Dapagliflozin suppresses diabetes-induced oxidative DNA damage and hypermethylation in mouse somatic cells.

Diabetes mellitus is a complex metabolic disorder resulting from the interplay of environmental, genetic, and epigenetic factors that increase the risk of cancer development. However, it is unclear whether the increased cancer risk is due to poor glycemic control or the use of some antidiabetic medications. Therefore, we investigated the genetic and epigenetic changes in somatic cells in a mouse model of diabetes and studied whether multiple exposures to the antidiabetic medication dapagliflozin influence these changes. We also elucidated the mechanism(s) of these ameliorations. The micronucleus test and modified comet assay were used to investigate bone marrow DNA damage and methylation changes. These assays revealed that dapagliflozin is non-genotoxic in the tested regimen, and oxidative DNA damage and hypermethylation were significantly higher in diabetic mice. Spectrophotometry also evaluated oxidative DNA damage and global DNA methylation, revealing similar significant alterations induced by diabetes. Conversely, the dapagliflozin-treated diabetic animals significantly reduced these changes. The expression of some genes involved in DNA repair and DNA methylation was disrupted considerably in the somatic cells of diabetic animals. In contrast, dapagliflozin treatment significantly restored these disruptions and enhanced DNA repair. The simultaneous effects of decreased oxidative DNA damage and hypermethylation levels suggest that dapagliflozin can be used as a safe antidiabetic drug to reduce DNA damage and hypermethylation in diabetes, demonstrating its usefulness in patients with diabetes to control hyperglycemia and decrease the development of its subsequent complications.

Aflatoxin B1 exposure exacerbates chemokine receptor expression in the BTBR T+ Itpr3tf/J Mouse Model, unveiling insights into autism spectrum disorder: A focus on brain and spleen.

OBJECTIVE: Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by significant difficulties in social interaction, communication, and repeated stereotypic behaviour. Aflatoxin B1 (AFB1) is the most potent and well-known mycotoxin in various food sources. Despite its propensity to generate significant biochemical and structural changes in human and animal tissues, the influence of AFB1 on ASD has yet to be thoroughly studied. Mounting evidence indicates that chemokine receptors play a crucial function in the central nervous system and are implicated in developing several neuroinflammatory disorders. Chemokine receptors in individuals with ASD were elevated in the anterior cingulate gyrus astrocytes, cerebellum, and brain. METHODS: The BTBR T+Itpr3tf/J (BTBR) mice are inbred strains that exhibit strong and consistently observed deficits in social interactions, characterized by excessive self-grooming and limited vocalization in social contexts. We examined the impact of AFB1 on CCR3-, CCR7-, CCR9-, CXCR3-, CXCR4-, and CXCR6-expressing I-A/I-E+ cells in the spleen of the BTBR mouse model of autism. We evaluated the mRNA levels of CCR3, CCR7, CCR9, CXCR3, CXCR4, and CXCR6 chemokine receptors in the brain. RESULTS: The exposure to AFB1 in BTBR mice resulted in a significant rise in the number of I-A/I-E+CCR3+, I-A/I-E+CCR7+, I-A/I-E+CCR9+, I-A/I-E+CXCR3+, I-A/I-E+CXCR4+, and I-A/I-E+CXCR6+ cells. Furthermore, exposure to AFB1 increased mRNA expression levels of CCR3, CCR7, CCR9, CXCR3, CXCR4, and CXCR6 in the brain. CONCLUSIONS: These findings highlight that AFB1 exposure increases the expression of chemokine receptors in BTBR mice, indicating the necessity for further research into AFB1's role in the development of ASD.

Saxagliptin, a selective dipeptidyl peptidase-4 inhibitor, alleviates somatic cell aneugenicity and clastogenicity in diabetic mice.

Diabetes-related complications are becoming increasingly common as the global prevalence of diabetes increases. Diabetes is also linked to a high risk of developing cancer. This raises the question of whether cancer vulnerability is caused by diabetes itself or the use of antidiabetic drugs. Chromosomal instability, a source of genetic modification involving either an altered chromosomal number or structure, is a hallmark of cancer. Saxagliptin has been approved by the FDA for diabetes treatment. However, the detailed in vivo effects of prolonged saxagliptin treatment on chromosomal instability have not yet been reported. In this study, streptozotocin was used to induce diabetes in mice, and both diabetic and non-diabetic mice received saxagliptin for five weeks. Fluorescence in situ hybridization was conducted in combination with a bone marrow micronucleus test for measuring chromosomal instability. Our results indicated that saxagliptin is neither mutagenic nor cytotoxic, under the given treatment regimen. Diabetic mice had a much higher incidence of micronuclei formation, and a centromeric DNA probe was present inside the majority of the induced micronuclei, indicating that most of these were caused by chromosome nondisjunction. Conversely, diabetic mice treated with saxagliptin exhibited a significant decrease in micronuclei induction, which were centromeric-positive and centromeric-negative. Diabetes also causes significant biochemical changes indicative of oxidative stress, such as increased lipid peroxidation and decreased reduced/oxidized glutathione ratio, which was reversed by saxagliptin administration. Overall, saxagliptin, the non-mutagenic antidiabetic drug, maintains chromosomal integrity in diabetes and reduces micronuclei formation by restoring redox imbalance, further indicating its usefulness in diabetic patients.

Dapagliflozin Mitigated Elevated Disomic and Diploid Sperm in a Mouse Model of Diabetes and Recover the Disrupted Ogg1, Parp1, and P53 Gene Expression.

Increases in numerical chromosomal syndromes were observed in children of diabetic mothers. However, the effects of diabetes on male reproduction, specifically numerical chromosomal aberrations (aneuploidy), have not been studied. Furthermore, despite the increasing use of dapagliflozin for diabetes treatment, no data exists on its ability to affect aneuploidy levels in germ cells. Thus, our investigation aimed to evaluate the effects of diabetes on spontaneous sperm aneuploidy and whether treatment with dapagliflozin influences the frequency of aneuploidy in the sperm of an experimental diabetic animal model. Our findings show that dapagliflozin has no aneugenic effects on the meiotic stages of spermatogenesis. In contrast, diabetes raised the frequency of aneuploidy, and dapagliflozin administration decreased the elevated levels of disomic and diploid sperm. The level of oxidative stress was markedly increased in diabetic mice, but were reduced by dapagliflozin treatment. Furthermore, the expression of some of DNA repair genes was disrupted in diabetic animals, whereas dapagliflozin therapy restored these disruptions and significantly enhanced DNA repair. Thus, dapagliflozin may effectively ameliorate diabetes-induced aneugenic effects on male meiosis and treating diabetic patients with dapagliflozin may effectively mitigate the transmission of diabetes-induced chromosomal defects to offspring.

Carfilzomib Mitigates Lipopolysaccharide/D-Galactosamine/Dimethylsulfoxide-Induced Acute Liver Failure in Mice.

Acute liver failure (ALF) is a disease accompanied by severe liver inflammation. No effective therapy is available yet apart from liver transplantation; therefore, developing novel treatments for ALF is urgently required. Inflammatory mediators released by NF-кB activation play an essential role in ALF. Proteasome inhibitors have many medical uses, such as reducing inflammation and NF-кB inhibition, which are believed to account for most of their repurposing effects. This study was undertaken to explore the possible protective effects and the underlying mechanisms of carfilzomib, a proteasome inhibitor, in a mouse model of ALF induced by lipopolysaccharide/D-galactosamine/dimethylsulfoxide (LPS/GalN/DMSO). Carfilzomib dose-dependently protected mice from LPS/GalN/DMSO-induced liver injury, as indicated by the decrease in serum alanine aminotransferase and aspartate aminotransferase levels. LPS/GalN/DMSO increased TNF-α, NF-кB, lipid peroxidation, NO, iNOS, cyclooxygenase-II, myeloperoxidase, and caspase-3 levels. Carfilzomib administration mitigated LPS/GalN/DMSO-induced liver damage by decreasing the elevated levels of TNF-α, NF-кB, lipid peroxidation, nitric oxide, iNOS, cyclooxygenase-II, myeloperoxidase, caspase-3, and histopathological changes. A restored glutathione level was also observed in the carfilzomib-treated LPS/GalN/DMSO mice. Our results demonstrate that carfilzomib protects against LPS/GalN/DMSO-induced ALF by inhibiting NF-кB, decreasing inflammatory mediators, oxidative/nitrosative stress, neutrophil recruitment, and apoptosis, suggesting that carfilzomib may be a potential therapeutic agent for ALF.

Impacts of the DPP-4 Inhibitor Saxagliptin and SGLT-2 Inhibitor Dapagliflozin on the Gonads of Diabetic Mice.

Diabetes mellitus is a metabolic disease that can cause systemic problems, including testicular dysfunction. Several diabetes medications have demonstrated potential adverse effects on the male reproductive system; however, the effects of saxagliptin and dapagliflozin have not been sufficiently examined. This investigation studied the impacts of saxagliptin and dapagliflozin treatments on the gonads in a male mouse model of diabetes. Testicular disturbances were assessed by sperm DNA damage, diakinesis-metaphase I chromosome examination, and spermiogram analysis. Our results showed more sperm DNA damage, more spermatocyte chromosome aberrations, lower sperm motility/count, and more sperm morphological anomalies in diabetic mice than in the control mice. Dapagliflozin significantly restored all examined measures to the control values in diabetic mice, unlike saxagliptin, which exacerbated the reduction in sperm count and motility. Both drugs significantly restored the gonadal redox imbalances in diabetic mice by decreasing reactive oxygen species accumulation and increasing glutathione levels. In conclusion, our study presents preliminary evidence for the safety and efficacy of dapagliflozin in alleviating testicular abnormalities induced by diabetes, making it a promising candidate drug for patients with diabetes in their reproductive age. As saxagliptin may have negative effects on fertility, its prescription should be avoided in young male diabetic patients.

Rituximab alleviates increased disomic sperm in DBA/1J mouse models of rheumatoid arthritis via restoration of redox imbalance.

Compared to the general population, patients with arthritis have a higher risk of fertility abnormalities, which have deleterious effects on both reproductive function and pregnancy outcomes, especially in patients wishing to conceive. These may be due to the disease itself or those of drug therapies. Despite the increasing use of rituximab in arthritis, limited data are available on its potential to induce aneuploidy in germ cells. Therefore, the aim of the current investigation was to determine if repeated treatment with rituximab affects the incidence of aneuploidy and redox imbalance in arthritic mouse sperm. Mice were treated with 250 mg/kg rituximab once weakly for 3 weeks, and then sperm were sampled 22 days after the last dose of rituximab. Fluorescence in situ hybridization assay with chromosome-specific DNA probes was used to evaluate the disomic/diploid sperm. Our results showed that rituximab had no aneuploidogenic effect on the meiotic stage of spermatogenesis. Conversely, arthritis induced a significantly high frequency of disomy, and treatment of arthritic mice with rituximab reduced the increased levels of disomic sperm. The occurrence of total diploidy was not significantly different in all groups. Reduced glutathione and8-hydroxydeoxyguanosine, markers of oxidative stress were significantly altered in arthritic animals, while rituximab treatment restored these changes. Additionally, arthritis severity was reduced after rituximab treatment. We conclude that rituximab may efficiently alleviate the arthritis-induced effects on male meiosis and avert the higher risk of abnormal reproductive outcomes. Therefore, treating arthritic patients with rituximab may efficiently inhibit the transmission of genetic anomalies induced by arthritis to future generations.

Aflatoxin B1 Exacerbates Genomic Instability and Apoptosis in the BTBR Autism Mouse Model via Dysregulating DNA Repair Pathway.

The pathophysiology of autism is influenced by a combination of environmental and genetic factors. Furthermore, individuals with autism appear to be at a higher risk of developing cancer. However, this is not fully understood. Aflatoxin B1 (AFB1) is a potent food pollutant carcinogen. The effects of AFB1 on genomic instability in autism have not yet been investigated. Hence, we have aimed to investigate whether repeated exposure to AFB1 causes alterations in genomic stability, a hallmark of cancer and apoptosis in the BTBR autism mouse model. The data revealed increased micronuclei generation, oxidative DNA strand breaks, and apoptosis in BTBR animals exposed to AFB1 when compared to unexposed animals. Lipid peroxidation in BTBR mice increased with a reduction in glutathione following AFB1 exposure, demonstrating an exacerbated redox imbalance. Furthermore, the expressions of some of DNA damage/repair- and apoptosis-related genes were also significantly dysregulated. Increases in the redox disturbance and dysregulation in the DNA damage/repair pathway are thus important determinants of susceptibility to AFB1-exacerbated genomic instability and apoptosis in BTBR mice. This investigation shows that AFB1-related genomic instability can accelerate the risk of cancer development. Moreover, approaches that ameliorate the redox balance and DNA damage/repair dysregulation may mitigate AFB1-caused genomic instability.

Aneugenic and clastogenic alterations in the DBA/IJ mouse model of rheumatoid arthritis treated with rituximab, an anti-CD20 antibody.

Rheumatoid arthritis (RA), an autoimmune disorder in which the immune system attacks healthy cells, is associated with elevated risk of lymphoma. Rituximab, a treatment for non-Hodgkin's lymphoma, has been approved as a treatment for RA. We studied the effects of rituximab on chromosomal stability in collagen-induced arthritis DBA/1J animal models. Micronucleus levels were increased in the mouse models, mainly due to chromosome loss, as detected by fluorescence in situ hybridization; rituximab-treated arthritic mice had significantly less micronucleus formation. Serum 8-hydroxydeoxyguanosine, a DNA oxidative stress marker, was increased in the mice models but reduced following rituximab administration.

Treatment with the anti-CD20 monoclonal antibody rituximab mitigates gonadal disruptions in the collagen-induced arthritis in male DBA/1 J mouse model.

Rheumatoid arthritis (RA), which is driven by persistent activation of the immune system, primarily affects the joints. Several reports have estimated the risk of gonadal disruptions in arthritic patients, with potential attributable risk factors such as treatments with the disease-modifying antirheumatic drugs and the influence of the disease itself. The FDA approved rituximab, a therapy for non-Hodgkin's lymphoma, for management of RA in February 2006. However, the influence of repeated treatment with rituximab on gonadal function in RA has not been reported yet. Thus, the aim of the presents study is to evaluate whether repeated treatment with the clinically relevant dose of rituximab may change the gonadal disruptions in collagen-induced arthritis in male DBA/1 J mouse, a model of RA. Testicular disruptions, as determined by the sperm DNA strand breaks, spermatocyte chromosomal analysis and spermiogram examination have been conducted by the use of standard techniques. Additionally, we aimed to test whether the anti-rheumatic effect of rituximab also decreases the cellular oxidant-antioxidant imbalance in arthritic male DBA/1 J mice. Repeated treatment of naïve control DBA/1 J mice with rituximab did not exhibit any significant deleterious effects. Moreover, repeated administration of rituximab to the arthritic DBA/1 J mice suppressed disease severity and decreased testicular disruptions. Rituximab treatment also diminished gonadal oxidative stress, through decreasing reactive oxygen species generation and restoring the reduced glutathione level in arthritic DBA/1 J mice. In conclusion, rituximab is a safe therapeutic agent and can mitigate gonadal disruptions induced by arthritis, which insinuates the importance for arthritic patients especially at reproductive age.

Multiple exposure to methylmercury aggravates DNA damage in the BTBR T + Itpr3 tf/J autistic mouse model: The role of DNA repair efficiency.

Environmental and genetic factors have been recognized to play major roles in the pathogenesis of autism. Here we examined the BTBR T+Itpr3tf/J (BTBR) mice's susceptibility, an autistic model, to the genotoxic effects and DNA repair dysregulation of methylmercury. Micronuclei formation and oxidative DNA damage were analyzed using the micronucleus/fluorescence in situ hybridization test and modified comet assay, respectively. The results showed higher centromeric-positive micronuclei and oxidative DNA damage in BTBR mice exposed to methylmercury than the unexposed mice, which indicates that mutagenesis aggravated in BTBR mice after methylmercury exposure. Lipid peroxides in BTBR mice were significantly elevated, with a decrease in reduced/oxidized glutathione ratio after methylmercury exposure, indicating an augmenting oxidant-antioxidant imbalance. The expression of several genes involved in DNA repair was markedly altered in BTBR mice after methylmercury exposure as evaluated via PCR array and RT-PCR analyses. Declining of the antioxidant defense and dysregulation in DNA repair process after methylmercury exposure may explain the aggravated genotoxic susceptibility of BTBR mice. Thus, autistic individuals exposed to methylmercury must be under regular medical follow-up through standard timetabled medical laboratory inquiry to allow for early recognition of any mutagenic changes. Additionally, strategies that elevate cellular antioxidants/DNA repair efficiency may counteract methylmercury-induced genotoxicity.

The MAP kinase inhibitor PD98059 reduces chromosomal instability in the autoimmune encephalomyelitis SJL/J-mouse model of multiple sclerosis.

Multiple sclerosis (MS), a disease in which the immune system attacks nerve cells, has been associated with both genetic and environmental risk factors. We observed increased micronucleus (MN) formation in SJL/J mouse experimental autoimmune encephalomyelitis (EAE), an animal model of MS. Most of these MN were due to chromosomal loss. Increased activation of MAP kinases, which leads to disruption of the mitotic spindle and improper segregation of chromosomes, is associated with MS. MAP kinase inhibitors, such as PD98059, may therefore be beneficial for MS. In the EAE model, PD98059 treatment reduced adverse effects, including MN formation, lipid peroxidation, and GSH oxidation. Interventions that mitigate chromosomal instability may have therapeutic value in MS.

3-Aminobenzamide alleviates elevated DNA damage and DNA methylation in a BTBR T+Itpr3tf/J mouse model of autism by enhancing repair gene expression.

Little is known about genetic and epigenetic alterations in autism spectrum disorder. Moreover, the efficiency of DNA repair in autism must be improved to correct these alterations. We examined whether 3-aminobenzamide (3-AB) could reverse these alterations. We conducted experiments to clarify the molecular mechanism underlying these ameliorations. An assessment of genetic and epigenetic alterations by a modified comet assay showed elevated levels of oxidative DNA strand breaks and DNA hypermethylation in BTBR T+Itpr3tf/J (BTBR) mice used as a model of autism. Oxidative DNA strand breaks and DNA methylation were further quantified fluorometrically, and the results showed similar changes. Conversely, 3-AB treated BTBR mice showed a significant reduction in these alterations compared with untreated mice. The expressions of 43 genes involved in DNA repair were altered in BTBR mice. RT2 Profiler PCR Array revealed significantly altered expression of seven genes, which was confirmed by RT-PCR analyses. 3-AB treatment relieved these disturbances and significantly improved Ogg1 and Rad1 up-regulation. Moreover, autism-like behaviors were also mitigated in BTBR animals by 3-AB treatment without alterations in locomotor activities. The simultaneous effects of reduced DNA damage and DNA methylation levels as well as the regulation of repair gene expression indicate the potential of 3-AB as a therapeutic agent to decrease the levels of DNA damage and DNA methylation in autistic patients. The current data may help in the development of therapies that ultimately provide a better quality of life for individuals suffering from autism.

Vorinostat is genotoxic and epigenotoxic in the mouse bone marrow cells at the human equivalent doses.

Vorinostat was approved as the first histone deacetylase inhibitor for the management of cutaneous T cell lymphoma. However, it's in vivo genetic and epigenetic effects on non-cancerous cells remain poorly understood. As genetic and epigenetic changes play a critical role in the pathogenesis of carcinogenesis, we investigated whether vorinostat induces genetic and epigenetic alterations in mouse bone marrow cells. Bone marrow cells were isolated 24 h following the last oral administration of vorinostat at the doses of 25, 50, or 100 mg/kg/day for five days (approximately equal to the recommended human doses). The cells were then used to assess clastogenicity and aneugenicity by the micronucleus test complemented by fluorescence in situ hybridization assay; DNA strand breaks, oxidative DNA strand breaks, and DNA methylation by the modified comet assay; apoptosis by annexin V/PI staining analysis and the occurrence of the hypodiploid DNA content; and DNA damage/repair gene expression by polymerase chain reaction (PCR) Array. The expression of the mRNA transcripts were also confirmed by real-time PCR and western blot analysis. Vorinostat caused structural chromosomal damage, numerical chromosomal abnormalities, DNA strand breaks, oxidative DNA strand breaks, DNA hypomethylation, and programed cell death in a dose-dependent manner. Furthermore, the expression of numerous genes implicated in DNA damage/repair were altered after vorinostat treatment. Accordingly, the genetic/epigenetic mechanism(s) of action of vorinostat may play a role in its carcinogenicity and support the continued study and development of new compounds with lower toxicity.

Evaluation of DNA repair efficiency in autistic children by molecular cytogenetic analysis and transcriptome profiling.

Data regarding DNA repair perturbations in autism, which might increase the risk of malignancy, are scarce. To evaluate whether DNA repair may be disrupted in autistic children, we assessed the incidence of endogenous basal DNA strand breaks as well as the efficiency of repairing DNA damage caused by γ-ray in lymphocytes isolated from autistic and healthy children. The incidence of DNA damage and the kinetics of DNA repair were determined by comet assay, while the incidence of residual DNA damage was evaluated by structural chromosomal aberration analysis. Transcriptome profiling of 84 genes associated with DNA damage and repair-signaling pathways was performed by RT² Profiler PCR Array. The array data were confirmed by RT-PCR and western blot studies. Our data indicate that the incidence of basal oxidative DNA strand breaks in autistic children was greater than that in nonautistic controls. Lymphocytes from autistic children displayed higher susceptibility to damage by γ-irradiation and slower repair rate than those from nonautistic children. Although the total unstable chromosomal aberrations were unaffected, lymphocytes from autistic children were more susceptible to chromosomal damage caused by γ-ray than those from nonautistic children. Transcriptomic analysis revealed that several genes associated with repair were downregulated in lymphocytes from autistic individuals and in those exposed to γ-irradiation. This may explain the increased oxidative DNA damage and reduced repair rate in lymphocytes from autistic individuals. These features may be related to the possible correlation between autism and the elevated risk of cancer and may explain the role of the disruption of the DNA repair process in the pathogenesis of autism.

Genetic and epigenetic alterations induced by the small-molecule panobinostat: A mechanistic study at the chromosome and gene levels.

Increasing evidence supports the role of genetic and epigenetic alterations in a wide variety of human diseases, including cancer. Assessment of these alterations is hence essential for estimating the hazardous effects of human exposure to medications. Panobinostat received US Food and Drug Administration's approval in 2015 for treatment of certain tumors and its usefulness as part of a strategy to treat other diseases, such as human immunodeficiency virus infection, is currently investigated. Nevertheless, no data on in vivo genotoxical and epigenotoxical effects of panobinostat are available. The aim of the current study was to assess the genotoxical and epigenotoxical properties of panobinostat in murine bone marrow cells. Molecular mechanisms underlying these alterations were also evaluated. We show that mice treated with panobinostat doses recommended for human developed numerical chromosomal abnormalities, structural chromosomal damage, oxidative DNA damage, and DNA hypomethylation. These effects were dose-dependent. Further, panobinostat altered the expression of 23 genes implicated in DNA damage, as determined by RT² Profiler polymerase chain reaction (PCR) array, and confirmed by quantitative real-time PCR and western blotting. Collectively, these findings indicate that panobinostat exposure induces aneugenicity, clastogenicity, oxidative DNA damage, DNA hypomethylation, and down-regulation of repair gene expression, which may be responsible for panobinostat-induced genotoxical and epigenotoxical effects. Considering the potential toxicity of panobinostat, the medicinal use of panobinostat must be weighed against the risk of tumorigenesis and the demonstrated toxicity profile of panobinostat may support further development of chemotherapeutic treatments with reduced toxicity. Diminishing the metabolic liabilities associated with panobinostat exposure, and simultaneous use of panobinostat with DNA repair enhancers, are examples of strategies for drug design to reduce panobinostat carcinogenicity.