| 1. |
- 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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| 2. |
- Cutas, Daniela, 1978, et al.
(författare)
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Legal imperialism in the regulation of stem cell research and therapy: the problem of extraterritorial jurisdiction
- 2010
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Ingår i: Capps BJ & Campbell AV (eds.). CONTESTED CELLS: Global Perspectives on the Stem Cell Debate. - London : Imperial College Press. - 9781848164376 ; , s. 95-119
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- Countries worldwide have very different national regulations on human embryonic stem (ES) cell research, informed by a range of ethical values. Some countries find reason to extend the applicability of their regulations on such research to its citizens when they visit other countries. Extraterritorial jurisdiction has recently been identified as a potential challenge towards global regulation of ES cell research. This chapter explores the implications and impact of extraterritorial jurisdiction and global regulation of ES cell research on researchers, clinicians and national health systems, and how this may affect patients. The authors argue that it would make ethical sense for ES cell restrictive countries to extend its regulations on ES cell research beyond its borders, because, if these countries really consider embryo destruction to be objectionable on the basis on the status of the embryo, then they ought to count it morally on par with murder (and thus have a moral imperative to protect embryos from the actions of its own citizens). However, doing so could lead to a legal situation that would result in substantial harm to central values in areas besides research, such as health care, the job market, basic freedom of movement, and strategic international finance and politics. Thus, it seems that restrictive extraterritorial jurisdiction in respect to ES cell research would be deeply problematic, given that the ethical permissibility of ES cell research is characterised by deep and wide disagreement.
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| 3. |
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| 4. |
- Carlsson, Stina K., 1982-
(författare)
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Effects of adenosine and acetylcholine on the lacrimal gland
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- A balanced tear film is essential for a healthy ocular surface. Insufficient tear production may result in dry eye, a common disorder in the elderly population. Dry eye causes significant discomfort in the patients and may lead to visual impairment and ocular infections. The lacrimal gland secretes water, proteins and electrolytes to the aqueous layer of the tear film. Lacrimal gland secretion is tightly regulated by e.g. neuronally released acetylcholine. The effect of acetylcholine on lacrimal gland secretion was recently found to be potentiated by adenosine. Adenosine is an important signaling molecule acting upon the adenosine receptors: A1, A2A, A2B and A3.The aim of this thesis was to study effects of adenosine and acetylcholine on intracellular signaling pathways and lacrimal gland secretion. Cholinergic stimulation of secretion was shown to be regulated by the mitogen activated protein kinase p38, a protein previously not known to be involved in exocrine secretion. p38 was activated in response to cholinergic stimulation and inhibition of p38 significantly diminished cholinergic secretion.When investigating adenosine effects, potentiation of cholinergic secretion was observed by activation of the A2B receptor in addition to the previously studied A1 receptor. An A2 receptor agonist increased cholinergic rabbit lacrimal gland protein secretion at several concentrations. The increase was inhibited by antagonism of the A2B receptor, but not the A2A receptor. When investigating the intracellular signaling pathways following adenosine and acetylcholine receptor activation, adenosine was shown to increase of cAMP levels. An additional increase in cAMP levels was observed after parallel adenosine and cholinergic receptor activation. Inhibition of Ca2+ release from the endoplasmic reticulum had inhibitory effects of cholinergic stimulation of secretion. In addition, the expression of adenosine receptors in a mouse model of autoimmune dry eye was investigated. The results showed a lymphocyte dependent upregulation of A2A receptors in diseased mice compared to controls.In conclusion, the results in this thesis provide significant contributions in the search of dry eye therapeutics through studies of adenosine and acetylcholine receptor activation.
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| 5. |
- 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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| 6. |
- Yu, Di, 1985-
(författare)
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Adenovirus for Cancer Therapy : With a Focus on its Surface Modification
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Adenovirus serotype 5 (Ad5) is widely used as an oncolytic agent for cancer therapy. However, its infectivity is highly dependent on the expression level of coxsackievirus-adenovirus receptor (CAR) on the surface of tumor cells. We engineered Ad5 virus with the protein transduction domain (PTD) from the HIV-1 Tat protein (Tat-PTD) inserted in the hypervariable region 5 (HVR5) of the hexon protein in the virus capsid. Tat-PTD-modified Ad5 shows a dramatically increased transduction level of CAR-negative cells and bypassed fiber-mediated transduction. It also overcomes the fiber-masking problem, which is caused by release of excess fiber proteins from infected cells. To achieve specific viral replication in neuroblastoma and neuroendocrine tumor cells, we identified the secretogranin III (SCG3) promoter and constructed an adenovirus Ad5PTD(ASH1-SCG3-E1A) wherein E1A gene expression is controlled by the SCG3 promoter and the achaete-scute complex homolog 1 (ASH1) enhancer. This virus shows selective and efficient killing of neuroblastoma cell lines in vitro, and delays human neuroblastoma xenograft tumor growth on nude mice. To further enhance the viral oncolytic efficacy, we also switched the fiber 5 to fiber 35 to generate Ad5PTDf35. This vector shows dramatically increased transduction capacity of primary human cell cultures including hematopoietic cells and their derivatives, pancreatic islets and exocrine cells, mesenchymal stem cells and primary tumor cells including primary cancer initiating cells. Ad5PTDf35-based adenovirus could be a useful platform for gene delivery and oncolytic virus development. Viral oncolysis alone cannot completely eradicate tumors. Therefore, we further armed the Ad5PTDf35-D24 virus with a secreted form of Helicobacter pylori Neutrophil Activating Protein (HP-NAP). Expression of HP-NAP recruits neutrophils to the site of infection, activates an innate immune response against tumor cells and provokes a Th1-type adaptive immune response. Established tumor on nude mice could be completely eradicated in some cases after treatment with this virus and the survival of mice was significantly prolonged.
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| 7. |
- Halldórsdóttir, Anna Margrét, 1973-
(författare)
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Genetic and Epigenetic Profiling of Mantle Cell Lymphoma and Chronic Lymphocytic Leukemia
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Mantle cell lymphoma (MCL) and chronic lymphocytic leukemia (CLL) both belong to the group of mature B-cell malignancies. However, MCL is typically clinically aggressive while the clinical course of CLL varies. CLL can be divided into prognostic subgroups based on IGHV mutational status and into multiple subsets based on closely homologous (stereotyped) B-cell receptors. In paper I we investigated 31 MCL cases using high-density 250K single-nucleotide polymorphism arrays and gene expression arrays. Although most copy-number aberrations (CNAs) were previously reported in MCL, a novel deletion was identified at 20q (16%) containing the candidate tumor suppressor gene ZFP64. A high proliferation gene expression signature was associated with poor prognosis, large CNAs, 7p gains and 9q losses. Losses at 1p/8p/13q/17p were associated with increased genomic complexity. In paper II we sequenced exons 4 to 8 of the TP53 gene in 119 MCL cases. 17p copy-number status was known from previous studies or determined by real-time quantitative polymerase chain reaction. TP53 mutations were detected in 14% of cases and were strongly associated with poor survival while 17p deletions were more common (32%) but did not predict survival. In papers III and IV we applied high-resolution genomic 27K methylation arrays to 20 MCL and 39 CLL samples. In paper III MCL displayed a homogenous methylation profile without correlation with the proliferation signature whereas MCL was clearly separated from CLL. Gene ontology analysis revealed enrichment of developmental genes, in particular homeobox transcription factor genes, among targets methylated in MCL. In paper IV we compared three different stereotyped CLL subsets: #1 (IGHV unmutated), #2 (IGHV3-21) and #4 (IGHV mutated). Many genes were differentially methylated between each two subsets and immune response genes (e.g. CD80 and CD86) were enriched among genes methylated in subset #1 but not in subsets #2/#4.In summary, CNAs were frequent and not random in MCL. Specific CNAs correlated with a high proliferation gene expression signature or genomic complexity. TP53 mutations predicted short survival whereas 17p deletions did not. A high proliferation signature was not associated with differential DNA methylation in MCL, which demonstrated a homogeneous methylation pattern. In contrast, genomic methylation patterns differed between MCL and CLL and between stereotyped CLL subsets.
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| 8. |
- Jörnsten, Rebecka, 1971, et al.
(författare)
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Network modeling of the transcriptional effects of copy number aberrations in glioblastoma
- 2011
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Ingår i: Molecular Systems Biology. - : EMBO. - 1744-4292. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- DNA copy number aberrations (CNAs) are a hallmark of cancer genomes. However, little is known about how such changes affect global gene expression. We develop a modeling framework, EPoC (Endogenous Perturbation analysis of Cancer), to (1) detect disease-driving CNAs and their effect on target mRNA expression, and to (2) stratify cancer patients into long- and short-term survivors. Our method constructs causal network models of gene expression by combining genome-wide DNA- and RNA-level data. Prognostic scores are obtained from a singular value decomposition of the networks. By applying EPoC to glioblastoma data from The Cancer Genome Atlas consortium, we demonstrate that the resulting network models contain known disease-relevant hub genes, reveal interesting candidate hubs, and uncover predictors of patient survival. Targeted validations in four glioblastoma cell lines support selected predictions, and implicate the p53-interacting protein Necdin in suppressing glioblastoma cell growth. We conclude that large-scale network modeling of the effects of CNAs on gene expression may provide insights into the biology of human cancer. Free software in MATLAB and R is provided.
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| 9. |
- Uhlén, Mathias, et al.
(författare)
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The human secretome
- 2019
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Ingår i: Science Signaling. - : American Association for the Advancement of Science (AAAS). - 1945-0877 .- 1937-9145. ; 12:609
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Tidskriftsartikel (refereegranskat)abstract
- The proteins secreted by human cells (collectively referred to as the secretome) are important not only for the basic understanding of human biology but also for the identification of potential targets for future diagnostics and therapies. Here, we present a comprehensive analysis of proteins predicted to be secreted in human cells, which provides information about their final localization in the human body, including the proteins actively secreted to peripheral blood. The analysis suggests that a large number of the proteins of the secretome are not secreted out of the cell, but instead are retained intracellularly, whereas another large group of proteins were identified that are predicted to be retained locally at the tissue of expression and not secreted into the blood. Proteins detected in the human blood by mass spectrometry-based proteomics and antibody-based immuno-assays are also presented with estimates of their concentrations in the blood. The results are presented in an updated version 19 of the Human Protein Atlas in which each gene encoding a secretome protein is annotated to provide an open-access knowledge resource of the human secretome, including body-wide expression data, spatial localization data down to the single-cell and subcellular levels, and data about the presence of proteins that are detectable in the blood.
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| 10. |
- Piening, B. D., et al.
(författare)
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Integrative Personal Omics Profiles during Periods of Weight Gain and Loss
- 2018
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Ingår i: Cell Systems. - : Elsevier BV. - 2405-4712 .- 2405-4720. ; 6:2
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Tidskriftsartikel (refereegranskat)abstract
- Advances in omics technologies now allow an unprecedented level of phenotyping for human diseases, including obesity, in which individual responses to excess weight are heterogeneous and unpredictable. To aid the development of better understanding of these phenotypes, we performed a controlled longitudinal weight perturbation study combining multiple omics strategies (genomics, transcriptomics, multiple proteomics assays, metabolomics, and microbiomics) during periods of weight gain and loss in humans. Results demonstrated that: (1) weight gain is associated with the activation of strong inflammatory and hypertrophic cardiomyopathy signatures in blood; (2) although weight loss reverses some changes, a number of signatures persist, indicative of long-term physiologic changes; (3) we observed omics signatures associated with insulin resistance that may serve as novel diagnostics; (4) specific biomolecules were highly individualized and stable in response to perturbations, potentially representing stable personalized markers. Most data are available open access and serve as a valuable resource for the community.
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