Immune regulatory mechanisms in endocrine autoimmune disorders

Ulike mekanismer eksisterer for at immunforsvaret skal lære å skille det som er eget fra det som er fremmed, også kjent som immunologisk toleranse. I noen tilfeller brytes denne toleransen, noe som kan føre til at enkelte utvikler autoimmune sykdommer. Det som er interessant er at pasienter med immunsvikt også ofte utvikler autoimmunitet. Dette kan tyde på at en feil i immunforsvarets regulerende mekanismer kan være en felles risikofaktor. Regulatoriske T celler (Tregs) har evnen til å dempe en immunrespons og er veldig viktige for å opprettholde toleranse. Et annet fellestrekk er sirkulerende autoantistoffer rettet mot cytokiner, som er funnet i både immunsvikt og autoimmunitet, men hvordan disse mekanismene virker sammen for å regulere toleranse er fortsatt uvisst. I denne avhandlingen har ulike metoder blitt brukt for å studere regulatoriske mekanismer i to modellsykdommer for autoimmunitet. I artikkel I undersøkte vi transkriptomet og proteomet til in vitro ekspanderte Tregs fra pasienter med den monogene sykdommen autoimmunt polyendokrint syndrom type 1 (APS-1). Vi brukte enkelt-celle og T celle reseptor (TCR) sekvensering for å se på forskjell i genuttrykk mellom pasienter og kontroller. Samlet viste resultatene få forskjeller i transkriptomet til ekspanderte Tregs og TCR repertoar mellom pasienter og kontroller. Analyse med flow- og massecytometri viste at CD4+CD25+FOXP3+CD127- populasjonen var signifikant redusert hos APS-1 pasienter sammenlignet med friske kontroller. For å undersøke Tregs sin evne til å dempe proliferering av responder celler, satte vi opp et flowcytometri-basert suppresjonsassay, som viste at ekspanderte Tregs fra APS-1 pasienter kan hemme celledeling like bra som ekspanderte Tregs isolert fra friske kontroller. I artikkel II ville vi undersøke frekvensen av Tregs i PBMC og den suppressive kapasiteten til ekspanderte Tregs i pasienter med den endokrine autoimmune sykdommen primær binyrebarksvikt, med polygent arvemønster. I tillegg har vi sett på forskjeller mellom Tregs fra disse pasientene og friske kontroller på transkripsjonsnivå og studert mitokondriell respirasjon. Ingen forskjeller ble funnet i frekvensen av Tregs i PBMC når pasienter og kontroller ble sammenlignet. Suppresjonsassayet viste at det ikke er forskjeller mellom pasienter med primær binyrebarksvikt og friske kontroller når det kommer til å dempe proliferering av responder celler. Vi gikk videre med RNA-sekvensering for å se om vi kunne finne forskjeller på gennivå. Igjen viste resultatene få forskjeller mellom pasienter og kontroller, men flere gener uttrykt i mitokondriene var oppregulert i pasientgruppen. For å studere mitokondriell funksjon, satte vi opp et Seahorse metabolisme assay for å måle raten av oksygenforbruk. Assayet viste at Tregs fra pasienter hadde en basal-, maksimal-, og ikke-mitokondriell respirasjon, i tillegg til proton lekkasje og reservekapasitet på nivå med målinger for friske kontroller. I artikkel III ønsket vi å finne ut om cytokin-autoantistoff-screening kan brukes til å identifisere pasienter med mono- eller oligogen årsak til sykdom. Et enzyme-linked immunosorbent assay (ELISA) ble satt opp for å måle autoantistoffer mot interleukin (IL) 22, mens et etablert radioaktivitetsassay ble brukt til å mål autoantistoffer mot interferon (IFN)-ω. Positive pasienter ble deretter sekvensert med et panel bestående av immunerelaterte gener etter standard protokoll fra Medisinsk avdeling, Haukeland Universitetssykehus. Med denne strategien identifiserte vi fire nye pasienter med APS-1 og to pasienter med sykdomsfremkallende mutasjoner i henholdsvis NFKB2 og CTLA4. I tillegg fant vi sjeldne varianter i IKZF2, LAT, LCK, STAT1, JAK3, RAG1, TERC, TNFAIP3 og en variant av AIRE som er assosiert med primær binyrebarksvikt, varianter som kan være forårsakende for de kliniske manifestasjonene hos pasientene. Resultatene fra denne avhandlingen bidrar med økt kunnskap om regulatoriske mekanismer involvert i immunologisk toleranse og viser at screening for autoantistoffer er et verdifullt diagnostisk verktøy. Videre åpner funnene for studier som karakteriserer monogene sykdommer med autoantistoffer mot cytokiner og andre regulatoriske mekanismer.The immune system is, through various regulatory mechanisms, taught to discriminate self from non-self, known as immunological tolerance. Sometimes breakdown of tolerance occur, leading to autoimmune disease in susceptible individuals. Of note, patients with immunodeficiencies also commonly present with autoimmune components, indicating that impaired regulatory mechanisms might be a shared risk factor. Cells with suppressive function, particularly regulatory T cells (Tregs), are crucial for regulation of tolerance and alterations in the Treg compartment can contribute to disease development. Another shared factor is circulating autoantibodies targeting cytokines, which have been described in both immunodeficiency and autoimmunity. However, details on how these mechanisms come together and regulate immune tolerance still remain elusive. In the present work, several approaches have been used to study immune regulatory mechanisms in human model disorders of autoimmunity. In paper I, we aimed at investigating the proteome and transcriptome of in vitro expanded Tregs from patients with the monogenic autoimmune polyendocrine syndrome type 1 (APS-1). Single-cell and T cell receptor (TCR) sequencing was performed to examine differential gene expression and analyze the TCR repertoire. Overall, the expanded Treg transcriptome and TCR repertoire were found to be similar between patients and controls. Multipanel flow and mass cytometry revealed a significantly reduced CD4+CD25+FOXP3+CD127- population in patients compared to healthy donors. An optimized flow cytometry based Treg suppression assay revealed that APS-1 expanded Tregs could suppress responder cell proliferation to the same degree as expanded Tregs from healthy controls. In paper II, we wanted to examine the frequency of Tregs in PBMC and explore in vitro expanded Treg suppressive function, transcriptome and mitochondrial function in autoimmune primary adrenal insufficiency (PAI), an endocrine autoimmune disorder with a polygenic inheritance pattern. No differences were observed in the frequency of Tregs in PBMC when comparing patients and healthy controls. The Treg suppression assay showed that expanded PAI Tregs were able to suppress proliferation of responder cells similar to expanded healthy control Tregs. However, the RNA sequencing revealed an upregulation of mitochondrial genes in the patient group. To further examine mitochondrial function, a Seahorse mitochondria stress test assay was performed. Patient cells displayed a basal, maximal and non-mitochondrial respiration, in addition to proton leak and spare capacity, comparable to that of healthy controls. In paper III, we wanted to examine whether screening of patients with known endocrine disorders for cytokine autoantibodies could identify individuals with a possible monogenic or oligogenic cause of disease. An enzyme-linked immunosorbent assay (ELISA) was optimized and performed to screen patients with autoimmune PAI for autoantibodies against interleukin (IL) 22, while an established radio-immuno assay (RIA) was used to screen patients with a variety of endocrine autoimmune disorders for autoantibodies against interferon (IFN)-ω. Positive patients were subsequently sequenced with an immune deficiency pipeline at Haukeland University Hospital. This strategy identified four new patients with APS-1 and two patients with disease-causing mutations in NFKB2 and CTLA4, respectively. Further, rare variants were detected in the genes IKZF2, LAT, LCK, STAT1, JAK3, RAG1, TERC, TNFAIP3 and one variant of AIRE associated with autoimmune PAI, all with potential roles as causative factors for the clinical manifestations seen in these patients. Together, the findings from this thesis furthers our knowledge on immune regulatory mechanisms in autoimmune disease and highlights the use of autoantibody screenings as a part of the diagnostic toolbox. Our findings open for future studies involving further characterization of monogenic diseases with cytokine autoantibodies and of immune regulatory mechanisms.Doktorgradsavhandlin

War, PowerPoint, and hypnotised chickens: Standards and templates at work in a military staff

This article discusses concepts to explore decision-making processes in a military headquarters. Military planning is commonly perceived as a systematic and structured approach to organising ways and means to achieve military ends. While standardised procedures and decision-making tools are crucial for large military organisations to function efficiently, these devices are not neutral. Routines within the staff organisation carry implicit beliefs shaping the perception of war as a managerial problem with an optimal solution that can be elicited through a process and presented in a bulleted list. By examining organisational outcomes as socio-material assemblages, this article sheds light on how daily routines influence potential solutions and shape what can and cannot be thought. Conventional approaches in organisational studies have either overlooked the role of organisational tools or studied them as a matter of technology adoption. The entanglement of the social and material in organisational life should be observed and described empirically to understand how order is reconstructed after it has broken down.&nbsp

Pharmacological treatment of the T-cell receptor signalling pathway in autoimmune Addison's disease - A pilotstudy with abatacept (CTLA4-Ig)

FARM399/05HMATF-FAR

Home-school collaboration in Namibian secondary schools

Bacheloroppgave, grunnskolelærerutdanning, 2017.Norsk: Denne bachelor-oppgaven handler om hvordan skole-hjem samarbeidet fungerer ved to ulike skoler i Namibia. Oppgaven belyser hvilke samarbeidsformer som utarter seg, så vel som hvilke nivåer av samarbeid som finner sted ved skolene. Gjennom å se på ulike oppfatninger av samarbeidet på skolene, forsøkes dette å bli forklart sett i lys av foreldrenes kapital og skolekultur. Undersøkelsene er gjort gjennom intervjuer og observasjoner. Resultatene viste at skole-hjem samarbeidet foregikk forholdsvis likt på begge skolene, men ble av informantene oppfattet forskjellig på skole 1 og skole 2. Hvorvidt samarbeidet ble oppfattet som godt, ble påvirket av foreldrenes kapital og skolekulturen ved de enkelte skolene.English: This Bachelor thesis contains a study of home-school collaboration at two different schools in Namibia. The paper describes forms of collaboration and which level of collaboration that takes place at the schools. The paper tries to understand the different opinions on the collaborations, by giving insights on the parental cultural capital as well as school culture. Insights in this study has been found through interviews and observations. The results show us that the home-school collaboration takes similar forms in both schools, but the opinions however, differed in school 1 and school 2. In what extend the collaboration was regarded as good, was influenced by parental cultural capital as well as the school culture

Stosowanie środków psychostymulujących w leczeniu wspomagającym i paliatywnym pacjentów z chorobą nowotworową

Środki psychostymulujące są od dawna stosowane w leczeniu pacjentów z chorobami psychicznymi oraz somatycznymi. Potrzeba kontrolowania i przeciwdziałania różnorodnym stresującym objawom występującym w przebiegu choroby nowotworowej oraz podczas jej leczenia sprawiła, że leki te włączono do leczenia wspomagającego i paliatywnego u pacjentów z nowotworami. Badano wykorzystanie psychostymulantów w leczeniu takich objawów jak zmęczenie w przebiegu choroby nowotworowej, zaburzenia funkcji poznawczych, depresja oraz sedacja. W poniższej pracy oceniono farmakologię metylfenidatu, amfetaminy i premoliny oraz innych psychostymulantów, takich jak kofeina czy też nowy środek somnolityczny - modafinil. W pracy wykorzystano badania oceniające skutki działania tych leków u pacjentów z chorobą nowotworową

Genetic variation and cognitive dysfunction in opioid-treated patients with cancer

Background and purpose The effects of single‐nucleotide polymorphisms (SNP s) on the cognitive function of opioid‐treated patients with cancer until now have not been explored, but they could potentially be related to poor functioning. This study aimed at identifying associations between SNP s of candidate genes, high opioid dose, and cognitive dysfunction. Methods Cross‐sectional multicenter study (European Pharmacogenetic Opioid Study, 2005–2008); 1586 patients; 113 SNP s from 41 genes. Inclusion criteria: cancer, age ≥18 year, opioid treatment, and available genetic data. Cognitive assessment by Mini‐Mental State Examination (MMSE ). Analyses: SNP s were rejected if violation of Hardy–Weinberg equilibrium (P < 0.0005), or minor allele frequency <5%; patients were randomly divided into discovery sample (2/3 for screening) and validation sample (1/3 for confirmatory test); false discovery rate of 10% for determining associations (Benjamini–Hochberg method). Co‐dominant, dominant, and recessive models were analyzed by Kruskal–Wallis and Mann–Whitney tests. Results In the co‐dominant model significant associations (P < 0.05) between MMSE scores and SNP s in the HTR 3E , TACR 1 , and IL 6 were observed in the discovery sample, but the replication in the validation sample did not confirm it. Associations between MMSE scores among patients receiving ≥400 mg morphine equivalent dose/day and SNP s in TNFRSF 1B , TLR 5 , HTR 2A , and ADRA 2A were observed, but they could not be confirmed in the validation sample. After correction for multiple testing, no SNP s were significant in the discovery sample. Dominant and recessive models also did not confirm significant associations. Conclusions The findings did not support influence of those SNP s analyzed to explain cognitive dysfunction in opioid‐treated patients with cancer

Regulatory T cells in autoimmune primary adrenal insufficiency

Primary adrenal insufficiency (PAI) is most often caused by an autoimmune destruction of the adrenal cortex resulting in failure to produce cortisol and aldosterone. The aetiology is thought to be a combination of genetic and environmental risk factors, leading to breakdown of immunological tolerance. Regulatory T cells (Tregs) are deficient in many autoimmune disorders, but it is not known whether they contribute to development of PAI. We aimed to investigate the frequency and function of naive and expanded Tregs in patients with PAI and polyendocrine syndromes compared to age- and gender-matched healthy controls. Flow cytometry was used to assess the frequency and characterize functional markers of blood Tregs in PAI (N = 15). Expanded Treg suppressive abilities were assessed with a flow cytometry based suppression assay (N = 20), while bulk RNA-sequencing was used to examine transcriptomic differences (N = 16) and oxygen consumption rate was measured by a Seahorse cell metabolic assay (N = 11). Our results showed that Treg frequency and suppressive capacity were similar between patients and controls. An increased expression of killer-cell leptin-like receptors and mitochondrial genes was revealed in PAI patients, but their expanded Tregs did not display signs of mitochondrial dysfunction. Our findings do not support a clear role for Tregs in the contribution of PAI development.publishedVersio

Is admittance to specialised palliative care among cancer patients related to sex, age and cancer diagnosis?:A nation-wide study from the Danish Palliative Care Database (DPD)

BACKGROUND: Specialised palliative care (SPC) takes place in specialised services for patients with complex symptoms and problems. Little is known about what determines the admission of patients to SPC and whether there are differences in relation to institution type. The aims of the study were to investigate whether cancer patients’ admittance to SPC in Denmark varied in relation to sex, age and diagnosis, and whether the patterns differed by type of institution (hospital-based palliative care team/unit, hospice, or both). METHODS: This was a register-based study of adult patients living in Denmark who died from cancer in 2010–2012. Data sources were the Danish Palliative Care Database, Danish Register of Causes of Death and Danish Cancer Registry. The associations between the explanatory variables (sex, age, diagnosis) and admittance to SPC were investigated using logistic regression. RESULTS: In the study population (N = 44,548) the overall admittance proportion to SPC was 37%. Higher odds of overall admittance to SPC were found for women (OR = 1.23; 1.17–1.28), younger patients (<40 compared with 80+ years old) (OR = 6.44; 5.19–7.99) and patients with sarcoma, pancreatic and stomach cancers, whereas the lowest were for patients with haematological malignancies. The higher admission found for women was most pronounced for hospices compared to hospital-based palliative care teams/units, whereas higher admission of younger patients was more pronounced for hospital-based palliative care teams/units. Patients with brain cancer were more often admitted to hospices, whereas patients with prostate cancer were more often admitted to hospital-based palliative care teams/units. CONCLUSION: It is unlikely that the variations in relation to sex, age and cancer diagnoses can be fully explained by differences in need. Future research should investigate whether the groups having the lowest admittance to SPC receive sufficient palliative care elsewhere

Patient reported upper gastro-intestinal symptoms associated with fractionated image-guided conformal radiotherapy for metastatic spinal cord compression

Background and purpose Palliative radiotherapy is given to sustain or improve quality of life for patients with advanced cancer. Radiotherapy may however result in symptomatic side effects, which may affect the patient negatively. This prospective longitudinal study of 30 patients aimed at investigating the incidence and severity of early toxicity, particularly focusing on dysphagia, esophagitis and mucositis, following fractionated radiotherapy for cervical and thoracic metastatic spinal cord compression (MSCC), as well as determining the relationship between esophageal dose and early upper gastro-intestinal symptoms. Materials and methods Thirty patients receiving radiotherapy of 3Gyx10 for MSCC were included in the study. Patients were assessed for a total of 7 weeks from onset of radiotherapy using the Edmonton Symptom Assessment System (ESAS) questionnaire. Upper gastro-intestinal symptoms and severity were assessed from the tenth and eleventh question section of the ESAS questionnaire of “other problems” and how much this affected them. The relationships between the mean and maximum esophageal doses and incidence of dysphagia, esophagitis or mucositis were estimated and dose response curves determined. Results Eleven patients reported esophageal symptoms (average duration eleven days, range 1–18 days). Incidence of esophageal toxicity in patients treated at Th8 or above was 79 percent, while no patients treated below Th8 reported any symptoms (p < 0.001). Furthermore, 2 out of 3 patients irradiated at the cervical region reported substantial changes in taste sensation. Risk of symptoms correlated with both mean and maximum esophageal dose and may be a useful tool in planning radiotherapy for MSCC, potentially reducing early upper gastro-intestinal toxicity