Biocompatibility of Electrochemically Reduced Graphene Oxide on biomedical grade CoCr alloy. In vivo response of graphene

Abstract

In this work, electrochemically reduced Graphene oxide (ErGO) films are formed on the biomedical grade CoCr alloy to study its possible application in biomedical field. Biocompatibility tests were carried out to assess the biomedical performance of ErGO on the CoCr alloy. The discrepancy of in vivo results found in the literature regarding the side effects of graphene led to perform a previous in vivo study with Graphene. The reduction of an aqueous suspension of graphene oxide was carried out by electrochemical methods to generate ErGO films on CoCr alloys. The direct electrodeposition process was performed by cyclic voltammetry (CV) at room temperature. Characterization of the ErGO on CoCr alloys was carried out by XPS. Electrochemical procedure led to the deposition of graphene-based films on the CoCr surfaces after the partial removal of the oxygenated functional groups present in the graphene network of starting graphene oxide (GO). Biocompatibility tests of ErGO on CoCr were evaluated in J774A.1 mouse macrophages cultures, which are the main cells involved in the primary response to foreign bodies playing a decisive role in the inflammatory reactions. In vivo tests were carried out by intraperitoneal inoculation of graphene nano particles in rats evaluating possible changes in the blood line and in different organs. CV revealed the reduction of the GO around -1,2 V. The XPS high-resolution C and O peaks mainly showed sp2 bonding and the presence of C=O and C-O residual groups covering the CoCr surface. Biocompatibility studies on mouse macrophages J774A.1 cell cultures measured by the ratio between lactate dehydrogenase and mitochondrial activities showed an enhancement in the biocompatibility on the CoCr with the ErGO films, a result that seems to become more evident as exposition time increased. Optical and SE images and vimentin expression (protein responsible for architecture of cytoplasm) after 48 h exposure in macrophage culture revealed a different cell distribution, morphology and vimentin expression on CoCr or ErGO on CoCr. Macrophages on the ErGO on CoCr surfaces were well-distributed, conserved the characteristic cell shape and vimentin expression was unaffected, results that show an improvement on the CoCr biocompatibility due to the of ErGO films on material surface. Conventional optical microscopy of in vivo experiments revealed that at 15 and 30 days after inoculation there were no visible histological alterations in liver, kidney, spleen or lung, but traces of particles were found in the peritoneal cavity. The blood analysis showed alterations indicative of the hepatic inflammatory process at both 15 and 30 days. Haematological changes were found at 30 days that consisted of alterations of the red series in the form of microcytosis with a higher concentration of mean haemoglobin as well as an alteration in coagulation due to a longer prothrombin and thromboplastin time.