Psychologie
The glucocorticoid (GC) cortisol, main mediator of the hypothalamic-pituitary-adrenal axis, has many implications in metabolism, stress response and the immune system. GC function is mediated mainly via the glucocorticoid receptor (GR) which binds as a transcription factor to glucocorticoid response elements (GREs). GCs are strong immunosuppressants and used to treat inflammatory and autoimmune diseases. Long-term usage can lead to several irreversible side effects which make improved understanding indispensable and warrant the adaptation of current drugs. Several large scale gene expression studies have been performed to gain insight into GC signalling. Nevertheless, studies at the proteomic level have not yet been made. The effects of cortisol on monocytes and macrophages were studied in the THP-1 cell line using 2D fluorescence difference gel electrophoresis (2D DIGE) combined with MALDI-TOF mass spectrometry. More than 50 cortisol-modulated proteins were identified which belonged to five functional groups: cytoskeleton, chaperones, immune response, metabolism, and transcription/translation. Multiple GREs were found in the promoters of their corresponding genes (+10 kb/-0.2 kb promoter regions including all alternative promoters available within the Database for Transcription Start Sites (DBTSS)). High quality GREs were observed mainly in cortisol modulated genes, corroborating the proteomics results. Differential regulation of selected immune response related proteins were confirmed by qPCR and immuno-blotting. All immune response related proteins (MX1, IFIT3, SYWC, STAT3, PMSE2, PRS7) which were induced by LPS were suppressed by cortisol and belong mainly to classical interferon target genes. Mx1 has been selected for detailed expression analysis since new isoforms have been identified by proteomics. FKBP51, known to be induced by cortisol, was identified as the strongest differentially expressed protein and contained the highest number of strict GREs. Genomic analysis of five alternative FKBP5 promoter regions suggested GC inducibility of all transcripts. 2D DIGE combined with 2D immunoblotting revealed the existence of several previously unknown FKBP51 isoforms, possibly resulting from these transcripts. Additionally multiple post-translational modifications were found, which could lead to different subcellular localization in monocytes and macrophages as seen by confocal microscopy. Similar results were obtained for the different cellular subsets of human peripheral blood mononuclear cells (PBMCs). FKBP51 was found to be constitutively phosphorylated with up to 8 phosphosites in CD19+ B lymphocytes. Differential Co-immunoprecipitation for cytoplasm and nucleus allowed us to identify new potential interaction partners. Nuclear FKBP51 was found to interact with myosin 9, whereas cytosolic FKBP51 with TRIM21 (synonym: Ro52, Sjögren`s syndrome antigen). The GR has been found to interact with THOC4 and YB1, two proteins implicated in mRNA processing and transcriptional regulation. We also applied proteomics to study rapid non-genomic effects of acute stress in a rat model. The nuclear proteome of the thymus was investigated after 15 min restraint stress and compared to the non-stressed control. Most of the identified proteins were transcriptional regulators found to be enriched in the nucleus probably to assist gene expression in an appropriate manner. The proteomic approach allowed us to further understand the cortisol mediated response in monocytes/macrophages. We identified several new target proteins, but we also found new protein variants and post-translational modifications which need further investigation. Detailed study of FKBP51 and GR indicated a complex regulation network which opened a new field of research. We identified new variants of the anti-viral response protein MX1, displaying differential expression and phosphorylation in the cellular compartments. Further, proteomics allowed us to follow the very early effects of acute stress, which happen prior to gene expression. The nuclear thymocyte proteome of restraint stressed rats revealed an active preparation for subsequent gene expression. Proteomics was successfully applied to study differential protein expression, to identify new protein variants and phosphorylation events as well as to follow translocation. New aspects for future research in the field of cortisol-mediated immune modulation have been added.
Das Stresshormon Cortisol zeigt einen starken zirkadianen Rhythmus mit hohen Cortisolwerten nach dem morgendlichen Erwachen und niedrigen Werten am Abend. Die vorliegende Arbeit legt die Grundlagen dafür, dass der Cortisolspiegel nach dem Erwachen (Cortisol Awakening Response) zukünftig Bestandteil einer multimodalen Diagnostik stressbezogener Erkrankungen werden kann. Zu diesem Zweck werden besonders messmethodische Aspekte des Cortisol Awakening Response (CAR) dargestellt und eingehend diskutiert. Der Einfluss verschiedener konfundierender Variablen wurde in einer quantitativen Metaanalyse untersucht. Ein gesonderter Abschnitt beschreibt verschiedene Möglichkeiten der statistischen Analyse des CAR. Zu diesem Zweck wurden verschiedene statistische Kennwerte generiert und deren Reliabilitäten und Interkorrelationen an einem empirischen Datensatz untersucht. In dieser Arbeit werden auch Normwerte für die einzelnen statistischen Kennwerte des CAR angegeben.
Fibromyalgia is a disorder of unknown etiology characterized by widespread, chronic musculoskeletal pain of at least three month duration and pressure hyperalgesia at specific tender points on clinical examination. The disorder is accompanied by a multitude of additional symptoms such as fatigue, sleep disturbances, morning stiffness, depression, and anxiety. In terms of biological disturbances, low cortisol concentrations have been repeatedly observed in blood and urine samples of fibromyalgia patients, both under basal and stress-induced conditions. The aim of this dissertation was to investigate the presence of low cortisol concentrations (hypocortisolism) and potential accompanying alterations on sympathetic and immunological levels in female fibromyalgia patients. Beside the expected hypocortisolism, a higher norepinephrine secretion and lower natural killer cell levels were found in the patient group compared to a control group consisting of healthy, age-matched women. In addition, an increased activity of some pro-inflammatory markers was observed thus leading to alterations in the balance of pro-/anti-inflammatory activity. The results underline the relevance of simultaneous investigations of interacting bodily systems for a better understanding of underlying biological mechanisms in stress-related disorders.
Objective: Only 20-25% of the variance for the two to four-fold increased risk of developing breast cancer among women with family histories of the disease can be explained by known gene mutations. Other factors must exist. Here, a familial breast cancer model is proposed in which overestimation of risk, general distress, and cancer-specific distress constitute the type of background stress sufficient to increase unrelated acute stress reactivity in women at familial risk for breast cancer. Furthermore, these stress reactions are thought to be associated with central adiposity, an independent well-established risk factor for breast cancer. Hence, stress through its hormonal correlates and possible associations with central adiposity may play a crucial role in the etiology of breast cancer in women at familial risk for the disease. Methods: Participants were 215 healthy working women with first-degree relatives diagnosed before (high familial risk) or after age 50 (low familial risk), or without breast cancer in first-degree relatives (no familial risk). Participants completed self-report measures of perceived lifetime breast cancer risk, intrusive thoughts and avoidance about breast cancer (Impact of Event Scale), negative affect (Profile of Mood States), and general distress (Brief Symptom Inventory). Anthropometric measurements were taken. Urine samples during work, home, and sleep were collected for assessment of cortisol responses in the naturalistic setting where work was conceptualized as the stressful time of the day. Results: A series of analyses indicated a gradient increase of cortisol levels in response to the work environment from no, low, to high familial risk of breast cancer. When adding breast cancer intrusions to the model with familial risk status predicting work cortisol levels, significant intrusion effects emerged rendering the familial risk group non-significant. However, due to a lack of association between intrusions and cortisol in the low and high familial risk group separately, as well as a significant difference between low and high familial risk on intrusions, but not on work cortisol levels, full mediation of familial risk group effects on work cortisol by intrusions could not be established. A separate analysis indicated increased levels of central but not general adiposity in women at high familial risk of breast cancer compared to the low and no risk groups. There were no significant associations between central adiposity and cortisol excretion. Conclusion: A hyperactive hypothalamus-pituitary-adrenal axis with a more pronounced excretion of its end product cortisol, as well as elevated levels of central but not overall adiposity in women at high familial risk for breast cancer may indicate an increased health risk which expands beyond that of increased breast cancer risk for these women.
In jüngerer Zeit wurde in der neuroendokrinologischen Forschung das Phänomen eines Hypocortisolismus bei verschiedenen Störungen, die mit Stress assoziiert sind, beschrieben. Insbesondere bei der Posttraumatischen Belastungsstörung (PTSD) wurde eine verringerte adrenale Aktivität berichtet. Aber auch bei Patienten mit verschiedenen körperlichen Beschwerden wurden ähnliche neuroendokrine Veränderungen gefunden. Dazu zählen unter anderem das Fibromyalgiesyndrom (FMS) und chronische Unterbauchbeschwerden (CUBB). Die Mechanismen, welche dem Hypocortisolismus zugrunde liegen, sind bislang sowohl für die PTSD als auch für stressabhängige körperliche Beschwerden nicht abschließend geklärt. Weiterhin besteht Unklarheit darüber, inwieweit eine Vergleichbarkeit dieser Mechanismen zwischen den verschiedenen Störungsbildern besteht. Die Entstehung und Aufrechterhaltung dieser Erkrankungen scheinen somit ein sehr komplexes Zusammenspiel verschiedener Faktoren darzustellen. Andererseits weisen die Überlappungen hinsichtlich symptomatologischer, psychologischer und endokrinologischer Variablen zwischen PTSD, FMS und CUBB auf die Existenz störungsübergreifender Subgruppen hin. In der vorliegenden Studie wurden psychologische und endokrinologische Auffälligkeiten bei PTSD, FMS und CUBB weiter untersucht. Vorrangiges Ziel war, zu überprüfen, inwieweit störungsübergreifende Subgruppen mit vergleichbaren psychoendokrinologischen Auffälligkeiten bestehen. Insgesamt wurden 59 Patientinnen mittels verschiedener endokrinologischer Tests untersucht und mit 30 gesunden Kontrollfrauen verglichen. Mit einer Clusteranalyse konnten drei unabhängige störungsübergreifende Subgruppen identifiziert werden, die sich hinsichtlich ihrer Reaktionen in den endokrinologischen Tests unterschieden. Es konnte somit gezeigt werden, dass es sich bei den untersuchten Störungsgruppen weder um eine Störungsfamilie mit identischen endokrinen Auffälligkeiten noch um isolierte, d.h. distinkte, von einander unabhängige Erkrankungen handelt. Vielmehr scheinen störungsübergreifende Subgruppen zu bestehen. Weitere Studien sollten die gefunden Muster replizieren und gegebenenfalls erweitern.