No abstract

Layered silicates (LS, clays) are a composite group of minerals whose industrial interest and technological applications are progressively expanding, spanning from catalysis to biomedicine. However, the compatibility of LS with biological systems is not clear, and mechanistic data about biophysicochemical interactions at the interface of LS and biomembranes are scarce. Here, cell membrane damage, assessed using red blood cells as model membranes, is revealed for kaolin (> 75 wt.% kaolinite, 1:1 layer structure) and bentonite (> 90 wt.% montmorillonite, 1:2 layer structure) particles. The high membranolytic capacity of bentonite (i.e., high ion‐exchanger LS) is the result of the combined contribution of both mineral surface features and sample‐specific cation exchange capacity (CEC). For kaolin (i.e., non‐ion‐exchanger LS), the capacity to damage membranes is primarily due to surface hydroxyl species, that is, silanols and aluminols, exposed at the crystal lattice boundaries. When kaolin is thermally disorganized into amorphous metakaolin, membrane damage is driven by a specific sub‐population of surface hydroxyl species, namely the “nearly free silanols”, previously evidenced on quartz. This study establishes the rationale underlying the interactions between LS particles and membranes and can set the basis for the understanding of interfacial phenomena of LS.

Crystalline silica (CS) is a well-known hazardous material that causes severe diseases including silicosis, lung cancer, and autoimmune diseases. However, the hazard associated to crystalline silica is extremely variable and depends on some specific characteristics, including crystal structure and surface chemistry. The crystalline silica polymorphs share the SiO2 stoichiometry and differentiate for crystal structure. The different crystal lattices in turn expose differently ordered hydroxyl groups at the crystal surface, i.e., the silanols. The nearly free silanols (NFS), a specific population of weakly interacting silanols, have been recently advanced as the key surface feature that governs recognition mechanisms between quartz and cell membrane, initiating toxicity. We showed here that the nearly free silanols occur on the other crystalline silica polymorphs and take part in the molecular interactions with biomembranes. A set of crystalline silica polymorphs, including quartz, cristobalite, tridymite, coesite, and stishovite, was physico-chemically characterized and the membranolytic activity was assessed using red blood cells as model membranes. Infrared spectroscopy in highly controlled conditions was used to profile the surface silanol topochemistry and the occurrence of surface nearly free silanols on crystalline silica polymorphs. All crystalline silica polymorphs, but stishovite were membranolytic. Notably, pristine stishovite did not exhibited surface nearly free silanols. The topochemistry of surface silanols was modulated by thermal treatments, and we showed that the occurrence of nearly free silanols paralleled the membranolytic activity for the crystalline silica polymorphs. These results provide a comprehensive understanding of the structure-activity relationship between nearly free silanols and membranolytic activity of crystalline silica polymorphs, offering a possible clue for interpreting the molecular mechanisms associated with silica hazard and bio-minero-chemical interfacial phenomena, including prebiotic chemistry.

<p>Respirable crystalline silica (RCS) is the leading cause of occupational respiratory disease worldwide. RCS is associated with silicosis, cancer, and autoimmune diseases [1]. Silica (SiO<sub>2</sub>) 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, <em>i.e.,</em> deciphering the exact molecular nature of the interaction between silica and biological matter.</p><p>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 <em>in vitro</em> and <em>in vivo</em> particle toxicity assessment.</p><p>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.</p><p><strong>References<br></strong>[1] Leung et al., <em>Lancet</em> 2012, <em>379</em>, 2008;<em> </em>[2] Pavan et al.,<em> </em><em>Proc. Natl. </em><em>Acad. </em><em>Sci USA</em> 2020, <em>117</em>, 27836; [3] Pavan et al., <em>sumbitted to ACS Central Science</em></p>

<p>Kaolin and bentonite, two clays mainly made of kaolinite and montmorillonite, respectively, are largely used in various industrial applications. However, their impact on human health has not been fully investigated and data about their mechanism of cellular toxicity are scarce. <em>In vivo</em> and <em>in vitro </em>studies showed that kaolin and bentonite particles can induce transient inflammation, alveolar proteinosis, and are cytotoxic to a variety of mammalian cells (Wiemann et al. 2020; Maciaszek et al. 2021). We recently demonstrated (Pavan et al. 2020) that a specific sub-population of surface silanols located at a well-defined intersilanol distance, <em>i.e.</em>, nearly-free silanols (NFS), is responsible for the membranolytic and inflammatory activity of quartz particles. We hypothesized that a similar structure-activity relationship may exist for kaolinite and montmorillonite particles, since they exhibit tetrahedral SiO<sub>2</sub> layers at their outer surface and hydroxyls groups, <em>i.e.</em>, silanols and aluminols, at the crystal lattice boundaries.</p><p>Four bentonite (> 90% montmorillonite) and kaolin (> 75% kaolinite) particles were characterized for their physico-chemical properties of toxicological interest and their capacity to damage cellular membranes was assessed using red blood cells as model of membranes. All bentonite and kaolin particles resulted highly membranolytic. As clay minerals may exchange cations with suspending medium and the structural integrity of biological membranes may be compromised by significant alteration of the medium ionic strength, the membranolytic activity of kaolin and bentonite leachates was assessed. Only bentonite leachates induced membrane damage with an effect that was dependent on each sample specific cation exchange capacity (CEC). A reduction or a complete abrogation of kaolin and bentonite membranolytic activity was observed when their surface was coated with dioleoyl lecithin, indicating that surface moieties play a key role for both kaolin and bentonite interactions with membranes. Investigations by IR spectroscopy of the surface-exposed hydroxyl groups revealed the occurrence of NFS, which vibrational feature was especially well defined for kaolin. Thermal treatments carried out on kaolin modified the relative intensity of NFS and its membranolytic activity, suggesting a relationship between NFS and membrane damage.</p><p>In conclusion, the capacity of kaolin particles to damage membranes appears related to kaolinite specific surface hydroxylated species. On the other hand, the mechanism of interaction of montmorillonite particles with membranes is function of both mineral surface features and CEC. These findings provide a preliminary understanding of the mechanism of interaction of clay minerals with biological membranes. This interaction may represent the triggering event of kaolin and bentonite adverse cellular effects.</p><p> </p><p>Bibliography</p><p>Maciaszek K. et al. (2022) An in vitro assessment of the toxicity of two-dimensional synthetic and natural layered silicates,Toxicol <em>In Vitro,</em> 78:105273</p><p>Pavan C. et al. (2020) Nearly free surface silanols are the critical molecular moieties that initiate the toxicity of silica particles, Proc Natl Acad Sci USA, 117 (45):27836</p><p>Wiemann M. et al. (2020) Lung toxicity analysis of nano-sized kaolin and bentonite: missing indications for a common grouping, Nanomaterials, 10 (2):204</p>