Mechanisms of Reduced Astrocyte Surface Coverage in Cortical Neuron-Glia Co-cultures on Nanoporous Gold Surfaces

Abstract

Nanoporous gold (np-Au) is a promising multifunctional material for neural electrodes. We have previously shown that np-Au nanotopography reduces astrocyte surface coverage (linked to undesirable gliosis) while maintaining high neuronal coverage in a cortical primary neuron-glia co-culture model as long as two weeks in vitro. Here, we investigate the potential influence of secreted soluble factors from cells grown on np-Au on the cell type-specific surface coverage of cells grown on conventional tissue culture plastic and test the hypothesis that secretion of factors is responsible for inhibiting astrocyte coverage on np-Au. In order to assess whether factors secreted from cells grown on np-Au surfaces reduced surface coverage by astrocytes, we seeded fresh primary rat neuron-glia co-cultures on conventional polystyrene culture dishes, but maintained the cells in conditioned media from co-cultures grown on np-Au surfaces. After one week in vitro, a preferential reduction in astrocyte surface coverage was not observed, suggesting that soluble factors are not playing a role. In contrast, four hours after cell seeding there were a significant number of non-adhered, yet still viable, cells for the cultures on np-Au surfaces. We hypothesize that the non-adherent cells are mainly astrocytes, because: (i) there was no difference in neuronal cell coverage between np-Au and pl-Au for long culture durations and (ii) neurons are post-mitotic and not expected to increase in number upon attaching to the surface. Overall, the results suggest that the np-Au topography leads to preferential neuronal attachment shortly after cell seeding and limits astrocyte-specific np-Au surface coverage at longer culture durations.

A Microfluidic Platform to Study Astrocyte Adhesion on Nanoporous Gold Thin Films

“…For pl-Au, the difference was lower at roughly 15%. The general trend of reduced cellular attachment with decreasing ligament width was consistent with what has been observed previously for reduced cell coverage on np-Au surfaces [ 35 ], suggesting that adhesion strength plays a role in cellular spreading.…”

“…While we attributed the differences in astrocyte detachment (adhesion strength) mainly to the substrate nanotopography, it is important to mention that other factors such as surface chemistry and mechanical stiffness can play an important role in dictating cellular adhesion strength [ 39 ]. In addition, as described in our previous study, for short incubation durations, the probability of cells forming focal adhesion to a pitted surface upon reaching the surface may play a role [ 35 ].…”

“…However, the cell adhesion behavior following short attachment duration was still significantly dependent on the coating morphology as shown in Figure 3 and Figure 4 . It is expected that the behavior in this transitional cell attachment regime was driven by a combination of the aforementioned FA formation and integrin clustering mechanisms as well as the kinetics of FA establishment on the complex nanotopography (composed of ligaments and voids) [ 35 ].…”

Nanoporous Metals with Structural Hierarchy: A Review

“…Local processes at the surface of the pores or at their interface with a fluid are exploited for applications in fields such as catalysis, [11,[47][48][49][50] sensing, [11,51,52] actuation (see below), optical switching, [140][141][142] electropumping for microfluidic devices, [143] integrated circuit contacting, [144] biological implants, [145][146][147][148] and as electrodes where a large surface area is needed. Local processes at the surface of the pores or at their interface with a fluid are exploited for applications in fields such as catalysis, [11,[47][48][49][50] sensing, [11,51,52] actuation (see below), optical switching, [140][141][142] electropumping for microfluidic devices, [143] integrated circuit contacting, [144] biological implants, [145][146][147][148] and as electrodes where a large surface area is needed.…”

Biomaterial Approaches to Modulate Reactive Astroglial Response

“…Nanoporous gold surfaces seeded with both astrocytes and neurons show selective reduction of astrocytic coverage while maintaining high neuronal coverage [Chapman et al, 2015]. This response was later revealed to be due to both an initial inhibition of astrocyte attachment and topographical cues provided by the nanoporous gold structure [Chapman et al, 2016]. Nanoporous alumina surfaces have also been studied, and it was found that pore size was important for astrocyte attachment, focal adhesions, and GFAP expression [Ganguly et al, 2018].…”