| 1. |
- Håkansson, Joakim, 1975, et al.
(författare)
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N-CAM exhibits a regulatory function in pathological angiogenesis in oxygen induced retinopathy.
- 2011
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Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 6:10
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Tidskriftsartikel (refereegranskat)abstract
- Diabetic retinopathy and retinopathy of prematurity are diseases caused by pathological angiogenesis in the retina as a consequence of local hypoxia. The underlying mechanism for epiretinal neovascularization (tuft formation), which contributes to blindness, has yet to be identified. Neural cell adhesion molecule (N-CAM) is expressed by Müller cells and astrocytes, which are in close contact with the retinal vasculature, during normal developmental angiogenesis.
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| 2. |
- Johannesson, Martina, et al.
(författare)
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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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Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 4:3
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Tidskriftsartikel (refereegranskat)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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| 3. |
- Kesavan, Gokul, et al.
(författare)
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Cdc42-mediated tubulogenesis controls cell specification.
- 2009
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Ingår i: Cell. - : Elsevier BV. - 1097-4172 .- 0092-8674. ; 139:4, s. 791-801
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Tidskriftsartikel (refereegranskat)abstract
- Understanding how cells polarize and coordinate tubulogenesis during organ formation is a central question in biology. Tubulogenesis often coincides with cell-lineage specification during organ development. Hence, an elementary question is whether these two processes are independently controlled, or whether proper cell specification depends on formation of tubes. To address these fundamental questions, we have studied the functional role of Cdc42 in pancreatic tubulogenesis. We present evidence that Cdc42 is essential for tube formation, specifically for initiating microlumen formation and later for maintaining apical cell polarity. Finally, we show that Cdc42 controls cell specification non-cell-autonomously by providing the correct microenvironment for proper control of cell-fate choices of multipotent progenitors. For a video summary of this article, see the PaperFlick file with the Supplemental Data available online.
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| 4. |
- Norrman, Karin, et al.
(författare)
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Quantitative comparison of constitutive promoters in human ES cells.
- 2010
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 5:8
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Constitutive promoters that ensure sustained and high level gene expression are basic research tools that have a wide range of applications, including studies of human embryology and drug discovery in human embryonic stem cells (hESCs). Numerous cellular/viral promoters that ensure sustained gene expression in various cell types have been identified but systematic comparison of their activities in hESCs is still lacking. METHODOLOGY/PRINCIPAL FINDINGS: We have quantitatively compared promoter activities of five commonly used constitutive promoters, including the human β-actin promoter (ACTB), cytomegalovirus (CMV), elongation factor-1α, (EF1α), phosphoglycerate kinase (PGK) and ubiquitinC (UbC) in hESCs. Lentiviral gene transfer was used to ensure stable integration of promoter-eGFP constructs into the hESCs genome. Promoter activities were quantitatively compared in long term culture of undifferentiated hESCs and in their differentiated progenies. CONCLUSION/SIGNIFICANCE: The ACTB, EF1α and PGK promoters showed stable activities during long term culture of undifferentiated hESCs. The ACTB promoter was superior by maintaining expression in 75-80% of the cells after 50 days in culture. During embryoid body (EB) differentiation, promoter activities of all five promoters decreased. Although the EF1α promoter was downregulated in approximately 50% of the cells, it was the most stable promoter during differentiation. Gene expression analysis of differentiated eGFP+ and eGFP- cells indicate that promoter activities might be restricted to specific cell lineages, suggesting the need to carefully select optimal promoters for constitutive gene expression in differentiated hESCs.
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| 5. |
- Rymo, Simin, 1959, et al.
(författare)
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A Two-Way Communication between Microglial Cells and Angiogenic Sprouts Regulates Angiogenesis in Aortic Ring Cultures
- 2011
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: Myeloid cells have been associated with physiological and pathological angiogenesis, but their exact functions in these processes remain poorly defined. Monocyte-derived tissue macrophages of the CNS, or microglial cells, invade the mammalian retina before it becomes vascularized. Recent studies correlate the presence of microglia in the developing CNS with vascular network formation, but it is not clear whether the effect is directly caused by microglia and their contact with the endothelium. Methodology/Principal Findings: We combined in vivo studies of the developing mouse retina with in vitro studies using the aortic ring model to address the role of microglia in developmental angiogenesis. Our in vivo analyses are consistent with previous findings that microglia are present at sites of endothelial tip-cell anastomosis, and genetic ablation of microglia caused a sparser vascular network associated with reduced number of filopodia-bearing sprouts. Addition of microglia in the aortic ring model was sufficient to stimulate vessel sprouting. The effect was independent of physical contact between microglia and endothelial cells, and could be partly mimicked using microglial cell-conditioned medium. Addition of VEGF-A promoted angiogenic sprouts of different morphology in comparison with the microglial cells, and inhibition of VEGF-A did not affect the microglia-induced angiogenic response, arguing that the proangiogenic factor(s) released by microglia is distinct from VEGF-A. Finally, microglia exhibited oriented migration towards the vessels in the aortic ring cultures. Conclusions/Significance: Microglia stimulate vessel sprouting in the aortic ring cultures via a soluble microglial-derived product(s), rather than direct contact with endothelial cells. The observed migration of microglia towards the growing sprouts suggests that their position near endothelial tip-cells could result from attractive cues secreted by the vessels. Our data reveals a two-way communication between microglia and vessels that depends on soluble factors and should extend the understanding of how microglia promote vascular network formation.
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| 6. |
- Wolfhagen Sand, Fredrik
(författare)
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ENDOTHELIAL- EPITHELIAL INTERACTIONS Blood vessels friends or foes?
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Diabetes affects enormous amounts of patients. In type 1 diabetes the insulin producing β cells in the islet of Langerhans are destroyed by the immune system. The β cells, which are located in the pancreas, regulate blood glucose levels. The lack of insulin production requires administration of insulin. This is accomplished by insulin injections. Even though the treatment methods have been improved, we are still not able to artificially regulate the blood glucose levels as evenly as the β cells. The fluctuations of the blood glucose levels results in the long term in a number of complication, among the diabetic retinopathy. Diabetic retinopathy is the leading cause of blindness among the younger population. Both to reduce the complications of diabetes and to find cures of the disease is important. By using a model for diabetic retinopathy in mice we have been able to show that lack of the molecule N-CAM is protecting against the pathological neovascularization affecting diabetic patients. The last 10 years have proven it possible to cure diabetes through transplantations of islets of Langerhans from cadaveric donors. The transplantations requires suppression of the immune response, this could lead to other complications. The amount of diabetic patients in need of transplantations far exceeds the availability of donors. The use of stem cells to produce insulin producing β cells might be a possible way. To be able to direct the stem cells into β cells it is important to understand how these cells form during embryo development. By the use of mice we have been able to study how blood vessels regulate the growth of the precursors of β cells. We have also been able to show that the formation of a tubular system in the pancreas is important for the regulation of which cell types that are to be formed. Our results can be used to develop better methods for the production of β cells from stem cells.
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| 7. |
- Wolfhagen Sand, Fredrik, et al.
(författare)
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Growth-limiting role of endothelial cells in endoderm development
- 2011
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Ingår i: Developmental Biology. - : Academic Press. - 0012-1606 .- 1095-564X. ; 352:2, s. 267-277
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Tidskriftsartikel (refereegranskat)abstract
- Endoderm development is dependent on inductive signals from different structures in close vicinity, including the notochord, lateral plate mesoderm and endothelial cells. Recently, we demonstrated that a functional vascular system is necessary for proper pancreas development, and that sphingosine-1-phosphate (S1P) exhibits the traits of a blood vessel-derived molecule involved in early pancreas morphogenesis. To examine whether S1P(1)-signaling plays a more general role in endoderm development, S1P(1)-deficient mice were analyzed. S1P(1) ablation results in compromised growth of several foregut-derived organs, including the stomach, dorsal and ventral pancreas and liver. Within the developing pancreas the reduction in organ size was due to deficient proliferation of Pdx1(+) pancreatic progenitors, whereas endocrine cell differentiation was unaffected. Ablation of endothelial cells in vitro did not mimic the S1P(1) phenotype, instead, increased organ size and hyperbranching were observed. Consistent with a negative role for endothelial cells in endoderm organ expansion, excessive vasculature was discovered in S1P(1)-deficient embryos. Altogether, our results show that endothelial cell hyperplasia negatively influences organ development in several foregut-derived organs.
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