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CURRICULUM FUNDAMENTAL

Exerccio e Stress Oxidativo Cardaco [48]

ANTNIO ASCENSO, JOS MAGALHES, JOS SOARES, JOS OLIVEIRA, JOS ALBERTO DUARTE
Faculdade de Cincias do Desporto e de Educao Fsica Universidade do Porto, Porto
Rev Port Cardiol 2003; 22 (5) : 651-678

RESUMO
O tecido muscular cardaco encontra-se
sujeito a um vasto conjunto de situaes
perturbadoras da sua homeostasia, entre os
quais o exerccio fsico. Se, por um lado, o
exerccio parece apresentar-se como uma
actividade recomendada do ponto de vista da
sade, constitui-se, por outro lado, como uma
agresso orgnica favorvel produo
acrescida de espcies reactivas de oxignio
(ERO). Associadas a mecanismos fundamentais
no metabolismo celular, estas ERO tm sido,
igualmente, relacionadas com a etiologia e
fisiopatologia de algumas patologias cardacas.
O tecido muscular cardaco possui uma taxa
metablica oxidativa elevada e uma actividade
das principais enzimas antioxidantes
relativamente baixa, o que parece torn-lo
susceptvel a fenmenos de leso tecidual por
stress oxidativo aps um perodo de exerccio
agudo. Contudo, quando repetido de forma
sistemtica, o exerccio poder constituir-se
como um importante estmulo para o
incremento dos diferentes sistemas
antioxidantes cardacos, dos quais se destacam
os da glutationa (GSH) e os associados
actividade de algumas enzimas antioxidantes
como a superxido dismutase (SOD) e a
glutationa peroxidase (GPX), que visam a
proteco do msculo cardaco. O treino de
resistncia parece, assim, induzir melhoria das
principais defesas antioxidantes, promovendo
efeitos positivos na funcionalidade cardaca e
protegendo o corao perante situaes
nefastas indutoras de stress oxidativo adicional.
No entanto, alguns dos mecanismos
relacionados com este efeito de tolerncia
cruzada perante diferentes estmulos indutores
de toxicidade cardaca parecem no encontrar-se, ainda, totalmente compreendidos.
Palavras-Chave
Stress oxidativo; Msculo cardaco; Exerccio agudo;
Treino; Tolerncia

ABSTRACT
Exercise and Cardiac Oxidative Stress
Cardiac muscle is frequently affected by many
stimuli responsible for loss of cell homeostasis,
including physical exercise. While exercise
has been presented as a recommended activity
for health reasons, it also provides favorable
conditions for additional production of reactive
oxygen species. These compounds are
associated with fundamental mechanisms of
cell metabolism but are also related to the
etiology and pathophysiology of some cardiac
diseases. Cardiac muscle tissue has a high
oxidative metabolic rate and relatively low
activity of the main antioxidant enzymes, which
could enhance its susceptibility to oxidative
injury after acute exercise. However, physical
training could be considered an important
stimulus for the different antioxidant systems
like glutathione and those related to the
activity of some important antioxidant enzymes
in myocardial protection such as superoxide
dismutase and glutathione peroxidase.
Endurance training seems to induce upregulation in some antioxidant defenses,
protecting cardiac muscle in potentially
harmful situations that induce additional
oxidative stress. Nevertheless, the mechanisms
related to this cross-tolerance effect of training
are not yet well understood.

Key words
Oxidative stress; Cardiac muscle; Acute exercise;
Training; Tolerance

Recebido para publicao: Outubro de 2002 Aceite para publicao: Fevereiro de 2003
Received for publication: October 2002 Accepted for publication: February 2003

651

652

INTRODUO

INTRODUCTION

os ltimos anos a prtica de exerccio fsico tem sofrido uma expanso massiva,
estando associada a objectivos distintos entre
os quais a melhoria da performance em competio e/ou a preveno da doena e promoo
da sade. De facto, existem hoje evidncias
claras que suportam a importncia do exerccio
regular na preveno e/ou no controlo de algumas doenas crnicas (1). No entanto, inevitavelmente, o aumento do consumo de oxignio
(O2) induzido pelo exerccio apresenta-se como
uma situao favorvel produo acrescida
de espcies reactivas de O2 (ERO) e, aparentemente, ao incremento do stress oxidativo a nvel celular, tecidual e orgnico (2, 3). Uma vez
que cerca de 2-5 % do O2 consumido pode resultar na produo de ERO, um incremento da
taxa metablica oxidativa cardaca decorrente
do exerccio fsico constitui-se como um factor
predisponente para a produo acrescida de
ERO a nvel mitocondrial, com alteraes no
estado redox celular e aumentos dos indicadores directos e indirectos de leso oxidativa
tecidual (4). Contudo, esta situao, repetida no
tempo, poder constituir um forte estmulo modelador dos diferentes sistemas antioxidantes
cardacos (5-8).
Parece consensual que o exerccio agudo
promove, a nvel cardaco, um incremento dos
indicadores de leso celular por stress oxidativo bem como, quando realizado regularmente,
alteraes crnicas no sistema de defesa antioxidante (para refs. ver 5, 6). Por outro lado, e uma vez
que o exerccio praticado de forma regular tem
sido utilizado como medida teraputica em
doentes portadores de patologias cardacas crnicas (9-11), importa analisar a sua utilidade clnica e as potenciais adaptaes cardacas a ele
associadas. De facto, alguns autores tm sugerido o treino de resistncia como um factor
susceptvel de induzir incrementos na tolerncia s situaes adversas que exageram o
stress oxidativo cardaco (9, 12-15). Deste modo,
tendo em considerao a sua potencial aplicabilidade clnica, o objectivo da presente reviso foi o de analisar o conjunto de trabalhos
publicados na literatura sobre as repercusses
cardacas agudas e crnicas induzidas pelo
exerccio, bem como sobre o efeito de tolerncia cruzada motivada pelo exerccio relativamente a alguns agentes indutores de alterao
da homeostasia cardaca.

n recent years there has been a massive expansion in physical exercise for different
purposes, from improvement of competitive
performance to disease prevention and promotion of a healthier lifestyle. Indeed, there is
now clear evidence to support the importance
of regular exercise in the prevention and/or
control of certain chronic diseases (1). However,
an inevitable consequence is that the increased oxygen (O2) consumption induced by
exercise produces favorable conditions for
increased generation of reactive oxygen species (ROS) and apparently an increase in oxidative stress at the cell, tissue and organic level
(2, 3)
. Since around 2-5 % of O2 consumed can
result in ROS generation, an increased cardiac
oxidative metabolic rate arising from physical
exercise becomes a predisposing factor for
increased ROS production at the mitochondrial
level, leading to alterations in the cells redox
state and increases in the direct and indirect
markers of tissue oxidative injury (4). However,
if this situation is repeated over time, it may
have a strong modulating effect on various
cardiac antioxidant systems (5-8).
There appears to be agreement that acute
exercise promotes an increase in markers of
cell damage from cardiac oxidative stress, as
well as chronic alterations in the antioxidant
defense system when practiced regularly (for
references, see (5, 6)). On the other hand, and
since regular exercise has been used as a therapeutic measure in patients with chronic heart
disease (9-11), it is important to analyze its clinical utility and the associated potential cardiac
adaptations. Some authors have suggested that
endurance training may lead to increased tolerance to adverse situations that greatly exacerbate cardiac oxidative stress (9, 12-15). Thus, in
view of its potential clinical applications, the
aim of this review was to analyze the works in
the literature on the acute and chronic cardiac
effects of exercise, as well as on the cross-tolerance effect produced by exercise on certain
agents responsible for changes in cardiac homeostasis.
CELLULAR OXIDATIVE STRESS
There appears to be consensus nowadays
that free radicals are involved in many biological processes, often playing a crucial role.
These compounds appear to be responsible for
numerous physiological processes at the gene-

STRESS OXIDATIVO CELULAR

Parece, hoje em dia, consensual que os radicais livres participam em inmeros processos
biolgicos, muitas vezes de forma decisiva. Estes compostos parecem ser responsveis por
numerosos processos fisiolgicos nos quais se
incluem a transduo do sinal, nas clulas em
geral e, particularmente no sistema cardiovascular, a regulao do tnus vascular e a agregao plaquetria (16-18). Contudo, ao nvel da
fisiopatologia de um leque variado de doenas,
especialmente as degenerativas, que os radicais livres ganham predominncia. De facto,
um crescente nmero de trabalhos sobre este
tpico tem sido desenvolvido em campos distintos, desde a bioqumica bsica at s reas
clnicas e aplicadas (para refs. ver 19).
Um radical livre , por definio, uma molcula ou um tomo portador de electres desemparelhados numa das suas rbitas, situao
que lhe confere instabilidade e reactividade
bioqumica (19-22). Os radicais livres adquirem a
estabilidade qumica interagindo com substncias no-radicais, ao cederem o electro desemparelhado a uma molcula estvel, formando um radical reduzido, ou aceitando um
electro a partir de uma molcula estvel, convertendo-a em radical oxidado. Os diferentes
radicais existentes nos sistemas biolgicos encontram-se, habitualmente, associados a 4 tomos, nomeadamente o de carbono (C), o de enxofre (S), o de azoto (N) e o oxignio (O) (23). Em
funo do tomo base que est na gnese da
formao dos radicais, assim se designam, genericamente, por radicais de C, S, N ou O.
Existem, no entanto, outros compostos celulares que, embora no possam, por definio, ser
designados radicais livres, por no possurem
qualquer electro desemparelhado na sua estrutura qumica, so extremamente reactivos e
potencialmente geradores de radicais livres.
Deste modo, habitual utilizar-se a denominao mais abrangente de espcies reactivas
(ER) para englobar quer os radicais livres quer
os compostos no radicais seus percursores.
Uma vez que durante o exerccio o aumento do
consumo de O2 parece ser um dos factores predisponentes produo de ER, ser dada uma
ateno particular s espcies reactivas de O2
(ERO). Destas, as mais referidas na literatura
so o radical superxido (O2-), o perxido de
hidrognio (H2O2) e o radical hidrxilo (OH).
O desequilbrio entre os mecanismos de
produo e de neutralizao das ERO deno-

ral cell level, including signal transduction

and, particularly the in cardiovascular system,
regulating vascular tone and platelet aggregation (16-18). However, it is in the pathophysiology
of a wide range of diseases, particularly degenerative disorders, that free radicals assume a
major role. A growing number of studies on
this topic have been carried out in various
fields, from basic biochemistry to clinical and
applied areas (for references, see (19)).
A free radical is defined as a molecule or
atom carrying an unpaired electron in one of
its orbits, which leads to instability and biochemical reactivity (19-22). Free radicals acquire
chemical stability by interacting with non-radical substances, by either giving up the unpaired electron to a stable molecule and forming a
reduced radical, or acquiring an electron from
a stable molecule, forming an oxidized radical.
The various radicals found in biological
systems are usually associated with four elements, namely carbon (C), sulfur (S), nitrogen
(N) and oxygen (O) (23). Depending on the atom
involved in the formation of radicals, they are
generally called C, S, N or O radicals. There
are, however, other cellular compounds that although not strictly considered free radicals,
since they do not have any unpaired electrons
in their chemical structure, are extremely reactive and may generate free radicals. In this
case, it is usual to employ the wider term
reactive species (RS) to cover both free radicals and the non-radical compounds that are
their precursors. Since during exercise increased O2 consumption seems to be a predisposing factor for RS generation, particular attention will be given to reactive oxygen species
(ROS). Of these, those most often referred to in
the literature are the superoxide radical (O2-),
hydrogen peroxide (H2O2) and the hydroxyl radical (OH).
An imbalance between ROS production and
neutralization mechanisms is generally known
as oxidative stress, a natural process that results from the inability of antioxidant systems
to cope with increased generation of ROS (4, 24).
There are various cellular sources capable
of producing ROS, the most frequently cited
being mitochondria and the mechanisms associated with the enzymes xanthine oxidase and
nicotinamide adenine dinucleotide phosphate
(NADPH) oxidase (2, 8, 25, 26).
Although the main function of mitochondria
is to produce energy, there is evidence that
points to their being a source of ROS during

653

mina-se, habitualmente, por stress oxidativo.

Este fenmeno natural resulta de uma incapacidade dos sistemas antioxidantes em combater
a produo adicional de ERO (4, 24).
Existem vrias fontes celulares passveis de
produzir ERO, sendo as mais referidas a mitocndria, os mecanismos associados s enzimas
xantina oxidase e nicotinamida adenina dinucletido fosfato (NADPH) oxidase (2, 8, 25, 26).
Apesar da principal funo da mitocndria
ser a produo de energia, existem evidncias
que apontam as mitocndrias como fonte de
ERO durante as reaces que antecedem a fosforilao oxidativa (para refs. ver 8, 25, 27, 28-31). A formao
de intermedirios da reduo tetravalente do
O2, assim como a fuga de electres para o O2, a
partir da cadeia transportadora de electres
(CTE), particularmente dos complexos I e III,
so os mecanismos apontados como responsveis pela produo de ERO na mitocndria
(Fig. 1) (para refs. ver 27).
A determinao e quantificao directa das
ERO requerem procedimentos experimentais
complexos, dado o seu curto perodo de vida.

the reactions that precede oxidative phosphorylation (for references, see (8, 25, 27, 28-31)). The formation
of intermediates from the tetravalent reduction
of oxygen, and the loss of electrons to O2 from
the electron transport chain (ETC), particularly
from complexes I and III, are the mechanisms
suggested as being responsible for ROS production in the mitochondrion (Fig. 1) (for references,
see (27))
.
The detection and direct quantification of
ROS require complex experimental procedures
since they are short-lived. The facility with
which they rapidly give up or capture electrons
from other compounds gives them a high biochemical instability and consequently extremely short lifetimes, making it difficult to
determine their conventrations accurately (for references, see (2))
. However, experimental studies on isolated mitochondria have used various methods
to determine both ROS production at different
metabolic stages and the different quantities of
O2- and H2O2 produced (for references, see (25)). For example, it has been estimated that cardiac muscle

AcCoA

NADH
L
R

LOO*
O2

II
*OH

III

O2-

IV
Fe

SOD

2+

O2
H2O

Fig. 1 Produo de ERO na mitocndria. CAT - catalase; GPX glutationa peroxidase; SOD - superxido dismutase; LOO - radical peroxil lipdico; QH - semiquinona. Adaptado de Ji e
Leichtweis (2).

654

Fig. 1 - ROS production in the

mitochondrion. CAT - catalase;
GPX - glutathione peroxidase;
SOD - superoxide dismutase;
LOO - lipid peroxyl radical; QH
- semiquinone. Adapted from Ji
and Leichtweis (2).

GPX
CAT
2H2O

2H2O

+ O2

A facilidade com que rapidamente cedem ou

captam electres a partir de outros compostos,
confere-lhes uma instabilidade bioqumica elevada com consequentes tempos de vida bastante reduzidos, tornando-se difcil a determinao precisa das suas concentraes (para refs. ver 2).
Contudo, vrios trabalhos experimentais em
mitocndrias isoladas tm utilizado diferentes
mtodos para determinar quer a produo de
ERO nas diferentes etapas metablicas, quer
as quantidades diferenciadas de O 2- e H 2O 2
produzidos (para refs. ver 25). Por exemplo, foi estimado
que as mitocndrias do tecido muscular cardaco geram 0.3-0.6 nmol O2-/min/mg de protena, o que representa cerca de 2 % do O 2
total consumido (para refs.ver 2, 31).
A hiptese de a mitocndria ser, efectivamente, uma das principais fontes de ERO,
contribuindo por isso para a ocorrncia de
stress oxidativo, parece tambm ser reforada
por algumas evidncias indirectas de leso mitocondrial. A presena de indicadores de peroxidao lipdica, nomeadamente de malondialdedo (MDA) (32) e de substncias reactivas ao
cido tiobarbitrico (TBARS) (30), as elevadas
taxas de oxidao de diclorofluorescina (DCFH)
evidenciadas em mitocndrias isoladas de diferentes tecidos sujeitos a situaes indutoras de
alterao da homeostasia celular (29, 33, 34), sugerem
o referido organelo, simultaneamente, como local de produo e alvo de ERO. Adicionalmente, as alteraes na actividade de algumas
enzimas antioxidantes mitocondriais, aps a
aplicao de estmulos crnicos, assim como
os distrbios no estado redox da glutationa, em
mitocndrias de diferentes tecidos aps estimulao aguda, suportam a teoria da produo
mitocondrial de ERO (para refs. ver 2).
Para alm das ERO, a mitocndria parece
ser, igualmente, um dos locais de produo de
outras ER no centradas no O2, tais como as
ER de azoto, de que exemplo o radical xido
ntrico (ON) (25). Os resultados de diversos estudos sugerem a presena da xido ntrico sintetase nas mitocndrias. Esta enzima, inicialmente estudada no endotlio vascular,
responsvel pela sntese de xido ntrico a partir do percursor L-arginina, tendo sido encontradas correlaes elevadas entre os nveis de
expresso endotelial desta enzima e o seu contedo mitocondrial no msculo cardaco e esqueltico, rim e crebro (para refs. ver 25). O ON e o
O 2- podem ainda reagir conjuntamente para
formar peroxinitrito (ONOO-). Este oxidante,
bastante potente, constitui-se, tambm, como

mitochondria generate 0.3-0.6 nmol O2-/min/mg

of protein, which represents around 2 % of total O2 consumed (for references, see (2, 31)).
The hypothesis that the mitochondrion is in
fact one of the main sources of ROS, thereby
contributing to oxidative stress, would also
seem to be borne out by indirect evidence of
mitochondrial damage. The presence of increased markers of lipid peroxidation, notably
malondialdehyde (MDA) (32) and thiobarbituric
acid reactive substances (TBARS) (30), together
with the high oxidation rates of dichlorofluorescin (DCFH) shown by isolated mitochondria
from different tissues subjected to situations
that induce changes in cell homeostasis (29, 33, 34),
suggest that this organelle is simultaneously a
production site for ROS and their target. Moreover, changes in the activity of certain mitochondrial antioxidant enzymes after application
of chronic stimuli, as well as disturbances in
the glutathione redox state in the mitochondria
of different tissues following acute stimulation,
support the theory of mitochondrial ROS production (for references, see (2)).
Besides ROS, the mitochondrion also appears to be a site for the production of other
RS, such as nitrogen RS, of which one example is the nitric oxide radical (NO) (25). The results of various studies suggest that nitric
oxide synthase is found in mitochondria. This
enzyme, initially studied in vascular endothelium, is responsible for the synthesis of nitric
oxide from its precursor L-arginine, there
being a strong correlation between endothelial
levels of expression of the enzyme and its mitochondrial content in cardiac and skeletal
muscle, the kidneys and the brain (for references, see (25)).
NO and O2- can also react together to form peroxynitrite (ONOO-). This highly potent oxidant
is another important aggressive agent against
various cell structures (for references, see (25, 31)).
Although the mitochondrion is in general
the principal production site of ROS, there are
other cellular sources that may be involved in
their generation. Some are active in normal
physiological conditions, while others are activated only in particular circumstances such as
ischemia/reperfusion (I/R), administration of
drugs and intense exercise (7).
One of the main extra-mitochondrial sources of ROS production is xanthine oxidase, the
enzyme responsible for the conversion of hypoxanthine (Hx) to xanthine and thence to uric
acid (35). The production of ROS by this enzyme

655

656

mais um importante agente agressor para as diferentes estruturas celulares (para refs. ver 25, 31).
Apesar da mitocndria se apresentar, de
uma forma geral, como o principal local de
produo ERO, existem outras potenciais fontes celulares para a sua formao. Se algumas
se encontram activas em condies fisiolgicas
normais, outras so activadas apenas em circunstncias especiais, tais como situaes de
isquemia-reperfuso (I-R), administrao de
frmacos ou exerccio intenso (7).
Uma das principais fontes extra-mitocondriais de produo de ERO encontra-se associada xantina oxidase, enzima responsvel
pela converso da hipoxantina (Hx) em xantina e desta em cido rico (35). A produo de
ERO por esta enzima tem sido relacionada, em
particular, com situaes de I-R e tambm com
situaes de degradao acentuada de nucletidos de adenina, nomeadamente de ATP, com
formao de AMP, IMP, inosina e Hx, decorrentes de taxas de utilizao de ATP superiores s da sua ressntese tecidual (36-38).
A xantina oxidase, localizada principalmente no endotlio vascular da maior parte dos
tecidos, incluindo o dos msculos esqueltico
e cardaco, pode existir sob forma de desidrogenase (XDH), a qual utiliza a nicotinamida
adenina dinucletido (NAD+) como receptor de
H+, ou sob a forma de oxidase (XO), utilizando
o O2 como receptor de electres, com consequente formao de O2- (37). Duas condies fundamentais para que se verifique a converso
da enzima XDH em XO so as disponibilidades elevadas de Hx, como substrato, e o aumento das concentraes intra-celulares de
clcio (Ca2+) pela activao de proteases citoplasmticas (para refs. ver 37, 38). Em repouso e na ausncia de qualquer estmulo indutor de isquemia tecidual, esta enzima actua como uma
desidrogenase. No entanto, em situaes de
exerccio intenso, em que a taxa de degradao
de nucletidos de adenina muito elevada, ou
em condies de diminuio do fluxo sanguneo e de O2 aos tecidos, p.e. durante alguns
tipos de intervenes cirrgicas, a XDH convertida em XO (38). Quando o fluxo de O2 restabelecido, na fase de reperfuso tecidual, a
XO oxida a HX e utiliza o O2 como aceitador
de electres formando radicais O2- (Fig. 2).
A formao de ERO pelos neutrfilos polimorfonucleares (NPM) apresenta-se como outro
dos mecanismos importantes na acentuao do
stress oxidativo aps uma situao de agresso
tecidual (para refs. ver 11, 25). Durante a resposta de fase

has been linked to I/R in particular, as well as

to situations of marked degradation of adenine
nucleotides, notably ATP, with the formation of
AMP, IMP, inosine and Hx, arising from a higher rate of ATP consumption than its regeneration in tissues (36-38).
Xanthine oxidase, mainly found in the vascular endothelium of most tissues, including
skeletal and cardiac muscle, may be in the
form of xanthine dehydrogenase (XDH), which
uses nicotinamide adenine dinucleotide
(NAD+) as the H+ receptor, or in the form of
xanthine oxidase (XO), using O2 as the electron receptor, with the consequent formation of
O2- (37). Two essential conditions for the conversion of XDH to XO are the availability of high
levels of Hx as substrate, and increased intracellular concentrations of calcium (Ca2+) by the
activation of cytoplasmic proteases (for references, see (37, 38)).
At rest and in the absence of any stimulus that
might induce tissue ischemia, this enzyme acts
as a dehydrogenase. However, in situations of
intense exercise, in which the degradation rate
of adenine nucleotides is high, or in conditions
of reduced blood flow and oxygen supply to
tissues, such as during certain types of surgical procedure, XDH is converted to XO (38).
When the oxygen supply is re-established during reperfusion, XO oxidizes the Hx and uses
O2 as the electron acceptor, forming O2- radicals (Fig. 2).
ROS production by polymorphonuclear
neutrophils (PMNs) is another important mechanism in increasing oxidative stress after tissue aggression (for references, see (11, 25)). During the acute
phase response, neutrophils are attracted to
the damaged region and release lysosomic
enzymes and ROS (11). In the presence of agents
that destabilize the neutrophil plasma membrane, the production of O2- radicals is stimulated by the action of the enzyme NADPH oxidase, which uses NADPH as cofactor, directly
reducing O2 (8, 25). Besides O2-, neutrophils have
also been reported as the source of a series of
other RS, including ON, H2O2 and hypochlorous acid (HOCl), to combat invading pathogens
(Fig. 3) (for references, see (8, 11, 25)). In fact, whereas this
inflammatory response is considered of crucial
importance in the repair and regeneration of
damaged tissue, the reactive species produced
by neutrophils can cause secondary damage by
degrading the surrounding tissue and thus aggravating the injury (for references, see (11)).
The cells making up different types of tis-

O2

AU
O2-

Clulas endoteliais

O2

AU

O2

Clulas endoteliais
HX

AU
XO

Clula muscular cardaca

O2
AMP

IMP

INO

AU

HX
XO

SOD

H2O2

O2
ADP

Protease b Protease a
OH-

ATP

Fe2+

XDH

aguda, os neutrfilos so atrados para a regio

lesada libertando enzimas lisossmicas e ERO (11).
Na presena de agentes perturbadores da estabilidade da membrana plasmtica dos neutrfilos, estimulada a produo de radicais O2por aco da enzima NADPH oxidase, a qual
utiliza o NADPH como cofactor, reduzindo directamente o O2 (8, 25). Para alm do O2-, os neutrfilos tm sido tambm referidos como fonte
de um conjunto de outras ER, incluindo o ON,
o H2O2 e o cido hipocloroso (HOCl) para combater os agentes patognicos invasores (Fig. 3)
(para refs. ver 8, 11, 25)
. De facto, enquanto esta resposta
inflamatria considerada de importncia cr-

Fig. 2 Papel da xantina oxidase

(XO) na produo de ERO no tecido muscular cardaco e nas clulas endoteliais. HX - hipoxantina; AU - cido rico; XDH xantina desidrogenase; INO - inosina; IMP - inosina monofosfato;
AMP, ADP, ATP - adenosina
mono, di e tri fosfato. Adaptado
de Ji e Leichtweis (2).
Fig. 2 Role of xanthine oxidase
(XO) in ROS production in cardiac muscle tissue and endothelial cells. Hx - hypoxanthine; UA
- uric acid; XDH - xanthine
dehydrogenase; INO - inosine;
IMP - inosine monophosphate;
AMP, ADP, ATP - adenosine
mono-, di- and triphosphate.
Adapted from Ji and Leichtweis (2).

sue have antioxidant enzymes that play a crucial role in protective mechanisms against oxidative stress, of which superoxide dismutase
(SOD), catalase (CAT) and glutathione peroxidase (GPX) are the most commonly cited. Each
of these enzymes is able to catalyze reactions
that lead to the production of less reactive species or the neutralization of ROS (39). In addition to these enzymes, there are other nonenzyme antioxidant substances, both
endogenous and exogenous, such as glutathione (GSH), vitamins C and E, and lipoic
acid, that play an important role in the neutralization or attenuation of the effects caused

NPM
NAPH
NAPD oxidase
O2

O2-

SOD

CI

MPO

HOCI

H2O2
Fe2+

OH*

Fig. 3 Formao de ERO nos

neutrfilos polimorfonucleares
(NPM). NADPH Ox - NADPH
oxidase; MPO - mieloperoxidase;
SOD - superxido dismutase.
Adaptado de Ji e Leichtweis (2).
Fig. 3 ROS formation in polymorphonuclear neutrophils
(PMNs). NADPH Ox - NADPH
oxidase; MPO - myeloperoxidase;
SOD - superoxide dismutase.
Adapted from Ji and Leichtweis (2).

657

658

tica na reparao e regenerao da leso tecidual, as espcies reactivas produzidas pelos

neutrfilos podem constituir-se como agentes
indutores de leso secundria, degradando o
tecido envolvente e contribuindo para acentuar
os processos de leso (para refs. ver 11).
As clulas que compem os diferentes tecidos possuem enzimas antioxidantes que participam de forma decisiva nos mecanismos de
proteco tecidual contra o stress oxidativo, das
quais a superxido dismutase (SOD), a catalase (CAT) e a glutationa peroxidase (GPX) so
as mais referidas. Cada uma destas enzimas
tem capacidade de catalisar reaces que conduzem produo de espcies menos reactivas
ou neutralizao das ERO (39). Para alm destas enzimas, existem outras substncias antioxidantes no enzimticas, endgenas e exgenas, tais como a glutationa (GSH), as vitaminas
C e E, o cido lipico, entre outras, que participam de forma determinante na neutralizao
e atenuao dos efeitos induzidos pela produo adicional de ERO (para refs. ver 40, 41-44).
A SOD promove a dismutao do radical
O2-, formando H2O2 e O2. Nos mamferos existem duas isoformas desta enzima, que so diferentes na sua localizao celular, assim como
no io metlico que se encontra ligado no seu
local activo. As referidas isoformas catalisam a
dismutao do anio O2- com uma eficincia similar (45). Assim, a Cu/Zn SOD encontra-se fundamentalmente no citosol, enquanto a Mn SOD
existe em quantidades mais elevadas na matriz
mitocondrial (39).
A CAT, cuja principal funo a de catalisar a decomposio do H 2O 2 em gua e O 2,
existe em concentraes mais elevadas nos peroxissomas do que nas mitocndrias e mais
abundante nos tecidos predominantemente oxidativos do ponto de vista metablico (39). Embora se verifique alguma sobreposio de funes entre a CAT e a GPX, as duas enzimas
diferem na sua afinidade pelo H2O2 como substrato (39). A GPX dos mamferos tem uma afinidade muito maior pelo H 2O 2 do que a CAT
(GPX Km=1M vs CAT Km=1mM), o que significa que com concentraes baixas de H2O2 a
GPX possui um papel mais activo na sua remoo celular. Efectivamente, a GPX, enzima
dependente da presena de selnio, catalisa a
reduo do H2O2 em gua ou de hidroperxidos
em aldedos, utilizando a glutationa reduzida
(GSH) como dador de electres. Localizada
quer no citosol, quer nas mitocndrias e apresentando cerca de 45 % e 55 % da sua activi-

by increased ROS production (for references, see (40-44)).

SOD promotes the dismutation of the O2- radical, forming H2O2 and O2. There are two isoforms of this enzyme in mammals, which differ
in their cell location, as well as in the metal
ion that is linked to its active site. These isoforms catalyze the dismutation of the O2- anion
with similar efficiency (45) . Cu/Zn-SOD is
mainly found in the cytosol, whereas Mn-SOD
is found in larger quantities in the mitochondrial matrix (39).
CAT, whose main function is to catalyze the
decomposition of H 2O 2 into water and O 2, is
present in greater concentrations in peroxisomes than in mitochondria and is more abundant in predominantly oxidative tissues (39). Although the functions of CAT and GPX overlap
to some extent, the two enzymes differ in their
affinity for H 2O 2 as substrate (39). Mammalian
GPX has a much greater affinity for H2O2 than
CAT (GPX Km=1 M vs. CAT Km= 1mM),
which means that with low H 2O 2 concentrations, GPX has a more active role in its removal from cells. GPX, which depends on the
presence of selenium, catalyzes the reduction
of H2O2 to water or hydroperoxides to aldehydes, using reduced glutathione (GSH) as electron donor. Located in either the cytosol or the
mitochondria, with around 45 % and 55 % of
its total activity respectively in each of these
cell compartments, this enzyme functions as
one of the main neutralizing mechanisms of
hydroperoxides from different cell sources (6).
As GSH is oxidized by GPX and forms glutathione disulfide (GSSG), cells must have a
way of regenerating GSH. This reaction is catalyzed by the enzyme glutathione reductase
(GR), which uses NADPH as cofactor and reducing agent, converting GSSG back into GSH
(Fig. 4).
Intracellular GSH levels depend on the effects of the enzymes GPX and GR on the cells
redox GSH/GSSG balance, as well as on mechanisms of intracellular GSH resynthesis and
uptake from the blood (for references, see (39, 46)). Although
this compound can also be produced in other
tissues, most de novo GSH synthesis takes
place in the liver. Particularly in situations of
increased oxidative stress, the liver releases
higher quantities of GSH into the blood, most
of the remaining tissues needing to import
GSH from the circulation via the -glutamyl
cycle, a process for which certain enzyme complexes are crucial (39). Due to the difficulty in
transporting this tripeptide across the plasma
membrane while preserving its original struc-

dade total em cada um dos referidos compartimentos celulares, respectivamente, esta enzima
assume-se como um dos principais neutralizadores de hidroperxidos provenientes das diferentes fontes celulares (6).
Devido ao facto da GSH ser oxidada pela
GPX e formar glutationa oxidada (GSSG), as
clulas devero possuir uma via de regenerao de GSH. Esta reaco catalisada pela
enzima glutationa redutase (GR), a qual utiliza
o NADPH como cofactor e agente redutor,
transformando a GSSG novamente em GSH
(Fig. 4).
Os nveis intra-celulares de GSH dependem
da interaco das enzimas GPX e GR sobre o
equilbrio redox celular GSH/GSSG, bem como
dos mecanismos de ressntese intra-celular e
de captao de GSH, a partir do sangue (para refs. ver
39, 46)
. Apesar deste composto poder ser igualmente sintetizado noutros tecidos, a maior
parte da sntese de novo de GSH ocorre no
fgado. Deste modo, particularmente em situaes de stress oxidativo adicional, o fgado exporta quantidades superiores de GSH para o
sangue, tendo a maioria dos restantes tecidos
necessidade de importar GSH da circulao
via ciclo g-glutamil, no qual interagem alguns
complexos enzimticos determinantes neste
processo (39). De facto, devido dificuldade de
transporte deste tripeptdeo atravs das membranas plasmticas mantendo a sua estrutura
original, torna-se necessria a sua cleavage
nos diferentes aminocidos que o compem
para subsequente translocao membranar,
passo que catalisado pela enzima -glutamiltranspeptidase (GT). Por sua vez, a sntese de
novo de GSH a nvel intracelular reiniciada
pela -glutamilcisteina-sintetase (GCS), que

GSSG
R

ture, it must be cleaved into its various component amino acids to enable subsequent transmembrane transport, a step that is catalyzed by
the -glutamyl transpeptidase (GT). De novo
synthesis of GSH at the intracellular level is in
turn re-initiated by -glutamylcysteine synthetase (GCS), the catalyst for the formation of
the initial peptide of the cycle, by associating
glutamate and cysteine. This step is a limiting
factor in the rate of GSH production and is
controlled by negative feedback from GSH.
The final reaction in GSH synthesis, the linking of glycine to the above dipeptide, is catalyzed by the enzyme GSH synthetase (GS). The
rate of intracellular GSH production in each
type of tissue depends on the activity of these
synthesizing enzymes (for a detailed review, see (39, 46-48)).
OXIDATIVE STRESS
AND HEART DISEASE
Besides the many recent studies that confirm the crucial importance of ROS in the stimulation and activation in the heart of signaling mechanisms that determine cell survival
and adaptation (49-51), the consequences of marked increases in their production, together with
variations in the levels of certain antioxidant
compounds, have been investigated in the context of heart disease (43, 52-55). Cardiac pathologies, particularly those with an ischemic
and/or I/R component (e.g. myocardial infarction), have been the subject of numerous extensive studies, although the mechanisms underlying their pathophysiology are still poorly
understood (42, 43, 56). Nevertheless, there is a good
deal of scientific evidence that ROS production systems and antioxidant systems are associated with the genesis, manifestation and dev-

NADP

GPX-Se

GR

GSH

NADPH

Vitamin E

Ascorbate

DHI A

Ascorbate

-LA

Vitamin E

Fig. 4 Ilustrao da interaco

entre o cido a-lipoico (a-LA),
glutationa (GSH), vitamina C (ascorbato) na reciclagem da vitamina E. Ascorbato (radical ascorbato); Vitamina E (radical
vitamina E), DHLA (cido dihidrolipoico; R (radicais livres);
GSSG (glutationa disulfido oxidada) Adaptado de Ji (6).
Fig. 4 Illustration of interaction
between -lipoic acid (-LA),
glutathione (GSH), and vitamin C
(ascorbate) in the recycling of vitamin E. Ascorbate (ascorbate
radical); Vitamin E (vitamin E
radical), DHLA (dihydrolipoic
acid); R (free radicals); GSSG
(oxidized glutathione). Adapted
from Ji (6).

659

catalisa a formao do peptdeo inicial do ciclo, associando o glutamato e a cisteina. Este

passo limitador da taxa de sntese de GSH e
controlado por feedback negativo da GSH. A
reaco final da sntese de GSH, correspondente ligao da glicina ao dipeptdeo anteriormente referido, catalisada pela enzima
GSH sintetase (GS). A taxa de sntese intracelular de GSH em cada tecido depende da actividade destas enzimas sintetizadoras (para reviso detalhada ver 39, 46-48)
.
STRESS OXIDATIVO
E PATOLOGIA CARDACA

660

Para alm dos inmeros trabalhos recentes

que confirmam a importncia decisiva das
ERO na estimulao e activao, a nvel cardaco, de mecanismos de sinalizao determinantes na sobrevivncia e adaptao celulares (49-51),
as consequncias da sua produo acrescida e
exagerada, assim como as variaes no contedo de alguns compostos antioxidantes tm
sido objecto de investigao no mbito da patologia cardaca (43, 52-55). De facto, as patologias
cardacas, particularmente aquelas com uma
componente isqumica e/ou de I-R (e.g. enfartos de miocrdio) tm sido alvo de numerosos
e extensivos estudos, embora os mecanismos
subjacentes sua fisiopatologia no se encontrem, ainda, bem compreendidos (42, 43, 56). Contudo, h numerosas evidncias cientficas que
associam os sistemas de produo de ERO e
os sistemas antioxidantes origem, manifestao e desenvolvimento destas patologias (42, 43, 55).
Os episdios de I-R do miocrdio representam um problema clnico relevante, associado,
por exemplo, realizao de angioplastias ou
de cirurgias de bypass coronrio. A leso do
tecido muscular cardaco induzida por fenmenos de I-R promove disfuno contrctil, arritmias, assim como danos irreversveis nos cardiomicitos (43). De facto, tem sido considerado
que estas alteraes esto parcialmente relacionadas com situaes de stress oxidativo celular (para refs. ver 42, 43, 55). Embora possa ficar em
aberto a eventual possibilidade do stress oxidativo ocorrer em paralelo ou ser uma consequncia do desenvolvimento das patologias
com uma componente de I-R, existem evidncias experimentais da produo de ERO e do
seu estreito envolvimento na origem dessas patologias e das disfunes cardacas (30, 42, 57, 58).
A anlise das alteraes nos indicadores de
leso oxidativa de algumas estruturas celulares
cardacas refora o papel da produo acresci-

elopment of these pathologies (42, 43, 55).

Episodes of myocardial I/R are a major
clinical problem, being associated for example
with angioplasty and coronary bypass surgery.
Cardiac muscle injury caused by I/R can lead
to contractile dysfunction and arrhythmias, as
well as irreversible damage to cardiomyocytes (43).
It is thought that these alterations can be related to situations of cellular oxidative stress (for references, see (42, 43, 55))
. Although it remains an open
question whether oxidative stress occurs in parallel with or as a consequence of the development of pathologies with an I/R component,
there is experimental evidence that ROS production is closely linked to the genesis of
these pathologies and cardiac dysfunctions (30, 42,
57, 58)
.
Analysis of changes in markers of oxidative
injury in certain cardiac cell structures confirms the role of increased ROS production in
the origin and development of this type of pathology and dysfunction. Increases in proteinoxidation markers resulting from I/R support
the hypothesis that oxidative injury to myofilaments may be one possible explanation for
changes in myocardial contractile function arising from exposure to this type of stimulus (59).
Furthermore, significant alterations in membrane phospholipid elements (e.g. cardiolipin)
have been described in cardiac mitochondria
subjected to I/R, thus compromising their selective permeability (60).
Another indirect experimental model used
to assess the importance of oxidative stress in
the etiology and pathophysiology of heart failure and myocardial infarction is the analysis of
tissue and plasma expression of certain antioxidant compounds (42). Studies have demonstrated a clear tendency for reduced antioxidant
capacity immediately following myocardial infarction (61, 62). Chamorro et al. (63) have shown the
important antioxidant role of serum uric acid
in reducing the severity of ischemic infarction,
observing a 12 % increase in the odds ratio for
a favorable clinical outcome for each mg/dl increase in serum levels of this antioxidant.
Although some experimental studies have
failed to demonstrate the cardioprotective effect of antioxidant supplements (42, 64), there is a
large body of work that suggests a relationship
between cardiac levels of certain antioxidants
and the extent of cardiac damage and dysfunction through I/R. For example, circadian variations in melatonin (65) and concentrations of lipoic acid (66) and vitamin E (56) , as well as

da de ERO na origem e desenvolvimento deste

tipo de patologias e disfunes. O incremento
de indicadores de oxidao proteica resultantes de situaes de I-R suporta a hiptese de
que a leso oxidativa dos miofila- mentos poder constituir-se como uma explicao plausvel para as alteraes da funo contrctil do
miocrdio decorrentes da exposio a este tipo
de estmulos (59). Adicionalmente, tm sido descritas em mitocndrias cardacas, sujeitas a IR, alteraes significativas nos elementos fosfolipdicos das membranas (e.g. cardiolipina),
comprometendo, assim, a sua permeabilidade
selectiva (60).
Um outro modelo experimental indirecto
utilizado para avaliar a importncia do stress
oxidativo na etiologia e fisiopatologia da insuficincia cardaca e do enfarto de miocrdio a
anlise da expresso tecidual e plasmtica de
alguns compostos antioxidantes (42). Alguns estudos demonstraram, imediatamente aps a
ocorrncia de um enfarto de miocrdio, um
perfil antioxidante tpico com uma tendncia
clara para uma reduo da capacidade antioxidante (61, 62). Chamorro e col (63) demonstraram a
importncia antioxidante do cido rico srico
na diminuio da severidade de enfartos isqumicos, tendo verificado um aumento de 12 %
no odds ratio para um outcome clnico favorvel por cada mg/dl de incremento dos nveis
sricos deste antioxidante.
Apesar de alguns estudos experimentais terem falhado na demonstrao do efeito cardioprotector da suplementao antioxidante (42, 64),
existe um vasto conjunto de trabalhos que sugerem uma relao entre os nveis cardacos de
alguns antioxidantes e a exuberncia da leso/disfuno cardaca por I-R. Por exemplo,
as variaes circadianas de melatonina (65), as
concentraes de cido lipoico (66), de vitamina
E (56) assim como a suplementao com combinaes de antioxidantes (-caroteno, vitamina
C e E) (67) parecem influenciar a resposta tecidual do msculo cardaco quando sujeito a fenmenos de I-R. Estudos in vitro revelaram um
efeito protector da metalotionina, um importante antioxidante pela sua aco na captao
de ies ferro e consequente reduo da formao de radicais OH, contra a aco txica do
H2O2 em cardiomicitos (68). A escassez de trabalhos com humanos que analisem a referida
relao prende-se com a bvia impossibilidade
de obteno de amostras de tecido cardaco
para anlise dos indicadores de stress oxidativo
e de leso oxidativa.

supplements of antioxidant combinations (carotene, vitamins C and E) (67), appear to influence the response of cardiac muscle tissue
subjected to I/R. In vitro studies have revealed
the protective effect of metallothionein, an important antioxidant due to its uptake of iron
ions and consequent reduction in the formation
of OH radicals, against the toxic action of
H2O2 in cardiomyocytes (68). The paucity of studies analyzing this relationship in humans reflects the understandable difficulty in obtaining samples of cardiac tissue in order to
analyze markers of oxidative stress and injury.
Against this background of direct and indirect evidence, in both humans and animal
models, there appears to be a clear relation
between oxidative stress and cardiac pathologies resulting from I/R. Thus, from the clinical point of view, any measures designed to
attenuate or minimize the harmful effects of
oxidative stress should be considered. Together
with other strategies, exercise has been used
as a preventive and therapeutic measure in
certain clinical areas, particularly in cardiovascular medicine. However, as with situations
of I/R, acute exercise can also be a vehicle for
oxidative stress due to the changes it causes in
cardiac metabolism, and should therefore be
recommended only under controlled circumstances for patients with heart disease (1, 69).
ACUTE EXERCISE
AND CARDIAC OXIDATIVE STRESS
Post-neonatal heart normally has a low rate
of cell growth and slow protein turnover. These
functional characteristics suggest that the myocardium may have limited ability to adapt to
acute and/or chronic oxidative stress (7). Since
the myocardium consists of predominantly aerobic tissue, it has a higher activity in absolute
terms of most antioxidant enzymes compared to
skeletal muscle (for references, see (7)). Even so, given the
high rates of oxygen consumption and ROS
production, cardiac muscles ability to neutralize ROS appears to be limited (7). Using the activity of the mitochondrial enzyme citrate
synthase (CS) as the criterion to estimate the
oxidative capacity of the myocardium, the activity ratios of various antioxidant enzymes to
CS would suggest that this tissue has limited
antioxidant potential compared, for example,
with the liver or the vastus lateralis and soleus
muscles (the latter possessing similar oxidative
metabolic characteristics to the myocardium)

661

Perante este quadro de evidncias directas

e indirectas demonstradas, quer em humanos,
quer atravs da utilizao de modelos de experimentao animal, parece clara a relao entre o stress oxidativo e as patologias cardacas
decorrentes de situaes de I-R. Deste modo,
todas as medidas que visem atenuar ou minimizar os efeitos deletrios decorrentes do stress
oxidativo devero ser considerados do ponto de
vista clnico. Conjuntamente com outras estratgias, o exerccio tem sido utilizado como medida preventiva e teraputica em algumas
reas clnicas em geral, e na medicina cardiovascular em particular. Contudo, semelhana
das situaes de I-R, tambm o exerccio
agudo, pelas alteraes metablicas cardacas
que induz um veculo de stress oxidativo, devendo a sua prescrio ser efectuada de forma
controlada em pacientes portadores de patologias cardacas (1, 69).
EXERCCIO AGUDO
E STRESS OXIDATIVO CARDACO

662

Os coraes ps-neonatais possuem, normalmente, baixas taxas de proliferao celular

e um turnover proteico lento. Estas caractersticas funcionais sugerem que o miocrdio
possa ter uma adaptabilidade limitada ao stress
oxidativo agudo e/ou crnico (7). Como tecido
predominantemente aerbio, o miocrdio possui uma actividade da maioria das enzimas antioxidantes, em termos absolutos, mais elevada,
comparativamente do tecido muscular esqueltico (para refs. ver 7). Contudo, no msculo cardaco,
tendo em considerao as elevadas taxas de
consumo de oxignio e de produo de ERO, a
sua capacidade de neutralizar as ERO parece
ser, ainda assim, limitada (7). De facto, utilizando a actividade da enzima mitocondrial citrato sintetase (CS) como critrio de estimativa
da capacidade oxidativa do miocrdio, os ratios da actividade das vrias enzimas antioxidantes sobre a CS parecem sugerir um potencial antioxidante limitado deste tecido quando
comparado, por exemplo, com o do fgado, com
o dos msculos vastus lateralis e soleus (este
possuidor de caractersticas metablicas oxidativas similares s do miocrdio) (Quadro I).
Esta baixa actividade relativa das enzimas
antioxidantes no corao atenuada com a frequente utilizao de suplementos de SOD e
CAT durante cirurgias cardacas, por forma a
aumentar a resistncia tecidual exposio a
ERO decorrentes dos fenmenos de I-R (para refs. ver 7).

(Table I).
This relatively low antioxidant enzyme activity in the heart is attenuated during heart
surgery by frequent administration of SOD and
CAT supplements in order to increase tissue
resistance to exposure to ROS resulting from
I/R (for references, see (7)). In the light of the above, cardiac muscle would indeed appear to be extremely susceptible to oxidative stress.
The ROS sources that appear to play a
more important role in the heart are the mitochondrion and mechanisms associated with
XO. However, catecholamines, peroxisomes
and PMNs are also reported as ROS generation
sites. The increase in the cardiac mitochondrial/cytosolic activity ratio of the antioxidant
enzymes SOD, CAT and GPX following an endurance training program highlights the role of
the mitochondrion as both agent and target of
oxidative stress (12). There are two characteristics of cardiac metabolism that greatly influence ROS production: i) the high volume
and density of mitochondria, which means a
larger number of ROS sources; and ii) the
equally high rate of O2 supply to mitochondria,
which may increase the rate of electron leakage from the ETC, with the consequent formation of O2- radicals.
Even at rest, the consumption of O 2/g of
cardiac muscle is higher than that of skeletal
muscle during intense exercise (5). During exercise, coronary blood flow increases more than
four-fold, and the myocardium has a remarkable capacity to extract O 2 from the blood.
While O2 is essential for the aerobic metabolism of cardiac muscle, a high cardiac oxidative metabolic rate resulting from physical exercise may be a predisposing factor for increased
production of partially reduced forms of O2 and
its reactive derivatives (2, 8, 20, 25, 39, 70).
During and after exercise, various mechanisms are activated in different organs and
systems in order to maintain or restore cell homeostasis. Alterations in intracellular concentrations of ATP with increases in ADP and
AMP levels, reductions in glycogen reserves,
changes in temperature and pH, loss of Ca2+
ion homeostasis and uncoupled mitochondrial
respiration, among others, can be important
stimuli for the increased formation of ROS in
the myocardium during and after acute exercise (71). Besides the mitochondria, these changes can also affect the conversion of XDH into
XO. It has been suggested that XO activity in
cardiac tissue is species-specific. For example,

Quadro I
Comparao da actividade das enzimas antioxidantes do miocrdio, fgado e msculo esqueltico
Tecido
Cu/Zn

Fgado
Miocrdio
Soleus
Vastus lateralis

500
65
ND
21

SOD
Mn

Cit.

GPX
Mit.

50
21
ND
8

550
150
ND
23

430
70
ND
17

CAT

GR

GST

CS

670
39
37
12

44
1.3
1.3
0.6

940
2.1
1.1
0.5

18
72
40
8

Actividades das Cu/Zn e Mn SOD, unidades/mg proteina; GPX citoslica e mitocondrial, nmol/min/mg proteina; CAT, K x 10-2/g msculo hmido;
unidade para a expresso da actividade das restantes enzimas mmol/min/g msculo hmido; ND, no determinado. Adaptado de Ji (7).

Table I
Comparison of antioxidant enzyme activity in myocardium, liver and skelectal muscle
Tissue
Cu/Zn

Liver
Myocardium
Soleus
Vastus lateralis

500
65
ND
21

SOD
Mn

50
21
ND
8

GPX
Cyto.
Mito.

550
150
ND
23

430
70
ND
17

CAT

GR

GST

CS

670
39
37
12

44
1.3
1.3
0.6

940
2.1
1.1
0.5

18
72
40
8

Cu/Zn and Mn SOD activity, units/mg protein; cytosolic and mitochondrial GPX, nmol/min/mg protein; CAT, K x 10-2/g wet muscle; unit for the expression
of the activity of the remaining enzymes, mol/min/g wet muscle; ND, no data. Adapted from Ji (7).

Em funo do anteriormente exposto, o msculo cardaco parece ser, de facto, bastante

susceptvel ao stress oxidativo.
As fontes de ERO que, a nvel cardaco,
parecem ter um papel mais determinante, so
a mitocndria e os mecanismos associados
XO. Contudo, as catecolaminas, os peroxissomas e os NPM so tambm referidos como locais de produo de ERO. O aumento da razo
mitocondrial/citoslica cardaca da actividade
das enzimas antioxidantes SOD, CAT e GPX
aps um programa de treino de resistncia refora o papel da mitocndria enquanto agente
e alvo do stress oxidativo (12). De facto, existem
duas caractersticas metablicas que influenciam, marcadamente, a produo de ERO no
corao: i) por um lado, o elevado volume e
densidade mitocondriais que se constitui como
um factor associado a um maior nmero de
fontes de ERO e, por outro lado, ii) o igualmente elevado fluxo mitocondrial de O2, o qual
poder favorecer a taxa de leakage ou fuga de
electres a partir da CTE, com consequente
formao de radicais O2-.
Mesmo em repouso, o consumo de O2/g de
tecido cardaco superior ao consumo de O2/g
de msculo esqueltico observado durante a
realizao de exerccios intensos (5). Durante o
exerccio, o fluxo sanguneo coronrio aumenta
mais de 4 vezes e o miocrdio possui uma capacidade notvel de extrair O2 do sangue. Se o
O2 essencial para a funcionalidade do metabolismo aerbio do tecido muscular cardaco,

rat, pig and dog hearts appear to present some

XO activity, unlike rabbit myocardium. However, in humans, this enzyme has been reported as playing a crucial role in cardiac muscle
damage resulting from I/R (for references, see (7)).
There is direct evidence that acute exercise
can induce increased ROS generation in the
myocardium, as confirmed by the results of a
study by Ohkuwa et al. (72), who found an increase in cardiac levels of the OH radical. The
increase in tissue metabolic rate obviously represents a major aggression against cardiac
muscle. Moreover, alterations in antioxidant
systems have been reported, as well as in myocardial injury markers, following acute exercise. Various authors (32, 72) have observed an increase in cardiac concentrations of GSH in
young rats subjected to a short period of intense exercise. This rise may be the result of
an increased rate of GSH outflow from the
liver during exercise and a concomitant increase in GT activity (72).
Venditti and Di Meo (9) found an increase in
cardiac markers of lipid peroxidation in rats
after a period of exhausting swimming with a
load equal to 2 % of the animals weight. This
exercise induced an increase in myocardial lipid peroxidation products, MDA and hydroperoxides, which supports the hypothesis that it
increases oxidative stress and that this in turn
causes structural damage to cardiomyocytes. In
the same study, a reduction in the integrity of
plasma membranes and the sarcoplasmic reti-

663

664

uma elevada taxa metablica oxidativa cardaca decorrente do exerccio fsico poder ser
um factor predisponente para produo acrescida de formas parcialmente reduzidas de O2 e
dos seus derivados reactivos (2, 8, 20, 25, 39, 70).
Durante e aps o exerccio so activados
diversos mecanismos nos diferentes rgos e
sistemas no sentido de manter ou readquirir a
homeostasia celular. Alteraes nas concentraes intracelulares de ATP com incrementos
nas concentraes de ADP e AMP, diminuies das reservas de glicognio, modificaes
na temperatura e pH, perda de homeostasia ao
io Ca 2+ e respirao mitocondrial no acoplada, podem constituir-se, entre outros, como
estmulos importantes para a formao adicional de ERO no miocrdio durante e aps o
exerccio agudo (71). Para alm das mitocndrias, estas alteraes podem tambm condicionar a converso da XDH em XO. Efectivamente, tem sido sugerido que a actividade da
XO no tecido cardaco especfica de cada espcie. Por exemplo, coraes de ratos, porcos e
ces parecem apresentar alguma actividade da
XO, ao contrrio do que acontece com o miocrdio de coelhos. Contudo, nos humanos, esta enzima tem sido referida como tendo um papel determinante nos processos de leso muscular
cardaca decorrentes de situaes de I-R (para refs. ver 7).
Existem evidncias directas de que o exerccio agudo pode induzir aumentos na formao de ERO no miocrdio, o que pode ser confirmado pelos resultados do estudo de Ohkuwa
e col (72) os quais verificaram um incremento
dos nveis cardacos do radical OH. O incremento da taxa metablica tecidual constitui-se,
naturalmente, como uma forte agresso para o
tecido muscular cardaco. Adicionalmente, tm
sido encontradas alteraes nos sistemas antioxidantes, assim como nos marcadores de leso no miocrdio aps a realizao de exerccio agudo. Vrios autores (32, 72) observaram um
incremento das concentraes cardacas de
GSH em ratos jovens sujeitos a um curto perodo de exerccio intenso. Este aumento poder ser resultado de uma taxa acrescida de
efluxo heptico de GSH durante o exerccio e
de um concomitante incremento da actividade
da GT (72).
Venditti e Di Meo (9) verificaram, em ratos,
um incremento dos marcadores cardacos de
peroxidao lipdica aps a realizao de um
perodo de natao at exausto com uma
carga correspondente a 2 % do peso dos ani-

culum was also evident following exercise.

GSH is a non-enzyme compound, with a
protein structure, which is crucial to tissue antioxidant activity in general, and the myocardium in particular. Concentrations of this tripeptide in cardiac muscle are approximately
1-2 mM, well below those found in other tissue, notably oxidative skeletal muscle (3 mM),
the liver (6-8 mM) and the kidneys (3-4 mM) (39, 73).
Nevertheless, since the activity of the main antioxidant enzymes of the myocardium appears to
be relatively low, GSH is the most important
antioxidant found in cardiac muscle (7).
The potential protective role of GSH in
against cardiac oxidative stress induced by
acute exercise has been studied in animals
subjected to the drug buthionine sulfoximine
(BSO), an inhibitor of the enzyme -glutamylcysteine synthetase (GCS) and therefore
a depletor of intracellular GSH (74, 75). Data from
these studies demonstrated that in contrast to
the control group, the rats with GSH deficiency
did not show a further reduction in cardiac
concentrations of total glutathione with acute
exercise, presenting a 50 % lower capacity to
withstand exhausting exercise (75). A recent
study by Khanna et al. (76) showed that the
systematic administration (150 mg/kg/day) of
-lipoic acid, an important compound in the
resynthesis of ascorbate from its radical semidehydroascorbate, protects cardiac tissue from
the GSH-depleting effect of acute exercise.
Furthermore, compared to the animals in the
control group, the hearts supplemented with lipoic acid had lower levels of lipid peroxidation, as assessed by the concentration of
TBARS in the cardiac homogenate.
Although GSH can be considered the most
efficacious cardiac antioxidant compound, the
role of exogenous antioxidants in protecting
cardiac tissue during and after intense exercise
has also been studied. Benderitter et al. (77)
analyzed the relationship in rats between the
variation in cardiac concentrations of vitamin
E and markers of lipid peroxidation (TBARS)
induced by exercise. The study found a fall in
vitamin E levels in animals subjected to exhausting exercise compared to those not exercised, with no alterations in TBARS levels in
the cardiac homogenate of the two groups, a
fact that would seem to reflect the contribution
of vitamin E in combating exercise-induced
cardiac lipid peroxidation.
Another response of cardiac muscle to

mais. De facto, o referido exerccio induziu um

aumento dos produtos da peroxidao lipdica
do miocrdio, MDA e hidroperxidos, suportando a hiptese de que aumenta o stress oxidativo e este, por sua vez, induz leses estruturais nos cardiomicitos. Nesse mesmo estudo
foi evidente uma diminuio da integridade
das membranas plasmtica e do retculo sarcoplasmtico aps o exerccio.
Um composto no enzimtico, de estrutura
proteica, determinante na actividade antioxidante tecidual em geral, e do miocrdio em
particular, a GSH. As concentraes deste
tripeptdeo no msculo cardaco so de, aproximadamente, 1-2 mM, claramente inferiores
s encontradas noutros tecidos, nomeadamente
no msculo esqueltico oxidativo (3 mM), no
fgado (6-8 mM) e no rim (3-4 mM) (39, 73). No
entanto, ainda assim, uma vez que a actividade
das principais enzimas antioxidantes do miocrdio parece ser relativamente baixa, a GSH
apresenta-se como o mais importante antioxidante existente no msculo cardaco (7).
O potencial papel protector da GSH no
combate ao stress oxidativo cardaco induzido
pelo exerccio agudo foi estudado em animais
sujeitos aco de um frmaco, a butionina
sulfoximina (BSO), inibidor da enzima -glutamilcisteina sintetase (GCS), e por consequncia, depletor intra-celular de GSH (74, 75). Alguns
dados destes estudos demonstraram que, ao
contrrio dos animais do grupo controlo, nos
ratos j deplecionados em GSH no se verificou uma diminuio acrescida das concentraes cardacas de glutationa total com o exerccio agudo, tendo estes apresentado uma
capacidade de resistncia ao esforo de longa
durao cerca de 50 % inferior (75). Um estudo
recente conduzido por Khanna e col (76) demonstrou que a administrao sistemtica (150
mg/kg/dia) de cido -lipoico, composto importante na ressntese de ascorbato a partir do
seu radical semidihidroascorbato, protege o tecido cardaco do efeito depletor de GSH pelo
exerccio agudo. Adicionalmente, quando comparados com os dos animais do grupo controlo,
os coraes suplementados em cido a-lipoico
diminuiram os seus nveis de peroxidao lipdica, avaliados pela concentrao de TBARS
no homogenizado cardaco.
Apesar da GSH poder ser considerada
como o mais eficaz composto antioxidante cardaco, o papel de alguns antioxidantes exgenos tem sido, igualmente, estudado na proteco do tecido cardaco durante e aps

stress induced by various stimuli, including

exercise, is elevation of heat shock proteins
(HSP). Several studies (for references, see (71, 78)) have demonstrated that acute intense exercise appears
to promote increased production of these proteins in cardiomyocytes, particularly those of
the 70 kDa family, HSP 70.
It therefore seems clear that cardiac muscle
tissue, when stimulated by acute exercise, presents increases in markers of cell damage due
to oxidative stress, notably lipid peroxidation
and protein and DNA oxidation. Despite direct
evidence of oxidative stress and lipid peroxidation, the GSH and vitamin E systems would
appear to play a crucial role in protecting the
heart in harmful situations arising from acute
exercise.
On the other hand, the potential adaptability of the myocardium faced with systematic
aggression presents a paradox, which appears
to be O2-dependent, and therefore leads us to
carefully analyze the effect of training, particularly endurance training, on oxidative stress in
this major organ.
TRAINING AND THE CARDIAC
ANTIOXIDANT SYSTEM
There are a growing number of experimental studies that associate certain types of intervention or stimulus, such as long-term manipulation of diet, genetic overexpression of
enzymes and antioxidant compounds, and supplementation and/or deficiency of antioxidants,
with a greater or lesser susceptibility to cardiac tissue damage when it is subjected to situations that induce oxidative stress. Kim et al. (79)
demonstrated in rats that chronic calorie restriction over a period of 18.5 months can alter
antioxidant metabolism at the cell level. At the
end of this period, the authors found an increase in the activity of the cytosolic antioxidant enzymes SOD, GPX and GSH S-transferase, but no significant increase in CAT
activity. These data may point to an effect of
this intervention on the hearts antioxidant
systems, and appear to demonstrate that prolonged systematic reduction of energy input,
paradoxically, improves the efficacy of the mechanisms that neutralize agents causing cardiac tissue damage through oxidative stress.
However, the mechanisms associated with increased efficacy of antioxidant defenses after
periods of restricted calorie intake are still not
completely understood (79). In addition, some

665

exerccio intenso. Benderitter e col (77) analisaram, em ratos, a relao entre a variao das
concentraes cardacas de vitamina E e dos
marcadores de peroxidao lipdica (TBARS)
induzidas pelo exerccio. Neste estudo, verificou-se uma diminuio das concentraes cardacas de vitamina E nos animais exercitados
at exausto, comparativamente aos no
exercitados, sem quaisquer alteraes nas concentraes de TBARS dos homogenizados cardacos nos dois grupos, dado que parece traduzir o contributo da vitamina E no combate
peroxidao lipdica cardaca induzida pelo
exerccio.
Outra das respostas do tecido muscular cardaco perante situaes de stress induzido por
diferentes estmulos, nos quais se encontra o
exerccio, a elevao das protenas de choque trmico (HSP). Vrios estudos (para refs. ver 71, 78)
demonstraram que o exerccio agudo intenso
parece promover incrementos na sntese destas
protenas nos cardiomicitos, nomeadamente
das HSP da famlia 70 kDa, as HSP 70.
Parece, portanto, evidente que o tecido
muscular cardaco, quando estimulado pelo
exerccio agudo, apresenta incrementos nos indicadores de leso celular por stress oxidativo,
nomeadamente de peroxidao lipdica, oxidao proteica e do DNA. Apesar das evidncias
directas da ocorrncia de stress oxidativo e de
peroxidao lipdica, os sistemas da GSH e da
vitamina E parecem assumir um papel determinante na proteco deste rgo contra situaes deletrias decorrentes do exerccio agudo.
Por outro lado, a eventual capacidade adaptativa do miocrdio perante a agresso sistemtica, parece sugerir a existncia de um paradoxo, O2 dependente, que, inevitavelmente, nos
remete para a anlise cuidada do efeito do
treino, particularmente o de resistncia, na hipottica alterao do efeito do stress oxidativo
neste importante rgo.
TREINO E SISTEMA ANTIOXIDANTE
CARDACO

666

crescente o nmero de trabalhos experimentais que associam alguns tipos de interveno e de estmulos, tais como, manipulaes de
longo termo na dieta, sobre-expresso gentica
de enzimas e compostos antioxidantes e a suplementao e/ou depleco de compostos antioxidantes, com a maior ou menor susceptibilidade leso do tecido cardaco quando
sujeito a situaes indutoras de agresso oxi-

studies have demonstrated that inducing endogenous overexpression of antioxidant compounds appears to produce a protective effect
in both the myocardium (80) and isolated cardiomyocytes (68) when subjected to intrinsic or
extrinsic factors that trigger oxidative injury.
Cardiac dysfunction and oxidative stress
caused by I/R (81), as well as severe hypertension (82), are significantly intensified in rats following administration of BSO, with a consequent reduction in levels of GSH synthesis.
Thus, since cardiac muscle tissue appears
capable of a series of adaptations when subjected to certain chronic stimuli, the role of regular endurance training in increasing cardiac tolerance to oxidative stress is a subject worthy of
investigation.
In general, most studies designed to investigate the influence of regular exercise on cardiac antioxidant activity focus on endurance
training programs, and there is little information available on the effect of sprint training in
regulating the hearts antioxidant defenses (20).
In one study, a predominantly anaerobic training program (with significant increase in activity of the glycolytic enzyme lactate dehydrogenase in all the skeletal muscles analyzed
except the soleus) caused an increase in the
cardiac activity of the GPX and GR enzymes
of the glutathione redox cycle, without however
inducing any changes in the activity of GSH Stransferase, SOD or GSH levels in the myocardium (83).
The number of studies related to the effect
of endurance training on the antioxidant
system and damage markers of cardiac oxidative stress is, on the other hand, much greater
(Table II).
As shown by Table II, there is clearly great
diversity and variability in the effects of endurance training in modulating the various
enzyme systems of cardiac antioxidant defense.
Direct comparison of the results is hampered
by methodological differences, of which the
following are the most important: i) characteristics of the training protocols used; ii) biochemical analytical techniques used; iii) tissue
sub-fractions studied; iv) time between the end
of the training program and sacrifice of the
animals; and v) type of markers and enzymes
studied.
Since cardiac antioxidant enzymes appear
to have a relatively low level of activity, the
importance suggested by Table II of these
enzymes in cardiac protection should be the
subject of more detailed analysis, given the

dativa. Kim e col (79) demonstraram, em ratos,

que uma situao crnica de restrio calrica,
perpetuada durante 18,5 meses, pode alterar o
metabolismo antioxidante a nvel celular. Aps
este perodo, os autores verificaram um aumento da actividade das enzimas antioxidantes
citoslicas SOD, GPX e GSH S-transferase,
no tendo sido evidente qualquer incremento
significativo na actividade da CAT. Estes dados
podero indiciar um provvel efeito da referida
interveno crnica nos sistemas antioxidantes
cardacos. Estes dados parecem demonstrar
que a diminuio sistemtica e prolongada do
input energtico promove, paradoxalmente,
uma melhoria da eficcia dos mecanismos de
neutralizao de agentes indutores de leso tecidual cardaca por stress oxidativo. Contudo,
os mecanismos associados ao incremento da
eficcia dos sistemas antioxidantes aps perodos de restrio calrica no se encontram,
ainda, totalmente compreendidos (79). Adicionalmente, alguns estudos tm demonstrado que a
induo da sobre-expresso endgena de compostos antioxidantes parece promover um
efeito de proteco quer do miocrdio (80) quer
de cardiomicitos isolados (68) perante factores
intrnsecos ou extrnsecos indutores de leso
oxidativa. Fenmenos de disfuno cardaca e
de stress oxidativo induzido por I-R (81), assim
como de hipertenso severa (82) foram significativamente intensificados aps a administrao
de BSO em ratos, com consequente diminuio
dos nveis de sntese de GSH.
Desta forma, e uma vez que o tecido muscular cardaco, quando sujeito a alguns estmulos de natureza crnica, parece sensvel a
um conjunto de adaptaes, o papel do treino
de resistncia sistemtico no eventual aumento
da tolerncia cardaca ao stress oxidativo poder ser um tpico de interesse a equacionar.
De uma forma geral, a maioria dos estudos
que pretenderam investigar a influncia do
treino sistemtico na actividade antioxidante
cardaca, recorreram a programas de treino de
resistncia. Efectivamente, a informao disponvel relativamente ao efeito do treino de velocidade na regulao das defesas antioxidantes
cardacas escassa (20). Num destes trabalhos,
um programa de treino predominantemente
anaerbio (aumento significativo da actividade
da enzima glicoltica lactato desidrogenase em
todos os msculos esquelticos analisados,
com excepo do soleus) promoveu um incremento da actividade cardaca das enzimas do
ciclo redox da glutationa, a GPX e a GR sem,

importance of GSH metabolism, the possible

role of other cellular protection mechanisms
(for example HSP), and the possible decrease
in ROS production resulting from endurance
training.
The idea that regular exercise can have beneficial effects, by preserving cardiac structure
and function through its positive influence on
antioxidant defenses, can be confirmed by
analysis of the results of various studies in
which endurance training: i) induced increases
in the activity of GPX, GR (9, 12, 13, 15), CAT (12, 13, 79),
SOD and GT (15) and reduced levels of lipid
peroxidation, as determined by MDA concentrations in tissue (9) and mitochondria (12, 79); and
ii) led to increases in cytosolic GSH levels (12)
and in the activity of glucose-6-phosphate
dehydrogenase (G6PDH) (13) , an essential
enzyme in the pentose cycle, one of the main
cellular sources of NADPH.
There is evidence that the intensity of endurance training and the duration of sessions
influence the antioxidant systems ability to
adapt. Powers et al. (84) observed that different
combinations of intensity (low, moderate and
high) and duration (30, 60 and 90 minutes/
day) produced different effects on the regulation of the antioxidant enzymes SOD, CAT and
GPX in the left ventricle. None of the above
combinations induced increases in CAT and
GPX activity in both ventricles, with SOD increasing after high intensity training for all durations (right and left ventricle) and after moderate intensity for 90 minutes/day (right
ventricle).
The adaptive response of the antioxidant
system, and consequently of the expression of
the various indirect markers of oxidative tissue
damage, would also appear to be specific, to
either the type of tissue (32, 85), or the different
antioxidant systems involved (86).
Some studies have demonstrated that endurance training can cause deregulation of certain antioxidant systems or maintain them unchanged. For example, Hong and Johnson (86),
despite observing an increase in cardiac SOD
activity in trained animals, also found an unexpected increase in TBARS concentrations
and reduced CAT, GPX and GR activity. This
finding appears to be in agreement with the results of other studies in which the cardiac activity of the main antioxidant enzymes did not
in general undergo any significant change with
endurance training (85, 87, 88). It should also be noted that in the work of Leeuwenburgh et al. (85),

667

Quadro II
Efeito do treino sistemtico nas defesas antioxidantes e nos marcadores de leso oxidativa cardacos
Estudo

Modelo utilizado

Tipo de treino

Efeito

Ratos

Corrida - 10 semanas

SOD
CAT
GPX
GR
TBARS

Ramires e Ji (15)

Ratos sujeitos a I-R e

suplemento de GSH (GSHs)

Corrida 10 semanas

SOD
CAT
GPX
GR
GT
GSH (GSHs)
GSH/GSSG (GSHs)

Demirel e col (89)

Ratos

Corrida 10 semanas

SOD
GPX
MDA
HSP 72

Leichtweis e col (90)

Ratos sujeitos a I-R

Natao intensa 8-9 semanas

GT
GSH
VO2 mitocondrial

Ji e col (14)

Ratos sujeitos a I-R

Natao/Corrida 5 semanas

GSH
GPX
GR

Powers e col (84)

Ratos

Corrida 10 semanas
30, 60, 90 minutos/dia
Intensidade baixa,
moderada, elevada

SOD (intensidade elevada para

todas as duraes)
CAT
GPX

Gore e col (87)

Ratos

Corrida 10 semanas

CS
Mn SOD
Cu-Zn SOD
GPX
Cu-Zn SOD RNAm

Leeuwenburgh e col (85)

Ratos

Corrida 10 semanas

CS
GPX
GR
Mn SOD
Cu-Zn SOD
MDA
GSH
GSSG
GSH/GSSG

Tiidus e Houston (88)

Ratos

Corrida 8 semanas

SOD
CAT
GPX

Lew e Quintanilha (13)

Ratos

Corrida 10 semanas

GPX
GR
G6PDH
CAT

Somani e col (12)

Ratos

Corrida 10 semanas

SOD
CAT
GPX
MDA mitocondrial

Kim e col (79)

Ratos

Corrida 18.5 meses

Venditti e Di Meo (9)

Ratos

Corrida 10 semanas

CAT
MDA mitocondrial
MDA
GPX
GR
RCI

Hong e Johnson (86)

668

Kihlstrom (92)

Ratos e murganhos

Natao e corrida 149-159horas/animal

Kihlstrom (93)

Ratos

219-229horas/animal

GSH
TBARS
TBARS
H-R TBARS
H-R CK leakage
GSH
GSSG
Ventrculo direito G6PDH
SOD
CAT
GR

CAT - catalase; CS - citrato sintetase; G6PDH - glucose-6-fosfato desidrogenase; GPX - glutationa peroxidase; GR - glutationa redutase;
GSH - glutationa reduzida; GSSG - glutationa oxidada; GT - -glutamil transpeptidase; HSP - protenas de choque trmico;
H-R - hipxia-reoxigenao; I-R - isquemia-reperfuso; MDA - malondialdedo; RCI - ndice de controlo respiratrio; SOD - superxido dismutase;
TBARS - substncias reactivas ao cido tiobarbitrico; VO2 - consumo de oxignio.
(aumento); (diminuio); (manuteno)

Table II
Effect of regular training on antioxidant defenses and cardiac markers of oxidative injury
Study

Model

Type of training

Effect

Rats

Running 10 weeks

SOD
CAT
GPX
GR
TBARS

Rats subjected to I/R and

GSH supplement (GSHs)

Running 10 weeks

SOD
CAT
GPX
GR
GT
GSH (GSHs)
GSH/GSSG (GSHs)

Rats

Running 10 weeks

SOD
GPX
MDA
HSP 72

Leichtweis et al (90)

Rats subjected to I/R

Rigorous swimming 8-9 weeks

GT
GSH
Mitocondrial VO2

Ji et al (14)

Rats subjected to I/R

Swimming/running 5 weeks

GSH
GPX
GR

Powers et al (84)

Rats

Running 10 weeks
30, 60, 90 minutes/day
Low, moderate and
high intensity

SOD (high intensity for all

durations)
CAT
GPX

Gore et al (87)

Rats

Running 10 weeks

CS
Mn SOD
Cu-Zn SOD
GPX
Cu-Zn SOD RNAm

Leeuwenburgh et al (85)

Rats

Running 10 weeks

CS
GPX
GR
Mn SOD
Cu-Zn SOD
MDA
GSH
GSSG
GSH/GSSG

Hong and Johnson

(86)

Ramires and Ji (15)

Demirel et al (89)

669

Tiidus and Houston (88)

Rats

Running 8 weeks

SOD
CAT
GPX

Lew and Quintanilha (13)

Rats

Running 10 weeks

GPX
GR
G6PDH
CAT

Somani et al (12)

Rats

Running 10 weeks

SOD
CAT
GPX
Mitochondrial MDA

Kim et al (79)

Rats

Running 18.5 months

Venditti and Di Meo (9)

Rats

Running 10 weeks

CAT
Mitochondrial MDA
MDA
GPX
GR
RCI

Kihlstrom (92)

Rats and mice

Swimming and running 149-159 hours/animal

Kihlstrom (93)

Rats

219-229 hours/animal

GSH
TBARS
TBARS
H-R TBARS
H-R CK leakage
GSH
GSSG
Right ventricular G6PDH
SOD
CAT
GR

CAT - catalase; CS - citrate synthase; G6PDH - glucose-6-phosphate dehydrogenase; GPX - glutathione peroxidase; GR - glutathione reductase;
GSH - reduced glutathione; GSSG - oxidized glutathione; GT - - glutamyl transpeptidase; HSP - heat shock proteins; H/R - hypoxia/reoxygenation;
I/R - ischemia/reperfusion; MDA - malondialdehyde; RCI - respiratory control index; SOD - superoxide dismutase;
TBARS - thiobarbituric acid reactive substances; VO2 - oxygen consumption.
(increase); (decrease); (no change)

670

contudo, induzir quaisquer alteraes na actividade da GSH S-transferase, da SOD e do

contedo de GSH do miocrdio (83).
O volume de trabalhos relativos ao efeito do
treino de resistncia no sistema antioxidante e
marcadores de leso por stress oxidativo cardacos , por outro lado, bem mais vasto (Quadro II).
Pela anlise do Quadro II, evidente a diversidade e variabilidade dos efeitos do treino
de resistncia na modelao dos diferentes sistemas enzimticos de defesa antioxidante cardacos. Aspectos de carcter metodolgico dificultam a comparao directa dos resultados.
Entre eles, podemos destacar as diferenas entre: i) as caractersticas dos protocolos de
treino implementados; ii) as tcnicas de anlise bioqumica utilizadas; iii) as sub-fraces
teciduais estudadas; iv) os tempos que medeiam o final do programa de treino e o sacrifcio dos animais e v) o tipo de marcadores e enzimas estudadas.
Uma vez que parece verificar-se uma reduzida actividade relativa das enzimas antioxidantes cardacas, de salientar que a impor-

training induced a significant decrease in the

GSH/GSSG ratio due to a greater increase in
GSSG concentrations in the trained group.
Gore et al. (87) found that although endurance
training did not produce any change in antioxidant enzyme activity, it did raise the cardiac expression of the mRNA of Cu-Zn SOD
by 112 %, which would suggest post-translation alterations.
The role of endurance training has also
been studied as a possible factor in increasing
the hearts tolerance to certain pathologies, in
particular the cross-tolerance effect of training
in various models that are generally used to induce cardiac muscle injury, of which one
example is ischemia/reperfusion.
With the aim of studying the effect of endurance training on the biochemical and physiological response of myocardium subjected to
I/R, Ji et al. (14) observed that in the trained
group, I/R did not produce a decrease in GSH
levels or in the GSH/GSSG ratio, as had been
found in the myocardium of untrained animals.
In addition, the activity of the cardiac enzymes
GPX and GR was lower in the trained group

tncia que a leitura do Quadro II pode sugerir

relativamente a estas enzimas na proteco
cardaca dever ser objecto de uma anlise
mais cuidada, dada a relevncia do metabolismo da GSH, o possvel papel de outros mecanismos de proteco celular, de que so
exemplo as HSP e uma eventual diminuio da
produo de ERO induzida pelo treino de resistncia.
A sugesto de que o exerccio regular pode
ter efeitos benficos na preservao da estrutura e funcionalidade cardaca, por influncia
positiva nas defesas antioxidantes, pode ser
confirmada atravs da anlise dos resultados
de diversos estudos, nos quais o treino de resistncia: i) induziu aumentos na actividade da
GPX, GR (9, 12, 13, 15), CAT (12, 13, 79), SOD e GT (15) e
diminuiu os nveis de peroxidao lipdica, determinados pela concentrao de MDA tecidual (9) e mitocondrial (12, 79); ii) promoveu incrementos do contedo citoslico de GSH (12) e da
actividade da glucose-6-fosfato desidrogenase
(G6PDH) (13), enzima fundamental do ciclo das
pentoses, uma das principais fontes celulares
de NADPH.
Existem evidncias que a intensidade do
treino de resistncia e a durao das sesses
influenciam a capacidade de adaptao do sistema antioxidante. Powers e col (84) verificaram
que a combinao de diferentes intensidades
(baixa, moderada e elevada) e duraes dirias
(30, 60 e 90 minutos/dia) do treino promoveu
efeitos diferenciados na regulao das enzimas
antioxidantes SOD, CAT e GPX no ventrculo
esquerdo. Nenhuma das combinaes referidas
induziu incrementos na actividade da CAT e
GPX em ambos os ventrculos, sendo que a da
SOD aumentou aps treino de intensidade elevada para todas as duraes dirias (ventrculo
direito e esquerdo) e aps treino de intensidade moderada de 90 minutos/dia (ventrculo
direito).
A resposta adaptativa do sistema antioxidante e, consequentemente, da expresso dos
diferentes indicadores indirectos de leso oxidativa tecidual parece ser, igualmente, especfica, quer do tipo de tecido (32, 85), quer dos diferentes sistemas antioxidantes em causa (86).
Alguns trabalhos demonstraram que o
treino de resistncia pode tambm promover
desregulao ou manter inalterados alguns sistemas antioxidantes. Por exemplo, Hong e
Johnson (86), apesar de terem verificado, em animais treinados, um incremento da actividade
da SOD cardaca, encontraram um aumento

than in the control group, although GSH levels

in the trained hearts were significantly higher
than in those of the untrained animals when
subjected to I/R. In the study of Demirel et al. (89),
although endurance training did not induce
any change in SOD or GPX activity, the authors found that the trained rats were less susceptible than untrained animals to I/R-induced
lipid peroxidation as assessed by left ventricular MDA concentrations. However, although
ventricular GSH content was not determined,
the authors propose that the hypothetical increase of this important polypeptide with training, confirmed by other studies (12, 14), together
with elevated levels of cardiac HSP, in this
case HSP 72 (for references, see (71, 78)), may explain the
decrease in lipid peroxidation in the myocardium following I/R.
This tolerance to I/R injury in animals subjected to endurance training or a GSH-rich
diet has also been studied by Ramires and Ji (15).
The authors concluded that endurance training
combined with administration of a GSH supplement during the last 17 days of training can
improve the tolerance of cardiac muscle to I/Rinduced injury. This important adaptation may
be the result of preserved homeostasis of cardiac GSH metabolism associated with an increase in antioxidant activity brought about by
training.
Leichtweis et al. (90) studied the effect of a
rigorous swim training program of 8-9 weeks
duration on mitochondrial function in the
hearts of rats subjected to I/R. The results demonstrated that training produced a decrease
in the cytosolic and mitochondrial content of
GSH and total GSH (TGSH), although it improved the ability of cardiac tissue to import
extracellular GSH through increased -GT activity. Furthermore, this study showed that intensive training regimes may diminish mitochondrial function following in vitro stimulation, as shown by the decrease in the rate of
state 3 and 4 mitochondrial respiration of the
trained animals compared to the control group.
Whereas reduced state 4 respiration may not
necessarily be an indication of dysfunction, the
dramatic reduction in respiratory state 3 would
appear to demonstrate clearly that the mitochondria of the trained hearts presented a reduced capacity for oxidative phosphorylation (90).
The authors state that, since the production of
reduced equivalents from the tricarboxylic acid
cycle did not diminish with this type of rigorous training, as shown by increases in cardiac

671

672

inesperado da concentrao de TBARS e uma

diminuio da actividade da CAT, GPX e GR.
Esta tendncia parece estar de acordo com os
resultados de alguns estudos nos quais a actividade cardaca das principais enzimas antioxidantes no sofreu, regra geral, qualquer alterao significativa com o treino de resistncia (85, 87, 88). De salientar, ainda, que no trabalho
de Leeuwenburgh e col (85) o treino induziu uma
diminuio significativa da razo GSH/GSSG,
pelo aumento superior da concentrao de
GSSG no grupo treinado. O estudo de Gore e
col (87) evidenciou que o treino de resistncia,
ainda que sem promover qualquer alterao na
actividade das enzimas antioxidantes, elevou a
expresso cardaca do RNAm da Cu-Zn SOD
em 112 %, sugerindo a ocorrncia de alteraes ps translacionais.
O papel do treino de resistncia tem sido,
igualmente, estudado enquanto potencial factor
indutor de aumento da tolerncia cardaca perante algumas situaes patolgicas. Ou seja,
vrios trabalhos tm estudado o efeito de tolerncia cruzada do treino perante alguns modelos habitualmente utilizados para induzir leso
muscular cardaca, de que exemplo a isquemia-reperfuso (I-R).
Com o objectivo de estudar o efeito do
treino de resistncia na resposta bioqumica e
fisiolgica do miocrdio perante a I-R, Ji e col (14)
verificaram que no grupo treinado, a I-R no
promoveu um decrscimo dos nveis de GSH
nem da razo GSH/GSSG, como se verificou
nos miocrdios de animais no treinados. Adicionalmente, a actividade das enzimas GPX e
GR cardacas foi inferior no grupo treinado
comparativamente com o grupo controlo, embora o contedo de GSH dos coraes treinados tenha sido significativamente superior ao
dos no treinados, quando sujeitos a I-R. Apesar de, no estudo de Demirel e col (89), o treino
de resistncia no ter induzido qualquer alterao na actividade da SOD e da GPX, os autores verificaram que os ratos treinados, comparativamente com os no treinados, eram
menos susceptveis peroxidao lipdica induzida por I-R, quando avaliada pela concentrao de MDA do ventrculo esquerdo. Contudo, embora no tendo determinado o
contedo ventricular de GSH, referem que um
hipottico aumento deste importante polipeptdeo com o treino, confirmado por outras investigaes (12, 14), associado elevao cardaca
das HSP, neste caso das HSP 72 (para refs. ver 71, 78),
poder justificar a diminuio dos nveis de
peroxidao lipdica no miocrdio aps I-R.

CS activity in the trained rats, the low response of O2 consumption in state 3 may be attributable to the possible decrease in the activity of NADH dehydrogenase (complex I)
and/or of other ETC enzyme complexes.
These changes in mitochondrial function
seem very different from those observed by Ji
and Mitchell (91), in which the fall in the respiratory control index was attributed to increased
state 4 respiration and not to reduced state 3
respiration. In this study, the increased O2 consumption in state 4 was interpreted as evidence
of damage to the inner mitochondrial membrane, with a consequent decrease in the proton
gradient, which may suggest different mechanisms associated with dysfunction of cardiac
mitochondrial respiration arising from different
types of exercise (90). The authors concluded that
intense endurance training has a negative effect
on cardiac mitochondrial respiratory function,
making mitochondria more susceptible to oxidative stress, and that this effect may be associated
with lower tissue reserves of GSH.
The effect of endurance training on the
hearts susceptibility to lipid peroxidation induced by perfusion with the oxidizing substance cumene hydroperoxide (CumOOH) was studied by
Kihlstrom (92). The results suggest that training
reduces the susceptibility of rat and mouse
myocardium to lipid peroxidation induced by
hydroperoxides. Another study conducted by
Kihlstrom (93) suggests suggestes GSH as an important mediator in the cardiac protective effect
of training in situations of tissue damage arising
from anoxia/reoxygenation (A/R). In this study,
despite decreases in the activity of enzymes
such as CAT and GR and the absence of change
in cardiac SOD activity with training, increases
in GSH levels and G6PDH activity in the myocardium, together with reduced release of creatine kinase following A/R, would suggest that
endurance training has a protective effect in situations of A/R. This interpretation of the data
was confirmed by the fall in tissue TBARS levels in the trained hearts subjected to A/R
when compared to the control group (93).
CONCLUSION
Despite some disagreement in the literature, regular exercise appears to have a positive influence on cardiac antioxidant systems
and to promote greater tolerance in the myocardium, with a consequent general improvement in its function, both at rest and when
subjected to stimuli that produce additional

Este efeito de tolerncia leso cardaca

induzida por I-R, em animais treinados em resistncia ou sujeitos a uma dieta rica em GSH,
foi igualmente estudado por Ramires e Ji (15).
Os autores concluram que o treino de resistncia, conjuntamente com a administrao de
um suplemento de GSH durante os ltimos 17
dias de treino, pode melhorar a tolerncia do
msculo cardaco leso induzida por I-R.
Esta importante adaptao poder ter resultado
da preservao da homeostasia do metabolismo
da GSH cardaca associada a um incremento
da actividade antioxidante induzida pelo
treino.
Leichtweis e col (90) estudaram o efeito de
um programa de treino de resistncia vigoroso
de natao, com 8-9 semanas de durao, na
funcionalidade mitocondrial de coraes de ratos sujeitos a I-R. Os resultados demonstraram
que o treino promoveu uma diminuio do contedo citoslico e mitocondrial de GSH e de
GSH total (TGSH), embora tivesse incrementado a capacidade do tecido cardaco de importar GSH extracelular pelo aumento da actividade da -GT. Adicionalmente, este estudo
mostrou que os regimes intensos de treino podero induzir diminuio da funo mitocondrial, aps estimulao in vitro, como pode ser
comprovado pela diminuio da taxa de respirao mitocondrial em estado 3 e 4 dos animais treinados comparativamente com os do
grupo controlo. Enquanto a diminuio da respirao em estado 4 pode no ter sido, necessariamente, indicadora de uma disfuno, a reduo dramtica na respirao em estado 3
parece demonstrar claramente que as mitocndrias dos coraes treinados apresentam uma
capacidade reduzida de fosforilao oxidativa (90).
Os autores referem que, uma vez que a produo de equivalentes reduzidos a partir do ciclo
do cido tricarboxlico no sofreu qualquer decrscimo com este tipo de treino vigoroso,
como provam os incrementos na actividade da
CS cardaca dos ratos treinados, a resposta atenuada do consumo de O2 em estado 3 poder
ser atribuda a uma eventual diminuio da actividade da NADH desidrogenase (complexo I)
e/ou de outros complexos enzimticos da CTE.
Estas alteraes na funo mitocondrial parecem ser claramente distintas das encontradas
por Ji e Mitchell (91), nas quais o decrscimo do
ndice de controlo respiratrio foi atribudo ao
aumento da respirao em estado 4 e no a
uma reduo da respirao em estado 3. Neste
estudo (91), o aumento do consumo de O2 em es-

oxidative stress. The GSH system, reduced

ROS production, and the synthesis of other potential protective agents in situations of stress
(such as HSP) appear to be the mechanisms in
which systematic endurance training has the
greatest modulating effect. Nevertheless, some
of the mechanisms associated with the crosstolerance effect of endurance training in relation to different stimuli that cause cardiac toxicity are not yet well understood.
LIST OF ABBREVIATIONS
ROS - Reactive oxygen species
DCFH - Dichlorofluorescin
GSH - Reduced glutathione
NO - Nitric oxide radical
GSSG - Oxidized glutathione
ONOO - Peroxynitrite
TGSH - Total glutathione
I/R - Ischemia/reperfusion
SOD - Superoxide dismutase
Hx - Hypoxanthine
CAT - Catalase
ATP - Adenosine triphosphate
GPX - Glutathione peroxidase
ADP - Adenosine diphosphate
GR - Glutathione reductase
AMP - Adenosine monophosphate
O2 - Molecular oxygen
IMP - Inosine monophosphate
O2- - Superoxide radical
XDH - Xanthine dehydrogenase
H2O2 - Hydrogen peroxide
XO - Xanthine oxidase
OH - Hydroxyl radical
Ca2+ - Calcium ion
C - Carbon
HOCl - Hypochlorous acid
S - Sulfur
GT - -glutamyl transpeptidase
N - Nitrogen
GCS - -glutamylcysteine synthetase
NAD - Nicotinamide adenine dinucleotide
GS - Glutathione synthetase
NADP - Nicotinamide adenine dinucleotide phosphate
HSP - Heat shock proteins
ETC - Electron transport chain
BSO - Buthionine sulfoximine
MDA - Malondialdehyde
mRNA - Messenger RNA
TBARS - Thiobarbituric acid reactive substances

673

tado 4 foi interpretado como uma evidncia de

leso na membrana interna mitocondrial, com
consequente diminuio do gradiente protnico, o que poder sugerir mecanismos distintos associados disfuno da respirao mitocondrial cardaca decorrentes da realizao de
diferentes tipos de exerccio (90). Os autores
concluram que o treino de resistncia intenso
afecta negativamente a funcionalidade respiratria mitocondrial cardaca, tornando as mitocndrias mais susceptveis ao stress oxidativo e
que este efeito poder estar associado a uma
menor reserva tecidual de GSH.
O efeito do treino de resistncia na susceptibilidade cardaca peroxidao lipdica induzida pela perfuso de uma substncia oxidante, o cumnio hidroperxido (CumOOH),
foi estudado por Kihlstrom (92). Os resultados
sugerem que o treino reduz a susceptibilidade
do miocrdio de ratos e ratinhos peroxidao
lipdica induzida por hidroperxidos. Uma investigao igualmente conduzida por Kihlstrom (93)
sugere a GSH como importante mediador na
proteco cardaca induzido pelo treino perante leses teciduais decorrentes de situaes
de anxia-reoxigenao (A-R). Neste estudo,
apesar da diminuio da actividade de enzimas
como a CAT e a GR e da inalterao da actividade da SOD cardacas com o treino, os incrementos do contedo de GSH e da actividade
da G6PDH do miocrdio, acompanhados da reduo da libertao de CK aps A-R, parecem
sugerir que o treino de resistncia tem um
efeito protector perante situaes de A-R. O
teor destes dados foi confirmado pela diminuio dos nveis de TBARS teciduais nos coraes treinados sujeitos a A-R, comparativamente com o respectivo grupo de controlo (93).
CONCLUSO

674

Apesar de algumas discrepncias na literatura, o exerccio regular parece influenciar positivamente os sistemas antioxidantes cardacos
e promover um aumento da tolerncia do miocrdio, com uma consequente melhoria geral
da sua funcionalidade, quer na situao de repouso, quer quando sujeito a estmulos indutores de stress oxidativo adicional. O sistema da
GSH, a eventual diminuio da produo de
ERO e, ainda, a induo da sntese de outros
potenciais agentes protectores perante situaes de stress, de que so exemplo as HSP, parecem ser os mecanismos sobre os quais o
treino de resistncia sistemtico parece ter um

maior efeito modelador. Contudo, alguns dos

mecanismos relacionados com o efeito de tolerncia cruzada do treino de resistncia relativamente aos diferentes estmulos indutores de
toxicidade cardaca no so, ainda, totalmente
conhecidos.
LISTA DE ABREVIATURAS
ERO - Espcies reactivas de oxignio
DCFH - Diclorofluorescina
GSH - Glutationa reduzida
ON - Radical xido ntrico
GSSG - Glutationa oxidada
ONOO- - Peroxinitrito
TGSH - Glutationa total
I-R - Isquemia-reperfuso
SOD - Superxido dismutase
Hx - Hipoxantina
CAT - Catalase
ATP - Adenosina trifosfato
GPX - Glutationa peroxidase
ADP - Adenosina difosfato
GR - Glutationa redutase
AMP - Adenosina monofosfato
O2 - Oxignio molecular
IMP - Inosina monofosfato
O2- - Radical superxido
XDH - Xantina desidrogenase
H2O2 - Perxido de hidrognio
XO - Xantina oxidase
OH - Radical hidrxilo
Ca2+ - Io clcio
C - Carbono
HOCl - cido hipocloroso
S - Enxofre
GT - -glutamil transpeptidase
N - Azoto
GCS - -glutamilcistena sintetase
NAD - Nicotinamida adenina dinucletido
GS - Glutationa sintetase
NADP - Nicotinamida adenina dinucletido fosfato
HSP - Protenas de choque trmico
CTE - Cadeia transportadora de electres
BSO - Butionina sulfoximina
MDA - Malondialdedo
RNAm - cido ribonucleico mensageiro
TBARS - Substncias reactivas ao cido tiobarbitrico

Pedido de separatas para:

Address for reprints:
ANTNIO ASCENSO
Faculdade de Cincias do Desporto e de Educao Fsica
Universidade do Porto
Gabinete de Biologia do Desporto
Rua Dr. Plcido Costa, 91
4200-450 PORTO
Tel: 225 074 700 Fax: 225 500 689
e-mail: aascensao@fcdef.up.pt

675

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XXV Congresso Portugus

de Cardiologia
Organizao:

Direco da Sociedade Portuguesa de Cardiologia

Presidente:

Mrio Freitas
Presidente do Programa Cientfico:

Victor M. Gil
Vilamoura, 27-31 de Maro de 2004
Data limite de envio das comunicaes:
11 de Janeiro de 2004

Secretariado:

Sociedade Portuguesa de Cardiologia

Campo Grande, 28, 13.
1700-093 LISBOA
Tel.: 21 781 7634 e-mail: congresso@mail.telepac.pt
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