Transcriptome sequencing supports a conservation of macrophage polarization in fish
Wentzel, Annelieke S. ; Petit, Jules ; Veen, Wouter G. van; Rosenbek Fink, Inge ; Scheer, Marleen H. ; Piazzon, Carla M. ; Forlenza, Maria ; Spaink, Herman P. ; Wiegertjes, Geert F. - \ 2020
Scientific Reports 10 (2020). - ISSN 2045-2322
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.
Beyond heat stress: Intestinal integrity disruption and mechanism-based intervention strategies
Lian, Puqiao ; Braber, Saskia ; Garssen, Johan ; Wichers, Harry J. ; Folkerts, Gert ; Fink-Gremmels, Johanna ; Varasteh, Soheil - \ 2020
Nutrients 12 (2020)3. - ISSN 2072-6643
Heat stress (HS) - Intestinal integrity - Nutritional supplements - Reactive oxygen species (ROS) - Resilience pathways
The current climate changes have increased the prevalence and intensity of heat stress (HS) conditions. One of the initial consequences of HS is the impairment of the intestinal epithelial barrier integrity due to hyperthermia and hypoxia following blood repartition, which often results in a leaky gut followed by penetration and transfer of luminal antigens, endotoxins, and pathogenic bacteria. Under extreme conditions, HS may culminate in the onset of “heat stroke”, a potential lethal condition if remaining untreated. HS-induced alterations of the gastrointestinal epithelium, which is associated with a leaky gut, are due to cellular oxidative stress, disruption of intestinal integrity, and increased production of pro-inflammatory cytokines. This review summarizes the possible resilience mechanisms based on in vitro and in vivo data and the potential interventions with a group of nutritional supplements, which may increase the resilience to HS-induced intestinal integrity disruption and maintain intestinal homeostasis.
Fungal artillery of zombie flies: infectious spore dispersal using a soft water cannon
Ruiter, Jolet de; Arnbjerg-Nielsen, Sif Fink ; Herren, Pascal ; Høier, Freja ; Fine Licht, Henrik H. De; Jensen, Kaare H. - \ 2019
Journal of the Royal Society, Interface 16 (2019)159. - ISSN 1742-5689 - 10 p.
biomimetic soft cannon - dispersal range - Entomophthora muscae - force-balance model - fungal spore ejection - high-speed videography
Dead sporulating female fly cadavers infected by the house fly-pathogenic fungus Entomophthora muscae are attractive to healthy male flies, which by their physical inspection may mechanically trigger spore release and by their movement create whirlwind airflows that covers them in infectious conidia. The fungal artillery of E. muscae protrudes outward from the fly cadaver, and consists of a plethora of micrometric stalks that each uses a liquid-based turgor pressure build-up to eject a jet of protoplasm and the initially attached spore. The biophysical processes that regulate the release and range of spores, however, are unknown. To study the physics of ejection, we design a biomimetic 'soft cannon' that consists of a millimetric elastomeric barrel filled with fluid and plugged with a projectile. We precisely control the maximum pressure leading up to the ejection, and study the cannon efficiency as a function of its geometry and wall elasticity. In particular, we predict that ejection velocity decreases with spore size. The calculated flight trajectories under aerodynamic drag predict that the minimum spore size required to traverse a quiescent layer of a few millimetres around the fly cadaver is approximately 10 µm. This corroborates with the natural size of E. muscae conidia (approx. 27 µm) being large enough to traverse the boundary layer but small enough (less than 40 µm) to be lifted by air currents. Based on this understanding, we show how the fungal spores are able to reach a new host.
Modeling phosphorus in rivers at the global scale : recent successes, remaining challenges, and near-term opportunities
Harrison, John A. ; Beusen, Arthur H.W. ; Fink, Gabriel ; Tang, Ting ; Strokal, Maryna ; Bouwman, Alexander F. ; Metson, Geneviève S. ; Vilmin, Lauriane - \ 2019
Current Opinion in Environmental Sustainability 36 (2019). - ISSN 1877-3435 - p. 68 - 77.
Understanding and mitigating the effects of phosphorus (P) overenrichment of waters globally, including the evaluation of the global Sustainability Development Goals, requires the use of global models. Such models quantitatively link land use, global population growth and climate to aquatic nutrient loading and biogeochemical cycling. Here we describe, compare, and contrast the existing global models capable of predicting P transport by rivers at a global scale. We highlight important insights gained from the development and application of these models, and identify important near-term opportunities for model improvements as well as additional insight to be gained through new model analysis.
Strategies for optimal fertiliser management of vegetable crops in Europe
Thompson, R.B. ; Voogt, W. ; Incrocci, L. ; Fink, M. ; Neve, S. de - \ 2018
In: 5th International Symposium on Ecologically Sound Fertilization Strategies for Field Vegetable Production. - International Society for Horticultural Science (Acta Horticulturae ) - ISBN 9789462611887 - p. 129 - 140.
Crop monitoring - Decision support systems - Simulation models - Soil analysis
In Europe a number of procedures are used to assist growers and advisors to determine optimal N fertiliser recommendations. The implementation of European Union (EU) legislation is encouraging the adoption of fertiliser recommendation schemes. The most widely used schemes are those based on soil testing or on the use of indices that estimate the soil nitrogen supply. Soil testing approaches that are in use, particularly in NW Europe are the Nmin, KNS and N-Expert systems; the latter is operated as a computer-based decision support system (DSS). The comprehensive RB209 Fertiliser Manual of England and Wales uses soil N supply indices, but soil analysis can also be used. Nitrogen balance calculations are widely used throughout Europe and form part of the KNS and N-Expert systems, and a number of other DSSs. The N balance considers the various soil N sources and treats mineral N fertiliser as a supplemental N source. The EU-Rotate_N simulation model is a comprehensive and versatile tool, developed for diverse European conditions, that is useful for scenario analysis simulations to stakeholders. Various DSS have been developed in different European countries, with different levels of complexity. There are a number of different DSS that calculate N fertiliser recommendations for particular cropping systems; some DSS calculate the requirements for other nutrients, and some also do so for irrigation which is particularly useful where fertigation is used. Sap analysis has been shown to be sensitive to crop nutrient status, for N and some other nutrients; currently, there is renewed interest in sap analysis. Proximal optical sensors are a promising approach for N management.
An overview of the diurnal cycle of the atmospheric boundary layer during the West African monsoon season : Results from the 2016 observational campaign
Kalthoff, Norbert ; Lohou, Fabienne ; Brooks, Barbara ; Jegede, Gbenga ; Adler, Bianca ; Babić, Karmen ; DIone, Cheikh ; Ajao, Adewale ; Amekudzi, Leonard K. ; Aryee, Jeffrey N.A. ; Ayoola, Muritala ; Bessardon, Geoffrey ; Danuor, Sylvester K. ; Handwerker, Jan ; Kohler, Martin ; Lothon, Marie ; Pedruzo-Bagazgoitia, Xabier ; Smith, Victoria ; Sunmonu, Lukman ; Wieser, Andreas ; Fink, Andreas H. ; Knippertz, Peter - \ 2018
Atmospheric Chemistry and Physics 18 (2018)4. - ISSN 1680-7316 - p. 2913 - 2928.
A ground-based field campaign was conducted in southern West Africa from mid-June to the end of July 2016 within the framework of the Dynamics-Aerosol-Chemistry-Cloud Interactions in West Africa (DACCIWA) project. It aimed to provide a high-quality comprehensive data set for process studies, in particular of interactions between low-level clouds (LLCs) and boundary-layer conditions. In this region missing observations are still a major issue. During the campaign, extensive remote sensing and in situ measurements were conducted at three supersites: Kumasi (Ghana), Savè (Benin) and Ile-Ife (Nigeria). Daily radiosoundings were performed at 06:00 UTC, and 15 intensive observation periods (IOPs) were performed during which additional radiosondes were launched, and remotely piloted aerial systems were operated. Extended stratiform LLCs form frequently in southern West Africa during the nighttime and persist long into the following day. They affect the radiation budget and hence the evolution of the atmospheric boundary layer and regional climate. The relevant parameters and processes governing the formation and dissolution of the LLCs are still not fully understood. This paper gives an overview of the diurnal cycles of the energy-balance components, near-surface temperature, humidity, wind speed and direction as well as of the conditions (LLCs, low-level jet) in the boundary layer at the supersites and relates them to synoptic-scale conditions (monsoon layer, harmattan layer, African easterly jet, tropospheric stratification) in the DACCIWA operational area. The characteristics of LLCs vary considerably from day to day, including a few almost cloud-free nights. During cloudy nights we found large differences in the LLCs' formation and dissolution times as well as in the cloud-base height. The differences exist at individual sites and also between the sites. The synoptic conditions are characterized by a monsoon layer with south-westerly winds, on average about 1.9 km deep, and easterly winds above; the depth and strength of the monsoon flow show great day-to-day variability. Within the monsoon layer, a nocturnal low-level jet forms in approximately the same layer as the LLC. Its strength and duration is highly variable from night to night. This unique data set will allow us to test some new hypotheses about the processes involved in the development of LLCs and their interaction with the boundary layer and can also be used for model evaluation.
Hydrological and Agricultural Impacts of Climate Change in the Vu Gia-Thu Bon River Basin in Central Vietnam
Laux, Patrick ; Fink, Manfred ; Waongo, Moussa ; Pedroso, Rui ; Salvini, G. ; Hoa Tran, Dang ; Quang Thinh, Dang ; Cullmann, Johannes ; Flügel, Wolfgang-Alvert ; Kunstmann, H. - \ 2017
In: Land Use and Climate Change Interactions in Central Vietnam / Nauditt, A., Ribbe, L., Singapore : Springer Science (Water Resources Development and Management ) - ISBN 9789811026232 - p. 123 - 142.
This paper summarizes some of the climate (change) impact modeling activities conducted in the Land useandClimate Changeinteractionsin Central Vietnam (LUCCi)project. The study area is the Vu Gia-Thu Bon (VGTB) river basin in Central Vietnam, which is characterized by recurrent floods during the rainy season, but also water shortages during the dry season. The impact modeling activities, such as the validation of the models are hindered by the scarcity of hydrometeorological data and an unfavorable distribution of the observation network, i.e., station data is available only for the lowlands. In total, two different process-based and distributed hydrological models are applied in concert with climate change and land use projections. Based on that, the magnitudes and return periods of extreme flows are estimated. The modeling results suggest increases of extreme high flows due to climate change. A multi-objective agro-economical model was developed for a typical irrigation scheme in the region in order to optimize the area for cropping, irrigation-techniques and schedules. The model results suggest the irrigation technique Alternate Wetting and Drying, which has the potential to increase the benefits for the farmers and help to mitigate greenhouse gases at the same time. In addition, the regional-scale crop model GLAM is applied for groundnut under rainfed conditions, which is capable to identify regions suitable for cropping in the future. The paper further synthesizes recommendations for local stakeholders in Central Vietnam.
Innate immunity of carp : fishing for receptors
Fink, Inge - \ 2017
Wageningen University. Promotor(en): Geert Wiegertjes; Huub Savelkoul, co-promotor(en): Maria Forlenza. - Wageningen : Wageningen University - ISBN 9789463430753 - 240
carp - cyprinus - immunity - platelets - macrophage activation - receptors - polarization - immunostimulation - immunology - karper - cyprinus - immuniteit - bloedplaatjes - macrofaag activering - receptoren - polarisatie - immunostimulatie - immunologie
Recent decades have seen a significant intensification of aquaculture leading to increased risk of infections with several pathogenic organisms. On economical and ethical grounds it is more appropriate to improve general welfare conditions and prevent infections rather than treating disease outbreaks once they have occurred. Immunostimulation through feed can provide more efficient and sustainable control of diseases in aquaculture through enhancing the immunocompetence of fish; however, the underlying mechanisms are poorly characterized. The overall aim of this thesis was to perform a molecular and functional characterization of how pathogen-associated molecular patterns (PAMPs), such as β-glucans, affect the innate immune response of carp and which receptors on carp leukocytes are likely candidates to play a role in sensing such PAMPs.
In chapter 1 we provide a framework for this thesis by introducing different classes of PAMPs, including β-glucans. These molecules were the centrepiece of an intra-European training network called NEMO (Protective immune modulation in warm water fish by feeding glucans), which this PhD project was part of. The scientific aim of the NEMO network was to develop a sustainable and cost-effective use of β-glucans as immunostimulants for aquaculture, using common carp as the model fish species, since on a global scale common carp is the most cultured fish species for food consumption. Our aims within the NEMO project entailed both the characterization of carp leukocytes and the characterization of candidate pattern recognition receptors (PRRs) that could play a role in sensing PAMPs and initiating immune responses. Chapter 1 therefore introduces the thrombocytes and macrophages pertinent to this thesis, as well as important classes of PRRs.
In our first experimental study, described in chapter 2, we investigated the relevance of thrombocytes for the immune system of carp. We found that thrombocytes from healthy carp express a large number of immune-relevant genes, among which several cytokines and Toll-like receptors (Tlrs). Furthermore, we dissected the role of thrombocytes during infections with two different, albeit related, protozoan parasites, Trypanoplasma borreli and Trypanosoma carassii, and found thrombocytes were massively depleted from blood and spleen of fish infected with T. borreli. The pathology of this infection is associated with elevated levels of tissue nitration, prompting us to investigate, ex vivo, the effect of nitric oxide on thrombocytes. Our studies revealed that nitric oxide can induce a clear and rapid apoptosis of thrombocytes from healthy carp, supporting a role for nitric oxide-mediated control of immune-relevant thrombocytes during infection with T. borreli. Thereby, this particular study provided an excellent example of interplay between pathogen and the innate immune system of carp.
We reviewed in chapter 3 another cell type central to innate immunity: the macrophage. We focused on the heterogeneity of macrophage activation states as these cells, at least in humans and mice, have the ability to polarize in several directions during an immune response. Based on the signals that lead to activation and the effector functions and cytokine profile as a result thereof, macrophages can be broadly divided into two types: classically activated macrophages induced in a T helper 1 (TH1) cytokine environment, and alternatively activated macrophages, induced in a TH2 cytokine environment. Mirroring the TH1–TH2 dichotomy, classically activated macrophages have also been termed M1, whereas alternatively activated macrophages have been termed M2. Classically activated macrophages are typically induced by stimulation with microbial ligands such as LPS in combination with pro-inflammatory cytokines such as IFNγ, and can be viewed as an extension of innate activated macrophages which are induced by microbial ligands only, thus are independent of cytokines. Alternatively activated macrophages are generated in the presence of IL4 and/or IL13. In addition to M1 and M2, one can distinguish regulatory macrophages, which are associated with the presence of the cytokine IL10. In this chapter, we reviewed the evidence of existence of polarized macrophages in teleost fish, among other things based on observations of the fundamentally different immune responses elicited by the parasites T. borreli and T. carassii.
We further investigated the polarization of carp macrophages in chapter 4, where we obtained gene signature profiles of carp macrophages via a transcriptome approach. Independently of cytokines, carp macrophages showed the ability to differentiate into cells with functional characteristics highly comparable to those of mammalian M1 and M2, consistent with a conserved ability of macrophages to polarize into distinct subsets. In addition to obtaining a global view of gene expression, our transcriptome approach identified gene signatures for M1 and M2 macrophages which appear conserved from fish to mammals. We selected a number of these interesting genes that were differentially regulated between M1 and M2 macrophages and discussed in detail five potential M1 markers; il1β, ptx3a, saa, nos2b, and il12a – as well as five potential M2 markers; cyr61, inhba, timp2, tgm2, and arg2. These transcriptome studies may pave the way for future studies of polarized macrophages during immune responses in fish. Furthermore, additional analyses of the datasets described in this chapter will undoubtedly lead to the characterization of more genes relevant to macrophage polarization and recognition of immunostimulants.
As part of the characterization of candidate PRRs that could play a role in sensing PAMPs and initiating immune responses, we studied the scavenger receptor Cd36 (chapter 5), which in mammals is expressed by many different (immune) cell types and plays a role in highly diverse processes, both homeostatic and pathologic. Among other things, it is often found associated with sensing of β-glucans and also with M2 macrophage activation, sparking our interest in this molecule in fish. We studied Cd36 in common carp as well as in zebrafish, a closely related cyprinid fish species. Whereas a single cd36 gene is present in zebrafish, carp was shown to have two paralogs of cd36. Although all genes show conserved synteny compared to mammalian CD36, unexpectedly we could not detect gene expression of cyprinid cd36 in macrophages or any other immune cell type or immune organ. Yet, because gene expression of cd36 was down-regulated during Mycobacterium marinum infection of zebrafish, and knockdown of cd36 in zebrafish embryos led to higher bacterial burden upon such infection, our data imply a role for Cd36 in immune responses of fish. Future studies are needed to clarify the exact mechanisms involved.
As characterization of candidate PRRs we also examined the Toll-like receptors Tlr1 and Tlr2 (chapter 6). We identified a full-length, expressed tlr1 gene, a tlr1 pseudogene, and a second tlr2 gene next to the tlr2 which had been described previously. Sequence, phylogenetic and synteny analyses supported the conserved nature of these genes, and three-dimensional modelling showed a good fit with the mammalian TLR1/TLR2 heterodimer including the potential to bind to the prototypical ligand Pam3CSK4. However, we were unable to demonstrate Tlr1/Tlr2-mediated ligand binding in transfected cell lines through NFκB activation, despite showing the expression and co-localization of Tlr1 and Tlr2. This prompted a discussion of methods available for studying ligand-binding properties of fish Tlrs.
Finally, we discuss in chapter 7 the findings of this thesis in the context of the NEMO project. We present the concept of trained immunity, which could provide the conceptual framework within which the immune-stimulating ability of compounds such as β-glucans could be explained. We discuss recent advances in the field of TLR research as well as that of macrophage polarization, and highlight immunometabolism as a new area of interest which may help to illuminate the molecular events occurring in immune cells during health and disease. In conclusion, we found that carp leukocytes, along with their pattern recognition receptors, are central players of the innate immune system of carp. Our findings contribute to the understanding of mechanisms of immunostimulation, and expect this will enable the valorisation and use of immunostimulants for sustainable aquaculture and improvement of fish health.
Characterizing microbiota-independent effects of oligosaccharides on intestinal epithelial cells : insight into the role of structure and size: Structure–activity relationships of non-digestible oligosaccharides
Akbari, Peyman ; Fink-Gremmels, Johanna ; Willems, Rianne H.A.M. ; Difilippo, Elisabetta ; Schols, Henk A. ; Schoterman, Margriet H.C. ; Garssen, Johan ; Braber, Saskia - \ 2017
European Journal of Nutrition 56 (2017)5. - ISSN 1436-6207 - p. 1919 - 1930.
Caco-2 cells - CXCL8 - Degree of polymerization - Intestinal permeability - Non-digestible oligosaccharides - Tight junctions
Purpose: The direct effects of galacto-oligosaccharides (GOS), including Vivinal® GOS syrup (VGOS) and purified Vivinal® GOS (PGOS), on the epithelial integrity and corresponding interleukin-8 (IL-8/CXCL8) release were examined in a Caco-2 cell model for intestinal barrier dysfunction. To investigate structure–activity relationships, the effects of individual DP fractions of VGOS were evaluated. Moreover, the obtained results with GOS were compared with Caco-2 monolayers incubated with fructo-oligosaccharides (FOS) and inulin. Methods: Caco-2 monolayers were pretreated (24 h) with or without specific oligosaccharides or DP fractions of VGOS (DP2 to DP6) before being exposed for 12 or 24 h to the fungal toxin deoxynivalenol (DON). Transepithelial electrical resistance and lucifer yellow permeability were measured to investigate barrier integrity. A calcium switch assay was used to study the reassembly of tight junction proteins. Release of CXCL8, a typical marker for inflammation, was quantified by ELISA. Results: In comparison with PGOS, FOS and inulin, VGOS showed the most pronounced protective effect on the DON-induced impairment of the monolayer integrity, acceleration of the tight junction reassembly and the subsequent CXCL8 release. DP2 and DP3 in concentrations occurring in VGOS prevented the DON-induced epithelial barrier disruption, which could be related to their high prevalence in VGOS. However, no effects of the separate DP GOS fractions were observed on CXCL8 release. Conclusions: This comparative study demonstrates the direct, microbiota-independent effects of oligosaccharides on the intestinal barrier function and shows the differences between individual galacto- and fructo-oligosaccharides. This microbiota-independent effect of oligosaccharides depends on the oligosaccharide structure, DP length and concentration.
Facing the challenge of a functional characterization of toll-like receptor (TLR)1 and TLR2 in common carp
Fink, I.R. ; Forlenza, M. ; Pietretti, D. ; Wiegertjes, G. - \ 2016
- p. 76 - 76.
Carp, TLR1, TLR2, sub-cellular localizatioan, ligand binding
Fish & Shellfish Immunology Special Issue ISFSI 2016
|Conservation of macrophage polarization in fish
Wentzel, A.S. ; Petit, J. ; Fink, I.R. ; Forlenza, M. ; Wiegertjes, G. - \ 2016
- p. 54 - 54.
Macrophages of higher vertebrates can express a range of activation states, with the extremes termed M1 and M2. Neither the evolutionary conservation of these activation states, nor the exact microbial and/or cytokine stimulants involved, have been examined in detail in lower vertebrates. We have shown that macrophages of teleost fish, including carp, have the ability to polarize into activation states typical of classical (M1) and alternative (M2) extremes upon stimulation with LPS, or exogenous cAMP, respectively. Upon this polarization, carp macrophages display functional profiles and several molecular markers that indicate the M1-M2 dichotomy could be an intrinsic property of macrophages which arose early in evolution. Owing to the more recent discoveries of IFN-y and lL-4/IL-13-like cytokines in teleost fish, we now can also study cytokine-dependent polarization of carp macrophages. Interferon-y amplifies LPS-induced polarisation into M1-like profiles of carp macrophages, including the induction of nitric oxide. Strikingly, IL-4/IL-13 appears to increase levels of arginase along with an activation profile that at least partly overlaps with that of cAMP-induced M2 macrophages. Thus, the chief macrophage M1 and M2 activation states appear to operate under the guidance of primordially conserved principles.
Molecular and functional characterization of Toll-like receptor (Tlr)1 and Tlr2 in common carp (Cyprinus carpio)
Fink, Inge R. ; Pietretti, Danilo ; Voogdt, Carlos G.P. ; Westphal, Adrie H. ; Savelkoul, Huub F.J. ; Forlenza, Maria ; Wiegertjes, Geert F. - \ 2016
Fish and Shellfish Immunology 56 (2016). - ISSN 1050-4648 - p. 70 - 83.
Heterodimer - Innate immunity - Teleost - TLR signalling - Toll-like receptor
Toll-like receptors (TLRs) are fundamental components of innate immunity that play significant roles in the defence against pathogen invasion. In this study, we present the molecular characterization of the full-length coding sequence of tlr1, tlr2a and tlr2b from common carp (Cyprinus carpio). Each is encoded within a single exon and contains a conserved number of leucine-rich repeats, a transmembrane region and an intracellular TIR domain for signalling. Indeed, sequence, phylogenetic and synteny analysis of carp tlr1, tlr2a and tlr2b support that these genes are orthologues of mammalian TLR1 and TLR2. The tlr genes are expressed in various immune organs and cell types. Furthermore, the carp sequences exhibited a good three-dimensional fit with the heterodimer structure of human TLR1-TLR2, including the potential to bind to the ligand Pam3CSK4. This supports the possible formation of carp Tlr1-Tlr2 heterodimers. However, we were unable to demonstrate Tlr1/Tlr2-mediated ligand binding in transfected cell lines through NF-κB activation, despite showing the expression and co-localization of Tlr1 and Tlr2. We discuss possible limitations when studying ligand-specific activation of NF-κB after expression of Tlr1 and/or Tlr2 in human but also fish cell lines and we propose alternative future strategies for studying ligand-binding properties of fish Tlrs.
Milk Oligosaccharide Variation in Sow Milk and Milk Oligosaccharide Fermentation in Piglet Intestine
Difilippo, Elisabetta ; Pan, Feipeng ; Logtenberg, Madelon ; Willems, Rianne ; Braber, Saskia ; Fink-Gremmels, Johanna ; Schols, Henk Arie ; Gruppen, Harry - \ 2016
Journal of Agricultural and Food Chemistry 64 (2016)10. - ISSN 0021-8561 - p. 2087 - 2093.
abundance - chromatography - cow - mass analysis - pigs - sugars - variation
Porcine milk oligosaccharides (PMOs) were analyzed in six colostrum and two mature milk samples from Dutch Landrace sows. In total, 35 PMOs were recognized of which 13 were new for the PMO literature: Neutral HexNAc-Hex, β4′-galactosyllactose, putative GalNAc(α/β1-3)Gal(β1-4)Glc, lacto-N-fucopentaose-II, lacto-N-tetraose, galactose substituted lacto-N-neohexaose, lacto-N-hexaose and difucosyl-lacto-N-hexaose, and acidic Neu5Ac(α2-6)GlcNAc(β1-3)Gal(β1-4)Glc, sialyllacto-N-tetraose-a and -b, Neu5Ac2-Hex3, and sialyllacto-N-fucopentaose-II. PMOs were analyzed using capillary electrophoresis with laser-induced florescence detection or mass spectrometry and using liquid chromatography with mass spectrometry. Interindividual variation regarding PMO presence and concentration was observed between porcine milks. Within a limited sample set, a 43% decrease of the major PMOs was found during a 1 w lactation period. Interestingly, while some PMOs decreased, some other PMOs increased in concentration. PMOs were also monitored in fecal samples of suckling piglets. In feces of 1-2 d old piglets, few intact PMOs were found, indicating considerable PMO fermentation at early stage of life.
In Vitro Fermentation of Porcine Milk Oligosaccharides and Galacto-oligosaccharides Using Piglet Fecal Inoculum
Difilippo, Elisabetta ; Pan, Feipeng ; Logtenberg, Madelon ; Willems, Rianne ; Braber, Saskia ; Fink-Gremmels, Johanna ; Schols, Henk A. ; Gruppen, Harry - \ 2016
Journal of Agricultural and Food Chemistry 64 (2016)10. - ISSN 0021-8561 - p. 2127 - 2133.
fibers - gas chromatography - human fermentation - liquid chromatography - nondigestible carbohydrates - organic acids - short-chain fatty acids - sugars
In this study, the in vitro fermentation by piglet fecal inoculum of galacto-oligosaccharides (GOS) and porcine milk oligosaccharides (PMOs) was investigated to identify possible preferences for individual oligosaccharide structures by piglet microbiota. First, acidic PMOs and GOS with degrees of polymerization 4-7 were depleted within 12 h of fermentation, whereas fucosylated and phosphorylated PMOs were partially resistant to fermentation. GOS structures containing β1-3 and β1-2 linkages were preferably fermented over GOS containing β1-4 and β1-6 linkages. Upon in vitro fermentation, acetate and butyrate were produced as the main organic acids. GOS fermentation by piglet inoculum showed a unique fermentation pattern with respect to preference of GOS size and organic acids production.
The piglet as a model for studying dietary components in infant diets : effects of galacto-oligosaccharides on intestinal functions
Alizadeh, A. ; Akbari, P. ; Difilippo, E. ; Schols, H.A. ; Ulfman, L.H. ; Schoterman, M.H.C. ; Garssen, J. ; Fink-Gremmels, J. ; Braber, S. - \ 2016
The British journal of nutrition 115 (2016)4. - ISSN 0007-1145 - p. 605 - 618.
Galacto-oligosaccharides - Gut microbiota - Immunomodulation - Intestinal integrity - Neonatal piglet models
Prebiotic oligosaccharides, including galacto-oligosaccharides (GOS), are used in infant formula to mimic human milk oligosaccharides, which are known to have an important role in the development of the intestinal microbiota and the immune system in neonates. The maturation of the intestines in piglets closely resembles that of human neonates and infants. Hence, a neonatal piglet model was used to study the multi-faceted effect of dietary GOS in early life. Naturally farrowed piglets were separated from the mother sow 24–48 h postpartum and received a milk replacer with or without the addition of GOS for 3 or 26 d, whereafter several indicators of intestinal colonisation and maturation were measured. Dietary GOS was readily fermented in the colon, leading to a decreased pH, an increase in butyric acid in caecum digesta and an increase in lactobacilli and bifidobacteria numbers at day 26. Histomorphological changes were observed in the intestines of piglets fed a GOS diet for 3 or 26 d. In turn, differences in the intestinal disaccharidase activity were observed between control and GOS-fed piglets. The mRNA expression of various tight junction proteins was up-regulated in the intestines of piglet fed a GOS diet and was not accompanied by an increase in protein expression. GOS also increased defensin porcine β-defensin-2 in the colon and secretory IgA levels in saliva. In conclusion, by applying a neonatal piglet model, it could be demonstrated that a GOS-supplemented milk replacer promotes the balance of the developing intestinal microbiota, improves the intestinal architecture and seems to stimulate the intestinal defence mechanism.
Polarization of immune responses in fish : The 'macrophages first' point of view
Wiegertjes, Geert F. ; Wentzel, Annelieke S. ; Spaink, Herman P. ; Elks, Philip M. ; Fink, Inge R. - \ 2016
Molecular Immunology 69 (2016). - ISSN 0161-5890 - p. 146 - 156.
Arginase - Fish - INOS - LPS - Macrophage polarization - Zebrafish
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.
Oligosaccharides in Urine, Blood, and Feces of Piglets Fed Milk Replacer Containing Galacto-oligosaccharides
Difilippo, Elisabetta ; Bettonvil, Monique ; Willems, Rianne ; Braber, Saskia ; Fink-Gremmels, Johanna ; Jeurink, Prescilla V. ; Schoterman, Margriet H.C. ; Gruppen, Harry ; Schols, Henk A. - \ 2015
Journal of Agricultural and Food Chemistry 63 (2015)50. - ISSN 0021-8561 - p. 10862 - 10872.
absorption - capillary electrophoresis - creatinine - fermentation - GOS - intestine - liquid chromatography - mass spectrometry - pig - prebiotics
Human milk oligosaccharides (HMOs) are absorbed into the blood (about 1% of the HMO intake) and subsequently excreted in urine, where they may protect the infant from pathogen infection. As dietary galacto-oligosaccharides (GOS) have partial structural similarities with HMOs, this study investigated the presence of GOS and oligosaccharides originating from milk replacer in blood serum, urine, and cecal and fecal samples of piglets, as a model for human infants. Using liquid chromatography-mass spectrometry and capillary electrophoresis with fluorescence detection, oligosaccharides originating from piglet diet including 3′-sialyllactose and specific GOS ranging from degree of polymerization 3 to 6 were detected in blood serum and in urine of piglets. In blood serum, GOS levels ranged from 16 to 23 μg/mL, representing about 0.1% of the GOS daily intake. In urine, approximately 0.85 g of GOS/g of creatinine was found. Cecum digesta and feces contained low amounts of oligosaccharides, suggesting an extensive GOS intestinal fermentation in piglets.
Immune-relevant thrombocytes of common carp undergo parasite-induced nitric oxide-mediated apoptosis
Fink, I.R. ; Ribeiro, C.M.S. ; Forlenza, M. ; Taverne-Thiele, J.J. ; Rombout, J.H.W.M. ; Savelkoul, H.F.J. ; Wiegertjes, G. - \ 2015
Developmental and Comparative Immunology 50 (2015)2. - ISSN 0145-305X - p. 146 - 154.
lymphocyte maturation factor - cyprinus-carpio - expression analysis - human platelets - functional-characterization - monoclonal-antibodies - cell-differentiation - teleost fish - genes - l.
Common carp thrombocytes account for 30–40% of peripheral blood leukocytes and are abundant in the healthy animals' spleen, the thrombopoietic organ. We show that, ex vivo, thrombocytes from healthy carp express a large number of immune-relevant genes, among which several cytokines and Toll-like receptors, clearly pointing at immune functions of carp thrombocytes. Few studies have described the role of fish thrombocytes during infection. Carp are natural host to two different but related protozoan parasites, Trypanoplasma borreli and Trypanosoma carassii, which reside in the blood and tissue fluids. We used the two parasites to undertake controlled studies on the role of fish thrombocytes during these infections. In vivo, but only during infection with T. borreli, thrombocytes were massively depleted from the blood and spleen leading to severe thrombocytopenia. Ex vivo, addition of nitric oxide induced a clear and rapid apoptosis of thrombocytes from healthy carp, supporting a role for nitric oxide-mediated control of immune-relevant thrombocytes during infection with T. borreli. The potential advantage for parasites to selectively deplete the host of thrombocytes via nitric oxide-induced apoptosis is discussed.
Comparison of Milk Oligosaccharides Pattern in Colostrum of Different Horse Breeds
Difilippo, E. ; Willems, H.A.M. ; Vendrig, J.C. ; Fink-Gremmels, J. ; Gruppen, H. ; Schols, H.A. - \ 2015
Journal of Agricultural and Food Chemistry 63 (2015)19. - ISSN 0021-8561 - p. 4805 - 4814.
neutral oligosaccharides - mass-spectrometry - bovine colostrum - fed babies - lactation - feces - acid
Colostrum oligosaccharides are known to exhibit prebiotic and immunomodulatory properties. Oligosaccharide composition is species-specific, and equine colostrum has been reported to contain unique oligosaccharides. Therefore, equine oligosaccharides (EMOS) from colostrum from different horse breeds were analyzed by CE-LIF, CE-MSn, HILIC-MSn, and exoglycosidase degradation. Sixteen EMOS were characterized and quantified, of which half were neutral and half were acidic. EMOS showed about 63% structural overlap with human milk oligosaccharides, known for their bioactivity. Seven EMOS were not reported before in equine oligosaccharides literature: neutral Gal(ß1–4)HexNAc, Gal(ß1–4)Hex-Hex, ß4'-galactosyllactose, and lactose-N-hexaose, as well as acidic 6'-Sialyl-Hex-Ac-HexNAc, sialyllacto-N-tetraose-a, and disialylacto-N-tetraose (isomer not further specified). In all colostrum samples, the average oligosaccharide concentration ranged from 2.12 to 4.63 g/L; with ß 6'and 3'- galactosyllactose, 3'-sialyllactose, and disialyllactose as the most abundant of all oligosaccharides (27–59, 16–37, 1–8, and 1–6%, respectively). Differences in presence and in abundance of specific EMOS were evident not only between the four breeds but also within the breed.
Molecular and functional characterization of the scavenger receptor CD36 in zebrafish and common carp
Fink, I.R. ; Benard, E.L. ; Hermsen, G.J. ; Meijer, A.H. ; Forlenza, M. ; Wiegertjes, G. - \ 2015
Molecular Immunology 63 (2015)2. - ISSN 0161-5890 - p. 381 - 393.
toll-like receptors - salmon salmo-salar - cyprinus-carpio - density-lipoprotein - neutrophilic granulocytes - monoclonal-antibodies - accessory molecules - innate immunity - gene family - in-vivo
CD36 is a scavenger receptor which has been studied closely in mammals where it is expressed by many different cell types and plays a role in highly diverse processes, both homeostatic and pathologic. It is among other things important in the innate immune system, in angiogenesis, and in clearance of apoptotic cells, and it is also involved in lipid metabolism and atherosclerosis. Recently, in the cephalochordate amphioxus a primitive CD36 family member was described, which was present before the divergence of CD36 from other scavenger receptor B family members, SCARB1 and SCARB2. Not much is known on the Cd36 molecule in teleost fish. We therefore studied Cd36 in both zebrafish and common carp, two closely related cyprinid fish species. Whereas a single cd36 gene is present in zebrafish, carp has two cd36 genes, and all show conserved synteny compared to mammalian CD36. The gene expression of carp cd36 is high in brain, ovary and testis but absent in immune organs. Although in mammals CD36 expression in erythrocytes, monocytes and macrophages is high, gene expression studies in leukocyte subtypes of adult carp and zebrafish larvae, including thrombocytes and macrophages provided no indication for any substantial expression of cd36 in immune cell types. Surprisingly, analysis of the cd36 promoter region does show the presence of several binding sites for transcription factors known to regulate immune responses. Overexpression of carp cd36 locates the receptor on the cell surface of mammalian cell lines consistent with the predicted topology of cyprinid Cd36 with a large extracellular domain, two transmembrane domains, and short cytoplasmic tails at both ends. Gene expression of cd36 is down-regulated during infection of zebrafish with Mycobacterium marinum, whereas knockdown of cd36 in zebrafish larvae led to higher bacterial burden upon such infection. We discuss the putative role for Cd36 in immune responses of fish in the context of other members of the scavenger receptor class B family.
Identification and functional characterization of nonmammalian Toll-like receptor 20
Pietretti, D. ; Scheer, M.H. ; Fink, I.R. ; Taverne, N. ; Savelkoul, H.F.J. ; Spaink, H.P. ; Forlenza, M. ; Wiegertjes, G. - \ 2014
Immunogenetics 66 (2014)2. - ISSN 0093-7711 - p. 123 - 141.
carp cyprinus-carpio - leucine-rich repeats - common carp - ictalurus-punctatus - monoclonal-antibodies - pathogen recognition - accessory molecules - expression analysis - sequence-analysis - channel catfish
Like other vertebrate Toll-like receptors (TLRs), the TLRs of teleost fish can be subdivided into six major families, each of which recognize a general class of molecular patterns. However, there also are a number of Tlrs with unknown function, the presence of which seems unique to the bony fish, among which is Tlr20. We identified full-length complementary DNA (cDNA) sequences for tlr20 of zebrafish and common carp, two closely related fish species. Zebrafish have six copies of tlr20, whereas carp express only a single copy. Both zebrafish Tlr20 (at least Tlr20a–d) and carp Tlr20 have 26 leucine-rich repeats (LRRs). Three-dimensional modeling indicates a best fit to the crystal structure of TLR8. Phylogenetic analyses place Tlr20 in the TLR11 family closest to Tlr11 and Tlr12, which sense ligands from protozoan parasites in the mouse. Conservation of genes on zebrafish chromosome 9, which carries tlr20, with genes on mouse chromosome 14, which carries tlr11, indicates Tlr11 could be a possible ortholog of Tlr20. Confocal microscopy suggests a subcellular localization of Tlr20 at the endoplasmatic reticulum. Although in vitro reporter assays could not identify a ligand unique to Tlr20, in vivo infection experiments indicate a role for Tlr20 in the immune response of carp to protozoan parasites (Trypanoplasma borreli). Carp tlr20 is mainly expressed in peripheral blood leukocytes (PBL) with B lymphocytes, in particular, expressing relatively high levels of Tlr20. In vitro stimulation of PBL with T. borreli induces an upregulation of tlr20, supportive of a role for Tlr20 in the immune response to protozoan parasites.
Chronic Allopurinol Treatment during the Last Trimester of Pregnancy in Sows: Effects on Low and Normal Birth Weight Offspring
Gieling, E.T. ; Antonides, A. ; Fink-Gremmels, J. ; Haar, K. ter; Kuller, W.I. ; Meijer, E. ; Nordquist, R.E. ; Stouten, J.M. ; Zeinstra, E. ; Staay, F.J. van der - \ 2014
PLoS ONE 9 (2014)1. - ISSN 1932-6203 - 15 p.
intrauterine growth-restriction - for-gestational-age - children born - placental insufficiency - attentional problems - newborn piglets - spleen weight - brain - memory - stress
Low-birth-weight (LBW) children are born with several risk factors for disease, morbidity and neonatal mortality, even if carried to term. Placental insufficiency leading to hypoxemia and reduced nutritional supply is the main cause for LBW. Brain damage and poor neurological outcome can be the consequence. LBW after being carried to term gives better chances for survival, but these children are still at risk for poor health and the development of cognitive impairments. Preventive therapies are not yet available. We studied the risk/efficacy of chronic prenatal treatment with the anti-oxidative drug allopurinol, as putative preventive treatment in piglets. LBW piglets served as a natural model for LBW. A cognitive holeboard test was applied to study the learning and memory abilities of these allopurinol treated piglets after weaning. Preliminary analysis of the plasma concentrations in sows and their piglets suggested that a daily dose of 15 mg.kg-1 resulted in effective plasma concentration of allopurinol in piglets. No adverse effects of chronic allopurinol treatment were found on farrowing, birth weight, open field behavior, learning abilities, relative brain, hippocampus and spleen weights. LBW piglets showed increased anxiety levels in an open field test, but cognitive performance was not affected by allopurinol treatment. LBW animals treated with allopurinol showed the largest postnatal compensatory body weight gain. In contrast to a previous study, no differences in learning abilities were found between LBW and normal-birth-weight piglets. This discrepancy might be attributable to experimental differences. Our results indicate that chronic prenatal allopurinol treatment during the third trimester of pregnancy is safe, as no adverse side effects were observed. Compensatory weight gain of treated piglets is a positive indication for the chronic prenatal use of allopurinol in these animals. Further studies are needed to assess the possible preventive effects of allopurinol on brain functions in LBW piglets.
A closer look at Toll-like receptor 4 (TLR4) and toll-like receptor 20 (TLR20) of common carp (Cyprinus carpio)
Pietretti, D. ; Forlenza, M. ; Fink, I.R. ; Wiegertjes, G.F. - \ 2013
Fish and Shellfish Immunology 34 (2013)6. - ISSN 1050-4648 - p. 1673 - 1673.
Toll-like receptors (TLRs) constitute an important class of pattern-recognition receptors, which recognize a multitude of pathogen-associated molecular patterns (PAMPs). We focused on TLR4 and TLR20 of common carp (Cyprinus carpio), their signaling pathway and their possible functions as activating receptors involved in innate immune responses to various pathogens or immunostimulants. So far the expression of TLR4 and TLR20 genes has been reported only for few fish species of the Cypriniformes (zebrafish, grass carp, common carp) and close relatives Siluriformes (channel catfish). Multiple fish TLR4 genes, created by a (recent) duplication rather than speciation event, seem to exist. It is possible that these paralogs have evolved to express different ligand specificities. In mammals TLR4 recognizes lipopolysaccharides from Gram-negative bacteria. The ligand(s) for fish TLR4 have not been confirmed. Multiple TLR20 genes also seem to exist in zebrafish, but not in carp. Our phylogenetic analysis indicates an ancestral relationship could exist between TLR20 and TLR11/TLR12, two TLRs found in mice but not in humans with ligand specificity for profilin from parasitic Toxoplasma gondii. At present, the ligand of TLR20 is unknown. We have cloned full-length coding sequences of carp TLR4a/b and carp TLR20 to study the function of these molecules. Expression analysis using real-time PCR show both TLR4 and TLR20 are mainly expressed in immune organs such as head kidney, gut and spleen with neutrophilic granulocytes as the primary source. We have also investigated the expression of TLR4 and of TLR20 after infection with protozoan parasites or after infection with spring vireamia of carp virus (SVCV). For TLR4, neither challenge with parasites nor with virus induced major changes in gene expression. However, we find high gene expression of TLR20 after infection with the protozoan parasites Trypanoplasma borreli or Trypanosoma carassii. We used the full-length sequences to create fluorescent protein-tagged TLRs for visualization of localization of these receptors by fluorescence microscopy. Our study suggests both receptors are located mainly in the cytoplasmic region. To define the unknown ligands of TLR4 and TLR20 we overexpressed these receptors in both human cell lines (HEK 293) and fish cell lines (EPC, CLC) stably transfected with a promoter of the transcription factor NF-¿B and a luciferase reporter gene. These studies may help to identify ligands for carp TLR4 a/b and TLR20.
Molecular cloning and cellular localization of the scavenger receptor SCARF1 in common carp
Østergaard, A.E. ; Fink, I.R. ; Sukinta, A. ; Yixian, L. ; Forlenza, M. ; Wiegertjes, G.F. - \ 2013
Fish and Shellfish Immunology 34 (2013)6. - ISSN 1050-4648 - p. 1727 - 1727.
The recognition of pathogens by the innate immune system relies on a wide range of germ-line encoded pattern recognition receptors of which some belong to the large superfamily of scavenger receptors (SRs). These receptors are expressed on cells surveying potential portals of entry, including macrophages, dendritic cells and endothelial cells and are involved in recognition of both endogenous and pathogen-derived ligands. Here we describe for the first time an evolutionarily conserved member of the SR family, named scavenger receptor class F-member1 (SCARF1), in the common carp (Cyprinus carpio). Orthologs of mammalian SCARF1 have been reported in nematodes and shown to be involved in recognition of e.g. beta-glucans. Also, SCARF1 has been shown to mediate an inflammatory cytokine response upon ligand binding whilst functioning as co-receptor for Toll-like receptors (TLRs). We found two SCARF1 genes in the common carp genome, confirming the hypothesis that carp has undergone an extra genome duplication event compared to zebrafish, where only one SCARF1 gene is found. The two SCARF1 genes in carp are 94% similar and show comparable mRNA expression levels. Sequence analysis of SCARF1 reveals a high level of amino acid conservation compared to the human orthologue. Phylogenetic analysis of carp SCARF1 with other vertebrate SCARF1 genes groups carp and zebrafish together with high bootstrap values. Synteny studies show conserved linkage with several neighbouring genes when comparing genomic regions from different species. Gene expression analysis of SCARF1 shows expression in several organs, whereas gene expression analysis of purified cell populations shows highest expression in endothelial cells, but also macrophages, granulocytes, thrombocytes and thymocytes show SCARF1 gene expression. In contrast, no expression was observed in B cells. Previously, our group has cloned TLR2 from common carp and we are now investigating the possible role of SCARF1 acting as an internalizing receptor that would facilitate TLR activation upon phagocytosis of ligands. The localization of SCARF1 was studied by confocal microscopy of human (HEK293) and fish (EPC) cell lines transfected with a fluorescently tagged receptor. In addition, antibodies directed against a V5-tagged version of SCARF1 were also used to detect SCARF1 localization. These studies showed that carp SCARF1 is expressed in the cytoplasm, but also on the cell membrane, which is comparable to human SCARF1 localization. We are performing phagocytosis studies in order to verify the internalizing ability of SCARF1 and its potential role in TLR signaling.
|Toll-like receptor-1 and -2 in common carp
Fink, I.R. ; Pietretti, D. ; Forlenza, M. ; Wiegertjes, G.F. - \ 2013
Fish and Shellfish Immunology 34 (2013)6. - ISSN 1050-4648 - p. 1706 - 1707.
|Research on herbal products for production animals in the Netherlands
Groot, M.J. ; Asseldonk, A.G.M. ; Fink-Gremmels, J. ; Lourens, S. ; Wagenaar, J.P. ; Kleijer Ligtenberg, G. - \ 2013
In: Proceedings of the 61st Annual Meeting of the Society for Medicinal Plant and Natural Product Research, 1-5 September 2013, Stuttgart, Germany. - Stuttgart : Georg Thieme Verlag KG - p. 1123 - 1123.
Cytotoxicity of surface-functionalized silicon and germanium nanoparticles: the dominant role of surface charges
Bhattacharjee, S. ; Rietjens, I.M.C.M. ; Singh, M.P. ; Atkins, T.M. ; Purkait, T.K. ; Xu, Z. ; Regli, S. ; Shukaliak, A. ; Clark, R.J. ; Mitchell, B.S. ; Alink, G.M. ; Marcelis, A.T.M. ; Fink, M.J. ; Veinot, J.G.C. ; Kauzlarich, S.M. ; Zuilhof, H. - \ 2013
Nanoscale 5 (2013). - ISSN 2040-3364 - p. 4870 - 4883.
block copolymer nanoparticles - cerium oxide nanoparticles - quantum dots - in-vitro - oxidative stress - gold nanoparticles - cellular toxicity - dependent endocytosis - hemolytic-activity - epithelial-cells
Although it is frequently hypothesized that surface (like surface charge) and physical characteristics (like particle size) play important roles in cellular interactions of nanoparticles (NPs), a systematic study probing this issue is missing. Hence, a comparative cytotoxicity study, quantifying nine different cellular endpoints, was performed with a broad series of monodisperse, well characterized silicon (Si) and germanium (Ge) NPs with various surface functionalizations. Human colonic adenocarcinoma Caco-2 and rat alveolar macrophage NR8383 cells were used to clarify the toxicity of this series of NPs. The surface coatings on the NPs appeared to dominate the cytotoxicity: the cationic NPs exhibited cytotoxicity, whereas the carboxylic acid-terminated and hydrophilic PEG- or dextran-terminated NPs did not. Within the cationic Si NPs, smaller Si NPs were more toxic than bigger ones. Manganese-doped (1% Mn) Si NPs did not show any added toxicity, which favors their further development for bioimaging. Iron-doped (1% Fe) Si NPs showed some added toxicity, which may be due to the leaching of Fe3+ ions from the core. A silica coating seemed to impart toxicity, in line with the reported toxicity of silica. Intracellular mitochondria seem to be the target for the toxic NPs since a dose-, surface charge- and size-dependent imbalance of the mitochondrial membrane potential was observed. Such an imbalance led to a series of other cellular events for cationic NPs, like decreased mitochondrial membrane potential (¿¿m) and ATP production, induction of ROS generation, increased cytoplasmic Ca2+ content, production of TNF-a and enhanced caspase-3 activity. Taken together, the results explain the toxicity of Si NPs/Ge NPs largely by their surface characteristics, provide insight into the mode of action underlying the observed cytotoxicity, and give directions on synthesizing biocompatible Si and Ge NPs, as this is crucial for bioimaging and other applications in for example the field of medicine.
|Studies into the formation of heterodimers of Toll-like receptor-1 and -2 in common carp
Fink, I.R. ; Østergaard, A.E. ; Forlenza, M. ; Wiegertjes, G.F. - \ 2012
In: 12th Congress of ISDCI program and abstracts, Fukuoka, 9-13 July 2012. - - p. 51 - 51.
|Pattern recognition receptors
Fink, I.R. - \ 2012
In: Fish Immunology Workshop 2012, Wageningen, 22-26 April 2012. -
Kinesin-3 and dynein cooperate in long-range retrograde endosome motility along a nonuniform microtubule array
Schuster, M. ; Kilaru, S. ; Fink, G. ; Collemare, J.A.R. ; Roger, Y. ; Steinberg, G. - \ 2011
Molecular Biology of the Cell 22 (2011)19. - ISSN 1059-1524 - p. 3645 - 3657.
fungus ustilago-maydis - tug-of-war - lipid-droplet transport - cytoplasmic dynein - molecular motors - intracellular-transport - vesicle transport - cargo transport - caenorhabditis-elegans - polarity orientation
The polarity of microtubules (MTs) determines the motors for intracellular motility, with kinesins moving to plus ends and dynein to minus ends. In elongated cells of Ustilago maydis, dynein is thought to move early endosomes (EEs) toward the septum (retrograde), whereas kinesin-3 transports them to the growing cell tip (anterograde). Occasionally, EEs run up to 90 mu m in one direction. The underlying MT array consists of unipolar MTs at both cell ends and antipolar bundles in the middle region of the cell. Cytoplasmic MT-organizing centers, labeled with gamma-tubulin ring complex protein, are distributed along the antipolar MTs but are absent from the unipolar regions. Dynein colocalizes with EEs for 10-20 mu m after they have left the cell tip. Inactivation of temperature-sensitive dynein abolishes EE motility within the unipolar MT array, whereas long-range motility is not impaired. In contrast, kinesin-3 is continuously present, and its inactivation stops long-range EE motility. This indicates that both motors participate in EE motility, with dynein transporting the organelles through the unipolar MT array near the cell ends, and kinesin-3 taking over at the beginning of the medial antipolar MT array. The cooperation of both motors mediates EE movements over the length of the entire cell.
|Molecular and functional characterization of toll-like receptor 4 in common carp (Cyprinus carpio)
Pietretti, D. ; Fink, I.R. ; Forlenza, M. ; Wiegertjes, G.F. - \ 2011
In: 15th international conference on disease of fish and shellfish Split, Croatia, 12 - 16 September, 2011. - - p. 72, O - 060.
|Molecular and functional characterization of toll-like receptor-1 in common carp (Cyprinus carpio)
Fink, I.R. ; Forlenza, M. ; Wiegertjes, G.F. - \ 2011
In: 15th international conference on disease of fish and shellfish, Split Croatia, 12 - 16 September, 2011. - - p. 70, O - 058.
|Heterogeneity of macrophage activation during parasite infections
Wiegertjes, G.F. ; Fink, I.R. ; Forlenza, M. - \ 2011
In: The 15th International Conference on Diseases of Fish and Shellfish organized by the EAFP, Split, Croatia, 12 - 16 September, 2011. - - p. 17 (O - 008).
|Molecular and functional characterization of scavenger receptor class F member 1 (SCARF1) and CD36 in common carp (Cyprinus carpio)
Østergaard, A.E. ; Fink, I.R. ; Sukinta, A. ; Wiegertjes, G.F. - \ 2011
In: Abstract book of the 15th International Conference on Disease of Fish and Shellfish, Split, Croatia, 12-16 September 2011. - - p. 71, O - 059.
|Functional evidence for toll-like receptors in modern bony fish
Wiegertjes, G.F. ; Ribeiro, C.M.S. ; Pietretti, D. ; Fink, I.R. ; Østergaard, A.E. ; Forlenza, M. ; Scheer, M.H. ; Savelkoul, H.F.J. - \ 2011
In: Proceedings of the 25th Annual EMDS Meeting, clinical and fundamental aspects of monocyte macrophage and DC plasticity, Brussels, Belgium, 22-24 september 2011. - - p. 21, B10 - 21, B10.
Heterogeneity of macrophage activation in fish
Forlenza, M. ; Fink, I.R. ; Raes, G. ; Wiegertjes, G.F. - \ 2011
Developmental and Comparative Immunology 35 (2011)12. - ISSN 0145-305X - p. 1246 - 1255.
tumor-necrosis-factor - cyprinus-carpio l. - trout oncorhynchus-mykiss - nitric-oxide synthase - toll-like receptors - interferon-gamma genes - carassius-auratus l. - expression analysis - rainbow-trout - factor-alpha
In this review, we focus on four different activation states of fish macrophages. In vitro, stimulation with microbial ligands induces the development of innate activated macrophages whereas classically activated macrophages can be induced by stimulation with LPS in combination with (recombinant) IFN¿. Both types of macrophages show elevated phagocytic activity, expression of pro-inflammatory cytokine genes and radical production. Alternatively activated macrophages require the cytokines IL-4/IL-13 for induction of, among others, arginase activity. Until in vitro studies identify the effects of putative IL-4 and IL-13 homologues on fish macrophages, arginase enzyme activity remains the most reliable marker for the presence of alternatively activated macrophages in fish. The best evidence for the existence of regulatory macrophages, associated with the presence of IL-10, comes from in vivo studies, for example during parasitic infections of carp. Altogether, differentially activated macrophages in fish largely resemble the phenotypes of mammalian macrophages. However, the presence of fish-specific ligand recognition by TLRs and of duplicated genes coding for proteins with particular activities, poses additional challenges for the characterization of phenotype-specific gene signatures and cell surface markers.
Adaptive braking by Ase1 prevents overlapping microtubules from sliding completely apart.
Braun, M. ; Lansky, Z. ; Fink, G. ; Ruhnow, F. ; Diez, S. ; Janson, M.E. - \ 2011
Nature Cell Biology 13 (2011). - ISSN 1465-7392 - p. 1259 - 1264.
fission yeast - spindle midzone - cross-linkers - protein - kinesin-14 - binding - motors - prc1 - organization - cytokinesis
Short regions of overlap between ends of antiparallel microtubules are central elements within bipolar microtubule arrays. Although their formation requires motors1, recent in vitro studies demonstrated that stable overlaps cannot be generated by molecular motors alone. Motors either slide microtubules along each other until complete separation2, 3, 4 or, in the presence of opposing motors, generate oscillatory movements5, 6, 7. Here, we show that Ase1, a member of the conserved MAP65/PRC1 family of microtubule-bundling proteins, enables the formation of stable antiparallel overlaps through adaptive braking of Kinesin-14-driven microtubule–microtubule sliding. As overlapping microtubules start to slide apart, Ase1 molecules become compacted in the shrinking overlap and the sliding velocity gradually decreases in a dose-dependent manner. Compaction is driven by moving microtubule ends that act as barriers to Ase1 diffusion. Quantitative modelling showed that the molecular off-rate of Ase1 is sufficiently low to enable persistent overlap stabilization over tens of minutes. The finding of adaptive braking demonstrates that sliding can be slowed down locally to stabilize overlaps at the centre of bipolar arrays, whereas sliding proceeds elsewhere to enable network self-organization.
Molecular characterization of Toll-like and scavenger receptors activated by B-glucans in common carp (Cyprinus carpio)
Fink, I.R. ; Oestergaard, A.E. ; Wiegertjes, G.F. - \ 2010
|Molecular characterization of TOLL-like receptor 4B in common carp
Pietretti, D. ; Fink, I.R. ; Oestergaard, A.E. ; Savelkoul, H.F.J. ; Wiegertjes, G.F. - \ 2010
In: Abstracts First EOFFI Symposium European Organisation of Fish Immunology, Viterbo, Italy, 23-27 May 2010. - - p. 85 - 85.
Biofilm formation and genotyping of Staphylococcus aureus bovine mastitis isolates: Evidence for lack of penicillin-resistance in Agr-type II strains
Melchior, M.B. ; Osch, M.H.J. ; Graat, R. ; Duijkeren, E. van; Mevius, D.J. ; Nielen, M. ; Gaastra, W. ; Fink, J. - \ 2009
Veterinary Microbiology 137 (2009)1-2. - ISSN 0378-1135 - p. 83 - 89.
polysaccharide intercellular adhesin - quaternary ammonium-compounds - sequence element is256 - somatic-cell count - antimicrobial susceptibility - genetic-variability - epidermidis - milk - infections - protein
The increasing evidence for a role of biofilm formation in bovine mastitis caused by Staphylococcus aureus led to further investigations on biofilm formation by S. aureus isolates from mastitis in two growth media (TSBg and bovine milk serum). The ability of 99 S. aureus strains that were recently isolated or obtained from a culture collection (historical strains) to form biofilm, in both growth media as well as the correlation of biofilm formation with the presence of the ica-, bap-, and IS257 genes are described. These genes have been correlated with biofilm formation by human S. aureus isolates. All strains were also genotyped with respect to their Agr-type and -subtype, and for the presence of the antibiotic resistance genes blaZ and smr by PCR. The prevalence of the Agr-types and the investigated genes and their correlation with biofilm formation were statistically evaluated. The Agr-type of a strain had a marked effect on the biofilm formation, by that strain, however in contrast to human isolates no significant effect of ica- and IS257 genes on biofilm formation was observed. The bap gene was not found in any of the investigated strains. The presence of biofilm related genes showed a high correlation with the Agr-type of the strains. The data give evidence for a very strong correlation of Agr-type I strains and penicillin-resistance in the bovine S. aureus mastitis strains; none of the Agr-type II strains was found to harbor penicillin-resistance genes. These data indicate that the most prevalent Agr-types in S. aureus bovine mastitis, Agr-type I and II, can be regarded as different subspecies, with different abilities for the formation of biofilm in bovine milk serum. The very high correlation between Agr-type II and penicillin-susceptibility strongly suggests that these strains are not able to accommodate blaZ genes
Fyto-V eindrapport, Ontwikkelen van fytotherapie als middel bij het reduceren van en/of behandelen van dierziekten
Groot, M.J. ; Noordam, M.Y. ; Kleter, G.A. ; Asseldonk, A.G.M. ; Kleijer-Ligtenberg, E. ; Halkes, S.B.A. ; Fink-Gremmels, J. ; Osch, H.H. van - \ 2009
Wageningen : RIKILT (Rapport / RIKILT, Instituut voor Voedselveiligheid 2008.010) - 1009
medicinale planten - veeartsenijkunde - vee - diervoedering - voedertoevoegingen - voedersupplementen - wetgeving - regelingen - nederland - europa - medicinal plants - veterinary medicine - livestock - animal feeding - feed additives - feed supplements - legislation - regulations - netherlands - europe
This report describes the current legislation in the Netherlands, Europe and in a number of other important countries regarding the use of herbs in animals and in humans; the bottlenecks in current legislation as noticed by the registration authorities, the industry and the animal production sector; the recommendations of the project group to solve these bottlenecks.
Redirection of flavonoid biosynthesis through the down-regulation of an anthocyanidin glucosyltransferase in ripening strawberry fruit
Griesser, M. ; Hoffmann, T. ; Bellido, M.L. ; Rosati, C. ; Fink, B. ; Kurtzer, R. ; Aharoni, A. ; Munoz-Blanco, J. ; Schwab, W. - \ 2008
Plant Physiology 146 (2008)4. - ISSN 0032-0889 - p. 1528 - 1539.
fragaria x ananassa - phenylalanine ammonia-lyase - udp-glucose - gene-expression - molecular characterization - arabidopsis-thaliana - oxidative stress - petunia-hybrida - pathway genes - cinnamic acid
Strawberry (Fragaria x ananassa) fruit contains several anthocyanins that give the ripe fruits their attractive red color. The enzyme that catalyzes the formation of the first stable intermediate in the anthocyanin pathway is anthocyanidin-3-O-glucosyltransferase. A putative glycosyltransferase sequence (FaGT1) was cloned from a strawberry fruit cDNA library and the recombinant FaGT1 transferred UDP-glucose to anthocyanidins and, to a lesser extent, flavonols, generating the respective 3-O-glucosides. Quantitative polymerase chain reaction revealed that transcripts of FaGT1 were almost undetectable in green fruits, but gene expression increased dramatically in both turning and ripe red fruit, corresponding closely to the accumulation of anthocyanins during fruit ripening. The expression of FaGT1 is fruit associated and negatively regulated by auxin. To elucidate the in planta function of FaGT1, Agrobacterium tumefaciens cells harboring an intron-hairpin construct of a partial FaGT1 sequence were injected into midsized ripening fruits. In about one-third of the injected fruits, this led to significant down-regulation of FaGT1 transcript levels that corresponded to reduced concentrations of anthocyanin pigments in ripe strawberry fruits. In contrast, significant levels of epiafzelechin-formed by anthocyanidin reductase (ANR) from pelargonidin-were identified in FaGT1-silenced fruits, indicating competition of FaGT1 and FaANR for the common anthocyanidin substrate. Thus, FaGT1 represents an important branching-point enzyme because it is channeling the flavonoid pathway to anthocyanins. These results demonstrate a method to redirect the anthocyanin biosynthesis into flavan-3-ol production to increase the levels of bioactive natural products or modify pigments in plant tissues.
|Genotyping and biofilm formation in different growth media of Staphylococcus aureus bovine mastitis isolates: evidence for lack of penicillin-resistance in Agr-type II strains
Melchior, M.B. ; Osch, M.H.J. ; Graat, R. ; Duijkeren, E. van; Mevius, D.J. ; Nielen, M. ; Gaastra, W. ; Fink-Gremmels, J. - \ 2008
Strategy and Culture in the International Co-ordination Mechanisms
Gusc, J.S. ; Bremmers, H.J. ; Zuraw, J. - \ 2004
In: Papers of participants of the Summer School on Intercultural Business Management in Central Eastern Europe, Vienna, Austria, 26-04-2003 t/m 10-05-2003. - Vienna, Austria : Institut für den Donauraum und Mitteleuropa - p. 59 - 70.
A revised secondary structure model for the internal transcribed spacer 2 of the green algae Scenedesmus and Desmodesmus and its implication for the phylogeny of these algae
Hannen, E.J. van; Fink, P. ; Lürling, M. - \ 2002
European Journal of Phycology 37 (2002). - ISSN 0967-0262 - p. 203 - 208.
Secondary structure analysis of 34 internal transcribed spacer 2 (ITS-2) sequences showed that the current model for the green algae Scenedesmus and Desmodesmus is not accurate. In particular, helix I of the currently used model showed considerable deviations from our new model. The newly proposed model is supported by many two-sided compensated base pair changes and fully compensated insertions in all four helices. Phylogenetic analysis by maximum parsimony based on the new alignment confirmed the recent division of the old genus Scenedesmus into the new genera Scenedesmus and Desmodesmus. However, the analysis was not able to show phylogenetic relationships within these two genera. Hence, the ITS-2 region alone is not suitable for clarifying the phylogeny of Scenedesmus and Desmodesmus and new regions have to be found for future sequence analyses
Health Council of the Netherlands. Deoxynivalenol (DON).
Koeman, J.H. ; Feron, V.J. ; Fink-Gremmels, J. ; Lowik, M.R.H. ; Mulder, G.J. ; Sauer, P.J.J. ; Waard, M.A. de; Stoppelaar, J.M. de; Theelen, R.M.C. ; Pijls, L.T.J. - \ 2001
The Hague, the Netherlands : Health Council of the Netherlands - ISBN 9789055493944 - 70
fusarium - toxinogene schimmels - risicoschatting - tarwe - vomitoxine - fusarium - toxinogenic fungi - risk assessment - wheat - vomitoxin
|Voedermiddelen en risico's voor de consument van dierlijke producten : advies van de Commissie Risicovolle Voedermiddelen
Baars, J.A. ; Fink, J. ; Emmels, J. ; Hofstra, H. ; Jongen, W. ; Mengelen, M.J.B. ; Verstegen, M.W.A. ; Wever, C.J.G. - \ 1999
Ede : IKC - 16 p.
|Infrared Photothermal Spectroscopy in the Science of Human Nutrition
Bicanic, D. ; Fink, T. ; Franko, M. ; Mocnik, G. ; Bovenkamp, P. van de; Veldhuizen, A. van; Gerkema, E. - \ 1999
In: Photoacoustic and Photothermal Phenomena / Scudieri, F., Bertolotti, M., Woodburry, New York : American Institute of Physics (American Institute of Physics (AIP) Conference Proceedings 463) - ISBN 9781563968051 - p. 637 - 639.
|Application of infrared photothermal spectroscopy in human nutrition.
Bicanic, D. ; Fink, T. ; Franko, M. ; Bovenkamp, P. van de; Veldhuizen, B. van; Gerkema, E. - \ 1998
In: 10th International Conference on Photoacoustic and Photothermal Phenomena, Rome, The Netherlands - p. 513 - 514.
CO2 laser photoacoustic spectroscopy and absolute absorption coefficients of short chain saturated fatty acid vapours.
Fink, T. ; Braun, R. ; Bicanic, D. - \ 1998
Instrumentation Science and Technology 26 (1998). - ISSN 1073-9149 - p. 189 - 202.
|Photoacoustic and photothermal methods as a tool to aid authenticity tests and quality assessment of foods
Bicanic, D. ; Dóka, O. ; Gibkes, J. ; Offermann, S. ; Dadarlat, D. ; Keyzer, C. ; Long, G. ; Fink, T. ; Gerkema, E. ; Bein, B. ; Boekel, T. van; Jalink, H. - \ 1996
Progress in Natural Science 6 (1996). - ISSN 1002-0071 - p. 573 - 576.
|New applications of gas photoacoustic spectroscopy in food/dairy sciences, horticulture and environmental technology: absorption of trans fatty acid vapours, observation of heat induced effects in ultrafiltrated cow milk protein concentrate and studies of flower pollen and contaminated (PAHC) soil.
Dóka, O. ; Bicanic, D. ; Fink, T. ; Alebic-Juretic, A. ; Mastrigt, S. van; Pol, P. van der - \ 1996
In: Digest 9th Topical Int. Conf. on Photoacoustic and Photothermal Phenomena, Nanjing China - p. 438 - 439.
|High temperature photoacoustic cells for recording infrared vapour phase spectra of saturated and unsaturated fatty acids.
Bicanic, D. ; Fink, T. ; Gerkema, E. ; Jalink, H. - \ 1996
Acustica - acta acustica 82 (1996). - ISSN 0001-7884 - p. S116 - S116.