Keeping bugs in check: The mucus layer as a critical component in maintaining intestinal homeostasis

In the mammalian gastrointestinal tract the close vicinity of abundant immune effector cells and trillions of commensal microbes requires sophisticated barrier and regulatory mechanisms to maintain vital host-microbial interactions and tissue homeostasis. During co-evolution of the host and its intestinal microbiota a protective multilayered barrier system was established to segregate the luminal microbes from the intestinal mucosa with its potent immune effector cells, limit bacterial translocation into host tissues to prevent tissue damage, while ensuring the vital functions of the intestinal mucosa and the luminal gut microbiota. In the present review we will focus on the different layers of protection in the intestinal tract that allow the successful mutualism between the microbiota and the potent effector cells of the intestinal innate and adaptive immune system. In particular, we will review some of the recent findings on the vital functions of the mucus layer and its site-specific adaptations to the changing quantities and complexities of the microbiota along the (gastro-) intestinal tract. Understanding the regulatory pathways that control the establishment of the mucus layer, but also its degradation during intestinal inflammation may be critical for designing novel strategies aimed at maintaining local tissue homeostasis and supporting remission from relapsing intestinal inflammation in patients with inflammatory bowel diseases.

Species interactions and random dispersal rather than habitat filtering drive community assembly during early plant succession

Theory on plant succession predicts a temporal increase in the complexity of spatial community structure and of competitive interactions: initially random occurrences of early colonising species shift towards spatially and competitively structured plant associations in later successional stages. Here we use long-term data on early plant succession in a German post mining area to disentangle the importance of random colonisation, habitat filtering, and competition on the temporal and spatial development of plant community structure. We used species co-occurrence analysis and a recently developed method for assessing competitive strength and hierarchies (transitive versus intransitive competitive orders) in multispecies communities. We found that species turnover decreased through time within interaction neighbourhoods, but increased through time outside interaction neighbourhoods. Successional change did not lead to modular community structure. After accounting for species richness effects, the strength of competitive interactions and the proportion of transitive competitive hierarchies increased through time. Although effects of habitat filtering were weak, random colonization and subsequent competitive interactions had strong effects on community structure. Because competitive strength and transitivity were poorly correlated with soil characteristics, there was little evidence for context dependent competitive strength associated with intransitive competitive hierarchies.

An ultrasensitive NanoLuc-based luminescence system for monitoring Plasmodium berghei throughout its life cycle

BACKGROUND: Bioluminescence imaging is widely used for cell-based assays and animal imaging studies, both in biomedical research and drug development. Its main advantages include its high-throughput applicability, affordability, high sensitivity, operational simplicity, and quantitative outputs. In malaria research, bioluminescence has been used for drug discovery in vivo and in vitro, exploring host-pathogen interactions, and studying multiple aspects of Plasmodium biology. While the number of fluorescent proteins available for imaging has undergone a great expansion over the last two decades, enabling simultaneous visualization of multiple molecular and cellular events, expansion of available luciferases has lagged behind. The most widely used bioluminescent probe in malaria research is the Photinus pyralis firefly luciferase, followed by the more recently introduced Click-beetle and Renilla luciferases. Ultra-sensitive imaging of Plasmodium at low parasite densities has not been previously achieved. With the purpose of overcoming these challenges, a Plasmodium berghei line expressing the novel ultra-bright luciferase enzyme NanoLuc, called PbNLuc has been generated, and is presented in this work. RESULTS: NanoLuc shows at least 150 times brighter signal than firefly luciferase in vitro, allowing single parasite detection in mosquito, liver, and sexual and asexual blood stages. As a proof-of-concept, the PbNLuc parasites were used to image parasite development in the mosquito, liver and blood stages of infection, and to specifically explore parasite liver stage egress, and pre-patency period in vivo. CONCLUSIONS: PbNLuc is a suitable parasite line for sensitive imaging of the entire Plasmodium life cycle. Its sensitivity makes it a promising line to be used as a reference for drug candidate testing, as well as the characterization of mutant parasites to explore the function of parasite proteins, host-parasite interactions, and the better understanding of Plasmodium biology. Since the substrate requirements of NanoLuc are different from those of firefly luciferase, dual bioluminescence imaging for the simultaneous characterization of two lines, or two separate biological processes, is possible, as demonstrated in this work.

Effects of the microenvironment on Fas/Fas ligand interactions and their implications for UV-induced immune suppression

Skin cancer is the most common malignancy in humans. Although highly treatable, non-melanoma skin cancer is commonly followed by other non-cutaneous malignancies. Ultraviolet radiation (UVR) acts as both tumor initiator and promoter, and also results in the suppression of specific immune responses. The systemic suppression of immune responses is initiated by DNA damage, which promotes IL-10 production, an important cytokine as anti-IL-10 can abrogate the suppression, and upregulates the pro-apoptotic proteins Fas and Fas ligand (FasL). FasL is a critical factor for UV-induced immune suppression, and the suppressor cell induced by UV expresses FasL. ^ We hypothesized that the microenvironment affects Fas/FasL interactions, and that these interactions are important to the phenomenon of UV induced immune suppression. To determine the effects of the interaction of FasL and IL-10, splenocytes isolated from C57Bl/6 mice were cultured in the presence or absence of IL-10 post-mitogenic activation. We determined that IL-10 protects from Fas-mediated apoptosis by lowering Fas sensitivity and lowering the levels of either Fas or FasL. This protection is stronger when IL-10 is given immediately after mitogenic activation, and does not increase any of the inhibitors of apoptosis studied. In vivo, splenocytes from UV-irradiated mice are resistant to Fas-mediated apoptosis and present very high levels of IL-10, lowered Fas sensitivity and lowered caspase cleavage despite higher expression of Fas and FasL than non-irradiated mice. ^ UV-induced immune suppression affects female mice preferentially, which led us to look at prolactin as a possible component of this suppression since this hormone has also been associated with increased skin carcinogenesis. The interaction of FasL and prolactin results in suppression of the delayed type hypersensitivity response to Candida albicans. This lack of response depends on FasL as is not seen in gld mice. Similar to UV-induced immune suppression, the suppression is caused by a Th2 deviation, and correlates with a significant increase in Fas expression. In the presence of UV, the effects of prolactin seemed to be protective, and UV actually restores the DTH response.^ Taken together, these observations suggest that the microenvironment dictates the outcome of the interaction of FasL with Fas going from promoting apoptosis to preventing apoptosis or mediating a Th2 deviation and suppression of a Th1 response. ^

Characterization of humoral immune responses against Treponema pallidum antigens

The spirochete Treponema pallidum subsp. pallidum is the causative agent of syphilis, a sexually transmitted disease with an estimated 12 million new cases per year worldwide. There is no vaccine currently available for the prevention of syphilis. In the present study, the T. pallidum hypothetical protein TP0693 was examined to determine its cellular location, and its potential for use as a vaccine candidate and immunodiagnostic for syphilis. TP0693 was demonstrated to be strongly reactive with sera from rabbits infected experimentally with T. pallidum for >25 days. Results from proteinase K digestion, immunofluorescence and immunoelectron microscopy were consistent with outer surface localization of TP0693. Serum reactivity against TP0693 was detected in only 68% of syphilis patients, which does not support its use as an immunodiagnostic for syphilis. Immunization of rabbits with TP0693 or three other outer membrane candidates did not alter the course of lesion development atter T. pallidum inoculation. We also examined the T. pallidum proteome by two-dimensional gel electrophoresis coupled with mass spectrometry analysis and immunoblotting. This approach resulted in the identification of 95 unique polypeptides, several of which were reactive with sera from infected rabbits and syphilis patients. The analyses described here enabled us to identify antigens potentially useful as vaccine candidates or diagnostic markers, and may provide insight into host-pathogen interactions during T. pallidum infection. ^

Fatty acid and stable isotope composition of calanoid copepods in the open eastern Atlantic Ocean during POLARSTERN cruise ANT-XXIX/1

The majority of global ocean production and total export production is attributed to oligotrophic oceanic regions due to their vast regional expanse. However, energy transfers, food-web structures and trophic relationships in these areas remain largely unknown. Regional and vertical inter- and intra-specific differences in trophic interactions and dietary preferences of calanoid copepods were investigated in four different regions in the open eastern Atlantic Ocean (38°N to 21°S) in October/November 2012 using a combination of fatty acid (FA) and stable isotope (SI) analyses. Mean carnivory indices (CI) based on FA trophic markers generally agreed with trophic positions (TP) derived from d15N analysis. Most copepods were classified as omnivorous (CI ~0.5, TP 1.8 to ~2.5) or carnivorous (CI >=0.7, TP >=2.9). Herbivorous copepods showed typical CIs of <=0.3. Geographical differences in d15N values of epi- (200-0 m) to mesopelagic (1000-200 m) copepods reflected corresponding spatial differences in baseline d15N of particulate organic matter from the upper 100 m. In contrast, species restricted to lower meso- and bathypelagic (2000-1000 m) layers did not show this regional trend. FA compositions were species-specific without distinct intra-specific vertical or spatial variations. Differences were only observed in the southernmost region influenced by the highly productive Benguela Current. Apparently, food availability and dietary composition were widely homogeneous throughout the oligotrophic oceanic regions of the tropical and subtropical Atlantic. Four major species clusters were identified by principal component analysis based on FA compositions. Vertically migrating species clustered with epi- to mesopelagic, non-migrating species, of which only Neocalanus gracilis was moderately enriched in lipids with 16% of dry mass (DM) and stored wax esters (WE) with 37% of total lipid (TL). All other species of this cluster had low lipid contents (< 10% DM) without WE. Of these, the tropical epipelagic Undinula vulgaris showed highest portions of bacterial markers. Rhincalanus cornutus, R. nasutus and Calanoides carinatus formed three separate clusters with species-specific lipid profiles, high lipid contents (>=41% DM), mainly accumulated as WE (>=79% TL). C. carinatus and R. nasutus were primarily herbivorous with almost no bacterial input. Despite deviating feeding strategies, R. nasutus clustered with deep-dwelling, carnivorous species, which had high amounts of lipids (>=37% DM) and WE (>=54% TL). Tropical and subtropical calanoid copepods exhibited a wide variety of life strategies, characterized by specialized feeding. This allows them, together with vertical habitat partitioning, to maintain high abundance and diversity in tropical oligotrophic open oceans, where they play an essential role in the energy flux and carbon cycling.

Learning Analytics and Interactions in Virtual Learning Environments. A Comparative Study of Typologies and their Relationship with Academic Performance

Analysis of learning data (learning analytics) is a new research field with high growth potential. The main objective of Learning analytics is the analysis of data (interactions being the basic data unit) generated in virtual learning environments, in order to maximize the outcomes of the learning process; however, a consensus has not been reached yet on which interactions must be measured and what is their influence on learning outcomes. This research is grounded on the study of e-learning interaction typologies and their relationship with students? academic performance, by means of a comparative study between different interaction typologies (based on the agents involved, frequency of use and participation mode). The main conclusions are a) that classifications based on agents offer a better explanation of academic performance; and b) that each of the three typologies are able to explain academic performance in terms of some of their components (student-teacher and student-student interactions, evaluating students interactions and active interactions, respectively), with the other components being nonrelevant.

Interactions of the chaperone Hsp104 with yeast Sup35 and mammalian PrP

[PSI+] is a genetic element in yeast for which a heritable change in phenotype appears to be caused by a heritable change in the conformational state of the Sup35 protein. The inheritance of [PSI+] and the physical state of Sup35 in vivo depend on the protein chaperone Hsp104 (heat shock protein 104). Although these observations provide a strong genetic argument in support of the “protein-only” or “prion” hypothesis for [PSI+], there is, as yet, no direct evidence of an interaction between the two proteins. We report that when purified Sup35 and Hsp104 are mixed, the circular dichroism (CD) spectrum differs from that predicted by the addition of the proteins’ individual spectra, and the ATPase activity of Hsp104 is inhibited. Similar results are obtained with two other amyloidogenic substrates, mammalian PrP and β-amyloid 1-42 peptide, but not with several control proteins. With a group of peptides that span the PrP protein sequence, those that produced the largest changes in CD spectra also caused the strongest inhibition of ATPase activity in Hsp104. Our observations suggest that (i) previously described genetic interactions between Hsp104 and [PSI+] are caused by direct interaction between Hsp104 and Sup35; (ii) Sup35 and PrP, the determinants of the yeast and mammalian prions, respectively, share structural features that lead to a specific interaction with Hsp104; and (iii) these interactions couple a change in structure to the ATPase activity of Hsp104.

Cnr interferes with dimerization of the replication protein α in phage-plasmid P4

DNA replication of phage-plasmid P4 in its host Escherichia coli depends on its replication protein α. In the plasmid state, P4 copy number is controlled by the regulator protein Cnr (copy number regulation). Mutations in α (αcr) that prevent regulation by Cnr cause P4 over-replication and cell death. Using the two-hybrid system in Saccharomyces cerevisiae and a system based on λ immunity in E.coli for in vivo detection of protein–protein interactions, we found that: (i) α protein interacts with Cnr, whereas αcr proteins do not; (ii) both α–α and αcr–αcr interactions occur and the interaction domain is located within the C-terminal of α; (iii) Cnr–Cnr interaction also occurs. Using an in vivo competition assay, we found that Cnr interferes with both α–α and αcr–αcr dimerization. Our data suggest that Cnr and α interact in at least two ways, which may have different functional roles in P4 replication control.

Detection of a major gene for resistance to fusiform rust disease in loblolly pine by genomic mapping.

Genomic mapping has been used to identify a region of the host genome that determines resistance to fusiform rust disease in loblolly pine where no discrete, simply inherited resistance factors had been previously found by conventional genetic analysis over four decades. A resistance locus, behaving as a single dominant gene, was mapped by association with genetic markers, even though the disease phenotype deviated from the expected Mendelian ratio. The complexity of forest pathosystems and the limitations of genetic analysis, based solely on phenotype, had led to an assumption that effective long-term disease resistance in trees should be polygenic. However, our data show that effective long-term resistance can be obtained from a single qualitative resistance gene, despite the presence of virulence in the pathogen population. Therefore, disease resistance in this endemic coevolved forest pathosystem is not exclusively polygenic. Genomic mapping now provides a powerful tool for characterizing the genetic basis of host pathogen interactions in forest trees and other undomesticated, organisms, where conventional genetic analysis often is limited or not feasible.

Role of IFN-gamma in immune responses to Candida albicans infections

Candida albicans is the most frequent etiologic agent that causes opportunistic fungal infections called candidiasis, a disease whose systemic manifestation could prove fatal and whose incidence is increasing as a result of an expanding immunocompromised population. Here we review the role of interferon-gamma (IFN-γ) in host protection against invasive candidiasis. This cytokine plays an essential role in both the innate and adaptive arms of the immune response to candidiasis. We focus on recent progress on host-pathogen interactions leading to the production of IFN-γ by host cells. IFN-γ is produced by CD4 Th1, CD8, γδ T, and natural killer (NK) cells, essentially in response to both IL-12 and/or IL-18; more recently, a subset of C. albicans-specific Th17 cells have been described to produce both IL-17 and IFN-γ. IFN-γ plays an important role in the regulation of the immune system as well as in the control of the infectious process, as it is required for optimal activation of phagocytes, collaborates in the generation of protective antibody response, and favors the development of a Th1 protective response.

In vivo imaging enters parasitology

In vivo infection routes of parasites have remained something of a "black box", in which only snapshot views of fixed tissues are available. Clearly, there exists a strong need for imaging approaches to visualise living parasites within intact organs and animals. In vivo imaging of fluorescent Plasmodium parasites now provides us with exciting insights into the infection process, from the bite of the infected mosquito to the invasion of liver cells, and alternative approaches using luciferase-expressing parasites have been used to monitor their dissemination in mice. This rapidly developing field will go a long way towards deepening our understanding of host-parasite interactions at different levels.

Gene-environment interactions between adult lead exposure and Apolipoprotein E4 on adult hippocampal neurogenesis and cognitive behavior in mice

Thesis (Ph.D.)--University of Washington, 2016-06

Current concepts of tumour metastasis

Background: Tumour metastasis remains the principal cause of treatment failure and poor prognosis in patients with cancer. Recent advances in our understanding of the biology of metastasis are providing novel potential targets for anti-cancer therapies. Aim: This paper reviews the current concepts in tumour metastasis. Methods: A review of Medline publications relating to the molecular biology and therapy of human tumour metastasis was conducted. Results and Discussion: Early metastasis models were based upon the premise of uninterrupted tumour growth, with the inevitable formation of distant metastases and eventual death of the patient. However, current research suggests that metastasis is an inefficient process governed by several rate-limiting steps, and that failure to negotiate these steps can lead to tumour dormancy. Successful metastatic tumour growth depends upon appropriate tumour-host microenvironment interactions and, ultimately, the development of vascularised metastases post-extravasation in the target organ. An understanding of the molecular mechanisms involved in this dynamic process will aid in the identification of therapeutic targets that may allow earlier diagnosis and more specific therapies for patients with metastasis.

Coordinations in business interactions between Hong Kong Chinese, Mainland Chinese and western actors

Hong Kong firms continue to play an important role in the development of mainland China trade. This paper studies the nature of coordinations in triadic interactions between Hong Kong Chinese intermediaries and mainland Chinese sellers, and buyers from the West, and their impact on relationships development. Contrary to popular belief, the results presented here identify that interactions between Hong Kong intermediaries and mainland Chinese sellers tend to include relationships based on price and authority. These interactions tend to be classical in nature, and serve to hinder the development of relationships. On the other hand, interactions between buyers from the West and Hong Kong intermediaries tend to be relational in nature. They include different forms of personal relationships, which tends to be positive for the development of relationships. A new plural form of international business relationship triad is also identified.