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Chrysotile asbestos is a carcinogenic mineral that has abundantly been used in industrial and consumer applications. The carcinogenicity of the fibers is partly governed by reactive Fe surface sites that catalyze the generation of highly toxic hydroxyl radicals (HO

The natural occurrence of asbestos and asbestos-like minerals (NOA) poses a risk to the environment and human health, notably when natural processes and anthropic activities promote fibre dispersion. Hundreds of potentially hazardous elongate mineral particles (EMPs, NIOSH 2011 definition) exist, and their toxicological profile is often unknown. We aim here to define a general approach, from field analysis to nano-structural investigation, to assess whether a fibrous mineral occurring in a specific site could pose a risk to human health. To evaluate the hazard associated with NOA, a multi-scale and multi-analytical integrated approach was adopted. Specifically, the geological factors that control the occurrence and distribution of NOA on site, and the mechanisms of formation and liberation of airborne fibres were investigated. In parallel, we explored the key bulk and surface properties of several natural mineral fibres and defined crystallographic, chemical, and morphological aspects that should be considered during hazard assessment. Also, the effect of standardized mechanical stress was used to quantitatively evaluate the potency of NOA-bearing exposed rock to generate inhalable fibres. This property was connected with the mineral characteristic, the rock fabric, and the rock erosion rate. Isolated fibre specimens were used to assess solubility in simulated body fluids, surface reactivity, and toxicological endpoints in vitro and in vivo. Taken together, these findings allowed us to build a multidimensional description of the hazard parameters of mineral fibre and paved the way for a science-based risk assessment in an unexplored NOA site.

Crystalline silica (CS) is a well-known toxic particle that may cause severe pathologies including silicosis, lung cancer, and several autoimmune diseases. [1a,b] The hazard associated to crystalline silica is extremely variable and depends on some specific characteristics, including crystal structure and surface chemistry.[1c] In particular, a specific family of surface silanols, called nearly free silanols (NFS), has been recently related to the interaction mechanisms occurring between quartz particles and cell membrane components, which initiate the lung inflammatory reaction. [2] Even though this phenomenon has been studied for decades on quartz, the research on other silica polymorphs was limited, also because of the relative low abundance of some polymorphs. The CS polymorphs, i.e., quartz, cristobalite, tridymite, coesite, and stishovite, share the SiO2 stoichiometry and differentiate for crystal structure. [3] Thus, the different crystal lattices expose differently ordered hydroxyl group patterns at the crystal surface. We proved that the NFS occur and take part in the molecular bio-interactions, not only on quartz, but also on the other CS polymorphs. Five high-purity samples representative of the five CS polymorphs were fully characterized by XRPD, Scanning Electron Microscopy, and IR spectroscopy. When CS polymorphs were contacted with model membranes (red blood cells), all of them were able to disrupt cell membranes, except stishovite, which was the only polymorph without NFS. By thermally modulating the topochemistry of surface silanols, it was possible to show that the membranolytic activity of the CS polymorphs quantitatively paralleled the occurrence of NFS. This observations confirmed the central role of NFS in regulating the interaction of silica with biomembranes. In conclusion, these results put the surface characteristics of CS particles in the foreground with respect to the crystal habit and provide a comprehensive understanding of the molecular mechanisms associated with silica hazard and bio-minero-chemical interfacial phenomena. [4][1] a) KAWASAKI, H. 2015, Inhal Toxicol, 27, 363-77; b) IARC 2012, IARC monographs on the evaluation of carcinogenic risks to humans, Lyon, World Health 543 Organisation c). BORM, P. J. A., et al. 2018, Part. Fibre. Toxicol., 15, 23;[2] PAVAN, C., et al., 2020, Proc Natl Acad Sci U S A, 117, 27836-27846.[3] GUTHRIE, G. D. & HEANEY, P. J. 1995, Scand J Work Environ Health, 21 Suppl 2, 5-8.[4] PAVAN. C., et al., 2023, accepted on Frontiers in Chemistry, DOI: 10.3389/fchem.2022.1092221.

Studies on naturally occurring asbestos (NOA) and on the relevant geo-environmental problems have been traditionally focused on metamorphic rocks (and, more recently, on magmatic rocks). Besides these 'primary' occurrences (i.e., those related to the in situ growth of NOA minerals), 'secondary', detrital NOA may occur in sediments, sedimentary rocks and soils derived from the erosion of 'primary' NOA bearing rocks.The occurrence of detrital NOA in sediments and soils is increasingly recognized worldwide. However, a few studies exist that investigate the 'sedimentology of NOA', i.e., the mechanisms underlying the genesis, transport, deposition and post-depositional modifications of detrital NOA particles in the different sedimentary environments. A better understanding of these mechanisms would give us the tools to predict the presence and possible concentration of detrital NOA in sediments and sedimentary rocks.The occurrence and distribution of detrital NOA within the Oligocene-Miocene succession of the southern Tertiary Piemonte Basin (NW Italy), will be investigated as a scientific development in the frame of the CARG project (Geological Mapping at 1:50,000 scale - sheet 195 Novi Ligure). This succession consists of stratigraphic units rich in ophiolite clasts and deposited in a variety of sedimentary environments, from continental to deep marine, thus representing an ideal study case.

Although epidemiological studies have suggested an association between asbestos exposure and systemic autoimmunity, the underlying mechanisms are poorly understood. Short asbestos fibers are often considered less harmful than long fibers but such a statement is still a matter of debates. This study aimed to compare the effects of short (SFA) and long (LFA) amosite fiber exposure on lung damage, autoimmunity and macrophage phenotype. Four months after lung exposure to 0.1 mg of fibers, BAL levels of lactate dehydrogenase, free DNA, CCL2, TIMP-1 and immunoglobulins A of LFA-exposed C57Bl/6 mice were increased when compared to fluids from control- and SFA-exposed mice. Effects in LFA-exposed mice were associated with lung fibrosis and autoimmunity including anti-double-strand DNA antibody production. Human monocyte-derived macrophages (MDMs) exposed to SFA or LFA at 20 µg/cm2 have a pro-inflammatory phenotype characterized by a significant increase of TNFα and IL-6 secretion. A decrease of efferocytosis capacities was also noted after SFA and LFA, whereas macrophage abilities to phagocyte fluorescent beads were unchanged when compared to control MDMs. SFA exposure induced IL-6 secretion and reduced the percentage of MDMs expressing MHCII and CD86 markers involved in antigen and T-lymphocyte stimulation. By contrast, NLRP3 inflammasome activation, evaluated through quantification of caspase-1 activity and IL-1β secretion is rather associated to LFA than SFA exposure. Our results demonstrated that only long-term exposure to LFA has induced significant lung damages and autoimmune effects supporting a worsened health effects of LFA in comparison to SFA.

The natural occurrence of asbestos and asbestos-like minerals (NOA) poses a risk to the environment and human health, notably when natural processes and anthropic activities promote fibre dispersion. Hundreds of potentially hazardous elongated mineral particles (EMPs, NIOSH 2011 definition) exist, and their toxicological profile is often unknown. We aim here to define a general approach, from field analysis to nano-structural investigation, to assess whether a fibrous mineral occurring in a specific site could pose a risk to human health. To evaluate the hazard associated with NOA, a multi-scale and multi-analytical integrated approach was adopted. Specifically, the geological factors that control the occurrence and distribution of NOA on site, and the mechanisms of formation and liberation of airborne fibres were investigated. In parallel, we explored the key bulk and surface properties of several natural mineral fibres and defined crystallographic, chemical, and morphological aspects that should be considered during hazard assessment. Also, the effect of standardized mechanical stress was used to simulate variation on morphometric parameters of investigated minerals and allowed us to quantitatively evaluate the potency to generate inhalable fibres. This property was connected with the mineral characteristic and the rock fabric. Isolated fibre specimens were used to assess solubility in simulated body fluids, surface reactivity, and toxicological endpoints in vitro and in vivo. Taken together, these findings allowed us to build a multidimensional description of the hazard parameters of a mineral fibre and paved the way for a science-based risk assessment in an unexplored NOA site.

Crystalline silica (CS) is a well-known human carcinogen and freshly fractured CS is strongly held to be more toxic than aged dust. Mechanical fracturing indeed generates on CS surface a specific family of Nearly-Free Silanols (NFS), a moiety that was proved to destabilize cell membranes and initiate inflammation in vivo. During milling, silica nanoparticles are also generated, however their contribution to the overall toxicity of quartz is still largely unexplored. Nanoquartz can be synthetized via bottom-up methods, but the surface chemistry of those crystals strongly differs from nanoparticles resulting from fracturing. We report here a top-down milling procedure to obtain a nanometric quartz that shares with fractured quartz the key surface properties relevant to toxicity. Fracturing procedure was optimized by coupling dry and wet milling steps, using water as a dispersing agent, and varying milling times and rotational speeds. We obtained a set of samples that would be classified as nanomaterials under EU CLP regulation (>50% of particles in number are < 100 nm) and exhibited a strong tendency to form submicrometric agglomerates. Deagglomeration with surfactants or simulated body fluids was negligible. Partial lattice amorphization and bimodal crystallite domain size were observed, as confirmed by the presence of two distinct domains of scattering with nanometric (< 50 nm) and submicrometric (0.8-1 µm) crystallite size. A moderate membranolytic activity, which nicely correlated with the amount of surface NFS, signalled that our nanoquartz may induce inflammation in vivo. Overall, a membranolytic nanoquartz for investigating the toxic activity of nanometric silica was obtained.

Occupational exposure to quartz dust is associated with fatal diseases. Quartz dusts generated by mechanical fracturing are characterized by a broad range of micrometric to nanometric particles. The contribution of this nanometric fraction to the overall toxicity of quartz is still largely unexplored, primarily because of the strong electrostatic adhesion forces that prevent isolation of the nano-fraction. Furthermore, fractured silica dust exhibits special surface features, namely Nearly Free Silanols (NFS), which impart membranolytic activity to quartz. Nanoquartz can be synthetized via bottom-up methods, but the surface chemistry of those crystals strongly differs from nanoparticles resulting from fracturing. We report here a top-down milling procedure to obtain a nanometric quartz that shares with fractured quartz the key surface properties relevant to toxicity. Ball milling was optimized by coupling dry and wet milling steps, using water as a dispersing agent, and varying milling times and rotational speeds. Nanoquartz with a strong tendency to form submicrometric agglomerates was obtained. Deagglomeration with surfactants or simulated body fluids was negligible. Partial lattice amorphization and bimodal crystallite domain size were observed. A moderate membranolytic activity, which correlated with the amount of NFS, signaled coherence with previous toxicological data. Membranolytic nanoquartz for toxicological investigations was obtained.

Exposure to asbestos and asbestos-like minerals has been related to the development of severe lung diseases, including cancer and malignant mesothelioma (MM). A high incidence of non-occupational MM was observed in New Caledonia (France) in people living in proximity of serpentinite outcrops, containing chrysotile and fibrous antigorite. Antigorite is a magnesium silicate, which shares with chrysotile asbestos the chemical formula. To achieve information on antigorite toxicity, we investigated the physico-minero-chemical features relevant for toxicity and cellular effects elicited on murine macrophages (MH-S) and alveolar epithelial cells (A549) of three fibrous antigorites (f-Atg) collected in a Caledonian nickel lateritic ore and subjected to supergene alteration. Field Atg were milled to obtain samples suitable for toxicological studies with a similar particle size distribution. UICC chrysotile (Ctl) and a non-fibrous antigorite (nf-Atg) were used as reference minerals. A high variability in toxicity was observed depending on shape, chemical alteration, and surface reactivity. The antigorites shared with Ctl a similar surface area (16.3, 12.1, 20.3, 13.4, and 15.6 m

Respirable crystalline silica (RCS) is the leading cause of occupational respiratory diseases worldwide. RCS is associated with inflammatory lung reactions, which can lead to silicosis, cancer, and autoimmune diseases. The extreme variability of silica specimens, including its amorphous forms, the surface heterogeneity, and variable toxicity effects, generated one of the most intriguing enigmas in particle toxicology, i.e., deciphering which physico-chemical features explain and predict the variable hazard of silica. Using a set of ad hoc prepared synthetic and natural quartz particles, we have identified a unique subfamily of surface silanols as a key initiator of the toxicity of silica particles. These moieties, namely the "nearly-free silanols" (NFS), appear on the surface of quartz when crystals are fractured, and their amount can be modulated by physico-chemical processes. The peculiar spatial arrangement of NFS was demonstrated as the most energetically favorable and selective for interacting zwitterionic phospholipids that make up cell membranes. The toxic activity of NFS was also confirmed with amorphous nanosilica synthesized at high temperatures, and our recent findings suggest that NFS could impart toxic properties to other silica polymorphs and hydroxylated surfaces, including aluminosilicates and engineered stones. Overall, NFS occurrence accounts for the origin and variability of the toxicity of silica, opening perspectives for controlling RCS hazard, and may contribute to understand other important interfacial phenomena of hydroxylated materials.

The pathogenicity of quartz involves lysosomal alteration in alveolar macrophages. This event triggers the inflammatory cascade that may lead to quartz-induced silicosis and eventually lung cancer. Experiments with synthetic quartz crystals recently showed that quartz dust is cytotoxic only when the atomic order of the crystal surfaces is upset by fracturing. Cytotoxicity was not observed when quartz had as-grown, unfractured surfaces. These findings raised questions on the potential impact of quartz surfaces on the phagolysosomal membrane upon internalization of the particles by macrophages. To gain insights on the surface-induced cytotoxicity of quartz, as-grown and fractured quartz particles in respirable size differing only in surface properties related to fracturing were prepared and physico-chemically characterized. Synthetic quartz particles were compared to a well-known toxic commercial quartz dust. Membranolysis was assessed on red blood cells, and quartz uptake, cell viability and effects on lysosomes were assessed on human PMA-differentiated THP-1 macrophages, upon exposing cells to increasing concentrations of quartz particles (10–250 µg/ml). All quartz samples were internalized, but only fractured quartz elicited cytotoxicity and phagolysosomal alterations. These effects were blunted when uptake was suppressed by incubating macrophages with particles at 4 °C. Membranolysis, but not cytotoxicity, was quenched when fractured quartz was incubated with cells in protein-supplemented medium. We propose that, upon internalization, the phagolysosome environment rapidly removes serum proteins from the quartz surface, restoring quartz membranolytic activity in the phagolysosomes. Our findings indicate that the cytotoxic activity of fractured quartz is elicited by promoting phagolysosomal membrane alteration.

The “Geological map of Naturally Occurring Asbestos of Piemonte region” at 1:250,000 scale aims at reviewing the distribution of NOA-bearing rocks and the individual occurrences of NOA minerals of Piemonte at the regional scale. A geo-lithological basemap was produced ad hoc through a reasoned simplification of the basic Geological Map of Piemonte and its geo-database (GeoPiemonteMap, Piana et al., 2017; also available as a WebGIS service at https://webgis.arpa.piemonte.it/Geoviewer2D/index.html?config=other-configs/geologia250k_config.json). Particular emphasis was given to the potentially NOA-bearing rocks, i.e., meta-ophiolites and sedimentary successions containing meta-ophiolite clasts. The map reports hundreds of punctual occurrences of NOA minerals, as well as the main former asbestos mining sites. The NOA occurrences dataset derives from a thorough revision of the regional geological literature, integrated with authors' original data acquired during several years of geological and geo-environmental surveys.The map is supported by a geo-database, compliant with the geo-datsabase of basic Geological Map of Piemonte. Each mapped NOA occurrence corresponds to an item of the geo-database, containing the following information: NOA mineral(s) species, locality and coordinates, occurrence type (i.e., in vein/in the rock matrix/detrital), host rock lithology, meso- and micro-scale description (e.g., vein thickness, associated minerals, etc.), NOA mineral identification method (e.g, optical microscopy, microRaman spectroscopy, XRD, etc...), bibliographic reference, and sample availability (i.e., presence of samples in university or museum collections).References:Piana et al. (2017), Journal of Maps 13: 395-405, DOI: 10.1080/17445647.2017.1316218

Studies on naturally occurring asbestos (NOA) and on the relevant geo-environmental problems have been traditionally focused on metamorphic rocks (and, more recently, on magmatic rocks). Besides these 'primary' occurrences (i.e., those realted to the in situ growth of NOA minerals), 'secondary', detrital NOA may occurr in sediments, sedimentary rocks and soils derived form the erosion of 'primary' NOA bearing rocks.The occurrence of detrital NOA in sediments and soils is increasingly recognized worldwide. However, a few studies exist that investigate the 'sedimentology of NOA', i.e., the mechanisms underlying the genesis, transport, deposition and post-depositional modifications of detrital NOA particles in the different sedimentary environments. A better understanding of these mechanisms would give us the tools to predict the presence and possible concentration of detrital NOA in sediments and sedimentary rocks.The occurrence and distribution of detrital NOA within the Oligocene-Miocene succession of the southern Tertiary Piemonte Basin (NW Italy), will be investigated as a scientific development in the frame of the CARG project (Geological Mapping at 1:50,000 scale - sheet 195 Novi Ligure). This succession consists of stratigraphic units rich in ophiolite clasts and deposited in a variety of sedimentary environments, from continental to deep marine, thus representing an ideal study case.

Respirable crystalline silica (RCS) is the leading cause of occupational respiratory disease worldwide. RCS is associated with silicosis, cancer, and autoimmune diseases [1]. Silica (SiO2) is a simple, yet structurally very complex oxide and tens of variable crystalline and amorphous forms exist with different structures and surfaces. Structural heterogeneity is reflected in variable toxic effects, and this in turn generates one of the most intriguing enigmas in particle toxicology, i.e., deciphering the exact molecular nature of the interaction between silica and biological matter.A large set of synthetic and natural, crystalline and amorphous, micrometric and nanometric silica particles were prepared, modified, and characterized. The interaction of these surface-modified silicas with membrane systems of decreasing molecular complexity, e.g., red blood cells, liposomes, and phospholipid supramolecular structures (PLS), was investigated and compared with the results of in vitro and in vivo particle toxicity assessment.A specific silanol (≡Si-OH) sub-group at silica surface, the “nearly free silanols” (NFS), was evidenced as the cause for the membranolytic and inflammatory effect of silica [2]. Silica powders with NFS-rich surfaces caused RBC membrane lysis, and selectively perturbed liposomes and adsorbed PLS. Specific amino groups exposed at the membrane surface are proposed as recognition epitopes for the selective interaction with NFS, which are in turn proposed as the molecular pattern that defines the interaction of silica with biomembranes [3]. Our findings open a new perspective for tailoring less toxic silica particles and for designing improved technological applications of silica. NFS and hydroxylated surface moieties may be also relevant for the toxicity of other respirable mineral dusts, suggesting a new paradigm for particle toxicity mechanism.References[1] Leung et al., Lancet 2012, 379, 2008; [2] Pavan et al., Proc. Natl. Acad. Sci USA 2020, 117, 27836; [3] Pavan et al., sumbitted to ACS Central Science

Exposure to asbestos is known for inducing inflammation, pulmonary fibrosis, lung cancer and malignant mesothelioma. Although several novel biomarkers and treatments are being tested, these chronic disorders are currently incurable, and the prognosis is particularly poor. The development of asbestos-induced cancer results from mutations, cell transformation and proliferation caused by reactive oxygen species and elevated levels of pro-inflammatory cytokines. Fiber-induced chronic inflammation also explains asbestosis, an interstitial lung disease characterized by uncontrolled matrix protein deposition leading to detrimental lung fibrosis. The investigation of the in-vivo molecular mechanisms driving asbestos pathogenicity is still a matter of debate. In this context, it is accepted that the physico-chemical properties of the fibers play a crucial role in causing adverse effects, and long fibers are still considered more toxic than short ones. Indeed, when fibers reach the alveolar space and migrate to the pleural/peritoneal cavity, long and thin fibers shows stronger inflammogenic, fibrogenic and tumorigenic effects than short fibers, in the long term. It is largely hold that short asbestos fibers are more easily cleared from the lungs and elicit a lower reactional and/or inflammatory effect. To further investigate this paradigm of toxicity, we compared the pro-inflammatory and pro-fibrogenic potential of short and long amosite fibers in in-vitro (J774 murine macrophages) and in-vivo (C57BL/6 mice) models. Surprisingly, our results demonstrated that short fibers were more prone to induce in-vitro cytotoxicity, accompanied by the release of pro-inflammatory biomarkers, in comparison to long fibers. On the contrary, the long fibers were significantly more inflammogenic and fibrogenic in the lungs of treated mice, while the short fibers were almost inert and did not induce acute and chronic inflammation and fibrosis. Furthermore, we observed that, while the long fibers were still present in the lungs of the animals 4 months after the exposure, the short amosite was substantially absent. These findings imply that the pulmonary deposition and a defect of clearance of the fibers play a crucial role in the development of in-vivo detrimental effects associated to long asbestos. This effect overcomes the mere in-vitro cytotoxic and inflammogenic potential of short fibers. Present results provide new insights into the mechanisms that drive asbestos toxicity, opening new perspectives for the development of reliable in vitro tests that fully predict health adverse effects associated to inorganic mineral fibers.

Volcanic ashes from the Pomici di Avellino (PdA) eruption, an Early Bronze Age (ca. 3.9 ka BP) Plinian event from Somma-Vesuvius, had a wide dispersal area including most of the Central-South regions of Italy. The finest fraction of volcanic ashes can impact the human respiratory apparatus and induce severe respiratory difficulties and pathologies. Particle size, shape and composition, as well as surface reactivity are the key properties that define health hazard of volcanic ashes. Recent studies evidenced that volcanic ashes could generate significant amounts of free radicals. The particle-derived free radicals can contribute, together with the reactive species (ROS) produced by the cells, to the onset of oxidative stress. To evaluate the potential health impact of volcanic ashes both in proximal and distal areas we examined some physical and chemical characteristics of PdA ashes that may play a role in the onset of adverse health effects. Specifically, the particle size distribution (PSD) and surface properties (the specific surface area (SSA), the mobilization of bio-accessible iron ions and the ability to generate hydroxyl radicals) of PdA ashes have been measured. PSD was obtained by automated image analysis coupled with electron microscopy. Bio-available iron was quantified using specific iron chelator and colorimetric reactions. The physisorption of N2 at 77 K (N2 BET method) was used to measure the specific surface area. Electron Paramagnetic Resonance (EPR) coupled with spin trapping technique was used to quantify the ·OH radical generation. The results show a high surface area value, the presence of an amount of removable iron, and a reactivity in the formation of hydroxyl radicals. Furthermore, the ·OH radical generation is continuous in time. In particular, PdA volcanic ashes reactivity is probably related to the presence of surface bio-accessible iron, which is able to generate free radicals. Thus, free radical generation could make Mt. Vesuvius ashes potentially toxic and threaten people’s health causing respiratory problems. Overall, the physico-chemical characteristics of ash particles from Somma-Vesuvius might pose a significant health hazard even in distal areas, where the transport and the inhalation of fine-grained ashes from explosive eruptions might cause respiratory diseases. In light of these new data, further investigation (e.g. crystalline silica amount) to assess Somma-Vesuvius ashes toxicity is required.

Crystalline silica (CS) is a well-known human toxicant and inhalation of the airborne particles with size lower than 4 µm is associated to severe occupational diseases, such as silicosis and lung cancer.1 The International Agency for the Research on Cancer (IARC) classified CS as carcinogenic to humans and freshly fractured CS is held to be more toxic than aged dust.1,2 Fracturing generates on CS a specific family Nearly-Free Silanols (NFS), which are able to destabilize cell membranes,3 and some nanometric particles. We aim here to create and assess the possible toxicological impact and the chemical characteristics of the nanometric fraction (nano-CS) formed when CS is fractured.A highly pure CS of synthetic origin4 (α-quartz, micrometric in size) was ball-milled to obtain ultrafine particles. We coupled a dry milling step and a wet milling step, using water as dispersing agent, and we generated particles with specific surface area (SSA) ranging from 37 to 60 m2/g. These SSA values signaled the generation of a relevant nanometric fraction. The increase in SSA paralleled the energy delivered to quartz during the milling, that exceeded by far the energies commonly used in industrial processing. Morphology, crystallinity, size, surface silanols, including quantification of NFS, and membranolytic activity toward red blood cells were assessed. The nano-CS samples exhibited: i) a partial lattice amorphization that increased with the increase of the milling energy; ii) the presence of two distinct domains of scattering that indicated the occurrence of crystallite with nanometric (< 50 nm) and submicrometric (0.8-1 µm) size; iii) a strong tendency to form micrometric agglomerates , which could be partially dispersed with ultrasounds and surfactants in water suspensions; and iv) a moderate membranolytic activity that correlated with the presence of NFS. We selected a nano-CS sample that would be classified as a nanomaterial under EU CLP regulation (>50% of particles in number are < 100 nm).5 The nano-CS sample will be used in the next future as a reference material to quantify the nanometric fraction of silica powders and assess the potential exposure to nano-CS in industrial hygiene context.In conclusion, the preparation and characterization of nano-CS was achieved and the physico-chemical characteristics that could be relevant for silica toxicity were assessed. The nano-CS reference material will be used to quantify nano-CS in industrial scenario, and to clarify the toxic activity of nanometric silica obtained by mechanical fracturing. [1] IARC, Monograph Vol. 100C, 2012[2] Turci et al, Part Fibre Toxicol, 2016, 13, 32.[3] Pavan et al, Proc. Natl. Amer. Soc. USA, 2020, 117 (45), 27836[4] Pastero et al., 2016, Cryst. Growth Des. 16, 4, 2394–2403.[5] SCoEaNIHR, E. S., 2010, 'Scientific Basis for the Definition of the Term “nanomaterial”', European Commission.

The occurrence of asbestos and asbestos-like minerals in natural sites may pose a risk to human health and the environment when rocks and soils are mobilized. Weathering and anthropic activities favour the liberation of potentially hazardous Elongated Mineral Particles (EMP, NIOSH 2011). The definition of EMP includes both asbestos and other fibrous minerals. The latter share several physical-chemical properties with asbestos, but their toxicological profile is still unknown. The assessment of risk requires the quantification of the occurrence and the estimation of the potential emissivity of EMP from the hosting matrix.We quantitatively described the potency of a rock to disperse EMPs in the environment with a quantitative parameter namely the “liberability factor” (Lf). Lf was measured for 40 meta-ophiolite fragments from the NOA-bearing units of Liguria (Voltri Group and Sestri-Voltaggio Zone) and Calabria (Gimigliano-Monte Reventino Unit, Southern Ligurian Domain). The mineral–petrographic characterization of these rocks showed the presence of veins of chrysotile, fibrous tremolite-actinolite, fibrous sepiolite and fibrous antigorite.By adapting the UNI EN 12457-2:2004 method for solid waste, we designed a weathering simulation test to quantify the EMPs and the fibres (according to the World Health Organization) possibly liberated by applying to the rock a standardized mechanical stress. Waterborne EMPs were filtered on membranes and counted by electron microscopy (SEM-EDS), by adapting the Italian Regional Agency for the Protection of the Environment (ARPA) procedure for waterborne asbestos (ARPA Piemonte, 2016). We obtained Lf values as the number of waterborne fibres suspended per unit volume of water (fibres/Litre). All analysed rock samples showed Lf values ranging from 30 Mf/L to 21’000 Mf/L. Chrysotile, tremolite, sepiolite, and antigorite, with asbestos-like habit, were detected.Lf proved to be a reliable, easy to use method for the characterization and prediction of EMP and fibre dispersion in the environment from NOA-bearing rocks subjected to a standardized mechanical stress. This study is part of the BRIC 2019 project (grant number ID 57.1) supported by INAIL (Italian National Institute for Insurance against Accidents at Work). ReferencesARPA Piemonte 2016. U.RP. M842 rev.03. Asbestos in water by Scanning Electron Microscopy. NIOSH, 2011. Asbestos fibers and other elongate mineral particles. Current Intelligence Bulletin 62. WHO, 1997. Determination of Airborne Fibre Number Concentrations. ISBN 92 4 154496 1

Inhaled crystalline silica causes inflammatory lung diseases, but the mechanism for its unique activity compared to other oxides remains unclear, preventing the development of potential therapeutics. Here, the molecular recognition mechanism between membrane epitopes and "nearly free silanols" (NFS), a specific subgroup of surface silanols, is identified and proposed as a novel broad explanation for particle toxicity in general. Silica samples having different bulk and surface properties, specifically different amounts of NFS, are tested with a set of membrane systems of decreasing molecular complexity and different charge. The results demonstrate that NFS content is the primary determinant of membrane disruption causing red blood cell lysis and changes in lipid order in zwitterionic, but not in negatively charged liposomes. NFS-rich silica strongly and irreversibly adsorbs zwitterionic self-assembled phospholipid structures. This selective interaction is corroborated by density functional theory and supports the hypothesis that NFS recognize membrane epitopes that exhibit a positive quaternary amino and negative phosphate group. These new findings define a new paradigm for deciphering particle-biomembrane interactions that will support safer design of materials and what types of treatments might interrupt particle-biomembrane interactions.

Collective behaviour in living systems is observed across many scales, from bacteria to insects, to fish shoals. Zebrafish have emerged as a model system amenable to laboratory study. Here we report a three-dimensional study of the collective dynamics of fifty zebrafish. We observed the emergence of collective behaviour changing between ordered to randomised, upon adaptation to new environmental conditions. We quantify the spatial and temporal correlation functions of the fish and identify two length scales, the persistence length and the nearest neighbour distance, that capture the essence of the behavioural changes. The ratio of the two length scales correlates robustly with the polarisation of collective motion that we explain with a reductionist model of self–propelled particles with alignment interactions., Author summary Groups of animals can display complex collective motion, which emerges from physical and social interactions amongst the individuals. A quantitative analysis of emergent collective behaviour in animals is often challenging, as it requires describing the movement of many individual animals. With an innovative 3D tracking system, we comprehensively characterized the motion of large groups of zebrafish (Danio rerio), a freshwater fish commonly used as a vertebrate model organism. We find that the different collective behaviours are captured by two physical scales: the length of persistent motion in a given direction and the typical nearest neighbour distance. Their ratio allows us to interpret the experimental results, in the light of a statistical mechanics model for swarming with persistent motion and local neighbourhood alignment.

Occupational exposure to quartz dust is associated with fatal diseases. Quartz dusts generated by mechanical fracturing are characterized by a broad range of micrometric to nanometric particles. The contribution of this nanometric fraction to the overall toxicity of quartz is still largely unexplored, primarily because of the strong electrostatic adhesion forces that prevent isolation of the nanofraction. Furthermore, fractured silica dust exhibits special surface features, namely nearly free silanols (NFS), which impart a membranolytic activity to quartz. Nanoquartz can be synthetized via bottom-up methods, but the surface chemistry of such crystals strongly differs from that of nanoparticles resulting from fracturing. Here, we report a top-down milling procedure to obtain a nanometric quartz that shares the key surface properties relevant to toxicity with fractured quartz. The ball milling was optimized by coupling the dry and wet milling steps, using water as a dispersing agent, and varying the milling times and rotational speeds. Nanoquartz with a strong tendency to form submicrometric agglomerates was obtained. The deagglomeration with surfactants or simulated body fluids was negligible. Partial lattice amorphization and a bimodal crystallite domain size were observed. A moderate membranolytic activity, which correlated with the number of NFS, signaled coherence with the previous toxicological data. A membranolytic nanoquartz for toxicological investigations was obtained.

Colloidal dispersions are prized as model systems to understand basic properties of materials, and are central to a wide range of industries from cosmetics to foods to agrichemicals. Among the key developments in using colloids to address challenges in condensed matter is to resolve the particle coordinates in 3D, allowing a level of analysis usually only possible in computer simulation. However in amorphous materials, relating mechanical properties to microscopic structure remains problematic. This makes it rather hard to understand, for example, mechanical failure. Here we address this challenge by studying the contacts and the forces between particles, as well as their positions. To do so, we use a colloidal model system (an emulsion) in which the interparticle forces and local stress can be linked to the microscopic structure. We demonstrate the potential of our method to reveal insights into the failure mechanisms of soft amorphous solids by determining local stress in a colloidal gel. In particular, we identify `force chains' of load-bearing droplets, and local stress anisotropy, and investigate their connection with locally rigid packings of the droplets., Comment: 12 pages

Asbestos inhalation is associated with fatal respiratory diseases and raises concerns from the perspective of workplace safety and environmental impacts. Asbestos and asbestos-like minerals naturally occur in rocks and may become airborne when outcrops or soils are disturbed by anthropic activities. In situ detection of these minerals is a crucial step for the risk evaluation of natural sites. We assess here whether a portable Raman spectrometer (pRS) may be used in the identification of asbestos and asbestos-like minerals at the mining front during exploitation. pRS performance was tested at three geologically different mining sites in Italy and New Caledonia and compared with a high-resolution micro-Raman spectrometer (HRS). About 80% of the overall in situ analyses at the mining front were successfully identified by pRS, even when intermixed phases or strongly disaggregated and altered samples were analyzed. Chrysotile and tremolite asbestos, asbestos-like antigorite, and balangeroite were correctly detected during surveys. The major difficulties faced during in situ pRS measurements were fluorescence emission and focussing the laser beam on non-cohesive bundles of fibers. pRS is adequate for discriminating asbestos and asbestos-like minerals in situ. pRS may support risk assessment of mining sites to better protect workers and environment.

Background: Silica continues to represent an intriguing topic of fundamental and applied research across various scientific fields, from geology to physics, chemistry, cell biology, and particle toxicology. The pathogenic activity of silica is variable, depending on the physico-chemical features of the particles. In the last 50 years, crystallinity and capacity to generate free radicals have been recognized as relevant features for silica toxicity. The ‘surface’ also plays an important role in silica toxicity, but this term has often been used in a very general way, without defining which properties of the surface are actually driving toxicity. How the chemical features (e.g., silanols and siloxanes) and configuration of the silica surface can trigger toxic responses remains incompletely understood. Main body: Recent developments in surface chemistry, cell biology and toxicology provide new avenues to improve our understanding of the molecular mechanisms of the adverse responses to silica particles. New physicochemical methods can finely characterize and quantify silanols at the surface of silica particles. Advanced computational modelling and atomic force microscopy offer unique opportunities to explore the intimate interactions between silica surface and membrane models or cells. In recent years, interdisciplinary research, using these tools, has built increasing evidence that surface silanols are critical determinants of the interaction between silica particles and biomolecules, membranes, cell systems, or animal models. It also has become clear that silanol configuration, and eventually biological responses, can be affected by impurities within the crystal structure, or coatings covering the particle surface. The discovery of new molecular targets of crystalline as well as amorphous silica particles in the immune system and in epithelial lung cells represents new possible toxicity pathways. Cellular recognition systems that detect specific features of the surface of silica particles have been identified. Conclusions: Interdisciplinary research bridging surface chemistry to toxicology is progressively solving the puzzling issue of the variable toxicity of silica. Further interdisciplinary research is ongoing to elucidate the intimate mechanisms of silica pathogenicity, to possibly mitigate or reduce surface reactivity. Keywords: Silica, Silicosis, Lung cancer, Auto-immune diseases, Surface reactivity, Silanol, Coating, Modelling, Spectroscopy, Atomic force microscopy

Anatase titanium dioxide nanoparticles (TiO2 NPs) are used in a large range of industrial applications mainly due to their photocatalytic properties. Before entering the lung, virtually all TiO2 NPs are exposed to some UV light, and lung toxicity of TiO2 NPs might be influenced by photoexcitation that is known to alter TiO2 surface properties. Although the TiO2 NPs toxicity has been extensively investigated, limited data are available regarding the toxicity of TiO2 NPs that have been pre-exposed to UV light, and their impact on humans remains unknown. In this study, five types of TiO2NPs with tailored physicochemical features were characterized and irradiated by UV for 30 min. Following irradiation, cytotoxicity, pro-inflammatory response, and oxidative stress on a human lung coculture system (A549 epithelial cells and macrophages differentiated from THP-1 cells) were assessed. The surface charge of all samples was less negative after UV irradiation of TiO2 NPs, and the average aggregate size was slightly increased. A higher cytotoxic effect was observed for preirradiated TiO2 NPs compared to nonirradiated samples. Preirradiation of TiO2 NPs had no significant impact on the pro-inflammatory response and oxidative stress as shown by a similar production of IL-8, TNF-α, and reactive oxygen species.

Anatase titanium dioxide nanoparticles (TiO

Exposure to natural occurrences of asbestos (NOA) and other potentially hazardous elongated mineral particles (EMPs) may pose a risk to human health and the environment. Weathering forces and anthropic activities may alter the cohesion of NOA-bearing outcrops and disperse EMPs in air, water, and soil. The current paradigm for fibre toxicity indicates that morphology and crystal chemistry are key parameters in determining the toxicological properties of a mineral. This work aims to assess and discuss the impact of sub-tropical supergene alteration and weathering on the morphology and the chemical composition of antigorite, a non-regulated serpentine that shares chemical composition with asbestos chrysotile. Antigorite naturally occurring in lateritic Ni ores of New Caledonia exhibits a unique asbestos-like habit at the microscopic scale. Standardized mechanical stress was performed on antigorites, selected to represent different cohesion states. The specimens produced a relevant amount of respirable fibres, between 32 and 42% (WHO counting criteria). PCA on chemical data and ternary diagrams show that all antigorites exhibit a similar Si content (from 2.05 to 2.09 a.f.u.) but were mainly differentiated by Mg and Ni content, ranging from 2.66 to 2.80 and 0.00 to 0.09 a.f.u., respectively. Si content in Caledonian antigorite is higher than Si in non-lateritic samples. This suggests that a main alteration process occurred after the obduction of the ultramafic protolith. The supergene alteration determined the Ni enrichment of lateritic deposits and is likely the main cause of the mineral alteration of antigorite under sub-tropical environments. Further, weathering processes prompt the disaggregation of altered antigorite causing the generation and dispersion of respirable, potentially hazardous, antigorite fibres in the environment.

Due to their ability to penetrate, deteriorate and discolour stone surfaces, rock-inhabiting black fungi represent a remarkable issue for cultural heritage conservation. Black microcolonial fungi (MCF) can also adapt to different environmental conditions, by converting from yeast-like morphology to a peculiar meristematic development with swollen cells (torulose hyphae, TH), to extremely thin structures (filamentous hyphae, FH). Furthermore, black MCF produce protective pigments: melanin, dark pigment particularly evident on light stone surfaces, and carotenoids. Black fungi produce melanin in critical, oligotrophic conditions as well as constitutively. Melanin function is mostly related to stress resistance and the ability of fungi to generate appressorial turgor to actively penetrate plant cells in pathogenic species. An involvement of melanins in stone surface penetration has been suggested, but not experimentally proved. In this work, we tested the role of hyphal melanisation in penetration mechanisms on the model black fungus Knufia petricola A95 in lab conditions. The wild-type and three mutants with introduced targeted mutations of polyketide-synthases (melanin production) and/or phytoene dehydrogenase (carotenoid synthesis) were inoculated on artificial carbonate pellets (pressed Carrara marble powder) of different porosity. After 5, 10, 17 and 27 weeks, hyphal penetration depth and spread were quantified on periodic acid Schiff-stained cross-sections of the pellets, collecting measurements separately for TH and FH. Droplet assay of the mutants on different media were conducted to determine the role of nutrients in the development of different fungal morphologies. In our in vitro study, the hyphal penetration depth, never exceeding 200 μm, was proven to be consistent with observed penetration patterns on stone heritage carbonate substrates. Pellet porosity affected penetration patterns of TH, which developed in voids of the more porous pellets, instead than actively opening new passageways. Oppositely, the thin diameter of FH allowed their penetration independently of substrate porosity. Instead, the long-hypothesized crucial role of melanin in black MCF hyphal penetration should be rejected. TH were developed within the pellets also by melanin deficient strains, and melanized strains showed an endolithic component of non-melanized TH. FH were non-melanized for all the strains, but deeply penetrated all pellet types, with higher penetration depth probably related to their potential exploratory (nutrient-seeking) role, while TH may be more related to a resistance to surface stress factors. In the melanin deficient strains, the absence of melanin caused an increased penetration rate of FH, hypothetically related to an earlier necessity to search for organic nutrients.

We study non-equilibrium analogues of surface phase transitions in a minimal model of active particles in contact with a purely repulsive potential barrier that mimics a thin porous membrane. Under conditions of bulk motility-induced phase separation, the interaction strength $\varepsilon_w$ of the barrier controls the affinity of the dense phase for the barrier region. We uncover clear signatures of a wetting phase transition as $\varepsilon_w$ is varied. In common with its equilibrium counterpart, the character of this transition depends on the system dimensionality: a continuous transition with large density fluctuations and gas bubbles is uncovered in 2d while 3d systems exhibit a sharp transition absent of large correlations.

Bioactive glasses are the materials of choice in the field of bone regeneration. Antioxidant properties of interest to limit inflammation and foreign body reactions have been conferred to bioactive glasses by the addition of appropriate ions (such as Ce or Sr). On the other hand, the antioxidant activity of bioactive glasses without specific ion/molecular doping has been occasionally cited in the literature but never investigated in depth. In the present study, three silica-based bioactive glasses have been developed and characterized for their surface properties (wettability, zeta potential, chemical composition, and reactivity) and radical scavenging activity in the presence/absence of cells. For the first time, the antioxidant activity of simple silica-based (SiO2–CaO–Na2O) bioactive glasses has been demonstrated.

The Fenton process activated by Zero Valent Iron (ZVI-Fenton) is shown here to effectively remove antibiotics reserved for hospital settings (specifically used to treat antibiotic-resistant infections) from wastewater, thereby helping in the fight against bacterial resistance. Effective degradation of cefazolin, imipenem and vancomycin in real urban wastewater was achieved at pH 5, which is quite near neutrality when compared with classic Fenton that works effectively at pH 3–4. The possibility to operate successfully at pH 5 has several advantages compared to operation at lower pH values: (i) lower reagent costs for pH adjustment; (ii) insignificant impact on wastewater conductivity, because lesser acid is required to acidify and lesser or no base for neutralization; (iii) undetectable release of dissolved Fe, which could otherwise be an issue for wastewater quality. The cost of reagents for the treatment ranges between 0.04 and 0.07 $ m−3, which looks very suitable for practical applications. The structures of the degradation intermediates of the studied antibiotics and their likely abundance suggest that, once the primary compound is eliminated, most of the potential to trigger antibiotic action has been removed. Application of the ZVI-Fenton technique to wastewater treatment could considerably lower the possibility for antibiotics to trigger the development of resistance in bacteria.