Publications by authors named "Raphael Reuten"

19 Publications

  • Page 1 of 1

Modeling Metastatic Colonization in a Decellularized Organ Scaffold-Based Perfusion Bioreactor.

Adv Healthc Mater 2022 01 17;11(1):e2100684. Epub 2021 Nov 17.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH), Ole Maaloes Vej 5, Copenhagen, 2200, Denmark.

Metastatic cancer spread is responsible for most cancer-related deaths. To colonize a new organ, invading cells adapt to, and remodel, the local extracellular matrix (ECM), a network of proteins and proteoglycans underpinning all tissues, and a critical regulator of homeostasis and disease. However, there is a major lack in tools to study cancer cell behavior within native 3D ECM. Here, an in-house designed bioreactor, where mouse organ ECM scaffolds are perfused and populated with cells that are challenged to colonize it, is presented. Using a specialized bioreactor chamber, it is possible to monitor cell behavior microscopically (e.g., proliferation, migration) within the organ scaffold. Cancer cells in this system recapitulate cell signaling observed in vivo and remodel complex native ECM. Moreover, the bioreactors are compatible with co-culturing cell types of different genetic origin comprising the normal and tumor microenvironment. This degree of experimental flexibility in an organ-specific and 3D context, opens new possibilities to study cell-cell and cell-ECM interplay and to model diseases in a controllable organ-specific system ex vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/adhm.202100684DOI Listing
January 2022

Organ-Specific, Fibroblast-Derived Matrix as a Tool for Studying Breast Cancer Metastasis.

Cancers (Basel) 2021 Jul 2;13(13). Epub 2021 Jul 2.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH), 2200 Copenhagen, Denmark.

During the metastatic process, breast cancer cells must come into contact with the extra-cellular matrix (ECM) at every step. The ECM provides both structural support and biochemical cues, and cell-ECM interactions can lead to changes in drug response. Here, we used fibroblast-derived ECM (FDM) to perform high throughput drug screening of 4T1 breast cancer cells on metastatic organ ECM (lung), and we see that drug response differs from treatment on plastic. The FDMs that we can produce from different organs are abundant in and contains a complex mixture of ECM proteins. We also show differences in ECM composition between the primary site and secondary organ sites. Furthermore, we show that global kinase signalling of 4T1 cells on the ECM is relatively unchanged between organs, while changes in signalling compared to plastic are significant. Our study highlights the importance of context when testing drug response in vitro, showing that consideration of the ECM is critically important.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/cancers13133331DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8269313PMC
July 2021

Decellularization of the Murine Cardiopulmonary Complex.

J Vis Exp 2021 05 30(171). Epub 2021 May 30.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH);

We present here a decellularization protocol for mouse heart and lungs. It produces structural ECM scaffolds that can be used to analyze ECM topology and composition. It is based on a microsurgical procedure designed to catheterize the trachea and aorta of a euthanized mouse to perfuse the heart and lungs with decellularizing agents. The decellularized cardiopulmonary complex can subsequently be immunostained to reveal the location of structural ECM proteins. The whole procedure can be completed in 4 days. The ECM scaffolds resulting from this protocol are free of dimensional distortions. The absence of cells enables structural examination of ECM structures down to submicron resolution in 3D. This protocol can be applied to healthy and diseased tissue from mice as young as 4-weeks old, including mouse models of fibrosis and cancer, opening the way to determine ECM remodeling associated with cardiopulmonary disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3791/61854DOI Listing
May 2021

Suppression of tumor-associated neutrophils by lorlatinib attenuates pancreatic cancer growth and improves treatment with immune checkpoint blockade.

Nat Commun 2021 06 7;12(1):3414. Epub 2021 Jun 7.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH), Copenhagen, Denmark.

Pancreatic ductal adenocarcinoma (PDAC) patients have a 5-year survival rate of only 8% largely due to late diagnosis and insufficient therapeutic options. Neutrophils are among the most abundant immune cell type within the PDAC tumor microenvironment (TME), and are associated with a poor clinical prognosis. However, despite recent advances in understanding neutrophil biology in cancer, therapies targeting tumor-associated neutrophils are lacking. Here, we demonstrate, using pre-clinical mouse models of PDAC, that lorlatinib attenuates PDAC progression by suppressing neutrophil development and mobilization, and by modulating tumor-promoting neutrophil functions within the TME. When combined, lorlatinib also improves the response to anti-PD-1 blockade resulting in more activated CD8 + T cells in PDAC tumors. In summary, this study identifies an effect of lorlatinib in modulating tumor-associated neutrophils, and demonstrates the potential of lorlatinib to treat PDAC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-23731-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8184753PMC
June 2021

Basement membrane stiffness determines metastases formation.

Nat Mater 2021 06 25;20(6):892-903. Epub 2021 Jan 25.

Center for Applied Tissue Engineering and Regenerative Medicine-CANTER, Munich University of Applied Sciences, Munich, Germany.

The basement membrane (BM) is a special type of extracellular matrix and presents the major barrier cancer cells have to overcome multiple times to form metastases. Here we show that BM stiffness is a major determinant of metastases formation in several tissues and identify netrin-4 (Net4) as a key regulator of BM stiffness. Mechanistically, our biophysical and functional analyses in combination with mathematical simulations show that Net4 softens the mechanical properties of native BMs by opening laminin node complexes, decreasing cancer cell potential to transmigrate this barrier despite creating bigger pores. Our results therefore reveal that BM stiffness is dominant over pore size, and that the mechanical properties of 'normal' BMs determine metastases formation and patient survival independent of cancer-mediated alterations. Thus, identifying individual Net4 protein levels within native BMs in major metastatic organs may have the potential to define patient survival even before tumour formation. The ratio of Net4 to laminin molecules determines BM stiffness, such that the more Net4, the softer the BM, thereby decreasing cancer cell invasion activity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41563-020-00894-0DOI Listing
June 2021

Laminin N-terminus α31 protein distribution in adult human tissues.

PLoS One 2020 2;15(12):e0239889. Epub 2020 Dec 2.

Institute of Life Course and Medical Sciences, University of Liverpool, Liverpool, United Kingdom.

Laminin N-terminus α31 (LaNt α31) is a netrin-like protein derived from alternative splicing of the laminin α3 gene. Although LaNt α31 has been demonstrated to influence corneal and skin epithelial cell function, its expression has not been investigated beyond these tissues. In this study, we used immunohistochemistry to characterise the distribution of this protein in a wide-array of human tissue sections in comparison to laminin α3. The data revealed widespread LaNt α31 expression. In epithelial tissue, LaNt α31 was present in the basal layer of the epidermis, throughout the epithelium of the digestive tract, and in much of the epithelium of the reproductive system. LaNt α31 was also found throughout the vasculature of most tissues, with enrichment in reticular-like fibres in the extracellular matrix surrounding large vessels. A similar matrix pattern was observed around the terminal ducts in the breast and around the alveolar epithelium in the lung, where basement membrane staining was also evident. Specific enrichment of LaNt α31 was identified in sub-populations of cells of the kidney, liver, pancreas, and spleen, with variations in intensity between different cell types in the collecting ducts and glomeruli of the kidney. Intriguingly, LaNt α31 immunoreactivity was also evident in neurons of the central nervous system, in the cerebellum, cerebral cortex, and spinal cord. Together these findings suggest that LaNt α31 may be functionally relevant in a wider range of tissue contexts than previously anticipated, and the data provides a valuable basis for investigation into this interesting protein.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0239889PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7710073PMC
January 2021

Smart Hydrogels for the Augmentation of Bone Regeneration by Endogenous Mesenchymal Progenitor Cell Recruitment.

Adv Sci (Weinh) 2020 Apr 5;7(7):1903395. Epub 2020 Feb 5.

Department of Obstetrics University Hospital Zurich University of Zurich Schmelzbergstr. 12 Zurich 8091 Switzerland.

The treatment of bone defects with recombinant bone morphogenetic protein-2 (BMP-2) requires high doses precluding broad clinical application. Here, a bioengineering approach is presented that strongly improves low-dose BMP-2-based bone regeneration by mobilizing healing-associated mesenchymal progenitor cells (MPCs). Smart synthetic hydrogels are used to trap and study endogenous MPCs trafficking to bone defects. Hydrogel-trapped and prospectively isolated MPCs differentiate into multiple lineages in vitro and form bone in vivo. In vitro screenings reveal that platelet-derived growth factor BB (PDGF-BB) strongly recruits prospective MPCs making it a promising candidate for the engineering of hydrogels that enrich endogenous MPCs in vivo. However, PDGF-BB inhibits BMP-2-mediated osteogenesis both in vitro and in vivo. In contrast, smart two-way dynamic release hydrogels with fast-release of PDGF-BB and sustained delivery of BMP-2 beneficially promote the healing of bone defects. Collectively, it is shown that modulating the dynamics of endogenous progenitor cells in vivo by smart synthetic hydrogels significantly improves bone healing and holds great potential for other advanced applications in regenerative medicine.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/advs.201903395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7141038PMC
April 2020

Author Correction: Decellularization and antibody staining of mouse tissues to map native extracellular matrix structures in 3D.

Nat Protoc 2020 06;15(6):2140

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH), Copenhagen, Denmark.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41596-020-0315-7DOI Listing
June 2020

AMD-Associated HTRA1 Variants Do Not Influence TGF-β Signaling in Microglia.

Adv Exp Med Biol 2019 ;1185:3-7

Laboratory for Experimental Immunology of the Eye, Department of Ophthalmology, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany.

Genetic variants of high-temperature requirement A serine peptidase 1 (HTRA1) and age-related maculopathy susceptibility 2 (ARMS2) are associated with age-related macular degeneration (AMD). One HTRA1 single nucleotide polymorphism (SNP) is situated in the promotor region (rs11200638) resulting in increased expression, while two synonymous SNPs are located in exon 1 (rs1049331:C > T, rs2293870:G > T). HtrA1 is known to inhibit transforming growth factor-β (TGF-β) signaling, a pathway regulating quiescence of microglia, the resident immune cells of the brain and retina. Microglia-mediated immune responses contribute to AMD pathogenesis. It is currently unclear whether AMD-associated HTRA1 variants influence TGF-β signaling and microglia phenotypes. Here, we show that an HtrA1 isoform carrying AMD-associated SNPs in exon 1 exhibits increased proteolytic activity. However, when incubating TGF-β-treated reactive microglia with HtrA1 protein variants, neither the wildtype nor the SNP-associated isoforms changed microglia activation in vitro.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-3-030-27378-1_1DOI Listing
February 2020

Decellularization and antibody staining of mouse tissues to map native extracellular matrix structures in 3D.

Nat Protoc 2019 12 8;14(12):3395-3425. Epub 2019 Nov 8.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen (UCPH), Copenhagen, Denmark.

The extracellular matrix (ECM) is a major regulator of homeostasis and disease, yet the 3D structure of the ECM remains poorly understood because of limitations in ECM visualization. We recently developed an ECM-specialized method termed in situ decellularization of tissues (ISDoT) to isolate native 3D ECM scaffolds from whole organs in which ECM structure and composition are preserved. Here, we present detailed surgical instructions to facilitate decellularization of 33 different mouse tissues and details of validated antibodies that enable the visualization of 35 mouse ECM proteins. Through mapping of these ECM proteins, the structure of the ECM can be determined and tissue structures visualized in detail. In this study, perfusion decellularization is presented for bones, skeletal muscle, tongue, salivary glands, stomach, duodenum, jejunum/ileum, large intestines, mesentery, liver, gallbladder, pancreas, trachea, bronchi, lungs, kidneys, urinary bladder, ovaries, uterine horn, cervix, adrenal gland, heart, arteries, veins, capillaries, lymph nodes, spleen, peripheral nerves, eye, outer ear, mammary glands, skin, and subcutaneous tissue. Decellularization, immunostaining, and imaging take 4-5 d.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41596-019-0225-8DOI Listing
December 2019

Interplay Between LOX Enzymes and Integrins in the Tumor Microenvironment.

Cancers (Basel) 2019 May 26;11(5). Epub 2019 May 26.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, 2200 Copenhagen, Denmark.

Members of the lysyl oxidase (LOX) family are secreted copper-dependent amine oxidases that catalyze the covalent crosslinking of collagens and elastin in the extracellular matrix (ECM), an essential process for the structural integrity of all tissues. LOX enzymes can also remodel the tumor microenvironment and have been implicated in all stages of tumor initiation and progression of many cancer types. Changes in the ECM can influence several cancer cell phenotypes. Integrin adhesion complexes (IACs) physically connect cells with their microenvironment. This review article summarizes the main findings on the role of LOX proteins in modulating the tumor microenvironment, with a particular focus on how ECM changes are integrated by IACs to modulate cells behavior. Finally, we discuss how the development of selective LOX inhibitors may lead to novel and effective therapies in cancer treatment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/cancers11050729DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6562985PMC
May 2019

Solution Structure of C. elegans UNC-6: A Nematode Paralogue of the Axon Guidance Protein Netrin-1.

Biophys J 2019 06 3;116(11):2121-2130. Epub 2019 May 3.

Alberta RNA Research and Training Institute, Department of Chemistry and Biochemistry, University of Lethbridge, Lethbridge, Alberta, Canada; Department of Microbiology, Immunology and Infectious Diseases, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada; DiscoveryLab and Li Ka Shing Institute of Virology, University of Alberta, Edmonton, Alberta, Canada. Electronic address:

UNCoordinated-6 (UNC-6) was the first member of the netrin family to be discovered in Caenorhabditis elegans. With homology to human netrin-1, it is a key signaling molecule involved in directing axon migration in nematodes. Similar to netrin-1, UNC-6 interacts with multiple receptors (UNC-5 and UNC-40, specifically) to guide axon migration in development. As a result of the distinct evolutionary path of UNC-6 compared to vertebrate netrins, we decided to employ an integrated approach to study its solution behavior and compare it to the high-resolution structure we previously published on vertebrate netrins. Dynamic light scattering and analytical ultracentrifugation on UNC-6 (with and without its C-domain) solubilized in a low-ionic strength buffer suggested that UNC-6 forms high-order oligomers. An increase in the buffer ionic strength resulted in a more homogeneous preparation of UNC-6, that was used for subsequent solution x-ray scattering experiments. Our biophysical analysis of UNC-6 ΔC solubilized in a high-ionic strength buffer suggested that it maintains a similar head-to-stalk arrangement as netrins -1 and -4. This phenomenon is thought to play a role in the signaling behavior of UNC-6 and its ability to move throughout the extracellular matrix.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bpj.2019.04.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6554493PMC
June 2019

Local VEGF-A blockade modulates the microenvironment of the corneal graft bed.

Am J Transplant 2019 09 3;19(9):2446-2456. Epub 2019 Apr 3.

Department of Ophthalmology, University Hospital of Cologne, Cologne, Germany.

The microenvironment plays an important role in several immunological processes. Vascular endothelial growth factor-A (VEGF-A) not only regulates angiogenesis, but is known as a modulator of the immune microenvironment. Modulating the site of transplantation might be beneficial for subsequent transplant survival. In this study, we therefore analyzed the effect that a local blockade of VEGF-A in the inflamed cornea as the graft receiving tissue has on the immune system. We used the murine model of suture-induced neovascularization and subsequent high-risk corneal transplantation, which is an optimal model for local drug application. Mice were treated with VEGFR1/R2 trap prior to transplantation. We analyzed corneal gene expression, as well as protein levels in the cornea and serum on the day of transplantation, 2 and 8 weeks later. Local VEGF depletion prior to transplantation increases the expression of pro-inflammatory as well as immune regulatory cytokines only in the corneal microenvironment, but not in the serum. Furthermore, local VEGFR1/R2 trap treatment significantly inhibits the infiltration of CD11c+ dendritic cells into the cornea. Subsequent increased corneal transplantation success was accompanied by a local upregulation of Foxp3 gene expression. This study demonstrates that locally restricted VEGF depletion increases transplantation success by modulating the receiving corneal microenvironment and inducing tolerogenic mechanisms.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/ajt.15331DOI Listing
September 2019

Structural decoding of netrin-4 reveals a regulatory function towards mature basement membranes.

Nat Commun 2016 11 30;7:13515. Epub 2016 Nov 30.

Institute for Dental Research and Oral Musculoskeletal Biology, Medical Faculty, University of Cologne, Joseph-Stelzmann-Strasse 52, Cologne 50931, Germany.

Netrins, a family of laminin-related molecules, have been proposed to act as guidance cues either during nervous system development or the establishment of the vascular system. This was clearly demonstrated for netrin-1 via its interaction with the receptors DCC and UNC5s. However, mainly based on shared homologies with netrin-1, netrin-4 was also proposed to play a role in neuronal outgrowth and developmental/pathological angiogenesis via interactions with netrin-1 receptors. Here, we present the high-resolution structure of netrin-4, which shows unique features in comparison with netrin-1, and show that it does not bind directly to any of the known netrin-1 receptors. We show that netrin-4 disrupts laminin networks and basement membranes (BMs) through high-affinity binding to the laminin γ1 chain. We hypothesize that this laminin-related function is essential for the previously described effects on axon growth promotion and angiogenesis. Our study unveils netrin-4 as a non-enzymatic extracellular matrix protein actively disrupting pre-existing BMs.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/ncomms13515DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5141367PMC
November 2016

Maltose-Binding Protein (MBP), a Secretion-Enhancing Tag for Mammalian Protein Expression Systems.

PLoS One 2016 30;11(3):e0152386. Epub 2016 Mar 30.

Institute for Dental Research and Oral Musculoskeletal Biology, Medical Faculty, University of Cologne, Cologne, Germany.

Recombinant proteins are commonly expressed in eukaryotic expression systems to ensure the formation of disulfide bridges and proper glycosylation. Although many proteins can be expressed easily, some proteins, sub-domains, and mutant protein versions can cause problems. Here, we investigated expression levels of recombinant extracellular, intracellular as well as transmembrane proteins tethered to different polypeptides in mammalian cell lines. Strikingly, fusion of proteins to the prokaryotic maltose-binding protein (MBP) generally enhanced protein production. MBP fusion proteins consistently exhibited the most robust increase in protein production in comparison to commonly used tags, e.g., the Fc, Glutathione S-transferase (GST), SlyD, and serum albumin (ser alb) tag. Moreover, proteins tethered to MBP revealed reduced numbers of dying cells upon transient transfection. In contrast to the Fc tag, MBP is a stable monomer and does not promote protein aggregation. Therefore, the MBP tag does not induce artificial dimerization of tethered proteins and provides a beneficial fusion tag for binding as well as cell adhesion studies. Using MBP we were able to secret a disease causing laminin β2 mutant protein (congenital nephrotic syndrome), which is normally retained in the endoplasmic reticulum. In summary, this study establishes MBP as a versatile expression tag for protein production in eukaryotic expression systems.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0152386PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4814134PMC
August 2016

Structural Decoding of the Netrin-1/UNC5 Interaction and its Therapeutical Implications in Cancers.

Cancer Cell 2016 Feb;29(2):173-85

Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, MB RT3 2N2, Canada. Electronic address:

Netrin-1 has been shown to be up-regulated in a fraction of human cancers as a mechanism to allow these tumors to escape the pro-apoptotic activity of some of its main dependence receptors, the UNC5 homologs (UNC5H). Here we identify the V-2 domain of netrin-1 to be important for its interaction with the Ig1/Ig2 domains of UNC5H2. We generate a humanized anti-netrin-1 antibody that disrupts the interaction between netrin-1 and UNC5H2 and triggers death of netrin-1-expressing tumor cells in vitro. We also present evidence that combining the anti-netrin-1 antibody with epidrugs such as decitabine could be effective in treating tumors showing no or modest netrin-1 expression. These results support that this antibody is a promising drug candidate.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ccell.2016.01.001DOI Listing
February 2016

Biophysical analysis of a lethal laminin alpha-1 mutation reveals altered self-interaction.

Matrix Biol 2016 Jan 26;49:93-105. Epub 2015 Jul 26.

Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba R3T 2N2, Canada. Electronic address:

Laminins are key basement membrane molecules that influence several biological activities and are linked to a number of diseases. They are secreted as heterotrimeric proteins consisting of one α, one β, and one γ chain, followed by their assembly into a polymer-like sheet at the basement membrane. Using sedimentation velocity, dynamic light scattering, and surface plasmon resonance experiments, we studied self-association of three laminin (LM) N-terminal fragments α-1 (hLM α-1N), α-5 (hLM α-5N) and β-3 (hLM β-3N) originating from the short arms of the human laminin αβγ heterotrimer. Corresponding studies of the hLM α-1N C49S mutant, equivalent to the larval lethal C56S mutant in zebrafish, have shown that this mutation causes enhanced self-association behavior, an observation that provides a plausible explanation for the inability of laminin bearing this mutation to fulfill functional roles in vivo, and hence for the deleterious pathological consequences of the mutation on lens function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.matbio.2015.06.005DOI Listing
January 2016

Ultraviolet-radiation-induced inflammation promotes angiotropism and metastasis in melanoma.

Nature 2014 Mar 26;507(7490):109-13. Epub 2014 Feb 26.

Laboratory of Experimental Dermatology, Department of Dermatology and Allergy, University of Bonn, 53115 Bonn, Germany.

Intermittent intense ultraviolet (UV) exposure represents an important aetiological factor in the development of malignant melanoma. The ability of UV radiation to cause tumour-initiating DNA mutations in melanocytes is now firmly established, but how the microenvironmental effects of UV radiation influence melanoma pathogenesis is not fully understood. Here we report that repetitive UV exposure of primary cutaneous melanomas in a genetically engineered mouse model promotes metastatic progression, independent of its tumour-initiating effects. UV irradiation enhanced the expansion of tumour cells along abluminal blood vessel surfaces and increased the number of lung metastases. This effect depended on the recruitment and activation of neutrophils, initiated by the release of high mobility group box 1 (HMGB1) from UV-damaged epidermal keratinocytes and driven by Toll-like receptor 4 (TLR4). The UV-induced neutrophilic inflammatory response stimulated angiogenesis and promoted the ability of melanoma cells to migrate towards endothelial cells and use selective motility cues on their surfaces. Our results not only reveal how UV irradiation of epidermal keratinocytes is sensed by the innate immune system, but also show that the resulting inflammatory response catalyses reciprocal melanoma-endothelial cell interactions leading to perivascular invasion, a phenomenon originally described as angiotropism in human melanomas by histopathologists. Angiotropism represents a hitherto underappreciated mechanism of metastasis that also increases the likelihood of intravasation and haematogenous dissemination. Consistent with our findings, ulcerated primary human melanomas with abundant neutrophils and reactive angiogenesis frequently show angiotropism and a high risk for metastases. Our work indicates that targeting the inflammation-induced phenotypic plasticity of melanoma cells and their association with endothelial cells represent rational strategies to specifically interfere with metastatic progression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nature13111DOI Listing
March 2014

Determination of a molecular shape for netrin-4 from hydrodynamic and small angle X-ray scattering measurements.

Matrix Biol 2012 Mar 24;31(2):135-40. Epub 2011 Dec 24.

Department of Chemistry, University of Manitoba, 144 Dysart Road, Winnipeg, Manitoba, Canada R3T 2N2.

As part of a continuing investigation of netrins, an emerging class of extracellular matrix proteins that are involved in axon guidance activity, we have used dynamic light scattering (DLS) and small angle X-ray scattering to investigate the solution conformation of a truncated version of netrin-4 (Δnetrin-4) that lacks the C-terminal portion. The protein is characterized by a hydrodynamic (Stokes) radius (r(H)) of 4.60 (±0.20) nm, a radius of gyration (r(G)) of 4.42 (±0.20) nm and a maximum particle dimension (D(max)) of 16nm. More detailed ab initio modeling of the SAXS data indicates an extended rod like conformation for Δnetrin-4 in solution-a concept supported by the excellent agreement observed between experimental parameter estimates and those calculated for the ab initio models for Δnetrin-4 by the HYDROPRO program.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.matbio.2011.11.004DOI Listing
March 2012
-->