaling, but little is known about transcriptional regulators mediating such signaling upon RS. Using a chemical screen, we identified protein arginine methyltransferase 5 (PRMT5) as a key mediator of RS-dependent induction of interferon-stimulated genes (ISGs). This response is also associated with reactivation of endogenous retroviruses (ERVs). Using quantitative mass spectrometry, we identify proteins with PRMT5-dependent symmetric dimethylarginine (SDMA) modification induced upon RS. Among these, we show that PRMT5 targets and modulates the activity of ZNF326, a zinc finger protein essential for ISG response. Our data demonstrate a role for PRMT5-mediated SDMA in the context of RS-induced transcriptional induction, affecting physiological homeostasis and cancer therapy.


Assuntos
Replicação do DNA , Imunidade Inata , Proteína-Arginina N-Metiltransferases , Proteína-Arginina N-Metiltransferases/metabolismo , Proteína-Arginina N-Metiltransferases/genética , Humanos , Transdução de Sinais , Arginina/metabolismo , Arginina/análogos & derivados , Estresse Fisiológico , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Dano ao DNA , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética
14.
Syst Biol Reprod Med ; 70(1): 124-130, 2024 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38833571

RESUMO

Many couples in contemporary societies suffer from infertility of unexplained origins (idiopathic). A promising treatment strategy within this context involves the administration to women of preparations containing lactic acid bacteria (Lactobacillus) and/or their metabolites. Recent investigations underscore the role of lactobacilli in sustaining female fertility and enhancing the effectiveness of assisted reproductive techniques. There have also been reports describing the effect of lactobacilli on sperm functions, but our knowledge in this domain remains uncertain. In this study, the effect of supernatant from Lactobacillus rhamnosus culture on mouse sperm viability and motility was tested. The protective properties of lactobacilli metabolites against hydrogen peroxide-induced DNA damage were also verified. It was shown that the metabolites have no effect on viability, motility, and genome integrity of spermatozoa, but in excessive concentrations they become toxic. The obtained results imply that probiotic and/or postbiotic preparations taken by women should not adversely affect the sperm of their partners, provided the dose is correctly selected.


Assuntos
Lacticaseibacillus rhamnosus , Motilidade dos Espermatozoides , Espermatozoides , Animais , Masculino , Espermatozoides/metabolismo , Espermatozoides/efeitos dos fármacos , Camundongos , Motilidade dos Espermatozoides/efeitos dos fármacos , Dano ao DNA , Probióticos , Sobrevivência Celular/efeitos dos fármacos , Lactobacillus
15.
Nat Commun ; 15(1): 4716, 2024 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-38830843

RESUMO

BRCA2 is a tumor suppressor protein responsible for safeguarding the cellular genome from replication stress and genotoxicity, but the specific mechanism(s) by which this is achieved to prevent early oncogenesis remains unclear. Here, we provide evidence that BRCA2 acts as a critical suppressor of head-on transcription-replication conflicts (HO-TRCs). Using Okazaki-fragment sequencing (Ok-seq) and computational analysis, we identified origins (dormant origins) that are activated near the transcription termination sites (TTS) of highly expressed, long genes in response to replication stress. Dormant origins are a source for HO-TRCs, and drug treatments that inhibit dormant origin firing led to a reduction in HO-TRCs, R-loop formation, and DNA damage. Using super-resolution microscopy, we showed that HO-TRC events track with elongating RNA polymerase II, but not with transcription initiation. Importantly, RNase H2 is recruited to sites of HO-TRCs in a BRCA2-dependent manner to help alleviate toxic R-loops associated with HO-TRCs. Collectively, our results provide a mechanistic basis for how BRCA2 shields against genomic instability by preventing HO-TRCs through both direct and indirect means occurring at predetermined genomic sites based on the pre-cancer transcriptome.


Assuntos
Proteína BRCA2 , Replicação do DNA , RNA Polimerase II , Ribonuclease H , Humanos , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Ribonuclease H/metabolismo , Ribonuclease H/genética , RNA Polimerase II/metabolismo , Transcrição Gênica , Terminação da Transcrição Genética , Dano ao DNA , Origem de Replicação , Estruturas R-Loop , Linhagem Celular Tumoral
16.
RNA Biol ; 21(1): 1-11, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38832821

RESUMO

LncRNA is a group of transcripts with a length exceeding 200 nucleotides that contribute to tumour development. Our research group found that LINC00052 expression was repressed during the formation of breast cancer (BC) multicellular spheroids. Intriguingly, LINC00052 precise role in BC remains uncertain. We explored LINC00052 expression in BC patients` RNA samples (TCGA) in silico, as well as in an in-house patient cohort, and inferred its cellular and molecular mechanisms. In vitro studies evaluated LINC00052 relevance in BC cells viability, cell cycle and DNA damage. Results. Bioinformatic RNAseq analysis of BC patients showed that LINC00052 is overexpressed in samples from all BC molecular subtypes. A similar LINC00052 expression pattern was observed in an in-house patient cohort. In addition, higher LINC00052 levels are related to better BC patient´s overall survival. Remarkably, MCF-7 and ZR-75-1 cells treated with estradiol showed increased LINC00052 expression compared to control, while these changes were not observed in MDA-MB-231 cells. In parallel, bioinformatic analyses indicated that LINC00052 influences DNA damage and cell cycle. MCF-7 cells with low LINC00052 levels exhibited increased cellular protection against DNA damage and diminished growth capacity. Furthermore, in cisplatin-resistant MCF-7 cells, LINC00052 expression was downregulated. Conclusion. This work shows that LINC00052 expression is associated with better BC patient survival. Remarkably, LINC00052 expression can be regulated by Estradiol. Additionally, assays suggest that LINC00052 could modulate MCF-7 cells growth and DNA damage repair. Overall, this study highlights the need for further research to unravel LINC00052 molecular mechanisms and potential clinical applications in BC.


Assuntos
Neoplasias da Mama , Biologia Computacional , Dano ao DNA , Regulação Neoplásica da Expressão Gênica , RNA Longo não Codificante , Humanos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Biologia Computacional/métodos , RNA Longo não Codificante/genética , Feminino , Ciclo Celular/genética , Proliferação de Células , Linhagem Celular Tumoral , Células MCF-7 , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Sobrevivência Celular/genética , Prognóstico , Perfilação da Expressão Gênica
17.
Proc Natl Acad Sci U S A ; 121(24): e2404383121, 2024 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-38843184

RESUMO

Transcription is extremely important for cellular processes but can be hindered by RNA polymerase II (RNAPII) pausing and stalling. Cockayne syndrome protein B (CSB) promotes the progression of paused RNAPII or initiates transcription-coupled nucleotide excision repair (TC-NER) to remove stalled RNAPII. However, the specific mechanism by which CSB initiates TC-NER upon damage remains unclear. In this study, we identified the indispensable role of the ARK2N-CK2 complex in the CSB-mediated initiation of TC-NER. The ARK2N-CK2 complex is recruited to damage sites through CSB and then phosphorylates CSB. Phosphorylation of CSB enhances its binding to stalled RNAPII, prolonging the association of CSB with chromatin and promoting CSA-mediated ubiquitination of stalled RNAPII. Consistent with this finding, Ark2n-/- mice exhibit a phenotype resembling Cockayne syndrome. These findings shed light on the pivotal role of the ARK2N-CK2 complex in governing the fate of RNAPII through CSB, bridging a critical gap necessary for initiating TC-NER.


Assuntos
Síndrome de Cockayne , DNA Helicases , Enzimas Reparadoras do DNA , Reparo do DNA , Proteínas de Ligação a Poli-ADP-Ribose , RNA Polimerase II , Enzimas Reparadoras do DNA/metabolismo , Enzimas Reparadoras do DNA/genética , RNA Polimerase II/metabolismo , RNA Polimerase II/genética , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/genética , Humanos , Animais , Camundongos , DNA Helicases/metabolismo , DNA Helicases/genética , Síndrome de Cockayne/genética , Síndrome de Cockayne/metabolismo , Transcrição Gênica , Fosforilação , Caseína Quinase II/metabolismo , Caseína Quinase II/genética , Camundongos Knockout , Dano ao DNA , ATPases Associadas a Diversas Atividades Celulares/metabolismo , ATPases Associadas a Diversas Atividades Celulares/genética , Cromatina/metabolismo , Ubiquitinação , Reparo por Excisão
18.
Nat Commun ; 15(1): 3684, 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38693181

RESUMO

The metal-nucleic acid nanocomposites, first termed metal-nucleic acid frameworks (MNFs) in this work, show extraordinary potential as functional nanomaterials. However, thus far, realized MNFs face limitations including harsh synthesis conditions, instability, and non-targeting. Herein, we discover that longer oligonucleotides can enhance the synthesis efficiency and stability of MNFs by increasing oligonucleotide folding and entanglement probabilities during the reaction. Besides, longer oligonucleotides provide upgraded metal ions binding conditions, facilitating MNFs to load macromolecular protein drugs at room temperature. Furthermore, longer oligonucleotides facilitate functional expansion of nucleotide sequences, enabling disease-targeted MNFs. As a proof-of-concept, we build an interferon regulatory factor-1(IRF-1) loaded Ca2+/(aptamer-deoxyribozyme) MNF to target regulate glucose transporter (GLUT-1) expression in human epidermal growth factor receptor-2 (HER-2) positive gastric cancer cells. This MNF nanodevice disrupts GSH/ROS homeostasis, suppresses DNA repair, and augments ROS-mediated DNA damage therapy, with tumor inhibition rate up to 90%. Our work signifies a significant advancement towards an era of universal MNF application.


Assuntos
Aptâmeros de Nucleotídeos , DNA Catalítico , Neoplasias Gástricas , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Humanos , Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/metabolismo , Linhagem Celular Tumoral , DNA Catalítico/metabolismo , DNA Catalítico/química , Animais , Receptor ErbB-2/metabolismo , Fator Regulador 1 de Interferon/metabolismo , Fator Regulador 1 de Interferon/genética , Espécies Reativas de Oxigênio/metabolismo , Camundongos , Reparo do DNA , Dano ao DNA , Glutationa/metabolismo , Glutationa/química , Ácidos Nucleicos/metabolismo , Ácidos Nucleicos/química
20.
Phys Med Biol ; 69(10)2024 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-38700988

RESUMO

Liew and Mairani commented on our paper 'Modeling for predicting survival fraction of cells after ultra-high dose rate irradiation' (Shiraishiet al2024aPhys. Med. Biol.69015017), which proposed a biophysical model to predict the dose-response curve of surviving cell fractions after ultra-high dose rate irradiation following conventional dose rate irradiation by considering DNA damage yields. They suggested the need to consider oxygen concentration in our prediction model and possible issues related to the data selection process used for the benchmarking test in our paper. In this reply, we discuss the limitations of both the present model and the available experimental data for determining the model's parameters. We also demonstrate that our proposed model can reproduce the experimental survival data even when using only the experimental DNA damage data measured reliably under normoxic conditions.


Assuntos
Sobrevivência Celular , Dano ao DNA , Relação Dose-Resposta à Radiação , Modelos Biológicos , Sobrevivência Celular/efeitos da radiação , Doses de Radiação , Humanos , Oxigênio/metabolismo
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