HDAC1 mRNA Research Articles

An Antisense Oligonucleotide Leads to Suppressed Transcription of Hdac2 and Long-Term Memory Enhancement

Knockout of the memory suppressor gene histone deacetylase 2 (Hdac2) in mice elicits cognitive enhancement, and drugs that block HDAC2 have potential as therapeutics for disorders affecting memory. Currently available HDAC2 catalytic activity inhibitors are not fully isoform specific and have short half-lives. Antisense oligonucleotides (ASOs) are drugs that elicit extremely long-lasting, specific inhibition through base pairing with RNA targets. We utilized an ASO to reduce Hdac2 messenger RNA (mRNA) in mice and determined its longevity, specificity, and mechanism of repression. A single injection of the Hdac2-targeted ASO in the central nervous system produced persistent reduction in HDAC2 protein and Hdac2 mRNA levels for 16 weeks. It enhanced object location memory for 8 weeks. RNA sequencing (RNA-seq) analysis of brain tissues revealed that the repression was specific to Hdac2 relative to related Hdac isoforms, and Hdac2 reduction caused alterations in the expression of genes involved in extracellular signal-regulated kinase (ERK) and memory-associated immune signaling pathways. Hdac2-targeted ASOs also suppress a nonpolyadenylated Hdac2 regulatory RNA and elicit direct transcriptional suppression of the Hdac2 gene through stalling RNA polymerase II. These findings identify transcriptional suppression of the target gene as a novel mechanism of action of ASOs.

A histone deacetylase 7-derived peptide promotes vascular regeneration via facilitating 14-3-3γ phosphorylation.

Histone deacetylase 7 (HDAC7) plays a pivotal role in the maintenance of the endothelium integrity. In this study, we demonstrated that the intron‐containing Hdac7 mRNA existed in the cytosol and that ribosomes bound to a short open reading frame (sORF) within the 5′‐terminal noncoding area of this Hdac7 mRNA in response to vascular endothelial growth factor (VEGF) stimulation in the isolated stem cell antigen‐1 positive (Sca1+) vascular progenitor cells (VPCs). A 7‐amino acid (7A) peptide has been demonstrated to be translated from the sORF in Sca1+‐VPCs in vitro and in vivo. The 7A peptide was shown to receive phosphate group from the activated mitogen‐activated protein kinase MEKK1 and transfer it to 14‐3‐3 gamma protein, forming an MEKK1‐7A‐14‐3‐3γ signal pathway downstream VEGF. The exogenous synthetic 7A peptide could increase Sca1+‐VPCs cell migration, re‐endothelialization in the femoral artery injury, and angiogenesis in hind limb ischemia. A Hd7‐7sFLAG transgenic mice line was generated as the loss‐of‐function model, in which the 7A peptide was replaced by a FLAG‐tagged scrabbled peptide. Loss of the endogenous 7A impaired Sca1+‐VPCs cell migration, re‐endothelialization of the injured femoral artery, and angiogenesis in ischemic tissues, which could be partially rescued by the addition of the exogenous 7A/7Ap peptide. This study provides evidence that sORFs can be alternatively translated and the derived peptides may play an important role in physiological processes including vascular remodeling.

Bacosides Encapsulated in Lactoferrin Conjugated PEG-PLA-PCL-OH Based Polymersomes Act as Epigenetic Modulator in Chemically Induced Amnesia.

The present study demonstrates the epigenetic mechanisms underlying the effect of Bacoside rich extract of Bacopa monniera-a nootropic herb, on scopolamine treated amnesic mice conferred via chromatin modifying enzymes. The focus of the work was to elucidate the modulation of the chromatin modifying enzymes: DNMT1, DNMT3a, DNMT3b, HDAC2, HDAC5 and CPB in scopolamine induced amnesic mice after treatment with bacoside rich extract of Bacopa monniera (BA) and BA encapsulated in lactoferrin conjugated PEG-PLA-PCL-OH based polymersomes (BAN). We observed remarkable difference between the results obtained after the treatment with BA and BAN. Interestingly BAN was found to be more efficient in downregulating DNA methylation and histone chain deacetylation. Scopolamine treatment showed up-regulation of DNMT1 expression in qRT-PCR by 3.14-fold as compared to the control, which was considerably decreased by 1.5-fold after treatment with BA and remarkably decreased 0.11-fold by BAN treatment. Scopolamine treatment up-regulated the expression of DNMT3a by 1.6-fold while for DNMT3b by 3.13-fold. In DNMT3a and DNMT3b the fold change decreased to 0.64 and 0.76 after BA treatment, whereas the BAN treatment further down-regulated to 0.32- and 0.63-fold, respectively. Similarly scopolamine up-regulated HDAC2 and HDAC5 by 3.12 fold and 3.64-fold, respectively. BA treatment reversed the changes by reducing HDAC2 mRNA to 0.89-fold and HDAC5 mRNA 0.83-fold. BAN further reduced expression of HDAC2 further to 0.39-fold and HDAC5 to 0.31-fold. On the other hand scopolamine down-regulated CBP mRNA expression by 0.28-fold and increased by 1.09 after BA treatment. BAN significantly increased the CPB expression by 1.65-fold as compared to BA treatment. These findings were consolidated by DNMT and HDAC enzyme activity assay, methylation in the promoter region of the memory related genes: ARC and BDNF and Dot blot assay for DNA methylation. The percent activity increase of DNMT and HDAC after scopolamine administration was 375.74 and 240.90 respectively. After treatment with BA the downfall in percent activity was observed as 167.99 in DMNT and 130.57 in HDAC. BAN treatment further decreased the percent enzyme activity of DNMT and HDAC significantly by 30.0 and 61.81 respectively. The potency of BAN in reversing the epigenetic changes of scopolamine induced amnesic mouse brain, can be attributed to the brain specific delivery of BA through polymersomes which are able to cross the blood brain barrier (BBB) via receptor mediated endocytosis.

P048 Mucosal macrophages express elevated levels of HDAC9 in inflamed and uninflamed mucosa of Crohn’s disease, but not ulcerative colitis

Abstract Background Histone deacetylases (HDACs) are a group of enzymes that control histone and non-histone deacetylation and influence inflammatory gene transcription. Certain members of the HDAC family control the function of macrophages and play an important role in immune response. In this study, we aimed to study the expression of HDACs in mucosal macrophages isolated from inflammatory bowel diseases (IBD) patients. Methods Both macroscopically inflamed and non-inflamed colon resection tissue were collected from 15 Crohn’s disease (CD) and nine ulcerative colitis (UC) patients operated on for therapy refractory disease. Of the CD patients, 53% had ileal and 47% ileocolonic disease. Of the UC patients, 44% had left-sided colitis and 56% pancolitis. Lamina propria was separated from the muscularis externa, and a targeted array for epigenetic enzymes was performed. To assess the relevance of HDAC9 gene expression in terms of protein level, immunofluorescence staining of HDAC9 protein was undertaken in tissue sections from inflamed and non-inflamed mucosa. CD68 was used as a pan-macrophage marker. Results From our array, expression of HDAC9 was significantly higher in the inflamed mucosa of CD patients compared with UC patients (p = 0.005). Gene expression of HDAC9 in non-inflamed mucosa from CD was elevated compared with non-inflamed mucosa from UC. In addition, in CD, HDAC9 mRNA level was increased in inflamed tissue in comparison to non-inflamed tissue (p = 0.046). In conjunction with the expression data, HDAC9 protein was found highly expressed in inflamed tissue. HDAC9 was predominantly localised in the cytoplasmic compartment of macrophages in non-inflamed tissue whilst HDAC9 localised to the nucleus of macrophages in inflamed tissue. Conclusion HDAC9 is member of class IIA HDAC superfamily that exerts pro-inflammatory properties. The inhibition of HDAC9 in experimental murine colitis clearly enhances regulatory T-cell function, suggesting a critical role for HDAC9 in breaching immune homeostasis (de Zoeten EF et al, 2009). We suggest here that HDAC9 can serve as an additional marker to distinguish CD from UC in tissue biopsies. Furthermore, we show for the first time that HDAC9 protein is expressed in mucosal macrophages of CD patients, indicating its potential in mediating macrophage inflammatory function in IBD. Further studies are currently being undertaken to elucidate the role of HDAC9 in CD pathogenesis.

Changes in the expression of genes involved in DNA methylation and histone modification in response to daily food availability times in zebra finches: epigenetic implications.

We hypothesised that daily food availability times serve as an 'epigenetic' factor and affect reproductive physiology in continuously reproducing species. This we tested by measuring mRNA expression of genes coding for enzymes involved in DNA methylation-demethylation (dnmt, tet) and histone modification (hat1, hdac) in the hypothalamus, liver and gonads of male and female zebra finches that were paired for a year under 12h light:12h dark conditions with food availability restricted to 4h in the morning (morning FA group) or evening (evening FA group), with controls provided with food ad libitum The overall hypothalamic and hepatic expression patterns of hat1 and hdac were similar but those of dnmt and tet were different between males and females. Irrespective of the timing of food availability, both hat1 and hdac mRNA levels were increased in the hypothalamus, but not in the liver, in which hat1 mRNA levels were increased in the morning FA group. While hypothalamic tet levels were higher in evening FA males, hepatic tet levels were higher in morning FA birds (tet1, only males). Gonadal expression levels similarly varied and showed sex differences. Histone-modifying genes did not show food availability effects, except for elevated testicular hdac3 levels. Similarly, testicular dnmt3b and tet2 mRNA levels were increased and decreased in morning and evening FA groups, respectively, whereas ovarian dnmt1 and tet2 levels were reduced in morning FA and tet1 levels were reduced in evening FA groups. The present results suggest that an enforced daily feeding schedule in the long term could serve as a conditioning environment that shapes overall hypothalamic regulation, and liver and gonadal function at the epigenetic level in diurnal vertebrates.

GPx1-mediated DNMT1 expression is involved in the blocking effects of selenium on OTA-induced cytotoxicity and DNA damage

Ochratoxin A (OTA) is a potent nephrotoxin. Selenium (Se) is an essential micronutrient for humans and animals, and plays a key role in antioxidant defense. To date, little is known about the effect of Se on OTA-induced DNA damage. In this study, the protective effects of Se (from selenomethionine) against OTA-induced cytotoxicity and DNA damage were investigated by using PK15 cells as a model. The results showed that OTA at 4.0 μg/mL induced cytotoxicity and DNA damage. Se at 0.5, 1, 2 and 4 μM significantly blocked OTA-induced cytotoxicity and DNA damage. Furthermore, Se blocked the increases of DNMT1, DNMT3a and HDAC1 mRNA and protein expression, reversed the decreases of glutathione peroxidase 1 (GPx1) mRNA and protein expression, and promoted the increases of SOCS3 mRNA and protein expression induced by OTA. Overexpression of GPx1 by pcDNA3.1-GPx1 inhibited the OTA-induced DNMT1 expression, promoted OTA-induced SOCS3 expression, and prevented the OTA-induced cytotoxicity and DNA damage. In contrast, knock-down of GPx1 by using a GPx1-specific siRNA had the opposite effects. The results suggest that GPx1-mediated DNMT1 expression is involved in the blocking effects of selenium on OTA-induced cytotoxicity and DNA damage.

HDAC4 Levels Control Sensibility toward Cisplatin in Gastric Cancer via the p53-p73/BIK Pathway.

Gastric cancer (GC) remains a health issue due to the low efficiency of therapies, such as cisplatin. This unsatisfactory situation highlights the necessity of finding factors impacting GC sensibility to therapies. We analyzed the cisplatin pangenomic response in cancer cells and found HDAC4 as a major epigenetic regulator being inhibited. HDAC4 mRNA repression was partly mediated by the cisplatin-induced expression of miR-140. At a functional level, HDAC4 inhibition favored cisplatin cytotoxicity and reduced tumor growth. Inversely, overexpression of HDAC4 inhibits cisplatin cytotoxicity. Importantly, HDAC4 expression was found to be elevated in gastric tumors compared to healthy tissues, and in particular in specific molecular subgroups. Furthermore, mutations in HDAC4 correlate with good prognosis. Pathway analysis of genes whose expression in patients correlated strongly with HDAC4 highlighted DNA damage, p53 stabilization, and apoptosis as processes downregulated by HDAC4. This was further confirmed by silencing of HDAC4, which favored cisplatin-induced apoptosis characterized by cleavage of caspase 3 and induction of proapoptotic genes, such as BIK, in part via a p53-dependent mechanism. Altogether, these results reveal HDAC4 as a resistance factor for cisplatin in GC cells that impacts on patients’ survival.

Early-life blockade of NMDA receptors induces epigenetic abnormalities in the adult medial prefrontal cortex: possible involvement in memory impairment in trace fear conditioning

RationaleSeveral findings indicate that early-life dysfunction of N-methyl-d-aspartate (NMDA) receptors might cause schizophrenia-like abnormalities in adulthood that might be induced by impairments in epigenetic regulation.ObjectivesIn the present study, we investigated whether postnatal blockade of NMDA receptors (within the first 3 weeks of life) by the competitive antagonist CGP 37849 (CGP) might affect some epigenetic markers in the adult medial prefrontal cortex (mPFC).MethodsHistone H3 phosphorylation at serine 10 (H3S10ph), histone H3 acetylation at lysine 9 or 14 (H3K9ac or H3K14ac, respectively), or expression of histone deacetylase (HDAC) 2, HDAC5, myocyte enhancer factor (MEF) 2D and activity-regulated cytoskeleton-associated protein (Arc) were analysed. Moreover, we also evaluated whether the deacetylase inhibitor sodium butyrate (SB; 1.2 mg/kg, ip) could prevent behavioural and neurochemical changes in the mPFC induced by CGP during memory retrieval in the trace fear conditioning paradigm.ResultsThe results showed that CGP administration increased the number of H3S10ph nuclei but did not affect H3K9ac and H3K14ac or HDAC2 protein levels. However, CGP administration altered the HDAC5 mRNA and protein levels and increased the mRNA and protein levels of MEF2D. CGP also increased Arc mRNA, which was correlated with an increase in the amount of Arc DNA bound to MEF2D. SB given 2 h after training prevented impairment of the freezing response and disruption of epigenetic markers (H3S10ph, HDAC5, MEF2D) and Arc expression during memory retrieval induced by CGP administration.ConclusionsThe early-life blockade of NMDA receptors impairs some epigenetic regulatory processes in the mPFC that are involved in fear memory formation.

Comparison of Epigenetic Modifier Genes in Bovine Adipose Tissue-Derived Stem Cell Based Embryos, as Donors, with In Vitro and Parthenogenesis Embryos.

Objective Regarding that undifferentiated mesenchymal stem cells, as donor cells, require less epigenetic reprogramming, possibility of using bovine adipose tissue-derived stem cells (BASCs) with low level of DNMTs and HDACs expression was evaluated. Materials and Methods In this experimental study, we examined gene expression of epigenetic modifiers including DNA methyltransferases (DNMT1, DNMT3A and DNMT3B) and histone deacetylases (HDAC1-3), as well as protein levels of histone H3 acetylation at lysine 9 (H3K9ac) and POU5F1 (also known as OCT4) at two stages of preimplantation development among in vitro fertilization (IVF), parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT) groups. Results The results revealed that developmental competence of IVF embryos was higher than SCNT embryos (P<0.05). In the PA and SCNT groups, DNMT1, HDAC2 and HDAC3 mRNA were overexpressed (P<0.05), and proteins levels of H3K9ac and POU5F1 were reduced at 6-8 cells and blastocyst stages compared to IVF (P<0.05). The mRNA expression of DNMT1 an<0.05) in both developmental stages (except HDAC1 in blastocyst stage). ConclusionThe SCNT embryos derived from BASCs have endured considerable nuclear reprogramming during early embryo development. Comparison of PA and SCNT blastocysts demonstrated that HDAC1 and DNMT1 may attribute to developmental competence variability of bovine embryos.

Cinnamic acid rescues behavioral deficits in a mouse model of traumatic brain injury by targeting miR-455-3p/HDAC2

AimsTraumatic brain injury (TBI) not only induces physiological disabilities but also leads to cognitive impairment. However, no effective therapeutic approach for TBI-related memory decline exists. In this study, we treated TBI mice with cinnamic acid (CNA) to detect whether CNA is able to rescue the memory deficits induced by TBI and to explore the potential mechanisms. Main methodsMice were divided into the following groups: the sham group, the TBI group, the TBI + CNA group and the CNA group. Basic physiological parameters, neurological severity score and brain water content were analyzed. The Morris water maze and inhibitory avoidance step-down task were used to determine learning and memory. Golgi staining was used to measure alterations in dendritic spines. Western blot analysis and a commercial kit were used to detect the content and activity of HDAC2. qPCR was used to detect the relative level of miR-455. Key findingsCNA did not affect physiological function but effectively restored neurological function and brain edema. CNA alleviated the memory impairments induced by TBI in both the Morris water maze and step-down task. CNA also recovered abnormalities in the synapses of TBI mice by suppressing the activity of HDAC2. Furthermore, CNA did not alter HDAC mRNA because it promoted the expression of miR-455-3p, a miRNA that regulates HDAC2 at the posttranscriptional level. SignificanceThe application of CNA effectively treats TBI-induced memory deficits by increasing miR-455-3p and by inhibiting HDAC2.

MicroRNA-365 functions as a mechanosensitive microRNA to inhibit end plate chondrocyte degeneration by targeting histone deacetylase 4.

End plate chondrocyte degeneration is a major cause of intervertebral disc degeneration. Mechanical biophysical forces, including intermittent cyclic mechanical tension (ICMT), exacerbate end plate chondrocyte degeneration. However, the underlying molecular mechanism of mechanical stretch-induced end plate chondrocyte degeneration is still unclear. This study sought to determine whether microRNAs (miRNAs) respond to mechanical stretch and play a role in regulating mechanically-induced end plate chondrocyte degeneration. We identified miR-365 as a mechanoresponsive miRNA in primary human end plate chondrocytes after ICMT application by miRNA microarray analysis. The expression of miR-365 was down-regulated in the disc samples obtained from patients with disc degeneration. We also found that the miR-365 stimulates chondrocyte proliferation but does not promote end plate chondrocyte death. Using bioinformatic analyses and subsequent confirmation by real-time RT-PCR, we identified multiple candidate target genes of miR-365 that responded to in vitro mechanical stimulation; among them, HDAC4 was fully characterized. Mutation of putative miR-365 binding sites in HDAC4 mRNA abolished miR-365 mediated repression of HDAC4 3'-untranslated region (3'UTR) luciferase reporter activity, suggesting that miR-365 binds to the HDAC4 3'UTR. Overexpression of miR-365 significantly decreased the HDAC4 protein level, suggesting that miR-365 acts as an endogenous attenuator of HDAC4 in human end plate chondrocytes. Further, perturbation of miR-365 expression also had a significant effect on the expression of COL2A and ACAN and on matrix degeneration. Overexpression of HDAC4 abolished miR-365 rescued end plate chondrocyte degeneration during ICMT application. Furthermore, we found that the wnt/β-catenin signal pathway was related to HDAC4 and promoted end plate chondrocyte degeneration. Overall, our results suggest that miR-365 is a mechanosensitive miRNA that regulates human chondrocyte degeneration by directly targeting HDAC4. We propose that therapeutic regulation of miR-365 may be an efficient anabolic strategy for inhibiting end plate chondrocyte degeneration.

Oncogenic enhancer of zeste homolog 2 is an actionable target in patients with non-small cell lung cancer.

BackgroundThe aims of this study were to investigate the link between enhancer of zeste homolog 2 (EZH2) and histone deacetylase (HDAC) in preclinical studies and in human lung cancer tissue microarrays.MethodsEnhancer of zeste homolog 2 and HDAC1 mRNA expression in two lung adenocarcinoma (LUAD) datasets (MDACC and TCGA) were correlated with patient outcomes. We evaluated the association of EZH2 and HDAC1 expression with response to the HDAC1 inhibitor, suberoylanilide hydroxamic acid (SAHA). The response to SAHA was assessed at baseline and after alteration of EZH2 or HDAC mRNA expression in LUAD cell lines.ResultsDirect correlation was found between EZH2 and HDAC1 expression (P < 0.0001). When EZH2 expression was knocked down‐ or upregulated, there was a corresponding decrease or increase in expression of HDAC expression, respectively. Cell lines with high EZH2 expression responded to SAHA treatment with a mean inhibition rate of 73.1% compared to 43.2% in cell lines with low EZH2 expression (P < 0.0001). This correlation was confirmed in non‐small cell lung cancer (NSCLC) specimens from MDACC (Spearman's correlation r = 0.416; P < 0.0001) and TCGA datasets (r = 0.221; P < 0.0001). Patients with high EZH2 and high HDAC1 expression in stage I NSCLC specimens of both datasets had the lowest survival compared to the patients with low expression of either or both markers.ConclusionOur findings show that overexpression of EZH2 is a negative prognostic indicator. Increased EZH2 expression predicts for response to HDAC inhibitors and thus could serve as a biomarker for selecting NSCLC patients for treatment with HDAC inhibitors.

Sin3a regulates the developmental progression through morula-to-blastocyst transition via Hdac1.

Suppressor interacting 3a (Sin3a) is a scaffold component of the chromatin repressive complex Sin3/histone deacetylase (Hdac). Sin3a has been shown as a hub gene driving preimplantation development in both mice and humans. However, its precise functions during preimplantation development remain unclear. Here, we show that the embryos arrested at morula stage upon specific depletion of Sin3a in mouse early embryos. Given the reduced cell number in Sin3a-depleted embryos, blocked cell proliferation is observed, likely because of the increased level of Trp53 acetylation at lysine 379. Moreover, we found that Sin3a depletion reduces Cdx2 and Tir Na Nog (Nanog), suggesting a failure of the first cell fate decision. In addition, we noted a striking increase of genome-wide DNA methylation, likely attributed to the increased nuclear DNA methyltransferase 1 observed in Sin3a-depleted embryos. Notably, RNA sequencing analyses showed 717 genes are differentially expressed, and Gene Ontology analysis of down-regulated genes (e.g., Hdac1) revealed top enriched terms involving protein deacetylation. Consistently, we confirmed a significant decrease of Hdac1 mRNA and protein abundance. Importantly, the development and Trp53 acetylation in Sin3a-depleted embryos could be rescued by expression of Hdac1 but not Hdac2. In summary, our results indicate a vital role of Sin3a in safeguarding the developmental progression through the morula-to-blastocyst transition via Hdac1.-Zhao, P., Li, S., Wang, H., Dang, Y., Wang, L., Liu, T., Wang, S., Li, X., Zhang, K. Sin3a regulates the developmental progression through morula-to-blastocyst transition via Hdac1.

Cyclic tensile strain promotes chondrogenesis of bone marrow-derived mesenchymal stem cells by increasing miR-365 expression

AimsThe chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) is critical for cartilage regeneration. Tissues constructed from BMSCs through cartilage tissue engineering still exhibit some histological, morphological, and biomechanical differences from normal cartilage tissues. Cyclic tensile strain (CTS) can increase chondrogenic gene expression and reduce hypertrophic gene expression in chondrocytes. miR-365 has been identified as a mechanoresponsive microRNA and is an important regulator of both chondrocyte hypertrophy and differentiation. Therefore, we hypothesized that CTS may promote the chondrogenesis of BMSCs by upregulating the expression of miR-365. MethodsBMSCs were subjected to CTS to investigate the effects and mechanism on chondrogenesis. An Agilent miRNA microarray was used to profile miRNAs in the CTS-treated BMSCs and 3D-cultured control BMSCs. miR-365 was shown to interact with HDAC4 mRNA through a luciferase reporter assay. An animal cartilage defect model was constructed and different groups of BMSCs were implanted to investigate their in vivo effect. Key findingsCTS promoted BMSC chondrogenesis. miR-365 was significantly upregulated in CTS-treated cells and played an important role in CTS-mediated chondrogenesis. Luciferase assays showed that HDAC4 is a direct target of miR-365. An in vivo study showed that CTS treatment and miR-365 overexpression could promote cartilage regeneration from BMSCs. SignificanceCTS can promote the expression of miR-365, a crucial mechanosensitive microRNA involved in the chondrogenesis of BMSCs, which directly inhibits the expression of HDAC4, in turn, enhancing the chondrogenesis of BMSCs.

Increased expression of the HDAC9 gene is associated with antiestrogen resistance of breast cancers.

Estrogens play a pivotal role in breast cancer etiology, and endocrine therapy remains the main first line treatment for estrogen receptor‐alpha (ERα)‐positive breast cancer. ER are transcription factors whose activity is finely regulated by various regulatory complexes, including histone deacetylases (HDACs). Here, we investigated the role of HDAC9 in ERα signaling and response to antiestrogens in breast cancer cells. Various Michigan Cancer Foundation‐7 (MCF7) breast cancer cell lines that overexpress class IIa HDAC9 or that are resistant to the partial antiestrogen 4‐hydroxy‐tamoxifen (OHTam) were used to study phenotypic changes in response to ER ligands by using transcriptomic and gene set enrichment analyses. Kaplan–Meier survival analyses were performed using public transcriptomic datasets from human breast cancer biopsies. In MCF7 breast cancer cells, HDAC9 decreased ERα mRNA and protein expression and inhibited its transcriptional activity. Conversely, HDAC9 mRNA was strongly overexpressed in OHTam‐resistant MCF7 cells and in ERα‐negative breast tumor cell lines. Moreover, HDAC9‐overexpressing cells were less sensitive to OHTam antiproliferative effects compared with parental MCF7 cells. Several genes (including MUC1, SMC3 and S100P) were similarly deregulated in OHTam‐resistant and in HDAC9‐overexpressing MCF7 cells. Finally, HDAC9 expression was positively associated with genes upregulated in endocrine therapy‐resistant breast cancers and high HDAC9 levels were associated with worse prognosis in patients treated with OHTam. These results demonstrate the complex interactions of class IIa HDAC9 with ERα signaling in breast cancer cells and its effect on the response to hormone therapy.

Effect of HDAC2/Inpp5f on neuropathic pain and cognitive function through regulating PI3K/Akt/GSK-3β signal pathway in rats with neuropathic pain.

The effect of histone deacetylase (HDAC)2/Inositol polyphosphate-5-phosphatase F (Inpp5f) on neuropathic pain and cognitive dysfunction through regulating PI3K/Akt/GSK-3β signal pathway in rats with neuropathic pain was investigated. A total of 80 SPF mature male SD rats were averagely randomized into the sham operation group, the model group, the HDAC2 intervention group (group A) and the Inpp5f intervention group (group B). The rat models of neuropathic pain were established in the model group, and groups A and B. At the 15th day after modeling, rats in group A were transfected with the interference vector of HDAC2, and rats in group B were transfected with the overexpression vector of Inpp5f. Rats in the four groups were observed before modeling, after modeling/before intervention and 3 days after intervention in terms of paw thermal withdrawal latency (PWL), paw withdrawal mechanical threshold (PWT) and changes in cognitive function (Morris water maze and passive avoidance task). Then the rats were sacrificed. RT-qPCR and western blot analysis were used to detect the levels of HDAC2 mRNA, Inpp5f mRNA, phosphorylated PI3K (p-PI3K), phosphorylated AKT (p-AKT), phosphorylated GSK-3β (p-GSK-3β) in rat brain tissue. Correlation of HDAC2 mRNA with Inpp5f mRNA expression levels was detected by Pearsons correlation analysis. Compared with the sham operation group, PWL was significantly lower while PWT was higher in the other 3 groups (P<0.05). Three days after intervention, PWL was significantly higher while PWT was significantly lower (P<0.05). Inhibiting the expression of HDAC2 or promoting the expression of Inpp5f can effectively improve cognitive function in rats (P<0.05). After intervention, compared with the sham operation group, rats in the other 3 groups had higher HDAC2 mRNA level and lower Inpp5f mRNA level (P<0.05). In conclusion, neuropathic pain can cause an increase in HDAC2 expression level and a decrease in Inpp5f expression level, and activate the PI3K/Akt/GSK-3β signal pathway. Inhibition of HDAC2 expression can inhibit the activation of PI3K/Akt/GSK-3β signal pathway through increasing Inpp5f expression, thus improving the condition and cognitive disorder of rats with neuropathic pain.

Reciprocal Relationship Between HDAC2 and P-Glycoprotein/MRP-1 and Their Role in Steroid Resistance in Childhood Nephrotic Syndrome.

Background: Reduced HDACs levels have been reported in steroid resistant chronic obstructive pulmonary disease and bronchial asthma patients. P-glycoprotein (P-gp) over expression in peripheral blood mononuclear cells (PBMCs) has been reported in patients with steroid resistant nephrotic syndrome (NS). Whether and how HDACs and P-gp are linked with each other is not clear, especially in NS patients.Aim: To evaluate mRNA expression of P-gp/MRP-1 and HDAC2 in PBMCs of steroid sensitive (SSNS) and steroid resistant nephrotic syndrome (SRNS) patients, and determine the relationship between expression of HDAC2 and P-gp/ MRP-1in NS patients.Methods: Twenty subjects (10 in each group), SSNS (mean age 7.54 ± 3.5 years), and SRNS (mean age 8.43 ± 3.8 years) were recruited. mRNA expression of HDAC2 and P-gp/MRP-1 was studied by quantitative real time PCR. PBMCs were treated with Theophylline, 1 μM, and Trichostatin A, 0.8 μM, for 48 h for induction and suppression of HDAC2, respectively.Results: At baseline, expression of P-gp (4.79 ± 0.10 vs. 2.13 ± 0.12, p < 0.0001) and MRP-1 (3.99 ± 0.08 vs. 1.99 ±0.11, p < 0.0001) on PBMCs were increased whereas, HDAC2 mRNA levels (2.97 ± 0.15 vs. 6.02 ± 0.13, p < 0.0001) were significantly decreased in SRNS as compared to that of SSNS patients. Compared to baseline, theophylline reduced mRNA expression of P-gp and MRP-1 (fold change 2.65 and 2.21, *p < 0.0001 in SRNS) (fold change 1.25, 1.24, *p < 0.0001 in SSNS), respectively. However, it increased the expression of HDAC2 (fold change 5.67, *p < 0.0001 in SRNS) (fold change 6.93, *p < 0.0001 in SSNS). Compared to baseline, TSA treatment increased mRNA levels of P-gp and MRP-1 (fold change 7.51, 7.31, *p < 0.0001 in SRNS) and (fold change 3.49, 3.35, *p < 0.0001 in SSNS), respectively. It significantly decreased the level of HDAC2 (fold change 1.50, *p < 0.0001 in SRNS) (fold change 2.53, *p < 0.0001 in SSNS) patients.Conclusion: Reduced HDAC2 and increased P-gp/MRP-1 activity may play a role in response to steroids in childhood NS. HDAC2 and P-gp/MRP-1 are in reciprocal relationship with each other.

Memory and plasticity impairment after binge drinking in adolescent rat hippocampus: GluN2A/GluN2B NMDA receptor subunits imbalance through HDAC2.

Ethanol (EtOH) induces cognitive impairment through modulation of synaptic plasticity notably in the hippocampus. The cellular mechanism(s) of these EtOH effects may range from synaptic signaling modulation to alterations of the epigenome. Previously, we reported that two binge-like exposures to EtOH (3g/kg, ip, 9h apart) in adolescent rats abolished long-term synaptic depression (LTD) in hippocampus slices, induced learning deficits, and increased N-methyl-d-aspartate (NMDA) receptor signaling through its GluN2B subunit after 48hours. Here, we tested the hypothesis of EtOH-induced epigenetic alterations leading to modulation of GluN2B and GluN2A NMDA receptor subunits. Forty-two days old rats were treated with EtOH or the histone deacetylase inhibitor (HDACi) sodium butyrate (NaB, 600mg/kg, ip) injected alone or 30minutes before EtOH. After 48hours, learning was tested with novel object recognition while synaptic plasticity and the role of GluN2A and GluN2B subunits in NMDA-fEPSP were measured in CA1 field of hippocampus slices. LTD and memory were impaired 48hours after EtOH and NMDA-fEPSP analysis unraveled changes in the GluN2A/GluN2B balance. These results were associated with an increase in histone deacetylase (HDAC) activity and HDAC2 mRNA and protein while Ac-H4K12 labelling was decreased. EtOH increases expression of HDAC2 and mRNA level for GluN2B subunit (but not GluN2A), while HDAC2 modulates the promoter of the gene encoding GluN2B. Interestingly, NaB pretreatment prevented all the cellular and memory-impairing effects of EtOH. In conclusion, the memory-impairing effects of two binge-like EtOH exposure involve NMDA receptor-dependent LTD deficits due to a GluN2A/GluN2B imbalance resulting from changes in GluN2B expression induced by HDAC2.

Increased Expression of miR-1 in Fast-Twitch Skeletal Muscle in Response to Resistance Exercise

Background: The response of microRNA (miR-1), upstream regulators (Myogenic Differentiation 1 (MyoD1) and Myocyte Enhancer Factor 2C (Mef2C)), and a downstream target (Histone Deacetylase 4 (Hdac4)) to an acute bout of Resistance Exercise (RE) in Extensor Digitorum Longus (EDL) and soleus muscles has remained elusive. Objectives: In this experimental study, we investigated the effect of an acute bout of RE on the expression of miR-1, MyoD1, Mef2C, and Hdac4 genes in the slow and fast-twitch muscle of rats. Methods: The current study was conducted at Tarbiat Modares University, Tehran, Iran, in 2017. Fifteen male Wistar rats were randomly divided into two groups, control (n = 5) and RE (n = 10). The RE protocol consisted of four sets of five repetitions of climbing a ladder with weights attached to the tails of rats. The soleus and EDL muscles of rats were collected at 3 h (n = 5) and 6 h (n = 5) post-RE. The real-time polymerase chain reaction was used to measure the levels of miR-1, MyoD1, Mef2C, and Hdac4 mRNA expression. Results: The miR-1 expression in EDL muscle was significantly lower at 3 h and 6 h post-exercise in the RE group than in the control group (P < 0.01). The miR-1 expression in soleus muscle was significantly lower at 3 h post-exercise in the RE group than in the control group (P < 0.007) but it was significantly higher at 6 h post-exercise (P < 0.036). The expression of MyoD1, Mef2C, and Hdac4 genes in the EDL muscle was higher in the RE group than in the control group (P < 0.01). The expression of these genes in response to RE had more fluctuations in soleus muscle. Conclusions: It can be concluded that miR-1 expression in extensor digitorum longus and soleus muscles of rats responds to resistance exercise in different manners and this coincides with a change in upstream regulators and downstream target.

Clinical significance of HDAC9 in hepatocellular carcinoma

In recent years, most related studies have found that chronic hepatitis B virus infection is the main cause of hepatocellular carcinoma (HCC), but the specific pathogenesis is still unclear. To investigate the function of HDAC in hepatocellular carcinoma (HCC), this study used qRT-PCR to determine the expression levels of miR-376a and HDAC9 mNRA in HCC and para-cancerous tissues. The clinical significance of HDAC9 in HCC was assessed in a study cohort containing 37 patients with HCC using immunohistochemistry. The expression level of miR-376a in liver cancer tissues was significantly lower than that in para-cancerous tissues, while the expression level of HDAC9 mRNA in liver cancer tissue was significantly higher than that in para-cancerous tissues. The expression of HDAC9 occurred mainly in the nucleus. There was a significant correlation between tumor differentiation and HDAC9. Survival analysis showed that HCC patients with higher HDAC9 expression had poorer prognosis, and subsequent multivariate analysis showed that HDAC9 expression level was an independent predictor. There was a definite correlation between HDAC9 and the expressions of AFP and Ki67. These results suggest that the expression level of HDAC9 in HCC is abnormally high while the expression level of miR-376a is significantly decreased, indicating that HDAC9 may be a potential prognostic indicator of HCC.