The effect of combined hypergravity and microgrooved surface topography on the behaviour of fibroblasts

Abstract

This study evaluated in vitro the differences in morphological behaviour between fibroblast cultured on smooth and micro-grooved substrata (groove depth: 1 mum, width: 1, 2, 5, 10 microm), which undergo artificial hypergravity by centrifugation (10, 24 and 50 g; or 1 g control). The aim of the study was to clarify which of these parameters was more important to determine cell behaviour. Morphological characteristics were investigated using scanning electron microscopy and fluorescence microscopy in order to obtain qualitative information on cell spreading and alignment. Confocal laser scanning microscopy visualised distribution of actin filaments and vinculin anchoring points through immunostaining. Finally, expression of collagen type I, fibronectin, and alpha(1)- and beta(1)-integrin were investigated by PCR. Microscopy and image analysis showed that the fibroblasts aligned along the groove direction on all textured surfaces. On the smooth substrata (control), cells spread out in a random fashion. The alignment of cells cultured on grooved surfaces increased with higher g-forces until a peak value at 25 g. An ANOVA was performed on the data, for all main parameters: topography, gravity force, and time. In this analysis, all parameters proved significant. In addition, most gene levels were reduced by hypergravity. Still, collagen type 1 and fibronectin are seemingly unaffected by time or force. From our data it is concluded that the fibroblasts primarily adjust their shape according to morphological environmental cues like substratum surface whilst a secondary, but significant, role is played by hypergravity forces.

The effect of different surface roughnesses on the differentiation of MC3T3-E1 mouse osteoblasts in vitro

“…However, cells cultured on wider microgrooves of either 5 μm or 10 μm in width attached to the substrate just the same as on smooth surfaces, and there was no acceleration of calcification. Loesberg et al (9,10) used the same size microgrooves as Matsuzaka et al (11) and suggested that cell differentiation was accelerated on microgrooves of 1 μm to 2 μm in width, because cells were suspended in the groove as a cellular bridge and exposed receptors existing on the cell membrane, making it easier to get molecular signals such as dexamethasone into the culture medium and cause (or induce) the cells to differentiate into mature osteoblasts.…”

Centrifuges for Microgravity Simulation. The Reduced Gravity Paradigm

“…Centrifuges are applied in various disciplines mainly in material and particle technology like for separation (Sahuquillo et al, 1999;Kuang et al, 2015), for nanomaterials classification (Wohlleben, 2012), casting (Huisman et al, 1994;Liu and Barnett, 2002;Dikel et al, 2005;Velhinho and Rocha, 2011), combustion (Merzhanov, 1995), but also in areas like geology (Gopal Madabhushi, 2004;Costa et al, 2016), life sciences in plants (Musgrave et al, 2009;Herranz et al, 2013;Hoson and Wakabayashi, 2015), single cells (Loesberg et al, 2006;Tavakolinejad et al, 2015), animals (van Loon, 2001;van Loon et al, 2005;Tateishi et al, 2015) and humans (Akima et al, 2005;van Loon et al, 2012;Goswami et al, 2015).…”

Cytoskeletal role in differential adhesion patterns of normal fibroblasts and breast cancer cells inside silicon microenvironments

“…Some of the aforementioned studies have focused on the critical role of cytoskeletal components, actin microfilaments and microtubules in cell alignment (Oakley et al 1997;Wojciakstothard et al 1995). Both mechanical loading and microgrooved surface topographies promote cytoskeletal reorganization and affect the behavior of fibroblast cells (Loesberg et al 2005(Loesberg et al , 2006. Cytoskeletal proteins play an important role in tissue engineering by mediating the cellular response to different substrates.…”