Publications by authors named "Annelieke S Wentzel"

7 Publications

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Transcriptome sequencing supports a conservation of macrophage polarization in fish.

Sci Rep 2020 08 10;10(1):13470. Epub 2020 Aug 10.

Aquaculture and Fisheries Group, Wageningen University and Research, De Elst 1, 6708 WD, Wageningen, The Netherlands.

Mammalian macrophages can adopt polarization states that, depending on the exact stimuli present in their extracellular environment, can lead to very different functions. Although these different polarization states have been shown primarily for macrophages of humans and mice, it is likely that polarized macrophages with corresponding phenotypes exist across mammals. Evidence of functional conservation in macrophages from teleost fish suggests that the same, or at least comparable polarization states should also be present in teleosts. However, corresponding transcriptional profiles of marker genes have not been reported thus far. In this study we confirm that macrophages from common carp can polarize into M1- and M2 phenotypes with conserved functions and corresponding transcriptional profiles compared to mammalian macrophages. Carp M1 macrophages show increased production of nitric oxide and a transcriptional profile with increased pro-inflammatory cytokines and mediators, including il6, il12 and saa. Carp M2 macrophages show increased arginase activity and a transcriptional profile with increased anti-inflammatory mediators, including cyr61, timp2b and tgm2b. Our RNA sequencing approach allowed us to list, in an unbiased manner, markers discriminating between M1 and M2 macrophages of teleost fish. We discuss the importance of our findings for the evaluation of immunostimulants for aquaculture and for the identification of gene targets to generate transgenic zebrafish for detailed studies on M1 and M2 macrophages. Above all, we discuss the striking degree of evolutionary conservation of macrophage polarization in a lower vertebrate.
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http://dx.doi.org/10.1038/s41598-020-70248-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418020PMC
August 2020

Fish Macrophages Show Distinct Metabolic Signatures Upon Polarization.

Front Immunol 2020 25;11:152. Epub 2020 Feb 25.

Aquaculture and Fisheries Group, Wageningen University & Research, Wageningen, Netherlands.

Macrophages play important roles in conditions ranging from host immune defense to tissue regeneration and polarize their functional phenotype accordingly. Next to differences in the use of L-arginine and the production of different cytokines, inflammatory M1 macrophages and anti-inflammatory M2 macrophages are also metabolically distinct. In mammals, M1 macrophages show metabolic reprogramming toward glycolysis, while M2 macrophages rely on oxidative phosphorylation to generate energy. The presence of polarized functional immune phenotypes conserved from mammals to fish led us to hypothesize that a similar metabolic reprogramming in polarized macrophages exists in carp. We studied mitochondrial function of M1 and M2 carp macrophages under basal and stressed conditions to determine oxidative capacity by real-time measurements of oxygen consumption and glycolytic capacity by measuring lactate-based acidification. In M1 macrophages, we found increased nitric oxide production and expression in addition to altered oxidative phosphorylation and glycolysis. In M2 macrophages, we found increased arginase activity, and both oxidative phosphorylation and glycolysis were similar to control macrophages. These results indicate that M1 and M2 carp macrophages show distinct metabolic signatures and indicate that metabolic reprogramming may occur in carp M1 macrophages. This immunometabolic reprogramming likely supports the inflammatory phenotype of polarized macrophages in teleost fish such as carp, similar to what has been shown in mammals.
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http://dx.doi.org/10.3389/fimmu.2020.00152DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7052297PMC
March 2021

The kinetics of cellular and humoral immune responses of common carp to presporogonic development of the myxozoan Sphaerospora molnari.

Parasit Vectors 2019 May 6;12(1):208. Epub 2019 May 6.

Institute of Parasitology, Biology, Centre of the Czech Academy of Sciences, České Budějovice, Czech Republic.

Background: Sphaerospora molnari is a myxozoan parasite causing skin and gill sphaerosporosis in common carp (Cyprinus carpio) in central Europe. For most myxozoans, little is known about the early development and the expansion of the infection in the fish host, prior to spore formation. A major reason for this lack of information is the absence of laboratory model organisms, whose life-cycle stages are available throughout the year.

Results: We have established a laboratory infection model for early proliferative stages of myxozoans, based on separation and intraperitoneal injection of motile and dividing S. molnari stages isolated from the blood of carp. In the present study we characterize the kinetics of the presporogonic development of S. molnari, while analyzing cellular host responses, cytokine and systemic immunoglobulin expression, over a 63-day period. Our study shows activation of innate immune responses followed by B cell-mediated immune responses. We observed rapid parasite efflux from the peritoneal cavity (< 40 hours), an initial covert infection period with a moderate proinflammatory response for about 1-2 weeks, followed by a period of parasite multiplication in the blood which peaked at 28 days post-infection (dpi) and was associated with a massive lymphocyte response. Our data further revealed a switch to a massive anti-inflammatory response (up to 1456-fold expression of il-10), a strong increase in the expression of IgM transcripts and increased number of IgM B lymphocytes, which produce specific antibodies for the elimination of most of the parasites from the fish at 35 dpi. However, despite the presence of these antibodies, S. molnari invades the liver 42 dpi, where an increase in parasite cell number and indistinguishable outer cell membranes are indicative of effective exploitation and disguise mechanisms. From 49 dpi onwards, the acute infection changes to a chronic one, with low parasite numbers remaining in the fish.

Conclusions: To our knowledge, this is the first time myxozoan early development and immune modulation mechanisms have been analyzed along with innate and adaptive immune responses of its fish host, in a controlled laboratory system. Our study adds important information on host-parasite interaction and co-evolutionary adaptation of early metazoans (Cnidaria) with basic vertebrate (fish) immune systems and the evolution of host adaptation and parasite immune evasion strategies.
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http://dx.doi.org/10.1186/s13071-019-3462-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6501462PMC
May 2019

Paralogs of Common Carp Granulocyte Colony-Stimulating Factor (G-CSF) Have Different Functions Regarding Development, Trafficking and Activation of Neutrophils.

Front Immunol 2019 19;10:255. Epub 2019 Feb 19.

Laboratory of Comparative Immunology, Department of Veterinary Medicine, Nihon University, Fujisawa, Japan.

Mammalian granulocyte colony-stimulating factor (G-CSF; CSF3) is a primary cytokine that promotes the development, mobilization, and activation of neutrophils and their precursors. Teleosts have been reported to possess two paralogs as a likely result of the teleost-wide whole genome duplication (WGD) event, but functional divergence of G-CSF paralogs remains poorly understood. Common carp are an allotetraploid species owing to an additional WGD event in the carp lineage and here, we report on genomic synteny, sequence similarity, and phylogeny of four common carp G-CSF paralogs ( and ; and ). and show differential and relatively high gene expression levels, while and show low basal gene expression levels in most tissues. All paralogs are expressed higher in macrophages than in other leukocyte sub-types and are highly up-regulated by treatment of macrophages with mitogens. Recombinant G-CSFa1 and G-CSFb1 both promoted the proliferation of kidney hematopoietic cells, while only G-CSFb1 induced the differentiation of kidney cells along the neutrophil-lineage. Colony-forming unit assays revealed that G-CSFb1 alone stimulates the formation of CFU-G colonies from head- and trunk-kidney whereas the combination of G-CSFa1 and G-CSFb1 stimulates the formation of both CFU-G and CFU-GM colonies. Recombinant G-CSFa1 and G-CSFb1 also exhibit chemotactic activity against kidney neutrophils and up-regulation of mRNA expression was highest in neutrophils after G-CSFb1 stimulation. Furthermore, G-CSFb1 more than G-CSFa1 induced priming of kidney neutrophils through up-regulation of a NADPH-oxidase component p47 . administration of G-CSF paralogs increased the number of circulating blood neutrophils of carp. Our findings demonstrate that gene duplications in teleosts can lead to functional divergence between paralogs and shed light on the sub-functionalization of G-CSF paralogs in cyprinid fish.
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http://dx.doi.org/10.3389/fimmu.2019.00255DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6389648PMC
October 2020

Carp Il10a and Il10b exert identical biological activities in vitro, but are differentially regulated in vivo.

Dev Comp Immunol 2017 02 29;67:350-360. Epub 2016 Aug 29.

Cell Biology and Immunology Group, Department of Animal Sciences, Wageningen University, 6708 WD, Wageningen, The Netherlands. Electronic address:

We recently reported on the functional characterization of carp Il10. We showed that carp Il10 is able to downregulate proinflammatory activities by carp phagocytes and promote B cell proliferation, differentiation and antibody production as well as proliferation of memory T cells. Taking advantage of the recent annotation of the carp genome, we completed the sequence of a second il10 paralogue, named il10b, the presence of which was expected owing to the recent (8 million years ago) fourth round of whole genome duplication that occurred in common carp. In the present study we closely compared the two Il10 paralogues and show that Il10a and Il10b have almost identical gene structure, synteny, protein sequence as well as bioactivity on phagocytes. Although the two il10 paralogues show a large overlap in tissue expression, il10b has a low constitutive expression and is highly upregulated upon infection, whereas il10a is higher expressed under basal conditions but its gene expression remains constant during viral and parasitic infections. This differential regulation is most likely due to the observed differences in their promoter regions. Altogether our results demonstrate that gene duplication in carp, although recent, led to sub-functionalization and expression divergence rather than functional redundancy of the Il10 paralogues, yet with very similar protein sequences.
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http://dx.doi.org/10.1016/j.dci.2016.08.016DOI Listing
February 2017

Polarization of immune responses in fish: The 'macrophages first' point of view.

Mol Immunol 2016 Jan 21;69:146-56. Epub 2015 Oct 21.

Cell Biology and Immunology Group, Wageningen Institute of Animal Sciences, Wageningen University, P.O. Box 338, 6700 AH Wageningen, The Netherlands.

In this review, we support taking polarized immune responses in teleost fish from a 'macrophage first' point of view, a hypothesis that reverts the dichotomous T helper (TH)1 and TH2 driving forces by building on the idea of conservation of innate immune responses in lower vertebrates. It is plausible that the initial trigger for macrophage polarization into M1 (inflammation) or M2 (healing) could rely only on sensing microbial/parasite infection or other innate danger signals, without the influence of adaptive immunity. Given the long and ongoing debate on the presence/absence of a typical TH1 cytokine environment and, in particular, TH2 cytokine environment in fish immune responses, it stands out that the presence of macrophages with polarized phenotypes, alike M1 and M2, have been relatively easy to demonstrate for fish. We summarize in short present knowledge in teleost fish on those cytokines considered most critical to the dichotomous development of TH1/M1 and TH2/M2 polarization, in particular, but not exclusively, interferon-γ and interleukin (IL)-4/IL-13. We review, in more detail, polarization of fish immune responses taken from the macrophage point of view for which we adopted the simple nomenclature of M1 and M2. We discuss inducible nitric oxide synthase, or NOS-2, as a reliable M1 marker and arginase-2 as a reliable M2 marker for teleost fish and discuss the value of these macrophage markers for the generation of zebrafish reporter lines to study M1/M2 polarization in vivo.
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http://dx.doi.org/10.1016/j.molimm.2015.09.026DOI Listing
January 2016

Cyprinid Herpesvirus 3 Il10 Inhibits Inflammatory Activities of Carp Macrophages and Promotes Proliferation of Igm+ B Cells and Memory T Cells in a Manner Similar to Carp Il10.

J Immunol 2015 Oct 14;195(8):3694-704. Epub 2015 Sep 14.

Cell Biology and Immunology Group, Department of Animal Sciences, Wageningen University, 6708WD Wageningen, the Netherlands; and

Cyprinid herpesvirus 3 (CyHV-3) is the causative agent of a lethal disease of carp and encodes for an Il10 homolog (ORF134). Our previous studies with a recombinant ORF134-deleted strain and the derived revertant strain suggested that cyprinid herpesvirus 3 Il10 (CyHV-3 Il10 [cyhv3Il10]) is not essential for viral replication in vitro, or virulence in vivo. In apparent contrast, cyhv3Il10 is one of the most abundant proteins of the CyHV-3 secretome and is structurally very similar to carp Il10 and also human IL10. To date, studies addressing the biological activity of cyhv3Il10 on cells of its natural host have not been performed. To address the apparent contradiction between the presence of a structurally conserved Il10 homolog in the genome of CyHV-3 and the lack of a clear phenotype in vivo using recombinant cyhv3Il10-deleted viruses, we used an in vitro approach to investigate in detail whether cyhv3Il10 exerts any biological activity on carp cells. In this study, we provide direct evidence that cyhv3Il10 is biologically active and, similarly to carp Il10, signals via a conserved Stat3 pathway modulating immune cells of its natural host, carp. In vitro, cyhv3Il10 deactivates phagocytes with a prominent effect on macrophages, while also promoting proliferation of Igm(+) B cells and memory T cells. Collectively, this study demonstrates a clear biological activity of cyhv3Il10 on cells of its natural host and indicates that cyhv3Il10 is a true viral ortholog of carp Il10. Furthermore, to our knowledge, this is the first report on biological activities of a nonmammalian viral Il10 homolog.
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http://dx.doi.org/10.4049/jimmunol.1500926DOI Listing
October 2015