scholarly journals Sustained Immunoparalysis in Endotoxin-Tolerized Monocytic Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Christina K. Weisheit ◽  
Alexandra Klüners ◽  
Lennart Wild ◽  
Alexandra Casalter ◽  
Stefanie Heilmann-Heimbach ◽  
...  
Keyword(s):  
Metabolic Pathways ◽  
Epigenetic Modification ◽  
Cellular Metabolism ◽  
Epigenetic Modifications ◽  
Lps Challenge ◽  
Monocyte Function ◽  
Immune Paralysis ◽  
Culture Models ◽  
Underlying Mechanisms

Sepsis is associated with a strong inflammatory reaction triggering a complex and prolonged immune response. Septic patients have been shown to develop sustained immunosuppression due to a reduced responsiveness of leukocytes to pathogens. Changes in cellular metabolism of leukocytes have been linked to this phenomenon and contribute to the ongoing immunological derangement. However, the underlying mechanisms of these phenomena are incompletely understood. In cell culture models, we mimicked LPS tolerance conditions to provide evidence that epigenetic modifications account for monocyte metabolic changes which cause immune paralysis in restimulated septic monocytes. In detail, we observed differential methylation of CpG sites related to metabolic activity in human PBMCs 18 h after septic challenge. The examination of changes in immune function and metabolic pathways was performed in LPS-tolerized monocytic THP-1 cells. Passaged THP-1 cells, inheriting initial LPS challenge, presented with dysregulation of cytokine expression and oxygen consumption for up to 7 days after the initial LPS treatment. Proinflammatory cytokine concentrations of TNFα and IL1β were significantly suppressed following a second LPS challenge (p<0.001) on day 7 after first LPS stimulation. However, the analysis of cellular metabolism did not reveal any noteworthy alterations between tolerant and nontolerant THP-1 monocytes. No quantitative differences in ATP and NADH synthesis or participating enzymes of energy metabolism occurred. Our data demonstrate that the function and epigenetic modifications of septic and tolerized monocytes can be examined in vitro with the help of our LPS model. Changes in CpG site methylation and monocyte function point to a correlation between epigenetic modification in metabolic pathways and reduced monocyte function under postseptic conditions.

scholarly journals Two-dimensional electrophoresis protein profiles of HL-60 and CCRF-CEM cell lines treated with epigenetic modification drugs

2019 ◽  
pp. 10-23
Author(s):  
Aziee Sudin ◽  
Haiyuni Mohd Yassim ◽  
Shafini Mohamed Yusoff ◽  
Shaharum Shamsuddin ◽  
Ridhwan Abdul Wahab ◽  
...  
Keyword(s):  
Cell Lines ◽  
Dna Methyltransferase ◽  
Epigenetic Modification ◽  
Trichostatin A ◽  
Lymphoblastic Leukemia ◽  
Genetic Alterations ◽  
Epigenetic Modifications ◽  
Hematopoietic Stem ◽  
2D Gel

Leukemia is classified as a malignant disease of hematopoietic stem cells (HSCs) that fails in cell differentiation but preserve their self-renewal. It is caused by genetic alterations and epigenetic modifications resulting in the activation or inactivation of particular genes for transcription. Epigenetic causes changes in gene expression without any alteration in the DNA sequence. The most common epigenetic modifications are DNA methylation and histone acetylation. 5-Azacitidine (5-Aza) is a DNA methytransferase inhibitor (DNMTi) that inhibits DNA methyltransferase enzymes resulting in hypomethylation. Trichostatin A (TSA) is a histone deacetylase inhibitor which inhibits deacetylation of both histone and non-histone proteins resulting in chromatin relaxation. This present study focused on the alteration of proteome profile on 2D gel electrophoresis (2-DE) induced by 5-Aza and TSA in HL-60 and CCRF-CEM cell lines as in vitro model to represent acute promyelocytic leukemia (APL) and T-lymphoblastic leukemia (T-ALL), respectively. Total proteins of untreated and 5-Aza/TSA-treated HL-60 and CCRF-CEM cell lines were extracted using urea/thiourea buffer and stained with Coomassie Blue. Comparative analysis of untreated and 5-Aza/TSA-treated HL-60 and CCRF-CEM was performed by PDQuest software. Qualitative analysis identified 190-659 protein spots detected in untreated, 5-Aza and TSA-treated HL-60 and CCRF-CEM. Quantitative comparison analysis was analyzed by over 2-fold change in 5-Aza/TSA-treated cells compared to untreated. One and eight upregulated proteins were detected in 5-Aza and TSA-treated HL-60, respectively. While five and one upregulated proteins were detected in 5-Aza and TSA-treated CCRF-CEM, respectively. These preliminary results suggested that 5-Aza and TSA induced proteome profiles alterations due to their inhibition effects in HL-60 and CCRF-CEM cell lines.

Rosuvastatin corrects oxidative stress and inflammation induced by LPS to attenuate cardiac injury by inhibiting the NLRP3/TLR4 pathway

2021 ◽  
Author(s):  
Guocheng Ren ◽  
Qiujie Zhou ◽  
Meili Lu ◽  
Hongxin Wang
Keyword(s):  
Oxidative Stress ◽  
Inflammatory Response ◽  
Cardiac Injury ◽  
H9c2 Cells ◽  
Sod Activity ◽  
Lps Challenge ◽  
Pathway Activation ◽  
Underlying Mechanisms

The aim of the current study was to evaluate whether rosuvastatin was effective in attenuating cardiac injury in lipopolysaccharide(LPS)-challenged mice and H9C2 cells and identify the underlying mechanisms, focusing on the NLRP3/TLR4 pathway. Cardiac injury, cardiac function, apoptosis, oxidative stress, inflammatory response and the NLRP3/TLR4 pathway were evaluated in both in vivo and in vitro studies. LPS-induced cardiomyocytes injury was markedly attenuated by rosuvastatin treatment. Apoptosis was clearly ameliorated in myocardial tissue and H9C2 cells cotreated with rosuvastatin. In addition, excessive oxidative stress was present, as indicated by increases in MDA content, NADPH activity and ROS production and decreased SOD activity after LPS challenge. Rosuvastatin improved all the indicators of oxidative stress, with a similar effect to NAC(ROS scavenger). Notably, LPS-exposed H9C2 cells and mice showed significant NLRP3 and TLR4/NF-κB pathway activation. Administration of rosuvastatin reduced the increases in expression of NLRP3, ASC, pro-caspase-1, TLR4, and p65 and decreased the contents of TNF-α, IL-1β, IL-18 and IL-6, with a similar effect as MCC950 (NLRP3 inhibitor). In conclusion, inhibition of the inflammatory response and oxidative stress contributes to cardioprotection of rosuvastatin on cardiac injury induced by LPS, and the effect of rosuvastatin was achieved by inactivation of the NF-κB/NLRP3 pathway

scholarly journals Sirtuin Lipoamidase Activity Is Conserved in Bacteria as a Regulator of Metabolic Enzyme Complexes

mBio ◽  
2017 ◽  
Vol 8 (5) ◽  
Author(s):  
Elizabeth A. Rowland ◽  
Todd M. Greco ◽  
Caroline K. Snowden ◽  
Anne L. McCabe ◽  
Thomas J. Silhavy ◽  
...  
Keyword(s):  
Metabolic Pathways ◽  
Tricarboxylic Acid Cycle ◽  
Cellular Metabolism ◽  
Gram Negative Bacteria ◽  
Gram Positive ◽  
E Coli ◽  
Metabolism Regulation ◽  
Entry Points ◽  
Enzyme Complexes

ABSTRACT Lipoic acid is an essential metabolic cofactor added as a posttranslational modification on several multimeric enzyme complexes. These protein complexes, evolutionarily conserved from bacteria to humans, are core regulators of cellular metabolism. While the multistep enzymatic process of adding lipoyl modifications has been well characterized in Escherichia coli, the enzyme required for the removal of these lipoyl moieties (i.e., a lipoamidase or delipoylase) has not yet been identified. Here, we describe our discovery of sirtuins as lipoamidases in bacteria and establish their conserved substrates. Specifically, by using a series of knockout, overexpression, biochemical, in vitro, proteomic, and functional assays, we determined the substrates of sirtuin CobB in E. coli as components of the pyruvate dehydrogenase (PDH), α-ketoglutarate dehydrogenase (KDH), and glycine cleavage (GCV) complexes. In vitro assays provided direct evidence for this specific CobB activity and its NAD+ dependence, a signature of all sirtuins. By designing a targeted quantitative mass spectrometry method, we further measured sirtuin-dependent, site-specific lipoylation on these substrates. The biological significance of CobB-modulated lipoylation was next established by its inhibition of both PDH and KDH activities. By restricting the carbon sources available to E. coli, we demonstrated that CobB regulates PDH and KDH under several growth conditions. Additionally, we found that SrtN, the sirtuin homolog in Gram-positive Bacillus subtilis, can also act as a lipoamidase. By demonstrating the evolutionary conservation of lipoamidase activity across sirtuin homologs, along with the conservation of common substrates, this work emphasizes the significance of protein lipoylation in regulating central metabolic processes. IMPORTANCE Here, we demonstrate that sirtuin lipoamidase activity exists in both Gram-positive and Gram-negative bacteria and establishing its conservation from bacteria to humans. Specifically, we discovered that CobB and SrtN act as lipoamidases in E. coli and B. subtilis, respectively. Intriguingly, not only is this sirtuin enzymatic activity conserved, but also the lipoylated substrates and functions are conserved, as bacterial sirtuins negatively regulate the lipoylation levels and activities of PDH and KDH. Considering that PDH and KDH regulate two carbon entry points into the tricarboxylic acid cycle, our finding highlights lipoylation as a conserved molecular toggle that regulates central metabolic pathways. Indeed, our findings from tests in which we limited nutrient availability support this. Furthermore, this study illustrates how the integration of technologies from different disciplines provides avenues to uncover enzymatic activities at the core of cellular metabolism regulation. IMPORTANCE Here, we demonstrate that sirtuin lipoamidase activity exists in both Gram-positive and Gram-negative bacteria and establishing its conservation from bacteria to humans. Specifically, we discovered that CobB and SrtN act as lipoamidases in E. coli and B. subtilis, respectively. Intriguingly, not only is this sirtuin enzymatic activity conserved, but also the lipoylated substrates and functions are conserved, as bacterial sirtuins negatively regulate the lipoylation levels and activities of PDH and KDH. Considering that PDH and KDH regulate two carbon entry points into the tricarboxylic acid cycle, our finding highlights lipoylation as a conserved molecular toggle that regulates central metabolic pathways. Indeed, our findings from tests in which we limited nutrient availability support this. Furthermore, this study illustrates how the integration of technologies from different disciplines provides avenues to uncover enzymatic activities at the core of cellular metabolism regulation.

scholarly journals Bigelovii A Protects against Lipopolysaccharide-Induced Acute Lung Injury by Blocking NF-κB and CCAAT/Enhancer-Binding ProteinδPathways

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Chunguang Yan ◽  
Fuqin Guan ◽  
Yanfei Shen ◽  
Huifang Tang ◽  
Dong Yuan ◽  
...  
Keyword(s):  
Acute Lung Injury ◽  
Lung Injury ◽  
P38 Mapk ◽  
Inflammatory Mediators ◽  
Neutrophil Infiltration ◽  
Therapeutic Approaches ◽  
Lps Challenge ◽  
Enhancer Binding Protein ◽  
Underlying Mechanisms

Optimal methods are applied to acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS), but the mortality rate is still high. Accordingly, further studies dedicated to identify novel therapeutic approaches to ALI are urgently needed. Bigelovii A is a new natural product and may exhibit anti-inflammatory activity. Therefore, we sought to investigate its effect on lipopolysaccharide- (LPS-) induced ALI and the underlying mechanisms. We found that LPS-induced ALI was significantly alleviated by Bigelovii A treatment, characterized by reduction of proinflammatory mediator production, neutrophil infiltration, and lung permeability. Furthermore, Bigelovii A also downregulated LPS-stimulated inflammatory mediator expressionsin vitro. Moreover, both NF-κB and CCAAT/enhancer-binding proteinδ(C/EBPδ) activation were obviously attenuated by Bigelovii A treatment. Additionally, phosphorylation of both p38 MAPK and ERK1/2 (upstream signals of C/EBPδactivation) in response to LPS challenge was also inhibited by Bigelovii A. Therefore, Bigelovii A could attenuate LPS-induced inflammation by suppression of NF-κB, inflammatory mediators, and p38 MAPK/ERK1/2—C/EBPδ, inflammatory mediators signaling pathways, which provide a novel theoretical basis for the possible application of Bigelovii A in clinic.

scholarly journals Garlic: Much More Than a Common Spice

Foods ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 1544
Author(s):  
Michael E. Netzel
Keyword(s):  
Antimicrobial Activities ◽  
Biological Properties ◽  
Organosulfur Compounds ◽  
Bioactive Components ◽  
Renal Protection ◽  
Biological Potential ◽  
Culture Models ◽  
Underlying Mechanisms ◽  
Cell Culture Models

Garlic is a widely consumed and popular spice with a characteristic “aroma” or odour. It contains a broad range of bioactive components such as organosulfur compounds, saponins and polyphenols, but can be also rich in vitamins and minerals. Numerous biological properties are attributed to garlic, from antimicrobial activities to neuro- and renal-protection. In addition, post-harvest treatment, storage and processing, such as fermentation and heat, can have a significant effect on garlic and its bioactive compounds, and subsequently alter its bioactive properties. Future studies are warranted to elucidate the “full” biological potential of garlic including well designed human clinical trials, detailed storage and processing studies as well as sophisticated in vitro cell culture models to better understand the underlying mechanisms of action.

scholarly journals Adenosine deaminase modulates metabolic remodeling and orchestrates joint destruction in rheumatoid arthritis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Sai Krishna Srimadh Bhagavatham ◽  
Prakash Khanchandani ◽  
Vishnu Kannan ◽  
Damodaram Potikuri ◽  
Divya Sridharan ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Adenosine Deaminase ◽  
Metabolic Pathways ◽  
Joint Destruction ◽  
Mediated Effects ◽  
Potential Marker ◽  
Metabolic Remodeling ◽  
Cytokine Levels ◽  
Culture Models

AbstractRheumatoid Arthritis (RA) is a chronic autoimmune disease associated with inflammation and joint remodeling. Adenosine deaminase (ADA), a risk factor in RA, degrades adenosine, an anti-inflammatory molecule, resulting in an inflammatory bias. We present an integrative analysis of clinical data, cytokines, serum metabolomics in RA patients and mechanistic studies on ADA-mediated effects on in vitro cell culture models. ADA activity differentiated patients into low and high ADA sets. The levels of the cytokines TNFα, IFNγ, IL-10, TGFβ and sRANKL were elevated in RA and more pronounced in high ADA sets. Serum metabolomic analysis shows altered metabolic pathways in RA which were distinct between low and high ADA sets. Comparative analysis with previous studies shows similar pathways are modulated by DMARDs and biologics. Random forest analysis distinguished RA from control by methyl-histidine and hydroxyisocaproic acid, while hexose-phosphate and fructose-6-phosphate distinguished high ADA from low ADA. The deregulated metabolic pathways of High ADA datasets significantly overlapped with high ADA expressing PBMCs GEO transcriptomics dataset. ADA induced the death of chondrocytes, synoviocyte proliferation, both inflammation in macrophages and their differentiation into osteoclasts and impaired differentiation of mesenchymal stem cells to osteoblasts and mineralization. PBMCs expressing elevated ADA had increased expression of cytokines and P2 receptors compared to synovial macrophages which has low expression of ADA. Our data demonstrates increased cytokine levels and distinct metabolic signatures of RA based on the ADA activity, suggests an important role for ADA in the pathophysiology of RA joints and as a potential marker and therapeutic target in RA patients.

Sub-clinical necrotizing enterocolitis-induced systemic immune suppression in neonatal preterm pigs

2021 ◽  
Author(s):  
Shuqiang Ren ◽  
Xiaoyu Pan ◽  
Yan Hui ◽  
Witold Kot ◽  
Fei Gao ◽  
...  
Keyword(s):  
Immune Responses ◽  
Preterm Infants ◽  
Necrotizing Enterocolitis ◽  
Immune Suppression ◽  
High Sensitivity ◽  
T Cell Subsets ◽  
Distal Small Intestine ◽  
Lps Challenge ◽  
Underlying Mechanisms

Preterm infants are at high risks of sepsis and necrotizing enterocolitis (NEC). Some develop sepsis shortly after suspected or confirmed NEC, implying that NEC may predispose to sepsis but the underlying mechanisms are unknown. Using NEC-sensitive preterm pigs as models, we investigated the immune status in animals following development of sub-clinical NEC-like lesions with variable severities. Caesarean-delivered preterm pigs were reared until day 5 or 9. Blood was analyzed for T cell subsets, neutrophil phagocytosis, transcriptomics and immune responses to in vitro LPS challenge. Gut tissues were used for histology and cytokine analyses. Pigs with/without macroscopic NEC lesions were scored as healthy, mild or severe NEC. Overall NEC incidence was similar on days 5 and 9 (61-62%) but with lower severity on day 9, implying gradual mucosal repair following the early phase of NEC. Pigs with NEC showed decreased goblet cell density and increased MPO+ and CD3+ cell infiltration in the distal small intestine or colon. Mild or severe NEC lesions had limited effects on circulating parameters on day 5. On day 9, pigs with NEC lesions (especially severe lesions) showed systemic immune suppression, as indicated by elevated Treg frequency, impaired neutrophil phagocytosis, low expression of genes related to innate immunity and Th1 polarization, and diminished LPS-induced immune responses. In conclusion, we shows evidence for NEC-induced systemic immune suppression, even with mild and sub-clinical NEC lesions. The results help to explain that preterm infants suffering from NEC may show high sensitivity to later secondary infections and sepsis.

scholarly journals Cadmium and Its Neurotoxic Effects

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Bo Wang ◽  
Yanli Du
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Prolonged Exposure ◽  
Epigenetic Modification ◽  
The Body ◽  
Functional Changes ◽  
Neurotoxic Effects ◽  
Underlying Mechanisms

Cadmium (Cd) is a heavy metal that has received considerable concern environmentally and occupationally. Cd has a long biological half-life mainly due to its low rate of excretion from the body. Thus, prolonged exposure to Cd will cause toxic effect due to its accumulation over time in a variety of tissues, including kidneys, liver, central nervous system (CNS), and peripheral neuronal systems. Cd can be uptaken from the nasal mucosa or olfactory pathways into the peripheral and central neurons; for the latter, Cd can increase the blood brain barrier (BBB) permeability. However, mechanisms underlying Cd neurotoxicity remain not completely understood. Effect of Cd neurotransmitter, oxidative damage, interaction with other metals such as cobalt and zinc, estrogen-like, effect and epigenetic modification may all be the underlying mechanisms. Here, we review thein vitroandin vivoevidence of neurotoxic effects of Cd. The available finding indicates the neurotoxic effects of Cd that was associated with both biochemical changes of the cell and functional changes of central nervous system, suggesting that neurotoxic effects may play a role in the systemic toxic effects of the exposure to Cd, particularly the long-term exposure.

84 Aging Model in Pig Oocytes as Viewed Through DNA Epigenetic Modification

2021 ◽  
Vol 99 (Supplement_1) ◽  
pp. 108-109
Author(s):  
Haley A Arena ◽  
Kimberly Sprungl ◽  
Skyla Reynolds ◽  
Brian D Whitaker
Keyword(s):  
In Vitro Maturation ◽  
In Vitro Model ◽  
Time Window ◽  
Epigenetic Modification ◽  
Trichostatin A ◽  
Epigenetic Modifications ◽  
Optimal Time ◽  
Vitro Model ◽  
Effects Of Aging

Abstract Oocytes of older animals are less likely to be fertilized during the optimal time window post ovulation, resulting in the potential diminished fertilization and embryonic development success. The activity of the epigenetic modifications during this period is a possible target to reverse these damaging effects of aging. The objective of this study was to study the effects of aging during in vitro oocyte maturation in pigs on epigenetic modifications. Oocytes (n = 54) were matured with or without Trichostatin A (TSA; 100 ng/mL), a known meiotic inhibitor, for 24 h, then for an additional 16 h without TSA or hormones for a total of 40 h. At the end of maturation, oocytes were denuded and their zona pellucida’s removed. Oocytes were stained with anti-5-methylcytosine (5mC, 1:500). Fluorescent images of the oocytes were acquired, images were analyzed using ImageJ, and data analysis was performed using ANOVA and Tukey’s test. Oocytes matured with TSA had significantly greater (P &lt; 0.05) levels of DNA methylation by the end of in vitro maturation compared to those not supplemented with TSA These results suggest that TSA can be used to develop an in vitro model to study the effects of epigenetic modifications in oocytes from aged livestock.

Phytosomal Curcumin Elicits Anti-tumor Properties Through Suppression of Angiogenesis, Cell Proliferation and Induction of Oxidative Stress in Colorectal Cancer

2019 ◽  
Vol 24 (39) ◽  
pp. 4626-4638 ◽  
Author(s):  
Reyhaneh Moradi-Marjaneh ◽  
Seyed M. Hassanian ◽  
Farzad Rahmani ◽  
Seyed H. Aghaee-Bakhtiari ◽  
Amir Avan ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Colorectal Cancer ◽  
Therapeutic Potential ◽  
Vegf Signaling ◽  
Anticancer Effects ◽  
Inhibition Of Angiogenesis ◽  
Underlying Mechanisms ◽  
Apoptotic Activity

Background: Colorectal cancer (CRC) is one of the most common causes of cancer-associated mortality in the world. Anti-tumor effect of curcumin has been shown in different cancers; however, the therapeutic potential of novel phytosomal curcumin, as well as the underlying molecular mechanism in CRC, has not yet been explored. Methods: The anti-proliferative, anti-migratory and apoptotic activity of phytosomal curcumin in CT26 cells was assessed by MTT assay, wound healing assay and Flow cytometry, respectively. Phytosomal curcumin was also tested for its in-vivo activity in a xenograft mouse model of CRC. In addition, oxidant/antioxidant activity was examined by DCFH-DA assay in vitro, measurement of malondialdehyde (MDA), Thiol and superoxidedismutase (SOD) and catalase (CAT) activity and also evaluation of expression levels of Nrf2 and GCLM by qRT-PCR in tumor tissues. In addition, the effect of phytosomal curcumin on angiogenesis was assessed by the measurement of VEGF-A and VEGFR-1 and VEGF signaling regulatory microRNAs (miRNAs) in tumor tissue. Results: Phytosomal curcumin exerts anti-proliferative, anti-migratory and apoptotic activity in-vitro. It also decreases tumor growth and augmented 5-fluorouracil (5-FU) anti-tumor effect in-vivo. In addition, our data showed that induction of oxidative stress and inhibition of angiogenesis through modulation of VEGF signaling regulatory miRNAs might be underlying mechanisms by which phytosomal curcumin exerted its antitumor effect. Conclusion: Our data confirmed this notion that phytosomal curcumin administrates anticancer effects and can be used as a complementary treatment in clinical settings.

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