Histone H3 Acetylation Levels Research Articles

BDNF levels are increased in peripheral blood of middle-aged amateur runners with no changes on histone H4 acetylation levels.

Our aim was to compare the basal levels of plasma brain-derived neurotrophic factor (BDNF) and global histone H4 acetylation in peripheral blood mononuclear cells (PBMCs) of healthy amateur runners (EXE group) with sedentary individuals (SED group) as well as to investigate the acute effect of a running race on these markers in the EXE group. Five days before the race, all participants were submitted to a basal blood collection. On the race day, two blood samples were collected in the EXE group before the running started and immediately at the end. In the basal period, a significant increase of plasma BDNF levels in the EXE individuals when compared to the SED group (p=0.036) was demonstrated, while no difference in global histone H4 acetylation levels was observed. These parameters were unaltered in the EXE group after the race. The increased levels of BDNF might be linked to healthy middle-aged runners' phenotype.

Effects of histone deacetylase inhibitor sodium butyrate on heroin seeking behavior in the nucleus accumbens in rats

Histone acetylation and other modifications of the chromatin are important regulators of gene expression and may contribute to drug-induced behaviors and neuroplasticity. Inhibition of histone deacetylases (HDAC) activity results in the change of some drug-induced behaviors，however, relatively little is known about the effects of HDAC inhibitors on heroin-seeking behavior. In the present study, male rats were trained to self-administer heroin under a FR1 schedule for consecutive 14 days, followed by 14 daily 2h extinction session in the operant chamber. After training, the heroin priming (250μg/kg) was introduced for the reinstatement of heroin-seeking behavior. Pretreatment with sodium butyrate (NaB) (200 or 400mg/kg, i.p.), an inhibitor of HDAC, failed to affect heroin self-administration. Additionally，systemic administration of NaB (400mg/kg, i.p.)increased significantly the reinstatement of heroin-seeking induced by heroin priming when NaB administered 12h, but not 6h before the reinstatement test. The same effect was observed after the intracerebroventricular injection of NaB (5μL, 100μg/μL). Moreover, the levels of histone H3 acetylation at lysine 18（H3K18）and H4 acetylation at lysine 5 or lysine 8（H4K5 or H4K8）in the accumbens nucleus core and shell were remarkably increased during the reinstatement and were further strengthened after intracerebroventricular injection of NaB. These results demonstrated that activation of histone acetylation may be involved in the heroin-seeking behavior, and identifying these epigenetic changes will be critical in proposing a novel pharmacological strategy for treating heroin addiction.

Epigenetic Variability of CD4+CD25+ Tregs Contributes to the Pathogenesis of Autoimmune Diseases

Autoimmune diseases are characterized by aberrant immune responses against healthy cells and tissues. However, the exact mechanisms underlying the development of these conditions remain unknown. CD4+CD25+ regulatory T cells (Tregs) are a subset of mature T cells which have an important role in maintaining immune homeostasis and preventing autoimmune diseases. Forkhead box p3 (Foxp3), a member of the fork head transcription factor family, is recognized as a marker of CD4+CD25+ Tregs. The decreased number and/or function of CD4+CD25+ Tregs in peripheral blood and related tissues has been demonstrated in systemic lupus erythematosus, systemic sclerosis, and other autoimmune diseases, which are at least partially regulated by epigenetic mechanisms. Epigenetics refers to the study of potentially heritable alterations in gene expression without underlying changes of the nucleotide sequence, mainly including DNA methylation, histone modification, and microRNAs (miRNAs). For example, DNA methylation status of CpG islands on the Foxp3 gene, which may be affected by normal aging and regulated by environmental factors, plays an important role in modulating the homeostasis of Foxp3 expression in Tregs. Foxp3 gene in Tregs also shows distinct acetylation and trimethylation levels of histone H3 and H4 when compared with effector T cells, leading to an open chromatin structure. MicroRNAs such as miR-155, miR-126, and miR-10a also exert an important influence on the differentiation, development, and immunological functions of Tregs. Aberrant epigenetic modifications affecting Foxp3 and other key genes in Tregs contribute to disease activity and tissue inflammation in autoimmune diseases, which holds great potential for providing novel targets for epigenetic therapies. Advances in research into the epigenetic regulation of CD4+CD25+ Tregs may also lead to the identification of new epigenetic biomarkers for diagnosis and prognosis.

Histone deacetylase inhibitor-induced emergence of synaptic δ-opioid receptors and behavioral antinociception in persistent neuropathic pain

The efficacy of opioids in patients with chronic neuropathic pain remains controversial. Although activation of δ-opioid receptors (DORs) in the brainstem reduces inflammation-induced persistent hyperalgesia, it is not effective under persistent neuropathic pain conditions and these clinical problems remain largely unknown. In this study, by using a chronic constriction injury (CCI) of the sciatic nerve in rats, we found that in the brainstem nucleus raphe magnus (NRM), DORs emerged on the surface membrane of central synaptic terminals on day 3 after CCI surgery and disappeared on day 14. Histone deacetylase (HDAC) inhibitors microinjected into the NRM in vivo increased the level of synaptosomal DOR protein and NRM infusion of DOR agonists producing an antinociceptive effect in a nerve growth factor (NGF) signaling-dependent manner. In vitro, in CCI rat slices incubated with HDAC inhibitors, DOR agonists significantly inhibited EPSCs. This effect was blocked by tyrosine receptor kinase A antagonists. Chromatin immunoprecipitation analysis revealed that NRM infusion of HDAC inhibitors in CCI rats increased the level of histone H4 acetylation at Ngf gene promoter regions. NGF was infused into the NRM or incubated CCI rat slices drove DORs to the surface membrane of synaptic terminals. Taken together, epigenetic upregulation of NGF activity by HDAC inhibitors in the NRM promotes the trafficking of DORs to pain-modulating neuronal synapses under neuropathic pain conditions, leading to δ-opioid analgesia. These findings indicate that therapeutic use of DOR agonists combined with HDAC inhibitors might be effective in chronic neuropathic pain managements.

Pits, a protein interacting with Ttk69 and Sin3A, has links to histone deacetylation.

Histone deacetylation plays an important role in transcriptional repression. Previous results showed that the genetic interaction between ttk and rpd3, which encodes a class I histone deacetylase, is required for tll repression. This study investigated the molecular mechanism by which Ttk69 recruits Rpd3. Using yeast two-hybrid screening and datamining, one novel protein was found that weakly interacts with Ttk69 and Sin3A, designated as Protein interacting with Ttk69 and Sin3A (Pits). Pits protein expressed in the early stages of embryos and bound to the region of the tor response element in vivo. Expanded tll expression patterns were observed in embryos lacking maternal pits activity and the expansion was not widened by reducing either maternal ttk or sin3A activity. However, in embryos with simultaneously reduced maternal pits and sin3A activities or maternal pits, sin3A and ttk activities, the proportions of the embryos with expanded tll expression were significantly increased. These results indicate that all three gene activities are involved in tll repression. Level of histone H3 acetylation in the tll proximal region was found to be elevated in embryo with reduced these three gene activities. In conclusion, Ttk69 causes the histone deacetylation-mediated repression of tll via the interaction of Pits and Sin3A.

Abstract P319: Chronic Ischemia Induces Global Changes in Placental Epigenetic Modifications

Preeclampsia is a common and serious obstetrical complication, hallmarked by new-onset hypertension and maternal endothelial dysfunction. Believed to result as a consequence of placental insufficiency and chronic placental ischemia, the symptoms of preeclampsia are caused by release of pathogenic factors from the placenta itself. A number of these factors have been identified, and it is likely that there are molecular mechanisms which remain obscure. The molecular regulation of even the known factors is often unclear. Here, we have used an established rodent model of placental ischemia which mimics many of the aspects of the human disorder to determine the direct effects of ischemia on global epigenetic modification in the placenta proper. In response to placental insufficiency, maternal blood pressure and rate of fetal demise increased significantly, while placental and fetal mass were decreased. Whole placental levels of histone H3 acetylation at K9 and K27 were increased ~2 and 3 fold respectively. Global methylation of cytosine from placental DNA was low in both groups (<1%), but there was ~50% increase in 5-mC in response to chronic ischemia. These results suggest that chronic placental ischemia induces global changes in epigenetic modification of chromatin in the placenta. This could be an important factor in the regulation of known and unknown pathogenic factors produced by the placenta in the preeclampsia patient.

Spirulina diet to lactating mothers protects the antioxidant system and reduces inflammation in post-natal brain after systemic inflammation

Objectives: This study concerns: (1) the long-term effects of peripheral lipopolysaccharide (LPS) in neonatal rats on inflammation and antioxidant parameters in brain and (2) the effects of a Spirulina-enriched diet given to lactating mothers on protective and inflammatory parameters in brains of suckling pups subjected to peripheral inflammation.Methods: Five-day old rat pups were treated with LPS (i.p. 2 mg/kg). After 3, 7, 30, and 65 days, mRNA, miRNA, and protein levels of pro-inflammatory cytokines and the Nuclear factor E2-related factor 2 (Nrf2)-system were examined. In a sub-group, a Spirulina-enriched diet was given to the mothers 24 hours before the pups were treated with LPS, then the effects on antioxidant and inflammatory parameters were evaluated.Results: The main findings were: (1) interleukin 1 beta (IL-1β) was upregulated in cortex 3, 7, and 30 days after LPS treatment, (2) Nrf2 and the catalytic subunit of γ-glutamylcysteinyl ligase were decreased in cortex 7 days after LPS in parallel with increased levels of phosphorylated p38 and decreased levels of histone H3 acetylation, and (3) a Spirulina-enriched diet to lactating mothers normalized both the increased IL-1β expression and the decreased antioxidant parameters after LPS. The protective effects of Spirulina were correlated with decreased levels of phosphorylated p38 and high levels of the antioxidant miRNA-146a.Discussion: A Spirulina diet given to lactating mothers can protect against neuroinflammation and decreased antioxidant defence in brain of suckling pups subjected to peripheral inflammation, possibly via decreased activation of p38 and high levels of the antioxidant miRNA-146a.

Phenylpyrrole-based HDAC inhibitors: synthesis, molecular modeling and biological studies.

Histone deacetylases (HDACs) regulate the expression and activity of numerous proteins involved in the initiation and progression of cancer. Currently, three hydroxamate-containing HDAC pan-inhibitors have been approved as antitumor agents. We herein present the development of a series of novel phenylpyrrole-based derivatives stemmed from combined computational and medicinal chemistry efforts to rationally modulate HDAC1/6 isoform selectivity. In vitro activity on HDAC1 and HDAC6 isoforms and the effects of selected analogs on histone H3 and α-tubulin acetylation levels were determined. Cell-based data evidenced, for selected compounds, a promising antitumor potential and low toxicity on normal cells. The newly developed compounds represent a valuable starting point for the development of novel anticancer agents.

A novel histone deacetylase inhibitor Chidamide induces G0/G1 arrest and apoptosis in myelodysplastic syndromes.

Chidamide as a newly designed and synthesized histone deacetylase inhibitor induces an antitumor effect in various cancer, and it has been used in several clinical trials such as peripheral T cell lymphoma (PTCL). Here we demonstrate that Chidamide was able to increase the acetylation levels of histone H3 and decrease HDAC activity in MDS cell lines(SKM-1,MUTZ-1)and AML cell line(KG-1). In vitro, at low concentration (<250nM) of Chidamide inhibited cell proliferation and delayed G0/G1 cell cycle progression by down-regulating CDK2 and regulating p-P53 and P21 protein expression. Meanwhile,it also induced cell apoptosis by down-regulating Bcl-2 and up-regulating cleaved Caspase-3 and Bax protein expression.The results of the present study demonstrates the potential utility of Chidamide for the treatment of Myelodysplastic syndromes.

Astrocyte Reactivity Following Blast Exposure Involves Aberrant Histone Acetylation

Blast induced neurotrauma (BINT) is a prevalent injury within military and civilian populations. The injury is characterized by persistent inflammation at the cellular level which manifests as a multitude of cognitive and functional impairments. Epigenetic regulation of transcription offers an important control mechanism for gene expression and cellular function which may underlie chronic inflammation and result in neurodegeneration. We hypothesize that altered histone acetylation patterns may be involved in blast induced inflammation and the chronic activation of glial cells. This study aimed to elucidate changes to histone acetylation occurring following injury and the roles these changes may have within the pathology. Sprague Dawley rats were subjected to either a 10 or 17 psi blast overpressure within an Advanced Blast Simulator (ABS). Sham animals underwent the same procedures without blast exposure. Memory impairments were measured using the Novel Object Recognition (NOR) test at 2 and 7 days post-injury. Tissues were collected at 7 days for Western blot and immunohistochemistry (IHC) analysis. Sham animals showed intact memory at each time point. The novel object discrimination decreased significantly between two and 7 days for each injury group (p < 0.05). This is indicative of the onset of memory impairment. Western blot analysis showed glial fibrillary acidic protein (GFAP), a known marker of activated astrocytes, was elevated in the prefrontal cortex (PFC) following blast exposure for both injury groups. Analysis of histone protein extract showed no changes in the level of any total histone proteins within the PFC. However, acetylation levels of histone H2b, H3, and H4 were decreased in both groups (p < 0.05). Co-localization immunofluorescence was used to further investigate any potential correlation between decreased histone acetylation and astrocyte activation. These experiments showed a similar decrease in H3 acetylation in astrocytes exposed to a 17 psi blast but not a 10 psi blast. Further investigation of gene expression by polymerase chain reaction (PCR) array, showed dysregulation of several cytokine and cytokine receptors that are involved in neuroinflammatory processes. We have shown aberrant histone acetylation patterns involved in blast induced astrogliosis and cognitive impairments. Further understanding of their role in the injury progression may lead to novel therapeutic targets.

Abnormal histone acetylation of CD8+ T cells in patients with severe aplastic anemia.

Severe aplastic anemia (SAA) is a rare autoimmune disease characterized by severe pancytopenia and bone marrow failure, which is caused by activated T lymphocytes. In the present study, we evaluated histone H3 acetylation levels of bone marrow CD8+ T cells in SAA patients, and analyzed its correlation with clinical condition parameters. We found that the percentages of CD8+ T cell histone H3 acetylation in patients with untreated SAA, recovering SAA (R-SAA) and normal control, were 1.21±0.08, 1.05±0.36, and 1.00±0.41, respectively, with no significant statistical differences. However, the amount of CD8+ T cell histone H3 acetylation from untreated SAA was 176.21±32.22μg/mg protein, which was significantly higher than that of complete response (CR)-SAA (104.29±62.06μg/mg protein) and normal control (133.94±56.27μg/mg protein) (P<0.05) groups. Moreover, histone H3 acetylation amount of CD8+ T cell was significantly and negatively correlated with absolute neutrophil count, proportion of reticulocytes, ratio of CD4+ to CD8+ T cell in peripheral blood, and percentage of bone marrow erythroid (P<0.05). To some extent, it also negatively correlated with hemoglobin level, platelet count, percentage of bone marrow granulocyte, and megakaryocyte count. Abnormal histone H3 acetylation of CD8+ T cells may thus play a role in the immune pathogenesis of SAA.

Effects of Sodium Butyrate Treatment on Histone Modifications and the Expression of Genes Related to Epigenetic Regulatory Mechanisms and Immune Response in European Sea Bass (Dicentrarchus Labrax) Fed a Plant-Based Diet.

Bacteria that inhabit the epithelium of the animals’ digestive tract provide the essential biochemical pathways for fermenting otherwise indigestible dietary fibers, leading to the production of short-chain fatty acids (SCFAs). Of the major SCFAs, butyrate has received particular attention due to its numerous positive effects on the health of the intestinal tract and peripheral tissues. The mechanisms of action of this four-carbon chain organic acid are different; many of these are related to its potent regulatory effect on gene expression since butyrate is a histone deacetylase inhibitor that play a predominant role in the epigenetic regulation of gene expression and cell function. In the present work, we investigated in the European sea bass (Dicentrarchus labrax) the effects of butyrate used as a feed additive on fish epigenetics as well as its regulatory role in mucosal protection and immune homeostasis through impact on gene expression. Seven target genes related to inflammatory response and reinforcement of the epithelial defense barrier [tnfα (tumor necrosis factor alpha) il1β, (interleukin 1beta), il-6, il-8, il-10, and muc2 (mucin 2)] and five target genes related to epigenetic modifications [dicer1(double-stranded RNA-specific endoribonuclease), ehmt2 (euchromatic histone-lysine-N-methyltransferase 2), pcgf2 (polycomb group ring finger 2), hdac11 (histone deacetylase-11), and jarid2a (jumonji)] were analyzed in fish intestine and liver. We also investigated the effect of dietary butyrate supplementation on histone acetylation, by performing an immunoblotting analysis on liver core histone extracts. Results of the eight-week-long feeding trial showed no significant differences in weight gain or SGR (specific growth rate) of sea bass that received 0.2% sodium butyrate supplementation in the diet in comparison to control fish that received a diet without Na-butyrate. Dietary butyrate led to a twofold increase in the acetylation level of histone H4 at lysine 8, but showed no effect on the histone H3 at Lys9. Moreover, two different isoforms of histone H3 that might correspond to the H3.1 and H3.2 isoforms previously found in terrestrial animals were separated on the immunoblots. The expression of four (il1 β, il8, irf1, and tnfα) out of seven analyzed genes related to mucosal protection and inflammatory response was significantly different between the two analyzed tissues but only il10 showed differences in expression due to the interaction between tissue and butyrate treatment. In addition, butyrate caused significant changes in vivo in the expression of genes related to epigenetic regulatory mechanisms such as hdac11, ehmt2, and dicer1. Statistical analysis by two-way ANOVA for these genes showed not only significant differences due to the butyrate treatment, but also due to the interaction between tissue and treatment.

Differential effect of treadmill exercise on histone deacetylase activity in rat striatum at different stages of development.

The study described herein aimed to evaluate the impact of exercise on histone acetylation markers in striatum from Wistar rats at different stages of development. Male Wistar rats were submitted to two different exercise protocols: a single session of treadmill (running 20min) or a moderate daily exercise protocol (running 20min for 2weeks). Striata of rats aged 39days postnatal (adolescents), 3months (young adults), and 20months (aged) were used. The single exercise session induced persistent effects on global HDAC activity only in the adolescent group, given that exercised rats showed decreased HDAC activity 1 and 18h after training, without effect on histone H4 acetylation levels. However, the moderate daily exercise did not alter any histone acetylation marker in adolescent and mature groups in any time point evaluated after training. In sum, our data suggest that exercise impacts striatal HDAC activity in an age- and protocol-dependent manner. Specifically, this response seems to be more evident during the adolescent period and might suffer a molecular adaptation in response to chronic training.

Treadmill exercise induces age and protocol-dependent epigenetic changes in prefrontal cortex of Wistar rats

Some studies have linked age-related beneficial effects of exercise and epigenetic mechanisms. Although, the impact of treadmill exercise on histone acetylation, histone and DNA methylation marks in aged cortices yet remains poorly understood. Considering the role of frontal cortex on brain functions, we investigated the potential of different exercise protocols, single session and daily exercise, to modulate epigenetic marks, namely global H4 acetylation, histone methyltransferase activity (HMT H3K27) and levels of DNA methytransferase (DNMT1 and DNMT3b) in prefrontal cortices from 3 and 21-months aged Wistar rats. The animals were submitted to two treadmill exercise protocols, single session (20min) or daily moderate (20min/day during 14days). The daily exercise protocol induced an increased in histone H4 acetylation levels in prefrontal cortices of 21-months-old rats, without any effects in young adult group. DNMT3b levels were increased in aged cortices of animals submitted to single session of exercise. These results indicate that prefrontal cortex is susceptible to epigenetic changes in a protocol dependent-manner and that H4 acetylation levels and DNMT3b content changes might be linked at least in part to exercise-induced effects on brain functions.

The Banana Transcriptional Repressor MaDEAR1 Negatively Regulates Cell Wall-Modifying Genes Involved in Fruit Ripening.

Ethylene plays an essential role in many biological processes including fruit ripening via modulation of ethylene signaling pathway. Ethylene Response Factors (ERFs) are key transcription factors (TFs) involved in ethylene perception and are divided into AP2, RAV, ERF, and DREB sub-families. Although a number of studies have implicated the involvement of DREB sub-family genes in stress responses, little is known about their roles in fruit ripening. In this study, we identified a DREB TF with a EAR motif, designated as MaDEAR1, which is a nucleus-localized transcriptional repressor. Expression analysis indicated that MaDEAR1 expression was repressed by ethylene, with reduced levels of histone H3 and H4 acetylation at its regulatory regions during fruit ripening. In addition, MaDEAR1 promoter activity was also suppressed in response to ethylene treatment. More importantly, MaDEAR1 directly binds to the DRE/CRT motifs in promoters of several cell wall-modifying genes including MaEXP1/3, MaPG1, MaXTH10, MaPL3, and MaPME3 associated with fruit softening during ripening and represses their activities. These data suggest that MaDEAR1 acts as a transcriptional repressor of cell wall-modifying genes, and may be negatively involved in ethylene-mediated ripening of banana fruit. Our findings provide new insights into the involvement of DREB TFs in the regulation of fruit ripening.

Mechanical stimulation orchestrates the osteogenic differentiation of human bone marrow stromal cells by regulating HDAC1.

Mechanical stimulation and histone deacetylases (HDACs) have essential roles in regulating the osteogenic differentiation of bone marrow stromal cells (BMSCs) and bone formation. However, little is known regarding what regulates HDAC expression and therefore the osteogenic differentiation of BMSCs during osteogenesis. In this study, we investigated whether mechanical loading regulates HDAC expression directly and examined the role of HDACs in mechanical loading-triggered osteogenic differentiation and bone formation. We first studied the microarrays of samples from patients with osteoporosis and found that the NOTCH pathway and skeletal development gene sets were downregulated in the BMSCs of patients with osteoporosis. Then we demonstrated that mechanical stimuli can regulate osteogenesis and bone formation both in vivo and in vitro. NOTCH signaling was upregulated during cyclic mechanical stretch (CMS)-induced osteogenic differentiation, whereas HDAC1 protein expression was downregulated. The perturbation of HDAC1 expression also had a significant effect on matrix mineralization and JAG1-mediated Notch signaling, suggesting that HDAC1 acts as an endogenous attenuator of Notch signaling in the mechanotransduction of BMSCs. Chromatin immunoprecipitation (ChIP) assay results suggest that HDAC1 modulates the CMS-induced histone H3 acetylation level at the JAG1 promoter. More importantly, we found an inhibitory role of Hdac1 in regulating bone formation in response to hindlimb unloading in mice, and pretreatment with an HDAC1 inhibitor partly rescued the osteoporosis caused by mechanical unloading. Our results demonstrate, for the first time, that mechanical stimulation orchestrates genes expression involved in the osteogenic differentiation of BMSCs via the direct regulation of HDAC1, and the therapeutic inhibition of HDAC1 may be an efficient strategy for enhancing bone formation under mechanical stimulation.

Epigenetic regulation of MdMYB1 is associated with paper bagging-induced red pigmentation of apples.

Paper-bagging treatment can transform non-transcribed MdMYB1 - 2 and MdMYB1 - 3 alleles into transcribed alleles through epigenetic regulations, resulting in the red pigmentation of a normally non-red apple cultivar 'Mutsu.' Anthocyanin biosynthesis in apples is regulated by MdMYB1/A/10, an R2R3-Type MYB gene. 'Mutsu,' a triploid apple cultivar harboring non-transcribed MdMYB1-2 and MdMYB1-3 alleles, retains green skin color under field conditions. However, it can show red/pink pigmentation under natural or artificial ultraviolet-B (UV-B) light exposure after paper-bagging and bag removal treatment. In the present study, we found that in 'Mutsu,' paper bagging-induced red pigmentation was due to the activation of non-transcribed MdMYB1-2/-3 alleles, which triggered the expression of downstream anthocyanin biosynthesis genes in a UV-B-dependent manner. By monitoring the epigenetic changes during UV-B-induced pigmentation, no significant differences in DNA methylation and histone modifications in the 5' upstream region of MdMYB1-2/-3 were recorded between the UV-B-treated fruit skin (red) and the fruit skin treated only by white light (green). In contrast, bag treatment lowered the DNA methylation in this region of MdMYB1-2/-3 alleles. Similarly, higher levels of histone H3 acetylation and trimethylation of H3 tail at lysine 4, and lower level of trimethylation of H3 tail at lysine 27 were observed in the 5' upstream region of MdMYB1-2/-3 in the skin of the fruit immediately after bag removal. These results suggest that bagging treatment can induce epigenetic changes, facilitating the binding of trans factor(s) to MdMYB1-2/-3 alleles, resulting in the activation of these MYBs after bag removal.

086 Combination of retinoid and histone deacetylase inhibitor produced an anti-tumor effect in cutaneous T-cell lymphoma by restoring tumor suppressor gene, retinoic acid receptorβ2, via histone acetylation

Retinoids exert anti-proliferative, differentiative, and apoptosis-inducing effects through their receptors. Retinoic acid receptor (RAR) β2 behaves as a tumor suppressor gene, and its expression is suppressible by DNA methylation in many malignancies. We aimed to determine whether combining a retinoid, Am 80, with a histone deacetylase inhibitor, MS-275, could suppress tumor growth in a RARβ2-negative human cutaneous T cell lymphoma (CTCL) cell lines and freshly isolated primary CTCL cells, and to elucidate the epigenetic mechanism behind the phenomena. SeAx cells were implanted subcutaneously in NOD-SCID mice which were randomly divided into four groups and treated with either Am80, MS-275 by oral gavage (five days/week), or a combination of the two agents. Cell proliferation assay, methylation-specific PCR, flow cytometric analysis of cell cycle and apoptosis and chromatin immunoprecipitation assay were employed.Quantitative PCR analysis revealed that RARβ2 gene expression was restored only by this combination rather than by either of the agents singly. Restored retinoid sensitivity was observed in combining retinoid with a histone deacetylase inhibitor significantly inhibited cell growth in vitro, suppressed subcutaneously transplanted tumor growth, and prolonged survival of tumor-bearing mice in vivo by more strongly inducing apoptosis and p21 expression in CTCL cells than either agent alone. In the combination treatment, the histone H4 acetylation level at lysine 12 and 16 in the promoter region increased after restoration of RARβ2 expression although the DNA methylation of RARβ2 remained unchanged. This is the first report of histone acetylation as the primary event in the restoration of RARβ2. Inducible RARβ2 expression may serve as a reliable predictor for tumor response in patients undergoing ‘epigenetic & differentiation’ therapy.

Mycobacterium tuberculosis EIS gene inhibits macrophage autophagy through up-regulation of IL-10 by increasing the acetylation of histone H3

Autophagy plays a crucial role in the progress of Mycobacterium tuberculosis (MTB) infection. Recently, MTB enhanced intracellular survival (EIS) protein was reported to be secreted from MTB cells and linked to the inhibition of autophagy and the intracellular persistence of the pathogen. Here, we investigated the mechanism of EIS-mediated inhibition of autophagy in a human phorbol myristate acetate (PMA)-treated THP-1 cell line as well as in murine macrophages. We confirmed that the presence of EIS led to the inhibition of rapamycin (Rapa)-induced autophagy, while IL-10 gene expression was increased and Akt/mTOR/p70S6K pathway was activated during the process. IL-10 gene silencing led to a significant recovery of EIS-mediated autophagy suppression and decreased activity of the Akt/mTOR/p70S6K pathway. IL-10 promoter activity was unaffected by EIS. Remarkably, EIS increased the acetylation level of histone H3 (Ac-H3), which binds to the SP1 and STAT3 region of the human IL-10 gene promoter sequence. Thus, EIS protein possibly increased IL-10 expression through the regulation of Ac-H3 of its promoter. Our data demonstrated that one possible mechanism of the MTB evasion of autophagy is that the EIS protein up-regulates IL-10 via Ac-H3 and thus activates Akt/mTOR/p70S6K pathway.

Long non-coding RNA SNHG5 suppresses gastric cancer progression by trapping MTA2 in the cytosol.

Recently, intriguing new roles for some small nucleolar RNA host genes (SNHGs) in cancer have emerged. In the present study, a panel of SNHGs was profiled to detect aberrantly expressed SNHGs in gastric cancer (GC). The expression of SNHG5 was significantly downregulated in GC and was significantly associated with the formation of a tumor embolus and with the tumor, node and metastasis stage. SNHG5 was a long non-coding RNA, which was a class of non-coding RNA transcripts longer than 200 nucleotides. SNHG5 suppressed GC cell proliferation and metastasis in vitro and in vivo. Furthermore, SNHG5 exerted its function through interacting with MTA2, preventing the translocation of MTA2 from the cytoplasm into the nucleus. SNHG5 overexpression led to significant increases in the acetylation levels of histone H3 and p53, indicating that SNHG5 might affect acetylation by trapping MTA2 in the cytosol, thereby interfering with the formation of the nucleosome remodeling and histone deacetylation complex. This study is the first to demonstrate that SNHG5 is a critical and powerful regulator that is involved in GC progression through trapping MTA2 in the cytosol. These results imply that SNHG5 may be a novel therapeutic target for the treatment of GC.