Výsledky projektu ROZŠÍŘENÁ STUDIE VLIVU CHELÁTORŮ ŽELEZA U MODELU AKUTNÍHO INFARKTU MYOKARDU

Iron (Fe) chelators are used clinically for the treatment of Fe overload disease. Iron also plays a role
in the pathology of many other conditions, and these potentially include the cardiotoxicity induced by
catecholamines such as isoprenaline (ISO). The current study examined the potential of Fe chelators to
prevent ISO cardiotoxicity. This was done as like other catecholamines, ISO contains the classical catechol
moiety that binds Fe and may form redox-active and cytotoxic Fe complexes. Studies in vitro used the
cardiomyocyte cell line, H9c2, which was treated with ISO in the presence or absence of the chelator,
desferrioxamine (DFO), or the lipophilic ligand, 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone
(PCTH). Both of these chelators were not cardiotoxic and significantly reduced ISO cardiotoxicity in
vitro. However, PCTH was far more effective than DFO, with the latter showing activity only at a high,
clinically unachievable concentration. Further studies in vitro showed that interaction of ISO with Fe(II)/
(III) did not increase cytotoxic radical generation, suggesting that this mechanism was not involved.
Studies in vivo were initiated using rats pretreated intravenously with DFO or PCTH before subcutaneous
administration of ISO (100 mg/kg). DFO at a clinically used dose (50 mg/kg) failed to reduce catecholamine
cardiotoxicity, while PCTH at an equimolar dose totally prevented catecholamine-induced mortality and
reduced cardiotoxicity. This study demonstrates that PCTH reduced ISO-induced cardiotoxicity in vitro
and in vivo, demonstrating that Fe plays a role, in part, in the pathology observed.

Coronary heart disease and in particular its most serious form – acute myocardial infarction (AMI) – represents
the most common cause of mortality in developed countries. Better prognosis may be achieved by understanding
the etiopathogenetic mechanisms of AMI. Therefore, a catecholamine model of myocardial injury, which has
appeared to be very similar to AMI in human in some aspect, was used. Male Wistar:Han rats were randomly
divided into two groups: control group (saline) and isoprenaline group (ISO; synthetic catecholamine,
100 mg.kg–1 subcutaneously [s.c.]). After 24 hours, functional parameters were measured, biochemical
markers in the blood and metals content in the heart tissue were analysed and histological examination was
performed. ISO caused marked myocardial injury that was associated with myocardial calcium overload. Close
correlation between myocardial impairment (i.e. serum TnT, stroke volume index and wet ventricles weight)
and the levels of myocardial calcium was observed. Direct reactive oxygen species (ROS) involvement was
documented only by non-significant increase in malonyldialdehyde 24 hours after ISO injury. Moreover, myocardial
element analysis revealed no significant changes as for the content of zinc and iron while selenium and
copper increased in the ISO group although it reached statistical significance only for the latter.

High levels of catecholamines are cardiotoxic and may trigger acute myocardial infarction (AMI). Similarly,
the synthetic catecholamine isoprenaline (ISO) evokes a pathological state similar to AMI. During
AMI there is a marked increase of free iron and copper which are crucial catalysts of reactive oxygen
species formation. Rutin, a natural flavonoid glycoside possessing free radical scavenging and iron/copper
chelating activity,may therefore be potentially useful in reduction of catecholamine cardiotoxicity aswas
previously demonstrated after its long-term peroral administration.
Male Wistar:Han rats received rutin (46 or 11.5mgkg−1 i.v.) alone or with necrogenic dose of ISO
(100mgkg−1 s.c.). Haemodynamic parameters were measured 24 h after drug application together with
analysis of blood, myocardial content of elements and histological examination. Results were confirmed
by cytotoxicity studies using cardiomyoblast cell line H9c2.
Rutin in a dose of 46mgkg−1 aggravated ISO-cardiotoxicity while the dose of 11mgkg−1 had no effect.
These unexpected results were in agreement with in vitro experiments, where co-incubation with larger
concentrations of rutin significantly augmented ISO cytotoxicity. Our results, in contrast to previous studies
in the literature, suggest that the reported positive effects of peroral administration of rutin were
unlikely to have been mediated by rutin per se but probably by its metabolite(s) or by some other, at this
moment, unknown adaptive mechanism(s), which merit further investigation.

Lactoferrin is recently under intense
investigation because of its proposed several pharmacologically
positive effects. Based on its iron-binding
properties and its physiological presence in the human
body, it may have a significant impact on pathological
conditions associated with iron-catalysed reactive
oxygen species (ROS). Its effect on a catecholamine
model of myocardial injury, which shares several
pathophysiological features with acute myocardial
infarction (AMI) in humans, was examined. Male
Wistar rats were randomly divided into four groups
according to the received medication: control (saline),
isoprenaline (ISO, 100 mg kg-1 s.c.), bovine lactoferrin
(La, 50 mg kg-1 i.v.) or a combination of
La ? ISO in the above-mentioned doses. After 24 h,
haemodynamic functional parameters were measured,
a sample of blood was withdrawn and the heart was
removed for analysis of various parameters. Lactoferrin
premedication reduced some impairment
caused by ISO (e.g. a stroke volume decrease, an
increase in peripheral resistance and calcium overload).
These positive effects were likely to have been
mediated by the positive inotropic effect of lactoferrin
and by inhibition of ROS formation due to chelation
of free iron. The failure of lactoferrin to provide
higher protection seems to be associated with the
complexity of catecholamine cardiotoxicity and with
its hydrophilic character.