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Chagas' disease or American trypanosomiasis, caused by Trypanosoma cruzi infection, is an endemic disease in Latin America, which has spread worldwide in the past years. The drugs presently used for treatment have shown limited efficacy due to the appearance of resistant parasites and severe side effects. Some of the most recent studies on anti-parasitic drugs have been focused on protein acetylation, a reversible reaction modulated by Acetyl Transferases (KATs) and Deacetylases (KDACs). We have previously reported the anti-parasite activity of resveratrol (RSV), an activator of KDACs type III (or sirtuins), and showed that this drug can reduce the growth of T. cruzi epimastigotes and the infectivity of trypomastigotes. Since RSV is now widely used in humans due to its beneficial effects as an antioxidant, it has become an attractive candidate as a repurposing drug. In this context, the aim of the present study was to evaluate the ability of this drug to protect three different types of host cells from parasite infection. RSV treatment before parasite infection reduced the percentage of infected cells by 50-70% depending on the cell type. Although the mammalian cell lines tested showed different sensitivity to RSV, apoptosis was not significantly affected, showing that RSV was able to protect cells from infection without the activation of this process. Since autophagy has been described as a key process in parasite invasion, we also monitored this process on host cells pretreated with RSV. The results showed that, at the concentrations and incubation times tested, autophagy was not induced in any of the cell types evaluated. Our results show a partial protective effect of RSV in vitro, which justifies extending studies to an in vivo model to elucidate the mechanism by which this effect occurs.

Histone deacetylase 6 (HDAC6) is known to deacetylate amino acid lysine in alpha-tubulin. However, the functional role of HDAC6 in the progression of cardiac disease remains uncertain. The functional role of HDAC6 in the hearts was examined using transgenic (TG) mice expressing either human wild-type HDAC6, deacetylase inactive HDAC6 (HDAC6supH216A, H611A/sup), and human HDAC6 replaced all serine or threonine residues with aspartic acid at N-terminal 1- 43 amino acids (HDAC6supNT-allD/sup) specifically in the hearts. Overexpression of wild-type HDAC6 significantly reduced acetylated tubulin levels, and overexpression of HDAC6supH216A, H611A/supsignificantly increased it in the mouse hearts. Detectable acetylated tubulin disappeared in HDAC6supNT-allD/supTG mouse hearts. Neither histological alteration nor alteration of cardiac function was observed in the HDAC6 TG mouse hearts. To analyze the role of HDAC6 and acetylated tubulin in disease conditions, we examined HDAC6 in isoprenaline-induced hypertrophy or pressure-overload hypertrophy (TAC). No obvious alteration in the heart weight/body weight ratio or gene expressions of hypertrophic markers between NTG and HDAC6supNT-allD/supmice was observed following treatment with isoprenaline. In contrast, a marked reduction in the shortening fraction and dilated chamber dilatation was detected in the HDAC6supNT-allD/supTG mouse hearts 2 weeks after TAC. A sustained low level of acetylated tubulin and acetylated cortactin was observed in the TAC HDAC6supNT-allD/supTG mouse hearts. Cardiac HDAC6 activity that can regulate acetylated levels of tubulin and cortactin may be critical factors involved in cardiac disease such as pressure-overload hypertrophy.

Early life stress (ELS) can cause long-term changes in gene expression, affect cognition, mood, and behavior, and increase susceptibility to post-traumatic stress disorder (PTSD) in adulthood, in which the histone acetylation plays a crucial role. Studies have found that environmental enrichment (EE) mitigated the unfavorable outcomes of ELS. However, the underlying mechanism of the histone acetylation is not yet completely clear. The purpose of this study was to explore the effect of EE on the histone acetylation after ELS. In this study, using single prolonged stress (SPS) paradigm in early adolescent rats explored the long-term effects of ELS on behavior, the activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs), as well as the acetylation levels of the lysine 9 site of histone H3 (H3K9) and lysine 12 site of histone H4 (H4K12) in the hippocampus and amygdala. Meanwhile, the protective effects of EE intervention were examined. We found that adult male rats exposed to ELS showed behavioral changes, including reduced locomotor activity, increased anxiety-like behaviors, impaired spatial learning and memory, enhanced contextual and cued fear memory, and the HATs/HDACs ratio and acetyl H3K9 (Ac-H3K9) and acetyl H4K12 (Ac-H4K12) were increased in the hippocampus and decreased in the amygdala. Furthermore, EE attenuated the behavioral abnormalities from ELS, possibly through down-regulating the activity of HATs in the hippocampus and up-regulating HDACs activities in the amygdala. These finding suggested that EE could ameliorate ELS-induced PTSD-like behaviors by regulating histone acetylation in the hippocampus and amygdala, reducing the susceptibility to PTSD in adulthood.

Since epigenetic regulation seemed likely to be involved in SV40 early transcription following infection, we have analyzed the organization of nucleosomes carrying histone modifications (acetyl-H3, acetyl-H4, H3K9me1, H3K9me3, H3K4me1, H3K4me3, H3K27me3, H4K20me1) at 30 min and 2 h post infection in SV40 minichromosomes prepared in the absence or presence of the transcription inhibitor dichloro-1-beta-d-ribofuranosyl benzimidazole. The former condition was used to determine how SV40 chromatin structure changed during early transcription, and the latter was used to determine the role of active transcription. The location of RNAPII was used as a marker to identify where histone modifications were most likely to be involved in regulation. Acetyl-H3 acted like epigenetic memory by being present at sites subsequently bound by RNAPII, while H3K9me1 and H3K27me3 were reorganized to the late side of the SV40 regulatory region apparently to repress late transcription. The organization of acetyl-H3 and H3K9me1 but not H3K27me3 required active transcription.

Epigenetic changes associated with histone modifications play an important role in the emergence and maintenance of the phenotype of various cancer types. In contrast to direct mutations in the main DNA sequence, these changes are reversible, which makes the development of inhibitors of enzymes of post-translational histone modifications one of the most promising strategies for the creation of anticancer drugs. To date, a wide variety of histone modifications have been found that play an important role in the regulation of chromatin state, gene expression, and other nuclear events. This review examines the main features of the most common and studied epigenetic histone modifications with a proven role in the pathogenesis of a wide range of malignant neoplasms: acetylation / deacetylation and methylation / demethylation of histone proteins, as well as the role of enzymes of the HAT / HDAC and HMT / HDMT families in the development of oncological pathologies. The data on the relationship between histone modifications and certain types of cancer are presented and discussed. Special attention is devoted to the consideration of various strategies for the development of epigenetic inhibitors. The main directions of the development of inhibitors of histone modifications are analyzed and effective strategies for their creation are identified and discussed. The most promising strategy is the use of multitarget drugs, which will affect multiple molecular targets of cancer. A critical analysis of the current status of approved epigenetic anticancer drugs has also been performed.

A modest autophagy benefits neuroprotection while an excessive autophagy leads to neuronal death after cerebral ischemia, but what governs an appropriate autophagy remains to be understood. Studies indicated that acetylation of histone H4 at lysine16 (H4K16ac) strongly modulated autophagic/lysosomal signaling pathway. Thus, this study was to investigate whether the autophagic neuronal injury could be alleviated by amending H4K16ac level after ischemic stroke. A rat model of middle cerebral artery occlusion (MCAO)/reperfusion was prepared to investigate dynamic variations between H4K16ac and autophagy at the penumbra. The results illustrated that the significantly elevated H4K16ac was coupled with dramatically promoted autophagic activity at 4 h after the insult, suggesting H4K16ac tightly controlled autophagic signaling. After that, H4K16ac level was altered by pretreatment with trichostatin A (TSA, a H4K16ac facilitator) and MG149 (a H4K16ac inhibitor), respectively. Four hours after MCAO/reperfusion, the penumbral tissues were obtained to detect the key proteins in autophagic/lysosomal pathway by western blot and immunofluorescence, respectively. Meanwhile, the infarct volume, neurological deficits, and neuron survival were assessed to evaluate the neurological outcomes. The results showed that TSA-promoted H4K16ac led to an excessively up-regulated autophagy resulting in autophagic/lysosomal dysfunction, as indicated by the accumulated autophagic substrates and exacerbated lysosomal inefficiency in neurons. By contrast, MG149-depressed H4K16ac significantly down-regulated autophagic activity and thereby restored the impaired autophagic flux. Consequently, the neurological injury was markedly alleviated in MCAO + MG149 group, compared with that in MCAO group. Our study suggests that the H4K16ac attenuation elicits neuroprotection against ischemic stroke by ameliorating autophagic/lysosomal dysfunction in neurons.

The optimization of the parameters involved in lignin extraction is crucial for obtaining a lignin with specific structural features for its further valorization. The aim of this work was to develop an eco-friendly organosolv protocol for tuning the acetylation degree of coconut shell lignins (CSLs) by using MgCl

Osmotic stress caused by drought and high salinity is a significant environmental threat that limits plant growth and agricultural yield. Redox regulation plays an important role in plant stress responses, but the mechanisms by which plants perceive and transduce redox signals are still underexplored. Here, we report a critical function for the thiol peroxidase GPX1 in osmotic stress response in rice, where it serves as a redox sensor and transducer. GPX1 is quickly oxidized upon exposure to osmotic stress and forms an intramolecular disulfide bond, which is required for the activation of bZIP68, a VRE-like basic leucine zipper (bZIP) transcription factor involved in the ABA-independent osmotic stress response pathway. The disulfide exchange between GPX1 and bZIP68 induces homo-tetramerization of bZIP68 and thus positively regulates osmotic stress response by regulating osmotic-responsive gene expression. Furthermore, we discovered that the nuclear translocation of GPX1 is regulated by its acetylation under osmotic stress. Taken together, our findings not only uncover the redox regulation of the GPX1-bZIP68 module during osmotic stress but also highlight the coordination of protein acetylation and redox signaling in plant osmotic stress responses.

There is a close connection between epigenetic regulation, cancer metabolism, and immunology. The combination of epigenetic therapy and immunotherapy provides a promising avenue for cancer management. As an epigenetic regulator of histone acetylation, panobinostat can induce histone acetylation and inhibit tumor cell proliferation, as well as regulate aerobic glycolysis and reprogram intratumoral immune cells. JQ1 is a BRD4 inhibitor that can suppress PD-L1 expression. Herein, we proposed a chemo-free, epigenetic-based combination therapy of panobinostat/JQ1 for metastatic colorectal cancer. A novel targeted binary-drug liposome was developed based on lactoferrin-mediated binding with the LRP-1 receptor. It was found that the tumor-targeted delivery was further enhanced by in situ formation of albumin corona. The lactoferrin modification and endogenous albumin adsorption contribute a dual-targeting effect on the receptors of both LRP-1 and SPARC that were overexpressed in tumor cells and immune cells (e.g., tumor-associated macrophages). The targeted liposomal therapy was effective to suppress the crosstalk between tumor metabolism and immune evasion via glycolysis inhibition and immune normalization. Consequently, lactic acid production was reduced and angiogenesis inhibited; TAM switched to an anti-tumor phenotype, and the anti-tumor function of the effector CD8+ T cells was reinforced. The strategy provides a potential method for remodeling the tumor immune microenvironment (TIME).

Brain EGR1 (early growth response protein 1) overexpression aggravates focal ischemic brain injury, but its role in intracerebral hemorrhage (ICH) induced cerebral injury remains obscure. In this study, a rat ICH model was established by injecting type VII collagenase into the brain, and EGR1 knockdown reversed the increase of hematoma area, neurological function score, brain water content, blood–brain barrier (BBB) permeability, inflammation, p300 and retinoid a X receptor-α (RXRα) protein levels, as well as RXRα acetylation level induced by ICH. EGR1 expression was up-regulated in primary brain microvascular endothelial cells (BMECs), neurons, and astrocytes after ICH induction, and the up-regulation was most significant in BMECs. We also found that EGR1 promoted RXRα acetylation level by regulating p300 in BMECs. Subsequently, the BMECs were treated with OGD (oxygen glucose deprivation) plus hemin to simulate ICH condition. And silencing EGR1 rescued the upregulation of cell inflammation and the reduction of cell viability and TEER (transendothelial electric resistance) caused by OGD plus hemin via p300-mediated RXRα acetylation. Furthermore, the STAT3/NF-κB pathway was activated after treatment with OGD plus hemin, which was suppressed by silencing EGR1. And treatment with Stattic (an inhibitor of STAT3) restrained the effect of OGD plus hemin on NF-κB pathway activity, inflammation, cell viability and TEER. In conclusion, EGR1 increased RXRα acetylation level by regulating p300, thereby aggravating brain damage in ICH rat model and dysfunction in BMECs, which might through the STAT3/NF-κB pathway.

SIRT6 belongs to the conserved NAD+-dependent deacetylase superfamily and mediates multiple biological and pathological processes. Targeting SIRT6 by allosteric modulators represents a novel direction for therapeutics, which can overcome the selectivity problem caused by the structural similarity of orthosteric sites among deacetylases. Here, developing a reversed allosteric strategy AlloReverse, we identified a cryptic allosteric site, Pocket Z, which was only induced by the bi-directional allosteric signal triggered upon orthosteric binding of NAD+. Based on Pocket Z, we discovered an SIRT6 allosteric inhibitor named JYQ-42. JYQ-42 selectively targets SIRT6 among other histone deacetylases and effectively inhibits SIRT6 deacetylation, with an IC50 of 2.33 μmol/L. JYQ-42 significantly suppresses SIRT6-mediated cancer cell migration and pro-inflammatory cytokine production. JYQ-42, to our knowledge, is the most potent and selective allosteric SIRT6 inhibitor. This study provides a novel strategy for allosteric drug design and will help in the challenging development of therapeutic agents that can selectively bind SIRT6.

Mitochondria are central to the metabolic circuitry that generates superoxide radicals/anions (O(2)(•−)) as a by-product of oxygen metabolism. By regulating superoxide levels, manganese superoxide dismutase plays important roles in numerous biochemical and molecular events essential for the survival of aerobic life. In this study, we used MitoParaquat (mPQ) to generate mitochondria-specific O(2)(•−) and stable isotope-resolved metabolomics tracing in primary human epidermal keratinocytes to investigate how O(2)(•−) generated in mitochondria regulates gene expression. The results reveal that isocitrate is blocked from conversion to α-ketoglutarate and that acetyl-coenzyme A (CoA) accumulates, which is consistent with a reduction in oxygen consumption rate and inactivation of isocitrate dehydrogenase (IDH) activity. Since acetyl-CoA is linked to histone acetylation and gene regulation, we determined the effect of mPQ on histone acetylation. The results demonstrate an increase in histone H3 acetylation at lysines 9 and 14. Suppression of IDH increased histone acetylation, providing a direct link between metabolism and epigenetic alterations. The activity of histone acetyltransferase p300 increased after mPQ treatment, which is consistent with histone acetylation. Importantly, mPQ selectively increased the nuclear levels and activity of the oxidative stress-sensitive nuclear factor erythroid 2-related factor 2. Together, the results establish a new paradigm that recognizes O(2)(•−) as an initiator of metabolic reprogramming that activates epigenetic regulation of gene transcription in response to mitochondrial dysfunction.

Biological redox signaling plays an important role in many diseases. Redox signaling involves reductive and oxidative mechanisms. Oxidative stress occurs when reductive mechanism overwhelms oxidative challenges. Cellular oxidative stress occurs when reactive oxygen/nitrogen species (RO/NS) exceed the cellular reductive/antioxidant capacity. Endogenously produced RO/NS from mitochondrial metabolic citric-acid-cycle coupled with electron-transport-chain or exogenous stimuli trigger cellular signaling events leading to homeostatic response or pathological damage. Recent evidence suggests that RO/NS also modulate epigenetic machinery driving gene expression. RO/NS affect DNA methylation/demethylation, histone acetylation/deacetylation or histone methylation/demethylation. Many health beneficial phytochemicals possess redox capability that counteract RO/NS either by directly scavenging the radicals or via inductive mechanism of cellular defense antioxidant/reductive enzymes. Amazingly, these phytochemicals also possess epigenetic modifying ability. This review summarizes the latest advances on the interactions between redox signaling, mitochondrial metabolism, epigenetics and redox active phytochemicals and the future challenges of integrating these events in human health.

β-Hydroxybutyl acid (βOHB), the most prevalent type of ketone in the human body, is involved in the pathogenesis of cognitive disorders, especially Alzheimer's dementia (AD), through a variety of mechanisms, such as enhancing mitochondrial metabolism, regulating signaling molecule, increasing histone acetylation, affecting the metabolism of Aβ and Tau proteins, inhibiting inflammation and lipid metabolism, and regulating intestinal microbes. Based on the above findings, clinical drug development in AD has begun to focus on βOHB.

Graphical abstract, Highlights • Cr(VI) can induce inflammatory, oxidative stress, and apoptosis in rat liver. • Cr(VI) induces inflammatory response in the liver by inhibiting deacetylation. • Oxidative stress caused by Cr(VI) is related to the inhibition of deacetylation. • Cr(VI) aggravates hepatocyte apoptosis by inhibiting deacetylation in rats. • Cr(VI) induces liver injury via inhibition of the deacetylation of Sirt1., Introduction Hexavalent chromium (Cr(VI)), one of the toxic heavy metals, poses a serious threat to human and animal health. Protein acetylation regulates the structure and function of most proteins in a variety of ways. However, the hepatotoxicity of Cr(VI) and whether it is related to deacetylation remains largely unknown. Objectives We aimed to explore the link between the deacetylation of silent information regulator two ortholog 1 (Sirt1) and hepatotoxicity induced by Cr(VI) exposure, and to better clarify the biological mechanism of liver injury induced by Cr(VI). Methods We established a model of liver injury of K2Cr2O7 by injecting rats intraperitoneally for 35 days continuously and adding resveratrol (Res) to further explore the link between deacetylation and hepatotoxicity. Results The results revealed that Cr(VI) induced inflammatory response and apoptosis in hepatocytes. Furthermore, Cr(VI) reduced Sirt1 expression and inhibited the deacetylation of Sirt1 to downstream key transcription factors, including nuclear factor erythroid 2-related factor 2 (Nrf2), Forkhead box O3 (FOXO3), and nuclear factor-kappa B (NF-κB). Conversely, when Res was administered as an activator of Sirt1, the deacetylation of Sirt1 was enhanced, and inflammatory response and apoptosis were significantly alleviated. Conclusion In summary, this work firstly demonstrates that Cr(VI) induces liver injury in rat by inhibiting the deacetylation of Sirt1, which is of positive significance for protecting the natural environment and animal health from chronic Cr poisoning.

Histone deacetylases (HDACs) inhibit the acetylation of crucial autophagy genes, thereby deregulating autophagy and autophagic cell death (ACD) and facilitating cancer cell survival. Vorinostat, a broad-spectrum pan-HDAC inhibitor, inhibits the deacetylation of key autophagic markers and thus interferes with ACD. Vorinostat-regulated ACD can have an autophagy-mediated, -associated or -dependent mechanism depending on the involvement of apoptosis. Molecular insights revealed that hyperactivation of the PIK3C3/VPS34–BECN1 complex increases lysosomal disparity and enhances mitophagy. These changes are followed by reduced mitochondrial biogenesis and by secondary signals that enable superactivated, nonselective or bulk autophagy, leading to ACD. Although the evidence is limited, this review focuses on molecular insights into vorinostat-regulated ACD and describes critical concepts for clinical translation.

Current study is intended to evaluate the expression and epigenetic variations of mitochondrial situins in 306 rheumatoid arthritis (RA) cases and compared with age/gender matched controls.The expression level was measured using the quantitative Real time PCR (qPCR) and epigenetic analysis was performed by measuring deacetylation activity. Oxidative stress was also measured in present study using the enzyme linked immunoassay (ELISA). The obtained results were evaluated by means of the student t-test, spearman correlation and ROC curve analysis.Expression analysis showed the significant downregulation of SIRT3 (p 0.0001), SIRT4 (p 0.0001) and SIRT5 (p 0.0001) in RA cases when compared with controls. Downregulation of mitochondrial sirtuins was significantly associated with positive anti-CCP status, increased ESR level and with increased CRP levels. Epigenetic analysis showed significant increased histone deacetylation in RA patients compared to controls. Co-expression analysis showed the significant negative association between expression level of mitochondrial sirtuins and deacytylation level (SIRT3 r = -0.438, p 0.0001; SIRT4 r = -0.424, p 0.0001; SIRT5 r = -0.282, p 0.0001). ROC curve analysis exhibited that downregulation of mitochondrial sirtuins (SIRT3 AUC = 0.91, p 0.001; SIRT4 AUC = 0.92, p 0.001; SIRT5 AUC = 0.85, p 0.001) was act as the good diagnostic marker for detection/diagnosis of arthritis.The results show that significant deregulation of mitochondrial sirtuins was associated with increased arthritis risk and can be act as an indicator of advance clinical outcome.

Multiple sequence alignment of homoserine-acetyltransferases, serine-acetyltransferases and homoserine-succinyltransferases show they all belong to MetX family, having evolved from a common ancestor by conserving the catalytic site and substrate binding residues. The discrimination in the substrate selection arises due to the presence of substrate-specific residues lining the substrate-binding pocket. Mutation of Ala59 and Gly62 to Gly and Pro respectively in homoserine-acetyltransferase from M. tuberculosis resulted in a serine-acetyltransferase like enzyme as it acetylated both l -homoserine and l -serine. Homoserine-acetyltransferase from M. tuberculosis when mutated at positon 322 where Leu was converted to Arg, resulted in succinylation over acetylation of l -homoserine. Our studies establish the importance of the substrate binding residues in determining the type of activity possessed by MetX family, despite all of them having the same catalytic triad Ser-Asp-His. Hence key residues at the substrate binding pocket dictate whether the given enzyme shows predominant transferase or hydrolase activity.

Sorafenib and lenvatinib are approved first-line targeted therapies for advanced liver cancer, but most patients develop acquired resistance. Herein, we found that sorafenib induced extensive acetylation changes towards a more energetic metabolic phenotype. Metabolic adaptation was mediated via acetylation of the Lys-491 (K491) residue of phosphoenolpyruvate carboxykinase isoform 2 (PCK2) (PCK2-K491) and Lys-473 (K473) residue of PCK1 (PCK1-K473) by the lysine acetyltransferase 8 (KAT8), resulting in isoenzyme transition from cytoplasmic PCK1 to mitochondrial PCK2. KAT8-catalyzed PCK2 acetylation at K491 impeded lysosomal degradation to increase the level of PCK2 in resistant cells. PCK2 inhibition in sorafenib-resistant cells significantly reversed drug resistance in vitro and in vivo. High levels of PCK2 predicted a shorter progression-free survival time in patients who received sorafenib treatment. Therefore, acetylation-induced isoenzyme transition from PCK1 to PCK2 contributes to resistance to systemic therapeutic drugs in liver cancer. PCK2 may be an emerging target for delaying tumor recurrence.

Mammalian hibernation is a period that involves substantial metabolic change in order to promote survival in harsh conditions, with animals typically relying on non-carbohydrate fuel stores during long bouts of torpor. However, the use and maintenance of carbohydrate fuel stores remains important during periods of arousal from torpor as well as when exiting hibernation. Gluconeogenesis plays a key role in maintaining glucose stores; however, little is known about this process within the muscles of hibernating mammals. Here, we used 13-lined ground squirrels (Ictidomys tridecemlineatus) as our model for mammalian hibernation, and showed that skeletal muscle fructose-1,6-bisphosphatase (FBPase; EC 3.1.3.11), the rate-limiting enzyme for the gluconeogenic pathway, was suppressed during torpor as compared to the euthermic control. A physical assessment of partially purified FBPase via exposure to increasing concentrations of the denaturant urea indicated that FBPase from the two conditions were structurally distinct. Western blot analysis suggests that the kinetic and physical differences between euthermic and torpid FBPase may be derived from differential acetylation, whereby increased acetylation of the torpid enzyme makes FBPase more rigid and less active. This study increases our understanding of skeletal muscle carbohydrate metabolism during mammalian hibernation and sets forth a potentially novel mechanism for the regulation of FBPase during environmental stress.

Dual modification of cassava starch was carried out using ultrasonication and acetylation by acetic acid by altering the sequence. The results revealed that the type of modification and sequence of modification for dual modified starches significantly affected the properties of starch. The swelling decreased for all the modified starches whereas solubility decreased for ultrasonicated starches but increased for acetylated starch and dual modified starch where acetylation was done after ultrasonication. The paste viscosities of all the modified starches were found to be significantly lower compared to native starch and the lowest viscosities were observed for dual modified starch where ultrasonication was done after acetylation. The resistant starch and slowly digestible starch content of the modified starches were significantly higher than in native starch, and the type of modification and sequence of modification for dual modified starches seemed to affect the digestibility of starches.

Since being discovered, alkali/urea has been widely used in the dissolution of natural polysaccharides and the preparation of functional materials such as hydrogels, fibers, films and nanoparticles. This work will focus on verifying the structural stability, homogeneous degradation and deacetylation of chitin in alkali-soluble systems. The chitin was dissolved in NaOH/urea solution and stored at different temperature. At the specific time, the structure, viscosity, acetylation degree (DA) and biocompatibility of chitin and prepared chitosan were determined. The results indicated that dissolution process did not affect the structure and bioactivity of chitin. However, with the increase of storage time and temperature, chitin undergone significant homogeneous deacetylation (DA from 99.5% to 33.2%) and degradation (viscosity from 9284 cP to 1538 cP), accompanying by changes in crystalline structure and thermal stability. Moreover, the processed chitins were no-toxic for the biomedicine applications. This work will provide new ideas for the application of alkali-soluble systems.

Non-invasive radiofrequency acts through an electromagnetic field that generates a thermal effect on adipose tissue without contact with the skin. The temperature in the subcutaneous reaches between 44°C to 45°C, which after 45 minutes of intervention causes apoptosis of adipocytes. The objective of this study is investigate the impact of non-invasive radiofrequency (RF) on global histone H3 acetylation and heat shock proteins (HSPs) 47 and 60 levels in the peripheral blood of healthy women. The participants were divided in control/sedentary (n = 10) and exercised/physically active (n = 10) groups and were submitted to 4 sessions of RF intervention (once a week; 45 min each session, total 4 sessions). In order to analyze the acute and long term effects, blood samples for biomarkers analysis were taken in 3 moments: before (T1), after the first session (T2) and after the last session (T3), while anthropometric measurements were taken before and after the treatment protocol. The intervention was not able to modulate the HSPs in both groups at any time, while a significant increase on global histone H3 levels was observed at T2 compared to T1 (p=0.047) in exercised group. Regarding anthropometric measurements, we observed a significant reduction in body mass and adipose mass in the exercised group in T3 when compared to T1. Non-invasive RF is able to improve anthropometric measurements in physically active healthy women. This response might be related to acute histone H3 hyperacetylation status and is not associated with the modulation of HSPs 47 and 60.

Protein lysine acetylation has emerged as a major regulatory post-translational modification in different organisms, present not only on histone proteins affecting chromatin structure and gene expression but also on nonhistone proteins involved in several cellular processes. The same scenario was observed in protozoan parasites after the description of their acetylomes, indicating that acetylation might regulate crucial biological processes in these parasites. The demonstration that glycolytic enzymes are regulated by acetylation in protozoans shows that this modification might regulate several other processes implicated in parasite survival and adaptation during the life cycle, opening the chance to explore the regulatory acetylation machinery of these parasites as drug targets for new treatment development.

Responsible for interpreting histone post-translational modifications (PTMs), epigenetic reader proteins have emerged as novel therapeutic targets for a wide range of diseases. Chemical probes have been critical in enabling target validation studies and have led to translational advances in cancer and inflammation-related pathologies. Here, we present the most recently reported probes of reader proteins that recognize acylated and methylated lysine. We will discuss challenges associated with achieving potent antagonism of reader domains and review ongoing efforts to overcome these hurdles, focusing on targeting strategies including the use of peptidomimetic ligands, allosteric modulators, and protein degraders.

Biological aging is attributed to progressive dysfunction in systems governing genetic and metabolic integrity. At the cellular level, aging is evident by accumulated DNA damage and mutation, reactive oxygen species, alternate lipid and protein modifications, alternate gene expression programs, and mitochondrial dysfunction (Sun et al., 2016; Nekhaeva et al., 2002; Kong et al., 2014). These effects sum to drive altered tissue morphology and organ dysfunction (Ballard and Edelberg, 2008; Han and Ren, 2010; Pina and Fitzpatrick, 1996). Protein-acylation has emerged as a critical mediator of age-dependent changes in these processes (Sun et al., 2016; Nekhaeva et al., 2002; Kong et al., 2014). Despite decades of research focus from academia and industry, heart failure remains a leading cause of death in the United States while the 5 year mortality rate for heart failure remains over 40% (Benjamin et al. 2019). Over 90% of heart failure deaths occur in patients over the age of 65 and heart failure is the leading cause of hospitalization in Medicare beneficiaries (Strait and Lakatta, 2012). In 1931, Cole and Koch discovered age-dependent accumulation of phosphates in skeletal muscle (Cole and Koch, 1931). These and similar findings provided supporting evidence for, now well accepted, theories linking metabolism and aging. Nearly two decades later, age-associated alterations in biochemical molecules were described in the heart (Kaufman and Poliakoff, 1950). From these small beginnings, the field has grown substantially in recent years. This growing research focus on cardiac aging has, in part, been driven by advances on multiple public health fronts that allow population level clinical presentation of aging related disorders. It is estimated that by 2030, 25% of the worldwide population will be over the age of 65 (Lakatta, 2002). This review provides an overview of acetylation-dependent regulation of biological processes related to cardiac aging and introduces emerging non-acetyl, acyl-lysine modifications in cardiac function and aging.

Reversible acylation of lysine ε-amino groups, e.g., acetylation, succinylation, maronylation, and myristoylation, is involved in basic physiological processes such as metabolism, cell signaling and aging. In this study, we developed a novel enrichment method for acylated peptides without the use of antibodies, in which endogenously acylated peptides are deacylated by recombinant lysine deacylases based on the enzyme-substrate relationship and enriched by N-hydroxysuccinimidyl chemistry for identification of the acylated sites by nanoscale liquid chromatography-tandem mass spectrometric analysis. To demonstrate the validity of this acylomics platform, we used it to identify acylated sites on chemically acylated model protein samples. We also applied it to the nuclei of HeLa cells to identify endogenous acylated sites.

Background and aims The thiol transferase glutaredoxin 1 controls redox signaling and cellular functions by regulating the S-glutathionylation status of critical protein thiols. Here we tested the hypothesis that by derepressing the expression of glutaredoxin 1, inhibition of histone deacetylase 2 prevents nutrient stress-induced protein S-glutathionylation and monocyte dysfunction and protects against atherosclerosis. Methods Using both a pharmacological inhibitor and shRNA-mediated knockdown of histone deacetylase 2, we determine the role of this deacetylase on glutaredoxin 1 expression and nutrient stress-induced inactivation of mitogen-activated protein kinase phosphatase 1 activity and monocyte and macrophage dysfunction. To assess whether histone deacetylase 2 inhibition in myeloid cells protects against atherosclerosis, we fed eight-week-old female and male HDAC2−/−MyeloidLDLR−/− mice and age and sex-matched LysMcretg/wtLDLR−/− control mice a high-calorie diet for 12 weeks and assessed monocyte function and atherosclerotic lesion size. Results Myeloid histone deacetylase 2 deficiency in high-calorie diet-fed LDLR−/− mice reduced atherosclerosis in males by 39% without affecting plasma lipid and lipoprotein profiles or blood glucose levels but had no effect on atherogenesis in female mice. Macrophage content in plaques of male mice was reduced by 31%. Histone deacetylase 2-deficient blood monocytes from male mice showed increased acetylation on histone 3, and increased Grx1 expression, and was associated with increased MKP-1 activity and reduced recruitment of monocyte-derived macrophages, whereas in females, myeloid HDAC2 deficiency had no effect on Grx1 expression, did not prevent nutrient stress-induced loss of MKP-1 activity in monocytes and was not atheroprotective. Conclusions Specific histone deacetylase 2 inhibitors may represent a potential novel therapeutic strategy for the prevention and treatment of atherosclerosis, but any benefits may be sexually dimorphic.

Age-related cognitive impairment is associated with diminished autophagy and progressively increased neuroinflammation. Histone acetylation has been shown to be a key process in sevoflurane-induced neurobehavioral abnormalities. Here, we investigated whether histone acetylation regulates the interaction between autophagy and the NLRP3 inflammasome in models of sevoflurane-induced cognitive impairment and explored the underlying molecular mechanisms. Aged C57BL/6 J mice and cultured primary hippocampal neurons were exposed to 3% sevoflurane for 2 h. Hippocampal tissue samples and hippocampal neurons were harvested. The processes of histone acetylation and autophagy and the activation of the NLRP3 inflammasome were observed using western blotting, immunofluorescence staining, and transmission electron microscopy. Suberoylanilide hydroxamic acid (SAHA), an inhibitor of histone deacetylases, increased histone H3 and H4 acetylation in both the mouse hippocampus and primary neurons. Concomitantly, sevoflurane upregulated components of the NLRP3 inflammasome (NLRP3, cleaved caspase-1, and IL-1β) by promoting autophagic degradation in the aging brain. Cognitive deficits and inadequate autophagy induced by sevoflurane were reversed and NLRP3 inflammasome activation was inhibited by SAHA. Treatment with 3-MA, an autophagy inhibitor, eliminated the neuroprotective effects of SAHA on improving cognition in mice, activating autophagy and downregulating the NLRP3 inflammasome. Based on these results, histone acetylation activates autophagy plays an important role in inhibiting the activation of the NLRP3 inflammasome to protect the host from excessive neuroinflammation and sevoflurane-induced cognitive dysfunction in the aging brain.

Polysaccharides were extracted by hot water and alkali in sequence from Dolichos lablab L. hull, and further purified by ion-exchange and gel columns. Hot water extracted D. lablab hull polysaccharide (DLHP) was rich in glucuronoxylan and pectin, and alkali extracted polysaccharide (DLHAP) mostly embraced glucuronoxylan. The structures of purified glucuronoxylans from DLHP and DLHAP were mainly analyzed by HPAEC-PAD, methylation combined with GC–MS, NMR and SEC-MALLS. DLHP-1 was identified as acetylated glucuronoxylan containing →4)-β-Xylp-(1→ backbone with substitution at O-2 site by α-GlcpA/4-O-methyl-α-GlcpA. The molar ratio of β-Xylp to α-GlcpA was 6.9:1, and acetylation was mainly at O-3 site of β-Xylp with acetylation degree of 21.5%. DLHP-1 and DLHP-2 had similar physicochemical properties, except for molecular weight (Mw). DLHAP-1 was the non-methylated glucuronoxylan almost without acetylation, and it had the molar ratio of β-Xylp to α-GlcpA of 5.6:1. Besides, DLHP-1 (Mw of 20.0 × 103 g mol−1) adopted semi-flexible chain, while DLHAP-1 (Mw of 15.4 × 103 g mol−1) showed flexible chain. These results provided a structural basis for study on polysaccharides from D. lablab hull, which was benefit for understanding biological activities and developing functional food or pharmaceuticals of D. lablab.

The present study investigates the effect of different acetylated cassava starch (ACS) concentrations on the physical and rheological properties of multicomponent emulsion-based products at specific pH values. The emulsion-based products were made by mixing 2% (w/v) prepared lipid droplets that were stabilized by either native or heated whey proteins, 0.01% (w/v) flaxseed gum and 0-6.0% (w/v) ACS. The results indicated that particle size, apparent viscosity and rheological moduli of multicomponent emulsion-based products were significantly enhanced with increasing addition amounts of ACS (P < 0.05). Moreover, the microscopic morphology showed that the addition of ACS contributed to the formation of a more compact, uniform, and continuous comb-like network. However, higher ACS concentration was prone to induce visibly larger aggregations and coarser textures, lending to some negative impact on visual appearance and overall acceptability. Moreover, acidic conditions could obviously promote droplet aggregation via electrostatic interactions, whereas neutral conditions had no effect on droplet aggregation. Additionally, when compared with native whey proteins, lipid droplets stabilized by their heated protein forms induced significantly higher apparent viscosities and rheological moduli of multicomponent emulsion-based products (P < 0.05). Our results potentially provide some information for the creation of multicomponent emulsion-based products with various desirable quality attributes.

Podocyte injury is associated with albuminuria and the progression of diabetic nephropathy (DN). MiR-34a, a p53-regulated miRNA, directly targets SIRT1 and contributed to DN progression. MiR-34a represses SIRT1 to activate p53 and establish a positive feedback loop. However, whether p53/miR-34a/SIRT1 signaling is activated in podocytes and contributes to DN pathogenesis remains elusive. In this study, we observed that serum miR-34a level was positively correlated with podocyte injury in DN patients. The expression of acetylated p53 and miR-34a was upregulated, SIRT1was downregulated in glomeruli from patients with DN and STZ induced diabetic mice, as well as in human podocytes treated with advanced glycation end (AGE). MiR-34a antagonism in vitro and vivo in STZ induced diabetic mice developed alleviated glomerulus injury as reflected by attenuated albuminuria, reduced podocyte loss and restored autophagic flux. In human podocyte, inhibition of AGE formation by pyridoxamine prevented miR-34a dependent repression of SIRT1, p53 acetylation and activate podocyte autophagy in a dose-dependent manner. MiR-34a overexpression increases acetylation of p53 by translational repression of SIRT1. SIRT1 overexpression also impacts AGE induced apoptosis through deacetylating p53, whereas silencing of SIRT1 by EX527 attenuated the cytoprotective functions of miR-34a knockdown. Moreover, blockade of p53 acetylation significantly rescued miR-34a-induced apoptosis through SIRT1 restoration. Collectively, we demonstrate that by activation of p53, AGE induced the transcription of miR-34a, miR-34a in turn repressed SIRT1 to activate p53, resulting in a positive-feedback loop and contributing to podocyte injury. Targeting modulation of p53/miR-34a/SIRT1 feedback by miR-34a knockdown or overexpression of SIRT1 could rescue podocyte injury during DN.

Histone variants are a universal means to alter the biochemical properties of nucleosomes, implementing local changes in chromatin structure. H2A.Z, one of the most conserved histone variants, is incorporated into chromatin by SWR1-type nucleosome remodelers. Here, we summarize recent advances toward understanding the transcription-regulatory roles of H2A.Z and of the remodeling enzymes that govern its dynamic chromatin incorporation. Tight transcriptional control guaranteed by H2A.Z nucleosomes depends on the context provided by other histone variants or chromatin modifications, such as histone acetylation. The functional cooperation of SWR1-type remodelers with NuA4 histone acetyltransferase complexes, a recurring theme during evolution, is structurally implemented by species-specific strategies.

SummaryCellular global translation is often measured using ribosome profiling or quantitative mass spectrometry, but these methods do not provide direct information at the level of elongating nascent polypeptide chains (NPCs) and associated co-translational events. Here we describe pSNAP, a method for proteome-wide profiling of NPCs by affinity enrichment of puromycin- and stable isotope-labeled polypeptides. pSNAP does not require ribosome purification and/or chemical labeling, and captures bona fide NPCs that characteristically exhibit protein N-terminus-biased positions. We applied pSNAP to evaluate the effect of silmitasertib, a potential molecular therapy for cancer, and revealed acute translational repression through casein kinase II and mTOR pathways. We also characterized modifications on NPCs and demonstrated that the combination of different types of modifications, such as acetylation and phosphorylation in the N-terminal region of histone H1.5, can modulate interactions with ribosome-associated factors. Thus, pSNAP provides a framework for dissecting co-translational regulations on a proteome-wide scale.

Background & Aims Lactate has recently been reported to accumulate in the livers of patients progressing from simple steatosis to non-alcoholic steatohepatitis (NASH). However, the underlying mechanism(s) of lactate accumulation and the role of lactate in the progression of non-alcoholic fatty liver disease (NAFLD) are essentially unknown. Methods We compared the acetylome in liver samples taken from healthy individuals, patients with simple steatosis and patients with NASH to identify potential targets of acetylation with a role in lactate metabolism. Interactions between the acetylated target and acetyltransferases were measured in multiple cell lines. An acetyltransferase inhibitor was injected into high-fat diet (HFD)-fed mice to determine the role of lactate on NAFLD progression in vivo. Results Hyperacetylation of lactate dehydrogenase B (LDHB) was found to be associated with lactate accumulation in NAFL and NASH livers in humans and mice. P300/CBP-associated factor (PCAF)-mediated acetylation of LDHB K82 was found to significantly decrease LDHB activity and impair hepatic lactate clearance, resulting in lactate accumulation. Acetylated LDHB induced lactate accumulation which exacerbated lipid deposition and inflammatory responses by activating histone hyperacetylation in HFD-induced NASH. The administration of embelin, a PCAF inhibitor, and the generation of an acetylation-deficient mutant of LDHB ameliorated NASH. Conclusion PCAF-dependent LDHB acetylation plays a key role in hepatic lipid accumulation and inflammatory responses by impairing lactate clearance; this process might be a potential therapeutic target for the treatment of NASH. Lay summary Lactate is known to accumulate in the livers of patients during the progression of non-alcoholic fatty liver disease (NAFLD); however, the underlying mechanism(s) of this accumulation and its importance in disease progression are unknown. Herein, we show that the acetylation of an enzyme involved in lactate metabolism leads to impaired lactate clearance and exacerbates NAFLD progression.

Shedding, loss of expression, or internalization of natural killer group 2, member D (NKG2D) ligands from the tumor cell surface leads to immune evasion, which is associated with poor prognosis in patients with cancer. In many cancers, matrix metalloproteinases cause the proteolytic shedding of NKG2D ligands. However, it remained unclear how to protect NKG2D ligands from shedding. Here, we showed that the shedding of the mouse NKG2D ligand Rae-1 can be prevented by two critical acetyltransferases, GCN5 and PCAF, which acetylate the lysine residues of Rae-1 to avoid shedding both in vitro and in vivo. In contrast, mutations at lysines 80 and 87 of Rae-1 abrogated this acetylation and thereby desensitized tumor cells to NKG2D-dependent immune surveillance. Notably, the protein levels of GCN5 correlated with the expression levels of the human NKG2D ligand ULPB1 in a human tumor tissue microarray and, more importantly, with prolonged overall survival in many cancers. Our results suggest that the acetylation of Rae-1 protein at lysines 80 and 87 by GCN5 and PCAF protects Rae-1 from shedding so as to activate NKG2D-dependent immune surveillance. This discovery may shed light on new targets for NKG2D immunotherapy in cancer treatment.

WDR35/IFT121 is an intraflagellar transport protein in primary cilia, which is associated with RagA, an mTORC1-activating protein. To elucidate the functions of the interaction between WDR35 and RagA in primary cilia, as well as mTOR signaling, we identified WDR35-interacting proteins using mass spectrometry. We found that WDR35 associates with CCT complex proteins including TCP1/CCT1, which act as molecular chaperones for α-tubulin folding. Immunostaining showed that acetylated α-tubulin was concentrated in the vicinity of primary cilia in 293T cells. In contrast, acetylated tubulin was dispersed in WDR35 partial knockout cells established from 293T cells. Similarly, scattered subcellular localization of acetylated tubulin was observed in RagA knockout cells. RagA was present in the primary cilia of NIH3T3 cells, and the GDP form of RagA exhibited strong binding to WDR35 and negative regulation of primary cilium formation. These results suggest that WDR35 is involved in the subcellular localization of acetylated tubulin in primary cilia via its interactions with TCP1 and/or RagA family proteins.

Biomolecules undergo liquid-liquid phase separation (LLPS), resulting in the formation of multicomponent protein-RNA membraneless organelles in cells. However, the physiological and pathological role of post-translational modifications (PTMs) on the biophysics of phase behavior is only beginning to be probed. To study the effect of PTMs on LLPS in silico, we extend our transferable coarse-grained model of intrinsically disordered proteins to include phosphorylated and acetylated amino acids. Using the parameters for modified amino acids available for fixed-charge atomistic force fields, we parameterize the size and atomistic hydropathy of the coarse-grained-modified amino acid beads and, hence, the interactions between the modified and natural amino acids. We then elucidate how the number and position of phosphorylated and acetylated residues alter the protein’s single-chain compactness and its propensity to phase separate. We show that both the number and the position of phosphorylated threonines/serines or acetylated lysines can serve as a molecular on/off switch for phase separation in the well-studied disordered regions of Fused in Sarcoma (FUS) and DDX3X, respectively. We also compare modified residues to their commonly used PTM mimics for their impact on chain properties. Importantly, we show that the model can predict and capture experimentally measured differences in the phase behavior for position-specific modifications, showing that the position of modifications can dictate phase separation. In sum, this model will be useful for studying LLPS of post-translationally modified intrinsically disordered proteins and predicting how modifications control phase behavior with position-specific resolution.

Multiple studies have found that increasing histone acetylation by means of histone deacetylase inhibitor (HDACi) treatment can ameliorate memory and rescue cognitive impairments, but their mode of action is not fully understood. In particular, it is unclear how HDACis, applied systemically and devoid of genomic target selectivity, would specifically improve memory-related molecular processes. One theory for such specificity is called cognitive epigenetic priming (CEP), according to which HDACis promote memory by facilitating the expression of neuroplasticity-related genes that have been stimulated by learning itself. In this review, we summarize the experimental evidence in support of CEP, describe newly discovered off-target effects of HDACis and highlight similarities between drug-induced and naturally occurring CEP. Understanding the underlying mechanisms of CEP is important in light of the preclinical premise of HDACis as cognitive enhancers.

Anaplastic thyroid carcinoma (ATC) is one of the most aggressive malignancies frequently associated with extrathyroidal extension and metastasis through pathways that remain unclear. Analysis of the cancer genome atlas (TCGA) database and an independent cohort showed that the expression of hematological and neurological expressed 1 (HN1) was higher in thyroid cancers than in normal tissues, and negatively correlated with progression-free survival. RT-PCR and immunohistochemistry revealed higher HN1 expression in ATC compared to healthy tissues and papillary thyroid carcinoma (PTC). HN1 knockdown attenuated migration and invasion of ATC cells, whereas HN1 overexpression increased migration and invasion of PTC cells. HN1 reduced the acetylation of α-tubulin and promoted progression through epithelial-mesenchymal transition of ATC cells and mouse xenografts. HN1 knockdown significantly attenuated TGF-β-induced mesenchymal phenotype, and inhibited tumor formation and growth of ATC xenografts in nude mice. Loss of STMN1 decreased the malignant potential of HN1, whereas HN1 knockdown in combination with STMN1 overexpression restored the aggressive properties of ATC cells. HN1 increased STMN1 mRNA expression, and prevented STMN1 ubiquitination and subsequent degradation. These results demonstrate that HN1 interacts with STMN1 and drives ATC aggressiveness.

We intended to characterize functional relevance of microRNA (miR)-224-3p in endothelial cell (EC) apoptosis and reactive oxygen species (ROS) accumulation in atherosclerosis, considering also the integral involvement of histone deacetylase 1 (HDAC1)-mediated hypoxia-inducible factor-1α (HIF1α) deacetylation. The binding affinity between miR-224-3p and Fos-like antigen 2 (FOSL2) was predicted and validated. Furthermore, we manipulated miR-224-3p, FOSL2, HDAC1, and HIF1α expression in oxidized low-density lipoprotein (ox-LDL)-induced ECs, aiming to clarify their effects on cell activities, inflammation, and ROS level. Additionally, we examined the impact of miR-224-3p on aortic atherosclerotic plaque and lesions in a high-fat-diet-induced atherosclerosis model in ApoE−/− mice. Clinical atherosclerotic samples and ox-LDL-induced human aortic ECs (HAECs) exhibited low HDAC1/miR-224-3p expression and high HIF1α/FOSL2 expression. miR-224-3p repressed EC cell apoptosis, inflammatory responses, and intracellular ROS levels through targeting FOSL2. HIF1α reduced miR-224-3p expression to accelerate EC apoptosis and ROS accumulation. Moreover, HDAC1 inhibited HIF1α expression by deacetylation, which in turn enhanced miR-224-3p expression to attenuate EC apoptosis and ROS accumulation. miR-224-3p overexpression reduced atherosclerotic lesions in vivo. In summary, HDAC1 overexpression may enhance the anti-atherosclerotic and endothelial-protective effects of miR-224-3p-mediated inhibition of FOSL2 by deacetylating HIF1α, underscoring a novel therapeutic insight against experimental atherosclerosis., Graphical Abstract, The study mainly addressed the suppressive role of HDAC1-mediated deacetylation of HIF1α in atherosclerosis progression, which was associated with miR-224-3p-mediated inhibition of FOSL2, thus highlighting a potential therapeutic action against atherosclerosis.

Alpha-synuclein (αS) is a membrane-binding protein found predominantly in neurons and erythrocytes. The protein remains unordered in aqueous solutions but folds into an α-helical structure when bound to membranes. Besides, it gets deposited as β-sheet rich aggregates in diseases known as synucleinopathies. The native αS has been reported to be acetylated at the N-terminus. Here, we compare the interfacial properties of the N-terminal acetylated αS (Ac-αS) with non-acetylated αS (NH2-αS) at the air-water interface. Both the protein forms are highly surface-active, with surface pressure reaching up to ~30 mN/m upon compression. The pressure-area isotherms obtained from the repeated compression-expansion cycles display large hysteresis suggesting self-assembly at higher surface pressures. The expansion isotherm is characterized by a rapid decrease in surface pressure followed by a slower transition phase starting around 15–17 mN/m. These data suggest that the compressed monolayer breaks into small clusters upon expansion, followed by these clusters' loosening. The circular dichroism spectroscopic analysis of the Blodgett-deposited films suggests the protein to be in largely α-helical conformation. The linear dichroism investigations suggest the protein to be anisotropically deposited. Blodgett deposition of the Langmuir films, therefore, is a rather simple method for preparing oriented monolayers of surface-active macromolecules.

In recent years, chitosan has attracted considerable interest in many fields due to its sufficient charge density under biological, non-hazardous conditions. Since chitosan originates from natural resources and has two different monomer units, its characterization must be carried out in a goal-oriented and precise manner. This work focuses on the characterization of chitosans most important parameters – solubility, crystallinity, degree of deacetylation (DD) and molecular weight – in a simple and convenient way. The DD was determined using Nuclear Magnetic Resonance spectroscopy (NMR), Particle Charge Detection (PCD), Fourier Transform Infrared spectroscopy (FTIR), CHN elemental analysis (CHN-EA) and conductometric/potentiometric titration with special attention to its physical state as solid or liquid. Investigation of DD by FTIR was successfully determined by calculating peak heights, peak areas and peak deconvolution from a linear combination of Gaussian and Lorentzian functions. Asymmetrical flow field flow fractionation with light scattering detection (AF4-LS) was applied in order to calculate molar masses and radii. In addition, pH-potentiometric titrations demonstrated a reproducible displacement of the point of zero charge (PZC) in form of a hysteresis depending on the titration direction. The DD affects the crystallinity, which was determined by deconvolution of the crystalline and amorphous domains.

Acetylsalicylic acid (ASA) and type 2 diabetes mellitus (T2DM) affect fibrin clot properties through fibrinogen acetylation or glycation. We aimed to identify glycation and acetylation sites on fibrinogen in plasma fibrin clot of T2DM patients with respect to effects of ASA and fibrin clot properties. In fibrin clots generated from plasma of 9 T2DM patients, we performed mass-spectrometric analysis of Ne-fructosyl-(FL), Ne-carboxyethyl-(CEL) and Ne-carboxymethyl-lysine (CML), and acetylation sites, before and after one-month administration of 75 mg/d ASA confirmed with determination of thromboxane B2 concentration (TXB2), along with clot permeability and lysis time, and thrombin generation. In the proteomic analysis, 216 proteins were identified. Among 10 glycation sites identified in α, 10 in β and 6 in γ fibrinogen chain, there were 17 FL, 5 CEL and 4 CML sites. Some of glycation sites in fibrinogen were previously reported to be involved in cross-linking by factor XIII (αK-208, αK-448 and αK-539) and plasmin cleavage (αK-81). There were 7 acetylation sites in α and β chains, and none in fibrinogen γ chain. Two acetylation sites were identical with FL sites (αK-195 and β-247), while one with CML site (βK-353). In 7 patients with low post-ASA TXB2, intensity of acetylation, as well as clot properties were unaffected by ASA. This study identifies glycation and acetylation sites on fibrinogen in plasma fibrin clot of T2DM and supports the view that low-dose ASA does not increase fibrinogen acetylation in T2DM. Our findings suggest that glycation may block sites previously identified to be acetylated in vitro.

The aim of this study was to investigate the primary structure of an acetylated Cyclocarya paliurus polysaccharide (Ac-CPP0.1) and its protective effect on H2O2-treated dendritic cells. The backbone of Ac-CPP0.1 was →3)-β-D-Galp-(1→, with some branches α-L-Araf-(1→ residues at O-6 and O-5, β-D-Galp-(1→ and 3,5,6)-β-D-Galf-(1 residues at O-4 and acetyl groups were substituted at the O-2 and O-6 positions of 3)-β-D-Galp-(1 residues. The CPP0.1 and Ac-CPP0.1 significantly increased the levels of superoxide dismutase, glutathione peroxidase and catalase on H2O2-treated dendritic cells. Meanwhile, both CPP0.1 and Ac-CPP0.1 up-regulated the expression of Nrf2 (NF-E2-related factor 2) and down-regulated the Keap1 (Kelch-like ECH-associated protein-1), but Ac-CPP0.1 had a better effect on antioxidant capacity. These results indicated that potential application of Ac-CPP0.1 as an antioxidant agent.