Epigenetic Action Research Articles

Resveratrol reverses male reproductive damage in rats exposed to nicotine during the intrauterine phase and breastfeeding.

Nicotine leads to reproductive changes culminating in male infertility and subfertility. Resveratrol, a polyphenol, is a biological modulator. Sirtuin 1 (SIRT1) protein can positively act on male reproduction, and its expression can be affected by nicotine and modulated by resveratrol. The capability of resveratrol to reverse the reproductive damage in adult male offspring, which was nicotine-exposed during the intrauterine phase and breastfeeding, was investigated. Four groups were established with male offspring born from nicotine-exposed and non-exposed rat dams during pregnancy and lactation. Forty-eight male Wistar rats were distributed into four groups: sham control (SC), resveratrol (R), nicotine (N), and nicotine + resveratrol (NR). Rat dams of the N and NR offspring were exposed to nicotine (2mg/kg/day) during pregnancy and lactation using a subcutaneously implanted minipump. The offspring of the R and NR groups received resveratrol (300mg/kg of body weight, gavage) for 63 days from puberty. At 114 days of age, the male rats were euthanized. Nicotine did not alter the body weight, biometry of reproductive organs, or quantitative sperm parameters of adult offspring but caused an evident worsening of all sperm qualitative parameters studied. Daily treatment with resveratrol from puberty up to adulthood improved all qualitative sperm parameters significantly, leading some of them close to the control values. Resveratrol also improved the morphological integrity and expression of SIRT1 in the seminiferous epithelium of nicotine-exposed offspring. Resveratrol reversed the male reproductive damage caused by nicotine. Nicotine crosses the blood-placental membrane and is present in the breast milk of mothers who smoke. Resveratrol restored the altered reproductive parameters in the male adult offspring that were nicotine-exposed during intrauterine life and breastfeeding. The epigenetic modulating action of resveratrol can be involved in this nicotine damage reversion. Resveratrol may be a promising candidate to be investigated regarding the adjuvant strategies in the treatment of male infertility.

Effect of titanium dioxide nanoparticles on histone modifications and histone modifying enzymes expression in human cell lines

Titanium dioxide (TiO2) nanoparticles are widely manufactured, with a range of applications in consumer products. Significant toxicity of TiO2 nanoparticles has, however, been recognized, suggesting considerable risk to human health. To evaluate fully their toxicity, assessment of the epigenetic action of these nanoparticles is critical. However, only few studies are available examining the capability of nanoparticles to alter epigenetic integrity. In the present study, the effect of TiO2 nanoparticles exposure on histone modifications, a major epigenetic mechanism, was investigated in human colorectal (Caco-2) and lung (NL20) epithelial cell lines. Histone H3 and H4 modifications were assessed by array analysis using the EpiQuickTM Histone H3 or H4 Modification Multiplex Assay. Seventeen histone modifications were identified with altered levels after exposure to TiO2 nanoparticles. Changes in several selected histone modifications (Caco-2 cells: H3cit, H3K9me3, H3K27me3, H3K36me3, H3K9ac, and H4K8ac; NL20 cells: H3K4me3, H3K9me3, H3K27me3, H3K9ac, and H3K18ac) were verified by Western blot analysis. The results also revealed aberrant expression of histone modifying enzymes in TiO2 exposed cells. Expression levels were determined by array analysis using the Human Epigenetic Chromatin Modification Enzymes RT2 Profiler™ PCR Array, with 12 genes identified in both Caco-2 cells and NL20 cells. qRT-PCR analysis confirmed the array results for several selected histone modifying enzyme genes (ASH1L, CARM1, EHMT2, HAT1, HDAC9, KMT2E, NCOA1, SETDB2, and USP16). The findings from this study clearly demonstrate the impact of TiO2 nanoparticles exposure on histone modification in two human cell lines, supporting potential involvement of this epigenetic mechanism in the toxicity of TiO2 nanoparticles. Hence, for complete assessment of potential risk from nanoparticle exposure, epigenetic studies are critical.

Colorectal cancer inhibition by BET inhibitor JQ1 is MYC-independent and not improved by nanoencapsulation

Bromodomain and extraterminal domain protein inhibitors (BETi) for cancer treatment did not convince during their first clinical trials. Their epigenetic mechanism of action is still not well understood, even if MYC is generally considered as its main downstream target. In this context, we intended to assess two new nanoformulations of the BETi JQ1 for the treatment of colorectal cancer (CRC). JQ1 was encapsulated at 10 mg/mL in lipid nanocapsules (LNC) or polymeric micelles (PM), both compatible for an intravenous administration. Their effect was compared with free JQ1 on several CRC cell lines in vitro and with daily intraperitoneal cyclodextrin (CD)-loaded JQ1 on the CT26 CRC tumor model in vivo. We showed that LNC preferentially accumulated in tumor, liver, and lymph nodes. LNC-JQ1 and CD-JQ1 similarly delayed tumor growth and increased median survival from 15 to 23 or 20.5 days. JQ1 altered MYC in only two among four CRC cell lines. This MYC-independence found in CT26 was confirmed in vivo by PCR and immunohistochemistry. The main explanation of the JQ1 anticancer effect was an increase in apoptosis. The investigation of its impact on the tumor microenvironment did not show significant effects. Finally, JQ1 association with irinotecan did not synergize in vivo with JQ1 nanoformulations. In conclusion, we demonstrated that the JQ1 anticancer effect was not improved by nanoencapsulation even if their tumor delivery was probably higher. MYC inhibition was not associated to JQ1 efficacy in the case of the CT26 CRC murine model.

Butyrate Restores the Fat/Lean Mass Ratio Balance and Energy Metabolism and Reinforces the Tight Junction-Mediated Intestinal Epithelial Barrier in Prediabetic Mice Independently of its Anti-Inflammatory and Epigenetic Actions

Butyrate Restores the Fat/Lean Mass Ratio Balance and Energy Metabolism and Reinforces the Tight Junction-Mediated Intestinal Epithelial Barrier in Prediabetic Mice Independently of its Anti-Inflammatory and Epigenetic Actions

Usefulness of Melatonin and Other Compounds as Antioxidants and Epidrugs in the Treatment of Head and Neck Cancer.

Along with genetic mutations, aberrant epigenetic alterations are the initiators of head and neck cancer carcinogenesis. Currently, several drugs are being developed to correct these epigenetic alterations, known as epidrugs. Some compounds with an antioxidant effect have been shown to be effective in preventing these malignant lesions and in minimizing the complications derived from cytotoxic treatment. Furthermore, in vitro and in vivo studies show a promising role in the treatment of head and neck squamous cell carcinoma (HNSCC). This is the case of supplements with DNA methylation inhibitory function (DNMTi), such as epigallocatechin gallate, sulforaphane, and folic acid; histone deacetylase inhibitors (HDACi), such as sodium butyrate and melatonin or histone acetyltransferase inhibitors (HATi), such as curcumin. The objective of this review is to describe the role of some antioxidants and their epigenetic mechanism of action, with special emphasis on melatonin and butyric acid given their organic production, in the prevention and treatment of HNSCC.

Electromagnetic Information Transfer (EMIT) by Ultra High Diluted (UHD) solutions: the suggestive hypothesis of an epigenetic action.

The aim of this work is to confirm the theoretical possibility of an epigenetic mechanism shared between EMIT and UHD. The presentation will be divided in three sections:
 1. Water aggregates with an electric dipole moment (UHD succussed solutions) as mediators of weak specific bioelectromagnetic signals on target stem cells.
 Recent experimental works confirm the developing concept of water mediated Electromagnetic Information Transfer (EMIT) of specific molecular signals, picked up from the source biological effector, on target stem cells with evident effect on their proliferation [1]. Similar Electromagnetic (EM) emission and consequences are also reported by the scientific literature on rotational excited aggregates with an electric dipole moment, created in polar liquids by Ultra High Diluted (UHD) or High Diluted (HD) succussed solutions. These aggregates are composed of solvent molecules only or a combination of these and solute particles [2]. 
 2. DNA mediated physiopathological effects of ELF EMFs 
 In detail, according to the International Agency for Research on Cancer (IARC), the extremely low-frequency (ELF) electromagnetic fields (EMFs) are classified as "possible carcinogenic" based on their effects [3-5], although most scientists agree that they are too weak to kill cells or to cause mutations and thus initiate cancer. Besides the prevailing paradigm of the environmentally-induced acute and chronic diseases involving either cell killing (cytotoxicity) or gene/chromosome mutations (genotoxicity), many studies concerning the biological and health consequences of ELF-EM exposure report that alteration of the expression of genetic information at the transcriptional, translational, or posttranslational levels has the potential to contribute to various diseases. 
 3. Epigenetic mechanism shared between EMIT and UHDs
 The latter referred mechanism, denoted as "epigenetic" (that affects gene expression rather than gene structure), is characterized by threshold-like action, multiple biochemical pathways and it needs chronic regular exposures to be effective [6]. Epigenetic factors affect one of four potential cell states, namely alteration of cell proliferation, cell differentiation, programmed cell death (apoptosis) or adaptive responses of differentiated cells, and probably they act as co-inductors of DNA damage rather than as a genotoxic agents per se. At the present time, studies on genomic and functional genetic are identifying many genes and gene variants that potentially modulate the fundamental molecular mechanisms underpinning both physiological and pathological processes.

Influence of Benzo(a)pyrene on Different Epigenetic Processes.

Epigenetic changes constitute one of the processes that is involved in the mechanisms of carcinogenicity. They include dysregulation of DNA methylation processes, disruption of post-translational patterns of histone modifications, and changes in the composition and/or organization of chromatin. Benzo(a)pyrene (BaP) influences DNA methylation and, depending on its concentrations, as well as the type of cell, tissue and organism it causes hypomethylation or hypermethylation. Moreover, the exposure to polyaromatic hydrocarbons (PAHs), including BaP in tobacco smoke results in an altered methylation status of the offsprings. Researches have indicated a potential relationship between toxicity of BaP and deregulation of the biotin homeostasis pathway that plays an important role in the process of carcinogenesis. Animal studies have shown that parental-induced BaP toxicity can be passed on to the F1 generation as studied on marine medaka (Oryzias melastigma), and the underlying mechanism is likely related to a disturbance in the circadian rhythm. In addition, ancestral exposure of fish to BaP may cause intergenerational osteotoxicity in non-exposed F3 offsprings. Epidemiological studies of lung cancer have indicated that exposure to BaP is associated with changes in methylation levels at 15 CpG; therefore, changes in DNA methylation may be considered as potential mediators of BaP-induced lung cancer. The mechanism of epigenetic changes induced by BaP are mainly due to the formation of CpG-BPDE adducts, between metabolite of BaP—BPDE and CpG, which leads to changes in the level of 5-methylcytosine. BaP also acts through inhibition of DNA methyltransferases activity, as well as by increasing histone deacetylases HDACs, i.e., HDAC2 and HDAC3 activity. The aim of this review is to discuss the mechanism of the epigenetic action of BaP on the basis of the latest publications.

Basics of Epigenetics and Role of Epigenetics in Diabetic Complications.

The term “Epigenetics” includes mechanisms by which genetic expression is altered without a change in the underlying DNA sequence. The changes caused by epigenetic mechanisms are inheritable and are one way in direction (irreversible) and also explains why there is differences in genetic expressions of monozygotic twins. The epigenetic mechanisms alter the genetic expressions through DNA methylation, posttranslational modifications (PTMs) of histone, and noncoding RNAs. DNA methylation and histone PTMs cause relaxation or condensation of chromatin units. The epigenetic actions of noncoding RNAs such as microRNAs, small nucleolar RNAs, small interfering RNAs, and long noncoding RNAs act by modifying transcription factors or by degrading target messenger RNAs and their translation factors. Various pathologies and environmental factors cause changes in the cellular epigenetic mechanisms and the epigenetic alterations occurring in diabetes mellitus (DM) are reviewed. DM causes hemodynamic changes and metabolic changes like hyperglycemia and dyslipidemia. These changes induce oxidative stress and activate intracellular signaling and kinases in the target cells. Epigenetic alterations cause chromatin remodeling and altered gene expression leading to inflammation, proliferation, atrophy, hypertrophy, etc.; thereby, diabetic complications such as neuropathy, nephropathy, vasculitis result in the corresponding target organ. When these epigenetic alterations persist for a longer period without intervention, the target cells attain “metabolic memory” meaning that these epigenetic mutations cannot be reversed even after attaining normal blood glucose levels. Thus, epigenetics, an insightful and efficient tool in genomic research, has started crawling into the research arena and needs to reach leaps and bounds for the better understanding of health and diseases.

New Highlights of Resveratrol: A Review of Properties against Ocular Diseases.

Eye diseases are currently a major public health concern due to the growing number of cases resulting from both an aging of populations and exogenous factors linked to our lifestyles. Thus, many treatments including surgical pharmacological approaches have emerged, and special attention has been paid to prevention, where diet plays a preponderant role. Recently, potential antioxidants such as resveratrol have received much attention as potential tools against various ocular diseases. In this review, we focus on the mechanisms of resveratrol against ocular diseases, in particular age-related macular degeneration, glaucoma, cataract, diabetic retinopathy, and vitreoretinopathy. We analyze, in relation to the different steps of each disease, the resveratrol properties at multiple levels, such as cellular and molecular signaling as well as physiological effects. We show and discuss the relationship to reactive oxygen species, the regulation of inflammatory process, and how resveratrol can prevent ocular diseases through a potential epigenetic action by the activation of sirtuin-1. Lastly, various new forms of resveratrol delivery are emerging at the same time as some clinical trials are raising more questions about the future of resveratrol as a potential tool for prevention or in therapeutic strategies against ocular diseases. More preclinical studies are required to provide further insights into RSV’s potential adjuvant activity.

Epigenetics and the endocannabinoid system signaling: An intricate interplay modulating neurodevelopment

The endocannabinoid (eCB) system is a complex system comprising endogenous cannabinoids (eCBs), their synthesis and degradation enzymes, and cannabinoid receptors. These elements crucially regulate several biological processes during neurodevelopment, such as proliferation, differentiation, and migration. Recently, eCBs were also reported to have an epigenetic action on genes that play key functions in the neurotransmitter signaling, consequently regulating their expression. In turn, epigenetic modifications (e.g. DNA methylation, histone modifications) may also modulate the function of eCB system’s elements. For example, the expression of the cnr gene in the central nervous system may be epigenetically regulated (e.g. DNA methylation, histone modifications), thus altering the function of the cannabinoid receptor type-1 (CB1R). Considering the importance of the eCB system during neurodevelopment, it is thus reasonable to expect that alterations in this interaction between the eCB system and epigenetic modifications may give rise to neurodevelopmental disorders.Here, we review key concepts related to the regulation of neuronal function by the eCB system and the different types of epigenetic modifications. In particular, we focus on the mechanisms involved in the intricate interplay between both signaling systems and how they control cell fate during neurodevelopment. Noteworthy, such mechanistic understanding assumes high relevance considering the implications of the dysregulation of key neurogenic processes towards the onset of neurodevelopment-related disorders.Moreover, considering the increasing popularity of cannabis and its synthetic derivatives among young adults, it becomes of utmost importance to understand how exogenous cannabinoids may epigenetically impact neurodevelopment.

A Review of the Functional Roles of the Zebrafish Aryl Hydrocarbon Receptors.

Over the last 2 decades, the zebrafish (Danio rerio) has emerged as a stellar model for unraveling molecular signaling events mediated by the aryl hydrocarbon receptor (AHR), an important ligand-activated receptor found in all eumetazoan animals. Zebrafish have 3 AHRs-AHR1a, AHR1b, and AHR2, and studies have demonstrated the diversity of both the endogenous and toxicological functions of the zebrafish AHRs. In this contemporary review, we first highlight the evolution of the zebrafish ahr genes, and the characteristics of the receptors including developmental and adult expression, their endogenous and inducible roles, and the predicted ligands from homology modeling studies. We then review the toxicity of a broad spectrum of AHR ligands across multiple life stages (early stage, and adult), discuss their transcriptomic and epigenetic mechanisms of action, and report on any known interactions between the AHRs and other signaling pathways. Through this article, we summarize the promising research that furthers our understanding of the complex AHR pathway through the extensive use of zebrafish as a model, coupled with a large array of molecular techniques. As much of the research has focused on the functions of AHR2 during development and the mechanism of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) toxicity, we illustrate the need to address the considerable knowledge gap in our understanding of both the mechanistic roles of AHR1a and AHR1b, and the diverse modes of toxicity of the various AHR ligands.

Impact of lipopolysaccharide administration on luteinizing hormone/choriogonadotropin receptor (Lhcgr) expression in mouse ovaries

Lipopolysaccharide (LPS) is isolated from the genital tract of animals suffering from uterine damage and ovarian dysfunction. This study provides direct molecular evidence about the mechanism through which endotoxins cause reproductive disorders. Granulosa cells and ovaries were collected from immature mice treated with eCG or with eCG and LPS injection intraperitoneally. Normal large antral follicles were observed in ovaries obtained from eCG and LPS coinjected mice, and the morphology of the ovaries was similar to that observed in the control group. These antral follicles were not deemed atretic because few TUNEL-positive cells were observed. However, the granulosa cells of large antral follicles did not acquire the ability to respond to hCG stimulation. The number of ovulated oocytes was significantly lower in LPS-injected mice after superovulation compared to mice that were not exposed to LPS. The low reactivity was caused by the limited expression of the Lhcgr gene, which encodes the LH receptor in granulosa cells as well as an LPS-induced increase in the level of Dnmt1 expression. The methylation rate of the Lhcgr promoter region was significantly higher in granulosa cells obtained from the LPS treatment group compared with the control group. Together, these findings demonstrated that the decrease in the expression of Lhcgr due to LPS was a result of the epigenetic regulatory action of LPS. Our studies suggest that ovarian follicular cysts that is characterized by bacterial infection in humans and animals, is closely connected to the level of methylation of the Lhcgr promoter region.

Transcription Factors Phox2a/2b Upregulate Expression of Noradrenergic and Dopaminergic Phenotypes in Aged Rat Brains.

The present study investigated the effects of forced overexpression of Phox2a/2b, two transcription factors, in the locus coeruleus (LC) of aged rats on noradrenergic and dopaminergic phenotypes in brains. Results showed that a significant increase in Phox2a/2b mRNA levels in the LC region was paralleled by marked enhancement in expression of DBH and TH per se. Furthermore, similar increases in TH protein levels were observed in the substantial nigra and striatum, as well as in the hippocampus and frontal cortex. Overexpression of Phox2 genes also significantly increased BrdU-positive cells in the hippocampal dentate gyrus and NE levels in the striatum. Moreover, this manipulation significantly improved the cognition behavior. The in vitro experiments revealed that norepinephrine treatments may increase the transcription of TH gene through the epigenetic action on the TH promoter. The results indicate that Phox2 genes may play an important role in improving the function of the noradrenergic and dopaminergic neurons in aged animals, and regulation of Phox2 gene expression may have therapeutic utility in aging or disorders involving degeneration of noradrenergic neurons.

Anticancer Activity of Rosmarinus officinalis L.: Mechanisms of Action and Therapeutic Potentials.

Alternative treatments for neoplastic diseases with new drugs are necessary because the clinical effectiveness of chemotherapy is often reduced by collateral effects. Several natural substances of plant origin have been demonstrated to be successful in the prevention and treatment of numerous tumors. Rosmarinus officinalis L. is a herb that is cultivated in diverse areas of the world. There is increasing attention being directed towards the pharmaceutical capacities of rosemary, utilized for its anti-inflammatory, anti-infective or anticancer action. The antitumor effect of rosemary has been related to diverse mechanisms, such as the antioxidant effect, antiangiogenic properties, epigenetic actions, regulation of the immune response and anti-inflammatory response, modification of specific metabolic pathways, and increased expression of onco-suppressor genes. In this review, we aim to report the results of preclinical studies dealing with the anticancer effects of rosemary, the molecular mechanisms related to these actions, and the interactions between rosemary and anticancer drugs. The prospect of utilizing rosemary as an agent in the treatment of different neoplastic diseases is discussed. However, although the use of rosemary in the therapy of neoplasms constitutes a fascinating field of study, large and controlled studies must be conducted to definitively clarify the real impact of this substance in clinical practice.

Anesthetic propofol epigenetically regulates breast cancer trastuzumab resistance through IL-6/miR-149-5p axis

Propofol, a common intravenous anesthetic, has been found to exert anti-cancer effects with inhibition of cancer cell proliferation, migration and invasion. We tested its possible action against HER2-overexpressing breast cancer cells that developed resistance against trastuzumab. Cell viability assay, ELISA for cytokines, mammosphere formation, quantitative RT-PCR for EMT/IL-6-targeting miRNAs and the in vivo experimental pulmonary metastasis model were performed to understand the epigenetic action of propofol. Propofol sensitized HER2 overexpressing cells to trastuzumab but such action was even more pronounced in resistant cells. Increased cytokines IL-6 as well as IL-8 were released by resistant cells, along with increased mammospheres and induction of EMT, all of which was inhibited by propofol. IL-6 targeting tumor suppressor miR-149-5p was found to be the novel miRNA that was up-regulated by propofol, resulting in the observed effects on cell viability, IL-6 production, mammospheres generation as well as EMT induction. Further, antagonizing miR-149-5p attenuated the propofol effects confirming the epigenetic activity of propofol through miR-149-5p regulation. Finally, in vivo validation in an experimental metastasis model conformed an inhibitory action of propofol against experimental lung metastasis and the essential mechanistic role of miR-149-5p/IL-6 loop. These results present a novel role of general anesthetic propofol against resistant breast cancer cells and the underlying epigenetic regulation of a tumor suppressor miRNA.

Flavonoids as Epigenetic Modulators for Prostate Cancer Prevention.

Prostate cancer (PCa) is a multifactorial disease with an unclear etiology. Due to its high prevalence, long latency, and slow progression, PCa is an ideal target for chemoprevention strategies. Many research studies have highlighted the positive effects of natural flavonoids on chronic diseases, including PCa. Different classes of dietary flavonoids exhibit anti-oxidative, anti-inflammatory, anti-mutagenic, anti-aging, cardioprotective, anti-viral/bacterial and anti-carcinogenic properties. We overviewed the most recent evidence of the antitumoral effects exerted by dietary flavonoids, with a special focus on their epigenetic action in PCa. Epigenetic alterations have been identified as key initiating events in several kinds of cancer. Many dietary flavonoids have been found to reverse DNA aberrations that promote neoplastic transformation, particularly for PCa. The epigenetic targets of the actions of flavonoids include oncogenes and tumor suppressor genes, indirectly controlled through the regulation of epigenetic enzymes such as DNA methyltransferase (DNMT), histone acetyltransferase (HAT), and histone deacetylase (HDAC). In addition, flavonoids were found capable of restoring miRNA and lncRNA expression that is altered during diseases. The optimization of the use of flavonoids as natural epigenetic modulators for chemoprevention and as a possible treatment of PCa and other kinds of cancers could represent a promising and valid strategy to inhibit carcinogenesis and fight cancer.

Klotho Pathways, Myelination Disorders, Neurodegenerative Diseases, and Epigenetic Drugs.

In this review we outline a rationale for identifying neuroprotectants aimed at inducing endogenous Klotho activity and expression, which is epigenetic action, by definition. Such an approach should promote remyelination and/or stimulate myelin repair by acting on mitochondrial function, thereby heralding a life-saving path forward for patients suffering from neuroinflammatory diseases. Disorders of myelin in the nervous system damage the transmission of signals, resulting in loss of vision, motion, sensation, and other functions depending on the affected nerves, currently with no effective treatment. Klotho genes and their single-pass transmembrane Klotho proteins are powerful governors of the threads of life and death, true to the origin of their name, Fates, in Greek mythology. Among its many important functions, Klotho is an obligatory co-receptor that binds, activates, and/or potentiates critical fibroblast growth factor activity. Since the discovery of Klotho a little over two decades ago, it has become ever more apparent that when Klotho pathways go awry, oxidative stress and mitochondrial dysfunction take over, and age-related chronic disorders are likely to follow. The physiological consequences can be wide ranging, potentially wreaking havoc on the brain, eye, kidney, muscle, and more. Central nervous system disorders, neurodegenerative in nature, and especially those affecting the myelin sheath, represent worthy targets for advancing therapies that act upon Klotho pathways. Current drugs for these diseases, even therapeutics that are disease modifying rather than treating only the symptoms, leave much room for improvement. It is thus no wonder that this topic has caught the attention of biomedical researchers around the world.

Effect of titanium dioxide nanoparticles on DNA methylation in multiple human cell lines

Nanoscale titanium dioxide (TiO2) is manufactured in wide scale, with a range of applications in consumer products. Significant toxicity of TiO2 nanoparticles has, however, been recognized, suggesting considerable risk to human health. To evaluate fully their toxicity, assessment of the epigenetic action of these nanoparticles is critical. However, only few studies are available examining capability of nanoparticles to alter epigenetic integrity. In the present study, the effect of TiO2 nanoparticles exposure on DNA methylation, a major epigenetic mechanism, was investigated in in vitro cellular model systems. A panel of cells relevant to portals of human exposure (Caco-2 (colorectal), HepG2 (liver), NL20 (lung), and A-431 (skin)) was exposed to TiO2 nanoparticles to assess effects on global methylation, gene-specific methylation, and expression levels of DNA methyltransferases, MBD2, and UHRF1. Global methylation was determined by enzyme-linked immunosorbent assay-based immunochemical analysis. Degree of promoter methylation across a defined panel of genes was evaluated using EpiTect Methyl II Signature PCR System Array technology. Expression of DNMT1, DNMT3a, DNMT3b, MBD2, and URHF1 was quantified by qRT-PCR. Decrease in global DNA methylation in cell lines Caco-2, HepG2, and A-431 exposed to TiO2 nanoparticles was shown. Across four cell lines, eight genes (CDKN1A, DNAJC15, GADD45A, GDF15, INSIG1, SCARA3, TP53, and BNIP3) were identified in which promotors were methylated after exposure. Altered expression of these genes is associated with disease etiology. The results also revealed aberrant expression of epigenetic regulatory genes involved in DNA methylation (DNMT1, DNMT3a, DNMT3b, MBD2, and UHRF1) in TiO2 exposed cells, which was cell type dependent. Findings from this study clearly demonstrate the impact of TiO2 nanoparticles exposure on DNA methylation in multiple cell types, supporting potential involvement of this epigenetic mechanism in the toxicity of TiO2 nanoparticles. Hence for complete assessment of potential risk from nanoparticle exposure, epigenetic studies are critical.

Histone lysine demethylase KDM5B maintains chronic myeloid leukemia via multiple epigenetic actions

The histone lysine demethylase KDM5 family is implicated in normal development and stem cell maintenance by epigenetic modulation of histone methylation status. Deregulation of the KDM5 family has been reported in various types of cancers, including hematological malignancies. However, their transcriptional regulatory roles in the context of leukemia remain unclear. Here, we find that KDM5B is strongly expressed in normal CD34+ hematopoietic stem/progenitor cells and chronic myeloid leukemia (CML) cells. Knockdown of KDM5B in K562 CML cells reduced leukemia colony-forming potential. Transcriptome profiling of KDM5B knockdown K562 cells revealed the deregulation of genes involved in myeloid differentiation and Toll-like receptor signaling. Through the integration of transcriptome and ChIP-seq profiling data, we show that KDM5B is enriched at the binding sites of the GATA and AP-1 transcription factor families, suggesting their collaborations in the regulation of transcription. Even though the binding of KDM5B substantially overlapped with H3K4me1 or H3K4me3 mark at gene promoters, only a small subset of the KDM5B targets showed differential expression in association with the histone demethylation activity. By characterizing the interacting proteins in K562 cells, we discovered that KDM5B recruits protein complexes involved in the mRNA processing machinery, implying an alternative epigenetic action mediated by KDM5B in gene regulation. Our study highlights the oncogenic functions of KDM5B in CML cells and suggests that KDM5B is vital to the transcriptional regulation via multiple epigenetic mechanisms.

Molecular mechanisms underlying eicosapentaenoic acid inhibition of HDAC1 and DNMT expression and activity in carcinoma cells

DNA methylation and histone acetylation, the most studied epigenetic changes, drive and maintain cancer phenotypes. DNA methyltransferase (DNMT) dysregulation promoted localized hypermethylation in CpG rich regions while upregulated histone deacetylases (HDAC) deacetylated histone tails. Both changes led to close chromatin conformation, suppressing transcription and silencing tumor suppressor genes. Consequently, HDAC and DNMT inhibitors appeared to reprogram the transcriptional circuit and potentiate anti-tumoral activity. Here, we report that eicosapentaenoic acid (EPA), a fatty acid with anti-cancer properties, inhibited HDAC1 and DNMT expression and activity, thus promoting tumor suppressor gene expression. In hepatocarcinoma cells (HCC) EPA bound and activated PPARγ thus downregulating HDAC1 which sequentially reduced expression of DNMT1, 3A and 3B. At the same time, activated PPARγ physically interacted with DNMT1 and HDAC1 in a CpG island on the Hic-1 gene to assemble PPARγ/DNMT1 and PPARγ/HDAC1 protein complexes, which exited from DNA. When EPA and PPARγ were no longer bound, the protein complexes separated into individual proteins. Consequently, DNMT1 and HDAC1 down-regulation and release from DNA inhibited their activities. Overall, EPA-bound PPARγ induced re-expression of the tumor suppressor gene Hic-1. In the present study PPARγ emerged as a master regulator acting synergistically through diverse targets and ways to reveal the epigenetic action of EPA as an HDAC1 and DNMT1 inhibitor.