The heavy subunit of ferritin stimulates NLRP3 inflammasomes in hepatic stellate cells through ICAM-1 to drive hepatic inflammation.

Serum ferritin concentrations increase during hepatic inflammation and correlate with the severity of chronic liver disease. Here, we report a molecular mechanism whereby the heavy subunit of ferritin (FTH) contributes to hepatic inflammation. We found that FTH induced activation of the NLRP3 inflammasome and secretion of the proinflammatory cytokine interleukin-1ß (IL-1ß) in primary rat hepatic stellate cells (HSCs) through intercellular adhesion molecule-1 (ICAM-1). FTH-ICAM-1 stimulated the expression of Il1b, NLRP3 inflammasome activation, and the processing and secretion of IL-1ß in a manner that depended on plasma membrane remodeling, clathrin-mediated endocytosis, and lysosomal destabilization. FTH-ICAM-1 signaling at early endosomes stimulated Il1b expression, implying that this endosomal signaling primed inflammasome activation in HSCs. In contrast, lysosomal destabilization was required for FTH-induced IL-1ß secretion, suggesting that lysosomal damage activated inflammasomes. FTH induced IL-1ß production in liver slices from wild-type mice but not in those from Icam1-/- or Nlrp3-/- mice. Thus, FTH signals through its receptor ICAM-1 on HSCs to activate the NLRP3 inflammasome. We speculate that this pathway contributes to hepatic inflammation, a key process that stimulates hepatic fibrogenesis associated with chronic liver disease.

Fine-tuning the standard setting of objective structured practical examinations in clinical anatomy.

The objective structured practical examination (OSPE) is a reliable assessment of practical skills in anatomy teaching. It is often administered as low-stake assessments to track progress at multiple time points in anatomy curricula. Standard-setting OSPEs to derive a pass mark and to ensure assessment quality and rigor is a complex task. This study compared standard-setting outcomes of clinical anatomy OSPEs determined by traditional criterion-referenced (Ebel) and norm-referenced ("mean minus standard deviation") methods in comparison to hybrid methods which apply both criterion-referenced and norm-referenced approaches in setting examination standards. The hybrid approaches utilized included the "Cohen method" and an adaptation of the "Taylor's method," which is an improvement on the Cohen method. These diverse standard-setting methods were applied retrospectively to 16 anatomy OSPEs conducted over 4 years for first- and second-year medical students in a graduate Doctor of Medicine Program at Griffith Medical School, Australia; and the pass marks, failure rates, and variances of failure rates were compared. The application of the adaptation of Taylor's method to standard set OSPEs produced pass marks and failure rates comparable to the Ebel method, whereas the variability of failure rates was higher with the Ebel method than with the Cohen and Taylor's methods. This underscores this study's adaptation of Taylor's method as a suitable alternative to the widely accepted but resource intensive, panel-based criterion-referenced standard-setting methods such as the Ebel method, where panelists with relevant expertise are unavailable, particularly for the multiple low-stakes OSPEs in an anatomy curriculum.

Adult schistosomes have an epithelial bacterial population distinct from the surrounding mammalian host blood.

BACKGROUND: Schistosomiasis is a neglected tropical parasitic and chronic disease affecting hundreds of millions of people. Adult schistosomes reside in the blood stream of the definitive mammalian host. These helminth parasites possess two epithelial surfaces, the tegument and the gastrodermis, both of which interact with the host during immune evasion and in nutrient uptake. METHODS: Female ARC Swiss mice (4-6 weeks old) were infected percutaneously with Schistosoma japonicum cercariae freshly shed from Oncomelania hupensis quadrasi snails (Philippines strain). Fluorescent in situ hybridisation (FISH) was performed by using fresh adult S. japonicum perfused from those infected mice. Adult S. japonicum worms were processed to isolate the tegument from the carcass containing the gastrodermis; blood and bile were collected individually from infected and uninfected mice. Total DNA extracted from all those samples were used for microbiome profiling. RESULTS: FISH and microbiome profiling showed the presence of bacterial populations on two epithelial surfaces of adult worms, suggesting they were distinct not only from the host blood but also from each other. Whereas microbial diversity was reduced overall in the parasite epithelial tissues when compared with that of host blood, specific bacterial taxa, including Anoxybacillus and Escherichia, were elevated on the tegument. Minimal differences were evident in the microbiome of host blood during an active infection, compared with that of control uninfected blood. However, sampling of bile from infected animals identified some differences compared with controls, including elevated levels of Limnohabitans, Clostridium and Curvibacter. CONCLUSIONS: Using FISH and microbial profiling, we were able to demonstrate, for the first time, that bacteria are presented on the epithelial surfaces of adult schistosomes. These schistosome surface-associated bacteria, which are distinct from the host blood microenvironment, should be considered as a new and important component of the host-schistosome interaction. The importance of individual bacterial species in relation to schistosome parasitism needs further elucidation.

Use of kinase inhibitors against schistosomes to improve and broaden praziquantel efficacy.

Praziquantel (PZQ) is the drug of choice for schistosomiasis. The potential drug resistance necessitates the search for adjunct or alternative therapies to PZQ. Previous functional genomics has shown that RNAi inhibition of Ca2+/calmodulin-dependent protein kinase II (CaMKII) gene in Schistosoma adult worms significantly improved the effectiveness of PZQ. Here we tested the in vitro efficacy of 15 selective and non-selective CaMK inhibitors against Schistosoma mansoni and showed that PZQ efficacy was improved against refractory juvenile parasites when combined with these CaMK inhibitors. By measuring CaMK activity and the mobility of adult S. mansoni, we identified two non-selective CaMK inhibitors, Staurosporine (STSP) and 1Naphthyl PP1 (1NAPP1), as promising candidates for further study. The impact of STSP and 1NAPP1 was investigated in mice infected with S. mansoni in the presence or absence of a sub-lethal dose of PZQ against 2- and 7-day-old schistosomula and adults. Treatment with STSP/PZQ induced a significant (47-68%) liver egg burden reduction compared with mice treated with PZQ alone. The findings indicate that the combination of STSP and PZQ dosages significantly improved anti-schistosomal activity compared to PZQ alone, demonstrating the potential of selective and non-selective CaMK/kinase inhibitors as a combination therapy with PZQ in treating schistosomiasis.

Murine Precision-Cut Liver Slices as an Ex Vivo Model of Liver Biology.

Understanding the mechanisms of liver injury, hepatic fibrosis, and cirrhosis that underlie chronic liver diseases (i.e., viral hepatitis, non-alcoholic fatty liver disease, metabolic liver disease, and liver cancer) requires experimental manipulation of animal models and in vitro cell cultures. Both techniques have limitations, such as the requirement of large numbers of animals for in vivo manipulation. However, in vitro cell cultures do not reproduce the structure and function of the multicellular hepatic environment. The use of precision-cut liver slices is a technique in which uniform slices of viable mouse liver are maintained in laboratory tissue culture for experimental manipulation. This technique occupies an experimental niche that exists between animal studies and in vitro cell culture methods. The presented protocol describes a straightforward and reliable method to isolate and culture precision-cut liver slices from mice. As an application of this technique, ex vivo liver slices are treated with bile acids to simulate cholestatic liver injury and ultimately assess the mechanisms of hepatic fibrogenesis.

Gene Expression in Developmental Stages of Schistosoma japonicum Provides Further Insight into the Importance of the Schistosome Insulin-Like Peptide.

We showed previously that the Schistosoma japonicum insulin-like peptide (SjILP) binds the worm insulin receptors, thereby, activating the parasite's insulin pathway and emphasizing its important role in regulating uptake of glucose, a nutrient essential for parasite survival. Here we show that SjILP is differentially expressed in the schistosome life cycle and is especially highly transcribed in eggs, miracidia, and adult female worms. RNA inference was employed to knockdown SjILP in adults in vitro, with suppression confirmed by significantly reduced protein production, declined adenosine diphosphate levels, and reduction in glucose consumption. Immunolocalization showed that SjILP is located to lateral gland cells of mature intra-ovular miracidia in the schistosome egg, and is distributed on the ciliated epithelium and internal cell masses of newly transformed miracidia. In schistosomula, SjILP is present on the tegument in two antero-lateral points, indicating highly polarized expression during cercarial transformation. Analysis of serum from S. japonicum-infected mice by ELISA using a recombinant form of SjILP as an antigen revealed IgG immunoreactivity to this molecule at 7 weeks post-infection indicating it is likely secreted from mature eggs into the host circulation. These findings provide further insights on ILP function in schistosomes and its essential roles in parasite survival and growth in different development stages.

Whole-genome sequence of the bovine blood fluke Schistosoma bovis supports interspecific hybridization with S. haematobium.

Mesenteric infection by the parasitic blood fluke Schistosoma bovis is a common veterinary problem in Africa and the Middle East and occasionally in the Mediterranean Region. The species also has the ability to form interspecific hybrids with the human parasite S. haematobium with natural hybridisation observed in West Africa, presenting possible zoonotic transmission. Additionally, this exchange of alleles between species may dramatically influence disease dynamics and parasite evolution. We have generated a 374 Mb assembly of the S. bovis genome using Illumina and PacBio-based technologies. Despite infecting different hosts and organs, the genome sequences of S. bovis and S. haematobium appeared strikingly similar with 97% sequence identity. The two species share 98% of protein-coding genes, with an average sequence identity of 97.3% at the amino acid level. Genome comparison identified large continuous parts of the genome (up to several 100 kb) showing almost 100% sequence identity between S. bovis and S. haematobium. It is unlikely that this is a result of genome conservation and provides further evidence of natural interspecific hybridization between S. bovis and S. haematobium. Our results suggest that foreign DNA obtained by interspecific hybridization was maintained in the population through multiple meiosis cycles and that hybrids were sexually reproductive, producing viable offspring. The S. bovis genome assembly forms a highly valuable resource for studying schistosome evolution and exploring genetic regions that are associated with species-specific phenotypic traits.

Whole-genome sequence of the oriental lung fluke Paragonimus westermani.

Background: Foodborne infections caused by lung flukes of the genus Paragonimus are a significant and widespread public health problem in tropical areas. Approximately 50 Paragonimus species have been reported to infect animals and humans, but Paragonimus westermani is responsible for the bulk of human disease. Despite their medical and economic importance, no genome sequence for any Paragonimus species is available. Results: We sequenced and assembled the genome of P. westermani, which is among the largest of the known pathogen genomes with an estimated size of 1.1 Gb. A 922.8 Mb genome assembly was generated from Illumina and Pacific Biosciences (PacBio) sequence data, covering 84% of the estimated genome size. The genome has a high proportion (45%) of repeat-derived DNA, particularly of the long interspersed element and long terminal repeat subtypes, and the expansion of these elements may explain some of the large size. We predicted 12,852 protein coding genes, showing a high level of conservation with related trematode species. The majority of proteins (80%) had homologs in the human liver fluke Opisthorchis viverrini, with an average sequence identity of 64.1%. Assembly of the P. westermani mitochondrial genome from long PacBio reads resulted in a single high-quality circularized 20.6 kb contig. The contig harbored a 6.9 kb region of non-coding repetitive DNA comprised of three distinct repeat units. Our results suggest that the region is highly polymorphic in P. westermani, possibly even within single worm isolates. Conclusions: The generated assembly represents the first Paragonimus genome sequence and will facilitate future molecular studies of this important, but neglected, parasite group.

Live imaging of collagen deposition during experimental hepatic schistosomiasis and recovery: a view on a dynamic process.

Hepatic fibrosis is the central cause of chronic clinical pathology resulting from infection by the blood flukes Schistosoma japonicum or S. mansoni. Much has been elucidated regarding the molecular, cellular and immunological responses that correspond to the formation of the granulomatous response to trapped schistosome eggs. A central feature of this Th2 response is the deposition of collagen around the periphery of the granuloma. To date, traditional histology and transcriptional methods have been used to quantify the deposition of collagen and to monitor the formation of the hepatic granuloma during experimental animal models of schistosomiasis. We have investigated the dynamic nature of granuloma formation through the use of a transgenic mouse model (B6.Collagen 1(A) luciferase mice (B6.Coll 1A-luc+)). With this model and whole-animal bioluminescence imaging, we followed the deposition of collagen during an active schistosome infection with Chinese and Philippines geographical strains of S. japonicum and after clearance of the adult parasites by the drug praziquantel. Individual mice were re-imaged over the time course to provide robust real-time quantitation of the development of chronic fibrotic disease. This model provides an improved method to follow the course of hepatic schistosomiasis-induced hepatic pathology and effectively supports the current dogma of the formation of hepatic fibrosis, originally elucidated from static traditional histology. This study demonstrates the first use of the B6.Coll 1A-luc+ mouse to monitor the dynamics of disease development and the treatment of pathogen-induced infection with the underlying pathology of fibrosis.

Calcium and Ca2+/Calmodulin-dependent kinase II as targets for helminth parasite control.

In eukaryotes, effective calcium homeostasis is critical for many key biological processes. There is an added level of complexity in parasites, particularly multicellular helminth worms, which modulate calcium levels while inhabiting the host microenvironment. Parasites ensure efficient calcium homeostasis through gene products, such as the calmodulin-dependent kinases (CaMK), the main focus of this review. The importance of CaMK is becoming increasingly apparent from recent functional studies of helminth and protozoan parasites. Investigations on the molecular regulation of calcium and the role of CaMK are important for both supplementing current drug regimens and finding new antiparasitic compounds. Whereas calcium regulators, including CaMK, are well characterised in mammalian systems, knowledge of their functional properties in parasites is increasing but is still in its infancy.

Suppression of Schistosoma japonicum Acetylcholinesterase Affects Parasite Growth and Development.

To further investigate the importance of Schistosoma japonicum acetylcholinesterase (SjAChE) in cholinergic signaling for parasite growth and development, we used RNA interference (RNAi) to knock-down its expression in adults and eggs in vitro. This resulted in its reduced transcription but also expression of other important genes involved both in cholinergic signaling and glucose uptake were impacted substantially. Significant decreases in AChE protein expression, AChE enzymatic activity, and glucose uptake were observed in the SjAChE-knockdown parasites compared with luciferase controls. In vaccine/challenge experiments, we found that immunization of mice with recombinant SjAChE (rSjAChE) expressed in Escherichia coli elicited reductions in male worm numbers (33%), liver granuloma density (41%), and reduced numbers of mature intestinal eggs (73%) in the vaccinated group compared with the control group. These results indicate AChE plays an important role in the metabolism of male worms, and impacts indirectly on female fecundity leading to increased numbers of immature eggs being released and reduced sizes of liver granulomas. Furthermore, cytokine analysis showed that immunization of mice with rSjAChE elicited a predominantly Th1-type immune response characterized by increased production of IFNγ in splenic CD4⁺ T cells of vaccinated mice. The study confirms the potential of SjAChE as a vaccine/drug candidate against zoonotic schistosomiasis japonica.

Taurocholate Induces Biliary Differentiation of Liver Progenitor Cells Causing Hepatic Stellate Cell Chemotaxis in the Ductular Reaction: Role in Pediatric Cystic Fibrosis Liver Disease.

Cystic fibrosis liver disease (CFLD) in children causes progressive fibrosis leading to biliary cirrhosis; however, its cause(s) and early pathogenesis are unclear. We hypothesized that a bile acid-induced ductular reaction (DR) drives fibrogenesis. The DR was evaluated by cytokeratin-7 immunohistochemistry in liver biopsies, staged for fibrosis, from 60 children with CFLD, and it demonstrated that the DR was significantly correlated with hepatic fibrosis stage and biliary taurocholate levels. To examine the mechanisms involved in DR induction, liver progenitor cells (LPCs) were treated with taurocholate, and key events in DR evolution were assessed: LPC proliferation, LPC biliary differentiation, and hepatic stellate cell (HSC) chemotaxis. Taurocholate induced a time-dependent increase in LPC proliferation and expression of genes associated with cholangiocyte differentiation (cytokeratin 19, connexin 43, integrin ß4, and γ-glutamyltranspeptidase), whereas the hepatocyte specification marker HNF4α was suppressed. Functional cholangiocyte differentiation was demonstrated via increased acetylated α-tubulin and SOX9 proteins, the number of primary cilia+ LPCs, and increased active γ-glutamyltranspeptidase enzyme secretion. Taurocholate induced LPCs to release MCP-1, MIP1α, and RANTES into conditioned medium causing HSC chemotaxis, which was inhibited by anti-MIP1α. Immunofluorescence confirmed chemokine expression localized to CK7+ DR and LPCs in CFLD liver biopsies. This study suggests that taurocholate is involved in initiating functional LPC biliary differentiation and the development of the DR, with subsequent induction of chemokines that drive HSC recruitment in CFLD.

Characterising granuloma regression and liver recovery in a murine model of schistosomiasis japonica.

For hepatic schistosomiasis the egg-induced granulomatous response and the development of extensive fibrosis are the main pathologies. We used a Schistosoma japonicum-infected mouse model to characterise the multi-cellular pathways associated with the recovery from hepatic fibrosis following clearance of the infection with the anti-schistosomal drug, praziquantel. In the recovering liver splenomegaly, granuloma density and liver fibrosis were all reduced. Inflammatory cell infiltration into the liver was evident, and the numbers of neutrophils, eosinophils and macrophages were significantly decreased. Transcriptomic analysis revealed the up-regulation of fatty acid metabolism genes and the identification of Peroxisome proliferator activated receptor alpha as the upstream regulator of liver recovery. The aryl hydrocarbon receptor signalling pathway which regulates xenobiotic metabolism was also differentially up-regulated. These findings provide a better understanding of the mechanisms associated with the regression of hepatic schistosomiasis.

Lysosome-associated membrane glycoprotein (LAMP)--preliminary study on a hidden antigen target for vaccination against schistosomiasis.

Our previously reported gene atlasing of schistosome tissues revealed transcripts that were highly enriched in the digestive tract of Schistosoma mansoni. From these, we selected two candidates, Sm-LAMP and Sm-NPC2 for testing as vaccine targets. The two molecules were selected on the basis of relatively high expression in the gastrodermis, their potentially important biological function, divergence from homologous molecules of the host and possible apical membrane expression in the gastrodermis. Bacterially expressed recombinant peptides corresponding to regions excluding trans-membrane domains of the selected vaccine targets were used in blinded vaccine trials in CBA mice using alum-CpG as adjuvant. Vaccine trials using the recombinant insoluble Sm-LAMP protein showed 16-25% significant reduction in total worm burden. Faecal egg count reduction was 52% and 60% in two trials, respectively, with similar results for the solubly expressed protein. Liver egg burden was reduced significantly (20% and 38%) with an insoluble recombinant Sm-LAMP in two trials, but not with the soluble recombinant form. Parasite fecundity was not affected by either Sm-LAMP protein preparations in the trials. It is concluded that Sm-LAMP may provide limited protection towards S. mansoni infections but could be used in combination with other vaccine candidates, to provide more comprehensive protection.

Suppression of the Insulin Receptors in Adult Schistosoma japonicum Impacts on Parasite Growth and Development: Further Evidence of Vaccine Potential.

To further investigate the importance of insulin signaling in the growth, development, sexual maturation and egg production of adult schistosomes, we have focused attention on the insulin receptors (SjIRs) of Schistosoma japonicum, which we have previously cloned and partially characterised. We now show, by Biolayer Interferometry, that human insulin can bind the L1 subdomain (insulin binding domain) of recombinant (r)SjIR1 and rSjIR2 (designated SjLD1 and SjLD2) produced using the Drosophila S2 protein expression system. We have then used RNA interference (RNAi) to knock down the expression of the SjIRs in adult S. japonicum in vitro and show that, in addition to their reduced transcription, the transcript levels of other important downstream genes within the insulin pathway, associated with glucose metabolism and schistosome fecundity, were also impacted substantially. Further, a significant decrease in glucose uptake was observed in the SjIR-knockdown worms compared with luciferase controls. In vaccine/challenge experiments, we found that rSjLD1 and rSjLD2 depressed female growth, intestinal granuloma density and faecal egg production in S. japonicum in mice presented with a low dose challenge infection. These data re-emphasize the potential of the SjIRs as veterinary transmission blocking vaccine candidates against zoonotic schistosomiasis japonica in China and the Philippines.

An ex vivo model for studying hepatic schistosomiasis and the effect of released protein from dying eggs.

BACKGROUND: We report the use of an ex vivo precision cut liver slice (PCLS) mouse model for studying hepatic schistosomiasis. In this system, liver tissue is unfixed, unfrozen, and alive for maintenance in culture and subsequent molecular analysis. METHODS AND FINDINGS: Using thick naive mouse liver tissue and sterile culture conditions, the addition of soluble egg antigen (SEA) derived from Schistosoma japonicum eggs, followed 4, 24 and 48 hrs time points. Tissue was collected for transcriptional analysis and supernatants collected to quantitate liver enzymes, cytokines and chemokines. No significant hepatotoxicity was demonstrated by supernatant liver enzymes due to the presence of SEA. A proinflammatory response was observed both at the transcriptional level and at the protein level by cytokine and chemokine bead assay. Key genes observed elevated transcription in response to the addition of SEA included: IL1-α and IL1-ß, IL6, all associated with inflammation. The recruitment of antigen presenting cells was reflected in increases in transcription of CD40, CCL4 and CSF1. Indications of tissue remodeling were seen in elevated gene expression of various Matrix MetalloProteinases (MMP3, 9, 10, 13) and delayed increases in TIMP1. Collagen deposition was significantly reduced in the presence of SEA as shown in COL1A1 expression by qPCR after 24 hrs culture. Cytokine and chemokine analysis of the culture supernatants confirmed the elevation of proteins including IL6, CCL3, CCL4 and CXCL5. CONCLUSIONS: This ex vivo model system for the synchronised delivery of parasite antigen to liver tissue provides an insight into the early phase of hepatic schistosomiasis, corresponding with the release of soluble proteins from dying schistosome eggs.

Transcriptional profiling of the oesophageal gland region of male worms of Schistosoma mansoni.

The intestinal tract of schistosomes opens at the mouth and leads into the foregut or oesophageal region that is lined with syncytium continuous with the apical cytoplasm of the tegument. The oesophagus is surrounded by a specialised gland, the oesophageal gland. This gland releases materials into the lumen of the oesophagus and the region is thought to initiate the lysis of erythrocytes and neutralisation of immune effectors of the host. The oesophageal region is present in the early invasive schistosomulum, a stage potentially targetable by anti-schistosome vaccines. We used a 44k oligonucleotide microarray to identify highly up-regulated genes in microdissected frozen sections of the oesophageal gland of male worms of S. mansoni. We show that 122 genes were up-regulated 2-fold or higher in the oesophageal gland compared with a whole male worm tissue control. The enriched genes included several associated with lipid metabolism and transmembrane transport as well as some micro-exon genes. Since the oesophageal gland is important in the initiation of digestion and the fact that it develops early after invasion of the mammalian host, further study of selected highly up-regulated functionally important genes in this tissue may reveal new anti-schistosome intervention targets for schistosomiasis control.

Gene Atlasing of digestive and reproductive tissues in Schistosoma mansoni.

BACKGROUND: While considerable genomic and transcriptomic data are available for Schistosoma mansoni, many of its genes lack significant annotation. A transcriptomic study of individual tissues and organs of schistosomes could play an important role in functional annotation of the unknown genes, particularly by providing rapid localisation data and thus giving insight into the potential roles of these molecules in parasite development, reproduction and homeostasis, and in the complex host-parasite interaction. METHODOLOGY/PRINCIPAL FINDINGS: Quantification of gene expression in tissues of S. mansoni was achieved by a combination of laser microdissection microscopy (LMM) and oligonucleotide microarray analysis. We compared the gene expression profile of the adult female gastrodermis and male and female reproductive tissues with whole worm controls. The results revealed a total of 393 genes (contigs) that were up-regulated two-fold or more in the gastrodermis, 4,450 in the ovary, 384 in the vitelline tissues of female parasites, and 2,171 in the testes. We have also supplemented these data with the identification of highly expressed genes in different regions of manually dissected male and female S. mansoni. Though relatively crude, this dissection strategy provides low resolution localisation data for critical regions of the adult parasites that are not amenable to LMM isolation. CONCLUSIONS: This is the first detailed transcriptomic study of the reproductive tissues and gastrodermis of S. mansoni. The results obtained will help direct future research on the functional aspects of these tissues, expediting the characterisation of currently unannotated gene products of S. mansoni and the discovery of new drug and vaccine targets.

Molecular analysis of zinc transporters in Schistosoma japonicum.

Members of the zinc-regulated transporter/iron-regulated transporter-like protein (ZIP) family of proteins transport metal ions across cell membranes. Genes encoding ZIPs are present in the genomes of schistosomes. Here, we describe molecular characterisation of six ZIPs (SjZIPA-F) from Schistosoma japonicum. Quantitative PCR analyses of these ZIPs through the lifecycle showed that each is expressed predominantly during the intramammalian stage and are particularly enriched in adult females. Using laser microdissected tissue as template, SjZIPA-D were transcriptionally enriched in female reproductive tissues, SjZIPE was not expressed in specific tissues and SjZIPF was expressed similarly in each tissue. Whole mount in situ hybridization revealed that SjZIPA and SjZIPB were localised to the oesophageal gland of adults and the vitellaria. We have shown that multiple ZIPs are expressed by schistosomes during the intramammalian parasitic phases and propose that the encoded products perform diverse cellular functions related to metal transport in different cells of S. japonicum.

Increased iron stores correlate with worse disease outcomes in a mouse model of schistosomiasis infection.

Schistosomiasis is a significant parasitic infection creating disease burden throughout many of the world's developing nations. Iron deficiency anemia is also a significant health burden resulting from both nutritional deficit as well as parasitic infection in these countries. In this study we investigated the relationships between the disease outcomes of Schistosoma japonicum infection and iron homeostasis. We aimed to determine if host iron status has an effect on schistosome maturation or egg production, and to investigate the response of iron regulatory genes to chronic schistosomiasis infection. Wild-type C57BL/6 and Transferrin Receptor 2 null mice were infected with S. japonicum, and sacrificed at the onset of chronic disease. Transferrin Receptor 2 null mice are a model of type 3 hereditary hemochromatosis and develop significant iron overload providing increased iron stores at the onset of infection. The infectivity of schistosomes and egg production was assessed along with the subsequent development of granulomas and fibrosis. The response of the iron regulatory gene Hepcidin to infection and the changes in iron status were assessed by real-time PCR and Western blotting. Our results show that Hepcidin levels responded to the changing iron status of the animals, but were not significantly influenced by the inflammatory response. We also show that with increased iron availability at the time of infection there was greater development of fibrosis around granulomas. In conclusion, our studies indicate that chronic inflammation may not be the primary cause of the anemia seen in schistosomiasis, and suggest that increased availability of iron, such as through iron supplementation, may actually lead to increased disease severity.