scholarly journals Oxidative damage, inflammation, genotoxic effect, and global DNA methylation caused by inhalation of formaldehyde and the purpose of melatonin

2020 ◽  
Vol 9 (6) ◽  
pp. 778-789
Author(s):  
Letícia Bernardini ◽  
Eduardo Barbosa ◽  
Mariele Feiffer Charão ◽  
Gabriela Goethel ◽  
Diana Muller ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Bone Marrow ◽  
Oxidative Damage ◽  
Bronchoalveolar Lavage ◽  
Bone Marrow Cells ◽  
Histological Study ◽  
Global Dna Methylation ◽  
Protective Effects ◽  
Global Methylation ◽  
Protein Thiols

Abstract Formaldehyde (FA) exposure has been proven to increase the risk of asthma and cancer. This study aimed to evaluate for 28 days the FA inhalation effects on oxidative stress, inflammation process, genotoxicity, and global DNA methylation in mice as well as to investigate the potential protective effects of melatonin. For that, analyses were performed on lung, liver and kidney tissues, blood, and bone marrow. Bronchoalveolar lavage was used to measure inflammatory parameters. Lipid peroxidation (TBARS), protein carbonyl (PCO), non-protein thiols (NPSH), catalase activity (CAT), comet assay, micronuclei (MN), and global methylation were determined. The exposure to 5-ppm FA resulted in oxidative damage to the lung, presenting a significant increase in TBARS and NO levels and a decrease in NPSH levels, besides an increase in inflammatory cells recruited for bronchoalveolar lavage. Likewise, in the liver tissue, the exposure to 5-ppm FA increased TBARS and PCO levels and decreased NPSH levels. In addition, FA significantly induced DNA damage, evidenced by the increase of % tail moment and MN frequency. The pretreatment of mice exposed to FA applying melatonin improved inflammatory and oxidative damage in lung and liver tissues and attenuated MN formation in bone marrow cells. The pulmonary histological study reinforced the results observed in biochemical parameters, demonstrating the potential beneficial role of melatonin. Therefore, our results demonstrated that FA exposure with repeated doses might induce oxidative damage, inflammatory, and genotoxic effects, and melatonin minimized the toxic effects caused by FA inhalation in mice.

scholarly journals Influence of tissue, age, and environmental quality on DNA methylation in Alligator mississippiensis

Reproduction ◽  
2014 ◽  
Vol 147 (4) ◽  
pp. 503-513 ◽  
Author(s):  
Benjamin B Parrott ◽  
John A Bowden ◽  
Satomi Kohno ◽  
Jessica A Cloy-McCoy ◽  
Matthew D Hale ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Whole Blood ◽  
Epigenetic Modification ◽  
Alligator Mississippiensis ◽  
Sentinel Species ◽  
Global Dna Methylation ◽  
American Alligator ◽  
Global Methylation ◽  
Contaminant Exposure ◽  
Liquid Chromatography Tandem Mass

Epigenetic modifications are key mediators of the interactions between the environment and an organism's genome. DNA methylation represents the best-studied epigenetic modification to date and is known to play key roles in regulating transcriptional activity and promoting chromosome stability. Our laboratory has previously demonstrated the utility of the American alligator (Alligator mississippiensis) as a sentinel species to investigate the persistent effects of environmental contaminant exposure on reproductive health. Here, we incorporate a liquid chromatography–tandem mass spectrometry method to directly measure the total (global) proportion of 5-methyl-2′-deoxycytidine (5mdC) in ovarian and whole blood DNA from alligators. Global DNA methylation in ovaries was significantly elevated in comparison with that of whole blood. However, DNA methylation appeared similar in juvenile alligators reared under controlled laboratory conditions but originating from three sites with dissimilar environmental qualities, indicating an absence of detectable site-of-origin effects on persistent levels of global 5mdC content. Analyses of tissues across individuals revealed a surprising lack of correlation between global methylation levels in blood and ovary. In addition, global DNA methylation in blood samples from juvenile alligators was elevated compared with those from adults, suggesting that age, as observed in mammals, may negatively influence global DNA methylation levels in alligators. To our knowledge, this is the first study examining global levels of DNA methylation in the American alligator and provides a reference point for future studies examining the interplay of epigenetics and environmental factors in a long-lived sentinel species.

DNMT3b’s Role in Hematopoietic Stem Cell Self-Renewal.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1406-1406
Author(s):  
Matthew J Boyer ◽  
Feng Xu ◽  
Hui Yu ◽  
Tao Cheng
Keyword(s):  
Dna Methylation ◽  
Bone Marrow ◽  
Stem Cell ◽  
Bone Marrow Transplantation ◽  
Peripheral Blood ◽  
Marrow Transplantation ◽  
Bone Marrow Cells ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Marrow Cells

Abstract DNA methylation is an epigenetic means of gene regulation and is carried out by a family of methyltransferases of which DNMT1 acts to maintain methylation marks following DNA replication and DNMT3a and DNMT3b methylate DNA de novo. DNMT3b has been shown to be essential for mammalian development and necessary for differentiation of germline and neural progenitor cells. Mutations of DNMT3b in humans lead to a rare autosomal recessive disorder characterized by immunodeficiency, centromeric instability, and facial abnormalities. We have shown by real-time, RT-PCR that DNMT3b mRNA is uniquely over-expressed by approximately 30-fold in immunophenotypically-defined longterm repopulating hematopoietic stem cells (HSCs) that are CD34−lineage−c-kit+Sca-1+ as compared to progenitor and differentiated cell types within the bone marrow and with respect to the other members of the DNMT family, namely DNMT1 and DNMT3a. To determine DNMT3b’s function in HSCs competitive bone marrow transplantation was undertaken. Isolated lineage− enriched bone marrow cells were transduced with a retroviral backbone based on the Murine Stem Cell Virus (MSCV) carrying either GFP and a short, hairpin RNA (shRNA) targeting DNMT3b or GFP alone. Following transduction 1×105 GFP+ cells along with 1×105 competitor cells were transplanted into 9.5 Gray irradiated congenic recipients. Two months following transplantation mice receiving bone marrow cells transduced with DNMT3b shRNA showed a significantly lower engraftment of donor cells as a percentage of total competitor cell engraftment in the peripheral blood as compared to those receiving cells transduced with GFP alone (24.8 vs 3.7, p<0.05) which persisted at 3 months (22.8 vs 1.5, p<0.05). Similarly, within the donor derviced cells in the peripheral blood there was a lower percentage of myeloid (CD11b+) cells at 2 and 3 months in the recipients of DNMT3b shRNA transduced cells as compared to controls. However there was no observed difference in the percentage of peripheral B (CD45R+) or T (CD3+) cells within the donor-derived cells. To determine the mechanism behind the observed engraftment defect with DNMT3b knockdown we cultured GFP+ transduced bone marrow cells in vitro with minimal cytokine support. As a control for our targeting methodology we also transduced bone marrow cells from mice harboring two floxed DNMT3b alleles with a MSCV carrying Cre recombinase and GFP. While lineage− bone marrow cells transduced with GFP alone increased 10-fold in number over two weeks of culture, cells in which DNMT3b was down regulated by shRNA or Cre-mediated recombination only doubled. Culture of lineage− bone marrow cells in methylcellulose medium by the colony-forming cell (CFC) assay revealed increases in the granulocytic and total number of colonies with DNMT3b knockdown or Cre-mediated recombination of DNMT3b similar to the increased myeloid engraftment of DNMT3b shRNA transduced cells observed 1 month following competitive bone marrow transplantation. However when 5,000 of these cells from the first CFC assay were sub-cultured there was a significant loss of colony forming ability within all lineages when DNMT3b was targeted by shRNA or Cre-mediated recombination. Taken together with the decreased engraftment of DNMT3b shRNA cells following competitive bone marrow transplantation, the observed limited proliferation in liquid culture and loss of colony forming ability during serial CFC assays is suggestive of a self-renewal defect of HSCs in the absence of DNMT3b, that was previously only reported in the absence of both DNMT3a and DNMT3b. Further elucidation of this proposed self-renewal defect is being undertaken and results of ongoing studies including long-term culture initiating cell (LTC-IC) assays and identification of genomic sites of DNA methylation within different hematopoietic subsets will also be presented.

Global and HOX Gene DNA Methylation In Normal Karyotype Acute Myeloid Leukemia: Clinical Implications and Molecular Correlations

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 231-231
Author(s):  
Stefan Deneberg ◽  
Andreas Lennartsson ◽  
Bertil Uggla ◽  
Verena Gaidzik ◽  
Andrea Corbacioglu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Bone Marrow ◽  
Hox Genes ◽  
Normal Karyotype ◽  
Global Methylation ◽  
Hox Gene ◽  
Mutational Status ◽  
Average Methylation ◽  
Wbc Count

Abstract Abstract 231 Background: DNA methylation induces gene silencing in a nonrandom fashion in many types of malignancies. In AML there is no consensus regarding the clinical implications of DNA methylation on a global level. Normal karyotype AML (NK AML) can be prognostically stratified by molecular mutations in genes such as FLT3 and NPM1. We have previously reported that increased gene promoter methylation levels may have positive prognostic implications in AML. Now we have focused and expanded our analysis in a homogenous group of NK AML in an effort to illuminate these issues. Methods and patients: We analyzed genome wide DNA methylation signatures from the diagnostic bone marrow samples of 58 de novo NK AML with the IlluminaHuman27 Methylation array, covering 27000 CpG sites, mainly located in the promoter regions of 15000 individual genes. Global methylation was defined as the average methylation values of all measured CpG sites in the specific sample. All patients were between 18 and 67 years of age and received standard induction chemotherapy. All were eligible for intensive consolidation therapy including allogeneic transplantation. FACS sorted normal bone marrow separated into four stages of myeloid differentiation were analyzed as normal controls. Methylation data were correlated to clinical outcomes and molecular mutational status of NPM1 and FLT3. Functional annotation analyses were performed on differentially methylated genes to find epigenetically perturbed pathways. Further molecular analysis of CEBPA, IDH1 and IDH2 is currently performed. Results and discussion: Global methylation levels varied substantially between AML samples but remained mainly unchanged during normal myeloid differentiation. Methylation levels were significantly higher in AML cases than in the normal myeloid progenitors (p<0.001). There were no correlations between methylation levels, age, white blood cell (WBC) count or bone marrow blast percentage at diagnosis. Increased global methylation correlated significantly to increased in vivo chemosensitivity and to patient survival. CR rates after one course of induction chemotherapy were 64% in the quartile of patients with the highest level of global DNA methylation compared to 32% in others (HR 3.9, p=0.04). Median overall survival of patients of the most methylated quartile was not reached and significantly longer than the 16 months median survival of the least methylated quartile (p=0.001) (see figure below). In a Cox Regression model stratified for FLT3-mutational status, methylation level was an independent prognostic factor for survival (p=0.01) together with WBC count (p=0.01). A functional annotation analysis revealed that NPM1 wild type samples had an enrichment of Homeobox (HOX) gene methylation as compared to NPM1 mutated cases (p=1.7×10−11), providing a mechanism for the previously described difference in HOX gene expression in NPM1 wt vs. NPM1 mutated AML. The average methylation levels of all HOX genes were higher in wild type NPM1 cases compared to NPM1 mutated cases (p=0.02). Additional gene expression array data in our cohort of patients also showed methylation levels of individual HOX genes to be correlated inversely with expression. HOX genes were also enriched in non-FLT3ITD cases compared to FLT3ITD-cases, however, not to a statistically significant degree. In conclusion we show that global DNA methylation levels are predictive of response to chemotherapy and an independent prognostic factor for survival in normal karyotype AML. Furthermore, our results suggest that HOX gene methylation may be the mechanism underlying the previously known difference in HOX-gene expression between NPM1 mutated and unmutated AML cases. Disclosures: Nahi: Jansen-Cilag: Honoraria; Celgene: Honoraria.

Dnmt3a Deletion and FLT3-ITD Cooperate in a Mouse Model of T-Lymphoblastic Leukemia (T-ALL).

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2428-2428
Author(s):  
Liubin Yang ◽  
Min Luo ◽  
Mira Jeong ◽  
Choladda V. Curry ◽  
Grant Anthony Challen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
T Cell ◽  
Bone Marrow Cells ◽  
Lymphoblastic Leukemia ◽  
Notch Pathway ◽  
Leukemic Transformation ◽  
Aberrant Dna Methylation

Abstract Abstract 2428 Aberrant DNA methylation repeatedly has been implicated in cancer development. DNA methyltransferase (DNMT) 3A, which mediates de novo DNA methylation, was found to be mutated in 20% of patients with acute myeloid leukemia and 10% of patients with myelodysplastic syndrome. Recently, mutations associated with myeloid malignancies such as DNMT3A and FLT3 have also been uncovered in patients with early T-cell precursor lymphoblastic leukemia (ETP-ALL) (Neumann et al., 2012; Van Vlierberghe et al., 2011; Zaremba et al., 2012). ETP-ALL is a type of very high-risk ALL associated with myeloid/stem cell gene expression signature and myeloid markers. We have demonstrated that Dnmt3a deletion in mouse causes increased self-renewal of hematopoietic stem cells and an impairment of differentiation (Challen et al., 2011). Dnmt3a loss also produces aberrant methylation associated with oncogenes and tumor suppressor genes. Yet, whether aberrant DNA methylation can drive leukemia remains unknown. As Dnmt3a deletion alone was insufficient for malignancy, secondary mutations are likely necessary for leukemic transformation. Because FLT3 internal tandem duplication (ITD) frequently co-exist with DNMT3A mutations in acute leukemias, we hypothesized that Dnmt3a-loss may cooperate with FLT3-ITD to promote leukemic transformation; and we established a mouse model to test this. Deletion of conditional Dnmt3a with Mx1-cre was induced by injections of pIpC. Subsequently, bone marrow from Dnmt3a-deleted (Dnmt3aKO) donor mice was transduced with MSCV-FLT3-ITD-GFP retrovirus or MSCV-GFP control and transplanted into lethally irradiated recipients. The mice were monitored monthly for development of malignancies by complete blood count and peripheral blood analysis by flow cytometry and followed for disease latency. Moribund mice were sacrificed and analyzed with peripheral blood smears, histology, and immunophenotyping. Dnmt3a deletion with overexpression of FLT3-ITD caused rapid onset T-ALL in 6/8 mice (n=6) with a median latency of 78 days compared to 121 days in WT mice (n=4) overexpressing FLT3-ITD (p&lt;0.0001 Log-rank Mantel-Cox Test) (See figure). Mice from both groups exhibited leukocytosis, splenomegaly, and thymomegaly with high GFP expression detected by FACS. Even after we transduced bone marrow cells enriched for myeloid progenitor and stem cells, Dnmt3a deletion again accelerated T-ALL with median survival of 89 days (n=9) versus 110 days in WT-FLT3-ITD (n=10) mice. T-ALL was observed in 2/4 WT-FLT3-ITD mice and 5/6 Dnmt3aKO-FLT3-ITD mice analyzed (p&lt;0.0001 Log-rank Mantel-Cox Test). By flow cytometry, two distinct types of T-ALL were observed in the bone marrow of Dnmt3a deleted leukemic mice: one was characterized by a double positive population (DP) of CD4+CD8+ lympoblasts (1/6) and another early immature T-cell-like type of CD4-CD8-CD44+CD25-CD11bloCD117+ lymphoblasts (4/6). Gene expression analysis by RT-PCR in the early immature T-ALL showed downregulation of Notch-pathway genes (such as Notch1, Notch 3, Deltex, Hes1) and upregulation of stem cell-associated genes Lyl1 and Scl1, suggesting an ETP-like T-ALL. The ETP-like ALL phenotype has not been seen in WT mice overexpressing FLT3-ITD. The opposite gene expression pattern was seen in the DP population with upregulation of Notch-pathway genes. Furthermore, the DP leukemia was transplantable to secondary recipients within 2 weeks. Whether ETP-like ALL can be transplanted is still under investigation. We are also currently studying the changes in global CpG methylation among the leukemias that have Dnmt3a loss, FLT3-ITD overexpression, and control and also anticipate data from transcriptome analysis by RNA-Seq. These data suggest that stem or progenitor bone marrow cells primed by early loss of Dnmt3a are transformed into DP T-ALL and ETP-like ALL fueled by the overexpression of the oncogene FLT3-ITD. The ETP-like ALL phenotype has not been seen previously in WT mice overexpressing FLT3-ITD, suggesting that Dnmt3a ablation is required. The Dnmt3a-deleted-FLT3-ITD mice with T-ALL is, to our knowledge, the first animal model of human immature T-cell leukemia. This model can enhance our understanding of the pathogenesis of ETP-like ALL with respect to aberrant DNA methylation and will serve as a powerful tool to test novel therapeutic strategies. Disclosures: No relevant conflicts of interest to declare.

Dnmt3a is Required For Aberrant Self-Renewal Driven by PML-RARA

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 477-477
Author(s):  
Christopher B Cole ◽  
Angela M. Verdoni ◽  
David H Spencer ◽  
Timothy J. Ley
Keyword(s):  
Dna Methylation ◽  
Bone Marrow ◽  
Progenitor Cells ◽  
Bone Marrow Cells ◽  
Ex Vivo ◽  
Myeloid Cells ◽  
Cd11b Expression ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Marrow Cells

We previously identified recurrent mutations in the DNA methyltransferase DNMT3A in patients with acute myeloid leukemia (AML). DNMT3A and the highly homologous gene DNMT3B encode the two methyltransferases that are primarily responsible for mediating de novo methylation of specific CpG residues during differentiation. Loss of Dnmt3a in hematopoietic stem cells impairs their ability to differentiate into committed progenitors (Challen et al Nat Gen 44:23, 2011). Importantly, DNMT3A mutations are mutually exclusive of the favorable prognosis AML-initiating translocations, including the t(15;17) translocation (which creates the PML-RARA fusion gene), and translocations involving MLL. PML-RARA has been shown to interact with DNMT3A in vitro (Di Croce et al Science 295:1079,2002), and to require DNMT3A to induce methylation and transcriptional silencing of a subset of specific target genes. These findings, and the lack of DNMT3A mutations in APL patients, suggest that PML-RARA may require functional DNMT3A to initiate leukemia. To investigate this possibility, we utilized a well-characterized transgenic mouse model (in a pure B6 background) in which expression of PML-RARA is driven in hematopoietic stem/progenitor cells by the mouse Cathepsin G locus (Ctsg-PML-RARA+/- mice). These mice spontaneously develop acute promyelocytic leukemia (APL) with high penetrance and long latency, and also exhibit a preleukemic phenotype marked by the accumulation of myeloid cells in bone marrow and spleen. In addition, myeloid progenitor cells derived from these mice have the ability to serially replate in methylcellulose cultures, demonstrating aberrant self-renewal. We generated Ctsg-PML-RARA+/- mice lacking Dnmt3a (PML-RARA+/- x Dnmt3a-/-) as well as mice in which conditional ablation of Dnmt3b in hematopoietic cells is driven by Vav-Cre (PML-RARA+/- x Dnmt3b fl/fl x Vav-Cre+). Loss of Dnmt3a completely abrogated the ex vivo replating ability of PML-RARA bone marrow (Figure 1). Although colonies from both PML-RARA+/- and PML-RARA+/- x Dnmt3a-/- mice appeared similar in morphology and number on the first plating, PML-RARA+/- x Dnmt3a-/- marrow ceased to form colonies with subsequent replating (see Figure), and cultured cells lost the expression of the myeloid marker CD11b. The same phenotype was also observed using bone marrow from both genotypes that was secondarily transplanted into wild type recipients, indicating that it is intrinsic to transplantable hematopoietic progenitors. Reintroduction of DNMT3A into bone marrow cells derived from PML-RARA+/- x Dnmt3a-/- mice with retroviral transduction restored replating ability and CD11b expression. Competitive repopulation experiments with PML-RARA+/- x Dnmt3a-/- marrow revealed a decreased contribution to peripheral lymphoid and myeloid cells at 4 weeks, relative to PML-RARA+/- or WT control animals. Finally, 12 weeks after transplantation, recipients of PML-RARA+/- x Dnmt3a-/- bone marrow did not display an accumulation of myeloid cells in the bone marrow and spleen. Importantly, bone marrow from PML-RARA+/- x Dnmt3b fl/fl x Vav-Cre+/- mice displayed no replating deficit or loss of CD11b expression ex vivo, indicating different functions for Dnmt3a versus Dnmt3b in this model. Finally, we interrogated the effect of Dnmt3a loss on bone marrow DNA methylation patterns using a liquid phase DNA capture technique that sampled ∼1.9 million mouse CpGs at >10x coverage. Loss of Dnmt3a caused a widespread loss of DNA methylation in whole bone marrow cells, with 36,000 CpGs that were highly methylated (methylation value >0.7) in the PML-RARA+/- and WT mice, but hypomethylated (methylation value <0.4) in Dnmt3a-/- and PML-RARA+/- x Dnmt3a-/- mice. Characterization of the effect of Dnmt3a loss on leukemia latency, penetrance, and phenotype in PML-RARA+/- mice is currently being defined in a tumor watch. In summary, we have demonstrated that PML-RARA requires functional Dnmt3a (but not Dnmt3b) to drive aberrant self-renewal of myeloid progenitors ex vivo, and that loss of Dnmt3a leads to widespread DNA hypomethylation in bone marrow cells, and abrogates preleukemic changes in mice expressing PML-RARA. This data may explain why DNMT3A mutations are not found in patients with APL initiated by PML-RARA. Disclosures: No relevant conflicts of interest to declare.

scholarly journals A maternal effect regulates global DNA methylation patterns

2020 ◽  
Author(s):  
Remco Loos ◽  
Valeria Carola ◽  
Enrica Audero ◽  
Elena Brini ◽  
Luisa Lo Iacono ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Maternal Effect ◽  
Cpg Island ◽  
Inbred Mouse ◽  
Mouse Strains ◽  
Global Dna Methylation ◽  
Global Methylation ◽  
Genome Methylation ◽  
Genetic And Environmental Factors ◽  
Methylation Patterns

AbstractVariation in DNA methylation between individuals has been shown to be influenced by both genetic and environmental factors. However, the relative impact of genetic and non-genetic factors on DNA methylation patterns across the mammalian genome has not been systematically studied. We performed whole-genome methylation analysis in two inbred mouse strains, revealing striking differences in the global distribution of DNA methylation. Although global methylation patterns were indistinguishable for most genomic features, a significant increase in the number of unmethylated CpG-island promoters and first exons was observed between strains. Experiments using F1 reciprocal hybrid strains demonstrated that the genotype of the mother dictated global DNA methylation patterns. Cross-fostering experiments ruled out a postnatal maternal effect on these differences and suggested that they were driven by a prenatal maternal effect, possibly via differential deposition of maternal gene products into the oocyte or uterine environment. These data demonstrate that maternal effects have a major impact on global DNA methylation patterns.

scholarly journals Protective Effects of Spirulina against Cyclophosphamide Induced Bone Marrow Toxicity in Mice

2021 ◽  
Vol 12 (1) ◽  
pp. 1085-1095
Keyword(s):  
Bone Marrow ◽  
Histological Analysis ◽  
Bone Marrow Cells ◽  
Mouse Bone Marrow ◽  
Bone Marrow Toxicity ◽  
Protective Effects ◽  
Wbc Count ◽  
Antioxidant Effects ◽  
Cyclophosphamide Administration ◽  
Normal Controls

Spirulina Platenesis (Sp) is rich in important compounds with antioxidant effects. This study investigates the treatment and prophylactic effect of Spirulina Platenesis (Sp) against Cyclophosphamide (CP) induced bone marrow toxicity. Fifty female BALB/C mice were randomly classified into 5 equal groups: (1) Normal controls; (2) CP group: injected with 40 mg/kg for 10 days; (3) Sp group: supplied by Sp (1000 mg/kg) for 10 days; (4) Treatment group: CP + Sp; (5) Prophylactic group: Sp + CP. After 10 days, At the end of the study period, all rats were killed, blood was withdrawn, and bone marrow (BM) was subjected to investigation. Cyclophosphamide administration caused rapid dimension in CBC parameter, Bone marrow WBC count and Erythropiotein concentration in blood that have been recovered with Sp administration. According to histological analysis of the bone marrow, spirulina impaired the proliferation and hypercellularity of immature myeloid elements in the bone marrow, which CP decreased. Obviously, spirulina may with antioxidative activity reduced the oxidative stress and toxicity induced by cyclophosphamide in mouse bone marrow cells.

scholarly journals The Long-Term Impact of Mild Traumatic Brain Injuries on Multiple Functional Outcomes and Epigenetics: A Pilot Study with College Students

2020 ◽  
Author(s):  
Hyunhwa Lee ◽  
Sungchul Lee ◽  
Ipuna Black ◽  
Laura Salado ◽  
Jonica Estrada ◽  
...  
Keyword(s):  
College Students ◽  
Dna Methylation ◽  
Pilot Study ◽  
Visual Function ◽  
Postural Balance ◽  
Sleep Disturbances ◽  
Global Dna Methylation ◽  
Balance Performance ◽  
Global Methylation

People who suffer a mild traumatic brain injury (mTBI) have heterogeneous symptoms and disease trajectories, which make it difficult to precisely diagnose and assess complications long-term. Insufficient information is available regarding how to precisely diagnose and assess mTBI. This study identified and compared deficits in cognitive, psychosocial, visual functions, and balance performance between college students with and without histories of mTBI. Global DNA methylation ratio (5-mC%) in blood was also compared as a peripheral epigenetic marker. Twenty-five volunteers participated in this pilot study, including 11 mTBI cases (27.3% females; mean age of 28.7 years, SD=5.92) and 14 healthy controls (64.3% females; mean age of 22.0, SD=4.13). All the participants were assessed for cognitive (by NIH toolbox&mdash;executive function, memory, and processing speed), psychological (by PROMIS&mdash;depression, anxiety, and sleep disturbances), visual function (by King-Devick and binocular accommodative tests), postural balance performance (by a force plate), and blood 5-mC% (global methylation) levels. Students with mTBI reported significantly poorer episodic memory, severe anxiety, and more sleep disturbance problems. They also had higher blood 5-mC% level (all p&rsquo;s&lt;.05). No significant differences were found in visual function and postural balance. These findings validate changes in cognitive, psychosocial, and global DNA methylation long after mTBI.

Dnmt3a-Deletion Accelerates FLT3-ITD Malignancies In Mice By Hypomethylation Of Enhancer Sites and Activating Stem Cell Programs; Implications For Therapy

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 595-595 ◽  
Author(s):  
Liubin Yang ◽  
Benjamin Rodriguez ◽  
Min Luo ◽  
Mira Jeong ◽  
David Ruau ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Mouse Model ◽  
Acute Leukemia ◽  
Bone Marrow Cells ◽  
Self Renewal ◽  
Expression Of Genes ◽  
Marrow Cells

Abstract The de novo DNA methyltransferase (DNMT) 3A is mutated in 50% of patients with mixed phenotype acute leukemia, 20% with acute myeloid leukemia (AML) and 18% with T-cell acute lymphoblastic leukemia (T-ALL). The mechanisms through which mutant DNMT3A contributes to hematologic malignancy are poorly understood. In mice, deletion of Dnmt3a in hematopoietic stem cells (HSCs) leads to abnormal DNA methylation and inhibition of differentiation, but is insufficient for leukemic transformation. To study the role of Dnmt3a in leukemia, we combined Dnmt3a-deletion with the activated FLT3 proto-oncogene (FLT3-ITD), a frequent co-mutation with DNMT3A in AML patients, to establish a murine model of Dnmt3a-associated malignancy. In mice transplanted with Dnmt3a-knockout (KO) or wild-type (WT) bone marrow cells transduced with a FLT3-ITD retrovirus, Dnmt3a-loss dramatically impacted the disease phenotype. Dnmt3aKO/ITD transplanted mice had significantly shortened survival (79 days vs. 116 days) and increased rate of acute leukemia compared to mice with ITD alone. The mice developed CD4+CD8+ Notch activation-associated T-ALL or myeloproliferative disease (MPD), or concurrently both, consistent with previous studies of FLT3-ITD in mice. To determine the leukemia-initiating population, we transplanted sorted HSC, myeloid, and lymphoid progenitors transduced with FLT3-ITD. All mice transplanted with HSC and myeloid progenitors succumbed to both malignancies. To uncover the mechanisms by which Dnmt3a-deletion accelerated acute leukemia, we analyzed changes in DNA methylation in T-ALL blasts by whole genome bisulfite sequencing. Compared to Dnmt3aWT/ITD, Dnmt3aKO/ITD blasts exhibited global hypomethylation, particularly at distal enhancer sites. These hypomethylated enhancer sites were associated with genes in signaling pathways, transcription regulators, and metabolic pathways in cancer (KEGG and GO Analysis). Transcriptome analysis showed that relative to Dnmt3aWT/ITD, the Dnmt3aKO/ITD blasts had 1577 significantly differentially expressed genes positively related to cancer, cellular growth, and proliferation, and negatively to apoptosis by Ingenuity Pathway Analysis (IPA). Surprisingly, we observed increased expression of genes related to HSCs and myeloid function and decreased expression of genes related to lymphocyte function. Human AML signature genes (Oncomine) were also upregulated in our mouse model. Predicted activated pathways include Myc, Nfe2l2, Eif4e, E2f1, Csf2, Cebpb, Vegf, Rxra, Ezh2, and Brd4 and inhibited pathways include tumor suppressors Rb, let7, Cdkn2a, and Tob1 (IPA). We did not observe changes in genomic copy number variation by chromosomal comparative hybridization (cCGH). To test whether Dnmt3a-deletion could functionally bestow stem cell properties on pre-leukemic cells, we examined self-renewal capabilities of malignant cells of Flt3+/ITD knock-in mouse (an ITD mutation knocked in to the endogenous murine Flt3 allele causing MPD). Remarkably, when Dnmt3aKO; Flt3+/ITD bone marrow cells were serially transplanted, MPD was seen in all recipients, compared to none in Dnmt3aWT; Flt3+/ITD transplanted mice (n=7). Further, we transplanted sorted CLP, CMP, GMP, MPP, ST-HSC, LT-HSC populations and observed myeloproliferation in transplanted non-stem (CMP, GMP, ST-HSC) and stem cell (LT-HSC) populations. This strongly suggests that Dnmt3aKO synergized with Flt3-ITD to confer stem cell self-renewal abilities to transformed progenitor and stem cells. Increasingly, decitabine is being used to treat patients with AML and MDS, but whether patients with DNMT3A mutations could benefit is unclear, so we examined the impact of decitabine treatment on the retroviral transduced Dnmt3aKO/ITD mice. Monthly treatment led to significantly increased survival of Dnmt3aKO/ITD mice from T-ALL and MPD and reduced presence of ITD-transduced KO cells. Together, we demonstrate that Dnmt3aKO accelerated malignancies induced by FLT3-ITD in mouse and may shed light on how DNMT3A mutations contribute to lymphoid and myeloid disease in patients. Dnmt3a deletion ignited multilineage and stem cell programs at the expense of lymphoid programs to accelerate disease, but was extinguishable by decitabine therapy. The findings from our mouse model can be used for the development and testing of targeted epigenetic therapy for DNMT3A-associated malignancies. Disclosures: No relevant conflicts of interest to declare.

Colorimetric and electrochemical quantification of global DNA methylation using a methyl cytosine-specific antibody

The Analyst ◽  
2017 ◽  
Vol 142 (11) ◽  
pp. 1900-1908 ◽  
Author(s):  
Md. Hakimul Haque ◽  
Ripon Bhattacharjee ◽  
Md. Nazmul Islam ◽  
Vinod Gopalan ◽  
Nam-Trung Nguyen ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Specific Antibody ◽  
Electrochemical Method ◽  
Global Dna Methylation ◽  
Global Methylation ◽  
Naked Eye ◽  
Methyl Cytosine

We report a simple colorimetric (naked-eye) and electrochemical method for the rapid, sensitive and specific quantification of global methylation levels using only 25 ng of input DNA.

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