| 1. |
- Ahlqvist, Emma, et al.
(author)
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High-resolution mapping of a complex disease, a model for rheumatoid arthritis, using heterogeneous stock mice
- 2011
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In: Human Molecular Genetics. - Oxford : Oxford University Press. - 0964-6906 .- 1460-2083. ; 20:15, s. 3031-3041
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Journal article (peer-reviewed)abstract
- Resolving the genetic basis of complex diseases like rheumatoid arthritis will require knowledge of the corresponding diseases in experimental animals to enable translational functional studies. Mapping of quantitative trait loci in mouse models of arthritis, such as collagen-induced arthritis (CIA), using F(2) crosses has been successful, but can resolve loci only to large chromosomal regions. Using an inbred-outbred cross design, we identified and fine-mapped CIA loci on a genome-wide scale. Heterogeneous stock mice were first intercrossed with an inbred strain, B10.Q, to introduce an arthritis permitting MHCII haplotype. Homozygous H2(q) mice were then selected to set up an F(3) generation with fixed major histocompatibility complex that was used for arthritis experiments. We identified 26 loci, 18 of which are novel, controlling arthritis traits such as incidence of disease, severity and time of onset and fine-mapped a number of previously mapped loci. © The Author 2011. Published by Oxford University Press. All rights reserved.
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| 2. |
- Ameri, Jacqueline, et al.
(author)
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FGF2 Specifies hESC-Derived Definitive Endoderm into Foregut/Midgut Cell Lineages in a Concentration-Dependent Manner.
- 2010
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In: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 28, s. 45-56
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Journal article (peer-reviewed)abstract
- Fibroblast growth factor (FGF) signaling controls axis formation during endoderm development. Studies in lower vertebrates have demonstrated that FGF2 primarily patterns the ventral foregut endoderm into liver and lung, whereas FGF4 exhibits broad anterior-posterior and left-right patterning activities. Furthermore, an inductive role of FGF2 during dorsal pancreas formation has been shown. However, whether FGF2 plays a similar role during human endoderm development remains unknown. Here, we show that FGF2 specifies hESC-derived definitive endoderm (DE) into different foregut lineages in a dosage-dependent manner. Specifically, increasing concentrations of FGF2 inhibits hepatocyte differentiation, whereas intermediate concentration of FGF2 promotes differentiation towards a pancreatic cell fate. At high FGF2 levels specification of midgut endoderm into small intestinal progenitors is increased at the expense of PDX1+ pancreatic progenitors. High FGF2 concentrations also promote differentiation towards an anterior foregut pulmonary cell fate. Finally, by dissecting the FGF receptor intracellular pathway that regulates pancreas specification, we demonstrate for the first time to our knowledge that induction of PDX1+ pancreatic progenitors relies on FGF2-mediated activation of the MAPK signaling pathway. Altogether, these observations suggest a broader gut endodermal patterning activity of FGF2 that corresponds to what has previously been advocated for FGF4, implying a functional switch from FGF4 to FGF2 during evolution. Thus, our results provide new knowledge of how cell fate specification of human DE is controlled - facts that will be of great value for future regenerative cell therapies.
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| 3. |
- Ehinger, Magnus, et al.
(author)
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Influence of CD4 or CD8 deficiency on collagen-induced arthritis
- 2001
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In: Immunology. - : Wiley. - 0019-2805 .- 1365-2567. ; 103:3, s. 291-300
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Journal article (peer-reviewed)abstract
- The role of T cells in the mouse collagen-induced arthritis (CIA) model for rheumatoid arthritis is not clarified, and different results have been reported concerning the role of CD4 and CD8 T cells. To address this issue, we have investigated B10.Q mice deficient for CD4 or CD8. The mice lacking CD4 were found to be less susceptible to disease, but not completely resistant, whereas the CD8 deficiency had no significant impact on the disease. No difference in the development of late occurring relapses was noted. Interestingly, the CD4-deficient mice had a severely reduced response to the glycosylated form of the immunodominant type II collagen (CII) 256–270 peptide whereas the response to the non-glycosylated peptide was not significantly different. Furthermore, CD4-deficient mice had lower antibody responses to CII, explaining the lower disease susceptibility. In comparison with previously reported results, it is apparent that the lack of CD4 molecules has a different impact on CIA if present on different genetic backgrounds, findings that could possibly be related to the occurrence of different disease pathways of CIA in different mouse strains.
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| 4. |
- Fischer, Yvonne, et al.
(author)
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NANOG reporter cell lines generated by gene targeting in human embryonic stem cells.
- 2010
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In: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 5:9
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Journal article (peer-reviewed)abstract
- BACKGROUND: Pluripotency and self-renewal of human embryonic stem cells (hESCs) is mediated by a complex interplay between extra- and intracellular signaling pathways, which regulate the expression of pluripotency-specific transcription factors. The homeodomain transcription factor NANOG plays a central role in maintaining hESC pluripotency, but the precise role and regulation of NANOG are not well defined. METHODOLOGY/PRINCIPAL FINDINGS: To facilitate the study of NANOG expression and regulation in viable hESC cultures, we generated fluorescent NANOG reporter cell lines by gene targeting in hESCs. In these reporter lines, the fluorescent reporter gene was co-expressed with endogenous NANOG and responded to experimental induction or repression of the NANOG promoter with appropriate changes in expression levels. Furthermore, NANOG reporter lines facilitated the separation of hESC populations based on NANOG expression levels and their subsequent characterization. Gene expression arrays on isolated hESC subpopulations revealed genes with differential expression in NANOG(high) and NANOG(low) hESCs, providing candidates for NANOG downstream targets hESCs. CONCLUSION/SIGNIFICANCE: The newly derived NANOG reporter hESC lines present novel tools to visualize NANOG expression in viable hESCs. In future applications, these reporter lines can be used to elucidate the function and regulation of NANOG in pluripotent hESCs.
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| 5. |
- Hansson, Ann-Sofie, et al.
(author)
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Relapsing polychondritis, induced in mice with matrilin 1, is an antibody- and complement-dependent disease
- 2004
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In: American Journal of Pathology. - New York, NY : Elsevier. - 0002-9440 .- 1525-2191. ; 164:3, s. 959-966
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Journal article (peer-reviewed)abstract
- Relapsing polychondritis is an autoimmune disease that affects cartilage in the ear, nose, and respiratory tract. A pathogenic immune response has been proposed and antibodies to several cartilage proteins are detected in sera from these patients. To investigate the role of the humoral immune response in relapsing polychondritis, we used the matrilin-1-induced relapsing polychondritis model. Mice deficient of B cells (muMT) and mice congenic at the complement factor 5, were immunized with matrilin-1, a cartilage-specific protein mainly detected in the tracheal cartilage. To investigate the binding properties and tissue selection of matrilin-1-specific antibodies we produced matrilin-1-specific B-cell hybridomas. Although 83% of the micro MT heterozygous mice developed respiratory distress and erosive chondritis in the respiratory tract, none of the B-cell-deficient mice were susceptible to disease. In addition, we show that complement factor 5 is important for the induction of matrilin-1-induced relapsing polychondritis. Monoclonal matrilin-1-specific antibodies injected into neonatal mice bound specifically to cartilage of the respiratory tract and adult B-cell-deficient mice injected with the same antibodies developed erosive chondritis in the respiratory tract. We conclude that relapsing polychondritis can be mediated by a pathway involving tissue-specific antibodies and complement activation.
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| 6. |
- Hjelmervik, Trond Ove R., et al.
(author)
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The influence of the NOD Nss1/Idd5 loci on sialadenitis and gene expression in salivary glands of congenic mice
- 2007
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In: Arthritis Research and Therapy. - : Springer Science and Business Media LLC. - 1478-6362 .- 1478-6354. ; 9:5
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Journal article (peer-reviewed)abstract
- The nonobese diabetic ( NOD) Nss1 and Idd5 loci have been associated with sialadenitis development in mice. In this study the NOD Nss1 and Idd5 loci were backcrossed onto the healthy control strain B10. Q by using the speed congenic breeding strategy, resulting in three congenic strains: B10. Q. Nss1, B10. Q. Nss1/Idd5 heterozygous and B10. Q. Nss1/Idd5 homozygous. We investigated the effects of the Nss1 and Idd5 loci on sialadenitis and gene expression in NOD congenic mice. One submandibular salivary gland from each mouse was used for histological analysis of sialadenitis, whereas the contralateral salivary gland was used for gene expression profiling with the Applied Biosystems Mouse Genome Survey chip v. 1.0. The results were validated using quantitative reverse transcriptase PCR. The NOD Nss1 and Idd5 loci had clear influence on the onset and progression of sialadenitis in congenic mice. Double congenic mice exhibited the most severe phenotype. We successfully identified several genes that are located in the NOD congenic regions to be differentially expressed between the congenic strains and the control strain. Several of these were found to be co-regulated, such as Stat1, complement component C1q genes and Tlr12. Also, a vast contingency of interferon-regulated genes ( such as Ltb, Irf7 and Irf8) and cytokine and chemokine genes ( such as Ccr7 and Ccl19) were differentially expressed between the congenic strains and the control strain. Over-representation of inflammatory signalling pathways was observed among the differentially expressed genes. We have found that the introgression of the NOD loci Nss1 and Idd5 on a healthy background caused sialadenitis in NOD congenic mouse strains, and we propose that genes within these loci are important factors in the pathogenesis. Furthermore, gene expression profiling has revealed several differentially expressed genes within and outside the NOD loci that are similar to genes found to be differentially expressed in patients with Sjogren's syndrome, and as such are interesting candidates for investigation to enhance our understanding of disease mechanisms and to develop future therapies.
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| 7. |
- Johannesson, Martina, et al.
(author)
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FGF4 and retinoic acid direct differentiation of hESCs into PDX1-expressing foregut endoderm in a time- and concentration-dependent manner.
- 2009
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In: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 4:3
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Journal article (peer-reviewed)abstract
- BACKGROUND: Retinoic acid (RA) and fibroblast growth factor 4 (FGF4) signaling control endoderm patterning and pancreas induction/expansion. Based on these findings, RA and FGFs, excluding FGF4, have frequently been used in differentiation protocols to direct differentiation of hESCs into endodermal and pancreatic cell types. In vivo, these signaling pathways act in a temporal and concentration-dependent manner. However, in vitro, the underlying basis for the time of addition of growth and differentiation factors (GDFs), including RA and FGFs, as well as the concentration is lacking. Thus, in order to develop robust and reliable differentiation protocols of ESCs into mature pancreatic cell types, including insulin-producing beta cells, it will be important to mechanistically understand each specification step. This includes differentiation of mesendoderm/definitive endoderm into foregut endoderm--the origin of pancreatic endoderm. METHODOLOGY/PRINCIPAL FINDINGS: Here, we provide data on the individual and combinatorial role of RA and FGF4 in directing differentiation of ActivinA (AA)-induced hESCs into PDX1-expressing cells. FGF4's ability to affect endoderm patterning and specification in vitro has so far not been tested. By testing out the optimal concentration and timing of addition of FGF4 and RA, we present a robust differentiation protocol that on average generates 32% PDX1(+) cells. Furthermore, we show that RA is required for converting AA-induced hESCs into PDX1(+) cells, and that part of the underlying mechanism involves FGF receptor signaling. Finally, further characterization of the PDX1(+) cells suggests that they represent foregut endoderm not yet committed to pancreatic, posterior stomach, or duodenal endoderm. CONCLUSION/SIGNIFICANCE: In conclusion, we show that RA and FGF4 jointly direct differentiation of PDX1(+) foregut endoderm in a robust and efficient manner. RA signaling mediated by the early induction of RARbeta through AA/Wnt3a is required for PDX1 expression. Part of RA's activity is mediated by FGF signaling.
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| 8. |
- Johannesson, Martina, et al.
(author)
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Gene expression profiling of arthritis using a QTL chip reveals a complex gene regulation of the Cia5 region in mice.
- 2005
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In: Genes and Immunity. - : Springer Science and Business Media LLC. - 1476-5470 .- 1466-4879. ; 6:7, s. 575-583
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Journal article (peer-reviewed)abstract
- One of the major quantitative trait loci (QTLs) associated with arthritis in crosses between B10.RIII and RIIIS/J mice is the Cia5 on chromosome 3. Early in the congenic mapping process it was clear that the locus was complex, consisting of several subloci with small effects. Therefore, we developed two novel strategies to dissect a QTL: the partial advanced intercross (PAI) strategy, with which we recently found the Cia5 region to consist of three loci, Cia5, Cia21 and Cia22, and now we introduce the QTL-chip strategy, where we have combined congenic mapping with a QTL-restricted expression profiling using a novel microarray design. The expression of QTL genes was compared between parental and congenic mice in lymph node, spleen and paw samples in five biological replicates and in dye-swapped experiments at three time points during the induction phase of arthritis. The QTL chip approach revealed 4 genes located in Cia21, differently expressed in lymph nodes, and 14 genes in Cia22, located within two clusters. One cluster contains six genes, differently expressed in spleen, and the second cluster contains eight genes, differently expressed in paws. We conclude the QTL-chip strategy to be valuable in the selection of candidate genes to be prioritized for further investigation.
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| 9. |
- Johannesson, Martina, et al.
(author)
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Genetics of autoimmune diseases: a multistep process.
- 2006
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In: Current Concepts in Autoimmunity and Chronic Inflammation (Current Topics in Microbiology and Immunology). - : Springer Berlin Heidelberg. - 0070-217X. - 9783540297130 ; :305
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Book chapter (other academic/artistic)abstract
- Abstract in UndeterminedIt has so far been difficult to identify genes behind polygenic autoimmune diseases such as rheumatoid arthritis (RA), multiple sclerosis (MS), and type I diabetes (T1D). With proper animal models, some of the complexity behind these diseases can be reduced. The use of linkage analysis and positional cloning of genes in animal models for RA resulted in the identification of one of the genes regulating severity of arthritis in rats and mice, the Ncf1 gene. The Ncf1 gene encodes for the Ncf1 protein that is involved in production of free oxygen radicals through the NADPH oxidase complex, which opens up a new pathway for therapeutic treatment of inflammatory diseases. in most cases, however, a quantitative trait locus (QTL) is the sum effect of several genes within and outside the QTL, which make positional cloning difficult. Here we will discuss the possibilities and difficulties of gene identification in animal models of autoimmune disorders.
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| 10. |
- Johannesson, Martina
(author)
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Human Embryonic Stem Cells: Directed Differentiation into Posterior Foregut Endoderm and a Functional Assay for Definitive Endoderm
- 2009
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Doctoral thesis (other academic/artistic)abstract
- A decade has passed since the first human embryonic stem cell (hESC) line was established, providing immense hope in curing diseases by cell replacement therapy. For instance, in the pancreas, the insulin-producing β-cells are damaged in diabetes patients, and a successful replacement of those cells could provide a cure. These cells can be replaced by organ donation, but owing to a profound scarcity of organ donors, there is a high demand in alternative sources of β-cells. By recapitulating the in vivo developmental pathway, extensively studied in various animal models, it has been demonstrated that hESC can be directed towards a β-cell fate. A complex network of signaling molecules derived from various tissues play multiple roles in the induction events that take place when directing fully pluripotent cells into their different cell fates in vivo, fates that can be followed by analyzing gene and protein expression profiles. The derivation into functional definitive endoderm (DE), the germ layer giving rise to lungs, intestines, liver and pancreas, is a prerequisite for further development towards a pancreatic fate. This thesis covers directed differentiation of hESC by specific factors as well as individual analysis of these. In the first study, hESC are sequentially differentiated first by Activin A (AA)-induction towards DE, and then by treatment with fibroblast growth factor 4 (FGF4) and retinoic acid (RA) to induce differentiation towards foregut endoderm. Importantly, FGF4 does not act as a posterior morphogen as can be observed in vivo, but rather plays an important role in cell survival. However, in combination with RA, which is known to induce dorsal pancreatic fates in vivo, a high level of PDX1+ posterior foregut endodermal cells, characterized by coexpression with SOX9, HNF6, and FOXA2, is obtained. In another study, we demonstrate a novel role of FGF2 as a broad patterning agent in hESC-differentiation. Low levels promote a hepatic fate, intermediate levels promote a pancreatic fate, whereas high levels promote lung and small intestinal fates. Importantly, with this protocol, a fraction of the PDX1+ cells were also positive for NKX6.1, indicating a putative pancreatic progenitor fate. HESC differentiation is most often followed by analyzing gene and protein expression, whereas functional assays to fully understand the in vivo potential of the derived cell types are rare. Here, we present the establishment of a functional in vivo assay in chick embryos for putative DE. The putative DE is derived by directed differentiation from mouse and human ESCs. Putative mouse and human DE is inserted in early chick embryos and integration into the chick endoderm is assayed 48 h later. Specifically, this thesis summarize the outcome of two important differentiation protocols that have provided knowledge on the individual roles of FGF4, RA and FGF2 in directing hESC towards a pancreatic fate, and provide a basis for further optimization towards obtaining pancreatic progenitors, endocrine progenitors, and ultimately fully functional β-cells for future cell replacement therapy.
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