Redox-regulation of PTPs; mechanisms and impact on PDGFR signaling

Protein tyrosine phosphatases (PTPs) are reversibly oxidized upon activation of platelet-derived growth factor receptor beta (PDGFβR). Dys-regulation of the PDGFβR signaling pathway is associated with several diseases, including cancers and cardiovascular disease, and is thus a known driver of disease progression. Ligand dependent PDGFβR phosphorylation stimulates cell proliferation and migration. The aim of this thesis was to elucidate redox-regulatory mechanisms of protein tyrosine phosphatases impacting on PDGFβR signaling. In Paper I, we analysed effects of mitochondria-derived ROS on PTP oxidation in models of hypoxia and hypoxia/re-oxygenation (H/R) in vitro and in vivo. We found an increase in PTP oxidation of multiple PTPs, including SHP-2, PTP1B and DEP-1, after exposure of NIH3T3 fibroblasts to H/R. An increase in total PTP oxidation and SHP-2 was also seen in rat cardiomyoblasts after H/R. Furthermore, H/R induced a delay of PDGFR dephosphorylation and also an antioxidant sensitive activation of downstream effectors ERK1/2. In addition, H/R enhanced PDGF-dependent cytoskeletal re-arrangements, which could be abolished by antioxidant treatment. Finally, we found an increase in total PTP oxidation and SHP2 oxidation in tissue extracts from an ex-vivo model of rat heart ischemia-reperfusion. In paper II, we studied expression and activity of PDGFβR pathway components in human pulmonary artery smooth muscle cells (hPASMC) subjected to hypoxia. We show that hypoxia- induced HIF-1α in hPASMC, both in vivo and in vitro, negatively regulate expression of PDGFβR associated PTPs, including PTP1B, DEP-1, TC-PTP and SHP2. The negatively regulation of these PDGFβR-associated PTPs occurred together with an enhanced PDGF receptor activation and an increase in both proliferation and migration of hPASMC. In paper III, we found that p66Shc dependent mitochondrial derived ROS contribute to inactivation of the PDGFβR associated PTPs PTP1B and SHP-2 upon ligand stimulation. In addition, deletion of p66Shc reduced downstream intracellular signaling after PDGF-BB stimulation. Furthermore, p66Shc KO cells displayed a decrease in migratory response to PDGF-BB treatment. In the final study paper IV, we studied the reactivation of oxidized PTPs and its impact on PDGFβR signaling. We showed that cells lacking expression of thioredoxin reductase 1 (TrxR1) displayed an increase in oxidation of PTP1B but not of SHP-2. Furthermore, in vivo oxidized PTP1B was re-activated by addition of Trx system components to cell lysates, whereas SHP-2 was not re-activated. Oxidized recombinant PTP1B was also re-activated by treatment with Trx system components while SHP-2 remained largely unaffected. Intriguingly, the Trx related protein TRP14 also reactivated PTP1B but not SHP-2. Furthermore, PDGFβR phosphorylation and signaling was enhanced in Txnrd1-/- fibroblasts leading to an enhanced proliferative response after PDGF-BB stimulation

Selective activation of oxidized PTP1B by the thioredoxin system modulates PDGF-ß receptor tyrosine kinase signaling

The inhibitory reversible oxidation of protein tyrosine phosphatases (PTPs) is an important regulatory mechanism in growth factor signaling. Studies on PTP oxidation have focused on pathways that increase or decrease reactive oxygen species levels and thereby affect PTP oxidation. The processes involved in reactivation of oxidized PTPs remain largely unknown. Here the role of the thioredoxin (Trx) system in reactivation of oxidized PTPs was analyzed using a combination of in vitro and cell-based assays. Cells lacking the major Trx reductase TrxR1 (Txnrd1-/-) displayed increased oxidation of PTP1B, whereas SHP2 oxidation was unchanged. Furthermore, in vivo-oxidized PTP1B was reduced by exogenously added Trx system components, whereas SHP2 oxidation remained unchanged. Trx1 reduced oxidized PTP1B in vitro but failed to reactivate oxidized SHP2. Interestingly, the alternative TrxR1 substrate TRP14 also reactivated oxidized PTP1B, but not SHP2. Txnrd1-depleted cells displayed increased phosphorylation of PDGF-ß receptor, and an enhanced mitogenic response, after PDGF-BB stimulation. The TrxR inhibitor auranofin also increased PDGF-ß receptor phosphorylation. This effect was not observed in cells specifically lacking PTP1B. Together these results demonstrate that the Trx system, including both Trx1 and TRP14, impacts differentially on the oxidation of individual PTPs, with a preference of PTP1B over SHP2 activation. The studies demonstrate a previously unrecognized pathway for selective redox-regulated control of receptor tyrosine kinase signaling

Neurotrophins and neurotrophin receptors in pulmonary sarcoidosis - granulomas as a source of expression

<p>Abstract</p> <p>Background</p> <p>Pulmonary sarcoidosis is an inflammatory disease, characterized by an accumulation of CD4⁺lymphocytes and the formation of non-caseating epithelioid cell granulomas in the lungs. The disease either resolves spontaneously or develops into a chronic disease with fibrosis. The neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) have been suggested to be important mediators of inflammation and mediate tissue remodelling. In support of this, we have recently reported enhanced NGF levels in the airways of patients with pulmonary sarcoidosis. However, less is known about levels of BDNF and NT-3, and moreover, knowledge in the cellular sources of neurotrophins and the distribution of the corresponding neurotrophin receptors in airway tissue in sarcoidosis is lacking.</p> <p>Methods</p> <p>The concentrations of NGF, BDNF and NT-3 in bronchoalveolar lavage fluid (BALF) of 41 patients with newly diagnosed pulmonary sarcoidosis and 27 healthy controls were determined with ELISA. The localization of neurotrophins and neurotrophin receptors were examined by immunohistochemistry on transbronchial lung biopsies from sarcoidosis patients.</p> <p>Results</p> <p>The sarcoidosis patients showed significantly enhanced NT-3 and NGF levels in BALF, whereas BDNF was undetectable in both patients and controls. NT-3 levels in BALF were found higher in patients with non-Löfgren sarcoidosis as compared to patients with Löfgren's syndrome, and in more advanced disease stage. Epithelioid cells and multinucleated giant cells within the sarcoid granulomas showed marked immunoreactivity for NGF, BDNF and NT-3. Also, immunoreactivity for the neurotrophin receptor TrkA, TrkB and TrkC, was found within the granulomas. In addition, alveolar macrophages showed positive immunoreactivity for NGF, BDNF and NT-3 as well as for TrkA, TrkB and TrkC.</p> <p>Conclusions</p> <p>This study provides evidence of enhanced neurotrophin levels locally within the airways of patients with sarcoidosis. Findings suggest that sarcoid granuloma cells and alveolar macrophages are possible cellular sources of, as well as targets for, neurotrophins in the airways of these patients.</p

Leadership in social work

The purpose of this paper is to increase our understanding of what constitutes a good leadership in social work. The questions I've chosen are: what competencies are important for a leader in social work? What values and/or ethical principles are important to have/follow as a leader in social work? To get an insight into how you look at leadership in social work in a Swedish context, I have made interviews with fifteen leaders, each and every one of them working within the field of social work. In the interviews I ask open-ended, yet somewhat directed questions, to figure out what characteristics and principles these leaders regard as the foundation of a good leadership. To broaden the studies knowledgebase I've also read, analyzed and summarized previous studies on the subject. I will use these studies to broaden my discussion by looking at similarities. The leaders have been given a chance to communicate what characterizes a good leader within social work. The main goal of the interviews has been to uncover what characteristics, competencies and ethical values/principles these leaders regard as important. The results clearly show that it's important with education and competencies within leadership. They also show the importance of other competencies such as and understanding of economics, psychology, behavioral sciences, conflict management, etc. Furthermore the interviews show that the softer values and ethical principles such as equal rights, empathy and having a salutogenic perspective are given more focus than education and competence. This study shows that leaders in social work praise the transformal leadership model and strive to act according to it themselves. The majority of Swedish leaders in my study don't think there's a difference between leadership in social work versus leadership in other professions. This separates Swedish and American leaders, where a majority of the latter think there's a difference.Syftet med denna uppsats är att skapa en ökad förståelse kring vad som är ett bra ledarskap inom socialt arbete. De frågeställningar jag valt är: vilka kompetenser är viktiga för en ledare inom socialt arbete? Vilka värderingar och/eller etiska principer är viktiga att ha/följa som ledare inom socialt arbete? För att få en inblick i hur man ser på ledarskap inom socialt arbete i en svensk kontext så valde jag att intervjua femton enhetschefer, samtliga verksamma inom socialt arbete i Sverige. Intervjuerna ställer öppna, men något riktade frågor för att ta reda på vilka egenskaper och principer dessa enhetschefer anser ligga till grund för ett bra ledarskap. För att skapa en starkare grund har jag även läst tidigare studier på ämnet, analyserat och sammanfattat de viktigaste punkterna i dessa för att sedan redovisa dem och återkomma till beröringspunkter i diskussionsdelen. Genom intervjuer har enhetscheferna fått delge sin syn på vad som kännetecknar en bra ledare inom socialt arbete. Intervjuerna har främst ämnat svara på frågor kring vilka egenskaper, kompetenser och etiska värderingar/principer som anses vara viktiga för en ledare inom socialt arbete. Intervjuerna visar tydligt att det enligt enhetscheferna är viktigt med utbildning och kompetens inom framförallt ledarskap men även inom ämnen som ekonomi, psykologi, beteendevetenskap, konflikthantering, m.m. Vidare visar intervjuerna att mjuka värden ges större utrymme än kompetens och intervjupersonerna lägger mer fokus vid etiska principer som alla människors lika värden, lyhördhet, empatisk förmåga och se saker på ett salutogent vis. Slutsatser som kan dras är att ledare inom socialt arbete i såväl Sverige som USA främst eftersträvar och lovordar det transformella ledarskapet och alla de kompetenser och etiska värderingar som detta innebär. Majoriteten av de svenska enhetschefer i min studie anser ej att det är någon skillnad på ledarskap inom socialt arbete kontra andra professioner, vilket skiljer sig från den amerikanska studie som lyfts i tidigare forskning

Attitude toward physical education amongst senior high school teachers : A qualitative investigation

Uppsatsen undersöker vilken inställning gymnasielärare har till ämnet idrott och hälsa. Tanken om detta har skapats från de historier som författaren har tagit del av, där lärare i ämnet idrott och hälsa har berättat hur andra lärare har haft en problematisk syn på ämnet och behandlat det därefter. För att få svar på ifall inställningen går att hitta på flera gymnasier, så används semi-strukturerade intervjuer som metod. Urvalet består av gymnasielärare på svenska gymnasieskolor som också utgör en heterogen grupp; det vill säga att där blandas ålder, kön, yrkeserfarenhet, ämnen de undervisar och är utbildade i samt vilken skola de tillhör. Svaren från intervjuerna presenteras i en resultatdel, och resultaten visar att majoriteten av de intervjuade, men även andra personer, verkar ha en problematisk syn på ämnet idrott och hälsa. I analysdelen så används professionsteorin, elevers syn, idrottsämnets historia samt läroplaner för att tolka resultatet. Slutsatsen är att skälen till en problematisk syn på ämnet kan vara en bristfällig förståelse för ämnets innehåll, en fördom från de gamla kursplanerna som inte har uppdaterats med innehållet i dagens kursplaner eller en oförståelse för att se praktisk erfarenhets hemhörighet i professionen

Studies on neurotrophins in inflammatory pulmonary diseases

Asthma, sarcoidosis and chronic obstructive pulmonary disease (COPD) are inflammatory pulmonary diseases, all being characterized by tissue inflammation, tissue damage (airway remodeling) and loss of lung function. In order to increase possibilities to find new biomarkers, drug targets and better treatment options for the patients, further research on the underlying mechanisms driving the inflammation and airway remodeling in these diseases are required. In the present thesis, a family of mediators, the neurotrophins, were studied to elucidate how these mediators are involved in the inflammatory and/or airway remodeling processes in asthma, sarcoidosis and COPD. The neurotrophin family consists of NGF, BDNF and NT-3, and these factors were initially characterized for their essential role as survival factors for nerve cells. However, they have been shown to display a far broader spectrum of functions, being mediators also involved in inflammation and tissue remodeling. Indeed, enhanced levels of neurotrophins have been found in several inflammatory conditions, including allergic diseases, such as asthma. The aim of this thesis was to elucidate if neurotrophin levels are altered in inflammatory pulmonary diseases other than asthma. Therefore, we set up studies to determine the protein levels of NGF, BDNF and NT-3 in bronchoalveolar lavage fluid (BALF) from patients with pulmonary sarcoidosis, patients with COPD, and relevant control subjects, including healthy non-smokers and current smokers with normal lung function. In addition, the aim was to study human bronchial smooth muscle cells, as possible target cells for the effects of the neurotrophins in the airways. We found that, similarly to asthma, sarcoidosis patients showed enhanced levels of both NGF and NT-3 in the airways as compared to healthy subjects. We also investigated the relation between levels of neurotrophins and clinical as well as inflammatory parameters, and found significant positive correlations between NGF and the concentration of lymphocytes and eosinophils, as well as inflammatory cytokines (IFN-γ, IL-4, IL-10, IL-12) in BALF of sarcoidosis patients. We found that the levels of NT-3 were higher in the sub-groups of patients with a higher risk of developing chronic disease and fibrosis, suggesting that NT-3 levels relate to the risk of developing chronic disease. To further elucidate cellular sources of, and possible targets for, neurotrophins, in the lungs of sarcoidosis patients, we performed immunohistochemistry on lung tissue biopsies. We found that granulomas expressed NGF and NT-3, as well as the corresponding neurotrophin receptors TrkA and TrkC, indicating that the granulomas are possible sources of the enhanced levels of neurotrophins in sarcoidosis, and that the neurotrophins may be able to mediate autocrine functions in the granuloma microenvironment. In contrast to sarcoidosis, COPD patients, as well as smokers with normal lung function, showed decreased levels of NGF and NT-3 in the airways as compared to healthy non-smokers. This indicated that cigarette-smoke exposure may impact neurotrophin content in human airways. Indeed, when exposing human lung fibroblasts in vitro to cigarette smoke extract, a down-regulation of neurotrophin expression was shown on both transcriptional- and protein levels. Studies on human bronchial smooth muscle cells revealed that they are possible target cells for the neurotrophins in the airways, as they express the neurotrophin receptors TrkA, -B, and C, and we showed that NGF, BDNF and NT-3 enhanced the secretion of the tissue degrading enzyme MMP-9, but not MMP-2. Additionally, BDNF and NT-3, but not NGF, stimulated cell migration. These results support the concept that neurotrophins have pro-fibrotic properties and may be involved in airway tissue remodeling, such as that seen in asthma. Taken together, the results of the present thesis contributes to a better understanding regarding the involvement of neurotrophins in inflammatory pulmonary diseases, and gives further support to the concept of neurotrophins as being mediators involved in airway inflammation and tissue remodeling