In this study, the cytoprotective effects of caffeine (CAF) and 8-(3-chlorostyryl)-caffeine
(CSC), A(2A) receptor antagonists, were tested against 6-OHDA-induced cytotoxicity,
in rat mesencephalic cells. Both drugs significantly increased the number of viable
cells, after their exposure to 6-OHDA, as measured by the MTT assay. While nitrite
levels in the cells were drastically increased by 6-OHDA, their concentrations were
brought toward normality after CAF or CSC, indicating that both drugs block 6-OHDA-induced
oxidative stress which leads to free radicals generation. A complete blockade of 6-OHDA-induced
lipid peroxidation, considered as a major source of DNA damage, was observed after
cells treatment with CAF or CSC. 6-OHDA decreased the number of normal cells while
increasing the number of apoptotic cells. In the CAF plus 6-OHDA group, a significant
recover in the number of viable cells and a decrease in the number of apoptotic cells
were seen, as compared to the group treated with 6-OHDA alone. A similar effect was
observed after cells exposure to CSC in the presence of 6-OHDA. Unexpectedly, while
a significant lower number of activated microglia was observed after cells exposure
to CAF plus 6-OHDA, this was not the case after cells exposure to CSC under the same
conditions. While CAF lowered the percentage of reactive astrocytes increased by 6-OHDA,
CSC presented no effect. The effects of these drugs were also examined on the releases
of myeloperoxidase (MPO), an inflammatory marker, and lactate dehydrogenase (LDH),
a marker for cytotoxicity, in human neutrophils, in vitro. CSC and CAF (0.1, 1 and
10 microg/ml) produced inhibitions of the MPO release from PMA-stimulated cells, ranging
from 45 to 83%. In addition, CSC and CAF (5, 50 and 100 microg/ml) did not show any
cytotoxicity in the range of concentrations used, as determined by the LDH assay.
All together, our results showed a strong neuroptrotection afforded by caffeine or
CSC, on rat mesencephalic cells exposed to 6-OHDA. Furthermore, CSC and caffeine actions,
inhibiting MPO as well as LDH releases, would contribute to their possible benefit
in the treatment of neurodegenerative diseases, including DP. These effects are partially
due to the ability of these A(2A) antagonists to decrease the cells free radicals
production and oxidative stress, that are major components of 6-OHDA-induced cytotoxicity.
2009 Elsevier Ltd. All rights reserved.