Histone Deacetylase Research Articles

Epigenetic modifications in transdifferentiation of adipose tissue-derived stromal cells into spontaneously beating cardiomyocyte-like cells

Abstract Background Adipose tissue-derived stromal vascular fraction (adipose SVF) contains pluripotent mesenchymal stem cells and rarely transdifferentiate into beating cardiomyocytes. We developed a simple culture protocol under which the adult murine inguinal adipose SVF reproductively transdifferentiates into beating cardiomyocyte-like cells (beating CM) without inductions in the primary culture. We also reported that Mef2c is a crucial factor in this conversion. However, the characteristics of beating CM and the factors that regulate the differentiation of adipose SVF toward the cardiac lineage are unknown. Purpose To investigate sequential changes in global gene expression profiles of adipose SVF in primary culture and determine the mechanism of differentiation into beating CM. Methods Adipose SVF cells were isolated from adult mice's inguinal subcutaneous fat pad and cultured using the previously established beating CM induction method (Sci Rep. 2021). At 6 time points (days 0, 3, 7, 14, 21, and 28) in primary culture, total RNA was extracted, and RNA-sequencing was performed (n = 3). The data was analyzed using Subio Platform and MetaCore software. To assess the candidate gene and proteins, adipose SVF cells were transduced with the gene using a lentivirus vector and supplemented with the specific inhibitors. The cardiac differentiation was evaluated by quantitative PCR on day 28. Results The beating CM was confirmed on days 14, 21, and 28 in the primary culture. Among prefiltered 14,574 genes, the expression level of 749 genes were significantly changed (153 genes upregulated vs. 596 genes downregulated) between days 7 and 14 (2-fold, P < 0.05), in which beating CM appeared. The GO molecular functions analysis showed that O-GluNAC transferase and histone deacetylase (HDAC)-associated genes were expressed dominantly before day 7, and KLF4-associated genes were expressed prominently after day 14 in primary culture. The expression of cardiac troponin T (Tnnt2) and myosin light chain ventricular type (Myl2) increased on day 3 and day 14, respectively. The HDAC7, an epigenetic regulator, decreased the expression on day 14 or later. Treatment with the HDAC inhibitor (150 nmol/L) increased the expression of Tnnt2 (5-fold vs. control) on day 28 in adipose SVF. Furthermore, treatment of Mef2c-transduced adipose SVFs with HDAC inhibitors increased expression of cardiac troponin T (29-fold vs. control, P < 0.05). Conclusion The time course analysis demonstrated that a remarkable change occurred in the regulation of transcription on day 14 in adipose SVF primary culture. Alterations in epigenetic modifications increased beating CM in adipose SVFs. Adipose SVF could be applied to new cardiac regeneration therapies by increasing the efficiency of introduction into the beating CM.

Anti-fibrotic effects of the sirt6-activator mdl-800 in cardiac stromal cells

Abstract Background Epigenetic changes are among the molecular substrates of cellular stress responses and tissue remodeling in heart failure progression. The sirtuin family of NAD+-dependent histone and non-histone protein deacetylases exert a protective anti-fibrotic role by negatively modulating the inflammatory response and by positive regulation of autophagy-related genes. Sirtuin6 (SIRT6) stands out as a key cardiac protector that can mitigate aging-induced cardiomyocyte senescence and cardiac hypertrophy. Cardiac stromal cells (CSCs) play a pivotal role in fibrosis and remodeling, regulating the composition and stiffness of the extracellular matrix (ECM). Autophagy enhancement has anti-fibrotic effects in CSCs, but the specific role of SIRT6 modulation in CSCs remains unexplored. Purpose To investigate the biological effects of MDL-800, a synthetic allosteric SIRT6 activator, on the stress response and fibrotic activation of CSCs. Methods CSCs were isolated from 4-week-old C57Bl6J mice by explant culture and stimulated with 10ng/ml of TGF-beta1 (TGFb1) up to 72h to induce fibrotic activation. CSCs were treated with MDL-800 up to 10µM and up to 72h, analyzed for cell viability, gene and protein expression levels. Results TGFb1 treatment led to a 0.64-fold reduction in SIRT6 gene expression after 24h. The MTS assay was used to assess a viability dose-response to MDL-800 up to 72h of treatment. The non-toxic concentrations of 2.5µM and 5µM were selected for further experiments (2.5µM vs CTR 0.97-fold and 5µM vs CTR 0.92-fold, normalized OD vs t0). Co-treatment of 5µM MDL-800 with TGFb1 for 72h resulted in a dose-dependent decrease in the expression of TGFb1-induced fibroblast activation markers, as confirmed by immunofluorescence staining for aSMA (TGFb1 vs CTR 2.21-fold, p<0.05; 5µM vs TGFb1 0.46-fold, p<0.05, N=4; mean fluorescence intensity). The anti-fibrotic effect mediated by MDL-800 was further validated by real-time PCR indicating a reduction in the mRNA expression of multiple fibrotic markers, including ACTA2 (TGFb1 vs CTR 1.67-fold, 5µM vs TGFB 0.29-fold; N=2), COL1a1 (TGFb1 vs CTR 1.32-fold, 5µM vs TGFb1 0.87-fold; N=2) COL3a1 (TGFb1 vs CTR 1.06-fold, 5µM vs TGFb1 0.43-fold; N=2) and POSTN (TGFb1 vs CTR 6.37-fold, 5µM vs TGFb1 0.23-fold; N=2). Treatment with 5µM MDL-800 demonstrated an increase in SIRT6 gene expression, suggesting a transcriptional modulation (TGFb1 vs CTR 0.95-fold, 5µM vs TGFb1 1.68-fold; N=2). Western Blot analyses showed an increase in protein levels of the autophagosome marker LC3-II following treatment with 5µM MDL-800 (TGFb1 vs CTR 0.95-fold; 5µM vs TGFb1 3.01-fold), suggesting that autophagy enhancement may be a potential mechanism for the observed anti-fibrotic effects. Conclusion MDL-800 treatment in CSCs is associated with a dose-dependent reduction of TGFb1-induced fibrotic markers and to increased levels of the autophagy marker, suggesting its potential use as an anti-fibrotic molecule for the heart.

Epigenetic modulation of sirt1 and sirt6 in cardiovascular diseases: unraveling autophagy regulation and endothelial function

Abstract Background Heart failure (HF) and cardiovascular diseases (CVD) still represent major causes of death. Dysmetabolic conditions led by epigenetic alterations play a major role as CVD triggers, expecially those related to the modulation and control of autophagy during the remodelling in HF. Accordingly, several studies have strengthened the role of epigenetics in regulating both cardiac/endothelial phenotype and function, demonstrating how sirtuins, a well-acknowledged family of histone deacetylase, can conjunct the control of cardiac fibrosis, angiogenesis and induction of autophagy. However, this link remains to be fully clarified and explored. Thus, novel drugs targeting autophagy trough epigenetic mechanisms are considered appealing as targets from a pharmacological standpoint in CVD. Purpose This project aims to develop and test two epigenetic modulators and allosteric activators targeting SIRT1 and -6 to further elucidate their role in autophagy regulation and in cardiac/endothelial biology. Methods Chemical synthesis of SIRT1 and SIRT6 activator isoform (SRT2104 and MDL800). Evaluation of epigenetic modulation (H3 histone acetylation H3K9) of endothelial cells (HUVEC). HUVEC survival, angiogenic and autophagy activity were assessed undergo hyperglycemic or hypoxic stress, co-treatment with the selected compounds. Results The specific activator of SIRT-1 (SRT2104) and SIRT-6 (MDL800) show epigenetic activity as they induce the deacetylation of histone H3 on lysine 9. Endothelial cell viability is not influenced by the treatment with both modulators in hyperglycemia and upon hypoxic condition. In HUVEC cultures underwent to hyperglycemia-based stress, both molecules can activate autophagy with an increase in LC3 II protein (1,4-fold HG SRT2104 and 2,8-fold HG MDL800), compared to control.The treatment with SRT2104 enhances HUVEC tube formation, suggesting the increased functional angiogenic ability even in presence of high levels of glucose. Interestingly, the treatment with SRT2104 impacts the transcriptional levels of other members of the sirtuins family, by increasing SIRT-2, 3 and 5. In conclusion, the epigenetic activators SRT2104 and MDL800 have a beneficial effect on endothelial cells by enhancing the autophagic process and stimulate neoangiogenesis.The implementation of these results will attempt to understand to what extent epigenetic drugs such as sirtuin activators, could boost the more pleiotropic and less targeted effects of the current cardiovascular drugs, and to evaluate their functional and phenotype-driven drug refinement.

Insufficient Effective Time of Suberanilohydroxamic Acid, a Deacetylase Inhibitor, Treatment Promotes PC3 Cell Growth.

Castration-resistant prostate cancer (CRPC) contributes mostly to prostate cancer-specific mortality, and conventional castration therapy is almost ineffective, new therapies are needed. As a new potential anti-cancer drug, histone deacetylases (HDACs) inhibitors were demonstrated to be effective in inhibiting drug-resistance cancers in preclinical studies, but the results from clinical trials on CRPC patients were disappointing, and the reasons are unknown. In this study, we investigated the effect of suberanilohydroxamic acid (SAHA), a broad-spectrum pan-HDAC inhibitor, on proliferation, apoptosis, cell cycle progression in PC3 cells, and found that, unlike significant inhibiting effects at high-dose, low-dose SAHA significantly promoted PC3 cell growth. Further colony formation assay showed that the inhibitory effect of SAHA is also dependent on the treatment time, high-dose SAHA also exhibited promoting effect on PC3 cells when the treatment time was insufficient. However, this effect was not observed in another CRPC cell line, DU145, or another HDAC inhibitor, Trichostatin A (TSA). Our results indicate that, instead of inhibitory effect, SAHA would promote PC3 cell growth if the dose is low or the treatment time is insufficient, but this effect has not been observed in other CRPC cell line or HDAC inhibitors.

Discovery of New Thiourea Derivatives as Potential SIRT Inhibitors

Objective: This study aimed to design and synthesize novel thiourea-based compounds targeting Sirtuins (SIRTs), a class of histone deacetylase enzymes implicated in various cellular processes, including cancer, and to evaluate their cytotoxic potential against colon cancer cells. Methods: Virtual modeling and molecular docking studies were conducted to design the compounds and assess their interaction with SIRT enzymes. Compounds showing acceptable docking scores were selected for organic synthesis. The final compounds (MA-1 to MA-6) were synthesized and characterized using FTIR, ¹H-NMR, and ¹³C-NMR spectroscopy. Cytotoxicity studies were performed on SW480 colon cancer cells to evaluate the biological activity of the synthesized compounds. Results: All synthesized compounds exhibited promising cytotoxicity against SW480 colon cancer cells, with MA-2 demonstrating a particularly strong effect. The half-maximal inhibition concentration (IC50) of the compounds was in the single-digit micromolar range, comparable to established SIRT inhibitors Selisistat and AGK2. Conclusion: The thiourea-based compounds synthesized in this study, especially MA-2, showed significant potential as SIRT inhibitors with cytotoxic activity against colon cancer cells. These results warrant further investigation into their therapeutic potential.

METTL14 Promotes Ischemic Stroke-induced Brain Injury by Stabilizing HDAC3 Expression in an m6A-IGF2BP3 Mechanism.

N6-methyladenosine (m6A) modification is the most abundant post-transcriptional modification of mRNAs and has been identified to play critical roles in ischemic stroke (IS). Herein, this study aimed to investigate the function and mechanism of Methyltransferase-like 14 (METTL14) methylase in cerebral IS. Murine BV-2 microglial cell OGD/R models and rat middle cerebral artery occlusion (MCAO) models were established to mimic IS-induced neuronal damage in vitro and brain injury in vivo. Levels of METTL14, Histone Deacetylase 3 (HDAC3) and cGAS-STING axis-related proteins were detected using qRT-PCR or western blotting. Cell proliferation and inflammation were assessed by CCK-8 assay, EdU assay and ELISA. Flow cytometry detected microglia polarization. Cell pyroptosis was analyzed by detecting related-protein markers by western blotting. The m6A modification was determined by methylated RNA immunoprecipitation assay. Brain injury was analyzed by evaluating infarct volume and neurologic score. METTL14 levels were higher in OGD/R-induced microglial cells, primary microglia and infarct brain tissues of rat MCAO models. Functionally, METTL14 silencing reversed OGD/R-induced proliferation inhibition, inflammation and pyroptosis in microglial cells and primary microglia in vitro, and ameliorated cerebral ischemic injury in rat MCAO models. Mechanistically, METTL14 induced HDAC3 m6A modification in an IGF2BP3-dependent manner, and could activate cGAS-STING pathway through HDAC3. Moreover, HDAC3 overexpression reversed the neuroprotective effects of METTL14 silencing. METTL14 silencing reversed ischemic stroke-induced brain injury by inducing HDAC3 m6A modification in an IGF2BP3-dependent mechanism, recommending a novel insight for ameliorating cerebral ischemic stroke.

Interplay Between the Circadian Clock and Sirtuins.

The circadian clock is an autonomous timekeeping system evolved by organisms to adapt to external changes, regulating a variety of important physiological and behavioral processes. Recent studies have shown that the sirtuin family of histone deacetylases is involved in regulating the expression of clock genes and plays an important role in maintaining the normal rhythm of clock gene expression and behavior. Moreover, sirtuins are regulated directly or indirectly by the circadian clock system. The mutual regulation between the circadian clock and sirtuins is likely involved in a variety of signal transduction and metabolism processes. In this review, we discuss the molecular mechanisms and research progress on the intertwined relationship between the circadian clock and sirtuins, mainly in mammals, highlighting sirtuins as molecular links between metabolic control and circadian rhythms and offering our perspectives on future developments in the field.

Magnolol Inhibits High Fructose-Induced Podocyte Inflammation via Downregulation of TKFC/Sp1/HDAC4/Notch1 Activation

Background/Objectives: High fructose has been implicated as an important trigger of kidney inflammation in patients and experimental models. Magnolol, isolated from Magnolia officinalis, has an anti-inflammatory effect, but its protective role in podocytes remains underexplored. This study explored the protective effects and underlying mechanism of magnolol against high fructose-induced podocyte inflammation. Methods: The effects of magnolol on high fructose-induced podocyte inflammation were assessed in male Sprague Dawley rats administered 10% (w/v) fructose water for 12 weeks and heat-sensitive human podocyte cell lines (HPCs) exposed to 5 mM fructose. Podocyte foot processes were examined using transmission electron microscopy. The expression levels of nephrin, podocin, tumor necrosis factor-α (TNF-α), Notch1 intracellular domain (NICD1), triokinase/FMN cyclase (TKFC), specificity protein 1 (Sp1) and histone deacetylase 4 (HDAC4) were determined by Western blot, immunofluorescence and real-time quantitative polymerase chain reaction (qRT-PCR). The chromatin immunoprecipitation (ChIP) assay was performed to evaluate the interaction between Sp1 and the promoter region of HDAC4. Results: Magnolol mitigated the impairment of glomerular filtration function in high fructose-fed rats. Besides, it significantly alleviated the inflammatory responses in glomeruli and HPCs, evidenced by decreased protein levels of TNF-α and NICD1. Increased protein levels of TKFC, Sp1 and HDAC4 were observed in high fructose-stimulated HPCs and rat glomeruli. TMP195, an HDAC4 inhibitor, reduced TNF-α and NICD1 protein levels in high fructose-exposed HPCs. The increased Sp1 was shown to associate with the promoter region of HDAC4, promoting HDAC4 protein expression in high fructose-exposed HPCs. The knockdown of TKFC in HPCs by TKFC siRNA decreased Sp1, HDAC4 and NICD1 protein levels, alleviating podocyte inflammatory response. Furthermore, magnolol inhibited TKFC/Sp1/HDAC4/Notch1 activation in vivo and in vitro. Conclusions: Magnolol attenuated high fructose-induced podocyte inflammation possibly through the suppression of TKFC/Sp1/HDAC4/Notch1 activation, providing new evidence for its potential role in podocyte protection.

Indolo[3,2-c]isoquinoline Hydroxamic Acid Derivatives as Novel Orally Topoisomerase-Histone Deacetylase Dual Inhibitors for NSCLC Therapy.

Based on the synergistic effects of topoisomerase (Top) inhibitors and histone deacetylase (HDAC) inhibitors in cancer therapy, a series of novel Top/HDAC dual inhibitors were designed and synthesized herein. The optimal compound 31 was identified to simultaneously inhibit both Tops and HDACs with potent antiproliferative activity against nonsmall cell lung cancer (NSCLC). Mechanistic studies indicated that compound 31 with increasing reactive oxygen species levels damages DNA, inhibiting cancer cell colony formation and migration and inducing both cancer cell apoptosis and cycle arrest. Noteworthily, compound 31 was orally active in the NSCLC xenograft model, and its antitumor efficacy (TGI = 77.5%, 100 mg/kg) was superior to that of HDAC inhibitor SAHA and SAHA in combination with the Top inhibitor irinotecan. Consequently, this work highlights the therapeutic potential of compound 31 as the Top/HDAC dual inhibitor in NSCLC therapy and provides valuable lead compounds for the further development of antitumor agents in solid tumor therapy.

Apigenin's Influence on Inflammatory and Epigenetic Responses in Rat Lungs After Radiotherapy.

The lung is a moderately radio-sensitive organ. When cells are damaged due to accidental radiation exposure or treatment, they release molecules that lead to the recruitment of immune cells, accumulating inflammatory cytokines at the site of damage. Apigenin (Api) is a natural flavonoid known for its anti-inflammatory properties. In this study, we investigated the radioprotective properties of Api in the lung. Thirty-six Wistar rats were randomly assigned to nine groups: control, radiation (Rad), CMC+Rad, Api10+Rad, and Api20+Rad. Api was administered with an intraperitoneal injection for 7 days, after which the rats were irradiated with 6 Gy whole-body X-ray. At 6 and 72 hours post-irradiation, the rats were euthanized, and their lung tissue was extracted. Radiation led to increased alveolar wall thickness and the infiltration of macrophages and lymphocytes. Furthermore, the expression levels of inflammatory factors such as a nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF-ĸB), Glycogen synthase kinase-3 beta (GSK-3β), transforming growth factor-beta1 (TGF-β1), and epigenetic factors including DNA methyltransferase 3a (DNMT3a) and Histone deacetylase 2 (HDAC2) were elevated in the lung tissue following radiation. Meanwhile, the expression level of IκB-α decreased. However, administration of Api (at both 10&20 mg/kg) reversed the adverse effects of radiation. Api administration mitigated radiation-induced lung damage by reversing inflammatory and epigenetic changes.

Butyrate Inhibits the HDAC8/NF-κB Pathway to Enhance Slc26a3 Expression and Improve the Intestinal Epithelial Barrier to Relieve Colitis.

Dietary fiber is known to promote the production of short-chain fatty acids (SCFAs) by gut bacteria, which can enhance intestinal epithelial barrier function and ameliorate intestinal inflammation in patients with inflammatory bowel disease (IBD). Interestingly, some IBD patients show reduced expression of solute carrier family member 3 (Slc26a3) in intestinal epithelial cells. The objective of this research was to investigate the interaction between SCFAs and Slc26a3 during colitis and assess how this interaction affects intestinal epithelial barrier function. We showed that butyrate alleviated colonic inflammation in a dose-dependent manner in a dextran sulfate sodium salt (DSS)-induced colitis model. Consistent with this, butyrate increased Slc26a3 and tight junction protein levels. In addition, butyrate inhibited histone deacetylase (HDAC) levels and significantly increased the expression of Slc26a3 by the acetylation of histones in Caco-2BBe cells. The utilization of a pan-HDAC inhibitor or inhibitors specific to certain classes of HDACs revealed that butyrate primarily suppressed HDAC8 to blunt the NF-κB pathways and enhance the expression of Slc26a3. Notably, we demonstrated that HDAC8 activation counteracted the beneficial effect of butyrate in DSS-induced colitis. Therefore, we concluded that butyrate improves the expression of Slc26a3 via inhibition of the HDAC8/NF-κB pathway, leading to increased intestinal epithelial barrier function.

Combined HDAC8 and checkpoint kinase inhibition induces tumor-selective synthetic lethality in preclinical models.

The elevated level of replication stress is an intrinsic characteristic of cancer cells. Targeting the mechanisms that maintain genome stability to further increase replication stress and thus induce severe genome instability has become a promising approach for cancer treatment. Here, we identify histone deacetylase 8 (HDAC8) as a drug target whose inactivation synergizes with the inhibition of checkpoint kinases to elicit substantial replication stress and compromise genome integrity selectively in cancer cells. We showed that simultaneous inhibition of HDAC8 and checkpoint kinases led to extensive replication fork collapse, irreversible cell-cycle arrest, and synergistic vulnerability in various cancer cells. The efficacy of the combination treatment was further validated in patient tumor-derived organoid (PDO) and xenograft mouse (PDX) models, providing important insights into patient-specific drug responses. Our data revealed that HDAC8 activity was essential for reducing the acetylation level of structural maintenance of chromosomes protein 3 (SMC3) ahead of replication forks and preventing R loop formation. HDAC8 inactivation resulted in slowed fork progression and checkpoint kinase activation. Our findings indicate that HDAC8 guards the integrity of the replicating genome, and the cancer-specific synthetic lethality between HDAC8 and checkpoint kinases provides a promising replication stress-targeting strategy for treating a broad range of cancers.

Histone deacetylase 2 and 3 of Sarcoptes scabiei: characterization of a potential drug target.

Scabies is a contagious zoonotic parasitic disease that causes a substantial risk to both human and animal health and results in significant financial losses. No vaccine is available for scabies, and drug resistance to the conventional treatment for the disease has increased. Histone deacetylase (HDAC) modifies proteins by removing acetyl moieties from histones, regulates transcription, and is crucial for the immune system and apoptotic processes. This study aimed to clone, express, and determine the immunoreactivity of HDAC-2 and HDAC-3 of scabies mites to investigate their potential as scabies drug targets. The effects of inhibitors on recombinant Sarcoptes scabiei HDAC-2 (rSsHDAC-2) and rSsHDAC-3, as well as on the survival rate and ultrastructure of scabies mites in vitro, were also verified. The findings showed that the inhibitors reduced the acetylase activity of rSsHDAC-2 and rSsHDAC-3. Additionally, these inhibitors could significantly reduce the survival rate of scabies mites, making structural alterations in the mites such as mitochondrial pyknosis and cytoplasmic vacuoles and reaching a fatality rate of 76.7% after 24 h of action. In conclusion, HDAC-2 and HDAC-3 were critical to the survival of scabies mites and might be targeted by medications. Furthermore, the effect of inhibitors on the survival rate and structure of isolated scabies mites provides a new direction for developing therapeutic drugs for scabies.IMPORTANCEIn this study, we successfully cloned and expressed recombinant SsHDAC-2 and SsHDAC-3 in a prokaryotic system and confirmed their acetylation-deacetylase activities. These results provide a solid experimental foundation for subsequent research on SsHDAC-2 and SsHDAC-3. Furthermore, we report for the first time the use of SsHDAC-2 and SsHDAC-3 as drug targets. We demonstrated that the inhibition of these HDACs by pharmacological agents can lead to structural damage in the parasite, thereby impacting the survival activity of the scabies mite. This finding opens up a novel therapeutic avenue for the treatment of scabies.

Distinct Regions within SAP25 Recruit O-Linked Glycosylation, DNA Demethylation, and Ubiquitin Ligase and Hydrolase Activities to the Sin3/HDAC Complex.

Sin3 is an evolutionarily conserved repressor protein complex mainly associated with histone deacetylase (HDAC) activity. Many proteins are part of Sin3/HDAC complexes, and the function of most of these members remains poorly understood. SAP25, a previously identified Sin3A associated protein of 25 kDa, has been proposed to participate in regulating gene expression programs involved in the immune response but the exact mechanism of this regulation is unclear. SAP25 is not expressed in HEK293 cells, which hence serve as a natural knockout system to decipher the molecular functions uniquely carried out by this Sin3/HDAC subunit. Using molecular, proteomic, protein engineering, and interaction network approaches, we show that SAP25 interacts with distinct enzymatic and regulatory protein complexes in addition to Sin3/HDAC. Additional proteins uniquely recovered from the Halo-SAP25 pull-downs included the SCF E3 ubiquitin ligase complex SKP1/FBXO3/CUL1 and the ubiquitin carboxyl-terminal hydrolase 11 (USP11). Furthermore, mutational analysis demonstrates that distinct regions of SAP25 participate in its interaction with USP11, OGT/TETs, and SCF(FBXO3). These results suggest that SAP25 may function as an adaptor protein to coordinate the assembly of different enzymatic complexes to control Sin3/HDAC-mediated gene expression. The data were deposited with the MASSIVE repository with the identifiers MSV000093576 and MSV000093553.

Anti-ovarian cancer migration and toxicity characteristics of a platinum(IV) pro-drug with axial HDAC inhibitor ligands in zebrafish models.

Ovarian cancer is the fifth leading cause of cancer related death in the United States. Cisplatin is a platinum-based anti-cancer drug used against ovarian cancer that enters malignant cells and then damages DNA causing cell death. Typically, ovarian cancer cells become resistant to cisplatin making it necessary to increase subsequent dosage, which usually leads to side-effects including irreversible damage to kidney and auditory system tissue. Ovarian cancer resistance is often associated with upregulation of histone deacetylase (HDAC) enzymes that cause DNA to adopt a closed configuration which reduces the ability of cisplatin to target and damage DNA. Compound B, a platinum(IV) complex with two axial phenylbutyrate (PBA) HDAC inhibitor ligands attached to a cisplatin core, can simultaneously inhibit HDAC activity and damage DNA causing decreased cancer cell viability in cisplatin-sensitive (A2780) and -resistant (A2780cis) ovarian cancer cell lines. However, compound B was not previously evaluated in vivo. As simultaneously inhibiting HDAC-mediated resistance with cisplatin treatment could potentiate the platinum drug's effect, we first confirmed the anti-cancer effect of compound B in the A2780 and A2780cis cell lines using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide spectrophotometric assay. Then, we used zebrafish embryo and transgenic animal models to comparatively analyze the effect of cisplatin, compound B, and controls on general organismal, auditory, and renal system toxicity, and cancer metastasis. We found that lower dosages of compound B (0.3 or 0.6 µM) than of cisplatin (2.0 µM) could cause similar or decreased levels of general, auditory, and renal tissue toxicity, and at 0.6 µM, compound B reduces cancer metastasis more than 2.0 µM cisplatin.

Interplay between acetylation and ubiquitination controls PSAT1 protein stability in lung adenocarcinoma

Serine is essential to maintain maximal growth and proliferation of cancer cells by providing adequate intermediate metabolites and energy. Phosphoserine aminotransferase 1 (PSAT1) is a key enzyme in de novo serine synthesis. However, little is known about the mechanisms underlying PSAT1 degradation. We found that acetylation was the switch that regulated the degradation of PSAT1 in lung adenocarcinoma (LUAD). Deacetylation of PSAT1 on Lys51 by histone deacetylase 7 (HDAC7) enhanced the interaction between PSAT1 and the deubiquitinase ubiquitin-specific processing protease 14 (USP14), leading to the deubiquitination and stabilization of PSAT1; while acetylation of PSAT1 promoted its interaction with the E3 ligase ubiquitination factor E4B (UBE4B), leading to proteasomal degradation. Acetylation of PSAT1 on Lys51 regulated serine metabolism and tumor proliferation in LUAD. Thus, acetylation and ubiquitination cooperatively regulated the protein homeostasis of PSAT1. In conclusion, our study reveals a key regulatory mechanism for maintaining PSAT1 protein homeostasis in LUAD.

Xylanase enhances gut microbiota-derived butyrate to exert immune-protective effects in a histone deacetylase-dependent manner

BackgroundCommensal bacteria in the intestine release enzymes to degrade and ferment dietary components, producing beneficial metabolites. However, the regulatory effects of microbial-derived enzymes on the intestinal microbiota composition and the influence on host health remain elusive. Xylanase can degrade xylan into oligosaccharides, showing wide application in feed industry.ResultsTo validate the immune-protective effects of xylanase, Nile tilapia was used as the model and fed with xylanase. The results showed that dietary xylanase improved the survival rate of Nile tilapia when they were challenged with Aeromonas hydrophila. The transcriptome analysis showed significant enrichment of genes related to interleukin-17d (il-17d) signaling pathway in the xylanase treatment group. High-throughput sequencing revealed that dietary xylanase altered the composition of the intestinal microbiota and directly promoted the proliferation of Allobaculum stercoricanis which could produce butyrate in vitro. Consequently, dietary xylanase supplementation increased the butyrate level in fish gut. Further experiment verified that butyrate supplementation enhanced the expression of il-17d and regenerating islet-derived 3 gamma (reg3g) in the gut. The knockdown experiment of il-17d confirmed that il-17d is necessary for butyrate to protect Nile tilapia from pathogen resistance. Flow cytometry analysis indicated that butyrate increased the abundance of IL-17D+ intestinal epithelial cells in fish. Mechanistically, butyrate functions as an HDAC3 inhibitor, enhancing il-17d expression and playing a crucial role in pathogen resistance.ConclusionDietary xylanase significantly altered the composition of intestinal microbiota and increased the content of butyrate in the intestine. Butyrate activated the transcription of il-17d in intestinal epithelial cells by inhibiting histone deacetylase 3, thereby protecting the Nile tilapia from pathogen infection. This study elucidated how microbial-derived xylanase regulates host immune function, providing a theoretical basis for the development and application of functional enzymes.7BVRPuQYn3eQdwFeU5j8vPVideo

Histone deacetylase inhibition disrupts the molecular signature of the glioblastoma secretome related to extracellular vesicle-associated proteins and targets RAB7a and RAB14 in vitro

Glioblastoma (GBM) is the most aggressive brain tumor with a poor prognosis. While Histone Deacetylase inhibitors have shown promising results in inhibiting cancer cell invasion and promoting apoptosis, their effects on GBM secretion, specifically focusing on extracellular vesicles (EVs) secretion, remain largely unexplored. Using label-free NANOLC-MS/MS methodology, we identified significant changes in the abundance of membrane traffic regulatory proteins in the secretome of U87MG cells after the treatment with the HDAC inhibitor Trichostatin A (TSA). In silico analysis showed that TSA treatment disrupted the secretion pattern of EVs-associated proteins and cellular signaling pathways, both qualitatively and quantitatively. Notably, RAB14/RAB7a interaction was only observed in the secretome of cells treated with TSA. In vitro assays revealed that TSA treatment of glioma cells increased EVs secretion and intracellular protein levels of RAB7a and RAB14 without affecting gene expression, suggesting a role of these two EVs-associated proteins in grade IV glioma cells. Additionally, an integrative approach using clinical data highlighted a correlation between DNA mutations affecting vesicle traffic coding-genes and clinical and phenotypic outcomes in glioma patients. These findings provide insights into the interplay between epigenetics and GBM intracellular trafficking, potentially leading to improved strategies for targeting and modifying the complex signaling network established between GBM cells and the tumor cell microenvironment.

Histone Deacetylases (HDACs) Roles in Inflammation-mediated Diseases; Current Knowledge.

The histone acetyl transferases (HATs) and histone deacetylases (HDACs), which are mostly recognized for their involvement in regulating chromatin remodeling via histone acetylation/deacetylation, have been shown to also change several non-histone proteins to regulate other cellular processes. Acetylation affects the activity or function of cytokine receptors, nuclear hormone receptors, intracellular signaling molecules, and transcription factors in connection to inflammation. Some small-molecule HDAC inhibitors are utilized as anticancer medications in clinical settings due to their capability to regulate cellular growth arrest, differentiation, and death. Here, we summarize our present knowledge of the innate and adaptive immunological pathways that classical HDAC enzymes control. The aim is to justify the targeted (or non-targeted) use of inhibitors against certain HDAC enzymes in inflammatory diseases such as arthritis, inflammatory bowel diseases (IBD), airways inflammation and neurological diseases.

Histone deacetylase 6 inhibition promotes microtubule acetylation and facilitates autophagosome-lysosome fusion in dystrophin-deficient mdx mice.

Duchenne muscular dystrophy is a progressive muscle-wasting disease caused by mutations in the dystrophin gene. Despite progress in dystrophin-targeted gene therapies, it is still a fatal disease requiring novel therapeutics that can be used synergistically or alternatively to emerging gene therapy. Defective autophagy and disorganized microtubule networks contribute to dystrophic pathogenesis, yet the mechanisms by which microtubule alterations regulate autophagy remain elusive. The present study was designed to uncover possible mechanisms underpinning the role of microtubules in regulating autophagy in dystrophic mice. Mdx mice were also supplemented with Tubastatin A, a pharmacological inhibitor of histone deacetylase 6, and pathophysiology was assessed. Mdx mice with a genetic deletion of the Nox-2 scaffolding subunit p47phox were used to assess redox dependence on tubulin acetylation. Our data show decreased acetylation of α-tubulin with enhanced histone deacetylase 6 expression. Tubastatin A increases tubulin acetylation and Q-SNARE complex formation but does not alter microtubule organization or density, indicating improved autophagosome-lysosome fusion. Tubastatin A increases the acetylation of peroxiredoxin and protects it from hyper-oxidation, hence modulating intracellular redox status in mdx mice. Tubastatin A reduces muscle damage and enhances force production. Genetic down regulation of Nox2 activity in the mdx mice promotes autophagosome maturation but not autolysosome formation. Our data highlight that autophagy is differentially regulated by redox and acetylation in mdx mice. By improving autophagy through promoting tubulin acetylation, Tubastatin A decreases the dystrophic phenotype and improves muscle function, suggesting a great potential for clinical translation and treating dystrophic patients.