Publications by authors named "Christina Eich"

18 Publications

  • Page 1 of 1

PLGA-Nanoparticles for Intracellular Delivery of the CRISPR-Complex to Elevate Fetal Globin Expression in Erythroid Cells.

Biomaterials 2021 Jan 7;268:120580. Epub 2020 Dec 7.

Translational Nanobiomaterials and Imaging, Department of Radiology, Leiden University Medical Center, the Netherlands. Electronic address:

Ex vivo gene editing of CD34 hematopoietic stem and progenitor cells (HSPCs) offers great opportunities to develop new treatments for a number of malignant and non-malignant diseases. Efficient gene-editing in HSPCs has been achieved using electroporation and/or viral transduction to deliver the CRISPR-complex, but cellular toxicity is a drawback of currently used methods. Nanoparticle (NP)-based gene-editing strategies can further enhance the gene-editing potential of HSPCs and provide a delivery system for in vivo application. Here, we developed CRISPR/Cas9-PLGA-NPs efficiently encapsulating Cas9 protein, single gRNA and a fluorescent probe. The initial 'burst' of Cas9 and gRNA release was followed by a sustained release pattern. CRISPR/Cas9-PLGA-NPs were taken up and processed by human HSPCs, without inducing cellular cytotoxicity. Upon escape from the lysosomal compartment, CRISPR/Cas9-PLGA-NPs-mediated gene editing of the γ-globin gene locus resulted in elevated expression of fetal hemoglobin (HbF) in primary erythroid cells. The development of CRISPR/Cas9-PLGA-NPs provides an attractive tool for the delivery of the CRISPR components to target HSPCs, and could provide the basis for in vivo treatment of hemoglobinopathies and other genetic diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.biomaterials.2020.120580DOI Listing
January 2021

Recent Advances in Rare-Earth-Doped Nanoparticles for NIR-II Imaging and Cancer Theranostics.

Front Chem 2020 17;8:496. Epub 2020 Jun 17.

Translational Nanobiomaterials and Imaging Group, Department of Radiology, Leiden University Medical Center, Leiden, Netherlands.

Fluorescence imaging in the second near infrared window (NIR-II, 1,000-1,700 nm) has been widely used in cancer diagnosis and treatment due to its high spatial resolution and deep tissue penetration depths. In this work, recent advances in rare-earth-doped nanoparticles (RENPs)-a novel kind of NIR-II nanoprobes-are presented. The main focus of this study is on the modification of RENPs and their applications in NIR-II and imaging and cancer theranostics. Finally, the perspectives and challenges of NIR-II RENPs are discussed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fchem.2020.00496DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7325968PMC
June 2020

Correction to: Ceramide and palmitic acid inhibit macrophage-mediated epithelial-mesenchymal transition in colorectal cancer.

Mol Cell Biochem 2020 Jun;469(1-2):179-180

Translational Nanobiomaterials and Imaging, Department of Radiology, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.

The third and fifth author's affiliation was published incorrectly in the original article. Also, the Figure 5 and the Acknowledgement section was published incorrectly. The corrected affiliation, Figure 5 and the Acknowledgement section are provided in this correction.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s11010-020-03735-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7645493PMC
June 2020

Ceramide and palmitic acid inhibit macrophage-mediated epithelial-mesenchymal transition in colorectal cancer.

Mol Cell Biochem 2020 May 28;468(1-2):153-168. Epub 2020 Mar 28.

Translational Nanobiomaterials and Imaging, Department of Radiology, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.

Accumulating evidence indicates that ceramide (Cer) and palmitic acid (PA) possess the ability to modulate switching of macrophage phenotypes and possess anti-tumorigenic effects; however, the underlying molecular mechanisms are largely unknown. The aim of the present study was to investigate whether Cer and PA could induce switching of macrophage polarization from the tumorigenic M2- towards the pro-inflammatory M1-phenotype, and whether this consequently altered the potential of colorectal cancer cells to undergo epithelial-mesenchymal transition (EMT), a hallmark of tumor progression. Our study showed that Cer- and PA-treated macrophages increased expression of the macrophage 1 (M1)-marker CD68 and secretion of IL-12 and attenuated expression of the macrophage 2 (M2)-marker CD163 and IL-10 secretion. Moreover, Cer and PA abolished M2 macrophage-induced EMT and migration of colorectal cancer cells. At the molecular level, this coincided with inhibition of SNAI1 and vimentin expression and upregulation of E-cadherin. Furthermore, Cer and PA attenuated expression levels of IL-10 in colorectal cancer cells co-cultured with M2 macrophages and downregulated STAT3 and NF-κB expression. For the first time, our findings suggest the presence of an IL-10-STAT3-NF-κB signaling axis in colorectal cancer cells co-cultured with M2 macrophages, mimicking the tumor microenvironment. Importantly, PA and Cer were powerful inhibitors of this signaling axis and, consequently, EMT of colorectal cancer cells. These results contribute to our understanding of the immunological mechanisms that underlie the anti-tumorigenic effects of lipids for future combination with drugs in the therapy of colorectal carcinoma.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s11010-020-03719-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145792PMC
May 2020

Cutting Edge: IL-6-Driven Immune Dysregulation Is Strictly Dependent on IL-6R α-Chain Expression.

J Immunol 2020 02 10;204(4):747-751. Epub 2020 Jan 10.

Institute for Molecular Medicine, University Medical Center of the Johannes Gutenberg-University Mainz, 55131 Mainz, Germany.

IL-6 binds to the IL-6R α-chain (IL-6Rα) and signals via the signal transducer gp130. Recently, IL-6 was found to also bind to the cell surface glycoprotein CD5, which would then engage gp130 in the absence of IL-6Rα. However, the biological relevance of this alternative pathway is under debate. In this study, we developed a mouse model, in which murine IL-6 is overexpressed in a CD11c-Cre-dependent manner. Transgenic mice developed a lethal immune dysregulation syndrome with increased numbers of Ly-6G neutrophils and Ly-6C monocytes/macrophages. IL-6 overexpression promoted activation of CD4 T cells while suppressing CD5 B-1a cell development. However, additional ablation of IL-6Rα protected IL-6-overexpressing mice from IL-6-triggered inflammation and fully phenocopied IL-6Rα-deficient mice without IL-6 overexpression. Mechanistically, IL-6Rα deficiency completely prevented downstream activation of STAT3 in response to IL-6. Altogether, our data clarify that IL-6Rα is the only biologically relevant receptor for IL-6 in mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.1900876DOI Listing
February 2020

Design, construction, and biological testing of an implantable porous trilayer scaffold for repairing osteoarthritic cartilage.

J Tissue Eng Regen Med 2020 02 26;14(2):355-368. Epub 2019 Dec 26.

Translational Nanobiomaterials and Imaging, Department of Radiology, Leiden University Medical Centre, Leiden, The Netherlands.

Various tissue engineering systems for cartilage repair have been designed and tested over the past two decades, leading to the development of many promising cartilage grafts. However, no one has yet succeeded in devising an optimal system to restore damaged articular cartilage. Here, the design, assembly, and biological testing of a porous, chitosan/collagen-based scaffold as an implant to repair damaged articular cartilage is reported. Its gradient composition and trilayer structure mimic variations in natural cartilage tissue. One of its layers includes hydroxyapatite, a bioactive component that facilitates the integration of growing tissue on local bone in the target area after scaffold implantation. The scaffold was evaluated for surface morphology; rheological performance (storage, loss, complex, and time-relaxation moduli at 1 kHz); physiological stability; in vitro activity and cytotoxicity (on a human chondrocyte C28 cell line); and in vivo performance (tissue growth and biodegradability), in a murine model of osteoarthritis. The scaffold was shown to be mechanically resistant and noncytotoxic, favored tissue growth in vivo, and remained stable for 35 days postimplantation in mice. These encouraging results highlight the potential of this porous chitosan/collagen scaffold for clinical applications in cartilage tissue engineering.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/term.3001DOI Listing
February 2020

In vivo single cell analysis reveals Gata2 dynamics in cells transitioning to hematopoietic fate.

J Exp Med 2018 01 7;215(1):233-248. Epub 2017 Dec 7.

Department of Cell Biology, Erasmus Stem Cell Institute, Erasmus Medical Center, Rotterdam, Netherlands

Cell fate is established through coordinated gene expression programs in individual cells. Regulatory networks that include the Gata2 transcription factor play central roles in hematopoietic fate establishment. Although Gata2 is essential to the embryonic development and function of hematopoietic stem cells that form the adult hierarchy, little is known about the in vivo expression dynamics of Gata2 in single cells. Here, we examine Gata2 expression in single aortic cells as they establish hematopoietic fate in mouse embryos. Time-lapse imaging reveals rapid pulsatile level changes in Gata2 reporter expression in cells undergoing endothelial-to-hematopoietic transition. Moreover, Gata2 reporter pulsatile expression is dramatically altered in aortic cells, which undergo fewer transitions and are reduced in hematopoietic potential. Our novel finding of dynamic pulsatile expression of suggests a highly unstable genetic state in single cells concomitant with their transition to hematopoietic fate. This reinforces the notion that threshold levels of Gata2 influence fate establishment and has implications for transcription factor-related hematologic dysfunctions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1084/jem.20170807DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748852PMC
January 2018

Effect of TLR ligands co-encapsulated with multiepitopic antigen in nanoliposomes targeted to human DCs via Fc receptor for cancer vaccines.

Immunobiology 2017 11 10;222(11):989-997. Epub 2017 Jun 10.

Department of Biochemistry and Molecular Biology, University of Barcelona, Diagonal 643, 08028 Barcelona, Spain. Electronic address:

Nanoliposomes (NLs) hold promise as new highly specific nanomedicine for anti-tumor vaccines, since they could be targeted to specific receptors on dendritic cell (DC) to induce maturation and activation and increase the anti-tumor immune response. Here we studied a NLs formulation targeted or not to FcR (the receptor for the IgG Fc fragment) for the treatment of androgen-responsive prostate cancer. Luteinizing-hormone-releasing hormone (LHRH) peptide (B- and T-cell epitopes), in tandem with a tetanus toxoid T-helper epitope (830-844 region) and several TLR (Toll-Like Receptor) ligands as adjuvants were co-encapsulated. Specific uptake in vitro of LHRH-TT liposomes targeted to the FcRs of human DCs was enhanced. DC maturation/activation, cytokine production and lymphocyte activation were consistently higher in targeted than non-targeted liposomes. Similar increase was observed as more adjuvants were administrated. Targeting to specific receptor and co-encapsulation of several TLR adjuvants are essential factors for the immune response in peptide based liposome vaccine.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.imbio.2017.06.002DOI Listing
November 2017

Controlled release of antigen and Toll-like receptor ligands from PLGA nanoparticles enhances immunogenicity.

Nanomedicine (Lond) 2017 Mar 9;12(5):491-510. Epub 2017 Feb 9.

Department of Tumor Immunology, Radboud Insititute for Molecular Life Sciences, Radboud University Medical Center, Postbox 9101, 6500 HB Nijmegen, The Netherlands.

Aim: Dendritic cells rapidly capture nanoparticles and induce a potent cellular immune response. It is yet unknown whether the immunological response induced by slow release of encapsulated versus soluble antigen and adjuvant is superior.

Materials & Methods: The kinetics of poly(lactic-co-glycolic acid) PLGA nanoparticles antigen release was studied by the DQ-bovine serum albumin (BSA) self-quenching antigen model. The immunological response induced was evaluated by means of dendritic cell activation/maturation markers, cytokine production and their ability to drive antigen-specific T-cell proliferation.

Results & Conclusion: PLGA-encapsulated antigen and adjuvant showed an enhanced T-cell response when compared with soluble vaccine components by increasing antigenicity and adjuvanticity. Although the kinetic profile followed the same pattern, encapsulation increased strength and duration of the response.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.2217/nnm-2016-0295DOI Listing
March 2017

Proteome Based Construction of the Lymphocyte Function-Associated Antigen 1 (LFA-1) Interactome in Human Dendritic Cells.

PLoS One 2016 18;11(2):e0149637. Epub 2016 Feb 18.

Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Centre, Nijmegen, The Netherlands.

The β2-integrin lymphocyte function-associated antigen 1 (LFA-1) plays an important role in the migration, adhesion and intercellular communication of dendritic cells (DCs). During the differentiation of human DCs from monocyte precursors, LFA-1 ligand binding capacity is completely lost, even though its expression levels were remained constant. Yet LFA-1-mediated adhesive capacity on DCs can be regained by exposing DCs to the chemokine CCL21, suggesting a high degree of regulation of LFA-1 activity during the course of DC differentiation. The molecular mechanisms underlying this regulation of LFA-1 function in DCs, however, remain elusive. To get more insight we attempted to identify specific LFA-1 binding partners that may play a role in regulating LFA-1 activity in DCs. We used highly sensitive label free quantitative mass-spectrometry to identify proteins co-immunoprecipitated (co-IP) with LFA-1 from ex vivo generated DCs. Among the potential binding partners we identified not only established components of integrin signalling pathways and cytoskeletal proteins, but also several novel LFA-1 binding partners including CD13, galectin-3, thrombospondin-1 and CD44. Further comparison to the LFA-1 interaction partners in monocytes indicated that DC differentiation was accompanied by an overall increase in LFA-1 associated proteins, in particular cytoskeletal, signalling and plasma membrane (PM) proteins. The here presented LFA-1 interactome composed of 78 proteins thus represents a valuable resource of potential regulators of LFA-1 function during the DC lifecycle.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0149637PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4758637PMC
July 2016

Changes in membrane sphingolipid composition modulate dynamics and adhesion of integrin nanoclusters.

Sci Rep 2016 Feb 12;6:20693. Epub 2016 Feb 12.

Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Postbox 9101, 6500 HB Nijmegen, The Netherlands.

Sphingolipids are essential constituents of the plasma membrane (PM) and play an important role in signal transduction by modulating clustering and dynamics of membrane receptors. Changes in lipid composition are therefore likely to influence receptor organisation and function, but how this precisely occurs is difficult to address given the intricacy of the PM lipid-network. Here, we combined biochemical assays and single molecule dynamic approaches to demonstrate that the local lipid environment regulates adhesion of integrin receptors by impacting on their lateral mobility. Induction of sphingomyelinase (SMase) activity reduced sphingomyelin (SM) levels by conversion to ceramide (Cer), resulting in impaired integrin adhesion and reduced integrin mobility. Dual-colour imaging of cortical actin in combination with single molecule tracking of integrins showed that this reduced mobility results from increased coupling to the actin cytoskeleton brought about by Cer formation. As such, our data emphasizes a critical role for the PM local lipid composition in regulating the lateral mobility of integrins and their ability to dynamically increase receptor density for efficient ligand binding in the process of cell adhesion.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep20693DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4751618PMC
February 2016

Functional and molecular characterization of mouse Gata2-independent hematopoietic progenitors.

Blood 2016 Mar 1;127(11):1426-37. Epub 2016 Feb 1.

Erasmus Medical Center Stem Cell Institute, Departments of Cell Biology and Genetics, and Centre for Inflammation Research, Queens Medical Research Institute, and.

The Gata2 transcription factor is a pivotal regulator of hematopoietic cell development and maintenance, highlighted by the fact that Gata2 haploinsufficiency has been identified as the cause of some familial cases of acute myelogenous leukemia/myelodysplastic syndrome and in MonoMac syndrome. Genetic deletion in mice has shown that Gata2 is pivotal to the embryonic generation of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs). It functions in the embryo during endothelial cell to hematopoietic cell transition to affect hematopoietic cluster, HPC, and HSC formation. Gata2 conditional deletion and overexpression studies show the importance of Gata2 levels in hematopoiesis, during all developmental stages. Although previous studies of cell populations phenotypically enriched in HPCs and HSCs show expression of Gata2, there has been no direct study of Gata2 expressing cells during normal hematopoiesis. In this study, we generate a Gata2Venus reporter mouse model with unperturbed Gata2 expression to examine the hematopoietic function and transcriptome of Gata2 expressing and nonexpressing cells. We show that all the HSCs are Gata2 expressing. However, not all HPCs in the aorta, vitelline and umbilical arteries, and fetal liver require or express Gata2. These Gata2-independent HPCs exhibit a different functional output and genetic program, including Ras and cyclic AMP response element-binding protein pathways and other Gata factors, compared with Gata2-dependent HPCs. Our results, indicating that Gata2 is of major importance in programming toward HSC fate but not in all cells with HPC fate, have implications for current reprogramming strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood-2015-10-673749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4797020PMC
March 2016

SMAD1 and SMAD5 Expression Is Coordinately Regulated by FLI1 and GATA2 during Endothelial Development.

Mol Cell Biol 2015 Jun 13;35(12):2165-72. Epub 2015 Apr 13.

Lowy Cancer Research Centre and the Prince of Wales Clinical School, The University of New South Wales, Sydney, NSW, Australia Department of Haematology, Prince of Wales Hospital, Sydney, NSW, Australia

The bone morphogenetic protein (BMP)/SMAD signaling pathway is a critical regulator of angiogenic sprouting and is involved in vascular development in the embryo. SMAD1 and SMAD5, the core mediators of BMP signaling, are vital for this activity, yet little is known about their transcriptional regulation in endothelial cells. Here, we have integrated multispecies sequence conservation, tissue-specific chromatin, in vitro reporter assay, and in vivo transgenic data to identify and validate Smad1+63 and the Smad5 promoter as tissue-specific cis-regulatory elements that are active in the developing endothelium. The activity of these elements in the endothelium was dependent on highly conserved ETS, GATA, and E-box motifs, and chromatin immunoprecipitation showed high levels of enrichment of FLI1, GATA2, and SCL at these sites in endothelial cell lines and E11 dorsal aortas in vivo. Knockdown of FLI1 and GATA2 but not SCL reduced the expression of SMAD1 and SMAD5 in endothelial cells in vitro. In contrast, CD31(+) cKit(-) endothelial cells harvested from embryonic day 9 (E9) aorta-gonad-mesonephros (AGM) regions of GATA2 null embryos showed reduced Smad1 but not Smad5 transcript levels. This is suggestive of a degree of in vivo selection where, in the case of reduced SMAD1 levels, endothelial cells with more robust SMAD5 expression have a selective advantage.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/MCB.00239-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4438244PMC
June 2015

Tracking targeted bimodal nanovaccines: immune responses and routing in cells, tissue, and whole organism.

Mol Pharm 2014 Dec 24;11(12):4299-313. Epub 2014 Oct 24.

Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center , Nijmegen, The Netherlands.

Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs), involved in the induction of immunity and currently exploited for antitumor immunotherapies. An optimized noninvasive imaging modality capable of determining and quantifying DC-targeted nanoparticle (NP) trajectories could provide valuable information regarding therapeutic vaccine outcome. Here, targeted poly(d,l-lactide-co-glycolide) nanoparticles (PLGA NPs) recognizing DC receptors were equipped with superparamagnetic iron oxide particles (SPIO) or gold nanoparticles with fluorescently labeled antigen. The fluorescent label allowed for rapid analysis and quantification of DC-specific uptake of targeted PLGA NPs in comparison to uptake by other cells. Transmission electron microscopy (TEM) showed that a fraction of the encapsulated antigen reached the lysosomal compartment of DCs, where SPIO and gold were already partially released. However, part of the PLGA NPs localized within the cytoplasm, as confirmed by confocal microscopy. DCs targeted with NPs carrying SPIO or fluorescent antigen were detected within lymph nodes as early as 1 h after injection by magnetic resonance imaging (MRI). Despite the fact that targeting did not markedly affect PLGA NP biodistribution on organism and tissue level, it increased delivery of NPs to DCs residing in peripheral lymph nodes and resulted in enhanced T cell proliferation. In conclusion, two imaging agents within a single carrier allows tracking of targeted PLGA NPs at the subcellular, cellular, and organismal levels, thereby facilitating the rational design of in vivo targeted vaccination strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/mp400717rDOI Listing
December 2014

Syntenin-1 and ezrin proteins link activated leukocyte cell adhesion molecule to the actin cytoskeleton.

J Biol Chem 2014 May 24;289(19):13445-60. Epub 2014 Mar 24.

From the Nanobiophysics, MIRA Institute for Biomedical Technology and Technical Medicine and MESA+ Institute for Nanotechnology, University of Twente, 7500AE Enschede, The Netherlands.

Activated leukocyte cell adhesion molecule (ALCAM) is a type I transmembrane protein member of the immunoglobulin superfamily of cell adhesion molecules. Involved in important pathophysiological processes such as the immune response, cancer metastasis, and neuronal development, ALCAM undergoes both homotypic interactions with other ALCAM molecules and heterotypic interactions with the surface receptor CD6 expressed at the T cell surface. Despite biochemical and biophysical evidence of a dynamic association between ALCAM and the actin cytoskeleton, no detailed information is available about how this association occurs at the molecular level. Here, we exploit a combination of complementary microscopy techniques, including FRET detected by fluorescence lifetime imaging microscopy and single-cell force spectroscopy, and we demonstrate the existence of a preformed ligand-independent supramolecular complex where ALCAM stably interacts with actin by binding to syntenin-1 and ezrin. Interaction with the ligand CD6 further enhances these multiple interactions. Altogether, our results propose a novel biophysical framework to understand the stabilizing role of the ALCAM supramolecular complex engaged to CD6 during dendritic cell-T cell interactions and provide novel information on the molecular players involved in the formation and signaling of the immunological synapse at the dendritic cell side.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M113.546754DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4036352PMC
May 2014

Lateral mobility of individual integrin nanoclusters orchestrates the onset for leukocyte adhesion.

Proc Natl Acad Sci U S A 2012 Mar 12;109(13):4869-74. Epub 2012 Mar 12.

BioNanophotonics group, Institute for Bioengineering of Catalonia and Centro de Investigación Biomédica en Red en Bioingeniería, Biomateriales y Nanomedicina, Baldiri Reixac 15-21, 08028 Barcelona, Spain.

Integrins are cell membrane adhesion receptors involved in morphogenesis, immunity, tissue healing, and metastasis. A central, yet unresolved question regarding the function of integrins is how these receptors regulate both their conformation and dynamic nanoscale organization on the membrane to generate adhesion-competent microclusters upon ligand binding. Here we exploit the high spatial (nanometer) accuracy and temporal resolution of single-dye tracking to dissect the relationship between conformational state, lateral mobility, and microclustering of the integrin receptor lymphocyte function-associated antigen 1 (LFA-1) expressed on immune cells. We recently showed that in quiescent monocytes, LFA-1 preorganizes in nanoclusters proximal to nanoscale raft components. We now show that these nanoclusters are primarily mobile on the cell surface with a small (ca. 5%) subset of conformational-active LFA-1 nanoclusters preanchored to the cytoskeleton. Lateral mobility resulted crucial for the formation of microclusters upon ligand binding and for stable adhesion under shear flow. Activation of high-affinity LFA-1 by extracellular Ca(2+) resulted in an eightfold increase on the percentage of immobile nanoclusters and cytoskeleton anchorage. Although having the ability to bind to their ligands, these active nanoclusters failed to support firm adhesion in static and low shear-flow conditions because mobility and clustering capacity were highly compromised. Altogether, our work demonstrates an intricate coupling between conformation and lateral diffusion of LFA-1 and further underscores the crucial role of mobility for the onset of LFA-1 mediated leukocyte adhesion.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.1116425109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3323969PMC
March 2012

Infection of dendritic cells with herpes simplex virus type 1 induces rapid degradation of CYTIP, thereby modulating adhesion and migration.

Blood 2011 Jul 11;118(1):107-15. Epub 2011 May 11.

Department of Immune Modulation at the Department of Dermatology, University Hospital Erlangen, Erlangen, Germany.

Immune responses require spatial and temporal coordinated interactions between different cell types within distinct microenvironments. This dynamic interplay depends on the competency of the involved cells, predominantly leukocytes, to actively migrate to defined sites of cellular encounters in various tissues. Because of their unique capacity to transport antigen from the periphery to secondary lymphoid tissues for the activation of naive T cells, dendritic cells (DCs) play a key role in the initiation and orchestration of adaptive immune responses. Therefore, pathogen-mediated interference with this process is a very effective way of immune evasion. CYTIP (cytohesin-interacting protein) is a key regulator of DC motility. It has previously been described to control LFA-1 deactivation and to regulate DC adherence. CYTIP expression is up-regulated during DC maturation, enabling their transition from the sessile to the motile state. Here, we demonstrate that on infection of human monocyte-derived DCs with herpes simplex virus type 1 (HSV-1), CYTIP is rapidly degraded and as a consequence β-2 integrins, predominantly LFA-1, are activated. Furthermore, we show that the impairment of migration in HSV-1-infected DCs is in part the result of this increased integrin-mediated adhesion. Thus, we propose a new mechanism of pathogen-interference with central aspects of leukocyte biology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood-2010-07-294363DOI Listing
July 2011

The lymphoid chemokine CCL21 triggers LFA-1 adhesive properties on human dendritic cells.

Immunol Cell Biol 2011 Mar 31;89(3):458-65. Epub 2010 Aug 31.

Department of Tumor Immunology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.

Dendritic cells (DCs) are the most potent APCs, involved in the induction of immunity and tolerance. Recently we showed that during differentiation of human DCs from monocyte precursors, Lymphocyte function-associated antigen-1 (LFA-1)-binding capacity is lost, although integrin expression levels were maintained constant, suggesting a different regulation mechanism of this integrin on different cell types. However, the exact role of LFA-1 in DC adhesion and migration remains obscure. Chemokines are potent regulators of integrin function, influencing migratory and adhesive properties of leukocytes. Here, we show that upon vaccination of cancer patients with human DCs, cells that have migrated in vivo into the lymph nodes upregulated the active form of LFA-1. We further show that exposure of human DCs to the lymphoid chemokine CCL21 specifically restores the high-affinity form of LFA-1 and induces binding to its ligand ICAM-1 under low shear stress. Our data indicate that on DCs LFA-1 may function as an inducible anchor during lymphatic transmigration or within the lymph nodes. A thorough understanding of the adhesive events during the DC life cycle will help to improve the outcome of DC-based antitumor clinical trials.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/icb.2010.103DOI Listing
March 2011