Determinants of biological activity of nanomaterials in innate immunity cells: the role of aspect ratio, environmental contaminants and protein corona

As defined by the European Union, “ ’Nanomaterial’ (NM) means a natural, incidental or manufactured material containing particles, in an unbound state or as an aggregate or agglomerate, where, for 50 % or more of the particles in the number size distribution, one or more external dimensions is in the size range 1 nm-100 nm ” (2011/696/UE). Given their peculiar physico-chemical features, nanostructured materials are largely used in many industrial fields (e.g. cosmetics, electronics, agriculture, biomedical) and their applications have astonishingly increased in the last fifteen years. Nanostructured materials are endowed with very large specific surface area that, besides making them very useful in many industrial processes, renders them very reactive towards the biological systems and, hence, potentially endowed with significant hazard for human health. For these reasons, in recent years, many studies have been focused on the identification of toxic properties of nanostructured materials, investigating, in particular, the mechanisms behind their toxic effects as well as their determinants of toxicity. This thesis investigates two types of nanostructured TiO2 materials, TiO2 nanoparticles (NP), which are yearly produced in tonnage quantities, and TiO2 nanofibres (NF), a relatively novel nanomaterial. Moreover, several preparations of MultiWalled Carbon Nanotubes (MWCNT), another nanomaterial widely present in many products, are also investigated.- Although many in vitro and in vivo studies have characterized the toxic properties of these materials, the identification of their determinants of toxicity is still incomplete. The aim of this thesis is to identify the structural determinants of toxicity, using several in vitro models. Specific fields of investigation have been a) the role of shape and the aspect ratio in the determination of biological effects of TiO2 nanofibres of different length; b) the synergistic effect of LPS and TiO2 NP on the expression of inflammatory markers and the role played therein by TLR-4; c) the role of functionalization and agglomeration in the biological effects of MWCNT. As far as biological effects elicited by TiO2 NF are concerned, the first part of the thesis demonstrates that long TiO2 nanofibres caused frustrated phagocytosis, cytotoxicity, hemolysis, oxidative stress and epithelial barrier perturbation. All these effects were mitigated by fibre shortening through ball-milling. However, short TiO2 NF exhibited enhanced ability to activate acute pro-inflammatory effects in macrophages, an effect dependent on phagocytosis. Therefore, aspect ratio reduction mitigated toxic effects, while enhanced macrophage activation, likely rendering the NF more prone to phagocytosis. These results suggest that, under in vivo conditions, short NF will be associated with acute inflammatory reaction, but will undergo a relatively rapid clearance, while long NF, although associated with a relatively smaller acute activation of innate immunity cells, are not expected to be removed efficiently and, therefore, may be associated to chronic inflammatory responses. As far as the relationship between the effects of TiO2 NP and LPS, investigated in the second part of the thesis, are concerned, TiO2 NP markedly enhanced macrophage activation by LPS through a TLR-4-dependent intracellular pathway. The adsorption of LPS onto the surface of TiO2 NP led to the formation of a specific bio-corona, suggesting that, when bound to TiO2 NP, LPS exerts a much more powerful pro-inflammatory effect. These data suggest that the inflammatory changes observed upon exposure to TiO2 NP may be due, at least in part, to their capability to bind LPS and, possibly, other TLR agonists, thus enhancing their biological activities. Finally, the last part of the thesis demonstrates that surface functionalization of MWCNT with amino or carboxylic groups mitigates the toxic effects of MWCNT in terms of macrophage activation and capability to perturb epithelial barriers. Interestingly, surface chemistry (in particular surface charge) influenced the protein adsorption onto the MWCNT surface, allowing to the formation of different protein coronae and the tendency to form agglomerates of different size. In particular functionalization a) changed the amount and the type of proteins adsorbed to MWCNT and b) enhanced the tendency of MWCNT to form large agglomerates. These data suggest that the different biological behavior of functionalized and pristine MWCNT may be due, at least in part, to the different tendency to form large agglomerates, which is significantly influenced by their different capability to interact with proteins contained in biological fluids. All together, these data demonstrate that the interaction between physico-chemical properties of nanostructured materials and the environment (cells + biological fluids) in which these materials are present is of pivotal importance for the understanding of the biological effects of NM. In particular, bio-persistence and the capability to elicit an effective inflammatory response are attributable to the interaction between NM and macrophages. However, the interaction NM-cells is heavily influenced by the formation at the nano-bio interface of specific bio-coronae that confer a novel biological identity to the nanostructured materials, setting the basis for their specific biological activities.

An examination of the role of the concept of the community within the current reorganisation of English local government

The thesis will examine the role and impact of the concept of the community within the structural reorganisation of English local government between 1992 and 1995. The methodological approach adopted within this thesis has been to compare the use, application and significance of the community with a case study of a specific local authority and its preparations for reorganisation. The authority in question was Wychavon District Council located in the County of Hereford and Worcester. The conclusions from this case study were then compared to the role and significance of the community in the reviews of other local authorities in England. This study produced two important results. These were that there was an established body of literature which argued that the community could be of value to local government and that the community should be identified by measuring individuals sense of belonging and feelings of attachment, as well as such daily activities as shopping and working (which help to stimulate these feelings). The then Conservative Government even instructed the specially appointed Commission to apply this particular interpretation of the community to their reviews, and to attempt to base any new unitary authorities upon the social and spatial area it created. The Conservative Government also gave the Commission a Community Index to assist with the identification of communities, and appointed the pollsters MORI to support the Commission with the task of identifying the emotional and more subjective senses of community. The Commission eventually came to rely entirely on the MORI polls, and whilst these polls attempted to faithfully apply the Governments interpretation of the community, they unfortunately produced small and often complex communities, which the Commission felt could not be applied to its reviews. This therefore led to the community becoming a secondary consideration to the factors of cost and efficiency. Furthermore the problematic nature of the community - that is, the production of small and complex communities - was repeated in this thesis' own survey of community identities in the District of Wychavon. In fact this authority's proposals for reorganisation were based almost entirely upon the factors of cost, size and efficiency.

The role of government research establishments : a study of the concept of public patronage for applied research and development

This thesis follows the argument that, to fully understand the current position of national research laboratories in Great Britain one needs to study the historical development of the government research establishment as a specific social institution. A particular model is outlined in which it is argued that institutional characteristics evolve through the continual interplay between internal development and environmental factors within a changing political and economic context, and that the continuous development of an institution depends on its ability to adapt to changes in its operational environment. Within this framework important historical precedents for formal government institutional support for applied research are identified. and the transition from private to public patronage documented. The emergence and consolidation of government research laboratories in Britain is described in detail. The subsequent relative decline of public laboratories is interpreted in terms of the undermining of a traditional role resulting in legitimation crisis. It is concluded that it is no longer feasible to consider the public research laboratory as a coherent institutional form, and that the future of each individual laboratory can only be considered in relation to the institutional needs of its own sphere of operation. Nevertheless the laboratories have been forced into decline in an essentially unplanned way which may have serious consequences for the maintenance of the scientific and technical infrastructures, necessary for material progress in the national context.