Citation: Khan, P.A.; Thoutam, A.K.; Gopal, V.; Gurumallesh, A.; Joshi, S.; Palaniappan, A.; Markocsan, N.; Manivasagam, G. Influence of Graphene Nanoplatelets on the Performance of Axial Suspension Plasma-Sprayed Hydroxyapatite Coatings. Bioengineering 2023, 10, 44. https://doi.org/10.3390/ bioengineering10010044 Academic Editor: Ali Zarrabi Received: 10 November 2022 Revised: 3 December 2022 Accepted: 8 December 2022 Published: 29 December 2022 Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). bioengineering Article Influence of Graphene Nanoplatelets on the Performance of Axial Suspension Plasma-Sprayed Hydroxyapatite Coatings Pearlin Amaan Khan 1 , Aravind Kumar Thoutam 2 , Vasanth Gopal 1,3 , Aswin Gurumallesh 1,3 , Shrikant Joshi 2 , Arunkumar Palaniappan 1 , Nicolaie Markocsan 2 and Geetha Manivasagam 1, * 1 Centre for Biomaterials, Cellular & Molecular Theranostics (CBCMT), Vellore Institute of Technology, Vellore 632014, India 2 Division of Subtractive and Additive Manufacturing, University West, 461 86 Trollhättan, Sweden 3 School of Advanced Sciences, Vellore Institute of Technology, Vellore 632014, India * Correspondence: geethamanivasagam@vit.ac.in; Tel.: +91-416-220-295 (ext. 2296) Abstract: Axial suspension plasma spraying (ASPS) is an alternative technique to atmospheric plasma spraying (APS), which uses a suspension of much finer powders (<5-micron particle size) as the feedstock. It can produce more refined microstructures than APS for biomedical implants. This paper highlights the influence of incorporated graphene nanoplatelets (GNPs) on the behavior of ASPS hydroxyapatite (HAp) coatings. The characterization of the ASPS coatings (HAp + varying GNP contents) was carried out using scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), confocal Raman microscopy (CRM), white light interferometry (WLI), and contact angle measurements. The evaluation of the mechanical properties such as the hardness, roughness, adhesion strength, and porosity was carried out, along with a fretting wear performance. Additionally, the biocompatibility of the Hap + GNP coatings was evaluated using cytotoxicity testing which revealed a decrease in the cell viability from 92.7% to 85.4%, with an increase in the GNP wt.%. The visualization of the cell’s components was carried out using SEM and Laser Scanning Microscopy. Furthermore, the changes in the genetic expression of the various cellular markers were assessed to analyze the epigenetic changes in human mesenchymal stem cells. The gene expression changes suggested that GNPs upregulated the proliferation marker and downregulated the pluripotent markers by a minimum of three folds. Keywords: plasma axial suspension plasma spraying; biocompatibility; Ti-6Al-4V; graphene nanoplatelets; hMSCs; hydroxyapatite 1. Introduction Atmospheric plasma spraying (APS) has been identified as one of the burgeoning orthopedic implant coating techniques alongside electrophoretic deposition (EPD), dip coating, anodization, powder coating, and galvanization [1] mainly due to its validation by the FDA (the Food and Drug Administration) in the USA [2]. APS involves a high-energy plasma arc that can melt the injected powder particles, which will then be propelled with a high velocity onto the target substrate. The deposition onto the substrate compels the particles to solidify into pancake-type splats [3]. Despite its efficacy in the coating, the APS is limited by its phase changes, leading to the generation of by-products such as tri- and tetra calcium phosphates when depositing HAp and producing thicker coatings (>50 μm)[4]. Thinner coatings (<50 μm) with a minimum of 45% of crystallinity is the ISO requisite for implant coatings [5,6]. Suspension plasma spraying (SPS) is a better alternative to APS to achieve the above. Finer ceramic particles with a nanometric to submicrometric size are suspended in a solvent, usually water, ethanol, or a mixture of them, and injected into the plasma plume. Due to the finer particle size feedstock and the inherent mechanisms responsible for the coating formation, SPS can yield coatings with unique microstructures, Bioengineering 2023, 10, 44. https://doi.org/10.3390/bioengineering10010044 https://www.mdpi.com/journal/bioengineering