1.
Abbas, Abdul-Karim, 1959, et al.
(författare)
Long-term potentiation and insult conditioning in hippocampal slices from young rats: a role for protein synthesis under chemical stress?
2010
Ingår i: The 10th Biennial Meeting of the Asia-Pacific Society for Neurochemistry (APSN), October 17-20, 2010, Phuket, Thailand.
Konferensbidrag (övrigt vetenskapligt/konstnärligt) abstract
We have previously demonstrated that in young rats (12-20-day-old) a sustained long-term potentiation (LTP) can still be induced under conditions of protein synthesis inhibition. It was therefore suggested that sufficient and necessary proteins were already available at the induction time to accomplish LTP maintenance for several hours. Against this background, we have questioned whether hippocampal slices subjected to certain insult conditions might be more sensitive to protein synthesis inhibitors. High K+ concentration has previously been reported to cause an amnesic effect in vivo as well as increasing protein turnover in vitro. We have here employed a K+ insult model under conditions when protein synthesis was inhibited. Recordings were obtained from hippocampal slices for up to 9 h, with or without a cocktail of protein synthesis inhibitors, containing cycloheximide (60 µM) and anisomycin (25 µM). High potassium (50 mM) was transiently applied (5-15 min) shortly after inducing LTP in one of two separate pathways stimulated alternatively. Additionally, an NMDA-receptor antagonist AP5 was supplied after LTP induction to minimize effects related to depolarization-induced glutamate release. Following elimination of all responses for about 30 min, both test and control responses partly recovered. The degree of remaining LTP, defined as test/control ratio, was reduced in both groups of slices (NMDA-independent depotentiation) but was significantly smaller in the drug-treated ones. We are also running an insult model based on oxidative stress, applying hydrogen peroxide (4-5 mM) before or after LTP induction; however, the results are still insufficient for a final conclusion. The potency of cycloheximide, anisomycin or cocktail of the drugs was verified by measurement of incorporation of [3H]-leucine into trichloracetic acid (TCA) precipitable macromolecules. Cycloheximide, anisomycin or cocktail, at concentrations used here caused 95%, 97% and 95% blocking effect, respectively. Our data confirm the idea that sufficient and necessary constitutive proteins are available in the young hippocampus to maintain LTP under conditions of protein synthesis inhibition. They also reveal that LTP in slices subjected to certain insult conditions early after the induction is sensitive to protein synthesis inhibition, probably due to increase in constitutive proteins turnover.
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4.
Lai, Kuei-Hung
(författare)
Studies on anti-leukemic terpenoids from medicinal mushrooms and marine sponges with ChemGPS-NP-based targets investigation of lead compounds
2017
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
This thesis investigates the anti-leukemic activity of terpenoids isolated from medicinal mushrooms and marine sponges, as well as their possible targets and mechanisms of action.In the first section, we focused on studying the triterpenoidal components of three triterpenoid-enriched medicinal mushrooms Antrodia cinnamomea, Ganoderma lucidum, and Poria cocos, which have been used in folk medicine for centuries and also developed into several contemporary marketed products. We isolated the major and characteristic triterpenoids from these mushrooms, together with six new lanostanoids (II-1–II-6). The anti-leukemic activity of the isolates was evaluated in vitro using MTT proliferative assay and seven of them exhibited potential anti-leukemic effect. The active lead compounds were further subjected to computational analyses utilizing the ChemGPS-NP tool. We established a database for the anti-leukemic relevant chemical space of triterpenoids isolated from these three medicinal mushrooms, which could be used as a reference database for further research on anti-leukemic triterpenoids. Our results indicated that the anti-leukemic effect of the active lead compounds was mediated not only through topoisomerases inhibition but also through inhibiting DNA polymerases.The second and third sections focused on isolation of anti-leukemic sesterterpenoids from sponges. The investigation of Carteriospongia sp. led to the isolation of two new scalarane-type sesterterpenoids (III-1 and III-2) and one known tetraprenyltoluquinol-related metabolite (III-3). All isolates exhibit an apoptotic mechanism of action against Molt 4 cells, found to be mediated through the disruption of the mitochondrial membrane potential (MMP) and inhibition of topoisomerase IIα expression. Detailed investigation of the apoptotic mechanism of action using molecular docking analysis revealed that compound III-1 might target Hsp90 protein. The apoptotic-inducing effect of III-3 was supported by in vivo experiment by suppressing the volume of xenograft tumor growth (47.58%) compared with the control.In the final section of this thesis we studied manoalide and its derivatives, sesterterpenoids isolated from the sponge Luffariella sp.. Manoalide has been studied as a potential anti-inflammatory agent for the last thirty years with more than 200 publications and 40 patents. However, the configurations at positions 24 and 25 were never revealed. In the current study, ten manoalide-type sesterterpenoids (IV-1–IV-10) were isolated from Luffariella sp. and their stereoisomers at positions 24 and 25 were identified and separated for the first time. The configuration at positions 24 and 25 showed to have a significant effect on the anti-leukemic activity of manoalide derivatives, with the 24R,25S-isomer exhibiting the most potent anti-leukemic activity. The apoptotic mechanism of action of compound IV-7 against Molt 4 cells was investigated, and the compound was found to trigger MMP disruption and intracellular reactive oxygen species (ROS) generation. Compound IV-7 also inhibited activity against both human topoisomerases, I and II. The in vivo experiment further supported the anti-leukemic effect of IV-7 with a 66.11% tumor volume suppression compared to the control.
5.
Bhandage, Amol K., 1988-
(författare)
Glutamate and GABA signalling components in the human brain and in immune cells
2016
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Glutamate and γ-aminobutyric acid (GABA) are the principal excitatory and inhibitory neurotransmitters in the central nervous system (CNS). They both can activate their ionotropic and metabotropic receptors. Glutamate activates ionotropic glutamate receptors (iGlu - AMPA, kainate and NMDA receptors) and GABA activates GABA-A receptors which are modulated by many types of drugs and substances including alcohol. Using real time quantitative polymerase chain reaction, I have shown that iGlu and/or GABA-A receptor subunits were expressed in the hippocampus dentate gyrus (HDG), orbitofrontal cortex (OFC), dorsolateral prefrontal cortex (DL-PFC), central amygdala (CeA), caudate and putamen of the human brain and their expression was altered by chronic excessive alcohol consumption. It indicates that excitatory and inhibitory neurotransmission may have been altered in the brain of human alcoholics. It is possible that changes in one type of neurotransmitter system may drive changes in another. These brain regions also play a role in brain reward system. Any changes in them may lead to changes in the normal brain functions.Apart from the CNS, glutamate and GABA are also present in the blood and can be synthesised by pancreatic islet cells and immune cells. They may act as immunomodulators of circulating immune cells and can affect immune function through glutamate and GABA receptors. I found that T cells from human, rat and mouse lymph nodes expressed the mRNAs and proteins for specific GABA-A receptor subunits. GABA-evoked transient and tonic currents recorded using the patch clamp technique demonstrate the functional GABA-A channel in T cells. Furthermore, the mRNAs for specific iGlu, GABA-A and GABA-B receptor subunits and chloride cotransporters were detected in peripheral blood mononuclear cells (PBMCs) from men, non-pregnant women, healthy and depressed pregnant women. The results indicate that the expression of iGlu, GABA-A and GABA-B receptors is related to gender, pregnancy and mental health and support the notion that glutamate and GABA receptors may modulate immune function. Intra- and interspecies variability exists in the expression and it is further influenced by physiological conditions.
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Rodríguez-Piñeiro, Ana María, et al.
(författare)
The colonic mucus protection depends on the microbiota
2015
Ingår i: Gut microbes. - : Informa UK Limited. - 1949-0976 .- 1949-0984. ; 6:5, s. 326-30
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt) abstract
The intestinal mucus is a pivotal part of our intestinal protection. It provides slow diffusion of protective molecules, trapping of luminal material as bacteria and smooth transport in the small intestine. In colon it restricts bacterial access to the epithelium limiting the responses to the enormous bacterial load present at this location. The development of these systems depends on the microbiota composition as seen in our recent study comparing the mucus phenotype in 2 colonies kept in different husbandries within the same SPF animal facility. One colony had impenetrable colonic mucus while the other colony had more penetrable mucus. The mucus phenotypes were transmitted via the microbiota and clear differences in its composition could be detected. Candidates associated with the different colonies were identified but the observed mucus difference could not be assigned to a specific bacterium.
8.
Klawonn, Anna, 1985-
(författare)
Molecular Mechanisms of Reward and Aversion
2017
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Various molecular pathways in the brain shape our understanding of good and bad, as well as our motivation to seek and avoid such stimuli. This work evolves around how systemic inflammation causes aversion; and why general unpleasant states such as sickness, stress, pain and nausea are encoded by our brain as undesirable; and contrary to these questions, how drugs of abuse can subjugate the motivational neurocircuitry of the brain. A common feature of these various disease states is involvement of the motivational neurocircuitry - from mesolimbic to striatonigral pathways. Having an intact motivational system is what helps us evade negative outcomes and approach natural positive reinforcers, which is essential for our survival. During disease-states the motivational neurocircuitry may be overthrown by the molecular mechanisms that originally were meant to aid us.In study I, to investigate how inflammation is perceived as aversive, we used a behavioral test based on Pavlovian place conditioning with the aversive inflammatory stimulus E. coli lipopolysaccharide (LPS). Using a combination of cell-type specific gene deletions, pharmacology, and chemogenetics, we uncovered that systemic inflammation triggered aversion by MyD88-dependent activation of the brain endothelium followed by COX1-mediated cerebral prostaglandin E2 (PGE2) synthesis. Moreover, we showed that inflammation-induced PGE2 targeted EP1 receptors on striatal dopamine D1 receptor–expressing neurons and that this signaling sequence induced aversion through GABA-mediated inhibition of dopaminergic cells. Finally, inflammation-induced aversion was not an indirect consequence of fever or anorexia but constituted an independent inflammatory symptom triggered by a unique molecular mechanism. Collectively, these findings demonstrate that PGE2-mediated modulation of the dopaminergic circuitry is a key mechanism underlying inflammation-induced aversion.In study II, we investigate the role of peripheral IFN-γ in LPS induced conditioned place aversion by employing a strategy based on global and cell-type specific gene deletions, combined with measures of gene-expression. LPS induced IFN-ɣ expression in the blood, and deletion of IFN-ɣ or its receptor prevented conditioned place aversion (CPA) to LPS. LPS increased the expression of chemokine Cxcl10 in the striatum of normal mice. This induction was absent in mice lacking IFN-ɣ receptors or Myd88 in blood brain barrier endothelial cells. Furthermore, inflammation-induced aversion was blocked in mice lacking Cxcl10 or its receptor Cxcr3. Finally, mice with a selective deletion of the IFN-ɣ receptor in brain endothelial cells did not develop inflammation-induced aversion. Collectively, these findings demonstrate that circulating IFN-ɣ binding to receptors on brain endothelial cells which induces Cxcl10, is a central link in the signaling chain eliciting inflammation-induced aversion.In study III, we explored the role of melanocortin 4 receptors (MC4Rs) in aversive processing using genetically modified mice in CPA to various stimuli. In normal mice, robust aversions were induced by systemic inflammation, nausea, pain and kappa opioid receptor-induced dysphoria. In sharp contrast, mice lacking MC4Rs displayed preference towards most of the aversive stimuli, but were indifferent to pain. The unusual flip from aversion to reward in mice lacking MC4Rs was dopamine-dependent and associated with a change from decreased to increased activity of the dopamine system. The responses to aversive stimuli were normalized when MC4Rs were re-expressed on dopamine D1 receptor-expressing cells or in the striatum of mice otherwise lacking MC4Rs. Furthermore, activation of arcuate nucleus proopiomelanocortin neurons projecting to the ventral striatum increased the activity of striatal neurons in a MC4R-dependent manner and elicited aversion. Our findings demonstrate that melanocortin signaling through striatal MC4Rs is critical for assigning negative motivational valence to harmful stimuli.The neurotransmitter acetylcholine has been implied in reward learning and drug addiction. However, the role of cholinergic receptor subtypes in such processes remains elusive. In study IV we investigated the function of muscarinic M4Rs on dopamine D1R expressing neurons and acetylcholinergic neurons, using transgenic mice in various reward-enforced behaviors and in a “waiting”-impulsivity test. Mice lacking M4-receptors from D1-receptor expressing neurons exhibited an escalated reward seeking phenotype towards cocaine and natural reward, in Pavlovian conditioning and an operant self-administration task, respectively. In addition, the M4-D1RCre mice showed impaired waiting impulsivity in the 5-choice-serial-reaction-time-task. On the contrary, mice without M4Rs in acetylcholinergic neurons were unable to learn positive reinforcement to natural reward and cocaine, in an operant runway paradigm and in Pavlovian conditioning. Immediate early gene expression mirrored the behavioral findings arising from M4R-D1R knockout, as cocaine induced cFos and FosB was significantly increased in the forebrain of M4-D1RCre mice, whereas it remained normal in the M4R-ChatCre mice. Our study illustrates that muscarinic M4Rs on specific neural populations, either cholinergic or D1R-expressing, are pivotal for learning processes related to both natural reward and drugs of abuse, with opposing functionality.
9.
Lindberg, Frida A, et al.
(författare)
SLC38A10 deficiency in male mice affect plasma levels of threonine and histidine
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Solute carriers belong to the biggest group of transporters in the human genome, but more knowledge is needed in order to fully understand their function and possible role as therapeutic targets. SLC38 is a family of amino acid transporters, commonly referred to as SNATs, consisting of 11 members. The tenth member, SLC38A10, is one of the least characterized members and is the focus of this study. By using a knockout mouse model, we studied the biological effects of SLC38A10 deficiency in vivo. We performed a transcriptomic analysis of whole brain and found seven differentially expressed genes in SLC38A10 deficient mice (Gm48159, Nr4a1, Tuba1c, Lrrc56, mt-Tp, Hbb-bt and Snord116/9). By measuring amino acids in plasma, we found lower levels of threonine and histidine in males, while no amino acids were altered in females, suggesting that SLC38A10-/- might affect sexes differently. Using RT-qPCR, we investigated the effect of SLC38A10 deficiency on mRNA expression of other SLC38 members, Mtor and Rps6kb1 in brain, liver, lung, muscle and kidney, but no differences were found. A relative telomere length measurement was also made, as a marker for cellular age, but no differences were found between the genotypes. We conclude that SLC38A10 might be important for keeping amino acid homeostasis in plasma, at least in males, but no major effects were seen on transcriptomic expression or telomere length in whole brain.
10.
Nowak, Christoph, 1986-
(författare)
Insulin Resistance : Causes, biomarkers and consequences
2017
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
The worldwide increasing number of persons affected by largely preventable diseases like diabetes demands better prevention and treatment. Insulin is required for effective utilisation of circulating nutrients. Impaired responsiveness to insulin (insulin resistance, IR) is a hallmark of type 2 diabetes and independently raises the risk of heart attack and stroke. The pathophysiology of IR is incompletely understood. High-throughput measurement of large numbers of circulating biomarkers may provide new insights beyond established risk factors.The aims of this thesis were to (i) use proteomics, metabolomics and genomics methods in large community samples to identify biomarkers of IR; (ii) assess biomarkers for risk prediction and insights into aetiology and consequences of IR; and (iii) use Mendelian randomisation analysis to assess causality.In Study I, analysis of 80 circulating proteins in 70-to-77-year-old Swedes identified cathepsin D as a biomarker for IR and highlighted a tentative causal effect of IR on raised plasma tissue plasminogen activator levels. In Study II, nontargeted fasting plasma metabolomics was used to discover 52 metabolites associated with glycaemic traits in non-diabetic 70-year-old men. Replication in independent samples of several thousand persons provided evidence for a causal effect of IR on reduced plasma oleic acid and palmitoleic acid levels. In Study III, nontargeted metabolomics in plasma samples obtained at three time points during an oral glucose challenge in 70-year-old men identified associations between a physiologic measure of IR and concentration changes in medium-chain acylcarnitines, monounsaturated fatty acids, bile acids and lysophosphatidylethanolamines. Study IV provided evidence in two large longitudinal cohorts for causal effects of type 2 diabetes and impaired insulin secretion on raised coronary artery disease risk.In conclusion, the Studies in this thesis provide new insights into the pathophysiology and adverse health consequences of IR and illustrate the value of combining traditional epidemiologic designs with recent molecular techniques and bioinformatics methods. The results provide limited evidence for the role of circulating proteins and small molecules in IR and require replication in separate studies and validation in experimental designs.
