Decreased DNA Damage Research Articles

1,25-Dihydroxyvitamin D exerts an antiaging role by activation of Nrf2-antioxidant signaling and inactivation of p16/p53-senescence signaling.

We tested the hypothesis that 1,25‐dihydroxyvitamin D3[1α,25(OH)2D3] has antiaging effects via upregulating nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2), reducing reactive oxygen species (ROS), decreasing DNA damage, reducing p16/Rb and p53/p21 signaling, increasing cell proliferation, and reducing cellular senescence and the senescence‐associated secretory phenotype (SASP). We demonstrated that 1,25(OH)2D3‐deficient [1α(OH)ase−/−] mice survived on average for only 3 months. Increased tissue oxidative stress and DNA damage, downregulated Bmi1 and upregulated p16, p53 and p21 expression levels, reduced cell proliferation, and induced cell senescence and the senescence‐associated secretory phenotype (SASP) were observed. Supplementation of 1α(OH)ase−/− mice with dietary calcium and phosphate, which normalized serum calcium and phosphorus, prolonged their average lifespan to more than 8 months with reduced oxidative stress and cellular senescence and SASP. However, supplementation with exogenous 1,25(OH)2D3 or with combined calcium/phosphate and the antioxidant N‐acetyl‐l‐cysteine prolonged their average lifespan to more than 16 months and nearly 14 months, respectively, largely rescuing the aging phenotypes. We demonstrated that 1,25(OH)2D3exerted an antioxidant role by transcriptional regulation of Nrf2 via the vitamin D receptor (VDR). Homozygous ablation of p16 or heterozygous ablation of p53 prolonged the average lifespan of 1α(OH)ase−/− mice on the normal diet from 3 to 6 months by enhancing cell proliferative ability and reducing cell senescence or apoptosis. This study suggests that 1,25(OH)2D3 plays a role in delaying aging by upregulating Nrf2, inhibiting oxidative stress and DNA damage，inactivating p53‐p21 and p16‐Rb signaling pathways, and inhibiting cell senescence and SASP.

Influence of chromatin compaction on simulated early radiation-induced DNA damage using Geant4-DNA.

In this work, we present simulated double-strand breaks (DSBs) obtained for two human cell nucleus geometries. The first cell nucleus represents fibroblasts, filled with DNA molecules in different compaction forms: heterochromatin or euchromatin only. The second one represents an endothelial cell nucleus, either filled with heterochromatin only or with a uniform distribution of 48% of heterochromatin and 52% of euchromatin, obtained from measurements carried out at IRSN. Protons and alpha particles of different energies were used as projectiles. Each cell nucleus model includes a multi-scale description of the DNA target from the molecular level to the whole human genome representation. The cell nucleus models were generated using an extended version of the DnaFabric software in which a new model of euchromatin was implemented in addition to the existing model of heterochromatin. Thus, each nucleus model contains the complete human genome (a total of 6 Gbp) in the G0/G1 phase of the cycle, filled with a continuous chromatin fiber per chromosome that can take into account the heterochromatin and the euchromatin compaction. These geometries were then exported to a simulation chain using the Monte Carlo toolkit Geant4-DNA to perform computations of the physical, physicochemical, and chemical stages, in order to evaluate the influence of chromatin compaction on DSB induction and the contribution of direct and indirect damage, as well as DSB complexity. More direct damage and less indirect damage were observed in the heterochromatin than in the euchromatin. Nevertheless, no difference in terms of DSB complexity was observed between those formed in the heterochromatin or the euchromatin models. Yields of DSB/Gy/Gbp show an increase when both heterochromatin and euchromatin models are taken into account, compared to when only heterochromatin is considered. The results presented indicate that the chromatin compaction decreases DNA damage generated by ionizing radiation and thus, DNA compaction should be considered for the simulation of DNA repair and other cellular outcomes.

Activation of Nrf2/Keap1 pathway by oral Dimethylfumarate administration alleviates oxidative stress and age-associated infertility might be delayed in the mouse ovary

BackgroundAge-associated infertility is a problem worldwide, and management of oxidative stress is known to be essential. Nuclear factor-E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1)-antioxidant response element (ARE) signaling pathway works as an essential defense mechanism against oxidative stress, and an oral drug Dimethylfumarate (DMF) is known to activate the pathway.MethodsWe tested the hypothesis that oral DMF could alleviate oxidative stress in the ovary, resulting in salvation of age-associated infertility in a mouse model of reproductive age, and we examined the effects of DMF administration. 20 mg/kg DMF was administrated to female mice from 32 to 48 weeks, and Nrf2 levels, antioxidant levels, ovarian reserve, DNA damage, and oxidative stress were examined.ResultsDMF administration resulted in elevated mRNA and protein levels of Nrf2, antioxidants, and telomere, and serum levels of Nrf2 and anti-mullerian hormone were also elevated. Results of TUNEL assay and Immunohistochemistry of mice ovarian tissues showed that DNA damage and oxidative stress were decreased by DMF administration, and significantly more oocytes were collected along with preservation of 60% more primordial follicles.ConclusionsOur data suggest that DMF administration activates the Nrf2/Keap1 pathway, elevate levels of antioxidants, and decrease DNA damage and oxidative stress, resulting in improved ovarian reserve in the mouse ovary.

Effect of cold atmospheric pressure plasma on DNA integrity in patients with asthenospermia

In this paper, we focused on the possible biomedical applications of the atmospheric pressure plasma, where we used plasma ignited in Argon (Ar) gas flow to study the effects of cold plasma on DNA integrity. Atmospheric pressure plasma, by needle, contains almost of equal amounts of positive and negative charges and has much potential for medical applications. A prospective study carried on 60 samples of human semen; each sample divided into 2 parts, before and after exposure to plasma, and then study the DNA integrity in both parts. Sperm DNA integrity was determined using a modified alkaline single cell gel electrophoresis (comet assay) and acridine orange test (AOT). The results showed that atmospheric pressure plasma give a significant (P< 0.05) decreasing in DNA damage for prepared sperm. Plasma for sperm preparation, decreases DNA damage.

GSE4 peptide suppresses oxidative and telomere deficiencies in ataxia telangiectasia patient cells.

Ataxia telangiectasia (AT) is a genetic disease caused by mutations in the ATM gene but the mechanisms underlying AT are not completely understood. Key functions of the ATM protein are to sense and regulate cellular redox status and to transduce DNA double-strand break signals to downstream effectors. ATM-deficient cells show increased ROS accumulation, activation of p38 protein kinase, and increased levels of DNA damage. GSE24.2 peptide and a short derivative GSE4 peptide corresponding to an internal domain of Dyskerin have proved to induce telomerase activity, decrease oxidative stress, and protect from DNA damage in dyskeratosis congenita (DC) cells. We have found that expression of GSE24.2 and GSE4 in human AT fibroblast is able to decrease DNA damage, detected by γ-H2A.X and 53BP1 foci. However, GSE24.2/GSE4 expression does not improve double-strand break signaling and repair caused by the lack of ATM activity. In contrast, they cause a decrease in 8-oxoguanine and OGG1-derived lesions, particularly at telomeres and mitochondrial DNA, as well as in reactive oxygen species, in parallel with increased expression of SOD1. These cells also showed lower levels of IL6 and decreased p38 phosphorylation, decreased senescence and increased ability to divide for longer times. Additionally, these cells are more resistant to treatment with H202 and the radiomimetic-drug bleomycin. Finally, we found shorter telomere length (TL) in AT cells, lower levels of TERT expression, and telomerase activity that were also partially reverted by GSE4. These observations suggest that GSE4 may be considered as a new therapy for the treatment of AT that counteracts the cellular effects of high ROS levels generated in AT cells and in addition increases telomerase activity contributing to increased cell proliferation.

BAP1 Status Determines the Sensitivity of Malignant Mesothelioma Cells to Gemcitabine Treatment.

Malignant mesothelioma (MMe) is a cancer with poor prognosis and resistance to standard treatments. Recent reports have highlighted the role of the BRCA1 associated protein 1 gene (BAP1) in the development of MMe. In this study, the chemosensitivity of human mesothelioma cell lines carrying BAP1 wild-type (WT), mutant and silenced was analysed. The BAP1 mutant cells were significantly less sensitive than BAP1 WT cell lines to the clinically relevant drug gemcitabine. Silencing of BAP1 significantly increased resistance of MMe cells to gemcitabine. Cell cycle analysis suggested that gemcitabine induced Sub-G1 phase accumulation of the BAP1 WT cells and increased in the S-phase in both BAP1 WT and mutant cells. Analysis of the role of BAP1 in apoptosis suggested that gemcitabine induced early apoptosis in both BAP1 WT and BAP1 mutant cells but with a much higher degree in the WT cells. Effects on the population of cells in late apoptosis, which can mark necrosis and necroptosis, could not be seen in the mutant cells, highlighting the possibility that BAP1 plays a role in several types of cell death. Significantly decreased DNA damage in the form of double-strand breaks was observed in gemcitabine-treated BAP1 mutant cells, compared to BAP1 WT cells under the same conditions. After BAP1 silencing, a significant decrease in DNA damage in the form of double-strand breaks was observed compared to cells transfected with scramble siRNA. Taken together, the results presented in this manuscript shed light on the role of BAP1 in the response of MMe cells to gemcitabine treatment and in particular in the control of the DNA damage response, therefore providing a potential route for more efficient MMe therapy.

Aclarubicin in subtoxic doses reduces doxorubicin cytotoxicity in human non-small cell lung adenocarcinoma (A549) and human hepatocellular carcinoma (HepG2) cells by decreasing DNA damage

In the study, the cytotoxicity and genotoxicity of aclarubicin (ACL) against A549 (human non-small cell lung adenocarcinoma) and HepG2 (human hepatocellular carcinoma) cell lines were evaluated and compared with that of doxorubicin (DOX). The effect of both anthracyclines in combination was also investigated. In order to get a deeper insight into the effectiveness of the drugs and their combination, their effects on the DNA damage and distribution of the cell cycle of A549 and HepG2 cells were investigated. After treatment with investigated compounds, apoptotic and necrotic morphological changes were estimated by double staining cells with orange acridine and ethidium bromide.The results showed that ACL was much more cytotoxic against lung (A549) and liver (HepG2) cancer cell lines than DOX. However, the drugs affected the cell cycle differently. ACL arrested cells in the G1 phase, while DOX arrested them in the G2/M phase. DOX and ACL at high concentrations are able to trigger apoptosis in both A549 and HepG2 cells. When the drugs were used in combination, subtoxic concentrations of ACL antagonized the cytotoxic effects of doxorubicin. Pre-incubation of cells with subtoxic concentrations of ACL reduced the level of DNA damage by DOX but increased DOX genotoxicity in the presence of verapamil.

The role of cytochrome P450 2E1 on ethanol-mediated oxidative stress and HIV replication in human monocyte-derived macrophages

BackgroundAlcohol consumption is considered to be a major health problem among people living with HIV/AIDS. Our previous reports have shown that ethanol reduced intracellular concentrations of antiretroviral drugs elvitegravir and darunavir in the HIV-1-infected U1 cell line. Ethanol also increased HIV-1 replication despite the presence of elvitegravir. Our previous finding has also shown that the levels of cytochrome P450 enzyme 2E1 (CYP2E1) and oxidative stress in blood monocytes were induced, while the concentration of alcohol in the plasma was reduced in HIV-1-infected alcohol users compared to uninfected alcohol users. However, the role of CYP2E1 in ethanol-enhanced oxidative stress and HIV-1 replication is still unclear. MethodsThis study examined the chronic effects (14 days) of ethanol on HIV viral load, oxidative DNA damage, expression of CYP2E1, expression of antioxidant enzymes (AOEs), expression of reactive oxygen species (ROS) in human monocyte-derived macrophages (MDM). Further, to evaluate the role of CYP2E1 in mediating ethanol-induced viral replication, CYP2E1 siRNA and CYP2E1 selective inhibitor were used in the HIV-1-infected U1 cell line following ethanol treatment. ResultsChronic ethanol exposure demonstrated an increase in oxidative DNA damage and CYP2E1 expression in both non-infected and HIV-1-infected MDM. Our results showed that ethanol chronic exposure increased HIV-1 replication by ~3-fold in HIV-1-infected MDM. This ethanol-enhanced HIV-1 replication was associated with an increased oxidative DNA damage, an increased expression of CYP2E1, and a decreased expression of antioxidant enzyme PRDX6. In HIV-1-infected U1 cell line, we observed a decreased viral replication (~30%) and a decreased DNA damage (~100%) after repression of CYP2E1 by siRNA, upon ethanol exposure. We also observed a decreased viral replication (~25%) after inhibition of CYP2E1 by using selective CYP2E1 inhibitor. ConclusionsThe data suggest that chronic ethanol exposure increases HIV-1 replication in MDM, at least in part, through CYP2E1-mediated oxidative stress. These results are clinically relevant to potentially find effective treatment strategies for HIV-1-infected alcohol users.

Epidermal Growth Factor Receptor - Mediated DNA Repair Regulates Hematopoietic Stem Cell Regeneration

Hematopoietic myelosuppression is the principal dose-limiting toxicity of curative chemotherapy and radiotherapy in patients with cancer. Both chemotherapy and radiotherapy deplete hematopoietic stem cells (HSCs) via induction of DNA damage. DNA damage contributes to both HSC dysfunction and risk for malignant transformation over time. Extrinsic signals capable of promoting DNA repair in HSCs following injury can potentially improve HSC function and may decrease risk for dysplasia and leukemia over time. Here, we show that treatment with epidermal growth factor (EGF) decreases DNA damage in murine HSCs following irradiation via activation of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and nonhomologous end joining (NHEJ) repair. Specifically, we observed that in vitro treatment of irradiated bone marrow (BM) ckit+sca-1+lin- (KSL) hematopoietic stem/progenitor cells (HSPCs) with 100 ng/ml EGF decreased HSPC DNA damage at 1 hour as measured by gamma-H2AX foci and Comet assay (p<0.0001, p<0.0001). Further, EGF treatment increased phosphorylation and nuclear localization of DNA-PKcs, which is integral to NHEJ repair, and increased phosphorylation of Akt in irradiated HSPCs. Inhibition of Akt or DNA-PKcs completely abrogated EGF - mediated DNA repair and EGF - mediated recovery of HSPCs following irradiation in vitro. In irradiated (500 cGy) C57BL/6 mice, subcutaneous administration of 10 μg/day of EGF significantly increased recovery of peripheral blood white blood cells and lymphocytes (p=0.03, p=0.04), as well as BM SLAM+KSL cells (HSCs) at day +14 (p=0.0004). Treatment with the DNA-PKcs inhibitor, NU7441, abrogated EGF - mediated recovery of peripheral blood WBCs in irradiated mice (p=0.007), suggesting that EGF - mediated mitigation of radiation injury in vivo was dependent on DNA-PKcs - mediated NHEJ repair. Importantly, EGF treatment of irradiated mice also substantially increased the survival of irradiated mice compared to irradiated control mice (p=0.009) and caused the recovery of long-term HSCs capable of 20 week competitive repopulation in congenic recipient mice (p=0.01, 20 weeks).Conversely, doxycycline inducible, cell - specific suppression of EGFR in hematopoietic cells using SCL-tTA;EGFR-DN mice caused a significant depressed regeneration of BM SLAM+KSL cells and KSL cells at day +10 after irradiation (p=0.005, p=0.02, respectively). SCL-tTA;EGFR-DN mice also displayed significantly decreased survival at day +30 compared to EGFR wild type mice (23.1% survival vs. 58.3% survival, p=0.04). Consistent with our hypothesis, BM HSPCs from EGFR-DN mice displayed increased DNA damage in response to total body irradiation, as measured by gamma H2AX foci and the Comet assay (p<0.0001, p=0.04). Coordinately, BM HSPCs from EGFR-DN mice displayed decreased phosphorylation of Akt and DNA-PKcs in response to irradiation in vitro (p<0.0001, p=0.004).These studies suggest that EGFR - mediated DNA repair is necessary for HSC and progenitor cell recovery following irradiation and that EGF treatment accelerates hematopoietic regeneration via augmentation of NHEJ repair in HSCs. EGF treatment following high dose chemotherapy and/or radiotherapy may increase HSC function and lessen near - term hematologic toxicities and morbidities. DisclosuresNo relevant conflicts of interest to declare.

Epidermal Growth Factor Rejuvenates Aging Hematopoietic Stem Cells

Aging hematopoietic stem cells display distinct abnormalities such as myeloid skewing, decreased repopulating capacity, and leukemia predisposition. Radiation exposure potentiates aging of the hematopoietic system, as evidenced by premature and persistent senescence of HSCs in mice exposed to moderate dose irradiation (Blood, 2014. 123(20): p. 3105-15). Our laboratory demonstrated that EGF promotes hematopoietic stem cell (HSC) regeneration after radiation injury (Nat Med, 2013. 19(3): p. 295-304). Based on these results, we hypothesized that EGF and EGFR signaling may have rejuvenating effects on aging HSCs.For comparisons of young and aged mice, we utilized 2-4 month old mice and >18-24 month old female C57BL/6 mice, respectively. We discovered that aged C57BL/6 mice have decreased levels of EGF in the peripheral blood compared to young mice (p= 0.02) and decreased expression of EGFR on bone marrow (BM) ckit+sca-1+lin- (KSL) stem/progenitor cells (p= 0.03). BM KSL cells from aged mice displayed increased DNA damage in culture with thrombopoietin, SCF and Flt-3 ligand (TSF) compared to young KSL cells (p< 0.0001). Aged BM KSL cells treated with 100 ng/ml EGF had decreased DNA damage compared to aged BM KSL cells treated with TSF only (p< 0.01) Treatment with EGF for 7 days increased the colony forming capacity of aged BM KSL cells (p< 0.01). Importantly, competitive repopulation assays revealed that 7 day in vitro treatment of aged BM KSL cells with EGF significantly increased primary donor cell engraftment (p< 0.01) at 16 weeks compared to aged BM KSL cells cultured with TSF alone.Based on these in vitro observations, we next tested whether systemic administration of a recombinant, pegylated EGF (pEGF), 20 mcg three times weekly for 4 weeks could alter the hematopoietic characteristics of aged C57BL/6 mice. pEGF treatment decreased BM myeloid skewing (p< 0.01) and increased CD3 T cell content (p< 0.01) in aged mice compared to saline-injected controls. Furthermore, competitive repopulation assays revealed that pEGF treatment of aged mice caused a significant increase in functional HSCs capable of competitive engraftment in recipient congenic mice. Total primary donor cell (p< 0.01), myeloid cell (p< 0.01), T cell (p< 0.01) and B cell (p= 0.01) engraftment were significantly increased in recipient mice transplanted with BM cells from aged, pEGF-treated donors compared to recipients of aged, saline-treated donors at 20 weeks. Additionally, secondary transplantation also resulted in increased donor engraftment (p< 0.01) and increased donor myeloid engraftment (p< 0.05) at 12 weeks. These results suggested that systemic administration of pEGF rejuvenates the repopulating ability of aged HSCs.We next sought to determine if deficiency in EGFR signaling could accelerate hematopoietic aging in mice. For this purpose, we utilized a doxycycline-inducible, hematopoietic cell specific EGFR dominant negative mutant model (SCL-tTA;EGFR-DN mice). Interestingly, adult (13 month old) SCL-tTA;EGFR-DN mice displayed increased myeloid skewing in the peripheral blood (p< 0.05) and BM cells from SCL-tTA;EGFR-DN mice generated significantly decreased colony forming cells, compared to age-matched, EGFR-expressing mice (p< 0.01). Mechanistically, BM KSL cells from 13 month old SCL-tTA;EGFR-DN mice demonstrated increased senescence (p< 0.01) and increased expression of p16 (p< 0.05) compared to young mice. In contrast, BM KSL cells from aged C57BL/6 mice that were treated with 100 ng/ml EGF (plus TSF) for 7 days displayed decreased senescence and decreased p16 expression compared to BM KSL cells from aged mice treated with TSF alone. Taken together, these studies suggest that EGF/EGFR signaling declines with age and that reactivation of EGFR signaling via EGF treatment can ameliorate clinically relevant features of hematopoietic aging, including HSC self-renewal capacity. DisclosuresNo relevant conflicts of interest to declare.

Critical Role for Apobec and Its Interacting Partners in Mediating Mutations and Cell Growth in Multiple Myeloma (MM)

The APOBEC family of cytidine deaminases include AID (activity induced deaminase) and 10 related APOBEC enzymes (A1,A2,A3A,A3B,A3C,A3D,A3F,A3G,A3H and A4). AID is well studied for its role in somatic hyper mutation and class switch recombination of immunoglobulin genes. APOBECs (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like) have been shown to have roles in mRNA editing and in antiviral immunity. Recently, a causal role for the AID/APOBECs in inducing somatic mutations in myeloma has been proposed and we have previously published that APOBEC signature mutations as a frequent event in Myeloma. We have also observed that APOBEC-mediated mutations may account for mutations associated with progression of smoldering myeloma to MM.We further investigated the role of APOBEC in genomic changes in MM and observed that APOBEC expression and activity is elevated in myeloma cell lines as well as patient samples. Using knockdown and over expression approaches, we showed that depletion of APOBECs in myeloma cell lines reduces genomic instability. Following APOBEC3G knock down we observed decreased DNA damage (by g-H2AX), decrease in acquisition of new copy number events over time, and reduced mutational load, strongly suggesting that inhibiting APOBECs could be a potential approach to reduce genome evolution in MM.We next investigated the effect of APOBEC inhibition on myeloma cell proliferation. We observed that Sh-RNA-based APOBEC knock down in MM1S and H929 MM cell lines, led to significant inhibition of MM cell proliferation, and induction of apoptotic cell death. Associated with APOBEC knockdown, we also observed increased levels of Cyclin-dependent kinase inhibitor 1B (p27Kip1) at both RNA and protein level. By immunoprecipitation we found that APOBEC3G interacts and inhibits the RNA binding protein DEAD-END 1 (DND1), thereby preventing it from inhibiting miR-221-mediated targeting of p27 transcripts. Knockdown of DND1, or over-expression of miR-221 in APOBEC-depleted cells rescued the cell proliferation defects with concomitant decrease in p27 levels. These results show that APOBCs bind to and sequester DND1, leading to miR-221-mediated depletion of p27. In the absence of APOBEC, DND1 prevents the degradation of p27 mRNA, leading to elevated p27 levels and inhibition of cell cycle, suggesting a role for APOBECs in regulating MM cell proliferation that might be independent of its RNA/DNA mutator function. Taken together, these results indicate a significant functional role for APOBECs both in genome evolution as well as cell growth in myeloma and may constitute an important therapeutic target. DisclosuresMunshi:OncoPep: Other: Board of director.

Dual actions of the antioxidant chlorophyllin, a glutathione transferase P1-1 inhibitor, in tumorigenesis and tumor progression.

Glutathione (GSH) and enzymes related to this antioxidant molecule are often overexpressed in tumor cells and may contribute to drug resistance. Blockade of glutathione transferases (GSTs) has been proposed to potentiate the efficacy of chemotherapeutic drugs in cancer. The aim of this study was to evaluate the effect of chlorophyllin that has antioxidant properties, and also interferes with the activity of GST P1-1, on breast cancers in vitro and in vivo. The in vivo studies were conducted using an N-methyl- N-nitrosourea (MNU)-induced chemical carcinogenesis model in laboratory rats. DNA damage, GST activity, and GSH levels were determined in liver and tumor tissues. Treatment with chlorophyllin increased the GSH levels in the liver and significantly decreased DNA damage in the blood, liver, and tumor tissues. Even though tumorigenesis was delayed in rats receiving chlorophyllin before MNU injections, once the tumors emerged, the progression of tumor appeared to be faster than in the animals that received the carcinogen only. Out of nine breast cell lines, GST P1-1 expression was detected in MCF-12A, MDA-MB-231, and HCC38. Concomitant incubation with chlorophyllin and docetaxel did not significantly affect cell proliferation and viability. Chlorophyllin displayed genoprotective effects that initially delayed tumorigenesis. However, once the tumors were established, it may act as a promoter that facilitates tumor growth, potentially by a mechanism independent of cell proliferation and viability. Our results underline the pros and cons of antioxidant treatment in cancer, even if it has a capacity to inhibit GST P1-1.

Hypovitaminosis D exacerbates the DNA damage load in human uterine fibroids, which is ameliorated by vitamin D3 treatment.

Uterine fibroids (UFs) are the most common benign neoplastic threat to women's health and associated with DNA damage and genomic instability. Hypovitaminosis D is a known risk factor for UFs, especially among African Americans. Vitamin D3 has been shown to effectively inhibit UF phenotype, but its mechanisms remain unclear. We hypothesize that Vitamin D3 ameliorates UFs by recovering the damaged DNA repair system, thus inhibits tumor progression. We compared the DNA damage status and Vitamin D receptor (VDR) expression between normal myometrial and UF primary cells. Unrepaired DNA double-strand breaks (DSBs) accumulated but VDR expression decreased in UFs. The RNA and protein levels of key DNA repair members belonging to DNA DSB sensors (MRE11, NBS1, RAD50), mediators and effectors (CHECK2, BRCA1, RAD51) were downregulated in UFs compared with myometrial cells. VDR KD induced DSB accumulation and DNA damage response (DDR) defects in myometrial cells. Using the DNA damage PCR array, the expression of many additional DNA repair genes was downregulated in VDR KD cells. Treatment of UF cells with Vitamin D3 (100 nM) significantly decreased DNA damage and restored DDR concomitant with VDR induction. Notably, the PCR array demonstrated that among 75 downregulated genes after VDR KD, 67 (89.3%) were upregulated after vitamin D3 treatment. These studies demonstrate a novel link between DNA damage and the vitamin D3/VDR axis in UFs. Vitamin D3 suppresses the UF phenotype through orchestrated targeting at multiple molecules in DNA repair pathways, thus offering novel mechanistic insights into the clinical effectiveness of vitamin D3 on UFs.

Novel therapeutic approaches to xeroderma pigmentosum.

The study of xeroderma pigmentosum has yielded unforeseen advances regarding how defects in the nucleotide excision repair pathway result in this devastating disease, but development of therapeutic strategies has trailed behind the mechanistic discoveries. This review aims to cover clinical presentation, molecular mechanisms and current management, and highlights more recent insights into targeting the deficiencies secondary to the DNA repair defects to prevent skin cancer and/or neurological degeneration. This review article discusses novel therapeutic approaches to xeroderma pigmentosum that focus on metabolic defects downstream of nucleotide excision repair. Current research demonstrates that specific sulfonylureas promote clearance of DNA damage and increase resistance to ultraviolet radiation in a cellular model of xeroderma pigmentosum. Moreover, nicotinamide attenuates the effects of ultraviolet radiation in cells, and caloric restriction decreases DNA damage burden in animal models of xeroderma pigmentosum. Clinical management of patients with xeroderma pigmentosum still focuses on preventative avoidance of sun exposure as opposed to therapies that would improve the patients' condition; thus, novel approaches to this disease are warranted.

Campylobacter jejuni promotes colorectal tumorigenesis through the action of cytolethal distending toxin

ObjectiveCampylobacter jejuni produces a genotoxin, cytolethal distending toxin (CDT), which has DNAse activity and causes DNA double-strand breaks. Although C. jejuni infection has been shown to promote intestinal inflammation, the...

Stimulation of AMPK prevents degeneration of photoreceptors and the retinal pigment epithelium

Retinal degenerative diseases are generally characterized by a permanent loss of light-sensitive retinal neurons known as photoreceptors, or their support cells, the retinal pigmented epithelium (RPE). Metabolic dysfunction has been implicated as a common mechanism of degeneration. In this study, we used the drug metformin in a gain-of-function approach to activate adenosine monophosphate-activated protein kinase (AMPK). We found that treatment protected photoreceptors and the RPE from acute injury and delayed inherited retinal degeneration. Protection was associated with decreased oxidative stress, decreased DNA damage, and increased mitochondrial energy production. To determine whether protection was a local or a systemic effect of metformin, we used AMPK retinal knockout mice and found that local expression of AMPK catalytic subunit α2 was required for metformin-induced protection. Our data demonstrate that increasing the activity of AMPK in retinal neurons or glia can delay or prevent degeneration of photoreceptors and the RPE from multiple types of cell-death triggers.

The nuclear receptor RXRA controls cellular senescence by regulating calcium signaling.

Calcium signaling is emerging as a key pathway controlling cellular senescence, a stable cell proliferation arrest playing a fundamental role in pathophysiological conditions, such as embryonic development, wound healing, cancer, and aging. However, how calcium signaling is regulated is still only partially understood. The inositol 1, 4, 5‐trisphosphate receptor type 2 (ITPR2), an endoplasmic reticulum calcium release channel, was recently shown to critically contribute to the implementation of senescence, but how ITPR2 expression is controlled is unclear. To gain insights into the regulation of ITPR2 expression, we performed an siRNA screen targeting 160 transcription factors and epigenetic regulators. Interestingly, we discovered that the retinoid X receptor alpha (RXRA), which belongs to the nuclear receptor family, represses ITPR2 expression and regulates calcium signaling though ITPR2 and the mitochondrial calcium uniporter (MCU). Knockdown of RXRA induces the production of reactive oxygen species (ROS) and DNA damage via the ITPR2‐MCU calcium signaling axis and consequently triggers cellular senescence by activating p53, whereas RXRA overexpression decreases DNA damage accumulation and then delays replicative senescence. Altogether, our work sheds light on a novel mechanism controlling calcium signaling and cellular senescence and provides new insights into the role of nuclear receptors.

Hibiscus acetosella extract protects against alkylating agent-induced DNA damage in mice

Hibiscus acetosella was shown to exert beneficial effects in humans and animal models however, the effects of this plant on DNA are unknown. The aim of this study was to determine the antigenotoxic and antimutagenic effects of H. acetosella extracts on alkylating agent methyl methanesulfonate (MMS) in vivo in mice. Initially, we performed analysis of phenolic compounds in extracts of H. acetosella by high-performance liquid chromatography (HPLC). Next, mice were divided into 8 groups and treated with distilled water or plant extract (0.1 ml/10 g) by gavage for 15 days, followed by intraperitoneal (ip) administration of saline solution or MMS (40 mg/Kg b.w) on day 16. Caffeic acid, following by gallic acid, gallocatechin, coumaric acid, and 3,4-dihydroxybenzoic acid were found to be present in extracts of H. acetosella leaves. In peripheral blood analysis of groups receiving pretreatment with H. acetosella at doses of 50 or 100 mg/kg plus MMS decreased DNA damage as evidenced by comet assay and Micronucleus assays relative to MMS alone. These results suggested that H. acetosella extracts exerted protective effects dose dependent against genotoxicity and mutagenicity induced by alkylating agents.

Small-for-pregnancy-age rats submitted to exercise: DNA damage in mothers and newborns, measured by the comet assay

BackgroundFetal impairment caused by a deleterious intrauterine environment may have long-term consequences, such as oxidative stress and genetic damage. Rats born as small-for-gestational-age (SPA) were submitted to exercise (swimming) before and during pregnancy. The animals exhibited glucose intolerance, reduced general adiposity, and increased maternal and offspring organ weight, showing the benefit of exercise for these rats. We hypothesised that regular exercise in SPA during gestation could prevent DNA damage in these animals and in their offspring, contributing to altered fetal programming of metabolism in the offspring. Severe diabetes was induced by streptozotocin treatment, to obtain SPA newborns. At adulthood, pregnant SPA rats were randomly distributed into two groups: exercised (SPAex – submitted to swimming program) or not-exercised (SPA – sedentary rats). Post-partum, blood was collected for analysis of DNA damage (comet assay) and oxidative stress. SPAex rats presented lower DNA damage levels, decreased lipid peroxidation, and a lower rate of newborns classified as large-for-pregnancy-age. DNA damage was also lower in SPAex newborns. We conclude that swimming applied to SPA pregnant rats contributes to decreased DNA damage and lipid peroxidation in the dams, and decreased DNA damage and macrosomia in their offspring.

Protective Effects of Ursolic Acid in the Kidneys of Diabetic Rats

Diabetes, a heteregenous metabolic and chronic disease, is a growing health problem in most countries. It has been claimed that diabetes is associated with the increased formation of free radicals and decreased in antioxidant potential. Oxidative stress formed in diabetes may cause DNA damage in the tissues. Ursolic acid, a well-known pentacylic triterpene, is commonly used in traditional Chinese medicine due to its beneficial health effects such as antioxidant, anticancer, and antiulcer properties. The aim of this study was to investigate the effects of ursolic acid in the kidneys of Wistar albino rats with streptozotocin-induced diabetes. DNA damage was evaluated in the kidney cells of rats using alkaline comet assays. Oxidative stress parameters such as CAT, SOD, GR, and GSH-Px enzyme activities and total GSH and MDA levels were also evaluated. Ursolic acid treatment was found to significantly decrease DNA damage, GR enzyme activities, and MDA levels, and significantly increase GSH levels and CAT, SOD and GSH-Px enzyme activities in diabetic rats. According to our results, it seems that ursolic acid may be beneficial against diabetes-induced renal damage.