Differential Expression of Chemokine and Matrix Re-Modelling Genes Is Associated with Contrasting Schistosome-Induced Hepatopathology in Murine Models

The pathological outcomes of schistosomiasis are largely dependent on the molecular and cellular mechanisms of the host immune response. In this study, we investigated the contribution of variations in host gene expression to the contrasting hepatic pathology observed between two inbred mouse strains following Schistosoma japonicum infection. Whole genome microarray analysis was employed in conjunction with histological and immunohistochemical analysis to define and compare the hepatic gene expression profiles and cellular composition associated with the hepatopathology observed in S. japonicum-infected BALB/c and CBA mice. We show that the transcriptional profiles differ significantly between the two mouse strains with high statistical confidence. We identified specific genes correlating with the more severe pathology associated with CBA mice, as well as genes which may confer the milder degree of pathology associated with BALB/c mice. In BALB/c mice, neutrophil genes exhibited striking increases in expression, which coincided with the significantly greater accumulation of neutrophils at granulomatous regions seen in histological sections of hepatic tissue. In contrast, up-regulated expression of the eosinophil chemokine CCL24 in CBA mice paralleled the cellular influx of eosinophils to the hepatic granulomas. Additionally, there was greater down-regulation of genes involved in metabolic processes in CBA mice, reflecting the more pronounced hepatic damage in these mice. Profibrotic genes showed similar levels of expression in both mouse strains, as did genes associated with Th1 and Th2 responses. However, imbalances in expression of matrix metalloproteinases (e.g. MMP12, MMP13) and tissue inhibitors of metalloproteinases (TIMP1) may contribute to the contrasting pathology observed in the two strains. Overall, these results provide a more complete picture of the molecular and cellular mechanisms which govern the pathological outcome of hepatic schistosomiasis. This improved understanding of the immunopathogenesis in the murine model schistosomiasis provides the basis for a better appreciation of the complexities associated with chronic human schistosomiasis

Christian Dotremont // Pierre Alechinsky

Well implemented criterion-referenced assessment (CRA) requires dedicated time and effort,\ud especially in describing realistic expectations of evidence of achievement to students in the form\ud of criteria sheets (or grading rubrics). It is also takes time out of delivering content to teach\ud students how to judge their own work using criteria sheets. In 2007, to engage third year\ud Microbiology students in using criteria sheets for the first time in their degree, we devised an\ud innovative assessment tutorial supported by online resources. We were sceptical of much of the\ud literature that reported ‘agreed’ characteristics of our predominantly gen Y cohort, because of\ud the older ages of the majority of authors. These authors claim gen Y has a propensity for digital\ud media, overconfidence in their own abilities and a collaborative orientation. We rejected this\ud stereotype when developing the tutorial. Evaluations by students were positive and there was no\ud dramatic change to grades for the unit. These results are similar to those in the literature for non\ud gen Y cohorts. This lends support to our claim that giving students control over their own\ud learning, irrespective of their generational label, is worth the time and effort

Levelling expectations across different years in an undergraduate degree to support work-related learning : evidence for a multi-pronged approach.

One of the ways in which university departments and faculties can enhance the quality of learning and\ud assessment is to develop a ‘well thought out criterion‐referenced assessment system’ (Biggs, 2003, p. 271). In\ud designing undergraduate degrees (courses) this entails making decisions about the levelling of expectations\ud across different years through devising objectives and their corresponding criteria and standards: a process of\ud alignment analogous to what happens in unit (subject) design. These decisions about levelling have important\ud repercussions in terms of supporting students’ work‐related learning, especially in relation to their ability to\ud cope with the increasing cognitive and skill demands made on them as they progress through their studies. They\ud also affect the accountability of teacher judgments of students’ responses to assessment tasks, achievement of\ud unit objectives and, ultimately, whether students are awarded their degrees and are sufficiently prepared for the\ud world of work.\ud Research reveals that this decision‐making process is rarely underpinned by an explicit educational rationale\ud (Morgan et al, 2002). The decision to implement criterion referenced assessment in an undergraduate\ud microbiology degree was the impetus for developing such a rationale because of the implications for alignment,\ud and therefore ‘levelling’ of expectations across different years of the degree. This paper provides supporting\ud evidence for a multi‐pronged approach to levelling, through backward mapping of two revised units (foundation\ud and exit year). This approach adheres to the principles of alignment while combining a work‐related approach\ud (via industry input) with the blended disciplinary and learner‐centred approaches proposed by Morgan et al.\ud (2002). It is suggested that this multi‐pronged approach has the potential for making expectations, especially\ud work‐related ones across different year levels of degrees, more explicit to students and future employers

Differential expression of ompA, ompB, pyk, nlpD and Cpn0585 genes between normal and interferon-gamma treated cultures of Chlamydia pneumoniae

A common feature of many chlamydial infections is that they are often\ud asymptomatic and may persist for long periods of time if left untreated. In\ud addition to the well recognized lytic stage of the chlamydial developmental\ud cycle, evidence is now emerging to support a persistent phase in the cycle in\ud which the reticulate bodies are morphologically abnormal, viable but\ud non-infectious and presumably also have altered gene expression patterns. We\ud used an RT-PCR approach to study the differential levels of gene transcription\ud for 14 genes (16SrRNA, ompA, ompB, omcB, 76 kDa, gseA, pmp1, gltX, hsp60, yaeT,\ud pyk, nlpD, Cpn0585, Cpn1046) between normal and IFN-gamma treated Chlamydia\ud pneumoniae cell cultures. Even though the level of morphologically abnormal\ud reticulate bodies in our IFN-gamma treated cultures was low (approximately 10%\ud morphologically discernible, although presumably a larger percentage were in the\ud persistent state but not yet morphologically altered) we identified five genes\ud (ompA, ompB, pyk, nlpD, Cpn0585) that were clearly upregulated when compared to\ud normal cultures. This gene transcript profile may be characteristic of a general\ud stress state in Chlamydia, induced by IFN-gamma treatment in this case, but\ud perhaps more widely induced in other in vitro and in vivo situations

A family of cathepsin B cysteine proteases expressed in the gut of the human hookworm, Necator americanus

mRNAs encoding cathepsin B-like cysteine proteases (CatBs) are abundantly expressed in the genomes of blood-feeding nematodes. Recombinant CatBs have been partially efficacious in vaccine trials in animal models of hookworm infection, supporting further investigation of these enzymes as new control tools. We recently described a family of four distinct CatBs (Na-CP-2, -3, -4, -5) from the human hookworm, Necator americanus. Here we show that these N. americanus CatBs form a robust clade with other hookworm CatBs and are most similar to intestinal CatBs from Haemonchus contortus. All four mRNAs (Na-cp-2, -3, -4 and -5) are up-regulated during the transition from a free-living larva to a blood-feeding adult worm and are also expressed in gut tissue of adult N. americanus that was dissected using laser microdissection microscopy. Recombinant Na-CP-3 was expressed in soluble, secreted form in the yeast Pichia pastoris, while Na-CP-2, -4 and -5 were expressed in insoluble inclusion bodies in Escherichia coli. Recombinant Na-CP-3 was not catalytically active when secreted by yeast but underwent auto-activation to an active enzyme at low pH in the presence of dextran sulphate. Activated Na-CP-3 digested gelatin and cleaved the fluorogenic substrate Z-Phe-Arg-aminomethylcoumarin (AMC) but not Z-Arg-Arg-AMC. Recombinant Na-CP-3 did not digest intact hemoglobin but digested globin fragments generated by prior hydrolysis with N. americanus aspartic hemoglobinases. Antibodies raised in mice to all four recombinant proteins showed minimal cross-reactivity with each other, and each antiserum bound to the intestine of adult N. americanus, supporting the intestinal expression of their mRNAs. These data show that N. americanus expresses a family of intestinal CatBs, many of which are likely to be involved in nutrient acquisition and therefore are potential targets for chemotherapies and vaccines

Peptide polymerisation facilitates incorporation into ISCOMs and increases antigen-specific IgG2a production

Synthetic peptides can be tailor-made to include any B or T epitopes desired from a single or multiple antigens or organisms. However, peptides in general are not very immunogenic and have not proven easy to incorporate into immunogenic vaccines. ISCOMs is an adjuvant system that has the capability not only to enhance the humoral immunogenicity of a protein but has also been shown to induce cell-mediated immune responses in animals. Synthetic peptide ISCOM vaccines are few because of the difficulty in incorporation of these peptides into ISCOMs. We have shown in this study that non-immunogenic peptides could be made immunogenic by polymerisation, and these polymers could be incorporated into ISCOMs to give highly immunogenic vaccines. Synthetic 20mer peptides containing known B and T-helper epitopes from the E7 protein of the cervical cancer associated human papillomavirus type 16 (HPV16 E7) have been used here as model immunogens. We have compared the humoral immunity induced by these peptides as polymers or as copolymers with a lipid binding 20mer peptide (LAP 20), with or without incorporation into ISCOMs. Unpolymerised peptide elicited no measurable antibody. When polymerised peptide was administered with CFA, or in phosphate-buffered saline (PBS) without adjuvant, or incorporated into ISCOMs, antibodies recognising both the immunising peptide and HPV16 E7 protein were produced. For equal quantities of administered peptide (5 μg), ISCOMs gave higher titres of antibody than CFA or PBS. Polymerised peptides induced high antigen-specific IgG2a:IgG1 ratios, which increased with multiple immunisations. These data indicate that polymerised peptides could be incorporated into ISCOMs to form efficient immunogens which may elicit a Th1 type response