Project description:Purpose: Identify genes implicated in resistance to H. polygyrus Method: RNA was extracted from duodenal tissue from C57BL/6 mice during primary and secondary H. polygyrus. Total RNA libraries were created using the Encore® Complete RNA-Seq Library Systems kit (NuGEN), following manufacturer’s instructions. Total RNA libraries were sequenced using the Illumina® HiSeq 2500. The raw Illumina reads were analysed as follows. First, the data quality was analysed using FastQC. Then the low quality bases were trimmed using Trimmomatic. The read pairs which passed the trimming quality filters were then aligned to mm10 (Ensembl version 75) using Tophat2. Counts were determined using htseq_count. Normalisation and statistical analysis was performed using edgeR. Differential gene analysis was calculated from naïve control group (A). Statistically significant genes with FDR < 0.05 are reported. Results: Following analysis, we identified many transcripts significantly different from the naïve control following primary and secondary H. polygyrus infection. Conclusions: The intestinal transcriptome of both primary and secondary infected mice differ significantly, identifying qualitative and quantitative differences between susceptible (primary infected) and resistant (secondary infected) mice.
Project description:The intestinal helminth parasite Heligmosomoides polygyrus initiates infection in mice by penetrating the duodenal mucosa, where it develops while surrounded by a multicellular granulomatous infiltrate before emerging into the intestinal lumen. We examined early H. polygyrus infection to assess the epithelial response to disruption of the mucosal barrier. Unexpectedly, intestinal stem cell markers, including Lgr5 and Olfm4, were completely lost in crypts overlying larvae-associated granulomas. We sought to identify the mechanism by which the H. polygyrus granuloma represses the activity of intestinal stem cells.
Project description:Immunity to intestinal helminth infections has been well studied, but the mechanism of helminth killing prior to expulsion remains unclear. Here we identify epithelial-cell-derived phospholipase A2 group 1B (PLA2g1B) as a host-derived endogenous anthelmintic. PLA2g1B is elevated in resistant mice and is responsible for killing tissue-embedded larvae. Despite comparable activities of other essential type-2-dependent immune mechanisms, Pla2g1b-/- mice failed to expel the intestinal helminths Heligmosomoides polygyrus or Nippostrongylus brasiliensis. Expression of Pla2g1b by epithelial cells was dependent upon intestinal microbiota, adaptive immunity, and common-gamma chain-dependent signaling. Notably, Pla2g1b was downregulated in susceptible mice and inhibited by IL-4R-signaling in vitro, uncoupling parasite killing from expulsion mechanisms. Resistance was restored in Pla2g1b-/- mice by treating infective H. polygyrus L3 larvae with PLA2g1B, which reduced larval phospholipid abundance. These findings uncover epithelial-cell-derived Pla2g1b as an essential mediator of helminth killing, highlighting a previously overlooked mechanism of anti-helminth immunity.
Project description:Heligmosomoides polygyrus larvae induce a strong intestinal granuloma response within their murine host, which has been associated with resistance. Immune cells, mostly alternatively activated macrophages and eosinophils, accumulate around the tissue encysted parasites to immobilize and damage/kill developing worms. In a one dose (bolus) experimental infection, infected C57Bl/6 mice are unable to clear parasites which results in chronic infection with high worm burdens. However, using a frequent dose trickle model of infection, we, like others, have found that C57Bl/6 mice can clear infection. The nanoString data included here measure the expression of key myeloid genes within the granuloma tissue from bolus and trickle infections. Our results highlight the importance of the granuloma in the host’s ability to clear H. polygyrus and emphasise the need to study this key tissue in more depth, rather than using correlates such as general intestinal or systemic responses.
Project description:Mouse bone marrow derived macrophages were stimulated with L3 larvae of the helminth Heligmosomoides polygyrus (Hp) (500 L3 /1 mio cells) in the presence or absence of immune serum (1:50 v:v) from challenge-Hp-infected mice.
Project description:Heligmosomoides polygyrus is a natural intestinal parasite of mice which exerts wide ranging modulatory effects on the immune system. This experiment was designed to investigate its abillity to modify intestinal epithelial cells, which form part of its natural niche. We tested gene expression in vitro, in differentiating organoids of small intestinal origin, exposed to cytokines and the released products of the parasite, termed HpES.
Project description:mRNA was isolated from duodenal tissue 3d after induction of acute pancreatitis in C57Bl/6 wild type mice to investigate transcriptional changes, untreated C57Bl/6 refer as controls we used microarrays to investigate the duodenal barrier function during acute pancreatitis
Project description:In this study, the composition of ES of male and female L4 stage Heligmosomoides polygyrus bakeri in the presence (cultured together) or absence (cultured alone) of the opposite sex was examined using mass spectrometry.