Imidazoline receptor antisera-selected protein: a unique modulator of neuronal differentiation.

The imidazoline I1 receptor (I1-R) is a novel receptor found primarily in the brain and nervous tissue where it modulates neurotransmission. It is named for its high affinity for compounds with an imidazoline structure such as the anti-hypertensive drugs, clonidine and moxonidine. The imidazoline receptor antisera-selected protein (IRAS) is the putative clone of the I1-R. IRAS has a unique structure, which does not resemble any other receptor protein. IRAS is present throughout the body with highest levels in the brain. There is a growing body of research examining the physiological roles of IRAS as an I1-R, in cell survival, migration and protein trafficking. However, there is little research into its neuronal functions. IRAS interacts with other membrane receptors: the mouse homologue of IRAS reorganises the actin cytoskeleton through interaction with the α5β1 fibronectin receptor. IRAS also binds insulin receptor substrate 4 and enhances insulin-induced extracellular signal-regulated kinase1/2 (ERK1/2) activation. Actin reorganisation and ERK1/2 activation are important for the development of neurites during neuronal differentiation. Therefore, the work described in this thesis aimed to investigate the effects of IRAS on neuronal differentiation. Studies reported in this thesis also aimed to investigate whether IRAS affected ERK1/2 signalling of other receptors involved in neuronal differentiation such as the NGF receptor, TrkA, and lysophospholipid receptors. The above aims were carried out in neuronal model PC12 cells transfected with either IRAS or a vector plasmid. Fluorescence microscopy and Western blotting techniques were used to examine the effect of IRAS on cell morphology and ERK1/2 signalling. The work described in this thesis found that IRAS reorganises the actin cytoskeleton and enhances growth cone development in PC12 cells. This study also shows that IRAS differentially enhances or inhibits NGF-induced PC12 cell differentiation depending on the presence or absence of serum in the media. In full-serum conditions, IRAS enhanced neurite outgrowth and this was accompanied by an increase in ERK1/2 activation. In serum-starved cells, IRAS inhibited neurite outgrowth with similar levels of ERK1/2 activation observed in vector- and IRAS-transfected cells. Finally, studies presented in this thesis found that IRAS enhances lysophosphatidic acid-induced ERK1/2 activation and that IRAS interacting with lysophospholipid receptor agonists present in serum is a potential mechanism by which it enhances NGF-induced ERK1/2 activation in full-serum conditions.Thesis (Ph.D.) - University of Adelaide, School of Medical Sciences, 200

(-)-Epigallocatechin-3-gallate (EGCG) maintains k-casein in its pre-fibrillar state without redirecting its aggregation pathway

The polyphenol (-)-epigallocatechin-3-gallate (EGCG) has recently attracted much research interest in the field of protein-misfolding diseases because of its potent anti-amyloid activity against amyloid-beta, alpha-synuclein and huntingtin, the amyloid-fibril-forming proteins involved in Alzheimer\u27s, Parkinson\u27s and Huntington\u27s diseases, respectively. EGCG redirects the aggregation of these polypeptides to a disordered off-folding pathway that results in the formation of non-toxic amorphous aggregates. whether this anti-fibril activity is specific to these disease-related target proteins or ismore generic remains to be established. In addition, the mechanism by which EGCG exerts its effects, as with all anti-amyloidogenic polyphenols, remains unclear. To address these aspects, we have investigated the ability of EGCG to inhibit amyloidogenesis of the generic model fibril-forming protein RCMkappa-CN (reduced and carboxymethylated kappa-casein) and thereby protect pheochromocytoma-12 cells from RCMkappa-CN amyloid-induced toxicity. We found that EGCG potently inhibits in vitro fibril formation byRCMkappa-CN [the IC50 for 50 uM RCMkappa-CN is 1 uM]. Biophysical studies reveal that EGCG prevents RCMkappa-CN fibril formation by stabilising RCMkappa-CN in its nativelike state rather than by redirecting its aggregation to the disordered, amorphous aggregation pathway. Thus, while it appears that EGCG is a generic inhibitor of amyloid-fibril formation, the mechanism by which it achieves this inhibition is specific to the target fibril-forming polypeptide. It is proposed that EGCG is directed to the amyloidogenic sheet-turn-sheet motif of monomeric RCMkappa-CN with high affinity by strong non-specific hydrophobic associations. Additional non-covalent pi-pi stacking interactions between the polyphenolic and aromatic residues common to the amyloidogenic sequence are also implicated

(-)-Epigallocatechin-3-gallate (EGCG) maintains k-casein in its pre-fibrillar state without redirecting its aggregation pathway

The polyphenol (-)-epigallocatechin-3-gallate (EGCG) has recently attracted much research interest in the field of protein-misfolding diseases because of its potent anti-amyloid activity against amyloid-beta, alpha-synuclein and huntingtin, the amyloid-fibril-forming proteins involved in Alzheimer\u27s, Parkinson\u27s and Huntington\u27s diseases, respectively. EGCG redirects the aggregation of these polypeptides to a disordered off-folding pathway that results in the formation of non-toxic amorphous aggregates. whether this anti-fibril activity is specific to these disease-related target proteins or ismore generic remains to be established. In addition, the mechanism by which EGCG exerts its effects, as with all anti-amyloidogenic polyphenols, remains unclear. To address these aspects, we have investigated the ability of EGCG to inhibit amyloidogenesis of the generic model fibril-forming protein RCMkappa-CN (reduced and carboxymethylated kappa-casein) and thereby protect pheochromocytoma-12 cells from RCMkappa-CN amyloid-induced toxicity. We found that EGCG potently inhibits in vitro fibril formation byRCMkappa-CN [the IC50 for 50 uM RCMkappa-CN is 1 uM]. Biophysical studies reveal that EGCG prevents RCMkappa-CN fibril formation by stabilising RCMkappa-CN in its nativelike state rather than by redirecting its aggregation to the disordered, amorphous aggregation pathway. Thus, while it appears that EGCG is a generic inhibitor of amyloid-fibril formation, the mechanism by which it achieves this inhibition is specific to the target fibril-forming polypeptide. It is proposed that EGCG is directed to the amyloidogenic sheet-turn-sheet motif of monomeric RCMkappa-CN with high affinity by strong non-specific hydrophobic associations. Additional non-covalent pi-pi stacking interactions between the polyphenolic and aromatic residues common to the amyloidogenic sequence are also implicated

More than One Way to be Happy: A Typology of Marital Happiness

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/100129/1/famp12028.pd

The amyloid fibril-forming properties of the amphibian antimicrobial peptide uperin 3.5

The amphibian skin is a vast resource for bioactive peptides, which form the basis of the animals' innate immune system. Key components of the secretions of the cutaneous glands are antimicrobial peptides (AMPs), which exert their cytotoxic effects often as a result of membrane disruption. It is becoming increasingly evident that there is a link between the mechanism of action of AMPs and amyloidogenic peptides and proteins. In this work, we demonstrate that the broad-spectrum amphibian AMP uperin 3.5, which has a random-coil structure in solution but adopts an α-helical structure in membrane-like environments, forms amyloid fibrils rapidly in solution at neutral pH. These fibrils are cytotoxic to model neuronal cells in a similar fashion to those formed by the proteins implicated in neurodegenerative diseases. The addition of small quantities of 2,2,2-trifluoroethanol accelerates fibril formation by uperin 3.5, and is correlated with a structural stabilisation induced by this co-solvent. Uperin 3.5 fibril formation and the associated cellular toxicity are inhibited by the polyphenol (-)-epigallocatechin-3-gallate (EGCG). Furthermore, EGCG rapidly dissociates fully formed uperin 3.5 fibrils. Ion mobility-mass spectrometry reveals that uperin 3.5 adopts various oligomeric states in solution. Combined, these observations imply that the mechanism of membrane permeability by uperin 3.5 is related to its fibril-forming properties.Antonio N. Calabrese, Yanqin Liu, Tianfang Wang, Ian F. Musgrave, Tara L. Pukala, Rico F. Tabor, Lisandra L. Martin, John A. Carver, John H. Bowi

Defective lung macrophage function in lung cancer +/- chronic obstructive pulmonary disease (COPD/emphysema)-mediated by cancer cell production of PGE2?

In chronic obstructive pulmonary disease (COPD/emphysema) we have shown a reduced ability of lung and alveolar (AM) macrophages to phagocytose apoptotic cells (defective ‘efferocytosis’), associated with evidence of secondary cellular necrosis and a resultant inflammatory response in the airway. It is unknown whether this defect is present in cancer (no COPD) and if so, whether this results from soluble mediators produced by cancer cells. We investigated efferocytosis in AM (26 controls, 15 healthy smokers, 37 COPD, 20 COPD+ non small cell lung cancer (NSCLC) and 8 patients with NSCLC without COPD) and tumor and tumor-free lung tissue macrophages (21 NSCLC with/13 without COPD). To investigate the effects of soluble mediators produced by lung cancer cells we then treated AM or U937 macrophages with cancer cell line supernatant and assessed their efferocytosis ability. We qualitatively identified Arachidonic Acid (AA) metabolites in cancer cells by LC-ESI-MSMS, and assessed the effects of COX inhibition (using indomethacin) on efferocytosis. Decreased efferocytosis was noted in all cancer/COPD groups in all compartments. Conditioned media from cancer cell cultures decreased the efferocytosis ability of both AM and U937 macrophages with the most pronounced effects occurring with supernatant from SCLC (an aggressive lung cancer type). AA metabolites identified in cancer cells included PGE2. The inhibitory effect of PGE2 on efferocytosis, and the involvement of the COX-2 pathway were shown. Efferocytosis is decreased in COPD/emphysema and lung cancer; the latter at least partially a result of inhibition by soluble mediators produced by cancer cells that include PGE2.Francis C. Dehle, Violet R. Mukaro, Craig Jurisevic, David Moffat, Jessica Ahern, Greg Hodge, Hubertus Jersmann, Paul N. Reynolds, Sandra Hodg

"Gå som en spårhund där sanningen trampade" : en studie i Tomas Tranströmers poetikk

Problemstillingen for denne avhandlingen er: Hva er Tomas Tranströmers poetikk slik denne dels nedfelles i diktningen og dels fremgår av forfatterens egne ord i intervjuer, brev og annen prosa? Jeg spør helt enkelt: Hva er Tranströmers dikteriske prosjekt, og hvordan gjør han det? I lys av problemstillingen er målet med avhandlingen å utforme en slags akademisk håndbok om forfatterskapet, dvs. å utforme og definere en inngangsport til Tranströmers poesi på en så poengtert og presis måte som mulig. Min utgangshypotese er at det dikteriske prosjektet dypest sett springer ut av en inderlig sannhetsstreben hos dikteren. Det er sannheten om den mangslungne eksistensen dikteren søker å nå i og med sin sentrallyriske diktning. At det forholder seg slik, er det første jeg gir belegg for. Dernest redegjør jeg for Tranströmers sannhetsforståelse rent generelt. Hva mener han med sin bruk av begrepet sannhet? Etter mitt syn har han en dobbeltsidig sannhetsforståelse: Sannhet blir dels betraktet som overflatefenomen og dels som dybdefenomen. Sannheten på et overflatenivå er at tilværelsen er formet, strukturert og bestemt av ulike typer grenser, slik som grensen mellom subjekt og objekt f.eks.. På et dyperliggende nivå er sannheten en annen: Det som på overflaten virker grenseinndelt, hører egentlig sammen på en eller annen måte. Sannhet innebærer altså generelt sett enten eksistensielle grenser eller overskridelser av de samme grensene. På bakgrunn av denne tosidige sannhetsforståelsen redegjør jeg i neste omgang for Tranströmers idémessige posisjon og verdensbilde. Et sentralt poeng her er å vise hvordan hans generelle sannhetsforståelse basert på begrepsparet grense-overskridelse preger hele hans virkelighetsforståelse for øvrig, om det så gjelder hans syn på religion, virkelighetens ontologi, tidsfenomenet, historien, mennesket, samfunnet eller kunsten. Kort sammenfattet argumenterer jeg for øvrig for at Tranströmers idémessige posisjon er grunnleggende romantisk. Verdensbildet er så vel idealistisk som monistisk, organisk og panteistisk, forklarer jeg. Det romantiske preget er imidlertid oppveid og balansert med en realistisk saklighet som hører 1900-tallet til. I siste del forsøker jeg å vise hvordan Tranströmers verdensbilde og idémessige posisjon blir omgjort til diktning. Jeg hevder her at Tranströmers diktning er preget av fem ulike poetisk grep som hver for seg og til sammen bidrar til å få frem hans monistiske, organiske og panteistiske verdensbilde