What have dendritic cells ever done for adjuvant design? Cellular and molecular methods for the rational development of vaccine adjuvants

Our new molecular understanding of immune priming states that dendritic cell activation is absolutely pivotal for expansion and differentiation of naïve T lymphocytes, and it follows that understanding DC activation is essential to understand and design vaccine adjuvants. This chapter describes how dendritic cells can be used as a core tool to provide detailed quantitative and predictive immunomics information about how adjuvants function. The role of distinct antigen, costimulation, and differentiation signals from activated DC in priming is explained. Four categories of input signals which control DC activation – direct pathogen detection, sensing of injury or cell death, indirect activation via endogenous proinflammatory mediators, and feedback from activated T cells – are compared and contrasted. Practical methods for studying adjuvants using DC are summarised and the importance of DC subset choice, simulating T cell feedback, and use of knockout cells is highlighted. Finally, five case studies are examined that illustrate the benefit of DC activation analysis for understanding vaccine adjuvant function.

Differentiation of neotropical ecosystems by modern soil phytolith assemblages and its implications for palaeoenvironmental and archaeological reconstructions

The interpretation of Neotropical fossil phytolith assemblages for palaeoenvironmental and archaeological reconstructions relies on the development of appropriate modern analogues. We analyzed modern phytolith assemblages from the soils of ten distinctive tropical vegetation communities in eastern lowland Bolivia, ranging from terra firme humid evergreen forest to seasonally-inundated savannah. Results show that broad ecosystems – evergreen tropical forest, semi-deciduous dry tropical forest, and savannah – can be clearly differentiated by examination of their phytolith spectra and the application of Principal Component Analysis (PCA). Differences in phytolith assemblages between particular vegetation communities within each of these ecosystems are more subtle, but can still be identified. Comparison of phytolith assemblages with pollen rain data and stable carbon isotope analyses from the same vegetation plots show that these proxies are not only complementary, but significantly improve taxonomic and ecosystem resolution, and therefore our ability to interpret palaeoenvironmental and archaeological records. Our data underline the utility of phytolith analyses for reconstructing Amazon Holocene vegetation histories and pre-Columbian land use, particularly the high spatial resolution possible with terrestrial soil-based phytolith studies.

Pollen-based differentiation of Amazonian rainforest communities and implications for lowland palaeoecology in tropical South America

An ongoing controversy in Amazonian palaeoecology is the manner in which Amazonian rainforest communities have responded to environmental change over the last glacial–interglacial cycle. Much of this controversy results from an inability to identify the floristic heterogeneity exhibited by rainforest communities within fossil pollen records. We apply multivariate (Principal Components Analysis) and classification (Unweighted Pair Group with Arithmetic Mean Agglomerative Classification) techniques to floral-biometric, modern pollen trap and lake sediment pollen data situated within different rainforest communities in the tropical lowlands of Amazonian Bolivia. Modern pollen rain analyses from artificial pollen traps show that evergreen terra firme (well-drained), evergreen terra firme liana, evergreen seasonally inundated, and evergreen riparian rainforests may be readily differentiated, floristically and palynologically. Analogue matching techniques, based on Euclidean distance measures, are employed to compare these pollen signatures with surface sediment pollen assemblages from five lakes: Laguna Bella Vista, Laguna Chaplin, and Laguna Huachi situated within the Madeira-Tapajós moist forest ecoregion, and Laguna Isirere and Laguna Loma Suarez, which are situated within forest patches in the Beni savanna ecoregion. The same numerical techniques are used to compare rainforest pollen trap signatures with the fossil pollen record of Laguna Chaplin.

Differentiation between Neotropical rainforest, dry forest and savannah ecosystems by their modern pollen spectra and implications for the fossil pollen record

Accurate differentiation between tropical forest and savannah ecosystems in the fossil pollen record is hampered by the combination of: i) poor taxonomic resolution in pollen identification, and ii) the high species diversity of many lowland tropical families, i.e. with many different growth forms living in numerous environmental settings. These barriers to interpreting the fossil record hinder our understanding of the past distributions of different Neotropical ecosystems and consequently cloud our knowledge of past climatic, biodiversity and carbon storage patterns. Modern pollen studies facilitate an improved understanding of how ecosystems are represented by the pollen their plants produce and therefore aid interpretation of fossil pollen records. To understand how to differentiate ecosystems palynologically, it is essential that a consistent sampling method is used across ecosystems. However, to date, modern pollen studies from tropical South America have employed a variety of methodologies (e.g. pollen traps, moss polsters, soil samples). In this paper, we present the first modern pollen study from the Neotropics to examine the modern pollen rain from moist evergreen tropical forest (METF), semi-deciduous dry tropical forest (SDTF) and wooded savannah (cerradão) using a consistent sampling methodology (pollen traps). Pollen rain was sampled annually in September for the years 1999–2001 from within permanent vegetation study plots in, or near, the Noel Kempff Mercado National Park (NKMNP), Bolivia. Comparison of the modern pollen rain within these plots with detailed floristic inventories allowed estimates of the relative pollen productivity and dispersal for individual taxa to be made (% pollen/% vegetation or ‘p/v’). The applicability of these data to interpreting fossil records from lake sediments was then explored by comparison with pollen assemblages obtained from five lake surface samples.

Palynological differentiation between genera of the Moraceae family and implications for Amazonian palaeoecology

The Moraceae family is one of the most abundant and ecologically important families in Neotropical rainforests and is very well-represented in Amazonian fossil pollen records. However, difficulty in differentiating palynologically between the genera within this family, or between the Moraceae and Urticaceae families, has limited the amount of palaeoecological information that can be extracted from these records. The aim of this paper is to analyse the morphological properties of pollen from Amazonian species of Moraceae in order to determine whether the pollen taxonomy of this family can be improved. Descriptive and morphometric methods are used to identify and differentiate key pollen types of the Moraceae (mulberry) and Urticaceae (nettle) families which are represented in Amazonian rainforest communities of Noel Kempff Mercado National Park (NKMNP), Northeast Bolivia. We demonstrate that Helicostylis, Brosimum, Pseudolmedia, Sorocea and Pourouma pollen can be identified in tropical pollen assemblages and present digital images of, and a taxonomic key to, the Moraceae pollen types of NKMNP. Indicator species, Maquira coriacea (riparian evergreen forest) and Brosimum gaudichaudii (open woodland and upland savanna communities), also exhibit unique pollen morphologies. The ability to recognise these ecologically important taxa in pollen records provides the potential for much more detailed and reliable Neotropical palaeovegetation reconstructions than have hitherto been possible. In particular, this improved taxonomic resolution holds promise for resolving long-standing controversies over the interpretation of key Amazonian Quaternary pollen records.

Applying Jackson’s methodological ideal-types: problems of differentiation and classification

In The Conduct of Inquiry in International Relations, Patrick Jackson situates methodologies in International Relations in relation to their underlying philosophical assumptions. One of his aims is to map International Relations debates in a way that ‘capture[s] current controversies’ (p. 40). This ambition is overstated: whilst Jackson’s typology is useful as a clarificatory tool, (re)classifying existing scholarship in International Relations is more problematic. One problem with Jackson’s approach is that he tends to run together the philosophical assumptions which decisively differentiate his methodologies (by stipulating a distinctive warrant for knowledge claims) and the explanatory strategies that are employed to generate such knowledge claims, suggesting that the latter are entailed by the former. In fact, the explanatory strategies which Jackson associates with each methodology reflect conventional practice in International Relations just as much as they reflect philosophical assumptions. This makes it more difficult to identify each methodology at work than Jackson implies. I illustrate this point through a critical analysis of Jackson’s controversial reclassification of Waltz as an analyticist, showing that whilst Jackson’s typology helps to expose inconsistencies in Waltz’s approach, it does not fully support the proposed reclassification. The conventional aspect of methodologies in International Relations also raises questions about the limits of Jackson’s ‘engaged pluralism’.

Repressor element 1 silencing transcription factor couples loss of pluripotency with neural induction and neural differentiation

Neural differentiation of embryonic stem cells (ESCs) requires coordinated repression of the pluripotency regulatory program and reciprocal activation of the neurogenic regulatory program. Upon neural induction, ESCs rapidly repress expression of pluripotency genes followed by staged activation of neural progenitor and differentiated neuronal and glial genes. The transcriptional factors that underlie maintenance of pluripotency are partially characterized whereas those underlying neural induction are much less explored, and the factors that coordinate these two developmental programs are completely unknown. One transcription factor, REST (repressor element 1 silencing transcription factor), has been linked with terminal differentiation of neural progenitors and more recently, and controversially, with control of pluripotency. Here, we show that in the absence of REST, coordination of pluripotency and neural induction is lost and there is a resultant delay in repression of pluripotency genes and a precocious activation of both neural progenitor and differentiated neuronal and glial genes. Furthermore, we show that REST is not required for production of radial glia-like progenitors but is required for their subsequent maintenance and differentiation into neurons, oligodendrocytes, and astrocytes. We propose that REST acts as a regulatory hub that coordinates timely repression of pluripotency with neural induction and neural differentiation.

Expression of the guanine nucleotide exchange factor, mr-gef, is regulated during the differentiation of specific subsets of telencephalic neurons

We have performed a screen combining subtractive hybridization with PCR to isolate genes that are regulated when neuroepithelial (NE) cells differentiate into neurons. From this screen, we have isolated a number of known genes that have not previously been associated with neurogenesis, together with several novel genes. Here we report that one of these genes, encoding a guanine nucleotide exchange factor (GEF), is regulated during the differentiation of distinct neuronal populations. We have cloned both rat and mouse GEF genes and shown that they are orthologs of the human gene, MR-GEF, which encodes a GEF that specifically activates the small GTPase, Rap1. We have therefore named the rat gene rat mr-gef (rmr-gef) and the mouse gene mouse mr-gef (mmr-gef). Here, we will collectively refer to these two rodent genes as mr-gef. Expression studies show that mr-gef is expressed by young neurons of the developing rodent CNS but not by progenitor cells in the ventricular zone (VZ). The expression pattern of mr-gef during early telencephalic neurogenesis is strikingly similar to that of GABA and the LIM homeobox gene Lhx6, a transcription factor expressed by GABAergic interneurons generated in the ventral telencephalon, some of which migrate into the cortex during development. These observations suggest that mr-gef encodes a protein that is part of a signaling pathway involved in telencephalic neurogenesis; particularly in the development of GABAergic interneurons.

Good and bad increases in ecological awareness: environmental differentiation revisited

We analyze a vertically differentiated market, assuming that conventional and green firms' products have different impacts on the environment. Heterogeneous consumers choose to be supplied by a conventional or a green firm, depending on their extra willingness to pay for a green product and the relative prices of the products in the market. We show that environmental awareness campaigns may have a negative impact on total welfare. This possibility is shown to exist without consumer misperceptions about the quality of green products and ruling out changes in the coverage and the structure of the market. Surprisingly, both conventional and green firms may benefit from heterogeneity-enhancing awareness campaigns, while social welfare is more likely to be enhanced by heterogeneity-reducing ones.

Cooperative R&D with endogenous technology differentiation

We study a nontournament R&D duopoly. Before the standard R&D investment and quantity-setting stages, we consider a stage in which firms choose their R&D technologies. Spillovers negatively depend on R&D technology differentiation. We show that, in equilibrium, firms will choose identical or very similar R&D processes. Such equilibria may entail less differentiation than would be dictated by social welfare maximization.

Regulating vertical relations in the presence of retailer differentiation costs

We discuss public policy towards vertical relations, comparing different types of contracts between a manufacturer and a maximum of two retailers. Together with (potential) price competition between the retailers, we study the role of a (sunk) differentiation cost paid by them in order to relax competition in the retail market and broaden the market potential of the distributed product. This non-price competition element in the downstream market is responsible for our conclusion that, unlike in standard policy guidelines and previous theoretical analysis, restrictions in intra-brand competition may deserve a permissive treatment even in the absence of inter-brand competition, if retailer differentiation is costly.

Genetic analysis reveals a novel locus in Vicia faba decoupling pigmentation in the flower from that in the extra-floral nectaries

The genetics of the stipule spot pigmentation (SSP) in faba bean (Vicia faba L.) was studied using four inbred lines, of which Disco/2 was zero-tannin (zt2) with colourless stipule spots, ILB938/2 was normal-tannin (ZT2) with colourless stipule spots, and both Aurora/2 and Mélodie/2 were ZT2 with coloured stipule spots. Crosses Mélodie/2 × ILB 938/2, Mélodie/2 × Disco/2, ILB 938/2 × Aurora/2 and ILB 938/2 × Disco/2 (A, B, C and D, respectively) were prepared, along with reciprocals and backcrosses, and advanced through single-seed descent. All F1 hybrid plants had pigmented stipule spots, and in the F2 generation, the segregation ratio fit 3 coloured:1 colourless in crosses A, B and C and 9:7 in cross D. In the F3 generation, the ratio fit 5:3 in crosses A and C and 25:39 in cross D, and in the F4 generation, 9:7 in cross A. SSP was linked to the zero-tannin characteristics (white flower) only in cross B. The results show that coloured stipule spot is dominant to colourless and that colouration is determined by two unlinked complementary recessive genes. We propose the symbols ssp2 for the gene associated with zt2 in Disco/2 and ssp1 for the gene not associated with tannin content in ILB938/2. The novel ssp1 locus was mapped at F5 in cross ‘A’ using Medicago truncatula-derived single-nucleotide polymorphism and was on chromosome 1 of faba bean, in a well-conserved region of M. truncatula chromosome 5 containing some candidate Myb and basic helix–loop–helix transcription factor genes.

Isotopic examination of links between diet, social differentiation and DISH at the post-medieval Carmelite Friary of Aalst, Belgium

Stable isotope ratios (δ13C and δ15N) were measured in human burials from the post-medieval (16th–18th c. AD) Carmelite friary burial grounds at Aalst, a town in Flanders, Belgium. Dietary patterns of 39 adult individuals were analyzed, from a mixed monastic and lay population buried in three different locations, reflecting groups with differing social status. The data show significant variation in the consumption of perhaps meat, but certainly also marine protein between females and males. This result represents a remarkable continuity with medieval dietary patterns, suggesting that the social and economic changes of the early modern period had a limited effect on everyday life. When both sexes were examined together, individuals buried in the cloister garth consumed significantly less marine protein compared to people buried in the church, likely reflecting social stratification. No statistical differences were observed between isotopic values from the church and the cloister alley, suggesting a similarly diverse diet of the monastic part of the buried population and that of the richer lay population. Finally, the hypothesis that diffuse idiopathic skeletal hyperostosis (DISH) is linked to a diet rich in animal protein was tested. No systematic or statistically significant differences between pathological and non-pathological bones from the same individuals affected with DISH were observed, and no statistical differences were found between individuals with DISH and individuals without DISH

Highly efficient neural differentiation of human somatic stem cells, isolated by minimally invasive periodontal surgery

Neural stem cells (NSCs) are potential sources for cell therapy of neurodegenerative diseases and for drug screening. Despite their potential benefits, ethical and practical considerations limit the application of NSCs derived from human embryonic stem cells (ES) or adult brain tissue. Thus, alternative sources are required to satisfy the criteria of ready accessibility, rapid expansion in chemically defined media and reliable induction to a neuronal fate. We isolated somatic stem cells from the human periodontium that were collected during minimally invasive periodontal access flap surgery as part of guided tissue regeneration therapy. These cells could be propagated as neurospheres in serum-free medium, which underscores their cranial neural crest cell origin. Culture in the presence of epidermal growth factor (EGF) and fibroblast growth factor-2 (FGF-2) under serum-free conditions resulted in large numbers of nestin-positive/Sox-2-positive NSCs. These periodontium-derived (pd) NSCs are highly proliferative and migrate in response to chemokines that have been described as inducing NSC migration. We used immunocytochemical techniques and RT-PCR analysis to assess neural differentiation after treatment of the expanded cells with a novel induction medium. Adherence to substrate, growth factor deprivation, and retinoic acid treatment led to the acquisition of neuronal morphology and stable expression of markers of neuronal differentiation by more than 90% of the cells. Thus, our novel method might provide nearly limitless numbers of neuronal precursors from a readily accessible autologous adult human source, which could be used as a platform for further experimental studies and has potential therapeutic implications.

Prolonged cultivation of hippocampal neural precursor cells shifts their differentiation potential and selects for aneuploid cells

Neural precursor cells (NPCs) are lineage-restricted neural stem cells with limited self-renewal, giving rise to a broad range of neural cell types such as neurons, astrocytes, and oligodendrocytes. Despite this developmental potential, the differentiation capacity of NPCs has been controversially discussed concerning the trespassing lineage boundaries, for instance resulting in hematopoietic competence. Assessing their in vitro plasticity, we isolated nestin+/Sox2+, NPCs from the adult murine hippocampus. In vitro-expanded adult NPCs were able to form neurospheres, self-renew, and differentiate into neuronal, astrocytic, and oligodendrocytic cells. Although NPCs cultivated in early passage efficiently gave rise to neuronal cells in a directed differentiation assay, extensively cultivated NPCs revealed reduced potential for ectodermal differentiation. We further observed successful differentiation of long-term cultured NPCs into osteogenic and adipogenic cell types, suggesting that NPCs underwent a fate switch during culture. NPCs cultivated for more than 12 passages were aneuploid (abnormal chromosome numbers such as 70 chromosomes). Furthermore, they showed growth factor-independent proliferation, a hallmark of tumorigenic transformation. In conclusion, our findings substantiate the lineage restriction of NPCs from adult mammalian hippocampus. Prolonged cultivation results, however, in enhanced differentiation potential, which may be attributed to transformation events leading to aneuploid cells.