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Elisabet Wallgard,
Anja Nitzsche,
Jimmy Larsson,
Xiaoyuan Guo,
Lothar C Dieterich,
Anna Dimberg,
Tommie Olofsson,
Fredrik C Pontén,
Taija Mäkinen, Mattias Kalén,
Mats Hellström
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ABSTRACT: Angiogenesis is implicated in many pathological conditions. The role of the proteins involved remains largely unknown, and few vascular-specific drug targets have been discovered. Previously, in a screen for angiogenesis regulators, we identified Paladin (mouse: X99384, human: KIAA1274), a protein containing predicted S/T/Y phosphatase domains.
We present a mouse knockout allele for Paladin with a β-galactosidase reporter, which in combination with Paladin antibodies demonstrate that Paladin is expressed in the vasculature. During mouse embryogenesis, Paladin is primarily expressed in capillary and venous endothelial cells. In adult mice Paladin is predominantly expressed in arterial pericytes and vascular smooth muscle cells. Paladin also displays vascular-restricted expression in human brain, astrocytomas, and glioblastomas.
Paladin, a novel putative phosphatase, displays a dynamic expression pattern in the vasculature. During embryonic stages it is broadly expressed in endothelial cells, while in the adult it is selectively expressed in arterial smooth muscle cells.
Developmental Dynamics 02/2012; 241(4):770-86. · 2.54 Impact Factor
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ABSTRACT: The Notch signaling pathway is essential for normal development due to its role in control of cell differentiation, proliferation and survival. It is also critically involved in tumorigenesis and cancer progression. A key enzyme in the activation of Notch signaling is the gamma-secretase protein complex and therefore, gamma-secretase inhibitors (GSIs)--originally developed for Alzheimer's disease--are now being evaluated in clinical trials for human malignancies. It is also clear that Notch plays an important role in angiogenesis driven by Vascular Endothelial Growth Factor A (VEGF-A)--a process instrumental for tumor growth and metastasis. The effect of GSIs on tumor vasculature has not been conclusively determined. Here we report that Compound X (CX), a GSI previously reported to potently inhibit Notch signaling in vitro and in vivo, promotes angiogenic sprouting in vitro and during developmental angiogenesis in mice. Furthermore, CX treatment suppresses tumor growth in a mouse model of renal carcinoma, leads to the formation of abnormal vessels and an increased tumor vascular density. Using a rabbit model of VEGF-A-driven angiogenesis in skeletal muscle, we demonstrate that CX treatment promotes abnormal blood vessel growth characterized by vessel occlusion, disrupted blood flow, and increased vascular leakage. Based on these findings, we propose a model for how GSIs and other Notch inhibitors disrupt tumor blood vessel perfusion, which might be useful for understanding this new class of anti-cancer agents.
PLoS ONE 01/2011; 6(4):e18709. · 4.09 Impact Factor
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Mattias Kalén,
Elisabet Wallgard,
Noomi Asker,
Aidas Nasevicius,
Elisabet Athley,
Erik Billgren,
Jon D Larson,
Shannon A Wadman,
Elizabeth Norseng,
Karl J Clark, [......],
Linda Karlsson-Lindahl,
Ann-Katrin Häger,
Holger Weber,
Hellmut Augustin,
Tore Samuelsson,
Chelsy K Kemmet,
Carly M Utesch,
Jeffrey J Essner,
Perry B Hackett,
Mats Hellström
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ABSTRACT: We combined reverse and chemical genetics to identify targets and compounds modulating blood vessel development. Through transcript profiling in mice, we identified 150 potentially druggable microvessel-enriched gene products. Orthologs of 50 of these were knocked down in a reverse genetic screen in zebrafish, demonstrating that 16 were necessary for developmental angiogenesis. In parallel, 1280 pharmacologically active compounds were screened in a human cell-based assay, identifying 28 compounds selectively inhibiting endothelial sprouting. Several links were revealed between the results of the reverse and chemical genetic screens, including the serine/threonine (S/T) phosphatases ppp1ca, ppp1cc, and ppp4c and an inhibitor of this gene family; Endothall. Our results suggest that the combination of reverse and chemical genetic screens, in vertebrates, is an efficient strategy for the identification of drug targets and compounds that modulate complex biological systems, such as angiogenesis.
Chemistry & biology 05/2009; 16(4):432-41. · 6.52 Impact Factor
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Mats Hellström,
Li-Kun Phng,
Jennifer J Hofmann,
Elisabet Wallgard,
Leigh Coultas,
Per Lindblom,
Jackelyn Alva,
Ann-Katrin Nilsson,
Linda Karlsson,
Nicholas Gaiano,
Keejung Yoon,
Janet Rossant,
M Luisa Iruela-Arispe, Mattias Kalén,
Holger Gerhardt,
Christer Betsholtz
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ABSTRACT: In sprouting angiogenesis, specialized endothelial tip cells lead the outgrowth of blood-vessel sprouts towards gradients of vascular endothelial growth factor (VEGF)-A. VEGF-A is also essential for the induction of endothelial tip cells, but it is not known how single tip cells are selected to lead each vessel sprout, and how tip-cell numbers are determined. Here we present evidence that delta-like 4 (Dll4)-Notch1 signalling regulates the formation of appropriate numbers of tip cells to control vessel sprouting and branching in the mouse retina. We show that inhibition of Notch signalling using gamma-secretase inhibitors, genetic inactivation of one allele of the endothelial Notch ligand Dll4, or endothelial-specific genetic deletion of Notch1, all promote increased numbers of tip cells. Conversely, activation of Notch by a soluble jagged1 peptide leads to fewer tip cells and vessel branches. Dll4 and reporters of Notch signalling are distributed in a mosaic pattern among endothelial cells of actively sprouting retinal vessels. At this location, Notch1-deleted endothelial cells preferentially assume tip-cell characteristics. Together, our results suggest that Dll4-Notch1 signalling between the endothelial cells within the angiogenic sprout serves to restrict tip-cell formation in response to VEGF, thereby establishing the adequate ratio between tip and stalk cells required for correct sprouting and branching patterns. This model offers an explanation for the dose-dependency and haploinsufficiency of the Dll4 gene, and indicates that modulators of Dll4 or Notch signalling, such as gamma-secretase inhibitors developed for Alzheimer's disease, might find usage as pharmacological regulators of angiogenesis.
Nature 03/2007; 445(7129):776-80. · 36.28 Impact Factor
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ABSTRACT: All blood capillaries consist of endothelial tubes surrounded by mural cells referred to as pericytes. The origin, recruitment, and function of the pericytes is poorly understood, but the importance of these cells is underscored by the severe cardiovascular defects in mice genetically devoid of factors regulating pericyte recruitment to embryonic vessels, and by the association between pericyte loss and microangiopathy in diabetes mellitus. A general problem in the study of pericytes is the shortage of markers for these cells. To identify new markers for pericytes, we have taken advantage of the platelet-derived growth factor (PDGF)-B knockout mouse model, in which developing blood vessels in the central nervous system are almost completely devoid of pericytes. Using cDNA microarrays, we analyzed the gene expression in PDGF-B null embryos in comparison with corresponding wild-type embryos and searched for down-regulated genes. The most down-regulated gene present on our microarray was RGS5, a member of the RGS family of GTPase-activating proteins for G proteins. In situ hybridization identified RGS5 expression in brain pericytes, and in pericytes and vascular smooth muscle cells in certain other, but not all, locations. Absence of RGS5 expression in PDGF-B and PDGFR beta-null embryos correlated with pericyte loss in these mice. Residual RGS5 expression in rare pericytes suggested that RGS5 is a pericyte marker expressed independently of PDGF-B/R beta signaling. With RGS5 as a proof-of-principle, our data demonstrate the usefulness of microarray analysis of mouse models for abnormal pericyte development in the identification of new pericyte-specific markers.
American Journal Of Pathology 04/2003; 162(3):721-9. · 4.89 Impact Factor
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ABSTRACT: Loss of pericytes from the capillary wall is a hallmark of diabetic retinopathy, however, the pathogenic significance of this phenomenon is unclear. In previous mouse gene knockout models leading to pericyte deficiency, prenatal lethality has so far precluded analysis of postnatal consequences in the retina. We now report that endothelium-restricted ablation of platelet-derived growth factor-B generates viable mice with extensive inter- and intra-individual variation in the density of pericytes throughout the CNS. We found a strong inverse correlation between pericyte density and the formation of a range of retinal microvascular abnormalities strongly reminiscent of those seen in diabetic humans. Proliferative retinopathy invariably developed when pericyte density was <50% of normal. Our data suggest that a reduction of the pericyte density is sufficient to cause retinopathy in mice, implying that pericyte loss may also be a causal pathogenic event in human diabetic retinopathy.
The EMBO Journal 08/2002; 21(16):4307-16. · 9.20 Impact Factor
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ABSTRACT: Microarray hybridization has rapidly evolved as an important tool for genomic studies and studies of gene regulation at the transcriptome level. Expression profiles from homogenous samples such as yeast and mammalian cell cultures are currently extending our understanding of biology, whereas analyses of multicellular organisms are more difficult because of tissue complexity. The combination of laser microdissection, RNA amplification, and microarray hybridization has the potential to provide expression profiles from selected populations of cells in vivo. In this article, we present and evaluate an experimental procedure for global gene expression analysis of slender embryonic structures using laser microbeam microdissection and laser pressure catapulting. As a proof of principle, expression profiles from 1000 cells in the mouse embryonic (E9.5) dorsal aorta were generated and compared with profiles for captured mesenchymal cells located one cell diameter further away from the aortic lumen. A number of genes were overexpressed in the aorta, including 11 previously known markers for blood vessels. Among the blood vessel markers were endoglin, tie-2, PDGFB, and integrin-beta1, that are important regulators of blood vessel formation. This demonstrates that microarray analysis of laser microbeam micro-dissected cells is sufficiently sensitive for identifying genes with regulative functions.
American Journal Of Pathology 04/2002; 160(3):801-13. · 4.89 Impact Factor
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ABSTRACT: The association of pericytes (PCs) to newly formed blood vessels has been suggested to regulate endothelial cell (EC) proliferation,
survival, migration, differentiation, and vascular branching. Here, we addressed these issues using PDGF-B– and PDGF receptor-β
(PDGFR-β)–deficient mice as in vivo models of brain angiogenesis in the absence of PCs. Quantitative morphological analysis
showed that these mutants have normal microvessel density, length, and number of branch points. However, absence of PCs correlates
with endothelial hyperplasia, increased capillary diameter, abnormal EC shape and ultrastructure, changed cellular distribution
of certain junctional proteins, and morphological signs of increased transendothelial permeability. Brain endothelial hyperplasia
was observed already at embryonic day (E) 11.5 and persisted throughout development. From E 13.5, vascular endothelial growth
factor-A (VEGF-A) and other genes responsive to metabolic stress became upregulated, suggesting that the abnormal microvessel
architecture has systemic metabolic consequences. VEGF-A upregulation correlated temporally with the occurrence of vascular
abnormalities in the placenta and dilation of the heart. Thus, although PC deficiency appears to have direct effects on EC
number before E 13.5, the subsequent increased VEGF-A levels may further abrogate microvessel architecture, promote vascular
permeability, and contribute to formation of the edematous phenotype observed in late gestation PDGF-B and PDGFR-β knock out
embryos.
The Journal of Cell Biology 04/2001; 153(3):543-554. · 10.26 Impact Factor
