The Role of Oxidative Stress in the Response of Endothelial Cells to Metals - Biologically Responsive Biomaterials for Tissue Engineering - page 77

reduced compared to cells cultured on PS (Fig. 4.5a ). Moreover, H 2 O 2 treatment of
endothelial cells grown on PS elevated catalase and SOD activity. It is known that
oxidative stress may increase the activity of antioxidant enzymes. The mechanism
of this induction may include changes in enzymatic activity of existing proteins and
de novo enzyme synthesis [ 58 ] . H 2 O 2 exposure induced a slight decrease in the
activity of both enzymes in endothelial cells grown on Ti6Al4V. This reduction in
SOD and catalase activity could be caused by a cumulative effect of the temporary
H 2 O 2 -induced and the permanent Ti6Al4V-induced oxidative stress. Consequently,
the facts demonstrated above indicated an elevated sensitivity of endothelial cells to
oxidative stress when growing in contact with Ti6Al4V and thus a reduced antioxi-
dant defence potential.
An animal study by Ozmen et al. [ 65 ] suggested a similar effect in vivo: it was
shown that 1 month after the insertion of titanium implants in rabbits an increase in
lipid peroxidation (a possible result of oxidative stress) in the tissues surrounding
the titanium implants occurred, whereas there was a decrease in the activities of
antioxidant enzymes (e.g. catalase, SOD, GPX). The same reactions, though to a
lesser extent, were seen in the tissues around implanted stainless steel but were
minimal in the case of implanted polyethylene. These observations indicate a state
of permanent oxidative stress at the surface of titanium implants, leading to exhaus-
tion of antioxidant enzymes in the surrounding tissues. In another study, increased
oxidative stress was observed in patients in the tissue surrounding stable and loose
total hip implants [ 40 ]. It was manifested by a low GSH/GSSH ratio and elevated
levels of MDA, a known product of lipid peroxidation, in patients compared to con-
trol individuals. Increased H 2 O 2 concentrations and decreased catalase activity were
also detected in the fibrotic capsule from a failed joint implant [ 89 ] . Endothelial
cells were also shown to undergo oxidative stress when growing in contact with a
NiTi alloy. However, this was attributed to the presence of the transition metal Ni in
the TiO 2 layer [ 68 ]. Vanadium, which is a component of the alloy used in this study,
is known to be toxic and to induce formation of ROS via Fenton and Haber-Weiss
reactions [ 99 ]. However, on analysis of the surface oxide film of the Ti6Al4V alloy
using X-ray photoelectron spectroscopy vanadium oxide was not detected [ 61 ] , but
the presence of traces of vanadium on the surface of titanium alloy cannot be com-
pletely excluded.
Another important function of endothelial cells is their role as the driving cell
type in angiogenesis. The influence of titanium degradation products on angiogenic
potential of endothelial cells has not yet been examined. In this study there were no
differences seen in angiogenic potential of HDMEC in an in vitro assay performed
on PS or Ti6Al4V (Fig. 4.6 ). It has to be stated, however, that the cells were not
growing in direct contact to the metal surface. Thus, the effects of oxidative stress
on Ti6Al4V could be masked by collagen and fibrin used in the assay. Addition of
0.5 mM H 2 O 2 , however, reduced the formation of tube-like structures on both mate-
rials. There are contradictory reports in the literature regarding the effects of H 2 O 2
on endothelial cells. In one study the low doses of H 2 O 2 (0.1 or 1 mM) induced pro-
liferative activity of endothelial cells and increased tube formation in an in vitro
angiogenesis assay. A higher concentration of H 2 O 2 (10 mM), in turn, reduced the