Cytocompatibility of Graphene Monolayer and Its Impact on Focal Cell Adhesion, Mitochondrial Morphology and Activity in BALB/3T3 Fibroblasts

Iwona Lasocka,Michał Skibniewski,Lidia Szulc-Dąbrowska,Ewa Skibniewska,Marie Hubalek Kalbacova,Iwona Pasternak,Karolina Gregorczyk-Zboroch

doi:10.3390/ma14030643

Iwona Lasocka, Michał Skibniewski + Show 5 more

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https://doi.org/10.3390/ma14030643

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Abstract

This study investigates the effect of graphene scaffold on morphology, viability, cytoskeleton, focal contacts, mitochondrial network morphology and activity in BALB/3T3 fibroblasts and provides new data on biocompatibility of the “graphene-family nanomaterials”. We used graphene monolayer applied onto glass cover slide by electrochemical delamination method and regular glass cover slide, as a reference. The morphology of fibroblasts growing on graphene was unaltered, and the cell viability was 95% compared to control cells on non-coated glass slide. There was no significant difference in the cell size (spreading) between both groups studied. Graphene platform significantly increased BALB/3T3 cell mitochondrial activity (WST-8 test) compared to glass substrate. To demonstrate the variability in focal contacts pattern, the effect of graphene on vinculin was examined, which revealed a significant increase in focal contact size comparing to control-glass slide. There was no disruption in mitochondrial network morphology, which was branched and well connected in relation to the control group. Evaluation of the JC-1 red/green fluorescence intensity ratio revealed similar levels of mitochondrial membrane potential in cells growing on graphene-coated and uncoated slides. These results indicate that graphene monolayer scaffold is cytocompatible with connective tissue cells examined and could be beneficial for tissue engineering therapy.

Highlights

Medical materials have to be tested in vitro to choose the most suitable of them for further studies including in vivo tests with animal models and clinical trials with human patients
The assessment of the in vitro cytotoxicity of the substance or scaffold is often a qualitative analysis based on the morphological examination of cell damage and growth after direct contact with the material [1,3,22]
There was no significant difference in the size of BALB/3T3 cells cultured on glass and graphene

Summary

Introduction

Medical materials have to be tested in vitro to choose the most suitable of them for further studies including in vivo tests with animal models and clinical trials with human patients. ISO (Organization of Standardization) 10993 standards are well established and give guidance on testing the biocompatibility of materials; especially, ISO 10993 Part 5 describes usual techniques to evaluate cytotoxicity of materials [1]. These methods and techniques have been adopted by researchers to evaluate the biocompatibility of nanomaterials [2]. There are many in vitro cytotoxicity tests, which detect the toxic potential of the material studied [3] Results of these tests could be expressed as cell viability, rate of proliferation or rate of metabolism. We suppose that graphene scaffold for fibroblasts or mesenchymal stem cells could be used in the future to aid reconstruction of damaged tissue caused by mechanical injuries, burns or as a result of chronic diseases

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