The influence of mineral particles on fibroblast behaviour: A comparative study

https://doi.org/10.1016/j.colsurfb.2018.04.013Get rights and content

Highlights

  • Minerals were characterised and the influence on fibroblasts behaviour was assessed.

  • FTIR and zeta potential show protein adsorption on the minerals from the serum.

  • Minerals can be grouped according to the time-dependent cellular response.

  • Calcium sulphates induced a significant change in the fibroblast morphology.

Abstract

Minerals are versatile tools utilised to modify and control the physical-chemical and functional properties of substrates. Those properties include ones directing cell fate; thus, minerals can potentially provide a direct and inexpensive method to manipulate cell behaviour. This paper shows how different minerals influence human dermal fibroblast behaviour depending on their properties. Different calcium carbonates, calcium sulphates, silica, silicates, and titanium dioxide were characterised using TEM, ATR-FTIR, and zeta potential measurements. Mineral-cell interactions were analysed through MTT assay, LDH assay, calcein AM staining, live cell imaging, immunofluorescence staining, western blot, and extra/intracellular calcium measurements. Results show that the interaction of the fibroblasts with the minerals was governed by a shared period of adaptation, followed by increased proliferation, growth inhibition, or increased toxicity. Properties such as size, ion release and chemical composition had a direct influence on the cells leading to cell agglomeration, morphological changes, and the possible formation of protein-mineral complexes. In addition, zeta potential and FTIR measurements of the minerals showed adsorption of the cell culture media onto the particles. This article provides fundamental insight into the mineral-fibroblast interactions, and makes it possible to arrange the minerals according to the time-dependent cellular response.

Introduction

Cell fate depends strongly on the physical-chemical properties of the extracellular environment. Cells are able to sense cues from their vicinity and respond with processes such as protein expression, cell differentiation, migration, proliferation, and death [[1], [2], [3]]. Therefore, it is possible to control cell decision-making through the modification of the extracellular features. Current standard methods for cell culture do not facilitate such modifications nor mimic native extracellular environment. Thus, the development of a cell-instructive platform would allow further understanding of the mechanisms underlying cellular processes, and potentially reduce the gap between in vitro and in vivo studies.

Minerals are utilised to modify and improve physical-chemical properties of substrates, including those relevant for the cell machinery, such as surface energy, roughness or porosity. In a race to develop smart environments for cell growth, minerals can provide the means to control and regulate cell behaviour. As particles, minerals can be used as additives in matrices, as fillers, surface coatings or porous packed structures [4]. The versatility of minerals lies in the easiness to manipulate the resulting properties of a system through changes to particle size, shape, hydrophobicity, and surface structure. Size, in particular, is relevant for cell behaviour since nanoparticles can be internalised by cells and potentially exert a toxic response. It is also possible to functionalise mineral surfaces through the adsorption of molecules.

Traditionally, the largest use of minerals, such as calcium carbonates, calcium sulphates, silica, silicates, and titanium dioxide, is in the paper industry, as fillers and in paper coatings [5]. Nowadays, the biological applications of minerals is rapidly expanding. The osteogenic potential of calcium-based minerals is an advantage in bone engineering where they are used as fillers, coatings, implants or scaffolds [[6], [7], [8], [9], [10]]. As part in composites, calcium carbonate provides improved stability for long term use. In contrast, calcium sulphates have a fast resorption rate that, together with its pH-dependent solubility [11], increases locally the concentration of calcium, which is a crucial ion in cell signalling [12]. Additionally, both calcium-based minerals support the encapsulation of biomacromolecules and induce a controlled drug or gene delivery [[13], [14], [15]]. Silica is used in the delivery of biomolecules, cell labelling, cell growth coatings, bone engineering, cancer treatment and in drug tracking studies [[16], [17], [18], [19], [20]]. Natural silica (diatomite) has excellent capabilities for drug delivery and particle functionalisation due to their highly ordered multiporous structures [21,22]. Silicates such as kaolin and zeolite induce the haemostatic response in the body. They are used as clotting agents in commercial wound healing dressings to stop haemorrhagic bleeding [23]. Until recently, biocompatibility studies related to kaolin showed increased toxicity [[24], [25], [26]], yet nowadays kaolin is a promising material for novel drug delivery systems [27,28]. In contrast, literature regarding the influence of talc on cells is limited and/or outdated [[29], [30], [31], [32], [33], [34]] regardless of its unique properties such as high thermal stability, low conductivity, high oil absorptivity, high aerophilicity and hydrophobicity [35,36]. Lastly, the photocatalytic properties of titanium dioxide is applied in photodynamic cancer treatments, coating of implants, drug delivery, cell labelling, biosensing, and genetic modification of cells [37].

Despite the research available on minerals in a biological environment, there are no guidelines to predict how specific cell lines will respond to new samples. Hence, the principles behind the influence of mineral properties on cell behaviour remain to be explored. This article assesses and compares the effect of selected minerals and their properties on cell behaviour. The screening of the minerals helps to elucidate possible future applications in cell-instructive platforms such as the paper-based cell growth platform [[38], [39], [40]].

Section snippets

Materials and methods

Table 1 shows the label, commercial name, material supplier, and general description of the minerals selected for this research. The selected minerals were: calcium carbonates of various sizes, calcium sulphates with different hydrated states, silica of both natural and synthetic origin, several silicates, and titanium dioxide. Commercial grades of minerals often contain additives such as dispersing agents to improve their dispersability in aqueous systems [4]. These additives may affect the

Results and discussion

In this section, first the mineral particles are characterised according to their morphology. Then, the effect of the cell culture media on the minerals is assessed, followed by discussion of the role of the mineral properties on the cellular response of fibroblasts.

Conclusion

In this article, human dermal fibroblasts were treated with different samples of calcium carbonates, calcium sulphates, silica, silicates, and titanium dioxide. Minerals were characterised, and their influence on the fibroblasts behaviour was assessed and compared to their properties. Results show that a shared initial cellular response to the minerals is followed by enhanced proliferation, growth inhibition, or cell damage. Therefore, minerals were grouped according to the cellular response as

Acknowledgments

Doctoral Network of Material Sciences from Åbo Akademi University for the financial support. The FunMat Consortia, more specifically the Laboratory of Prof. John Eriksson for providing the facilities for the cellular studies. All the imaging was possible thanks to the Cell Imaging Core at the Turku Centre for Biotechnology in Turku, Finland.

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