| 1. |
- Nowak, Christoph, 1986-
(författare)
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Insulin Resistance : Causes, biomarkers and consequences
- 2017
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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.
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| 2. |
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| 3. |
- Senkowski, Wojciech
(författare)
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High-throughput screening using multicellular tumor spheroids to reveal and exploit tumor-specific vulnerabilities
- 2017
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- High-throughput drug screening (HTS) in live cells is often a vital part of the preclinical anticancer drug discovery process. So far, two-dimensional (2D) monolayer cell cultures have been the most prevalent model in HTS endeavors. However, 2D cell cultures often fail to recapitulate the complex microenvironments of in vivo tumors. Monolayer cultures are highly proliferative and generally do not contain quiescent cells, thought to be one of the main reasons for the anticancer therapy failure in clinic. Thus, there is a need for in vitro cellular models that would increase predictive value of preclinical research results. The utilization of more complex three-dimensional (3D) cell cultures, such as multicellular tumor spheroids (MCTS), which contain both proliferating and quiescent cells, has therefore been proposed. However, difficult handling and high costs still pose significant hurdles for application of MCTS for HTS.In this work, we aimed to develop novel assays to apply MCTS for HTS and drug evaluation. We also set out to identify cellular processes that could be targeted to selectively eradicate quiescent cancer cells. In Paper I, we developed a novel MCTS-based HTS assay and found that nutrient-deprived and hypoxic cancer cells are selectively vulnerable to treatment with inhibitors of mitochondrial oxidative phosphorylation (OXPHOS). We also identified nitazoxanide, an FDA-approved anthelmintic agent, to act as an OXPHOS inhibitor and to potentiate the effects of standard chemotherapy in vivo. Subsequently, in Paper II we applied the high-throughput gene-expression profiling method for MCTS-based drug screening. This led to discovery that quiescent cells up-regulate the mevalonate pathway upon OXPHOS inhibition and that the combination of OXPHOS inhibitors and mevalonate pathway inhibitors (statins) results in synergistic toxicity in this cell population. In Paper III, we developed a novel spheroid-based drug combination-screening platform and identified a set of molecules that synergize with nitazoxanide to eradicate quiescent cancer cells. Finally, in Paper IV, we applied our MCTS-based methods to evaluate the effects of phosphodiesterase (PDE) inhibitors in PDE3A-expressing cell lines.In summary, this work illustrates how MCTS-based HTS yields potential to reveal and exploit previously unrecognized tumor-specific vulnerabilities. It also underscores the importance of cell culture conditions in preclinical drug discovery endeavors.
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| 4. |
- Bjermo, Helena, 1981-
(författare)
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Dietary Fatty Acids and Inflammation : Observational and Interventional Studies
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Dietary fat quality influences the risk of type 2 diabetes and cardiovascular disease. A low-grade inflammation is suggested to contribute to the disease development, often accompanied by obesity. Whereas n-3 polyunsaturated fatty acids (PUFA) have been considered anti-inflammatory, n-6 PUFA have been proposed to act pro-inflammatory. Saturated fatty acids (SFA) act pro-inflammatory in vitro. This thesis aimed to investigate effects of different fatty acids on low-grade inflammation in observational and interventional studies. In Paper I and II, fatty acid composition in serum cholesterol esters was used as objective marker of dietary fat quality and related to serum C-reactive protein (CRP) and other circulating inflammatory markers in two population-based cohorts, conducted in middle-aged men and elderly men and women, respectively. In Paper III and IV, the impact of diets differing in fat quality on inflammation and oxidative stress was investigated in randomised controlled studies, in subjects with metabolic syndrome and abdominal obesity. In Paper I and II, a low proportion of linoleic acid (18:2 n-6) in serum was associated with higher CRP concentrations, indicating that a low intake of vegetable fats may be related to low-grade inflammation. High CRP concentrations were also associated with high proportions of palmitoleic (16:1) and oleic (18:1) acids and high stearoyl coenzymeA desaturase index, possibly reflecting altered fat metabolism and/or high SFA intake in this population. When comparing two high-fat diets rich in either saturated or monounsaturated fat, and two low-fat diets with or without long-chain n-3 PUFA supplementation during 12 weeks (Paper III), no differences in inflammation or oxidative stress markers were observed. Moreover, a 10-week intervention (Paper IV) with high linoleic acid intake showed no adverse effects on inflammation or oxidative stress. Instead, interleukin-1 receptor antagonist and tumor necrosis factor receptor-2 decreased after linoleic acid intake compared with a diet high in SFA. The results in this thesis indicate that dietary n-6 PUFA found in vegetable fats is associated with lower inflammation marker levels, and to some extent reduces systemic inflammation when compared with SFA. Supplementation of n-3 PUFA did not exert any systemic anti-inflammatory effects, maybe due to a relatively low dose.
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| 5. |
- Latini, Francesco, M.D. 1982-
(författare)
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Significance of white matter anatomy in interpreting features and behaviour of low-grade gliomas and implications for surgical treatment
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Diffuse gliomas are extremely heterogeneous tumours characterized by slow growth but extensive infiltration. Their kinetic features reflect the complex interaction over time with the surrounding brain, influencing treatment planning and outcome. Indeed, resection of diffuse gliomas present a surgical challenge due to their invasiveness and the preferential location in eloquent regions. White matter bundles are the main eloquent limit to surgical resection, but this anatomical-functional information cannot be predicted preoperatively on the individual level. The incomplete description of the human brain connectome, the complex application of pathological/lesion model to the brain connectomic organization, and the underestimated role of white matter anatomy in radiological classification systems are among the major limitations for the comprehension of the glioma/white matter interaction. The overall aim of this thesis was to explore a new approach and new techniques to study the glioma/white matter interaction. A combination of white matter dissection and diffusion tensor tractography (DTT) was used to describe the connectomic organization of two major temporo-occipital connections, the inferior and the middle longitudinal fasciculus. This information was applied to patients with diffuse gliomas, demonstrating how white matter analysis was important to decode patient specific cognitive and language impairment. A new classification system for diffuse gliomas, the Brain-Grid, was created, merging local radiological anatomy with a DTT atlas for infiltration analysis. This standardized radiological tool provided information on subcortical extension (tumour invasiveness), speed, and preferential direction of glioma progression. Applied to a larger cohort of patients, differences were detected between diffuse gliomas subtypes. Tumour invasiveness and the preferential location, type, and extent of white matter involvement differed, impacting overall survival. Regional differences in white matter infiltration were detected among five major white matter bundles, and possible favourable morphological and diffusion features were investigated with transmission electron microscopy and DTT. Fibre diameter, myelin thickness, and the organization of the white matter fibres were different in regions with high infiltration frequency, providing a possible link to the preferential location of diffuse gliomas. Finally, the white matter connectivity, tumour-induced neuroplasticity, clinical and demographic information, preoperative assessment (neuropsychological and language evaluation) were compared with intraoperative findings during awake surgery. Neuropsychological impairment was associated with more invasive tumours and a higher risk of the intraoperative finding of eloquent tumour. The pattern of early cortical neuroplasticity seemed exhausted at the time of diagnosis, with age as a factor predicting the neuroplasticity potential. The combined use of these new techniques revealed new insights into the glioma/white matter interaction. The results provided in this thesis, describe a new way to structure the multidisciplinary perioperative management of these patients. This new information may improve the functional outcome at the individual level, resulting in prolonged survival for adults with diffuse gliomas.
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| 6. |
- Berg, Marie, 1955
(författare)
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Genuine Caring in Caring for the Genuine : Childbearing and high risk as experienced by women and midwives
- 2002
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The experience of pregnancy and childbirth is a central life event with special implications for women at high risk. This thesis describes the meaning of pregnancy, childbirth and midwifery care in four qualitative interview studies based on the lifeworld theory. Women were interviewed during pregnancy and within one week after childbirth. Midwives were interviewed concerning midwifery care for women at high risk. In an intervention study, childbirth experience as reported through a post partum questionnaire was compared between women receiving standard care and women who had formulated a birth plan preceded by a questionnaire on their expectations and feelings about childbirth.The findings emphasise that childbearing women at high risk live in an extremely vulnerable situation. The vulnerability is obvious in the use of an individual birth plan, where negative feelings become more frequent in women at high risk than in those with normal pregnancy and childbirth. During pregnancy the women feel a moral commitment towards the child, including feelings of objectification and of exaggerated responsibility. During an obstetrically complicated childbirth the essential meaning is the women’s desire to be recognised and affirmed as individual persons. Like women with normal pregnancy and childbirth, they need an emotionally present midwife who sees, give trust and supports.Good midwifery care of childbearing women at high risk is synthesised as "genuine caring in caring for the genuine". The ethos of caring constitutes the basis of caring. Women’s transition during pregnancy and childbirth is described as a genuinely natural process. Midwives have a special responsibility to encourage and preserve this process within women at high risk. The caring relationship is the core and the most essential tool in the care. Distinctive features in the midwifery care are embodied knowledge, physical as well as emotional presence, sensitivity, a mutual dialogue including shared control between midwife and woman, and confirmation and support of the genuine in each woman. The midwifery care is a struggle and a balance between natural and medical perspectives.
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| 7. |
- Aulin, Cecilia, 1979-
(författare)
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Extracellular Matrix Based Materials for Tissue Engineering
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The extracellular matrix is (ECM) is a network of large, structural proteins and polysaccharides, important for cellular behavior, tissue development and maintenance. Present thesis describes work exploring ECM as scaffolds for tissue engineering by manipulating cells cultured in vitro or by influencing ECM expression in vivo. By culturing cells on polymer meshes under dynamic culture conditions, deposition of a complex ECM could be achieved, but with low yields. Since the major part of synthesized ECM diffused into the medium the rate limiting step of deposition was investigated. This quantitative analysis showed that the real rate limiting factor is the low proportion of new proteins which are deposited as functional ECM. It is suggested that cells are pre-embedded in for example collagen gels to increase the steric retention and hence functional deposition. The possibility to induce endogenous ECM formation and tissue regeneration by implantation of growth factors in a carrier material was investigated. Bone morphogenetic protein-2 (BMP-2) is a growth factor known to be involved in growth and differentiation of bone and cartilage tissue. The BMP-2 processing and secretion was examined in two cell systems representing endochondral (chondrocytes) and intramembranous (mesenchymal stem cells) bone formation. It was discovered that chondrocytes are more efficient in producing BMP-2 compared to MSC. The role of the antagonist noggin was also investigated and was found to affect the stability of BMP-2 and modulate its effect. Finally, an injectable gel of the ECM component hyaluronan has been evaluated as delivery vehicle in cartilage regeneration. The hyaluronan hydrogel system showed promising results as a versatile biomaterial for cartilage regeneration, could easily be placed intraarticulary and can be used for both cell based and cell free therapies.
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| 8. |
- Bhandage, Amol K., 1988-
(författare)
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Glutamate and GABA signalling components in the human brain and in immune cells
- 2016
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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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| 9. |
- Bin Kaderi, Mohamed Arifin, 1978-
(författare)
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Assessment of Novel Molecular Prognostic Markers in Chronic Lymphocytic Leukemia
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The clinical course of chronic lymphocytic leukemia (CLL) is highly heterogeneous, which has prompted the search for biomarkers that can predict prognosis in this disease. The IGHV gene mutation status and certain genomic aberrations have been identified as reliable prognostic markers of clinical outcome for this disorder. However, the search for more feasible prognostic markers in CLL is still being pursued. Recently, certain single nucleotide polymorphisms (SNPs) in the GNAS1, BCL2 and MDM2 genes and the RNA expression levels of the LPL, ZAP70, TCL1, CLLU1 and MCL1 genes were suggested as novel prognostic markers in CLL. In papers I-III, we performed genotyping analyses of the GNAS1 T393C, BCL2 -938C>A and MDM2 SNP309 polymorphisms in 268-418 CLL patients and related the genotypes with clinical data. Association studies between the polymorphisms and established prognostic markers (i.e. IGHV mutation status, genomic aberrations, CD38 expression) were also performed. Our studies did not find any significant relationship between these SNPs with either clinical outcome or other known prognostic markers in CLL. In paper IV, we measured the RNA expression levels of LPL, ZAP70, TCL1, CLLU1 and MCL1 in 252 CLL cases and correlated these levels with clinical outcome. Here, we verified that high expression of all these RNA-based markers, except MCL1, were associated with an unfavourable prognosis. We also confirmed a close relationship between IGHV mutation status and the RNA-based markers, especially for LPL and CLLU1 expression. Among the RNA-based markers, multivariate analysis revealed LPL expression as the strongest independent prognostic marker for overall survival and time to treatment. Furthermore, the RNA-based markers could add further prognostic information to established markers in subgroups of patients, with LPL expression status giving the most significant results. In summary, data from papers I-III could not verify the GNAS1 T393C, BCL2 -938C>A and MDM2 SNP309 polymorphisms as prognostic markers in CLL. Future SNP markers must hence be confirmed in large, independent cohorts before being proposed as prognostic marker in CLL. In paper IV, we conclude that LPL expression appears to be the strongest among the RNA-based markers for CLL prognostication. Further efforts to standardize LPL quantification are required before it can be applied in the clinical laboratory to predict clinical outcome in this disease.
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| 10. |
- Centanni, Maddalena, 1994-
(författare)
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Model-based evaluation of biomarkers for dose-individualization in oncology
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In contemporary cancer care, several issues are garnering increasing attention. First, significant inter-individual variability among patients challenges the effectiveness of a uniform dosing approach. Second, the escalating costs of treatments necessitate careful consideration when selecting doses and other clinical modalities, including biomarkers, while balancing economic constraints. The objective of this thesis was to evaluate techniques for tailoring doses and guiding clinical decisions for cancer patients through the development and implementation of various models, with the aim of improving treatment outcomes in terms of both efficacy and safety. Through a model-based framework integrating sunitinib pharmacokinetics (PK), adverse events, biomarkers, tumor dynamics and their correlation with overall survival, different treatment schedules and biomarkers for dose individualization were explored. Based on the proposed threshold values, neutrophil count (ANC) and the biomarker sVEGFR-3 were identified as offering the best balance between safety and efficacy for sunitinib in gastro-intestinal stromal tumors (GIST) and could thus serve as viable guides for dose individualization in clinical practice. Given its routine measurement, dose adjustments guided by ANC may be preferable in clinical settings. The feasibility of utilizing diastolic blood pressure (dBP) for personalized dose optimization of tyrosine-kinase inhibitors in clinical settings is constrained due to its reliance on repeated measurements taken at consistent intervals. For axitinib and sunitinib, model-based predictions using multiple clinical measurements were more accurate than single sample measurements. For drugs with high unexplained inter-individual variability (IIV), low residual variability (RUV), and low inter-occasional variability (IOV), therapeutic drug monitoring (TDM) provided a more accurate measure of exposure. Conversely, for drugs with low IIV and high RUV and IOV, pharmacogenetic profiling was more suitable. However, the prevalence of pharmacogenetic subtypes and the challenge of measuring exposure metrics like AUC through limited sampling also influence these approaches.This research further emphasizes how model structure affects the outcomes of cost-effectiveness analyses and consequently the potential implications for regulatory decisions. Although creating mechanistic models for these analyses demands substantial initial effort, the growing need for model-based analyses in drug approval is likely to make these models more accessible for future compounds. Moreover, such models are expected to be more biologically plausible and therefore more reflective of reality and offer flexibility for exploring alternative dosages with limited additional effort.Using model-based assessments, the relationship between the PK and PK-pharmacodynamic (PKPD) profiles of adverse events arising from therapies for acute lymphocytic leukemia were established. For PEG-asparaginase, the PK model categorized 93% of patients who experienced inactivation against PEG-asparaginase as having an increased clearance, and 86% of patients who did not experience hypersensitivity as maintaining stable clearance throughout their asparaginase treatment. This approach marks a potential method for predicting inactivation by identifying early changes in clearance. For vincristine, model-informed precision dosing was shown to reduce the incidence of vincristine-induced peripheral neuropathy (VIPN) from 62.1% to 53.9%, though the clinical impact remains modest.
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