Evaluación y caracterización de la sirtuina 2.3 de Giardia duodenalis (CdSir2.3)

La giardiasis es la enfermedad gastrointestinal de mayor incidencia mundial, causada por el protozoario Giardia duodenalis, para la cual no se cuenta con una vacuna o tratamiento eficiente. En aras de buscar nuevos blancos farmacológicos contra este parásito, se han estudiado las enzimas del metabolismo energético, como las sirtuinas, deacetilasas dependientes del dinucleótido de adenina y nicotinamida (NAD). Previamente se identificó a GdSir2.1 y GdSir2.2 como deacetilasas dependientes de NAD, con localizaciones subcelulares diferentes. En este trabajo se estudió otro candidato a sirtuina (GdSir2.3) mediante herramientas bioinformáticas para la identificación de características típicas de la familia sirtuina en la secuencia del candidato, y experimentales como la obtención de la proteína recombinante 6xHis-GdSir2.3 que demostró actividad deacetilasa dependiente de NAD y que sirvió como antígeno en la producción de los IgY - α -6xHis-GdSir2.3 para la localización subcelular de la proteína endógena en G. duodenalis. Lo anterior concuerda con otros estudios donde se señala a GdSir2.3 como un importante regulador de la enquistación, debido a su aumento de expresión durante esta etapa del ciclo de vida, constituyéndola como un blanco farmacológico promisorio para el control de esta parasitemia.

Giardiasis is the gastrointestinal disease with the highest incidence worldwide, caused by the protozoan Giardia duodenalis, for which there is no vaccine or efficient treatment. In order to find new pharmacological targets against this parasite, energy metabolism enzymes such as sirtuins, deacetylases dependent on the nicotinamide adenine dinucleotide (NAD), have been studied. GdSir2.1 and GdSir2.2 were previously identified as NAD-dependent deacetylases, with different subcellular locations. In this work, another candidate for sirtuin (GdSir2.3) was studied using bioinformatic tools for the identification of typical characteristics of the sirtuin family in the sequence of the candidate; and experimental ones such as obtaining the recombinant protein 6xHis-GdSir2.3 that demonstrated NAD-dependent deacetylase activity; and that it served as an antigen in the production of IgY - α - 6xHis-GdSir2.3 for the subcellular localization of the endogenous protein in G. duodenalis. The foregoing is consistent with other studies where GdSir2.3 is indicated as an important regulator of encyst due to its increased expression during this stage of the life cycle, constituting it as a promising drug target for the control of this parasitaemia.

A giardíase é a doença gastrointestinal de maior incidência no mundo, causada pelo protozoário Giardia duodenalis, para a qual não existe vacina ou tratamento eficaz. Com o objetivo de encontrar novos alvos farmacológicos contra esse parasita, têm sido estudadas enzimas do metabolismo energético, como as sirtuínas, desacetilases dependentes do dinucleotídeo adenina nicotinamida (NAD). GdSir2.1 e GdSir2.2 foram previamente identificados como desacetilases dependentes de NAD, com diferentes localizações subcelulares. Neste trabalho, outro candidato a sirtuin (GdSir2.3) foi estudado usando ferramentas de bioinformática para a identificação de características típicas da família sirtuin na sequência do candidato; e experimentais, como a obtenção da proteína recombinante 6xHis-GdSir2.3 que demonstrou atividade desacetilase dependente de NAD; e que serviu como antígeno na produção de IgY - α - 6xHis-GdSir2.3 para a localização subcelular da proteína endógena em G. duodenalis. O exposto é consistente com outros estudos em que o GdSir2.3 é apontado como um importante regulador de encisto devido à sua expressão aumentada durante esta fase do ciclo de vida, constituindo-se como um alvo promissor para o controle dessa parasitemia.

Role of histone deacetylase in sodium butyrate-induced reduction of intestinal ischemia-reperfusion injury in mice: relationship with oxidative stress and cell apoptosis / 中华麻醉学杂志

Objective:To evaluate the role of histone deacetylase (HDAC) in sodium butyrate-induced reduction of intestinal ischemia-reperfusion (I/R) injury in mice and the relationship with oxidative stress and cell apoptosis.Methods:Twenty-four SPF healthy male C57BL mice, aged 6-8 weeks, weighing 22-25 g, were divided into 4 groups ( n=6 each) according to the random number table method: sham operation group (S group), intestinal I/R group (IR group), intestinal I/R+ sodium butyrate group (IN group) and intestinal I/R+ ITSA-1+ sodium butyrate group (INI group). In IR, IN and INI groups, the superior mesenteric artery was clamped for 45 min, followed by reperfusion for 2 h to prepare the model of intestinal I/R injury, while the superior mesenteric artery was only isolated without ligation in S group.One week before preparation of the model, sodium butyrate 500 mg/kg was intragastrically administered once a day in IN group and INI group, the HDAC activator ITSA-1 0.5 mg/kg was intraperitoneally injected three times a week in INI group, and the equal volume of normal saline was given instead in the other groups.The mice were sacrificed at 2 h of reperfusion and small intestinal tissues were obtained for microscopic examination of the pathological changes which were assessed using Chiu′s score and for determination of the content of MDA (by enzyme-linked immunosorbent assay) and expression of cleaved caspase-3 (by Western blot). Results:Compared with S group, Chiu′s score was significantly increased, and the expression of cleaved caspase-3 was up-regulated in IR, IN and INI groups, the content of MDA in small intestinal tissues was significantly increased in IR and INI groups ( P<0.05). Compared with IR group, Chiu′s score was significantly decreased in IN and INI groups, and the content of MDA was significantly decreased, and the expression of cleaved caspase-3 was down-regulated in IN group ( P<0.05). Compared with IN group, Chiu′s score and content of MDA were significantly increased, and the expression of cleaved caspase-3 was up-regulated in INI group ( P<0.05). Conclusions:HDAC is involved in sodium butyrate-induced reduction of intestinal I/R injury in mice, which is related to the inhibition of oxidative stress and cell apoptosis.

Role of histone deacetylase 6 in reduction of intestinal ischemia-reperfusion injury by sodium butyrate in mice / 中华麻醉学杂志

Objective:To evaluate the role of histone deacetylase 6 (HDAC6) in reduction of intestinal ischemia-reperfusion (I/R) injury by sodium butyrate in mice.Methods:Twenty-four SPF healthy adult male C57BL/6 mice, aged 8-10 weeks, weighing 22-25 g, were divided into 4 groups ( n=6 each) by the random number table method: sham operation group (S group), intestinal I/R group (I/R group), intestinal I/R + sodium butyrate group (I/R+ SB group), and intestinal I/R + ITSA-1+ sodium butyrate group (I/R+ I+ SB group). The model of intestinal I/R injury was established by clipping superior mesenteric artery for 45 min followed by 120 min of reperfusion in anesthetized animals.In I/R+ I+ SB group, the HDACs activator ITSA-1 0.5 mg/kg was intraperitoneally injected at 6, 3 and 1 days before ischemia.Sodium butyrate 500 mg/kg was given by intragastric administration every day one week before ischemia in I/R+ SB group and I/R+ I+ SB group, and the equal volume of normal saline was given in S group and I/R group.At 120 min of reperfusion, the mice were sacrificed and their small intestine tissues were obtained.The levels of diamine oxidase (DAO) in serum and intestinal tissues were detected by enzyme-linked immunosorbent assay.The pathological changes of small intestinal tissues were observed with a light microscope, and intestinal damage was assessed and scored according to Chiu.The expression of microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ), P62 and HDAC6 was determined by Western blot.The contents of histone H3 (H3) and acetylated histone H3 (Ac-H3) in small intestinal tissues were determined by enzyme-linked immunosorbent assay. Results:Compared with S group, the Chiu′s score, levels of DAO in serum and small intestinal tissues were significantly increased, the expression of LC3 Ⅱ and HDAC6 was up-regulated, P62 expression was down-regulated, H3 content was increased, and AC-H3 content was decreased in I/R group ( P<0.05). Compared with I/R group, the Chiu′s score, levels of DAO in serum and small intestinal tissues were significantly decreased, the expression of LC3 Ⅱ and HDAC6 was down-regulated, P62 expression was up-regulated, H3 content was decreased, and AC-H3 content was increased in I/R+ SB group ( P<0.05). Compared with I/R+ SB group, the Chiu′s score and levels of DAO in serum and small intestinal tissues were significantly increased, the expression of LC3 Ⅱ and HDAC6 was up-regulated, P62 expression was down-regulated, H3 content was increased, and AC-H3 content was decreased in I/R+ I+ SB group ( P<0.05). Conclusions:Sodium butyrate can alleviate intestinal I/R injury by inhibition of HDAC6 activity in mice, and the mechanism may be related to inhibition of autophagy and promotion of H3 acetylation.

Role of histone deacetylase 6 in epithelial-mesenchymal transition of renal tubular epithelial cells and activation of renal interstitial fibroblasts / 中华肾脏病杂志

Objective:To investigate the role and mechanism of (histone deacetylase 6, HDAC6) in the epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells and the activation of renal interstitial fibroblasts.Methods:Human renal tubular epithelial cells (HK-2) and rat renal interstitial fibroblast (NRK-49F) were cultured in vitro, and divided into 4 groups: control group, Tubastatin A (TA) group (treated with 10 μmol/L HDAC6 inhibitor TA for 36 h), transforming growth factor-β1 (TGF-β1) group (10 ng/ml TGF-β1 for 36 h), and TGF-β1+TA group (treated with 10 ng/ml TGF-β1 and 10 μmol/L TA for 36 h). The expression levels of fibronectin, α-smooth muscle actin (α-SMA), collagen I, E-cadherin, HDAC6, acetyl histone H3, histone H3, acetyl α-tubulin, α-tubulin, TGF-β receptor (TGF-βR) 1, p-Smad3, Smad3, connective tissue growth factor (CTGF), epidermal growth factor receptor (EGFR) and p-EGFR in HK-2 and NRK-49F cell samples were detected by Western blotting, and quantitative analysis was performed according to gray level. Results:(1) In HK-2 cells stimulated by TGF-β1, TA decreased the expression of fibronectin, α-SMA, collagen I, and increased the expression of epithelial cell marker E-cadherin. Meanwhile, TA decreased the expression of HDAC6 and increased the expression levels of acetyl histone H3 and acetyl α-tubulin (all P<0.05). (2) Compared with the TGF-β1 group, the expressions of TGF-βR1, p-Smad3, CTGF and p-EGFR in TGF-β1+TA group were decreased (all P<0.05), while the total protein levels of Smad3 and EGFR were not significantly different (both P>0.05). (3) In NRK-49F cells stimulated by TGF-β1, TA decreased the expressions of fibronectin, α-SMA, collagen I, TGF-βR1 and p-Smad3 (all P<0.05). Conclusions:Blockade of HDAC6 by TA may inhibit the EMT of renal tubular epithelial cells and the activation of renal interstitial fibroblasts via regulating multiple signaling pathways including TGF-β/Smad3, CTGF and EGFR.

Role of HDAC3 in high glucose hypoxia/reoxygenation injury to primary rat cardiomyocytes: the relationship with autophagy / 中华麻醉学杂志

Objective:To evaluate the role of histone deacetylase 3 (HDAC3) in high glucose hypoxia/reoxygenation (H/R) injury to primary rat cardiomyocytes and the relationship with autophagy.Methods:The primary cardiomyocytes extracted from newborn Sprague-Dawley rats, aged about 1-3 days, were divided into 5 groups ( n=24 each) according to the random number table method: control group (C group, glucose concentration 5.5 mmol/L), H/R group, high glucose group (H group, glucose concentration 30 mmol/L), high glucose H/R group (HH/R group), and high glucose H/R + HDAC3 inhibitor RGFP966 group (HH/R+ RG group). Fifty percent glucose injection was used to prepare high-glucose medium (final concentration 30 mmol/L). Cells were cultured in a hypoxic environment (5% CO 2-0.9% O 2-94.1% N 2) for 6 h, followed by reoxygenation in a normoxic environment for 2 h to establish the cardiomyocyte H/R model in H/R group.RGFP966 at a final concentration of 10 μmol/L was added at 24 h before H/R in HH/R+ RG group.At 2 h of reoxygenation, the cell viability was measured using CCK-8 kit, the activity of lactic dehydrogenase (LDH) in the cell supernatant was determined using enzyme-linked immunosorbent assay, the level of autophagy was detected with a confocal microscope after cells were transfected with autophagy double-labeled adenovirus (mRFP-GFP-LC3), and the expression of HDAC3, p62, LC3 Ⅱ and LC3 Ⅰ was detected using Western blot.LC3Ⅱ/LC3Ⅰ ratio was calculated. Results:Compared with group C, the cell viability was significantly decreased, and the activity of LDH in supernatant was increased in H/R and H groups, the number of autophagosomes was significantly increased, the expression of HDAC3 in cardiomyocytes was up-regulated, the expression of p62 was down-regulated, and the LC3 Ⅱ/I ratio was increased in group H/R, and the number of autophagosomes was significantly decreased, the expression of HDAC3 and p62 in cardiomyocytes was up-regulated, and the LC3 Ⅱ/I ratio was decreased in group H ( P<0.05). Compared with group H/R, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, the number of autophagosomes was decreased, the expression of HDAC3 and p62 in cardiomyocytes was up-regulated, and the LC3 Ⅱ/I ratio was decreased in group HH/R ( P<0.05). Compared with group H, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, the number of autophagosomes was increased, the expression of HDAC3 and p62 in cardiomyocytes was up-regulated, and the LC3 Ⅱ/I ratio was increased in group HH/R ( P<0.05). Compared with group HH/R, the cell viability was significantly increased, the activity of LDH in supernatant was decreased, the number of autophagosomes was increased, the expression of HDAC3 and p62 in cardiomyocytes was down-regulated, and the LC3 Ⅱ/I ratio was increased in group HH/R+ RG ( P<0.05). Conclusion:Up-regulation of HDAC3 expression is involved in high glucose H/R injury to primary rat cardiomyocytes, which is related to decreasing the level of autophagy.

Role of HDAC6 in spinal dorsal horn in dexmedetomidine-induced reduction of neuropathic pain in rats / 中华麻醉学杂志

Objective:To evaluate the role of histone deacetylase 6 (HDAC6) in spinal dorsal horn in dexmedetomidine-induced reduction of neuropathic pain (NP) in rats.Methods:Forty clean-grade healthy male Sprague-Dawley rats, aged 7-9 weeks, weighing 190-240 g, were divided into 5 groups ( n=8 each) using a random number table method: control group (group C), sham operation group (group SH), group NP, dexmedetomidine group (group D), and specific HDAC6 inhibitor ACY-1215 plus dexmedetomidine group (group AD). The animals were commonly fed without any treatment in group C. The sciatic nerve was only isolated but not ligated in group SH.The animals were anesthetized with intraperitoneal 10% chloral hydrate 350 mg/kg.NP was induced by chronic constrictive injury (CCI). The right sciatic nerve was exposed, and 4 loose ligatures were placed on the sciatic nerve at 1-mm intervals with 4-0 silk thread in NP and D groups.In group D, dexmedetomidine 40 μg/kg was intraperitoneally injected once a day starting from the end of operation until the animals were sacrificed.In group AD, ACY-1215 25 mg/kg was intraperitoneally injected every day immediately before CCI, and dexmedetomidine 40 μg/kg was intraperitoneally injected daily after CCI until 15 days after CCI.The equal volume of solvent was given instead of dexmedetomidine in S and NP groups.The mechanical paw withdrawal threshold to von Frey filament stimulation (MWT) and thermal paw withdrawal latency (TWL) were measured at 1 day before CCI (baseline, T 0) and 3, 6, 9, 12 and 15 days after CCI (T 1-5). The rats were then sacrificed, and the dorsal horn tissues of L 4-6 spinal cord were obtained for determination of the expression of myeloid differentiation factor 88 (MyD88) and nuclear factor kappa B (NF-κB) p65 by Western blot. Results:Compared with group C and group SH, the MWT was significantly decreased, and the TWL was shortened at T 1-5, and the expression of MyD88 and NF-κB p65 in the spinal dorsal horn was up-regulated in NP, D and AD groups ( P<0.05). Compared with group NP, the MWT was significantly increased, and the TWL was prolonged at T 1-5, the expression of MyD88 and NF-κB p65 in the spinal dorsal horn was down-regulated in group D ( P<0.05), and no significant change was found in the parameters mentioned above in group AD ( P>0.05). Compared with group D, MWT was significantly decreased, and TWL was shortened at T 1-5, and the expression of MyD88 and NF-κB p65 was up-regulated in the dorsal horn of the spinal cord in group AD ( P<0.05). Conclusion:HDAC6 in spinal dorsal horn is involved in dexmedetomidine-induced reduction of NP in rats, which is related to inhibiting MyD88/NF-κB signaling pathway.

Effect of SIRT3 overexpression on hypoxia-reoxygenation injury to hippocampal neurons of mice exposed to high glucose: relationship with SOD2 / 中华麻醉学杂志

Objective:To evaluate the effect of sirtuin 3 (SIRT3) overexpression on hypoxia-reoxygenation (H/R) injury to hippocampal neurons of mice exposed to high glucose and its relationship with SOD2.Methods:The normally cultured HT22 neurons at the logarithmic phase were selected and divided into 3 groups ( n=12 each) using a random number table method: high-glucose normoxia group (HG group), high glucose+ H/R group (HHR group) and high glucose+ H/R+ SIRT3 overexpression group (HHR+ SIRT3 group). To establish high glucose model, the neurons in 3 groups were cultured in high-glucose culture medium (glucose concentration of 50 mmol/L) for 8 h. In HHR and HHR+ SIRT3 groups, the cells were exposed to glucose-free and hypoxia for 6 h and then cultured in the high-glucose normoxic environment for 24 h to establish the high glucose and HR injury model.In HHR+ SIRT3 group, the neurons were transfected with SIRT3 overexpressed lentivirus.The cell viability was recorded by the cell counting kit-8 assay, reactive oxygen species (ROS) content was detected by flow cytometry, mitochondrial malonaldehyde (MDA) content, superoxide dismutase (SOD) activity, catalase (CAT) activity and adenosine triphosphate (ATP) content were determined by colorimetry, mitochondrial membrane potential (MMP) was detected by JC-1 probe, and the expression of nuclear respiratory factor 1 (NRF1), mitochondrial transcription factor A (TFAM), SIRT3, SOD2 and acetylated SOD2 (ac-SOD2) was detected by Western blot. Results:Compared with HG group, cell viability, SOD activity, CAT activity, ATP content, MMP, NRF1 and the expression of TFAM and SIRT3 were significantly decreased, and ROS content, MDA content and ac-SOD2/SOD2 ratio were increased in group HHR and group HHR+ SIRT3 ( P<0.05). Compared with HHR group, cell viability, SOD activity, CAT activity, ATP content, MMP, NRF1 and the expression of TFAM and SIRT3 were significantly increased, and ROS content, MDA content and ac-SOD2 /SOD2 ratio were decreased in HHR+ SIRT3 group ( P<0.05). Conclusion:SIRT3 overexpression can alleviate hypoxia-reoxygenation (H/R) injury to hippocampal neurons of mice incubated in high glucose medium, and the mechanism is related to activation of SOD2 deacetylation.

Molecular mechanisms of RPD3 family members in regulating plant development and environmental responses / 生物工程学报

Lysine acetylation is one of the major post-translational modifications and plays critical roles in regulating gene expression and protein function. Histone deacetylases (HDACs) are responsible for the removal of acetyl groups from the lysines of both histone and non-histone proteins. The RPD3 family is the most widely studied HDACs. This article summarizes the regulatory mechanisms of Arabidopsis RPD3 family in several growth and development processes, which provide a reference for studying the mechanisms of RPD3 family members in regulating plant development. Moreover, this review may provide ideas and clues for exploring the functions of other members of HDACs family.

Nicotinamide activates latent HIV-1 ex vivo in ART suppressed individuals, revealing higher potency than the association of two methyltransferase inhibitors, chaetocin and BIX01294

ABSTRACT Background: Latent HIV-1 is a major hurdle in obtaining HIV-1 sustained virological remission (SVR). Here we explored histone deacetylation inhibition property of nicotinamide (NAM; n = 17) for the first time in comparison to a combination of methyltransferase inhibitors (MTIs; Chaetocin and BIX01294; n = 25) to reactivate latent HIV ex vivo in CD8-depleted PBMCs from antiretroviral treated aviremic individuals. Results: NAM reactivated HIV-1 from 13/17 (76.4%) samples compared to 20/25 (80.0%) using MTIs with mean viral load (VLs) of 4.32 and 3.22 log10 RNA copies/mL, respectively (p = 0.004). Mean purging time after NAM and MTIs stimulation was 5.1 and 6.75 days, respectively (p = 0.73). Viral purging in autologous cultures exhibited blunted HIV recovery with fluctuating VLs followed by a complete viral extinction when expanded in allogenic system. Electron microscopy from five supernatants revealed anomalous viral particles, with lack of complete viral genomes when characterized by ultradeep sequencing through metagenomics approach (n = 4). Conclusion: NAM alone was more potent HIV-1 activator than combination of MTIs, with potential of clinical use.

Selectively enhancing radiosensitivity of cancer cells

The radiotherapy modulators used in clinic have disadvantages of high toxicity and low selectivity. For the first time, we used the

Molecular cloning and subcellular localization of six HDACs and their roles in response to salt and drought stress in kenaf (Hibiscus cannabinus L)

BACKGROUND: Histone acetylation is an important epigenetic modification that regulates gene activity in response to stress. Histone acetylation levels are reversibly regulated by histone acetyltransferases (HATs) and histone deacetylases (HDACs). The imperative roles of HDACs in gene transcription, transcriptional regulation, growth and responses to stressful environment have been widely investigated in Arabidopsis. However, data regarding HDACs in kenaf crop has not been disclosed yet. RESULTS: In this study, six HDACs genes (HcHDA2, HcHDA6, HcHDA8, HcHDA9, HcHDA19, and HcSRT2) were isolated and characterized. Phylogenetic tree revealed that these HcHDACs shared high degree of sequence homology with those of Gossypium arboreum. Subcellular localization analysis showed that GFP-tagged HcHDA2 and HcHDA8 were predominantly localized in the nucleus, HcHDA6 and HcHDA19 in nucleus and cytosol. The HcHDA9 was found in both nucleus and plasma membranes. Real-time quantitative PCR showed that the six HcHDACs genes were expressed with distinct expression patterns across plant tissues. Furthermore, we determined differential accumulation of HcHDACs transcripts under salt and drought treatments, indicating that these enzymes may participate in the biological process under stress in kenaf. Finally, we showed that the levels of histone H3 and H4 acetylation were modulated by salt and drought stress in kenaf. CONCLUSIONS: We have isolated and characterized six HDACs genes from kenaf. These data showed that HDACs are imperative players for growth and development as well abiotic stress responses in kenaf.

Gestation and breastfeeding in schistosomotic mice differentially alters the expression of histone deacetylases (HDACs) in adult offspring

BACKGROUND Breastfeeding or gestation in schistosomotic mothers can cause long-term alterations in the immune response of offspring. OBJECTIVES Evaluate the expression of histone deacetylases (HDACs) (all classes), the production of cytokines by T and B lymphocytes and macrophages, and the frequency of CD4+CD25+FoxP3+-cells in adult offspring born and/or suckled by schistosomotic mothers. METHODS We harvested splenocytes from offspring born to (BIM), suckled by (SIM), or born to/suckled by (BSIM) schistosomotic mothers and animals from noninfected mothers (Control) at seven-weeks old and cultured them with/without Concanavalin A. HDAC expression was evaluated by real-time quantitative polymerase chain reaction (qPCR), and cytokines and membrane markers were evaluated by fluorescence-activated cell sorting (FACS). FINDINGS Compared to Control, BIM mice showed increased expression of HDAC9 and frequency of CD4+IL-10+-cells. The SIM group had increased expression of HDAC1, HDAC2, HDAC6, HDAC7, HDAC10, Sirt2, Sirt5, Sirt6, and Sirt7. The BSIM group only had increased HDAC10 expression. The SIM and BSIM groups exhibited decreased frequencies of CD4+IL-4+-cells and CD4+CD25+FoxP3+-cells, along with a higher frequency of CD14+IL-10+-cells and an increase in CD45R/B220+IL-10+-cells. The BSIM group also showed a high frequency of CD4+IL10+-cells. MAIN CONCLUSIONS Breastfeeding induced the expression of HDACs from various classes involved in reducing inflammatory responses. However, gestation enhanced the expression of a single HDAC and breastfeeding or gestation appears to favour multiple IL-10-dependent pathways, but not cells with a regulatory phenotype.

Role of HDAC3 in renal injury induced by myocardial ischemia-reperfusion in diabetic rats / 中华麻醉学杂志

Objective@#To evaluate the role of histone deacetylase 3 (HDAC3) in renal injury induced by myocardial ischemia-reperfusion (I/R) in diabetic rats.@*Methods@#SPF healthy adult male Sprague-Dawley rats, weighing 210-220 g, were used in this study.Diabetes mellitus was induced by intraperitoneal injection of 1% streptozotocin 60 mg/kg.Eighteen diabetic rats were divided into 3 groups (n=6 each) using a random number table method: sham operation group (group S), group I/R and HDAC3 inhibitor RGFP966 group (RG). The myocardial I/R model was established by ligation of the left anterior descending coronary artery for 30 min followed by 120-min reperfusion.RGFP966 10 mg/kg was intraperitoneally injected at 1 h before myocardial ischemia in group RG.The right internal carotid artery was isolated at 120 min of reperfusion for measurement of serum lactate dehydrogenase (LDH), creatine kinase-MB (CK-MB), creatinine (Cr) and urea nitrogen concentrations.Renal tissues were obtained for examination of the pathological changes and for determination of the expression of HDAC3, silent information regulator 1 (SIRT1) and interleukin-1β (IL-1β). The damage to the renal tubules was scored.@*Results@#Compared with S group, the serum LDH, CK-MB, Cr and urea nitrogen concentrations and renal tubular damage score were significantly increased, the expression of HDAC3 and IL-1β was up-regulated, and the expression of SIRT1 was down-regulated in group I/R (P<0.05). Compared with group I/R, the serum LDH, CK-MB, Cr and urea nitrogen concentrations and renal tubular damage score were significantly decreased, the expression of HDAC3 and IL-1β was down-regulated, and the expression of SIRT1 was up-regulated in group RG (P<0.05).@*Conclusion@#Up-regulated expression of HDAC3 is involved in renal injury induced by myocardial I/R in diabetic rats.

Effect of apicidin on glioblastoma U87 cells and its regulation of OCT-4 gene expression / 肿瘤研究与临床

Objective@#To investigate the effect of histone deacetylase inhibitor apicidin on the glioblastoma U87 cells and its regulation of OCT-4 gene expression.@*Methods@#Glioblastoma U87 cells were treated with different concentrations of apicidin, and dimethyl sulfoxide instead of apicidin was negative control. Methyl thiazolyl tetrazolium (MTT) assay was used to detect the proliferative ability of U87 cells treated by apicidin. The cell apoptosis was observed under the fluorescence microscope, and the cell cycle was detected by using flow cytometry. Reverse transcription-polymerase chain reaction and Western blot was used to detect the expression of mRNA and protein of U87 cells, respectively relative to the expression of GAPDH.@*Results@#MTT assay results showed that apicidin inhibited U87 cells proliferation in a dose-dependent and time-dependent manner, and half of the inhibitory concentration of cell proliferation at 48 h was (1.74±0.13) μmol/L. The cell proportion of U87 cells in S-phase of the negative control, 0.2, 0.5, and 1.0 μmol/L apicidin was (32.68±0.49)%, (33.73±0.76)%, (42.92±0.56)%, and (56.95±0.53)%, respectively after 48 h apicidin administration (P < 0.05), while the proportion of G1 and G2 phase cells was decreased. The karyopyknosis and other apoptotic changes were detected in U87 cells after 48 h treatment of 1.0 μmol/L apicidin under the confocal fluorescence microscope. Western blot and RT-PCR showed that the mRNA and protein relative levels of U87 cells OCT-4 were reduced after 1.0 μmol/L apicidin treatment for 48 h compared with the negative control group (mRNA: 72.44±0.00 vs. 56.66±0.23; protein: 86.59±0.19 vs. 56.04±0.15, both P < 0.01).@*Conclusions@#Apicidin can inhibit the growth of glioblastoma U87 cells, induce cell cycle arrest and apoptosis. Its mechanism may be related to the expression of OCT-4 inhibited by apicidin.

Role of HDAC3 in renal injury induced by myocardial ischemia-reperfusion in diabetic rats / 中华麻醉学杂志

Objective To evaluate the role of histone deacetylase 3 (HDAC3) in renal injury induced by myocardial ischemia-reperfusion (I/R) in diabetic rats.Methods SPF healthy adult male Sprague-Dawley rats,weighing 210-220 g,were used in this study.Diabetes mellitus was induced by intraperitoneal injection of 1％ streptozotocin 60 mg/kg.Eighteen diabetic rats were divided into 3 groups (n =6each) using a random number table method:sham operation group (group S),group I/R and HDAC3 inhibitor RGFP966 group (RG).The myocardial I/R model was established by ligation of the left anterior descending coronary artery for 30 min followed by 120-min reperfusion.RGFP966 10 mg/kg was intraperitoneally injected at 1 h before myocardial ischemia in group RG.The right internal carotid artery was isolated at 120 min of reperfusion for measurement of serum lactate dehydrogenase (LDH),creatine kinase-MB (CKMB),creatinine (Cr) and urea nitrogen concentrations.Renal tissues were obtained for examination of the pathological changes and for determination of the expression of HDAC3,silent information regulator 1 (SIRT1) and interleukin-1β (IL-1β).The damage to the renal tubules was scored.Results Compared with S group,the serum LDH,CK-MB,Cr and urea nitrogen concentrations and renal tubular damage score were significantly increased,the expression of HDAC3 and IL-1β was up-regulated,and the expression of SIRT1 was down-regulated in group I/R (P＜0.05).Compared with group I/R,the serum LDH,CK-MB,Cr and urea nitrogen concentrations and renal tubular damage score were significantly decreased,the expression of HDAC3 and IL-1β was down-regulated,and the expression of SIRT1 was up-regulated in group RG (P＜0.05).Conclusion Up-regulated expression of HDAC3 is involved in renal injury induced by myocardial I/R in diabetic rats.

Effect and mechanism of suberoylanilide hydroxamic acid on fear extinction in mice with chronic so-cial defeat stress / 中华行为医学与脑科学杂志

Objective To investigate the effect and mechanism of suberoylanilide hydroxamic acid (SAHA) on the fear extinction in mice with chronic social defeat stress (SD). Methods Fifty-six male C57BL/6J mice aged 7-8 weeks were randomly divided into control group,social defeat group,control-SAHA group and social defeat-SAHA group to investigate the effect of SAHA and social defeat group,social defeat-AAV BDNF group and social defeat-AAV blank group to investigate the effect of BDNF. Fear extinction in mice was evaluated by fear conditioning test (FC). The levels of BDNF and HDAC2 in mice hippocampus were detected by Western blot (WB). The expression of BDNF-overexpressing virus in hippocampus of mice was detected by immunofluorescence assay. Results (1) Compared with control group,fear extinction in the social defeat group was significantly decreased (P<0. 05). Compared with control group, the level of HDAC2(0. 50±0. 02) in the social defeat group was significantly increased (P<0. 001),while the level of BDNF(0. 16 ± 0. 03) was significantly decreased (P<0. 001) in the social defeat group. ( 2) After using SAHA,fear extinction of mice significantly improved (P<0. 05). Compared with control group,the level of HDAC2 (0. 26±0. 02) in the control-SAHA group was significantly decreased(P<0. 001),and the level of BDNF (0. 40±0. 03) was significantly increased (P<0. 001). Compared with social defeat group,the level of HDAC2 (0. 39±0. 03) in the social defeat-SAHA group was significantly decreased (P<0. 001),and the lev-el of BDNF (0. 28±0. 01) was significantly increased (P<0. 001). (3)After injection BDNF-overexpressing virus,fear extinction was significantly improved(P<0. 05). Conclusion SAHA can enhance fear extinction in mice with chronic social defeat stress and its mechanism may be related to the up-regulation of BDNF ex-pression in hippocampus by inhibiting HDAC2 in hippocampal.

Effect of histone deacetylase and its inhibitor on tumor / 国际肿瘤学杂志

The occurrence and progress of tumor is the result of the interaction of heredity and epigenetics. Histone deacetylation modification,as an important epigenetic modification,plays an important role in tumorigenesis and development. The abnormal expression of histone deacetylase in normal tissues and cells promotes the development of tumor and is related to the proliferation and apoptosis,angiogenesis,metastasis and drug resistance of tumor cells,and becomes a new target of tumor therapy. Histone deacetylase inhibitors as anti-tumor drugs have a good prospect of application.

Role of silent information regulator 1 in hepatic ischemia-reperfusion injury / 临床肝胆病杂志

Hepatic ischemia-reperfusion injury has always been a difficult problem in liver transplantation and liver tumor resection. The involvement of multiple mechanisms makes it particularly complex, among which energy expenditure during ischemia and oxidative stress induced by reperfusion are the main mechanisms leading to hepatic ischemia-reperfusion injury and may cause cell death and even liver failure. Silent information regulators are a class of nicotinamide adenine dinucleotide-dependent deacetylases which can lead to deacetylation of transcription factors including histones and nonhistones, and they are playing an important regulatory role in cell apoptosis, inflammatory response, energy balance, and oxidative stress. There are 7 types of silent information regulators in mammals, i.e., Sirt1-Sirt7, among which Sirt1 can reduce hepatocyte stress, regulate cell metabolic pathways, and thus alleviate the degree of hepatic ischemia-reperfusion injury through various signaling pathways. This article reviews the role of Sirt1-related signaling pathways in hepatic ischemia-reperfusion injury.

Effect of amitriptyline on phosphorylation of HDAC5 in basolateral amygdale of rats with neuropathic pain / 中华麻醉学杂志

Objective To evaluate the effect of amitriptyline on the phosphorylation of histone deacetylase 5 (HDAC5) in the basolateral amygdala (BLA) of rats with neuropathic pain.Methods Thirty healthy male Wistar rats,weighing 250-300 g,were divided into 3 groups (n=10 each) using a random number table method:sham operation group (S group),neuropathic pain group (NP group) and amitriptyline group (A group).Spared nerve injury was produced by exposing the sciatic nerve and its branches and ligation and transection of tibial nerve and common fibular nerve in anesthetized rats.Amitriptyline 10 mg/kg was intraperitoneally injected every day on 14-35 days after establishing the model in group A,while the equal volume of normal saline was given instead of amitriptyline in S and NP groups.The mechanical paw withdrawal threshold (MWT) was measured on 3,7,14,21,28 and 35 days after establishing the model in each group.The forced swimming test was performed on day 36 after establishing the model,and immobility time,climbing time and swimming time were recorded.The rats were then sacrificed,and brain tissues in BLA were obtained for determination of the expression of HDAC5 and phosphorylated HDAC5 (p-HDAC5) (by Western blot) and expressionof HDAC5 mRNA (by real-time quantitative polymerase chain reaction).Results Compared with group S,the MWT was significantly decreased at each time point,the immobility time was prolonged,and the swimming time and climbing time were shortened in group NP,and the MWT was significantly decreased on days 14,21 and 28 after establishing the model,the expression of p-HDAC5 was down-regulated,and the expression of HDAC5 mRNA was up-regulated in group A (P＜0.05).Compared with group NP,the MWT was significantly increased on days 21,28 and 35 after establishing the model,the immobility time was shortened,the climbing time was prolonged,the expression of p-HDAC5 was up-regulated,and the expression of HDAC5 mRNA was down-regulated in group A (P＜0.05or 0.01).Conclusion The mechanism by which amitriptyline improves depression is associated with promoting the phosphorylation of HDAC5 in BLA of rats with NP.

Advances in epigenetic mechanism and drug research of reversing resistance to endocrine therapy in breast cancer / 肿瘤

Breast cancer is now the most common malignant tumor in Chinese women. Approximately 70% of breast cancers express estrogen receptor (ER) which plays a critical role in the development of breast cancer. Endocrine therapy has become one of the most effective treatments for ER-positive breast cancer. At present, the endocrine therapy drugs are mainly divided into three categories: selective ER modulators, selective ER down-regulators and aromatase inhibitors. The adjuvant endocrine therapy with these drugs can reduce the risk of breast cancer recurrence by nearly 40%. However, the resistance of tumor cells to endocrine therapy is a major factor limiting the success of breast cancer treatment. The known mechanisms of endocrine therapy resistance include the concentration of estrogen increased in tumor microenvironment, ER gene mutation, up-regulation of multiple molecular signal pathway, and epigenetic mechanisms. Overcoming endocrine therapy resistance is essential for further improving the benefit rate of endocrine therapy. In this review, the progresses of epigenetic mechanisms (including DNA methylation and histone modification) and epigenetic drugs (including DNAmethyltransferases inhibitors and histone deacetylases inhibitors) of endocrine therapy resistance in ER-positive breast cancer are introduced, in order to further understand the mechanism and therapeutics of endocrine therapy resistance in breast cancer.