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Nature Biotechnology Jun 2022High-redox-potential reactive oxygen species and reactive nitrogen species (ROS/RNS), generated by NADPH oxidase-2 (NOX2), myeloperoxidase (MPO) and related enzymes, are...
High-redox-potential reactive oxygen species and reactive nitrogen species (ROS/RNS), generated by NADPH oxidase-2 (NOX2), myeloperoxidase (MPO) and related enzymes, are key effector molecules of innate immunity. High-redox-potential radicals are difficult to distinguish by imaging from less potent ROS/RNS functioning as background biological signaling molecules. Here we present 4-[F]fluoro-1-naphthol ([F]4FN), a redox-tuned radiopharmaceutical that selectively binds proteins and cells when oxidized by products of human MPO plus HO, but not HO alone, and can be detected using positron emission tomography (PET). Activating HL-60 neutrophil-like human cells with phorbol ester (PMA) caused [F]4FN retention five-fold over unstimulated cells. An MPO-specific inhibitor (4-ABAH) blocked cellular retention by more than 95%. [F]4FN PET/CT imaging discriminated inflammatory foci in vivo in three murine models of activated innate immunity: endotoxin-induced toxic shock, PMA-induced contact dermatitis and lipopolysaccharide-induced ankle arthritis. 4-ABAH and Cybb (Nox2) gene deletion strongly abrogated [F]4FN retention in vivo. Thus, [F]4FN shows promise as a robust reporter of innate immunity activation by PET/CT.
Topics: Animals; Humans; Hydrogen Peroxide; Immunity, Innate; Mice; NADPH Oxidases; Oxidation-Reduction; Positron Emission Tomography Computed Tomography; Reactive Oxygen Species
PubMed: 35190688
DOI: 10.1038/s41587-021-01169-y -
Veterinary Research Communications Sep 2023Physic nut Jatropha curcas cake/meal obtained after oil extraction has a high protein content, however, the presence of antinutrients (trypsin inhibitor, lectin and...
Physic nut Jatropha curcas cake/meal obtained after oil extraction has a high protein content, however, the presence of antinutrients (trypsin inhibitor, lectin and phytate) and toxic compounds (phorbol esters) limit their use as an alternative feedstuff. Thus, the detoxification process in cake/meal is necessary to allow their inclusion in fish diets. The present study aimed to evaluate the effects of solvent and extrusion-treated jatropha cake (SETJC) in Nile tilapia (Oreochromis niloticus) diets on growth, body composition, nutrient utilization, metabolic and hematological responses, and digestibility of experimental diets. Five experimental diets were formulated to be isonitrogenous (28.50% digestible protein) and isoenergetic (13.39 MJ/kg digestible energy) with graded levels of SETJC (0, 3, 6, 9, and 12%). The experimental design was completely randomized with five treatments and four replicates. The detoxification treatments reduced the phorbol esters (PE) of jatropha cake by 96% (0.58 mg/g of PE before and 0.023 mg/g of PE after treatments). Increased levels of SETJC depressed growth, feed efficiency, and protein digestibility. A similar trend was observed for hematological and biochemistry parameters. Aspartate and alanine aminotransferase, as well as phosphorus and magnesium concentrations in the fillets, increased at the highest levels of SETJC. Thus, the data of the present study suggests that the residual content, different structural forms of phorbol ester and its biological activity, as well as some antinutritional factors, can influence negatively the growth, metabolism and digestibility of experimental diets for Nile tilapia.
Topics: Animals; Cichlids; Jatropha; Animal Feed; Solvents; Diet; Phorbol Esters; Seeds
PubMed: 36729277
DOI: 10.1007/s11259-023-10076-3 -
Photochemistry and Photobiology Jan 2022Microvesicle particles (MVP) are bioactive subcellular particles which have been recently implicated in the keratinocyte response to many environmental stressors...
Microvesicle particles (MVP) are bioactive subcellular particles which have been recently implicated in the keratinocyte response to many environmental stressors including ultraviolet B radiation (UVB). Previous studies have demonstrated that UVB generates high levels of MVP in a process involving the platelet-activating factor receptor (PAFR) and the enzyme acid sphingomyelinase (aSMase). Yet the fluences of UVB needed to generate MVP are usually above those commonly encountered. Using models including human epithelial cell lines in vitro, human skin explants ex vivo and murine studies in vivo, the present studies indicate that pretreatment of epithelial cells/skin with PAFR agonist/phorbol ester can synergize with low fluences of UVB to generate high levels of MVP. These studies indicate the possibility that MVP could play a role in combinatorial pathologic processes involving UVB.
Topics: Animals; Cell Line; Cell-Derived Microparticles; Humans; Keratinocytes; Mice; Skin; Ultraviolet Rays
PubMed: 34324709
DOI: 10.1111/php.13495 -
Experimental and Therapeutic Medicine Jan 2020The aim of the present study was to explore the possible role of microRNA-144-5p (miR-144-5p) in rheumatoid arthritis (RA) and the associated mechanism. Following the...
The aim of the present study was to explore the possible role of microRNA-144-5p (miR-144-5p) in rheumatoid arthritis (RA) and the associated mechanism. Following the induction of THP-1 cell differentiation into macrophages by phorbol ester (100 ng/ml) treatment, an inflammatory model of RA was established by treating the THP-1 macrophages with 1 µg/ml lipopolysaccharide (LPS). The level of miR-144-5p was subsequently measured using reverse transcription-quantitative PCR, which found that the expression of miR-144-5p was significantly reduced in LPS-treated THP-1 macrophages. Bioinformatics analysis and a dual-luciferase reporter assay were used to predict and confirm TLR2 as a direct target of miR-144-5p, respectively. Toll-like receptor 2 (TLR2) was then validated as a target gene of miR-144-5p. The effects of miR-144-5p upregulation and TLR2 silencing on LPS-treated THP-1 macrophages were then determined by transfection with miR-144-5p mimic and TLR2-siRNA, respectively. Cell viability was subsequently measured using a Cell Counting Kit-8 assay, whilst the expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and IL-8 secreted by THP-1 macrophages was measured using ELISA. Western blotting was performed to measure p65 phosphorylation (p-p65) in the NK-κB signaling pathway. It was found that miR-144-5p overexpression reduced macrophage cell viability, reduced the expression of TNF-α, IL-6 and IL-8, and reduced the expression of TLR2 and p-p65 compared with the control group. Likewise, TLR2 silencing also reduced macrophage cell viability and reduced the expression of TNF-α, IL-6 and IL-8 in THP-1 macrophages. In conclusion, the data from the present study suggested that miR-144-5p overexpression reduced THP-1 macrophage cell viability and inhibited the expression of TNF-α, IL-6 and IL-8 in cells, possibly by inhibiting the expression of TLR2 and suppressing the activation of NK-κB signaling. Therefore, miR-144-5p may serve as a novel therapeutic target for the treatment of RA.
PubMed: 31853295
DOI: 10.3892/etm.2019.8218 -
Journal of Biomolecular Structure &... 2022Munc13-1 is a presynaptic active zone protein that plays a critical role in priming the synaptic vesicle and releasing neurotransmitters in the brain. Munc13-1 acts as a...
Munc13-1 is a presynaptic active zone protein that plays a critical role in priming the synaptic vesicle and releasing neurotransmitters in the brain. Munc13-1 acts as a scaffold and is activated when diacylglycerol (DAG)/phorbol ester binds to its C1 domain in the plasma membrane. Our previous studies showed that bryostatin 1 activated the Munc13-1, but resveratrol inhibited the phorbol ester-induced Munc13-1 activity. To gain structural insights into the binding of the ligand into Munc13-1 C1 in the membrane, we conducted 1.0 μs molecular dynamics (MD) simulation on Munc13-1 C1-ligand-lipid ternary system using phorbol 13-acetate, bryostatin 1 and resveratrol as ligands. Munc13-1 C1 shows higher conformational stability and less mobility along membrane with phorbol 13-acetate and bryostatin 1 than with resveratrol. Bryostatin 1 and phorbol ester remained in the protein active site, but resveratrol moved out of Munc13-1 C1 during the MD simulation. While bryostatin 1-bound Munc13-1 C1 showed two different positioning in the membrane, phorbol 13-acetate and resveratrol-bound Munc13-1 C1 only showed one positioning. Phorbol 13-acetate formed hydrogen bond with Ala-574 and Gly-589. Bryostatin 1 had more hydrogen bonds with Trp-588 and Arg-592 than with other residues. Resveratrol formed hydrogen bond with Ile-590. This study suggests that different ligands control Munc13-1 C1's mobility and positioning in the membrane differently. Ligand also has a critical role in the interaction between Munc13-1 C1 and lipid membrane. Our results provide structural basis of the pharmacological activity of the ligands and highlight the importance of membrane in Munc13-1 activity.Communicated by Ramaswamy H. Sarma.
Topics: Molecular Dynamics Simulation; Ligands; Resveratrol; Phorbol Esters; Lipids
PubMed: 34779746
DOI: 10.1080/07391102.2021.2001375 -
Scientific Reports Feb 2023Macrophages (MQs) pro-inflammatory phenotype is triggered by gliadin peptides. Akkermansia muciniphila (A. muciniphila) showed to enhance the anti-inflammatory phenotype...
Macrophages (MQs) pro-inflammatory phenotype is triggered by gliadin peptides. Akkermansia muciniphila (A. muciniphila) showed to enhance the anti-inflammatory phenotype of MQs. This study aimed to investigate the anti-inflammatory effects of A. muciniphila, on gliadin stimulated THP-1 derived macrophages. THP-1 cell line monocytes were differentiated into MQs by phorbol 12-myristate 13-acetate (PMA). MQs were treated with A. muciniphila before and after stimulation with gliadin (pre- and post-treat). CD11b, as a marker of macrophage differentiation, and CD206 and CD80, as M1 and M2 markers, were evaluated by flow cytometry technique. The mRNA expression of TGF-β, IL-6, and IL-10 and protein levels of IL-10 and TNF-α were measured by RT-PCR and ELISA techniques, respectively. Results show an increased percentage of M1 phenotype and release of proinflammatory cytokines (like TNF-α and IL-6) by macrophages upon incubation with gliadin. Pre- and post-treatment of gliadin-stimulated macrophages with A. muciniphila induced M2 phenotype associated with decreased proinflammatory (IL-6, TNF-α) and increased anti-inflammatory (IL-10, TGF-β) cytokines expression relative to the group that was treated with gliadin alone. This study suggests the potential beneficial effect of A. muciniphila on gliadin-stimulated MQs and the importance of future studies focusing on their exact mechanism of action on these cells.
Topics: Interleukin-10; Gliadin; Tumor Necrosis Factor-alpha; Interleukin-6; Macrophages; Cytokines; Anti-Inflammatory Agents; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta
PubMed: 36828897
DOI: 10.1038/s41598-023-30266-y -
Inflammopharmacology Oct 2020Metformin, a potent AMPK activator is the most commonly used drug for diabetes. According to recent reports, metformin lowers the risk of diabetic complications and...
Metformin, a potent AMPK activator is the most commonly used drug for diabetes. According to recent reports, metformin lowers the risk of diabetic complications and inflammatory diseases. We found the expression levels of AMPK subunits including PRKAA1, PRKAA2, PRKAB1 and PRKAB2 are decreased in skin biopsies of dermatitis patients from multiple datasets. Interestingly, metformin treatment ameliorates dermatitis symptom in animal model of dermatitis using O-tetradecanoylphorbol-13-acetate (TPA). Especially, the levels of epidermis and dermis thickness were decreased by metformin. We found NFκB activity as well as of gene expression associated with collagen synthesis are attenuated by metformin treatment. These results suggest that metformin treatment alleviates animal model of dermatitis.
Topics: AMP-Activated Protein Kinases; Animals; Collagen; Dermatitis; Disease Models, Animal; Enzyme Activators; Gene Expression Regulation; Humans; Male; Metformin; Mice, Inbred C57BL; NF-kappa B; Skin; Tetradecanoylphorbol Acetate
PubMed: 32347398
DOI: 10.1007/s10787-020-00704-8 -
Psychoneuroendocrinology Feb 2021This study examined whether early life adversity (ELA) limited to infancy was associated with an increase in circulating levels of proinflammatory cytokines and cellular...
This study examined whether early life adversity (ELA) limited to infancy was associated with an increase in circulating levels of proinflammatory cytokines and cellular cytokine responses to three stimulants [lipopolysaccharide (LPS), phytohemagglutinin (PHA), and phorbol myristate acetate plus ionomycin (PMA/IO)]. Participants were previously institutionalized (PI) youth (N = 45, 56 % female) who had spent their first years in institutional care (e.g., orphanages, baby homes) before being adopted into well-resourced homes (median age at adoption = 13 mos) and non-adopted comparisons (NA; N = 38, 55 % female). Their age range was 13.3-21.2 years (M = 16.3 years). This analysis followed up an earlier report on these youth (Reid et al., 2019a) that identified an increase in terminally differentiated CD8 + CD57 T cells among the PI relative to the NA youth. Cytokine levels in circulation were not highly correlated and thus examined separately. PI youth had higher circulating levels of Tumor Necrosis Factor-alpha (TNFα), but not Interleukin-1β (IL-1β) or Interleukin-6 (IL-6). Cytokine responses to in vitro activation within each stimulant condition were highly correlated and were thus combined to generate an index of the inflammatory reaction to each stimulant. Using Multivariate Analysis of Covariance, there was a highly significant multivariate effect of group, which was carried primarily by the PMA/IO condition, with PI youth exhibiting a larger inflammatory response than NA youth. Tests of mediation showed that both the early rearing effects on circulating TNFα and the composite inflammatory index of PMA/IO responsiveness were mediated in the statistical model by the percentage of CD8 + CD57+ TEMRA cells in circulation, a marker of replicative senescence in T cells. Sex differences were also found in circulating levels of IL-6 and TNFα, with males having higher levels than females.
Topics: Adolescent; Adult; Cytokines; Female; Humans; Infant; Interleukin-6; Male; Orphanages; T-Lymphocytes; Tetradecanoylphorbol Acetate; Tumor Necrosis Factor-alpha; Young Adult
PubMed: 33278786
DOI: 10.1016/j.psyneuen.2020.105065 -
Bulletin of Experimental Biology and... Apr 2021We analyzed functional status of blood leukocytes in diabetes mellitus and after addition of glucose in vitro. To this end, generation of ROS and reactive halogen...
We analyzed functional status of blood leukocytes in diabetes mellitus and after addition of glucose in vitro. To this end, generation of ROS and reactive halogen species by monocytes and neutrophils from patients with diabetes mellitus and healthy donors was assayed using lucigenin- and luminol-dependent chemiluminescence after stimulation with phorbol 12-myristate 13-acetate or opsonized zymosan in vitro. Formation of neutrophil extracellular traps was evaluated in the blood after addition of glucose. In comparison with donors, leukocytes from patients with diabetes mellitus were primed and this effect can be modeled by addition of glucose to the blood in vitro. Addition of glucose to donor blood also triggered the formation of neutrophil extracellular traps.
Topics: Diabetes Mellitus; Extracellular Traps; Humans; Hyperglycemia; Leukocytes; Luminescent Measurements; Neutrophils; Reactive Oxygen Species; Tetradecanoylphorbol Acetate; Zymosan
PubMed: 33893954
DOI: 10.1007/s10517-021-05147-x -
Frontiers in Cell and Developmental... 2022Protein kinase C (PKC) isozymes transduce myriad signals within the cell in response to the generation of second messengers from membrane phospholipids. The conventional... (Review)
Review
Protein kinase C (PKC) isozymes transduce myriad signals within the cell in response to the generation of second messengers from membrane phospholipids. The conventional isozyme PKCγ reversibly binds Ca and diacylglycerol, which leads to an open, active conformation. PKCγ expression is typically restricted to neurons, but evidence for its expression in certain cancers has emerged. PKC isozymes have been labeled as oncogenes since the discovery that they bind tumor-promoting phorbol esters, however, studies of cancer-associated PKC mutations and clinical trial data showing that PKC inhibitors have worsened patient survival have reframed PKC as a tumor suppressor. Aberrant expression of PKCγ in certain cancers suggests a role outside the brain, although whether PKCγ also acts as a tumor suppressor remains to be established. On the other hand, PKCγ variants associated with spinocerebellar ataxia type 14 (SCA14), a neurodegenerative disorder characterized by Purkinje cell degeneration, enhance basal activity while preventing phorbol ester-mediated degradation. Although the basis for SCA14 Purkinje cell degeneration remains unknown, studies have revealed how altered PKCγ activity rewires cerebellar signaling to drive SCA14. Importantly, enhanced basal activity of SCA14-associated mutants inversely correlates with age of onset, supporting that enhanced PKCγ activity drives SCA14. Thus, PKCγ activity should likely be inhibited in SCA14, whereas restoring PKC activity should be the goal in cancer therapies. This review describes how PKCγ activity can be lost or gained in disease and the overarching need for a PKC structure as a powerful tool to predict the effect of PKCγ mutations in disease.
PubMed: 35800893
DOI: 10.3389/fcell.2022.929510