Publications by authors named "Annemieke van der Wal"

24 Publications

  • Page 1 of 1

Effect of the amount of organic trigger compounds, nitrogen and soil microbial biomass on the magnitude of priming of soil organic matter.

PLoS One 2019 16;14(5):e0216730. Epub 2019 May 16.

National Institute for Public Health and the Environment, Bilthoven, The Netherlands.

Priming effects (PEs) are defined as short-term changes in the turnover of soil organic matter (SOM) caused by the addition of easily degradable organic compounds to the soil. PEs are ubiquitous but the direction (acceleration or retardation of SOM decomposition) and magnitude are not easy to predict. It has been suggested that the ratio between the amount of added PE-triggering substrate to the size of initial soil microbial biomass is an important factor influencing PEs. However, this is mainly based on comparison of different studies and not on direct experimentation. The aim of the current study is to examine the impact of glucose-to-microbial biomass ratios on PEs for three different ecosystems. We did this by adding three different amounts of 13C-glucose with or without addition of mineral N (NH4NO3) to soils collected from arable lands, grasslands and forests. The addition of 13C-glucose was equivalent to 15%, 50% and 200% of microbial biomass C. After one month of incubation, glucose had induced positive PEs for almost all the treatments, with differences in magnitude related to the soil origin and the amount of glucose added. For arable and forest soils, the primed C increased with increasing amount of glucose added, whereas for grassland soils this relationship was negative. We found positive correlations between glucose-derived C and primed C and the strength of these correlations was different among the three ecosystems considered. Generally, additions of mineral N next to glucose (C:N = 15:1) had little effect on the flux of substrate-derived C and primed C. Overall, our study does not support the hypothesis that the trigger-substrate to microbial biomass ratio can be an important predictor of PEs. Rather our results indicate that the amount of energy obtained from decomposing trigger substrates is an important factor for the magnitude of PEs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0216730PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6522013PMC
January 2020

Antifungal Rhizosphere Bacteria Can increase as Response to the Presence of Saprotrophic Fungi.

PLoS One 2015 22;10(9):e0137988. Epub 2015 Sep 22.

Department of Microbial Ecology, Netherlands Institute of Ecology (NIOO-KNAW), Wageningen, 6708 PB, the Netherlands.

Knowledge on the factors that determine the composition of bacterial communities in the vicinity of roots (rhizosphere) is essential to understand plant-soil interactions. Plant species identity, plant growth stage and soil properties have been indicated as major determinants of rhizosphere bacterial community composition. Here we show that the presence of saprotrophic fungi can be an additional factor steering rhizosphere bacterial community composition and functioning. We studied the impact of presence of two common fungal rhizosphere inhabitants (Mucor hiemalis and Trichoderma harzianum) on the composition of cultivable bacterial communities developing in the rhizosphere of Carex arenaria (sand sedge) in sand microcosms. Identification and phenotypic characterization of bacterial isolates revealed clear shifts in the rhizosphere bacterial community composition by the presence of two fungal strains (M. hiemalis BHB1 and T. harzianum PvdG2), whereas another M. hiemalis strain did not show this effect. Presence of both M. hiemalis BHB1 and T. harzianum PvdG2 resulted in a significant increase of chitinolytic and (in vitro) antifungal bacteria. The latter was most pronounced for M. hiemalis BHB1, an isolate from Carex roots, which stimulated the development of the bacterial genera Achromobacter and Stenotrophomonas. In vitro tests showed that these genera were strongly antagonistic against M. hiemalis but also against the plant-pathogenic fungus Rhizoctonia solani. The most likely explanation for fungal-induced shifts in the composition of rhizosphere bacteria is that bacteria are being selected which are successful in competing with fungi for root exudates. Based on the results we propose that measures increasing saprotrophic fungi in agricultural soils should be explored as an alternative approach to enhance natural biocontrol against soil-borne plant-pathogenic fungi, namely by stimulating indigenous antifungal rhizosphere bacteria.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0137988PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4578881PMC
May 2016

Explaining bacterial dispersion on leaf surfaces with an individual-based model (PHYLLOSIM).

PLoS One 2013 4;8(10):e75633. Epub 2013 Oct 4.

Department of Microbial Ecology, Netherlands Institute of Ecology (NIOO-KNAW), Wageningen, The Netherlands.

We developed the individual-based model PHYLLOSIM to explain observed variation in the size of bacterial clusters on plant leaf surfaces (the phyllosphere). Specifically, we tested how different 'waterscapes' impacted the diffusion of nutrients from the leaf interior to the surface and the growth of individual bacteria on these nutrients. In the 'null' model or more complex 'patchy' models, the surface was covered with a continuous water film or with water drops of equal or different volumes, respectively. While these models predicted the growth of individual bacterial immigrants into clusters of variable sizes, they were unable to reproduce experimentally derived, previously published patterns of dispersion which were characterized by a much larger variation in cluster sizes and a disproportionate occurrence of clusters consisting of only one or two bacteria. The fit of model predictions to experimental data was about equally poor (<5%) regardless of whether the water films were continuous or patchy. Only by allowing individual bacteria to detach from developing clusters and re-attach elsewhere to start a new cluster, did PHYLLOSIM come much closer to reproducing experimental observations. The goodness of fit including detachment increased to about 70-80% for all waterscapes. Predictions of this 'detachment' model were further supported by the visualization and quantification of bacterial detachment and attachment events at an agarose-water interface. Thus, both model and experiment suggest that detachment of bacterial cells from clusters is an important mechanism underlying bacterial exploration of the phyllosphere.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0075633PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3790818PMC
July 2014

Cyst expansion and regression in a mouse model of polycystic kidney disease.

Kidney Int 2013 Jun 6;83(6):1099-108. Epub 2013 Mar 6.

Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands.

Autosomal-dominant polycystic kidney disease is characterized by progressive cyst formation and fibrosis in the kidneys. Here we describe an orthologous Pkd1(nl,nl) mouse model, with reduced expression of the normal Pkd1 transcript, on a fixed genetic background of equal parts C57Bl/6 and 129Ola/Hsd mice (B6Ola-Pkd1(nl,nl)). In these mice, the first cysts develop from mature proximal tubules around birth. Subsequently, larger cysts become visible at day 7, followed by distal tubule and collecting duct cyst formation, and progressive cystic enlargement to develop into large cystic kidneys within 4 weeks. Interestingly, cyst expansion was followed by renal volume regression due to cyst collapse. This was accompanied by focal formation of fibrotic areas, an increased expression of genes involved in matrix remodeling and subsequently an increase in infiltrating immune cells. After an initial increase in blood urea within the first 4 weeks, renal function remained stable over time and the mice were able to survive up to a year. Also, in kidneys of ADPKD patients collapsed cysts were observed, in addition to massive fibrosis and immune infiltrates. Thus, B6Ola-Pkd1(nl,nl) mice show regression of cysts and renal volume that is not accompanied by a reduction in blood urea levels.
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http://dx.doi.org/10.1038/ki.2013.13DOI Listing
June 2013

Accelerated antibody-mediated graft loss of rodent pancreatic islets after pretreatment with dexamethasone-treated immature donor dendritic cells.

Transplantation 2012 Nov;94(9):903-10

Department of Pathology, Leiden University Medical Center, Leiden, the Netherlands.

Background: Allogeneic islets of Langerhans transplantation is hampered in its success as a curative treatment of type 1 diabetes by the absence of potent, specific, and nontoxic immunosuppressive drugs. Here, we assessed whether donor bone marrow-derived dexamethasone-treated dendritic cells (dexDCs) could prolong islet allograft survival in a full major histocompatibility complex mismatch rat model.

Methods: Rodent allogeneic islet transplantation was performed from DA rats to Lewis rats and vice versa. Permanently immature dendritic cells were generated from the bone marrow of DA and Lewis rats by treatment with dexamethasone. Animals were either vehicle or donor dexDCs pretreated. Serum was used to monitor glucose, C-peptide, and alloreactive antibodies.

Results: The transplantation of DA islets into Lewis recipients showed direct graft failure with reduced numbers of β-cells when rats were pretreated with donor dexDCs. In the reverse model (Lewis islets into DA recipients), dexDC-treated DA recipients even showed a significantly accelerated rejection of Lewis islets. Immunohistochemical analysis of allograft tissue of dexDC-treated recipients showed a predominant natural killer cell infiltration and a presence of antibody reactivity in the absence of complement deposition. Alloreactive antibodies were solely found in dexDC-treated recipients.

Conclusion: Our study shows that pretreatment with donor-derived dexDCs induces an antibody-mediated rejection in this islet transplantation rodent model.
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http://dx.doi.org/10.1097/TP.0b013e31826acd01DOI Listing
November 2012

A thready affair: linking fungal diversity and community dynamics to terrestrial decomposition processes.

FEMS Microbiol Rev 2013 Jul 10;37(4):477-94. Epub 2012 Oct 10.

Department of Microbial Ecology, Netherlands Institute of Ecology, Wageningen, The Netherlands.

Filamentous fungi are critical to the decomposition of terrestrial organic matter and, consequently, in the global carbon cycle. In particular, their contribution to degradation of recalcitrant lignocellulose complexes has been widely studied. In this review, we focus on the functioning of terrestrial fungal decomposers and examine the factors that affect their activities and community dynamics. In relation to this, impacts of global warming and increased N deposition are discussed. We also address the contribution of fungal decomposer studies to the development of general community ecological concepts such as diversity-functioning relationships, succession, priority effects and home-field advantage. Finally, we indicate several research directions that will lead to a more complete understanding of the ecological roles of terrestrial decomposer fungi such as their importance in turnover of rhizodeposits, the consequences of interactions with other organisms and niche differentiation.
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http://dx.doi.org/10.1111/1574-6976.12001DOI Listing
July 2013

Controls on coarse wood decay in temperate tree species: birth of the LOGLIFE experiment.

Ambio 2012 ;41 Suppl 3:231-45

Department of Ecological Science, VU University, Amsterdam, The Netherlands.

Dead wood provides a huge terrestrial carbon stock and a habitat to wide-ranging organisms during its decay. Our brief review highlights that, in order to understand environmental change impacts on these functions, we need to quantify the contributions of different interacting biotic and abiotic drivers to wood decomposition. LOGLIFE is a new long-term 'common-garden' experiment to disentangle the effects of species' wood traits and site-related environmental drivers on wood decomposition dynamics and its associated diversity of microbial and invertebrate communities. This experiment is firmly rooted in pioneering experiments under the directorship of Terry Callaghan at Abisko Research Station, Sweden. LOGLIFE features two contrasting forest sites in the Netherlands, each hosting a similar set of coarse logs and branches of 10 tree species. LOGLIFE welcomes other researchers to test further questions concerning coarse wood decay that will also help to optimise forest management in view of carbon sequestration and biodiversity conservation.
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http://dx.doi.org/10.1007/s13280-012-0304-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3535053PMC
October 2012

Dose-dependent effects of sirolimus on mTOR signaling and polycystic kidney disease.

J Am Soc Nephrol 2012 May 16;23(5):842-53. Epub 2012 Feb 16.

Department of Human and Clinical Genetics, Leiden University Medical Center, The Netherlands.

Inhibition of the mammalian target of rapamycin (mTOR) shows beneficial effects in animal models of polycystic kidney disease (PKD); however, two clinical trials in patients with autosomal dominant PKD failed to demonstrate a short-term benefit in either the early or progressive stages of disease. The stage of disease during treatment and the dose of mTOR inhibitors may account for these differing results. Here, we studied the effects of a conventional low dose and a higher dose of sirolimus (blood levels of 3 ng/ml and 30-60 ng/ml, respectively) on mTOR activity and renal cystic disease in two Pkd1-mutant mouse models at different stages of the disease. When initiated at early but not late stages of disease, high-dose treatment strongly reduced mTOR signaling in renal tissues, inhibited cystogenesis, accelerated cyst regression, and abrogated fibrosis and the infiltration of immune cells. In contrast, low-dose treatment did not significantly reduce renal cystic disease. Levels of p-S6Rp(Ser240/244), which marks mTOR activity, varied between kidneys; severity of the renal cystic phenotype correlated with the level of mTOR activity. Taken together, these data suggest that long-term treatment with conventional doses of sirolimus is insufficient to inhibit mTOR activity in renal cystic tissue. Mechanisms to increase bioavailability or to target mTOR inhibitors more specifically to kidneys, alone or in combination with other compounds, may improve the potential for these therapies in PKD.
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http://dx.doi.org/10.1681/ASN.2011040340DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3338286PMC
May 2012

No apparent costs for facultative antibiotic production by the soil bacterium Pseudomonas fluorescens Pf0-1.

PLoS One 2011 16;6(11):e27266. Epub 2011 Nov 16.

Department of Microbial Ecology, Netherlands Institute of Ecology (NIOO-KNAW), Wageningen, The Netherlands.

Background: Many soil-inhabiting bacteria are known to produce secondary metabolites that can suppress microorganisms competing for the same resources. The production of antimicrobial compounds is expected to incur fitness costs for the producing bacteria. Such costs form the basis for models on the co-existence of antibiotic-producing and non-antibiotic producing strains. However, so far studies quantifying the costs of antibiotic production by bacteria are scarce. The current study reports on possible costs, for antibiotic production by Pseudomonas fluorescens Pf0-1, a soil bacterium that is induced to produce a broad-spectrum antibiotic when it is confronted with non-related bacterial competitors or supernatants of their cultures.

Methodology And Principal Findings: We measured the possible cost of antibiotic production for Pseudomonas fluorescens Pf0-1 by monitoring changes in growth rate with and without induction of antibiotic production by supernatant of a bacterial competitor, namely Pedobacter sp.. Experiments were performed in liquid as well as on semi-solid media under nutrient-limited conditions that are expected to most clearly reveal fitness costs. Our results did not reveal any significant costs for production of antibiotics by Pseudomonas fluorescens Pf0-1. Comparison of growth rates of the antibiotic-producing wild-type cells with those of non-antibiotic producing mutants did not reveal costs of antibiotic production either.

Significance: Based on our findings we propose that the facultative production of antibiotics might not be selected to mitigate metabolic costs, but instead might be advantageous because it limits the risk of competitors evolving resistance, or even the risk of competitors feeding on the compounds produced.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0027266PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3217935PMC
March 2012

Elimination of severe albuminuria in aging hypertensive rats by exchange of 2 chromosomes in double-consomic rats.

Hypertension 2011 Aug 31;58(2):219-24. Epub 2011 May 31.

Department of Clinical Pharmacology and Toxicology, Charité-Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany.

The inherited nephron deficit and progressive albuminuria development observed in hypertensive Munich Wistar Frömter (MWF) rats are influenced by quantitative trait loci on rat chromosome (RNO) 6 and RNO8. Previous studies in young MWF rats suggested that the nephron deficit represents a cause for glomerular hypertrophy preceding onset of albuminuria at 8 weeks and demonstrated a simultaneous induction of the podocyte stress marker desmin and podoplanin loss in podocytes. Here we investigated the separate genetic influence of RNO6 and RNO8 on early glomerular changes and subsequent albuminuria in single-consomic MWF rats in which RNO6 (MWF-6(SHR)) and RNO8 (MWF-8(SHR)) were replaced by the respective spontaneously hypertensive rat (SHR) chromosome. Furthermore, we tested the role of synergistic effects between both chromosomes in a double-consomic MWF-6(SHR)8(SHR) strain. Increased glomerular, extramesangial desmin expressions at 6 and albuminuria at 8 weeks were significantly reduced in single- and double-consomics (P<0.05 versus MWF, respectively). MWF-6(SHR)8(SHR) rats demonstrated the lowest desmin expression and glomerular volume (P<0.05 versus MWF, MWF-6(SHR), and MWF-8(SHR), respectively), indicating synergistic effects between RNO6 and RNO8. A significant and similar loss of podoplanin was only seen in MWF and MWF-6(SHR) rats but not in MWF-8(SHR) and MWF-6(SHR)8(SHR) rats (P<0.02, respectively); this refutes a mandatory coupling of desmin induction and podoplanin loss in podocytes preceding albuminuria and reveals a genetic link between RNO8 and loss of podoplanin protein. Long-term follow up in MWF-6(SHR)8(SHR) rats demonstrates the relevance of the absence of glomerular changes in young animals, because double-consomics demonstrate a complete suppression of progressive albuminuria and kidney damage compared with MWF rats despite similar blood pressures.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.111.170621DOI Listing
August 2011

Altered Hippo signalling in polycystic kidney disease.

J Pathol 2011 May 7;224(1):133-42. Epub 2011 Mar 7.

Department of Human Genetics, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

Autosomal dominant polycystic kidney disease (ADPKD) is characterized by progressive deterioration of renal function and formation of cysts, and is an important cause of end-stage renal disease. Previously we showed that tubular epithelial injury accelerates cyst formation in inducible Pkd1-deletion mice. In these mice, expression of the planar cell polarity (PCP) component Four-jointed (Fjx1) is decreased during epithelial repair, while in control mice Fjx1 expression is increased and may be required during tissue regeneration. In cystic kidneys, however, Fjx1 expression is also increased. Besides a PCP component, Four-jointed is also implicated in the Hippo-signalling pathway. This pathway is involved in organ size control by regulating proliferation and apoptosis. The role of Hippo signalling, together with the opposing expression pattern of Fjx1 during epithelial repair and at cystic stages, triggered us to investigate the activity of the Hippo pathway during these processes. Therefore, we examined its final effector molecule, the transcriptional co-activator Yes-associated protein (YAP) and observed that during tissue repair, YAP expression was not different between Pkd1-deletion mice and controls, ie during tissue regeneration YAP expression was increased and predominantly localized in the cytoplasm but normalized after tissue repair. At a later stage, however, in cystic epithelia and epithelia of dilated tubules, strong nuclear YAP accumulation was observed, accompanied by up-regulation of the YAP transcriptional targets Birc-3, Ctgf, InhbA, and Fjx1. Altered activity of the Hippo pathway was confirmed in renal tissues from human ADPKD and ARPKD patients, as well as in cystic renal tumours. Our data strengthen the concept that during epithelial repair Four-jointed is involved in PCP signalling, while in cystic kidneys it is related to Hippo signalling and cyst growth.
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http://dx.doi.org/10.1002/path.2856DOI Listing
May 2011

Curcumin inhibits cystogenesis by simultaneous interference of multiple signaling pathways: in vivo evidence from a Pkd1-deletion model.

Am J Physiol Renal Physiol 2011 May 23;300(5):F1193-202. Epub 2011 Feb 23.

Department of Human Genetics, Leiden Univ. Medical Center, The Netherlands.

Autosomal dominant polycystic kidney disease (ADPKD) caused by mutations in either the PKD1 or PKD2 gene is a major cause of end-stage renal failure. A number of compounds targeting specific signaling pathways were able to inhibit cystogenesis in rodent models and are currently being tested in clinical trials. However, given the complex signaling in ADPKD, an ideal therapy would likely have to comprise several pathways at once. Therefore, multitarget compounds may provide promising therapeutic interventions for the treatment of ADPKD. To test this hypothesis, we treated Pkd1-deletion mice with diferuloylmethane (curcumin), a compound without appreciable side effects and known to modulate several pathways that are also altered in ADPKD, e.g., mammalian target of rapamycin (mTOR) and Wnt signaling. After conditional inactivation of Pkd1, mTOR signaling was indeed elevated in cystic kidneys. Interestingly, also activation of signal transducers and activator of transcription 3 (STAT3) strongly correlated with cyst progression. Both pathways were effectively inhibited in vitro by curcumin. Importantly, Pkd1-deletion mice that were treated with curcumin and killed at an early stage of PKD displayed improved renal histology and reduced STAT3 activation, proliferation index, cystic index, and kidney weight/body weight ratios. In addition, renal failure was significantly postponed in mice with severe PKD. These data suggest that multitarget compounds hold promising potential for safe and effective treatment of ADPKD.
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http://dx.doi.org/10.1152/ajprenal.00419.2010DOI Listing
May 2011

Modelling sugar diffusion across plant leaf cuticles: the effect of free water on substrate availability to phyllosphere bacteria.

Environ Microbiol 2011 Mar 22;13(3):792-7. Epub 2010 Nov 22.

Department of Microbial Ecology, Netherlands Institute of Ecology (NIOO-KNAW), Heteren, The Netherlands.

We present a continuous model for the diffusion of sugars across intact plant leaf cuticles. It is based on the flow of sugars from a source, representing the leaf apoplast, to a sink, in the shape of a hemispherical drop of water on the outside of the cuticle. Flow is a function of the difference between sugar concentrations C(Source) and C(Sink) , permeability P of the cuticle, volume V(Sink) of the water drop, as well as its contact angle α with the cuticle surface. Using a bacterial bioreporter for fructose, and a two-compartment experimental set-up consisting of isolated cuticles of walnut (Juglans regia) carrying water droplets while floating on solutions with increasing concentrations of fructose, we determined a value of 1 × 10⁻⁶ m h⁻¹ for P. Using this value, we explored different scenarios for the leaching of sugars across plant leaf cuticles to reveal in quantitative terms how diffusion takes longer when V(Sink) increases, P decreases or α increases. Bacterial growth was modelled as a function of changes in P, α and V(Sink) and was consistent with observations or suggestions from the literature in relation to the availability of free water on leaves. These results are discussed in the light of bacteria as ecosystem engineers, i.e. with the ability to modify the plant leaf surface environment in favour of their own survival, e.g. by increasing cuticle leakage or leaf wetness. Our model represents a first step towards a more comprehensive model which will enhance our quantitative understanding of the factors that play a role in nutrient availability to bacterial colonizers of the phyllosphere, or plant leaf surface.
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http://dx.doi.org/10.1111/j.1462-2920.2010.02382.xDOI Listing
March 2011

Pkd1-inactivation in vascular smooth muscle cells and adaptation to hypertension.

Lab Invest 2011 Jan 20;91(1):24-32. Epub 2010 Sep 20.

Center for Human and Clinical Genetics, Leiden University Medical Center, Leiden, The Netherlands.

Autosomal dominant polycystic kidney disease (ADPKD) is a multisystem disorder characterized by renal, hepatic and pancreatic cyst formation and cardiovascular complications. The condition is caused by mutations in the PKD1 or PKD2 gene. In mice with reduced expression of Pkd1, dissecting aneurysms with prominent media thickening have been seen. To study the effect of selective disruption of Pkd1 in vascular smooth muscle cells (SMCs), we have generated mice in which a floxed part of the Pkd1 gene was deleted by Cre under the control of the SM22 promotor (SM22-Pkd1(del/del) mice). Cre activity was confirmed by X-gal staining using lacZ expressing Cre reporter mice (R26R), and quantitative PCR indicated that in the aorta Pkd1 gene expression was strongly reduced, whereas Pkd2 levels remained unaltered. Histopathological analysis revealed cyst formation in pancreas, liver and kidneys as the result of extravascular Cre activity in pancreatic ducts, bile ducts and in the glomerular Bowman's capsule. Remarkably, we did not find any spontaneous gross structural blood vessel abnormalities in mice with somatic Pkd1 gene disruption in SMCs or simultaneous disruption of Pkd1 in SMCs and endothelial cells (ECs). Extensive isometric myographic analysis of the aorta did not reveal differences in response to KCl, acetylcholine, phenylephrin or serotonin, except for a significant increase in contractility induced by phenylephrin on arteries from 40 weeks old Pkd1(del/+) germ-line mice. However, SM22-Pkd1(del/del) mice showed significantly reduced decrease in heart rate on angiotensin II-induced hypertension. The present findings further demonstrate in vivo, that adaptation to hypertension is altered in SM22-Pkd1(del/del) mice.
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http://dx.doi.org/10.1038/labinvest.2010.159DOI Listing
January 2011

Elevated TGFbeta-Smad signalling in experimental Pkd1 models and human patients with polycystic kidney disease.

J Pathol 2010 Sep;222(1):21-31

Centre for Human and Clinical Genetics, Leiden University Medical Centre, Leiden, The Netherlands.

Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited renal disease characterized by many fluid-filled cysts and interstitial fibrosis in the kidneys, leading to chronic renal failure. During cystogenesis the renal tubules undergo extensive structural alterations that are accompanied by altered cellular signalling, directly and/or indirectly regulated by the PKD1 and PKD2 proteins. Since transforming growth factor (TGF)-beta signalling modulates cell proliferation, differentiation, apoptosis, adhesion and migration of various cell types, we studied the activation of this signalling pathway in Pkd1-mutant mouse models at different stages of the disease. Therefore, we analysed expression of the TGFbeta-Smad signalling pathway and its target genes in different Pkd1 mutant mouse models in various stages of polycystic disease. Nuclear accumulation of P-Smad2 in cyst lining epithelial cells was not observed in the initiation phase but was observed at mild and more advanced stages of PKD. This coincides with mild fibrosis and increased mRNA levels of TGFbeta target genes, such as fibronectin, collagen type I, plasminogen activator inhibitor 1 and matrix metalloproteinase-2. At this stage many interstitial fibroblasts were found around cysts, which also showed nuclear localization for P-Smad2. However, bone morphogenetic protein (BMP) signalling, which can antagonize TGFbeta signalling, is not affected, since nuclear expression of P-Smad1/5/8 and expression of the BMP target gene, inhibitor of DNA binding/differential-1 (ID-1) is not altered compared to wild-type controls. Also, human kidneys with progressive ADPKD showed increased nuclear localization of P-Smad2, while in general expression of P-Smad1/5/8 was weak. These results exclude TGFbeta signalling at the initiation of cystogenesis, but indicate an important role during cyst progression and in fibrogenesis of progressive ADPKD.
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http://dx.doi.org/10.1002/path.2734DOI Listing
September 2010

Annexin A1 regulates TGF-beta signaling and promotes metastasis formation of basal-like breast cancer cells.

Proc Natl Acad Sci U S A 2010 Apr 22;107(14):6340-5. Epub 2010 Mar 22.

Division of Toxicology, Leiden/Amsterdam Center for Drug Research, Leiden University, 2300 RA, Leiden, The Netherlands.

Annexin A1 (AnxA1) is a candidate regulator of the epithelial- to mesenchymal (EMT)-like phenotypic switch, a pivotal event in breast cancer progression. We show here that AnxA1 expression is associated with a highly invasive basal-like breast cancer subtype both in a panel of human breast cancer cell lines as in breast cancer patients and that AnxA1 is functionally related to breast cancer progression. AnxA1 knockdown in invasive basal-like breast cancer cells reduced the number of spontaneous lung metastasis, whereas additional expression of AnxA1 enhanced metastatic spread. AnxA1 promotes metastasis formation by enhancing TGFbeta/Smad signaling and actin reorganization, which facilitates an EMT-like switch, thereby allowing efficient cell migration and invasion of metastatic breast cancer cells.
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http://dx.doi.org/10.1073/pnas.0913360107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2852023PMC
April 2010

Sdhd and SDHD/H19 knockout mice do not develop paraganglioma or pheochromocytoma.

PLoS One 2009 Nov 24;4(11):e7987. Epub 2009 Nov 24.

Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands.

Background: Mitochondrial succinate dehydrogenase (SDH) is a component of both the tricarboxylic acid cycle and the electron transport chain. Mutations of SDHD, the first protein of intermediary metabolism shown to be involved in tumorigenesis, lead to the human tumors paraganglioma (PGL) and pheochromocytoma (PC). SDHD is remarkable in showing an 'imprinted' tumor suppressor phenotype. Mutations of SDHD show a very high penetrance in man and we postulated that knockout of Sdhd would lead to the development of PGL/PC, probably in aged mice.

Methodology/principal Findings: We generated a conventional knockout of Sdhd in the mouse, removing the entire third exon. We also crossed this mouse with a knockout of H19, a postulated imprinted modifier gene of Sdhd tumorigenesis, to evaluate if loss of these genes together would lead to the initiation or enhancement of tumor development. Homozygous knockout of Sdhd results in embryonic lethality. No paraganglioma or other tumor development was seen in Sdhd KO mice followed for their entire lifespan, in sharp contrast to the highly penetrant phenotype in humans. Heterozygous Sdhd KO mice did not show hyperplasia of paraganglioma-related tissues such as the carotid body or of the adrenal medulla, or any genotype-related pathology, with similar body and organ weights to wildtype mice. A cohort of Sdhd/H19 KO mice developed several cases of profound cardiac hypertrophy, but showed no evidence of PGL/PC.

Conclusions: Knockout of Sdhd in the mouse does not result in a disease phenotype. H19 may not be an initiator of PGL/PC tumorigenesis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0007987PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2776493PMC
November 2009

Toxic tubular injury in kidneys from Pkd1-deletion mice accelerates cystogenesis accompanied by dysregulated planar cell polarity and canonical Wnt signaling pathways.

Hum Mol Genet 2009 Jul 28;18(14):2532-42. Epub 2009 Apr 28.

Department of Human Genetics, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

Autosomal dominant polycystic kidney disease (ADPKD) is characterized by large fluid-filled cysts and progressive deterioration of renal function necessitating renal replacement therapy. Previously, we generated a tamoxifen-inducible, kidney epithelium-specific Pkd1-deletion mouse model and showed that inactivation of the Pkd1 gene induces rapid cyst formation in developing kidneys and a slow onset of disease in adult mice. Therefore, we hypothesized that injury-induced tubular epithelial cell proliferation may accelerate cyst formation in the kidneys of adult Pkd1-deletion mice. Mice were treated with the nephrotoxicant 1,2-dichlorovinyl-cysteine (DCVC) after Pkd1-gene inactivation, which indeed accelerated cyst formation significantly. After the increased proliferation during tissue regeneration, proliferation decreased to basal levels in Pkd1-deletion mice just as in DCVC-treated controls. However, in severe cystic kidneys, 10-14 weeks after injury, proliferation increased again. This biphasic response suggests that unrestricted cell proliferation after injury is not the underlying mechanism for cyst formation. Aberrant planar cell polarity (PCP) signaling and increased canonical Wnt signaling are suggested to be involved in cyst formation. Indeed, we show here that in Pkd1 conditional deletion mice expression of the PCP component Four-jointed (Fjx1) is decreased while its expression is required during tissue regeneration. In addition, we show that altered centrosome position and the activation of canonical Wnt signaling are early effects of Pkd1-gene disruption. This suggests that additional stimuli or events are required to trigger the process of cyst formation. We propose that during tissue repair, the integrity of the newly formed Pkd1-deficient cells is modified rendering them susceptible to subsequent cyst formation.
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http://dx.doi.org/10.1093/hmg/ddp190DOI Listing
July 2009

Genetic predisposition for glomerulonephritis-induced glomerulosclerosis in rats is linked to chromosome 1.

Physiol Genomics 2008 Oct 5;35(2):173-81. Epub 2008 Aug 5.

Pathology, Leiden University Medical Center, Leiden, The Netherlands.

Genetic factors influence renal disease progression, and several loci have been linked to the spontaneous development of proteinuria and glomerulosclerosis in animal models. However, the role of genetic susceptibility in glomerulonephritis-induced progressive glomerulosclerosis is unknown. In a rat model of mesangial proliferative glomerulonephritis, anti-Thy-1 glomerulonephritis (antiThy1GN), Lewis/Maastricht (Lew/Maa) rats exhibit progression to glomerulosclerosis, whereas in genetically related Lewis/Møllegard (Lew/Moll) rats, glomerular lesions are repaired within 3 wk. The genetic factors underlying this strain-related difference are not known. To identify novel quantitative trait loci (QTL) involved in progression or repair in Lewis rats, 145 female backcross rats [F1(Lew/Maa x Lew/Moll) x Lew/Maa] were studied. After induction of antiThy1GN proteinuria, we determined mesangial activation, the percentage of microaneurysms, and the glomerular damage score for each animal; a genome scan using 187 microsatellite markers was performed. QTL mapping revealed a significant QTL for glomerular damage score on chromosome 1 with a logarithm of odds (LOD) score of 3.9. Homozygosity for Lew/Maa DNA in this region was associated with a higher percentage of damaged glomeruli on day 21. Furthermore, suggestive linkage was found for the percentage of glomeruli with microaneurysms on day 3 on chromosome 1, 6, and 11; for mesangial activation on day 7 on chromosome 18, while proteinuria was suggestively linked to chromosome 5 (day 0), 4 (day 3), and 6 (day 7). This study identifies a QTL on rat chromosome 1 that is significantly linked to progressive glomerulosclerosis after acute glomerulonephritis.
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http://dx.doi.org/10.1152/physiolgenomics.00268.2007DOI Listing
October 2008

Kidney-specific inactivation of the Pkd1 gene induces rapid cyst formation in developing kidneys and a slow onset of disease in adult mice.

Hum Mol Genet 2007 Dec 11;16(24):3188-96. Epub 2007 Oct 11.

Department of Human Genetics, Leiden University Medical Center, Leiden 2300RC, The Netherlands.

Autosomal dominant polycystic kidney disease, caused by mutations in the PKD1 gene, is characterized by progressive deterioration of kidney function due to the formation of thousands of cysts leading to kidney failure in mid-life or later. How cysts develop and grow is currently unknown, although extensive research revealed a plethora of cellular changes in cyst lining cells. We have constructed a tamoxifen-inducible, kidney epithelium-specific Pkd1-deletion mouse model. Upon administration of tamoxifen to these mice, a genomic fragment containing exons 2-11 of the Pkd1-gene is specifically deleted in the kidneys and cysts are formed. Interestingly, the timing of Pkd1-deletion has strong effects on the phenotype. At 1 month upon gene disruption, adult mice develop only a very mild cystic phenotype showing some small cysts and dilated tubules. Young mice, however, show massive cyst formation. In these mice, at the moment of gene disruption, cell proliferation takes place to elongate the nephron. Our data indicate that Pkd1 gene deficiency does not initiate sufficient autonomous cell proliferation leading to cyst formation and that additional stimuli are required. Furthermore, we show that one germ-line mutation of Pkd1 is already associated with increased proliferation.
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http://dx.doi.org/10.1093/hmg/ddm299DOI Listing
December 2007

Identification of a human CD8+ regulatory T cell subset that mediates suppression through the chemokine CC chemokine ligand 4.

Proc Natl Acad Sci U S A 2007 May 2;104(19):8029-34. Epub 2007 May 2.

Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, 2333 ZA, Leiden, The Netherlands.

Regulatory T cells (Treg) comprise multiple subsets and are important in controlling immunity and inflammation. However, the induction and mode of action of the various distinct Treg subsets remain ill defined, particularly in humans. Here, we describe a human CD8+ lymphocyte activation gene-3 (LAG-3)+CD25+FoxP3+ Treg subset, which suppresses T cells partly through the secretion of CC chemokine ligand 4 (CCL4), which can inhibit T cell activation by interfering with T cell receptor signaling. CD8+ Tregs are expanded by antigen in in vivo-primed donors, and can be detected in pathogen-infected human tissue. This CD8+LAG-3+CD25+FoxP3+CCL4+ Treg subset thus may play a role in immunoregulation in humans, including infectious diseases.
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http://dx.doi.org/10.1073/pnas.0702257104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1876566PMC
May 2007

Chimerism occurs twice as often in lupus nephritis as in normal kidneys.

Arthritis Rheum 2006 Sep;54(9):2944-50

Leiden University Medical Center, Leiden, The Netherlands.

Objective: Systemic lupus erythematosus (SLE) is an immune-mediated disease that particularly affects the kidneys, causing lupus nephritis. In experimental mouse models, lupus nephritis can be mimicked by inducing a chimeric state through the injection of parental T cells in offspring. In humans, pregnancy-induced chimerism may play a role in the pathogenesis of autoimmune diseases such as SLE, but it is likely that only certain chimeric cells have pathogenic potential. In this study, we investigated whether the distribution of chimeric cells is different in the kidneys of women with SLE from that in normal kidneys, and we examined the phenotype of chimeric cells in women with SLE.

Methods: The presence of chimeric cells was investigated by in situ hybridization targeting the Y chromosome in 57 renal biopsy samples from 49 women with lupus nephritis. Fifty-one kidney autopsy specimens without histomorphologic lesions served as controls. Double-staining for the Y chromosome in combination with CD3 and CD34 markers was performed in 5 kidney specimens with lupus nephritis to identify the phenotype of the chimeric cells.

Results: Y chromosome-positive cells were found in 27 of 49 patients with lupus nephritis and in 13 of 51 normal controls (P < 0.01). Both CD3+ and CD34+ chimeric cells were identified in lupus nephritis kidney specimens.

Conclusion: Chimeric cells are present significantly more often in kidneys with lupus nephritis than in normal kidneys, and some of these chimeric cells are T cells. This finding is interesting in light of experimental models demonstrating that lupus nephritis is initiated by chimeric T cells.
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http://dx.doi.org/10.1002/art.22038DOI Listing
September 2006

Increased activity of activator protein-1 transcription factor components ATF2, c-Jun, and c-Fos in human and mouse autosomal dominant polycystic kidney disease.

J Am Soc Nephrol 2005 Sep 27;16(9):2724-31. Epub 2005 Jul 27.

Department of Human Genetics, Leiden University Medical Center, Leiden, The Netherlands.

Autosomal dominant polycystic kidney disease is a common inherited disorder that predominantly manifests with the formation of fluid-filled cysts in both kidneys. The disease can be accounted for by a mutation in either the PKD1 or the PKD2 gene. It was demonstrated previously that aberrant expression of the PKD1 gene product, polycystin-1, results in modification of activator protein-1 (AP-1) transcription factor activity in cultured renal epithelial cells. Here, it is reported that activity of the AP-1 components c-Jun, ATF2, and c-Fos is altered in renal cystic tissue of patients with autosomal dominant polycystic kidney disease and of hypomorphic Pkd1 mice with polycystic kidney disease. Data were obtained using immunohistochemical and Western blot analysis. Significant upregulation of Thr71- and Thr69/71-phosphorylated ATF2 and Ser73-phosphorylated c-Jun and increased c-Fos were detected in small cysts and (dilated) ducts and tubules surrounded by fibrotic interstitium. The data indicate that various AP-1 components are constitutively activated in polycystic kidney disease and suggest that aberrant AP-1 activity is relevant for cyst formation.
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http://dx.doi.org/10.1681/ASN.2004110913DOI Listing
September 2005

The absence of delayed graft function is predicted by the presence of manganese-superoxide dismutase in distal tubules of renal allografts.

Transplantation 2005 Apr;79(8):946-52

Department of Nephrology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands.

Background: Acute tubular necrosis (ATN) in renal allograft biopsies correlates poorly with delayed graft function (DGF). Factors involved in the pathogenesis of DGF were evaluated in biopsies in an attempt to refine the recognition of DGF.

Methods: Anti-cubulin and anti-AE-1/AE-3 antibodies identified proximal and distal tubules, respectively. The terminal deoxynucleotide transferase-mediated dUTP nick-end labeling technique and active caspase-3 staining were used to demonstrate apoptosis. Antibodies against superoxide dismutase (SOD) were used as markers of the protective tubular response. Tubular regeneration was evaluated using anti-ki 67 and antivimentin antibodies.

Results: Of a total of 40 biopsies, 9 were associated with DGF. ATN was seen in 16 biopsies; 5 were associated with DGF. The finding of ATN in the biopsy of a graft predicted DGF in only 56% of cases. Absence of distal caspase-3 staining predicted the absence of ATN in 87% of cases. The presence of caspase-3 predicted ATN in 54% of cases. The detection of manganese-SOD in distal tubules predicts the absence of DGF in 76% of the cases.

Conclusions: The use of immunohistochemical staining on posttransplant renal biopsies improved its predictive value with respect to ATN and DGF: The absence of active caspase-3 in distal tubular epithelium predicts the absence of ATN in 87% of cases, whereas its presence predicts ATN in 54% of cases. The presence of manganese-SOD in distal tubules predicts the absence of DGF in 76% of cases.
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http://dx.doi.org/10.1097/01.tp.0000156166.60218.c7DOI Listing
April 2005