Search Results (1,543)

Concerns about radiation exposure following the Fukushima Nuclear Power Plant accident continue to grow, and health risks associated with medical radiation have also become an important issue. Therefore, identifying agents that can mitigate radiation-related health effects is necessary. We focused on the antioxidant 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (tempol) and investigated its radioprotective mechanisms. HeLa and TIG-3 cells were irradiated with X-rays, γ-rays, or heavy-ion beams. The effect of tempol on reactive oxygen species (ROS) production was evaluated using fluorescence-activated cell sorting (FACS) analysis. DNA double-strand break (DSB) formation was assessed by γ-H2AX immunofluorescence staining. In mice, γ-H2AX formation in the thymus and duodenum were evaluated after acute or chronic γ-ray exposure. Inflammatory responses were analyzed through macrophage infiltration and TNF mRNA expression, while apoptosis was measured using Annexin V staining. Tempol suppressed ROS production and γ-H2AX foci formation following irradiation. It also reduced γ-H2AX induction in mouse tissues. Activated macrophage infiltration and TNF expression in the duodenum tended to decrease in tempol-treated mice, whereas apoptotic levels showed no significant differences. Notably, tempol more effectively inhibited γ-H2AX formation during chronic irradiation than acute exposure. These findings suggest that tempol mitigates radiation-induced inflammation and reduces DNA damage, supporting its potential as a radioprotective agent. Full article

Radiotherapy (RT) is a standard treatment for lung cancer; however, radiation-induced toxicities such as pneumonitis and fibrosis limit dose escalation and tumor control. Therefore, improved RT approaches are needed. This study investigated the radiation response of human ex vivo normal lung tissue using the three-dimensional EpiAlveolar™ model. Tissue models were irradiated with broad-beam (BB) and two spatially fractionated microbeam radiation therapy (MRT) dose metrics: equivalent uniform dose (MRT-EUD) and valley dose (MRT-valley). Our findings show that ex vivo lung tissue is able to tolerate peak doses of 36 Gy following MRT-EUD. On day 21, models effectively repaired significant DNA double-strand break (DSB) damage seen in the MRT-EUD-irradiated peak regions. In contrast, persistent unresolved DSBs were detected in MRT-valley-irradiated models 21 days post irradiation. Prolonged culture time resulted in cell loss and a reduction in epithelial cell layers. A significant upregulation of the pro-inflammatory cytokine IL6 was observed in both BB and MRT-EUD groups at 21 days. Fibrotic collagen deposition was detected in one BB-irradiated model but was absent in remaining BB- and MRT-treated tissues. Further investigation is required to clarify the potential and suitability of EpiAlveolar™ models for studying radiation-induced lung injury. Full article

Background/Objectives: Acute radiotherapy-induced skin toxicity is a common complication in breast cancer treatment, with marked interindividual variability not fully explained by clinical factors. This study investigated the contribution of germline mutations in DNA repair and tumor suppressor genes to acute radiodermatitis in a homogeneous cohort treated with hypofractionated intensity-modulated radiotherapy with inverse planning, with adjustment for potential lifestyle confounders. Methods: Mutations were grouped into four functional categories: homologous recombination repair (HRR), Fanconi anemia (FA), DNA damage response (DDR), and tumor suppressor (TS) genes. Ordinal logistic regression models adjusted for clinical covariates evaluated pooled and functional category-specific mutation effects. Results: Overall, any mutation significantly increased the risk of higher-grade acute radiodermatitis (OR = 2.24, p = 0.003), an effect driven primarily by HRR and FA mutations, as exclusion of these mutations rendered the association non-significant (OR = 1.785, p = 0.064). Functional category-based analyses showed that HRR (OR = 2.60, p = 0.002) and FA (OR = 2.62, p = 0.002) mutations were the strongest predictors, reflecting overlapping roles in double-strand break and interstrand crosslink repair. DDR and TS mutations showed no significant effect. Conclusions: These results highlight the key role of high-fidelity DNA repair in normal tissue radiosensitivity and demonstrate that functional genetic stratification has diagnostic value as a pre-treatment predictive biomarker framework, enabling identification of patients at increased risk of acute skin toxicity and supporting personalized radiotherapy planning. Full article

Poly(ADP-ribose) polymerase (PARP) inhibitors exploit defects in homologous recombination (HR) but show limited and heterogeneous efficacy in non-small-cell lung cancer (NSCLC), where canonical HR deficiency is uncommon. Identifying alternative molecular determinants that modulate PARP inhibitor sensitivity therefore remains an important objective. In this study, we examined the role of the NDR/Hippo-associated cofactor human MOB2 (hMOB2) in shaping PARP inhibitor responses in lung cancer cells. hMOB2 was depleted by siRNA in A549 and H1299 cell lines, and cell viability, long-term survival, DNA damage, and apoptosis were assessed using WST-1 assays, clonogenic assays, Western blotting, immunofluorescence, comet assays, and caspase-3 activity assays. p53 dependency was evaluated using p53-null H1299 cells and p53 reconstitution via retroviral transduction. hMOB2 depletion sensitized A549 cells to olaparib and rucaparib, resulting in a marked reduction in long-term clonogenic survival. This effect was associated with enhanced p53 phosphorylation, persistent γH2AX accumulation, increased DNA strand breaks, and caspase-3-dependent apoptosis, while hMOB2 loss alone was not intrinsically cytotoxic. Sensitization required functional p53, as it was absent in p53-null cells but restored upon p53 re-expression. These findings suggest that hMOB2 contributes to PARP inhibitor responses in lung cancer cells and underscore the complexity of PARP inhibitor sensitivity beyond classical HR deficiency. Full article

Background/Objectives: TP53 mutation or deletion status is important for determining cellular responses to DNA-damaging drugs. Oxaliplatin (OXA) is combined with the fluoropyrimidine (FP) drug 5-fluorouracil (5-FU) in the FOLFOX regimen used to treat advanced colorectal cancer (CRC). However, the effects of TP53 deletion on 5-FU + OXA synergy are not well known. We investigated potential synergy between OXA and 5-FU and compared it with OXA synergy with a novel polymeric FP, CF10, in four cell lines harboring either wild-type (WT) or TP53-null status. Methods: Using CompuSyn and the highest single agent (HSA) models, we compared synergy between CF10 and OXA (COXA) and between 5-FU and OXA (FOXA). Cell cycle analysis was performed, as was Western blot quantification of canonical DNA damage pathway proteins. Likewise, immunofluorescent and confocal analysis allowed us to compare topoisomerase 1 cleavage complex and double-strand DNA break formation. Results: COXA synergy displayed minimal TP53 dependence with greatly improved potency compared to FOXA. COXA synergy resulted from OXA increasing: (i) Topoisomerase 1 (Top1) cleavage complex formation; (ii) DNA double-strand breaks (DSBs), and (iii) Checkpoint Kinase 1 and 2 (p-Chk1/2) phosphorylation, consistent with increased replication stress. Additionally, increased S-phase entry in TP53-null cells enhanced synergy between CF10, 5-FU, and OXA as S-phase drugs. Conclusions: Our results demonstrate that OXA synergizes with CF10 more effectively than with 5-FU through enhanced replication stress in both WT and TP53-null cells by causing greater Top1-mediated DNA double-strand breaks. Our studies provide a foundation for further testing of this combination in an orthotopic liver metastatic setting and eventual clinical development. Full article

Natural compounds are increasingly recognized as valuable sources of pharmacologically active agents for cancer therapy. Among them, plant-derived triterpenoids attract attention due to their structural diversity and broad biological activity. Roburic acid (RA), a tetracyclic triterpenoid, has previously been shown to exert antiproliferative effects in colorectal cancer (CRC) cells with limited cytotoxicity. In the present study, we investigated the cellular mechanisms underlying RA activity in CRC cells, focusing on cell cycle regulation, mitochondrial function, apoptosis, oxidative stress, and DNA integrity. RA treatment markedly suppressed CRC cell proliferation, resulting in G₀/G₁ cell cycle arrest and downregulation of key proliferation markers. Mitochondrial analysis revealed an early reduction in mitochondrial membrane potential (MMP) following RA exposure, indicating mitochondrial dysfunction. Importantly, these effects occurred in the absence of intracellular reactive oxygen species (ROS) generation and without induction of DNA strand breaks, demonstrating a non-pro-oxidant and non-genotoxic profile of RA. Apoptotic features were observed mainly at higher concentrations and after prolonged exposure and were strongly dependent on cell line and assay type. Overall, RA limits CRC cell growth predominantly through cytostatic mechanisms, including cell cycle arrest and mitochondrial modulation, while apoptosis is a secondary, context-dependent response. The lack of oxidative stress and genotoxicity distinguishes RA from many conventional cytotoxic agents and supports its further investigation as a non-genotoxic anticancer compound. Full article

Eukaryotic DNA is wrapped around octamers of four core histones, forming nucleosomes. Histone post-translational modifications (PTMs) influence chromatin structure and the recruitment of regulatory factors, thereby affecting gene expression and DNA repair, including the response to DNA double-strand breaks (DSBs). Here, we describe a robust chromatin immunoprecipitation protocol combined with micrococcal nuclease digestion and DNA sequencing (MNase-ChIP-seq) to map histone modifications and their genome-wide distribution after the induction of a single DSB by the HO endonuclease in Saccharomyces cerevisiae. We validate the method by detecting changes in histone H3 methylation following HO transcriptional activation and DSB induction. This protocol enables reliable analysis of histone PTMs across mutant strains or stress conditions, supporting studies of chromatin dynamics in yeast. Full article

Background/Objectives: DNA polymerase ε (Pol ε), encoded by POLE1, plays a pivotal role in high-fidelity DNA replication and in coordinating DNA repair. While pathogenic exonuclease-domain variants are well established in cancer, biallelic POLE1 variants remain largely unexplored in non-malignant human cells. Methods: Here, we analyzed primary fibroblasts derived from a skin biopsy of a compound-heterozygous patient carrying two POLE1 variants. Western blot analysis confirmed detectable Pol ε protein levels, indicating preserved protein expression despite the underlying variants. Results: Nevertheless, functional alterations were observed across multiple independent assays. Compared with healthy control fibroblasts, this patient-derived Pol ε fibroblast line exhibited reduced clonogenic survival following ionizing radiation. Surviving fractions were consistently lower across radiation doses from 2 to 4 Gy, with an approximately twofold reduction at 2 Gy and progressively greater differences at higher doses. The isoeffect dose corresponding to 10% survival was reduced relative to pooled control fibroblasts. In addition, chromosomal breakage was increased, supporting altered processing of radiation-induced DNA damage in this cellular model. Live-cell imaging and senescence assays revealed delayed proliferation and an increased proportion of senescent or senescence-like cells under baseline and genotoxic stress conditions, including enhanced senescence-associated β-galactosidase activity. Flow-cytometric analysis demonstrated S phase accumulation and G2/M arrest, consistent with replication stress and cell-cycle perturbation. Immunofluorescence staining revealed increased γH2AX foci, consistent with persistent DNA double strand breaks. RAD51 foci formation was not reduced; instead, increased RAD51 recruitment was observed under combined cisplatin and irradiation treatment, arguing against a primary defect in RAD51-mediated homologous recombination. POLE1-variant fibroblasts also showed impaired proliferative recovery, reduced wound closure, increased γH2AX accumulation following cisplatin exposure, suggesting heightened susceptibility to DNA crosslinking stress. Conclusions: Collectively, these findings provide the first functional characterization of a patient-derived POLE1-variant fibroblast cell line and indicate that altered Pol ε function may influence cellular responses to genotoxic stress. While based on primary fibroblasts from a single compound-heterozygous patient, validation in additional patient-derived or isogenic models will be required to determine the broader relevance of these findings. Full article

Sugar beet (Beta vulgaris L.) is one of the most important sugar crops and potential energy crops in China. The utilization of its heterosis is crucial for breaking through the bottlenecks in yield and quality, while the fertility identification of binary male-sterile lines is the core link to ensure the purity of hybrid seeds. Due to its indeterminate inflorescence, artificial emasculation of sugar beet is not feasible, which significantly increases the difficulty in hybrid seed production. To rapidly and accurately identify the fertility composition of monogerm binary male-sterile lines of sugar beet, ensure the maternal line purity in sugar beet hybrid seed production, and improve breeding efficiency, this study conducted fertility identification using molecular marker technology with 7 monogerm binary male-sterile line germplasm resources (297 plants) provided by three research institutions in different regions of China. Genomic DNA was extracted from young sugar beet leaves by the CTAB method. The cytoplasmic fertility types were identified using the TR1 primer, and the fertility gene composition at the nuclear Rf1 locus was verified by the s17 molecular marker combined with Hap II and Hind III double digestion. The results showed that in the cytoplasmic fertility identification, the proportion of S-type cytoplasm in Lines 2 to 7 reached 100%, indicating stable sterility without maintainer line contamination; Line 1 had 93.33% S-type cytoplasm, mixed with 6.67% N-type cytoplasm. For the nuclear Rf1 locus identification, 93.27% (277 plants) of the tested materials yielded the target 1800 bp band by PCR amplification, which were preliminarily identified as homozygous recessive type. Among them, Lines 1 to 3 all showed a single 1800 bp band pattern, indicating homozygous and consistent nuclear fertility genotypes; 20 plants (6.73%) in Lines 4 to 7 exhibited a composite 1800/1300 bp band pattern, suggesting the presence of restorer allele contamination in some lines. Genotype analysis based on 35 enzyme-digested verification samples revealed that the 4/4 genotype had the highest proportion. This study realized the rapid and accurate identification of cytoplasmic and nuclear fertility in monogerm binary male-sterile lines of sugar beet through molecular marker technology, clarified the fertility purity status of 7 germplasm resources, and verified the application value of this technology in the fertility identification of sugar beet binary male-sterile lines. These results provide a scientific basis and technical support for controlling maternal line purity and improving breeding efficiency in sugar beet hybrid seed production. Full article

Oxidative stress (OS) and sperm DNA fragmentation (SDF) are complementary contributors to male infertility. OS characterizes a compromised seminal redox status, whereas SDF quantifies downstream genomic damage. Human sperm are highly susceptible to redox damage due to lipid-rich membranes and disrupted post-meiotic DNA-repair capacity. Excess reactive oxygen species (ROS) can cause lipid peroxidation, oxidative base lesions, and DNA strand breaks that impair fertilization, embryo development, and pregnancy outcomes. This review explains how OS promotes genomic instability and summarizes the main laboratory assays that assess redox status and SDF in semen. These include direct ROS chemiluminescence assay, oxidation–reduction potential, total antioxidant capacity/ferric reducing antioxidant power, and lipid peroxidation biomarkers, alongside SDF platforms (Sperm Chromatin Structure Assay, terminal deoxynucleotidyl transferase dUTP nick-end labeling, alkaline/neutral Comet, and sperm chromatin dispersion). Additionally, guideline-aligned indications are highlighted to clarify the conditions for testing OS and SDF. OS testing is most relevant in men with leukocytospermia or suspected genital tract infection or inflammation, including dysbiosis; in cases of major modifiable exposures such as smoking or heat; and for early monitoring after treatment. SDF testing is particularly informative in couples with recurrent pregnancy loss and in unexplained infertility with normal semen parameters. Combined OS and SDF testing is recommended in clinical varicocele, repeated in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) failure, poor embryo development, and follow-up after targeted therapy. Management centers on treating infection and inflammation, improving lifestyle and environmental factors, considering varicocelectomy when indicated, using targeted antioxidant therapy in men with documented OS, and selectively applying sperm selection technologies or testicular sperm for ICSI when SDF remains high. Priorities include assay standardization, etiologic attribution of DNA damage, and trials testing OS/SDF-guided pathways with live birth as the primary endpoint. When used selectively and in the appropriate context, OS and SDF testing can help refine diagnosis, improve counseling, and help personalize care of infertile couples. Full article

DNA double-strand breaks (DSBs) are primarily repaired in eukaryotic cells through two pathways: homologous recombination (HR) and non-homologous end joining (NHEJ). The thermotolerant yeast Kluyveromyces marxianus is recognized for its highly active NHEJ pathway, making it a suitable model organism for studying the role of NHEJ in DSB repair. To induce DSBs in K. marxianus DMKU3-1042, an expression cassette containing the gene encoding the endonuclease RsaI was integrated into the LYS1 locus of both the wild-type and NHEJ-deficient KU70 mutant strains. This cassette is regulated by the galactose-inducible promoter GAL10. Cells expressing RsaI and grown in galactose medium exhibited an elongated, rod-shaped morphology under a microscope. Following RsaI expression, the viability of transformed KU70 cells decreased during the first three hours of culture in liquid medium and then partially recovered after six hours of incubation. In contrast, the KU70 mutant cells failed to produce viable survivors. Pulsed-field gel electrophoresis analysis revealed distinct chromosomal separation patterns among various RsaI-transformed KU70 cells. These findings demonstrate that the repair of RsaI-induced DSBs in K. marxianus DMKU3-1042 results in new strains with several forms of rearranged chromosomes. Full article

Human preimplantation embryo arrest (PREMBA) represents a significant clinical hurdle in assisted reproductive technology (ART), in which approximately 10% of in vitro fertilized (IVF) embryos arrest at the cleavage stages. Whole-exome sequencing (WES) studies have discovered numerous genetic mutations associated with preimplantation embryo arrest. These mutations often disrupt critical biological milestones such as maternal mRNA clearance (BTG4, ZFP36L2, ZAR1), subcortical maternal complex (TLE6, PADI6, OOEP, NLRP2, NLRP5, NLRP7, KHDC3L), DNA double-strand break formation and homologous recombination (REC114, TOP6BL, MEI1, MEI4, TRIP13), spindle assembly (TUBB8 and TUBA4A) and cell cycle and checkpoints (FBXO43, MOS, CHEK1, TRIP13, CDC20), as well as nuclear transport and translational regulation (KPNA7, DDOST). However, the cause of most clinical cases remains genetically unexplained. Studies investigating these unexplained arrests have uncovered widespread multi-omics abnormalities, including transcriptional arrest, DNA hypermethylation, higher chromatin accessibility, aberrant histone modification, chromosomal aneuploidy and senescent-like states. This review provides a comprehensive overview of the molecular mechanisms underlying PREMBA, categorized into those that are attributable to known genetic mutations and those with unexplained reasons. Full article

Colorectal cancer (CRC) is traditionally considered a “cold tumor” characterized by low immunogenicity and limited responsiveness to immune checkpoint inhibitors (ICIs). However, recent findings reveal that cytotoxic modalities can reprogram this immunologically inert landscape. This review integrates these evolving concepts to guide the optimization of future treatments. Radiotherapy induces extensive DNA double-strand breaks, which may generate de novo mutations through error-prone repair while simultaneously exposing cryptic antigens via increased transcriptional instability, alternative splicing, and enhanced proteasomal processing. Chemoradiation also amplifies epigenetic and epitranscriptomic sources of neoepitope diversity, including RNA editing and stress-induced splicing alterations, expanding the immunopeptidome beyond canonical mutation-driven neoantigens. These changes collectively enhance antigen presentation and facilitate T-cell priming. Chemotherapy further reduces immunosuppressive cell populations and promotes dendritic cell activation, creating a permissive milieu for subsequent immune engagement. Clinically, the VOLTAGE studies demonstrated that long-course chemoradiotherapy can sensitize even mismatch repair–proficient rectal cancers to PD-1 blockade, yielding clinically meaningful pathological responses. In contrast, mismatch repair–deficient rectal tumors may respond completely to ICIs alone. Short-course radiotherapy combined with chemotherapy and ICIs has also shown encouraging activity in the setting of total neoadjuvant therapy. Collectively, these findings support a paradigm in which radiotherapy, chemotherapy, and epigenetic/epitranscriptomic alterations—including RNA editing—act as potent modulators of tumor antigenicity. By expanding the neoantigen repertoire and reshaping the tumor microenvironment, these strategies can transform CRC from a cold tumor into one that is increasingly responsive to immunotherapy. Full article

Proton therapy offers superior dose localization, yet the biological effects of low-energy protons relevant to superficial tissues remain underexplored. We report the design and validation of a proton irradiation setup developed at the Tandem Accelerator of NCSR “Demokritos” for controlled radiobiological experiments. Monte Carlo simulations using Geant4 and Monte Carlo Damage Simulation (MCDS—Monte Carlo Damage Simulation) were used to determine proton energy spectra, linear energy transfer (LET), and predicted DNA damage yields. A single layer (15–20 μm in thickness) of human keratinocytes (HaCaT) was irradiated at doses from 0.65 to 3.65 Gy, and γ-H2AX foci were quantified as markers of tracks including one or more DNA double-strand breaks. The system achieved a uniform dose rate of 0.37 Gy/min, as calculated with Geant4, with a mean proton energy of 4.1 MeV (LET ≈ 8 keV/μm). A strong correlation (R² = 0.93) was observed between proton dose and γH2AX foci per nucleus (~10 foci/Gy), reflecting damage-inducing proton tracks rather than individual DNA double-strand breaks. At higher doses, an increased fraction of cells exhibited pan-nuclear γH2AX staining, characterized by a diffuse γH2AX signal throughout the nucleus and commonly associated with extensive or clustered DNA damage and global chromatin phosphorylation. These responses are consistent with the well-established dense ionization patterns produced by low-energy protons, as indicated by the LET spectrum and supported by MCDS-predicted clustered damage yields. While the γH2AX assay does not directly resolve simple versus complex DNA lesions, the agreement between Monte Carlo modeling and the observed cellular stress responses indicates that the irradiation platform reliably reproduces the expected biological signatures of low-energy proton exposure. Consequently, the developed system provides a robust experimental tool for systematic investigations of cellular radiosensitivity and radiotoxicity, with potential applications in skin dosimetry and radioprotection. Full article

Parkinson’s disease (PD) is a complex neurodegenerative disorder influenced by age, genetic predispositions, and environmental exposures, with a growing global incidence. This review aims to summarize findings from ATSDR Toxicological Profiles, EPA Risk Assessments, and other sources of peer-reviewed literature to examine the potential associations between PD and select metals, pesticides, and chlorinated organic compounds. Additionally, it explores using computational toxicology methods to elucidate the interactions between specific chemicals, associated genes, and their possible roles in PD. A total of 29 substances were identified to be neurotoxic with direct or probable association with PD. Risk of disease onset or symptom exacerbation of PD has been linked to exposures to neurodegenerative metals, pesticides, chlorinated organic compounds, and other environmental toxicants, alongside intrinsic factors such as genetic predisposition and aging. Supporting evidence from neurotoxicological studies directly or possibly associated with PD are summarized in referenced toxicological profiles and EPA risk assessments. Genotoxic endpoints evaluated in exposure-induced neurodegeneration including oxidative stress, DNA strand breaks, mitochondrial dysfunction, impaired DNA repair, and telomere alterations may play a critical role in linking environmental exposures to PD pathogenesis. Although these endpoints represent imperative data gaps between environmental and genetic risk factors for PD, isolating individual substances may not be necessary for prevention, as many co-occur at contaminated sites or within certain occupations. Further research is needed to clarify causal relationships between environmental exposure and genotoxic endpoints seen in neurodegenerative processes that can also be seen in PD for consideration in the development of preventive and therapeutic strategies. Full article