43 results on '"Van Hoof D"'
Search Results
2. Stem cell therapies (revision number 29)
- Author
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Atkinson, A, primary and van Hoof, D, additional
- Published
- 2016
- Full Text
- View/download PDF
3. Stem cell therapies (revision number 27)
- Author
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Atkinson, A, primary and van Hoof, D, additional
- Published
- 2012
- Full Text
- View/download PDF
4. Preparation of a Methaemocyanin of Astacus Leptodactylus Regenerable with Hydroxylamine
- Author
-
Witters, R., Van Hoof, D., Deleersnijder, W., Tahon, J.-P., Lontie, R., and Linzen, Bernt, editor
- Published
- 1986
- Full Text
- View/download PDF
5. Plangebied Noorderweg 27 te Noordwijk, gemeente Marum; archeologisch vooronderzoek: een verkennend veldonderzoek
- Author
-
drs. B.I. van Hoof & D. van den Berg and RAAP Archeologisch Adviesbureau
- Subjects
Archaeology ,booronderzoek ,archeologie - Published
- 2015
6. Plangebied Akkersplein te Heerenveen Gemeente Heerenveen Archeologisch vooronderzoek: een bureauen inventariserend veldonderzoek
- Author
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drs. B.I. van Hoof & D. van den Berg and RAAP Archeologisch Adviesbureau
- Subjects
Archaeology ,booronderzoek ,archeologie - Published
- 2013
7. Plangebied Akkersplein te Heerenveen : gemeente Heerenveen : archeologisch vooronderzoek: een bureau- en inventariserend veldonderzoek
- Author
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Hoof, B.I. van, Berg, D. van den, [auteurs: B.I. van Hoof & D. van den Berg], and Weesp : RAAP Archeologisch Adviesbureau
- Subjects
Archaeology ,Heerenveen ,archeologie - Abstract
Met lit. opg
- Published
- 2011
8. Plangebied compressorstation Scheemda, gemeente Scheemda : archeologisch vooronderzoek: een inventariserend veldonderzoek
- Author
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Hoof, B.I. van, Berg, D. van den, Zwet, E.J.M. van der, Perger, T.M., B.I. van Hoof, D. van den Berg, E.J.M. van der Zwet, T.M. Perger., and RAAP Archeologisch Adviesbureau
- Subjects
prospectie ,Archaeology ,archeologie - Abstract
Lit.opg.
- Published
- 2008
9. FosfoProteómica cuantitativa en células madre embrionarias humanas
- Author
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Muñoz, J., Pinkse, M.W., Van Hoof, D., Mohammed, S., Mummery, C.L., Heck, Albert, and Krijgsveld, J.
- Subjects
Sistemas biológicos ,Biotecnología ,Proteómica - Published
- 2008
10. Plangebied De Finne 2 te Ysbrechtum, gemeente Sneek : archeologisch vooronderzoek: een bureau- en inventariserend veldonderzoek
- Author
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Hoof, B.I. van, Berg, D. van den, [auteurs: B.I. van Hoof & D. van den Berg], RAAP Archeologisch Adviesbureau, and Weesp : RAAP Archeologisch Adviesbureau
- Subjects
prospectie ,Archaeology ,Ysbrechtum ,archeologie - Abstract
Met lit. opg
- Published
- 2008
11. Konkelswei 25 Oosterbierum : Gemeente Franekeradiel : archeologisch vooronderzoek: een bureau- en inventariserend veldonderzoek
- Author
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Hoof, B.I. van, Berg, D. van den, B.I. van Hoof & D. van den Berg, Amsterdam : RAAP Archeologisch Adviesbureau, and RAAP Archeologisch Adviesbureau
- Subjects
prospectie ,Archaeology ,Oosterbierum ,archeologie - Abstract
Met lit. opg
- Published
- 2007
12. An experimental correction for arginine-to-proline conversion artifacts in SILAC-based quantitative proteomics
- Author
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van Hoof, D., Pinkse, M.W.H., Ward-van Oostwaard, D., Mummery, C.L., Heck, A.J.R., Krijgsveld, J., Biomoleculaire Massaspectrometrie, Massaspectrometrie, Dep Scheikunde, and Dep Farmaceutische wetenschappen
- Subjects
Farmacie(FARM) - Published
- 2007
13. Embryonic stem cell proteomics
- Author
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van Hoof, D., Mummery, C.L., Heck, A.J.R., Krijgsveld, J., Biomoleculaire Massaspectrometrie, Massaspectrometrie, and Dep Scheikunde
- Subjects
Farmacie/Biofarmaceutische wetenschappen (FARM) ,Other medical specialities ,Geneeskunde (GENK) ,Farmacie(FARM) ,Geneeskunde(GENK) ,General [Econometric and Statistical Methods] ,Algemeen onderzoek - Published
- 2006
14. Proteomic Analysis of Stem Cell Differentiation and Early Development
- Author
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van Hoof, D., primary, Krijgsveld, J., additional, and Mummery, C., additional
- Published
- 2012
- Full Text
- View/download PDF
15. The reaction of nitrite with the haemocyanin of Astacus leptodactylus
- Author
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Tahon, J P, Van Hoof, D, Vinckier, C, Witters, R, De Ley, M, and Lontie, R
- Abstract
The reaction of nitrite at pH 5.7 with deoxyhaemocyanin of Astacus leptodactylus yielded methaemocyanin in two one-electron steps, as nitrite was reduced to NO. This methaemocyanin could be almost fully regenerated by an anaerobic treatment with HONH2, in contrast with the methaemocyanin prepared with H2O2. A destruction of active sites on treating oxyhaemocyanin with HONH2 explains the partial regeneration of methaemocyanin under air, as traces of H2O2 are formed in the autoxidation of HONH2. The reaction rate of nitrite with deoxyhaemocyanin is almost 15 times that with oxyhaemocyanin. The slope of -1.0 for the logarithm of the pseudo-first-order rate constants plotted against pH indicates that HNO2 is the reacting species. Methaemocyanin was e.p.r.-undetectable, but a binuclear signal was observed at g = 2 on binding nitrite to methaemocyanin. This signal disappeared with a pKa of 6.50, suggesting that a mu-aquo bridging ligand, which can be replaced by nitrite, is deprotonated to a mu-hydroxo bridging ligand, which resists substitution by nitrite. The intensity of this triplet e.p.r. signal allowed the determination of the association constant of nitrite to the active site of Astacus methaemocyanin and yielded a value of 237 M-1 at pH 5.7. The interpretation by some authors of nitrosylhaemocyanin as a nitrite derivative of semimethaemocyanin is contradicted by this rapid reaction of nitrite with copper(I) in deoxyhaemocyanin and in semi-methaemocyanin and by the low binding constant of nitrite to the active site of methaemocyanin.
- Published
- 1988
- Full Text
- View/download PDF
16. The reaction of hydroxyurea with oxyhaemocyanin and methaemocyanin of the crayfish Astacus leptodactylus and the snail Helix pomatia
- Author
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Van Hoof, D, primary, Witters, R, additional, and Lontie, R, additional
- Published
- 1988
- Full Text
- View/download PDF
17. The Beneluxa Initiative domain task force health technology assessment: a comparison of member countries' past health technology assessments.
- Author
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Vreman RA, van Hoof D, Nachtnebel A, Daems J, van de Casteele M, Fogarty E, Adams R, and Timmers L
- Subjects
- Retrospective Studies, Netherlands, Austria, Technology Assessment, Biomedical methods, Biosimilar Pharmaceuticals
- Abstract
Objective: This study aimed to compare assessments between Beneluxa Initiative member countries' assessments and identify alignments and divergences., Methods: A retrospective comparative analysis was performed that investigated (i) number and type of assessed indications (for Austria (AT), Belgium (BE), Ireland (IE), and the Netherlands (NL)); (ii) added benefit conclusions (for BE, IE, and NL); and (iii) the main arguments underlying differences in conclusions (for BE, IE, and NL). Data were retrieved directly from agency representatives and from public HTA reports. European Medicines Agency approved indications were included for drugs assessed between 2016 and 2020, excluding veterinary drugs, generics, and biosimilars., Results: Only 44 (10 percent) of the 444 included indications were assessed by all four member countries. Between any pair of two countries, the overlap was higher, from 63 (AT-NL) to 188 (BE-IE). Added benefit conclusions matched exactly in 62-74 percent of the indications, depending on the countries compared. In the remaining cases, most often a difference of one added benefit level was observed (e.g., higher vs. equal relative effect). Contradictory outcomes were very rare: only three cases were observed (lower vs. higher effect). When assessing the underlying arguments for seven cases with different outcomes, differences were attributable to slight differences in weighing of evidence and uncertainties rather than disagreement on aspects within the assessment itself., Conclusions: Despite high variability in European HTA procedures, collaboration on HTA between the Beneluxa Initiative member countries is very feasible and would likely not result in added benefit conclusions that would be very different from added benefit conclusions in national procedures.
- Published
- 2023
- Full Text
- View/download PDF
18. Abuse liability assessment of the JUUL system in two nicotine concentrations compared to combustible cigarette, nicotine gum and comparator electronic nicotine delivery system.
- Author
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Goldenson NI, Buchhalter AR, Augustson EM, Rubinstein ML, Van Hoof D, and Henningfield JE
- Subjects
- Adult, Cross-Over Studies, Female, Flavoring Agents, Humans, Male, Nicotine blood, Smokers, Taste, Electronic Nicotine Delivery Systems, Nicotine Chewing Gum, Tobacco Products, Tobacco Use Disorder
- Abstract
Background: To assess the abuse liability of the JUUL System (JS) in 5.0 % (59 mg/mL) and 3.0 % (35 mg/mL) nicotine concentrations., Methods: Adult smokers (N = 146; 45.9 % female; mean age = 41.29 years) were randomized to one of four study flavor arms and then to a within-subjects cross-over sequence for five test product categories: (1) JS 5.0 % nicotine concentration; (2) JS 3.0 % nicotine; (3) usual brand (UB) cigarette; (4) 4 mg mint nicotine gum; (5) comparator ENDS (VUSE Alto 5.0 % nicotine). Products were tested by ad libitum use (5 min for ENDS and cigarette; 30 min for gum); nicotine pharmacokinetic (PK) parameters and subjective effects were assessed following use., Results: Maximum plasma nicotine concentration (C
max-BL ), rate of plasma nicotine rise and total nicotine exposure (AUC0-60-BL ) of UB cigarette were significantly greater than all other test products. The comparator ENDS was significantly greater than 5.0 % and 3.0 % JS and nicotine gum on Cmax-BL , rate of plasma nicotine rise, and AUC0-60-BL ; Cmax-BL of JS 5.0 % was significantly greater than JS 3.0 % and nicotine gum. Product liking and satisfying effects were significantly highest for the UB cigarette; JS products and comparator ENDS did not significantly differ and were rated higher than nicotine gum on most subjective measures., Conclusions: These results suggest that the abuse liability of both 5.0 % and 3.0 % JS is: (1) substantially lower than UB cigarette; (2) somewhat lower than comparator ENDS; and (3) higher than nicotine gum. Additionally, the abuse liability of JS 5.0 % is somewhat higher than JS 3.0 %., (Copyright © 2020 Juul Labs, Inc. Published by Elsevier B.V. All rights reserved.)- Published
- 2020
- Full Text
- View/download PDF
19. Optimization and testing of dried antibody tube: The EuroFlow LST and PIDOT tubes as examples.
- Author
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van der Velden VHJ, Flores-Montero J, Perez-Andres M, Martin-Ayuso M, Crespo O, Blanco E, Kalina T, Philippé J, Bonroy C, de Bie M, Te Marvelde J, Teodosio C, Corral Mateos A, Kanderová V, van der Burg M, Van Hoof D, van Dongen JJM, and Orfao A
- Subjects
- Humans, Hematologic Neoplasms diagnosis, Immunophenotyping methods
- Abstract
Within EuroFlow, we recently developed screening tubes for hematological malignancies and immune deficiencies. Pipetting of antibodies for such 8-color 12-marker tubes however is time-consuming and prone to operational mistakes. We therefore evaluated dried formats of the lymphocytosis screening tube (LST) and of the primary immune deficiency orientation tube (PIDOT). Both tubes were evaluated on normal and/or on patient samples, comparing the mean fluorescence intensity of specific lymphocyte populations. Our data show that the dried tubes and liquid counterparts give highly comparable staining results, particularly when analyzed in multidimensional plots. In addition, the use of dried tubes may result in a reduced staining variability between different samples and thereby contributes to the generation of more robust data. Therefore, by using ready-to-use reagents in a dried single test tube format, the laboratory efficiency and quality will be improved., (Copyright © 2017 Erasmus MC, University Medical Center Rotterdam. Published by Elsevier B.V. All rights reserved.)
- Published
- 2019
- Full Text
- View/download PDF
20. Phosphorylation of NEUROG3 Links Endocrine Differentiation to the Cell Cycle in Pancreatic Progenitors.
- Author
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Krentz NAJ, van Hoof D, Li Z, Watanabe A, Tang M, Nian C, German MS, and Lynn FC
- Subjects
- Animals, Basic Helix-Loop-Helix Transcription Factors genetics, Endocrine Cells metabolism, Gene Expression Regulation, Developmental physiology, Humans, Islets of Langerhans cytology, Mice, Nerve Tissue Proteins genetics, Phosphorylation physiology, Stem Cells metabolism, Basic Helix-Loop-Helix Transcription Factors metabolism, Cell Cycle physiology, Cell Differentiation physiology, Nerve Tissue Proteins metabolism, Pancreas cytology, Pancreas metabolism, Stem Cells cytology
- Abstract
During pancreatic development, proliferating pancreatic progenitors activate the proendocrine transcription factor neurogenin 3 (NEUROG3), exit the cell cycle, and differentiate into islet cells. The mechanisms that direct robust NEUROG3 expression within a subset of progenitor cells control the size of the endocrine population. Here we demonstrate that NEUROG3 is phosphorylated within the nucleus on serine 183, which catalyzes its hyperphosphorylation and proteosomal degradation. During progression through the progenitor cell cycle, NEUROG3 phosphorylation is driven by the actions of cyclin-dependent kinases 2 and 4/6 at G
1 /S cell-cycle checkpoint. Using models of mouse and human pancreas development, we show that lengthening of the G1 phase of the pancreatic progenitor cell cycle is essential for proper induction of NEUROG3 and initiation of endocrine cell differentiation. In sum, these studies demonstrate that progenitor cell-cycle G1 lengthening, through its actions on stabilization of NEUROG3, is an essential variable in normal endocrine cell genesis., (Copyright © 2017 Elsevier Inc. All rights reserved.)- Published
- 2017
- Full Text
- View/download PDF
21. PEGylated IL-10 Activates Kupffer Cells to Control Hypercholesterolemia.
- Author
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Chan IH, Van Hoof D, Abramova M, Bilardello M, Mar E, Jorgensen B, McCauley S, Bal H, Oft M, Van Vlasselaer P, and Mumm JB
- Subjects
- Adolescent, Adult, Aged, Aged, 80 and over, Animals, Cholesterol immunology, Female, Humans, Hypercholesterolemia blood, Hypercholesterolemia immunology, Immunologic Factors chemistry, Immunologic Factors pharmacology, Interleukin-10 chemistry, Interleukin-10 pharmacology, Kupffer Cells immunology, Male, Mice, Inbred C57BL, Middle Aged, Polyethylene Glycols chemistry, Polyethylene Glycols pharmacology, Polyethylene Glycols therapeutic use, Recombinant Proteins chemistry, Recombinant Proteins pharmacology, Recombinant Proteins therapeutic use, Young Adult, Cholesterol blood, Hypercholesterolemia drug therapy, Immunologic Factors therapeutic use, Interleukin-10 therapeutic use, Kupffer Cells drug effects, Phagocytosis drug effects
- Abstract
Interleukin-10 (IL-10) is a multifunctional cytokine that exerts potent context specific immunostimulatory and immunosuppressive effects. We have investigated the mechanism by which PEGylated rIL-10 regulates plasma cholesterol in mice and humans. In agreement with previous work on rIL-10, we report that PEGylated rIL-10 harnesses the myeloid immune system to control total plasma cholesterol levels. We have discovered that PEG-rMuIL-10's dramatic lowering of plasma cholesterol is dependent on phagocytotic cells. In particular, PEG-rHuIL-10 enhances cholesterol uptake by Kupffer cells. In addition, removal of phagocytotic cells dramatically increases plasma cholesterol levels, suggesting for the first time that immunological cells are implicitly involved in regulating total cholesterol levels. These data suggest that treatment with PEG-rIL-10 potentiates endogenous cholesterol regulating cell populations not currently targeted by standard of care therapeutics. Furthermore, we show that IL-10's increase of Kupffer cell cholesterol phagocytosis is concomitant with decreases in liver cholesterol and triglycerides. This leads to the reversal of early periportal liver fibrosis and facilitates the restoration of liver health. These data recommend PEG-rIL-10 for evaluation in the treatment of fatty liver disease and preventing its progression to non-alcoholic steatohepatitis. In direct confirmation of our in vivo findings in the treatment of hypercholesterolemic mice with PEG-rMuIL-10, we report that treatment of hypercholesterolemic cancer patients with PEG-rHuIL-10 lowers total plasma cholesterol by up to 50%. Taken together these data suggest that PEG-rIL-10's cholesterol regulating biology is consistent between mice and humans.
- Published
- 2016
- Full Text
- View/download PDF
22. Simultaneous flow cytometric analysis of IFN-γ and CD4 mRNA and protein expression kinetics in human peripheral blood mononuclear cells during activation.
- Author
-
Van Hoof D, Lomas W, Hanley MB, and Park E
- Subjects
- Gene Expression Regulation, Humans, Kinetics, CD4 Antigens biosynthesis, Flow Cytometry methods, Interferon-gamma biosynthesis, Leukocytes, Mononuclear metabolism, RNA, Messenger biosynthesis
- Abstract
The application of fluorescently-labeled antibodies for flow cytometric identification and characterization of specific cell types within heterogeneous populations by their protein expression profile is well established. While detection of proteins is informative, concomitant transcript analysis in the same cells would provide a more complete and comprehensive view of intracellular signaling events. We recently reported on the efficient detection of RNA in suspension cells for flow cytometric analysis. The improved RNA flow cytometry procedure described here allows for the specific labeling of multiple RNA species, and is compatible with antibody-based targeting of extracellular and intracellular antigens for multiplexing purposes. To show proof of concept, human peripheral blood mononuclear cells were stimulated with phorbol 12-myristate 13-acetate and ionomycin for a maximum of 5 h, during which their CD4 and interferon-gamma (IFN-γ) transcript and protein levels were monitored. Substantial and increasing numbers of IFN-γ mRNA+ cells were detected within 30 min after initiation of induction, while IFN-γ protein+ cells could only be discerned at 1 h and beyond. Surprisingly, resting lymphocytes contained less CD4 mRNA but more of the protein per cell compared with monocytes, revealing a difference in the relationship of transcript and protein levels in these two cell types. We additionally applied monensin, which is commonly used to block cytokine secretion, and found that IFN-γ mRNA can still be analyzed consistently using the improved RNA flow cytometry staining method. Notably, a subset of IFN-γ mRNA(-)/protein+ cells that were not observed in the absence of monensin became apparent at the 5-h mark. This subset probably represents cells that have accumulated IFN-γ protein, but no longer transcribe mRNA. Collectively, the results described here exemplify how the improved RNA flow cytometry labeling procedure can be applied to simultaneously assess mRNA and protein dynamics to gain insight into the regulation of gene transcription and translation in individual cells., (© 2014 International Society for Advancement of Cytometry.)
- Published
- 2014
- Full Text
- View/download PDF
23. Directed differentiation of human pluripotent stem cells along the pancreatic endocrine lineage.
- Author
-
Van Hoof D and Liku ME
- Subjects
- Activins physiology, Animals, CHO Cells, Cell Culture Techniques, Cell Lineage, Cricetinae, Culture Media, Conditioned, Humans, Pancreas cytology, Pluripotent Stem Cells cytology, Cell Differentiation, Pluripotent Stem Cells physiology
- Abstract
Many research groups are engaged in using human pluripotent stem cells (hPSCs) to generate surrogate pancreatic β-cells for transplantation into diabetic patients. However, to our knowledge, there is no report on the successful generation of glucose-responsive insulin-producing β-cells from hPSCs in vitro. Below, we outline a method that is based on published protocols as well as our own experience by which one can differentiate hPSCs along the pancreatic lineage to generate insulin-producing β-cell-like cells. The protocol, which spans five distinct stages, is an attempt to recapitulate the derivation of pancreatic β-cells in vitro as they form in the developing embryo. We included details on materials and techniques, suggest ways to customize it to your hPSC line of choice, added notes on how to monitor and analyze the cells during differentiation, and indicate what results can be expected.
- Published
- 2013
- Full Text
- View/download PDF
24. Chromatin insulator elements block transgene silencing in engineered human embryonic stem cell lines at a defined chromosome 13 locus.
- Author
-
Macarthur CC, Xue H, Van Hoof D, Lieu PT, Dudas M, Fontes A, Swistowski A, Touboul T, Seerke R, Laurent LC, Loring JF, German MS, Zeng X, Rao MS, Lakshmipathy U, Chesnut JD, and Liu Y
- Subjects
- Cell Differentiation, Cell Line, Transformed, Cell Lineage, Chromosomes, Human, Pair 13, Genes, Reporter, Genetic Loci, Genetic Vectors, Green Fluorescent Proteins genetics, Humans, Integrases genetics, Integrases metabolism, Peptide Elongation Factor 1 genetics, Promoter Regions, Genetic, Transgenes, Chromatin genetics, Embryonic Stem Cells cytology, Embryonic Stem Cells metabolism, Genetic Engineering methods, Insulator Elements genetics, Recombination, Genetic
- Abstract
Lineage reporters of human embryonic stem cell (hESC) lines are useful for differentiation studies and drug screening. Previously, we created reporter lines driven by an elongation factor 1 alpha (EF1α) promoter at a chromosome 13q32.3 locus in the hESC line WA09 and an abnormal hESC line BG01V in a site-specific manner. Expression of reporters in these lines was maintained in long-term culture at undifferentiated state. However, when these cells were differentiated into specific lineages, reduction in reporter expression was observed, indicating transgene silencing. To develop an efficient and reliable genetic engineering strategy in hESCs, we used chromatin insulator elements to flank single-copy transgenes and integrated the combined expression constructs via PhiC31/R4 integrase-mediated recombination technology to the chromosome 13 locus precisely. Two copies of cHS4 double-insulator sequences were placed adjacent to both 5' and 3' of the promoter reporter constructs. The green fluorescent protein (GFP) gene was driven by EF1α or CMV early enhancer/chicken β actin (CAG) promoter. In the engineered hESC lines, for both insulated CAG-GFP and EF1α-GFP, constitutive expression at the chromosome 13 locus was maintained during prolonged culture and in directed differentiation assays toward diverse types of neurons, pancreatic endoderm, and mesodermal progeny. In particular, described here is the first normal hESC fluorescent reporter line that robustly expresses GFP in both the undifferentiated state and throughout dopaminergic lineage differentiation. The dual strategy of utilizing insulator sequences and integration at the constitutive chromosome 13 locus ensures appropriate transgene expression. This is a valuable tool for lineage development study, gain- and loss-of-function experiments, and human disease modeling using hESCs.
- Published
- 2012
- Full Text
- View/download PDF
25. Differentiation of human embryonic stem cells into pancreatic endoderm in patterned size-controlled clusters.
- Author
-
Van Hoof D, Mendelsohn AD, Seerke R, Desai TA, and German MS
- Subjects
- Cell Size, Embryonic Stem Cells metabolism, Endoderm metabolism, Gene Expression, Homeodomain Proteins genetics, Homeodomain Proteins metabolism, Humans, Insulin-Secreting Cells cytology, Insulin-Secreting Cells metabolism, Pancreas growth & development, Pancreas metabolism, Trans-Activators genetics, Trans-Activators metabolism, Cell Culture Techniques methods, Cell Differentiation, Embryonic Stem Cells cytology, Endoderm cytology, Pancreas cytology
- Abstract
Pancreatic β-cells function optimally when clustered in islet-like structures. However, nutrient and oxygen deprivation limits the viability of cells at the core of excessively large clusters. Hence, production of functional β-cells from human embryonic stem cells (hESCs) for patients with diabetes would benefit from the growth and differentiation of these cells in size-controlled aggregates. In this study, we controlled cluster size by seeding hESCs onto glass cover slips patterned by the covalent microcontact-printing of laminin in circular patches of 120 μm in diameter. These were used as substrates to grow and differentiate hESCs first into SOX17-positive/SOX7-negative definitive endoderm, after which many clusters released and formed uniformly sized three-dimensional clusters. Both released clusters and those that remained attached differentiated into HNF1β-positive primitive gut tube-like cells with high efficiency. Further differentiation yielded pancreatic endoderm-like cells that co-expressed PDX1 and NKX6.1. Controlling aggregate size allows efficient production of uniformly-clustered pancreatic endocrine precursors for in vivo engraftment or further in vitro maturation., (Copyright © 2011 Elsevier B.V. All rights reserved.)
- Published
- 2011
- Full Text
- View/download PDF
26. Identification of cell surface proteins for antibody-based selection of human embryonic stem cell-derived cardiomyocytes.
- Author
-
Van Hoof D, Dormeyer W, Braam SR, Passier R, Monshouwer-Kloots J, Ward-van Oostwaard D, Heck AJ, Krijgsveld J, and Mummery CL
- Subjects
- Antibodies metabolism, Biomarkers metabolism, Blotting, Western, Cell Differentiation, Cell Separation, Embryonic Stem Cells cytology, Fibrillins, Glycoproteins metabolism, Humans, Isotope Labeling, Mass Spectrometry, Membrane Proteins metabolism, Microfilament Proteins metabolism, Microscopy, Fluorescence, Myocytes, Cardiac cytology, Biomarkers analysis, Embryonic Stem Cells metabolism, Membrane Proteins analysis, Myocytes, Cardiac metabolism, Proteomics methods
- Abstract
The absence of identified cell surface proteins and corresponding antibodies to most differentiated derivatives of human embryonic stem cells (hESCs) has largely limited selection of specific cell types from mixed cell populations to genetic approaches. Here, we describe the use of mass spectrometry (MS)-based proteomics on cell membrane proteins isolated from hESCs that were differentiated into cardiomyocytes to identify candidate proteins for this particular lineage. Quantitative MS distinguished cardiomyocyte-specific plasma membrane proteins that were highly enriched or detected only in cardiomyocytes derived from hESCs and human fetal hearts compared with a heterogeneous pool of hESC-derived differentiated cells. For several candidates, cardiomyocyte-specific expression and cell surface localization were verified by conventional antibody-based methodologies. Using an antibody against elastin microfibril interfacer 2 (EMILIN2), we demonstrate that cardiomyocytes can be sorted from live cell populations. Besides showing that MS-based membrane proteomics is a powerful tool to identify candidate proteins that allow purification of specific cell lineages from heterogeneous populations, this approach generated a plasma membrane proteome profile suggesting signaling pathways that control cell behavior.
- Published
- 2010
- Full Text
- View/download PDF
27. Derivation of insulin-producing cells from human embryonic stem cells.
- Author
-
Van Hoof D, D'Amour KA, and German MS
- Subjects
- Cell Differentiation, Diabetes Mellitus etiology, Diabetes Mellitus therapy, Endoderm cytology, Humans, Insulin genetics, Insulin metabolism, Transcription Factors metabolism, Embryonic Stem Cells cytology, Insulin-Secreting Cells cytology
- Abstract
The potential of pluripotent human cells, such as human embryonic stem cells (hESCs) and induced pluripotent stem (iPS) cells, to differentiate into any adult cell type makes them ideally suited for the generation of various somatic cells and tissues in vitro. This remarkable differentiation capacity permits analyzing aspects of human embryonic development in the laboratory, as well as generating specialized adult human cells for screening drugs, and for replacing tissues damaged by injury or degenerative diseases, such as diabetes. Understanding and controlling the fundamental processes that drive the differentiation of specialized cells are the keys to the eventual application of this technology to patients. In this review, we discuss the different protocols developed that are aimed at deriving beta-cells from hESCs. Despite many differences, successful strategies share a general adherence to the normal differentiation pathway through definitive endoderm. Mimicking normal pancreagenesis offers the best strategy for producing glucose-responsive insulin-producing cells in vitro for people with diabetes.
- Published
- 2009
- Full Text
- View/download PDF
28. Phosphorylation dynamics during early differentiation of human embryonic stem cells.
- Author
-
Van Hoof D, Muñoz J, Braam SR, Pinkse MW, Linding R, Heck AJ, Mummery CL, and Krijgsveld J
- Subjects
- Bone Morphogenetic Proteins pharmacology, CDC2 Protein Kinase metabolism, Cell Differentiation drug effects, Cell Differentiation physiology, Cyclin-Dependent Kinase 2 metabolism, Embryonic Stem Cells drug effects, HeLa Cells, Humans, Phosphoproteins drug effects, Phosphorylation drug effects, Phosphorylation physiology, Pluripotent Stem Cells drug effects, Proteome drug effects, SOXB1 Transcription Factors antagonists & inhibitors, SOXB1 Transcription Factors metabolism, Signal Transduction drug effects, Signal Transduction physiology, Embryonic Stem Cells metabolism, Phosphoproteins metabolism, Pluripotent Stem Cells metabolism, Proteome metabolism
- Abstract
Pluripotent stem cells self-renew indefinitely and possess characteristic protein-protein networks that remodel during differentiation. How this occurs is poorly understood. Using quantitative mass spectrometry, we analyzed the (phospho)proteome of human embryonic stem cells (hESCs) during differentiation induced by bone morphogenetic protein (BMP) and removal of hESC growth factors. Of 5222 proteins identified, 1399 were phosphorylated on 3067 residues. Approximately 50% of these phosphosites were regulated within 1 hr of differentiation induction, revealing a complex interplay of phosphorylation networks spanning different signaling pathways and kinase activities. Among the phosphorylated proteins was the pluripotency-associated protein SOX2, which was SUMOylated as a result of phosphorylation. Using the data to predict kinase-substrate relationships, we reconstructed the hESC kinome; CDK1/2 emerged as central in controlling self-renewal and lineage specification. The findings provide new insights into how hESCs exit the pluripotent state and present the hESC (phospho)proteome resource as a complement to existing pluripotency network databases.
- Published
- 2009
- Full Text
- View/download PDF
29. Feeder-free monolayer cultures of human embryonic stem cells express an epithelial plasma membrane protein profile.
- Author
-
Van Hoof D, Braam SR, Dormeyer W, Ward-van Oostwaard D, Heck AJ, Krijgsveld J, and Mummery CL
- Subjects
- Antigens, Differentiation metabolism, Cell Adhesion, Cell Differentiation, Cells, Cultured, Coculture Techniques methods, Collagen metabolism, Drug Combinations, Embryonic Stem Cells metabolism, Epithelial Cells metabolism, Humans, Laminin metabolism, Mass Spectrometry, Microscopy, Fluorescence, Proteoglycans metabolism, Cell Membrane metabolism, Embryonic Stem Cells cytology, Epithelial Cells cytology, Membrane Proteins metabolism
- Abstract
Human embryonic stem cells (hESCs) are often cocultured on mitotically inactive fibroblast feeder cells to maintain their undifferentiated state. Under these growth conditions, hESCs form multilayered colonies of morphologically heterogeneous cells surrounded by flattened mesenchymal cells. In contrast, hESCs grown in feeder cell-conditioned medium on Matrigel instead tend to grow as monolayers with uniform morphology. Using mass spectrometry and immunofluorescence microscopy, we showed that hESCs under these conditions primarily express proteins belonging to epithelium-related cell-cell adhesion complexes, including adherens junctions, tight junctions, desmosomes, and gap junctions. This indicates that monolayers of hESCs cultured under feeder-free conditions retain a homogeneous epithelial phenotype similar to that of the upper central cell layer of colonies maintained on feeder cells. Notably, feeder-free hESCs also coexpressed vimentin, which is usually associated with mesenchyme, suggesting that these cells may have undergone epithelium-to-mesenchyme transitions, indicating differentiation. However, if grown on a "soft" substrate (Hydrogel), intracellular vimentin levels were substantially reduced. Moreover, when hESCs were transferred back to feeder cells, expression of vimentin was again absent from the epithelial cell population. These results imply that on tissue culture substrates, vimentin expression is most likely a stress-induced response, unrelated to differentiation. Disclosure of potential conflicts of interest is found at the end of this article.
- Published
- 2008
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30. A practical guide for the identification of membrane and plasma membrane proteins in human embryonic stem cells and human embryonal carcinoma cells.
- Author
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Dormeyer W, van Hoof D, Mummery CL, Krijgsveld J, and Heck AJ
- Subjects
- Humans, Membrane Proteins chemistry, Membrane Proteins metabolism, Protein Denaturation, Trypsin metabolism, Cell Membrane metabolism, Embryonal Carcinoma Stem Cells metabolism, Embryonic Stem Cells metabolism, Membrane Proteins isolation & purification, Proteomics methods
- Abstract
The identification of (plasma) membrane proteins in cells can provide valuable insights into the regulation of their biological processes. Pluripotent cells such as human embryonic stem cells and embryonal carcinoma cells are capable of unlimited self-renewal and share many of the biological mechanisms that regulate proliferation and differentiation. The comparison of their membrane proteomes will help unravel the biological principles of pluripotency, and the identification of biomarker proteins in their plasma membranes is considered a crucial step to fully exploit pluripotent cells for therapeutic purposes. For these tasks, membrane proteomics is the method of choice, but as indicated by the scarce identification of membrane and plasma membrane proteins in global proteomic surveys it is not an easy task. In this minireview, we first describe the general challenges of membrane proteomics. We then review current sample preparation steps and discuss protocols that we found particularly beneficial for the identification of large numbers of (plasma) membrane proteins in human tumour- and embryo-derived stem cells. Our optimized assembled protocol led to the identification of a large number of membrane proteins. However, as the composition of cells and membranes is highly variable we still recommend adapting the sample preparation protocol for each individual system.
- Published
- 2008
- Full Text
- View/download PDF
31. Proteomics and human embryonic stem cells.
- Author
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Van Hoof D, Heck AJ, Krijgsveld J, and Mummery CL
- Subjects
- Humans, Mass Spectrometry, Proteins analysis, Embryonic Stem Cells, Proteomics methods
- Abstract
The derivation of human embryonic stem cells (hESCs) brought cell therapy-based regenerative medicine significantly closer to clinical application. However, expansion of undifferentiated cells and their directed differentiation in vitro have proven difficult to control. This is mainly because of a lack of knowledge of the intracellular signaling events that direct these complex processes. Additionally, extracellular factors, either secreted by feeder cells that support self-renewal and maintain pluripotency or present in serum supplementing proprietary culture media, that influence hESC behavior are largely unknown. Xeno-free media that effectively support long-term hESC self-renewal and differentiation to specific types of specialized cells are only slowly becoming available. Microarray-based transcriptome analyses have produced valuable gene expression profiles of hESCs and indicated changes in transcription that occur during differentiation. However, proteins are the actual effectors of these events and changes in their levels do not always match changes in their corresponding mRNA. Furthermore, information on posttranslational modifications that influence the activity of pivotal proteins is still largely missing. Over the years, mass spectrometry has experienced major breakthroughs in high-throughput identification of proteins and posttranslational modifications in cells under different conditions. Mass spectrometry-based proteomic techniques are being applied with increasing frequency to analyze hESCs, as well as media conditioned by feeder cells, and have generated proteome profiles that not only support, but also complement, existing microarray data. In this review, the various proteomic studies on hESCs and feeder cells are discussed. In a meta-analysis, comparison of published data sets distinguished 32 intracellular proteins and 16 plasma membrane proteins that are present in multiple hESC lines but not in differentiated cells, which were therefore likely to include proteins important for hESCs. In addition, 13 and 24 proteins, respectively, were commonly found in different feeder cell lines of mouse and human origin, some of which may be extracellular signaling molecules that play a key role in the undifferentiated propagation of hESCs. These findings underscore the power of mass spectrometry-based techniques to identify novel proteins associated with hESCs by studying these cells in an unbiased, discovery-oriented manner on a proteome-wide scale.
- Published
- 2008
- Full Text
- View/download PDF
32. Plasma membrane proteomics of human embryonic stem cells and human embryonal carcinoma cells.
- Author
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Dormeyer W, van Hoof D, Braam SR, Heck AJ, Mummery CL, and Krijgsveld J
- Subjects
- Animals, Cell Membrane metabolism, Chromatography, Liquid, Embryonal Carcinoma Stem Cells metabolism, Gene Expression Profiling, Humans, Membrane Proteins genetics, Mice, Organelles metabolism, Proteomics, Species Specificity, Tandem Mass Spectrometry, Embryonic Stem Cells metabolism, Membrane Proteins metabolism
- Abstract
Human embryonic stem cells (hESCs) are of immense interest in regenerative medicine as they can self-renew indefinitely and can give rise to any adult cell type. Human embryonal carcinoma cells (hECCs) are the malignant counterparts of hESCs found in testis tumors. hESCs that have acquired chromosomal abnormalities in culture are essentially indistinguishable from hECC. Direct comparison of karyotypically normal hESCs with hECCs could lead to understanding differences between their mechanisms of growth control and contribute to implementing safe therapeutic use of stem cells without the development of germ cell cancer. While several comparisons of hECCs and hESCs have been reported, their cell surface proteomes are largely unknown, partly because plasma membrane proteomics is still a major challenge. Here, we present a strategy for the identification of plasma membrane proteins that has been optimized for application to the relatively small numbers of stem cells normally available, and that does not require tedious cell fractionation. The method led to the identification of 237 and 219 specific plasma membrane proteins in the hESC line HUES-7 and the hECC line NT2/D1, respectively. In addition to known stemness-associated cell surface markers like ALP, CD9, and CTNNB, a large number of receptors, transporters, signal transducers, and cell-cell adhesion proteins were identified. Our study revealed that several Hedgehog and Wnt pathway members are differentially expressed in hESCs and hECCs including NPC1, FZD2, FZD6, FZD7, LRP6, and SEMA4D, which play a pivotal role in stem cell self-renewal and cancer growth. Various proteins encoded on chromosome 12p, duplicated in testicular cancer, were uniquely identified in hECCs. These included GAPDH, LDHB, YARS2, CLSTN3, CSDA, LRP6, NDUFA9, and NOL1, which are known to be upregulated in testicular cancer. Distinct HLA molecules were revealed on the surface of hESCs and hECCs, despite their low abundance. Results were compared with genomic and proteomic data sets reported previously for mouse ESCs, hECCs, and germ cell tumors. Our data provides a surface signature for HUES-7 and NT2/D1 cells and distinguishes normal hESCs from hECCs, helping explain their 'benign' versus 'malignant' nature.
- Published
- 2008
- Full Text
- View/download PDF
33. An experimental correction for arginine-to-proline conversion artifacts in SILAC-based quantitative proteomics.
- Author
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Van Hoof D, Pinkse MW, Oostwaard DW, Mummery CL, Heck AJ, and Krijgsveld J
- Subjects
- Carbon Isotopes chemistry, Cell Culture Techniques, Cell Line, Humans, Nitrogen Isotopes chemistry, Proteomics standards, Sensitivity and Specificity, Arginine chemistry, Artifacts, Isotope Labeling methods, Mass Spectrometry standards, Peptides chemistry, Proline chemistry, Proteomics methods
- Published
- 2007
- Full Text
- View/download PDF
34. Concise review: trends in stem cell proteomics.
- Author
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Baharvand H, Fathi A, van Hoof D, and Salekdeh GH
- Subjects
- Bodily Secretions chemistry, Bodily Secretions metabolism, Cells, Cultured, Electrophoresis, Gel, Two-Dimensional, Gene Expression Profiling, Humans, Mass Spectrometry, Membrane Proteins analysis, Models, Biological, Protein Array Analysis, Protein Processing, Post-Translational, Proteome metabolism, Transplants, Proteome analysis, Proteomics trends, Stem Cells metabolism
- Abstract
Gene expression analyses of stem cells (SCs) will help to uncover or further define signaling pathways and molecular mechanisms involved in the maintenance of self-renewal, pluripotency, and/or multipotency. In recent years, proteomic approaches have produced a wealth of data identifying proteins and mechanisms involved in SC proliferation and differentiation. Although many proteomics techniques have been developed and improved in peptide and protein separation, as well as mass spectrometry, several important issues, including sample heterogeneity, post-translational modifications, protein-protein interaction, and high-throughput quantification of hydrophobic and low-abundance proteins, still remain to be addressed and require further technical optimization. This review summarizes the methodologies used and the information gathered with proteome analyses of SCs, and it discusses biological and technical challenges for proteomic study of SCs. Disclosure of potential conflicts of interest is found at the end of this article.
- Published
- 2007
- Full Text
- View/download PDF
35. Embryonic stem cell proteomics.
- Author
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Van Hoof D, Mummery CL, Heck AJ, and Krijgsveld J
- Subjects
- Animals, Cell Culture Techniques, Cell Differentiation, Coculture Techniques, Embryo, Mammalian cytology, Gene Expression Profiling, Humans, Mass Spectrometry methods, Mice, Pluripotent Stem Cells chemistry, Proteomics methods
- Abstract
Human embryonic stem cells potentially represent an unlimited source of cells and tissues for regenerative medicine. Understanding signaling events that drive proliferation and specialization of these cells into various differentiated derivatives is of utmost importance for controlling their behavior in vitro. Major progress has been made in unraveling these signaling events with large-scale studies at the transcriptional level, but analysis of protein expression, interaction and modification has been more limited, since it requires different strategies. Recent advances in mass spectrometry-based proteomics indicate that proteome characterization can contribute significantly to our understanding of embryonic stem cell biology. In this article, we review mass spectrometry-based studies of human and mouse embryonic stem cells and their differentiated progeny, as well as studies of conditioned media that have been reported to support self-renewal of the undifferentiated cells in the absence of the more commonly used feeder cells. In addition, we make concise comparisons with related transcriptome profiling reports.
- Published
- 2006
- Full Text
- View/download PDF
36. A quest for human and mouse embryonic stem cell-specific proteins.
- Author
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Van Hoof D, Passier R, Ward-Van Oostwaard D, Pinkse MW, Heck AJ, Mummery CL, and Krijgsveld J
- Subjects
- Animals, Biomarkers, Blotting, Western, Cell Differentiation, Cell Line, Cells, Cultured, Flow Cytometry, Humans, Mice, Microscopy, Fluorescence, Models, Biological, Embryo, Mammalian cytology, Proteomics, Stem Cells metabolism
- Abstract
Embryonic stem cells (ESCs) are of immense interest as they can proliferate indefinitely in vitro and give rise to any adult cell type, serving as a potentially unlimited source for tissue replacement in regenerative medicine. Extensive analyses of numerous human and mouse ESC lines have shown generic similarities and differences at both the transcriptional and functional level. However, comprehensive proteome analyses are missing or are restricted to mouse ESCs. Here we have used an extensive proteomic approach to search for ESC-specific proteins by analyzing the differential protein expression profiles of human and mouse ESCs and their differentiated derivatives. The data sets comprise 1,775 non-redundant proteins identified in human ESCs, 1,532 in differentiated human ESCs, 1,871 in mouse ESCs, and 1,552 in differentiated mouse ESCs with a false positive rate of <0.2%. Comparison of the data sets distinguished 191 proteins exclusively identified in both human and mouse ESCs but not in their differentiated derivatives. Besides well known ESC benchmarks, this subset included many uncharacterized proteins, some of which may be novel ESC-specific markers. To complement the mass spectrometric approach, differential expression of a selection of these proteins was confirmed by Western blotting, immunofluorescence confocal microscopy, and fluorescence-activated cell sorting. Additionally two other independently isolated and cultured human ESC lines as well as their differentiated derivatives were monitored for differential expression of selected proteins. Some of these proteins were identified exclusively in ESCs of all three human lines and may thus serve as generic ESC markers. Our wide scale proteomic approach enabled us to screen thousands of proteins rapidly and select putative ESC-associated proteins for further analysis. Validation by three independent conventional protein analysis techniques shows that our methodology is robust, provides an excellent tool to characterize ESCs at the protein level, and may disclose novel ESC-specific benchmarks.
- Published
- 2006
- Full Text
- View/download PDF
37. Sequence analysis of the non-recurring C-terminal domains shows that insect lipoprotein receptors constitute a distinct group of LDL receptor family members.
- Author
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Rodenburg KW, Smolenaars MM, Van Hoof D, and Van der Horst DJ
- Subjects
- Amino Acid Motifs, Amino Acid Sequence, Animals, Insect Proteins classification, Insect Proteins physiology, Molecular Sequence Data, Multigene Family, Phylogeny, Protein Structure, Tertiary physiology, Receptors, LDL classification, Receptors, LDL physiology, Sequence Alignment, Sequence Analysis, Protein, Sequence Homology, Amino Acid, Insect Proteins chemistry, Insecta metabolism, Receptors, LDL chemistry
- Abstract
Lipoprotein-mediated delivery of lipids in mammals involves endocytic receptors of the low density lipoprotein (LDL) receptor (LDLR) family. In contrast, in insects, the lipoprotein, lipophorin (Lp), functions as a reusable lipid shuttle in lipid delivery, and these animals, therefore, were not supposed to use endocytic receptors. However, recent data indicate additional endocytic uptake of Lp, mediated by a Lp receptor (LpR) of the LDLR family. The two N-terminal domains of LDLR family members are involved in ligand binding and dissociation, respectively, and are composed of a mosaic of multiple repeats. The three C-terminal domains, viz., the optional O-linked glycosylation domain, the transmembrane domain, and the intracellular domain, are of a non-repetitive sequence. The present classification of newly discovered LDLR family members, including the LpRs, bears no relevance to physiological function. Therefore, as a novel approach, the C-terminal domains of LDLR family members across the entire animal kingdom were used to perform a sequence comparison analysis in combination with a phylogenetic tree analysis. The LpRs appeared to segregate into a specific group distinct from the groups encompassing the other family members, and each of the three C-terminal domains of the insect receptors is composed of unique set of sequence motifs. Based on conservation of sequence motifs and organization of these motifs in the domains, LpR resembles most the groups of the LDLRs, very low density lipoprotein (VLDL) receptors, and vitellogenin receptors. However, in sequence aspects in which LpR deviates from these three receptor groups, it most notably resembles LDLR-related protein-2, or megalin. These features might explain the functional differences disclosed between insect and mammalian lipoprotein receptors.
- Published
- 2006
- Full Text
- View/download PDF
38. Intracellular fate of LDL receptor family members depends on the cooperation between their ligand-binding and EGF domains.
- Author
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Van Hoof D, Rodenburg KW, and Van der Horst DJ
- Subjects
- Animals, Asparagine chemistry, Blotting, Western, CHO Cells, Cell Membrane metabolism, Cricetinae, DNA, Complementary metabolism, Endocytosis, ErbB Receptors metabolism, Histidine chemistry, Hydrogen-Ion Concentration, Ligands, Lipoproteins chemistry, Locusta migratoria, Microscopy, Fluorescence, Models, Chemical, Models, Molecular, Mutation, Phenotype, Protein Structure, Tertiary, Receptors, LDL genetics, Receptors, LDL metabolism, Receptors, Lipoprotein chemistry, Time Factors, Transfection, Transferrin chemistry, Receptors, LDL chemistry
- Abstract
The insect low-density lipoprotein (LDL) receptor (LDLR) homologue LpR mediates endocytosis of an insect lipoprotein (lipophorin) that is structurally related to LDL. Despite these similarities, lipophorin and LDL follow distinct intracellular routes upon endocytosis by their receptors. Whereas LDL is degraded in lysosomes, lipophorin is recycled in a transferrin-like manner. We constructed several hybrid receptors composed of Locusta migratoria LpR and human LDLR regions to identify the domains implicated in LpR-mediated ligand recycling. Additionally, the triadic His562 residue of LDLR, which is putatively involved in ligand uncoupling, was mutated to Asn, corresponding to Asn643 in LpR, to analyse the role of the His triad in receptor functioning. The familial hypercholesterolaemia (FH) class 5 mutants LDLR(H562Y) and LDLR(H190Y) were also analysed in vitro. Fluorescence microscopic investigation and quantification suggest that LpR-mediated ligand recycling involves cooperation between the ligand-binding domain and epidermal growth factor (EGF) domain of LpR, whereas its cytosolic tail does not harbour motifs that affect this process. LDLR residue His562 appears to be essential for LDLR recycling after ligand endocytosis but not for constitutive receptor recycling. Like LDLR(H562N), LDLR(H562Y) did not recycle bound ligand; moreover, the intracellular distribution of both mutant receptors after ligand incubation coincides with that of a lysosomal marker. The LDLR mutant characterization in vitro suggests that LDLR FH class 5 mutations might be divided into two subclasses.
- Published
- 2005
- Full Text
- View/download PDF
39. Receptor-mediated endocytosis and intracellular trafficking of lipoproteins and transferrin in insect cells.
- Author
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Van Hoof D, Rodenburg KW, and Van der Horst DJ
- Subjects
- Animals, Cell Line, Drosophila metabolism, Fat Body metabolism, Gene Expression, Humans, Protein Transport, Receptors, LDL genetics, Receptors, Transferrin genetics, Recombinant Proteins metabolism, Spodoptera metabolism, Endocytosis physiology, Insecta metabolism, Lipoproteins metabolism, Receptors, LDL physiology, Receptors, Transferrin physiology, Transferrin metabolism
- Abstract
While the intracellular pathways of ligands after receptor-mediated endocytosis have been studied extensively in mammalian cells, in insect cells these pathways are largely unknown. We transfected Drosophila Schneider line 2 (S2) cells with the human low-density lipoprotein (LDL) receptor (LDLR) and transferrin (Tf) receptor (TfR), and used endocytosis of LDL and Tf as markers. After endocytosis in mammalian cells, LDL is degraded in lysosomes, whereas Tf is recycled. Fluorescence microscopy analysis revealed that LDL and Tf are internalized by S2 cells transfected with LDLR or TfR, respectively. In transfectants simultaneously expressing LDLR and TfR, both ligands colocalize in endosomes immediately after endocytic uptake, and their location remained unchanged after a chase. Similar results were obtained with Spodoptera frugiperda Sf9 cells that were transfected with TfR, suggesting that Tf is retained intracellularly by both cell lines. The insect lipoprotein, lipophorin, is recycled upon lipophorin receptor (LpR)-mediated endocytosis by mammalian cells, however, not after endocytosis by LpR-expressing S2 transfectants, suggesting that this recycling mechanism is cell-type specific. LpR is endogenously expressed by fat body tissue of Locusta migratoria for a limited period after an ecdysis. A chase following endocytosis of labeled lipophorin by isolated fat body tissue at this developmental stage resulted in a significant decrease of lipophorin-containing vesicles, indicative of recycling of the ligand.
- Published
- 2005
- Full Text
- View/download PDF
40. Cardiomyocytes derived from stem cells.
- Author
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Van Laake LW, Van Hoof D, and Mummery CL
- Subjects
- Animals, Cell Cycle, Cell Differentiation, Cell Transplantation methods, Heart Failure therapy, Humans, Mice, Myocardium pathology, Myocytes, Cardiac cytology, Myocytes, Cardiac transplantation, Stem Cells cytology
- Abstract
One way to restore failing heart function following myocardial infarction would be to replace lost or damaged cardiac cells by local or systemic injection. The sources of replacement cells presently discussed include embryonic stem cells, hematopoietic and non-hematopoietic stem cells from bone marrow or cord blood and small stem cell populations thought to reside in the heart itself or in skeletal muscle. Here we review this area of stem cell research with focus particularly on recent laboratory advances towards producing cardiomyocytes from embryonic stem cells. We conclude that embryonic stem cells and cardiac progenitors in the heart itself are the only proven sources of cardiomyocytes and that reported clinical effects of bone marrow stem currently undergoing validation are likely mediated by other mechanisms.
- Published
- 2005
- Full Text
- View/download PDF
41. Lipophorin receptor-mediated lipoprotein endocytosis in insect fat body cells.
- Author
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Van Hoof D, Rodenburg KW, and van der Horst DJ
- Subjects
- Animals, Carrier Proteins metabolism, Female, Fluorescent Antibody Technique, Humans, Larva metabolism, Ligands, Lipoproteins, HDL metabolism, Male, Molecular Chaperones metabolism, Endocytosis, Fat Body cytology, Fat Body metabolism, Grasshoppers metabolism, Lipoproteins metabolism, Receptors, Cytoplasmic and Nuclear metabolism
- Abstract
High-density lipophorin (HDLp) in the circulation of insects is able to selectively deliver lipids to target tissues in a nonendocytic manner. In Locusta migratoria, a member of the LDL receptor family has been identified and shown to mediate endocytosis of HDLp in mammalian cells transfected with the cDNA of this receptor. This insect lipophorin receptor (iLR) is temporally expressed in fat body tissue of young adult as well as larval locusts, as shown by Western blot analysis. Fluorescence microscopy revealed that fat body cells internalize fluorescently labeled HDLp and human receptor-associated protein only when iLR is expressed. Expression of iLR is down-regulated on Day 4 after an ecdysis. Consequently, HDLp is no longer internalized. By starving adult locusts immediately after ecdysis, we were able to prolong iLR expression. In addition, expression of the receptor was induced by starving adults after down-regulation of iLR. These results suggest that iLR mediates endocytosis of HDLp in fat body cells, and that expression of iLR is regulated by the demand of fat body tissue for lipids.
- Published
- 2003
- Full Text
- View/download PDF
42. Insect lipoprotein follows a transferrin-like recycling pathway that is mediated by the insect LDL receptor homologue.
- Author
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Van Hoof D, Rodenburg KW, and Van der Horst DJ
- Subjects
- Animals, CHO Cells, Cell Compartmentation physiology, Cricetinae, Endosomes metabolism, Fluorescent Antibody Technique, Humans, Ionophores pharmacology, LDL-Receptor Related Protein-Associated Protein metabolism, Monensin pharmacology, Organelles metabolism, Protein Transport physiology, Transferrin metabolism, Transport Vesicles metabolism, Carrier Proteins metabolism, Endocytosis physiology, Insecta metabolism, Lipoproteins metabolism, Lipoproteins, LDL metabolism, Mammals metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Receptors, LDL metabolism
- Abstract
The lipoprotein of insects, high-density lipophorin (HDLp), is homologous to that of mammalian low-density lipoprotein (LDL) with respect to its apolipoprotein structure. Moreover, an endocytic receptor for HDLp has been identified (insect lipophorin receptor, iLR) that is homologus to the LDL receptor. We transfected LDL-receptor-expressing CHO cells with iLR cDNA to study the endocytic uptake and intracellular pathways of LDL and HDLp simultaneously. Our studies provide evidence that these mammalian and insect lipoproteins follow distinct intracellular routes after receptor-mediated endocytosis. Multicolour imaging and immunofluorescence was used to visualize the intracellular trafficking of fluorescently labeled ligands in these cells. Upon internalization, which can be completely inhibited by human receptor-associated protein (RAP), mammalian and insect lipoproteins share endocytic vesicles. Subsequently, however, HDLp evacuates the LDL-containing endosomes. In contrast to LDL, which is completely degraded in lysosomes after dissociating from its receptor, both HDLp and iLR converge in a nonlysosomal juxtanuclear compartment. Colocalization studies with transferrin identified this organelle as the endocytic recycling compartment via which iron-depleted transferrin exits the cell. Fluorescently labeled RAP is also transported to this recycling organelle upon receptor-mediated endocytosis by iLR. Internalized HDLp eventually exits the cell via the recycling compartment, a process that can be blocked by monensin, and is re-secreted with a t(1/2) of approximately 13 minutes. From these observations, we conclude that HDLp is the first non-exchangeable apolipoprotein-containing lipoprotein that follows a transferrin-like recycling pathway despite the similarities between mammalian and insect lipoproteins and their receptors.
- Published
- 2002
- Full Text
- View/download PDF
43. Alternative lipid mobilization: the insect shuttle system.
- Author
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van der Horst DJ, van Hoof D, van Marrewijk WJ, and Rodenburg KW
- Subjects
- Animals, Apolipoproteins chemistry, Apolipoproteins metabolism, Carrier Proteins chemistry, Cell Line, Fatty Acid-Binding Protein 7, Fatty Acid-Binding Proteins, Flight, Animal physiology, Humans, Lipoproteins chemistry, Lipoproteins metabolism, Protein Conformation, Receptors, LDL chemistry, Receptors, LDL metabolism, Carrier Proteins metabolism, Endocytosis physiology, Insecta physiology, Lipid Metabolism, Neoplasm Proteins, Tumor Suppressor Proteins
- Abstract
Lipid mobilization in long-distance flying insects has revealed a novel concept for lipid transport in the circulatory system during exercise. Similar to energy generation for sustained locomotion in mammals, the work accomplished by non-stop flight activity is powered by oxidation of free fatty acids (FFA) derived from endogenous reserves of triacylglycerol. The transport form of the lipid, however, is diacylglycerol (DAG), which is delivered to the flight muscles associated with lipoproteins. In the insect system, the multifunctional lipoprotein, high-density lipophorin (HDLp) is loaded with DAG while additionally, multiple copies of the exchangeable apolipoprotein, apoLp-III, associate with the expanding particle. As a result, lipid-enriched low-density lipophorin (LDLp) is formed. At the flight muscles, LDLp-carried DAG is hydrolyzed and FFA are imported into the muscle cells for energy generation. The depletion of DAG from LDLp results in the recovery of both HDLp and apoLp-III, HDLp, identified which are reutilized for another cycle of DAG transport. A receptor for as a novel member of the vertebrate low-density lipoprotein (LDL) receptor family, does not seem to be involved in the lipophorin shuttle mechanism operative during flight activity. In addition, endocytosis of HDLp mediated by the insect receptor does not seem to follow the classical mammalian LDL pathway. Many structural elements of the lipid mobilization system in insects are similar to those in mammals. Domain structures of apoLp-I and apoLp-II, the non-exchangeable apolipoprotein components of HDLp, are related to apoB 100. ApoLp-III is a bundle of five amphipathic alpha-helices that binds to a lipid surface very similar to the four-helix bundle of the N-terminal domain of human apoE. Despite these similarities, the functioning of the insect lipoprotein in energy transport during flight activity is intriguingly different, since the TAG-rich mammalian lipoproteins play no role as a carrier of mobilized lipids during exercise and besides, these lipoproteins are not functioning as a reusable shuttle for lipid transport. On the other hand, the deviant behavior of similar molecules in a different biological system may provide a useful alternative model for studying the molecular basis of processes related to human disorders and disease.
- Published
- 2002
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