Beta Secretion Research Articles

Molecular Mapping of Hydrogen Sulfide Targets in Normal Human Keratinocytes.

Although sulfur-rich thermal waters have ancestrally been used in the context of dermatological conditions, a global mapping of the molecular effects exerted by H2S on human keratinocytes is still lacking. To fill this knowledge gap, we subjected cultured human keratinocytes to distinct amounts of the non-gaseous hydrogen sulfur donor NaHS. We first checked that H2S accumulated in the cytoplasm of keratinocytes under our experimental conditions andused a combination of proteomics, genomics and biochemical approaches to unravel functionally relevant H2S targets in human keratinocytes. We found that the identified targets fall into two main categories: (i) the oxidative stress response molecules superoxide dismutase 2 (SOD2), NAD(P)H quinone dehydrogenase 1 (NQO1) and culin 3 (CUL3) and (ii) the chemokines interleukin-8 (IL-8) and CXCL2. Interestingly, NaHS also stimulated the caspase-1 inflammasome pathway, leading to increased secretion of the pro-inflammatory molecule interleukin-18 (IL-18). Interestingly, the secretion of interleukin-1 beta (IL-1β) was only modestly impacted by NaHS exposure despite a significant accumulation of IL-1β pro-form. Finally, we observed that NaHS significantly hampered the growth of human keratinocyte progenitors and stem cells cultured under clonogenic conditions or as epidermal cell sheets. We conclude that H2S exerts specific molecular effects on normal human keratinocytes.

Antibacterial and Antifungal Agent, Triclosan (TCS) Alters Secretion of Interleukin beta and Interleukin 6 from Human Immune Cells

Triclosan (TCS) is an antimicrobial compound that is widely used in personal hygiene products such as mouthwash and toothpaste. TCS can be ingested or absorbed through the skin and has been found in human blood, breast milk, and urine. Interleukin (IL)‐6 and IL‐1 beta (IL‐1β) are important pro‐inflammatory cytokines produced by lymphocytes, monocytes, and other cells. Both regulate cell growth, tissue repair, and immune function, increased levels of each have been associated with a number of diseases including rheumatoid arthritis and certain cancers. TCS has been shown to inhibit the lytic function of human natural killer (NK) lymphocytes and to decrease expression of key cell surface proteins on NK cells. Here we examine whether TCS alters the secretion of IL‐1β and IL‐6 from human immune cell preparations. Human peripheral blood mononuclear cells (PBMCs) and monocyte‐depleted (MD)‐PBMCs were exposed to TCS at concentrations of 0–5 μM. Cytokine secretion was measured at 24 h, 48 h, and 6 days using enzyme‐linked immunosorbent assay (ELISA). In this study, both IL‐1β and IL‐6 secretion were altered at one or more concentration of TCS at one or more length of exposure. IL‐1β secretion was increased at 1, 2.5 or 5 μM TCS. Effects of 24 h exposure to 1, 2.5 and 5 μM TCS on IL‐1β secretion from PBMCs treated with selective MAPK pathway component inhibitors were also measured. These results indicate that TCS has the capacity to disrupt secretion of these two important pro‐inflammatory cytokines. Additionally the data show that TCS‐induced increases in IL‐β secretion were dependent on p38 MAPK.Support or Funding InformationSupported by NIH grant U54CA163066.

Deficiency of TREK-1 potassium channel exacerbates blood-brain barrier damage and neuroinflammation after intracerebral hemorrhage in mice

BackgroundIntracerebral hemorrhage (ICH) is a devastating medical emergency with high mortality and severe neurological deficit. ICH-related poor outcomes are due to a combination of pathological processes that could be complicated by secondary insults. TWIK-related K+ channel 1 (TREK-1) is a two-pore-domain potassium channel that is highly expressed in the mammalian nervous system. Previous studies have shown that TREK-1 channels play important roles in various central nervous system diseases. However, its role in the secondary injuries after intracerebral hemorrhage remains unknown. In this study, we explored the function of TREK-1 in secondary blood-brain barrier injuries and neuroinflammation after intracerebral hemorrhage in mice.MethodsAdult male TREK-1−/− mice and WT mice were subjected to a collagenase-induced ICH model. Immunostaining, western blot, and enzyme-linked immunosorbent assay were used to assess inflammatory infiltration and neuronal death. Blood-brain barrier compromise was assessed using electron microscopy and Evans Blue dye injection on days 1 and 3 after intracerebral hemorrhage. Magnetic resonance imaging and behavioral assessments were conducted to evaluate the neurologic damage and recovery after intracerebral hemorrhage.ResultsGenetic deficiency of TREK-1 channel exacerbated blood-brain barrier impairment and promoted cerebral edema after intracerebral hemorrhage. Meanwhile, TREK-1 deficiency aggravated focal inflammatory featured by the increased recruitment of microglia and neutrophils, the enhanced secretion of proinflammatory factors interleukin-1 beta (IL-1β), tumor necrosis factor alpha (TNF-α), and cell adhesion molecules (CAMs). Furthermore, TREK-1 deficiency promoted neuronal injury and neurological impairment.ConclusionsThese results establish the first in vivo evidence for the protective role of TREK-1 in blood-brain barrier injury and neuroinflammation after intracerebral hemorrhage. TREK-1 may thereby be harnessed to a potential therapeutical target for the treatment of intracerebral hemorrhage.

Regulation of interleukin-1 beta secretion from macrophages via modulation of potassium ion (K+ ) channel activity.

A causal relationship exists between macrophage cholesterol levels and inflammation, for example, Interleukin-1β (IL-1β) secretion. A decrease in intracellular K+ is essential for inflammasome activation/IL-1β secretion and, herein, we examined the hypothesis that cellular cholesterol affects K+ -channel activity and K+ -efflux using mouse peritoneal macrophages (MPMs) and human/THP1 macrophages. An increase in cellular cholesterol led to a significant increase in K+ currents (>350% in both MPM and THP1). Enhancing cholesterol efflux returned K+ currents back to basal levels with corresponding increase in intracellular K+ (11.2-14.5%) and reduced IL-1β secretion (32-62%). These data demonstrate a novel mechanism by which cellular cholesterol modulates inflammation/inflammasome via regulation of K+ -channel activity and intracellular K+ levels. Attenuation of IL-1β secretion by Nateglinide/Repaglinide further suggests involvement of Kir6 channels.

Nonsteroidal Anti-Inflammatory Drugs Prevent Vincristine-Dependent Cancer-Associated Fibroblasts Formation.

Vincristine is used in the clinical treatment of colon cancer, especially in patients diagnosed in the advanced phase of cancer development. Unfortunately, similar to other agents used during antitumor therapy, vincristine might induce chemoresistance. Studies of this process focus mainly on the analysis of the molecular mechanisms within cancer, usually ignoring the role of stromal cells. Our present findings confirm that vincristine stimulates the secretion of tumor growth factors class beta and interleukin-6 from cancer-associated fibroblasts as a result of paracrine stimulation by cancer cells. Based on alterations in morphology, modulation of capillary formation, and changes in endothelial and mesenchymal marker profile, our findings demonstrate that higher levels of tumor growth factor-βs and interleukin-6 enhance cancer-associated fibroblast-like cell formation through endothelial–mesenchymal transition and that nonsteroidal anti-inflammatory drug treatment (aspirin and ibuprofen) is able to inhibit this phenomenon. The process appears to be regulated by the rate of microtubule polymerization, depending on β-tubulin composition. While higher levels of tubulin-β2 and tubulin-β4 caused slowed polymerization and reduced the level of factors secreted to the extracellular matrix, tubulin-β3 induced the opposite effect. We conclude that nonsteroidal anti-inflammatory drugs should be considered for use during vincristine monotherapy in the treatment of patients diagnosed with colorectal cancer.

The cyanobacterial neurotoxin β-N-methylamino-l-alanine prevents addition of heparan sulfate to glypican-1 and increases processing of amyloid precursor protein in dividing neuronal cells

The neurotoxin β-N-methylamino-l-alanine replaces l-serine in proteins and produces Alzheimer-like pathology. In proteoglycans, e.g. glypican-1, this should preclude substitution with heparan sulfate chains. Reduced release of heparan sulfate should increase β-secretase activity and processing of amyloid precursor protein. Cultured cells were treated with β-N-methylamino-l-alanine during the growth-phase and the effect on heparan sulfate substitution and amyloid precursor protein processing was evaluated using antibodies specific for heparan sulfate, the N- and C-termini of the C-terminal fragment of β-cleaved amyloid precursor protein, and amyloid beta followed by immunofluorescence microscopy, flow cytometry or SDS-PAGE. Mouse fibroblasts, N2a neuroblastoma cells and human neural stem cells released less heparan sulfate when grown in the presence of β-N-methylamino-l-alanine. Cells expressing a recombinant, anchor-less glypican-1 secreted heparan sulfate-deficient glypican-1. There was increased processing of amyloid precursor protein in N2a cells when grown in the presence of the neurotoxin. The degradation products accumulated in cytoplasmic clusters. Secretion of amyloid beta increased approx. 3-fold. Human neural stem cells also developed cytoplasmic clusters containing degradation products of amyloid precursor protein. When non-dividing mouse N2a cells or cortical neurons were exposed to β-N-methylamino-l-alanine there was no effect on heparan sulfate substitution in glypican-1 or on amyloid precursor protein processing.

Discovery of GSK8612, a Highly Selective and Potent TBK1 Inhibitor.

The serine/threonine protein kinase TBK1 (Tank-binding Kinase-1) is a noncanonical member of the IkB kinase (IKK) family. This kinase regulates signaling pathways in innate immunity, oncogenesis, energy homeostasis, autophagy, and neuroinflammation. Herein, we report the discovery and characterization of a novel potent and highly selective TBK1 inhibitor, GSK8612. In cellular assays, this small molecule inhibited toll-like receptor (TLR)3-induced interferon regulatory factor (IRF)3 phosphorylation in Ramos cells and type I interferon (IFN) secretion in primary human mononuclear cells. In THP1 cells, GSK8612 was able to inhibit secretion of interferon beta (IFNβ) in response to dsDNA and cGAMP, the natural ligand for STING. GSK8612 is a TBK1 small molecule inhibitor displaying an excellent selectivity profile and therefore represents an ideal probe to further dissect the biology of TBK1 in models of immunity, neuroinflammation, obesity, or cancer.

Abstract TMP25: Targeting Myeloid Cell-Specific Pyruvate Kinase M2 Improves Stroke Outcome following Transient Cerebral Ischemia in the Comorbid Condition of Hyperlipidemia

Background: Activated leukocytes exhibit a high level of aerobic glycolysis compared to quiescent leukocytes. Dimeric form of pyruvate kinase M2 (PKM2) regulates aerobic glycolysis and transcription of pro-inflammatory mediators. By following the stroke therapy academic industry roundtable (STAIR) guidelines, we tested a novel hypothesis that manipulating metabolic plasticity of myeloid cells, by targeting PKM2, will improve stroke outcome. Methods: Using a well-characterized PKM2 activator (ML-265, inhibits PKM2 dimerization and stabilizes tetramer) in standardized neutrophil in vitro assays, we evaluated the mechanistic role of PKM2 in modulating post-stroke neutrophil function. Transient cerebral ischemia was induced using the intraluminal filament model of MCAO. Quantitative assessment of stroke outcome was evaluated (up to day 7) by measuring infarct volume, cerebral blood flow, survival rate, neurological/sensorimotor deficits, and thrombo-inflammation. Results: Treatment with ML-265 resulted in significantly reduced PKM2 nuclear translocation, IL-1 beta secretion and NETs formation in post-stroke peripheral neutrophils. Myeloid specific PKM2 -/- mice (male/female) on wild-type and hyperlipidemic background (PKM2 fl/fl LysM Cre and PKM2 fl/fl LysM Cre Apoe -/- ) exhibited significantly smaller infarcts and improved sensorimotor/neurological outcomes concomitant with enhanced survival rate and reduced post ischemic thrombo-inflammation. Using laser speckle imaging, we found significantly improved post-reperfusion cerebral blood flow in myeloid specific PKM2 -/- mice. In microfluidics flow chamber assay, human and mouse whole blood pretreated with ML-265 formed smaller thrombi when perfused over collagen at an arterial shear rate. Conclusions: Our data suggest that PKM2 regulates post-stroke neutrophil function and thrombo-inflammation. Targeting PKM2 may be explored as a novel strategy to improve stroke outcome.

Serotype b of Aggregatibacter actinomycetemcomitans triggers pro-inflammatory responses and amyloid beta secretion in hippocampal cells: a novel link between periodontitis and Alzheimer´s disease?

ABSTRACTIntroduction: Previous reports have proposed that Periodontal disease (PDis) predisposes to Alzheimer’s disease (AD), both highly prevalent pathologies among the elderly. The bacteria Aggregatibacter actinomycetemcomitans (Aa), associated with the most aggressive forms of PDis, are classified in different serotypes with distinct virulence according to the antigenicity of their lipopolysaccharide (LPS). Methods: Here, we determined the effects of purified LPS, from serotypes a, b or c of Aa, on primary cultures of microglia or mixed hippocampal cells. Results: We found that both culture types exhibited higher levels of inflammatory cytokines (IL-1β, IL-6 and TNFα) when treated with serotype b-LPS, compared with controls, as quantified by qPCR and/or ELISA. Also, cultures treated with serotype a-LPS displayed increased mRNA levels of the modulatory cytokines IL-4 and IL-10. Mixed hippocampal cultures treated with serotype b-LPS exhibited severe neuronal morphological changes and displayed increased levels of secreted Aβ1-42 peptide. These results indicate that LPS from different Aa serotypes triggers discriminatory immune responses, which differentially affect primary hippocampal cells. Conclusion: Altogether, our results show that treatment with serotype b-LPS triggers the secretion of proinflammatory cytokines by microglia, induces neurite shrinking, and increases the extracellular Aβ1-42 levels, all features strongly associated with the etiology of AD.

Decreased NAD Activates STAT3 and Integrin Pathways to Drive Epithelial-Mesenchymal Transition

Nicotinamide adenine dinucleotide (NAD) plays an essential role in all aspects of human life. NAD levels decrease as humans age, and supplementation with NAD precursors plays a protective role against aging and associated disease. Less is known about the effects of decreased NAD on cellular processes, which is the basis for understanding the relationship between cellular NAD levels and aging-associated disease. In the present study, cellular NAD levels were decreased by overexpression of CD38, a NAD hydrolase, or by treating cells with FK866, an inhibitor of nicotinamide phosphoribosyltransferase (NAMPT). Quantitative proteomics revealed that declining NAD levels downregulated proteins associated with primary metabolism and suppressed cell growth in culture and nude mice. Decreased glutathione synthesis caused a 4-fold increase in cellular reactive oxygen species levels, and more importantly upregulated proteins related to movement and adhesion. In turn, this significantly changed cell morphology and caused cells to undergo epithelial to mesenchymal transition (EMT). Secretomic analysis also showed that decreased NAD triggered interleukin-6 and transforming growth factor beta (TGFβ) secretion, which activated integrin-β-catenin, TGFβ-MAPK, and inflammation signaling pathways to sustain the signaling required for EMT. We further revealed that decreased NAD inactivated sirtuin 1, resulting in increased signal transducer and activator of transcription 3 (STAT3) acetylation and phosphorylation, and STAT3 activation. Repletion of nicotinamide or nicotinic acid inactivated STAT3 and reversed EMT, as did STAT3 inhibition. Taken together, these results indicate that decreased NAD activates multiple signaling pathways to promote EMT and suggests that age-dependent decreases in NAD may contribute to tumor progression. Consequently, repletion of NAD precursors has potential benefits for inhibiting cancer progression.

A new theory of depression based on the serotonin/kynurenine relationship and the hypothalamicpituitary- adrenal axis

The serotonergic and immunological hypothesis of depression proposes that certain types of excessive stress distort the relationship between the activities of the innate immune and central nervous systems, so that the stress caused by an infection, or excessive psychological stress, activate toll-like receptors such as the TLR-4, the transcription factor NF-kB, the inflammasome NLRP3, as well as the secretion of interleukin-1 beta (IL-1β), interleukin-6 (IL-6) and other factors of the innate immune response, causing first, the general symptoms of the disease which appear with any infection, but also those characteristic of depressive illness such as dysphoria and anhedonia. The evidence indicates that, if the stimulus persists or recurs within 24 hours, the indole-2, 3-dioxygenase enzyme (IDO) of the kynurenine metabolic pathway, which increases the synthesis of quinolinic acid, is activated with an associated reduction of serotonin synthesis. Quinolinic acid activates NMDA receptors in the central nervous system and stimulates the secretion of interleukins IL-6 and 1L-1β, among others, promoting hyper-activity of the HPA axis and reinforcing a bias of the tryptophan metabolism to produce quinolinic acid, and interleukins by the innate immune system, further reducing the synthesis of serotonin and consolidating the depressive process. We discuss the evidence showing that this process can be initiated by either interleukin stimulated by an infection or some vaccines or excessive psychological stress that activates the HPA axis together with said innate immune response, causing a process of aseptic inflammation in the central nervous system.

Oregano Essential Oil Attenuates RAW264.7 Cells from Lipopolysaccharide-Induced Inflammatory Response through Regulating NADPH Oxidase Activation-Driven Oxidative Stress.

Oregano is an aromatic plant widely distributed throughout the Mediterranean area and in Asia. Recent studies have revealed that the anti-inflammatory effect of essential oil in this plant. However, the mechanisms underlying the therapeutic potential have not been well elucidated. This study determined whether oregano essential oil (OEO) exerts an anti-inflammatory effect on lipopolysaccharide (LPS)-treated murine macrophage cells (RAW264.7 cells) in vitro and elucidated the possible underlying molecular mechanisms. The results showed that OEO (2.5–10 μg/mL) inhibited the expression and secretion of interleukin-1 beta (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α) in RAW264.7 cells treated with LPS (1 μg/mL). Consistent with the pro-inflammatory gene expression, the OEO treatment efficiently reduced the LPS-induced activation of mitogen-activated protein kinase, protein kinase B, and nuclear factor κB in RAW264.7 cells. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibition in Nox2 protein-silenced cells attenuated the mRNA expression of IL-1β, IL-6, and TNF-α in the LPS-induced RAW264.7 cells. The OEO inhibited the LPS-induced elevation of NADPH oxidase and oxidative stress. This result suggests that LPS induces RAW264.7 cell inflammation through the NADPH oxidase-mediated production of reactive oxygen species (ROS). In conclusion, OEO protects against the LPS-induced RAW264.7 cell inflammatory response through the NADPH oxidase/ROS pathway.

Anti-Infectivity against Herpes Simplex Virus and Selected Microbes and Anti-Inflammatory Activities of Compounds Isolated from Eucalyptus globulus Labill.

Herpes simplex virus (HSV) causes numerous mild-to-serious human diseases, including mucocutaneous herpes infections and life-threatening herpes encephalitis. Moreover, herpes viral lesions can be complicated by inflammation and secondary bacterial infections. The development of resistance to antiviral drugs along with the undesirable side effects of these drugs are relevant argue for the development of new anti-HSV drugs with diverse mechanisms of action. Eucalyptus extracts have been used for decades to combat various infectious diseases. We isolated and studied 12 pure compounds and one mixture of two constitutional isomers from the leaves and twigs of E. globulus. The structures were identified by spectroscopic methods (NMR, HR-MS, IR) and all of them were tested for antiherpetic activity against the replication of antigen types HSV-1 and HSV-2. Tereticornate A (12) (IC50: 0.96 μg/mL; selectivity index CC50/IC50: 218.8) showed the strongest activity in the anti-HSV-1 assay, even greater than acyclovir (IC50: 1.92 μg/mL; selectivity index CC50/IC50: 109.4), a standard antiviral drug. Cypellocarpin C (5) (EC50: 0.73 μg/mL; selectivity index CC50/EC50: 287.7) showed the most potent anti-HSV-2 activity, also more intensive than acyclovir (EC50: 1.75 μg/mL; selectivity index CC50/EC50: 120.0). The antimicrobial activity of the isolated compounds was also evaluated against the bacteria Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Pseudomonas aeruginosa and the yeast Candida albicans. The anti-inflammatory potential was examined using LPS-stimulated THP-1-XBlue™-MD2-CD14 and THP-1 macrophages and focusing on the influences of the NF-κB/AP-1 activity and the secretion of pro-inflammatory cytokines IL-1β and TNF-α.

Dengue Virus Induces the Release of sCD40L and Changes in Levels of Membranal CD42b and CD40L Molecules in Human Platelets.

Platelets are considered as significant players in innate and adaptive immune responses. The adhesion molecules they express, including P-selectin, CD40L, and CD42b, facilitate interactions with many cellular effectors. Upon interacting with a pathogen, platelets rapidly express and enhance their adhesion molecules, and secrete cytokines and chemokines. A similar phenomenon occurs after exposure of platelets to thrombin, an agonist extensively used for in vitro activation of these cells. It was recently reported that the dengue virus not only interacts with platelets but possibly infects them, which triggers an increased expression of adhesion molecule P-selectin as well as secretion of IL-1β. In the present study, surface molecules of platelets like CD40L, CD42b, CD62P, and MHC class I were evaluated at 4 h of interaction with dengue virus serotype 2 (DENV-2), finding that DENV-2 induced a sharp rise in the membrane expression of all these molecules. At 2 and 4 h of DENV-2 stimulation of platelets, a significantly greater secretion of soluble CD40L (sCD40L) was found (versus basal levels) as well as cytokines such as GM-CSF, IL-6, IL-8, IL-10, and TNF-α. Compared to basal, DENV-2 elicited more than two-fold increase in these cytokines. Compared to the thrombin-induced response, the level generated by DENV-2 was much higher for GM-CSF, IL-6, and TNF-α. All these events induced by DENV end up in conspicuous morphological changes observed in platelets by confocal microscopy and transmission electron microscopy, very different from those elicited by thrombin in a more physiological scenery.

Generation of a Novel Gene Therapy Vector Encoding Beta-Cell–Specific Insulin for Diabetes Treatment

Insulin injection is the main treatment modality especially for patients with type 1 diabetes. Despite the development of several insulin analogs, recent meta-analysis indicated that rapid and long acting insulin analogs provide little benefit compared to conventional insulins regarding to glycemic control. Therefore, gene therapy methods are investigated to develop a novel treatment approach to reconstitute a natural endogenous insulin expression profile in diabetic patients. For this purpose, Multisite Gateway Technology was employed to generate a transfer vector carrying lentiviral backbone with minimal insulin promotor hooked up to proinsulin gene sequence. Following confirmation of the transfer plasmid by restriction enzyme digestion and DNA sequence analysis, high-titer LentiINS vectors at a concentration of 10e9 TU/ml were successfully produced by CaPO4 cotransfection of 293T cells with packaging and transfer plasmids. While transduction of NIT1 mouse pancreatic beta cell lines produced 3 fold increase in insulin gene expression compared to controls, no insulin expression was detected in 293T kidney cell line. These results indicate that we successfully constructed an HIV-based lentiviral gene therapy vector being capable of beta cell-specific insulin gene expression and secretion (TUBITAK 215S820). Disclosure E.O. Sahin: None. F. Erendor: None. M. Balci: None. S. Sanlioglu: None.

Characterization of phospholipids from Pacific saury ( Cololabis saira ) viscera and their neuroprotective activity

Abstract Pacific saury (Cololabis saira) is an important marine commercial fish in the world. The crude Pacific saury viscera (PSV) phospholipids were extracted with supercritical carbon dioxide and organic solvent method using ethanol, yielding 24/100 g phospholipids and 21/100 g (dry weight), respectively. Their fatty acid profiles were obtained using gas chromatography. Docosahexaenoic acid (DHA) and palmitic acid were the major fatty acids. High-performance liquid chromatography equipped with an evaporation scattered light detector was used to quantitatively analyse the major component of the phospholipid extract. Phosphatidylcholine (89 mg/g), phosphatidylinositol (53 mg/g) and phosphatidylethanolamine (40 mg/g) were the main phospholipids, while phosphatidylserine, lysophosphatidylcholine and lysophosphatidylcholine were the minor phospholipids at 5, 15 and 19 mg/g, respectively. The inhibitory effects of PSV phospholipids and the PC fraction, obtained using silica gel column chromatography, on the secretion of amyloid beta (Aβ)1–42 in the supernatant of Chinese hamster ovary cells stably transfected with amyloid precursor protein and presenilin 1 were studied, with soybean phospholipids as the positive control. The results showed that, compared with the model control group, the Aβ1–42 was decreased 69% with PSV phospholipid, 62% with PC fraction and 36% with soybean phospholipids. Taken together, PSV phospholipids were rich in n-3 fatty acids, especially DHA, and may have potential for the treatment of neurodegenerative diseases such as Alzheimer disease.

P32: Magnesium sulfate attenuates cell cytotoxicity in an in‐vitro model of funisitis through inhibition of P2X7 receptor‐mediated IL1 beta secretion

P32: Magnesium sulfate attenuates cell cytotoxicity in an in‐vitro model of funisitis through inhibition of P2X7 receptor‐mediated IL1 beta secretion

Dysfunction of various organelles provokes multiple cell death after quantum dot exposure.

Quantum dots (QDs) are different from the materials with the micrometer scale. Owing to the superiority in fluorescence and optical stability, QDs act as possible diagnostic and therapeutic tools for application in biomedical field. However, potential threats of QDs to human health hamper their wide utilization in life sciences. It has been reported that oxidative stress and inflammation are involved in toxicity caused by QDs. Recently, accumulating research unveiled that disturbance of subcellular structures plays a magnificent role in cytotoxicity of QDs. Diverse organelles would collapse during QD treatment, including DNA damage, endoplasmic reticulum stress, mitochondrial dysfunction and lysosomal rupture. Different forms of cellular end points on the basis of recent research have been concluded. Apart from apoptosis and autophagy, a new form of cell death termed pyroptosis, which is finely orchestrated by inflammasome complex and gasdermin family with secretion of interleukin-1 beta and interleukin-18, was also summarized. Finally, several potential cellular signaling pathways were also listed. Activation of Toll-like receptor-4/myeloid differentiation primary response 88, nuclear factor kappa-light-chain-enhancer of activated B cells and NACHT, LRR and PYD domains-containing protein 3 inflammasome pathways by QD exposure is associated with regulation of cellular processes. With the development of QDs, toxicity evaluation is far behind its development, where specific mechanisms of toxic effects are not clearly defined. Further studies concerned with this promising area are urgently required.

Defining how mitochondrial DNA stress regulates innate immune responses

Abstract Polymerase gamma (pol γ) is nuclear-encoded protein responsible for mtDNA replication. POLG mutations are recognized as a significant cause of primary mitochondrial diseases and are implicated in a range of disorders. Several mouse models of POLG-related disease have been reported, the most well studied is the POLG-mutator mouse. Animals homozygous for the mutant alleles exhibit elevated mtDNA instability, and POLG-mutator mice exhibit pathology that mirrors various aspects of human POLG-related diseases. The literature has few descriptions about the immune system of POLG-mutator mice or patients with POLG-related disorders, despite the fact that aberrant immune responses have been implicated in the pathology of diseases with similar phenotypes. We have found that mitochondrial dysfunction, resulting from POLG mutation, alters innate immune responses at baseline and after challenge, which we hypothesize may exacerbate pathology in POLG-related mitochondrial disorders. To explore this hypothesis, we have stimulated macrophages from WT and POLG-mutator mice with various innate immune stimuli, and have uncovered that POLG-mutator macrophages display alterations in type I interferon responses, but strikingly, exhibit profound decreases in inflammasome-dependent interleukin-1 beta (IL-1b) secretion. Ongoing work will determine how mitochondrial dysfunction impairs inflammasome activation and IL-1b secretion in order to appreciate how aberrant innate immune responses influence POLG-related disease progression.

Effects of selective serotonin-reuptake inhibitors (SSRIs) on human villous trophoblasts syncytialization

Selective serotonin-reuptake inhibitors (SSRIs) are the most commonly prescribed antidepressants during pregnancy. The human placenta is a highly specialized organ supporting normal growth and development of the fetus. Therefore, this study aims to analyze the effects of SSRIs on villous cytotrophoblasts cells, using BeWo cells and human placental trophoblast cells in primary culture. The SSRIs fluoxetine and its metabolite norfluoxetine, sertraline and venlafaxine did not affect BeWo cell proliferation and viability, nor the percentage of M30-positive (apoptotic) primary trophoblast cells. None of the SSRIs affected basal or forskolin-stimulated BeWo cell fusion, whereas sertraline and venlafaxine increased the fusion of primary villous trophoblasts. Sertraline and venlafaxine also modified human chorionic gonadotropin beta (β-hCG) secretion by BeWo cells, whereas none of the SSRIs affected β-hCG secretion in primary trophoblasts. Norfluoxetine increased CGB (chorionic gonadotropin beta) and GJA1 (gap junction protein alpha 1) levels of gene expression (biomarkers of syncytialization) in BeWo cells, whereas in primary trophoblasts none of the SSRIs tested affected the expression of these genes. This study shows that SSRIs affect villous trophoblast syncytialization in a structure- and concentration-dependent manner and suggests that certain SSRIs may compromise placental health. In addition, it highlights the importance of using primary trophoblast cells instead of “trophoblast -like” cell lines to assess the effects of medications on human villous trophoblast function.