Impact of the Training Program on Lipid Profile and Cardiac Health

The aim of this study was to investigate the effects of training programs on serum lipid profile and myocardial oxidative stress. Male Wistar rats (2 mo-old) were divided into three groups (n=8): sedentary (S), loadless trained (T) and trained-overload 2% body weight (TL). T and TL were trained through swimming for 9 weeks. T and TL rats had increased myocardial lipoperoxide (TBA) and lipid hydroperoxide (HP), whereas HP was higher in TL than in T animals. Superoxide dismutase (SOD) activities were lowest in TL. Myocardial glutathione peroxidase (GSH-Px) was lower in TL than in T and S rats. TL decreased HDL-cholesterol and increased LDL-cholesterol. The serum lactate dehydrogenase and TBA were increased, while SOD and GSHPx activities were decreased in TL rats. Loadless training was able to improve HDL-cholesterol and to reduce LDL-cholesterol. In conclusion, the loadless training program induced beneficial effects on lipid profile, while overload training induced dyslipidemic profile that was associated with serum oxidative stress. The overload training program was deleterious relative to loadless training program, increasing myocardial oxidative stress. Key terms: Training, lipid profile, oxidative stress, heart, serum


INTRODUCTION
Exercise has been recommended as a therapeutic measure with a significant role in preventive medicine and cardiac health (15,24,33).However, it is questionable whether all training programs are really safe with respect to cardiac function.Regular aerobic training appears to enhance myocardial performance (30): Isolated exercise sessions may elicit acute transient cardiovascular responses (34), and exhaustive exercise may contribute to alterations in heart processes, resulting in further injury (17,19).
Reactive oxygen species (ROS) and free radicals are mediators of several forms of tissue damage, such as ischemic injuries (1) and cardiac damage (7,8,9,12,29).Antioxidant defense cell systems protect against or minimize oxidative damage induced by ROS, but there is conflicting information on the effects of exercise and training in these antioxidant defenses (17,18,19,28,35).
Although usually ascribed to dyslipidemic profile and atheroschlerosis (1), myocardial dysfunction has been reported in the absence of increased serum lipids (5,10,20).Because there is a relationship between oxidative stress and myocardial disease with or without dyslipidemic profile, studies on the effects of different types of training programs on lipid profile and myocardial oxidative stress are of particular interest.
Thus, the aim of this study was to investigate the effects of the training program on lipid profile and myocardial oxidative stress.

MATERIALS AND METHODS
The Ethical Committee for Conduction of Animal Studies at the Faculty of Medicine, University of São Paulo State (UNESP), approved the experimental protocol and all animals were cared for in accordance with the principles and guidelines of the Canadian Council on Animal Care, as outlined in the "Guide to the Care and Use of Experimental Animals."Male Wistar rats (24 animals, 2 months old) were fed a Purina rodent chow (3074 SIF, Purina Ltd., Campinas, S.P., Brazil) and water ad libitum.They were housed in groups of four animals, in polypropylene cages in an environmentally-controlled clean-air room, with a temperature of 22 ± 3ºC, 12 h light/ dark cycle and a relative humidity of 60 ± 5%.The rats were randomly divided into three groups (n-8): sedentary-control group (S), loadless trained (T), and trained overload 2% body weight (TL).Swimming was performed in two 100 cm high plastic containers, and the water temperature was 32 ± 1 o C.
Swimming was selected as a model for exercise performance because it appears to belong to the natural behavior of rodents and humans.It is less stressful and can avoid foot injury as in the case of a treadmill, which may generate ROS unrelated to exercise (28).The trained groups swam in the morning, four animals each group, five days/week, for nine weeks in progressive depths of 10-50 cm during the first week.The training time was increased by 15 min each week (35).At the fourth week the trained animals swam for 60 min/day and were divided into T and TL groups until the end of the study.The load was attached to the thorax and updated once a week.Loading 2% body weight was used to increase exercise intensity (28).At the end of the 9-week experimental period, all animals were sacrificed by decapitation (35).The sedentary-control group was sacrificed at rest at the same time as the others animals.
The heart was removed, and the cardiac adipose tissue was discarded.The heart was weighed, and the left ventricle (LV) was separated and weighed.In this experiment the biochemical determinations were carried out on the LV (myocytes) free of external adipose tissue.Samples of 200 mg of LV were homogenized in 5 ml of a cold phosphate buffer (0.1M, pH 7.4).Tissue homogenates were prepared in a motordriven Teflon glass Potter Elvehjem, tissue homogenizer (1 min, 100 rpm) immersed in iced water.The homogenate was centrifuged at 10,000 rpm for 15 min.The supernatant was used for protein, lipoperoxide (TBA), lipid hydroperoxide (HP), glutathione peroxidase (GSH-Px, E.C.1.11.1.9.) and superoxide dismutase (SOD, E.C.1.15.1.1.)determinations.
Results are presented as means ± standard deviation (SD).Significance of difference was tested by analysis of variance (ANOVA) and Tukey test to compare the treatment groups with each other, not just the control.The probability of 0.05 was chosen as the significant level (25).

RESULTS
TL decreased the body weight gain and the final body weight (bw), but no significant alterations were observed in heart weight (hw) and hw/bw ratio of TL in relation to S and T animals (Table I).No significant changes were observed in body weight gain, final body weight, hw and hw/bw ratio of T group from S. Table II shows that training induced a significant increase in TBA and HP myocardial levels, whereas    I).Serum LDH activities were significantly increased in TL animals, indicating cellular damage and reflecting the stress on endothelial cells (Table III).
In several organs, cell membrane damage is followed by the release of a number of cytoplasmic enzymes to the blood, a phenomenon that provides the basis for clinical diagnosis (14).Enzymes are liberated from tissues as soon as a physical-chemical alteration has occurred.Since LDH activity is used for myocardial disease diagnosis, we can assume that the training program of TL group was sufficiently elevated to induce myocardial alterations, suggesting that the animals of this group were over-trained.
In recent years, a considerable interest has been shown in the deleterious action of oxygen radicals on the heart (7,8,9,12,29).This concept may be regarded as a paradox because myocardial function is strictly dependent upon oxygen delivery and use.However, in some situations, oxygen is converted to ROS, which can elicit widespread damage to cells such as lipoperoxidation (22).In this study, two indices of lipoperoxidation were measured.This involves the abstraction of hydrogen from fatty acid side chains by ROS, forming conjugated dienes that oxidize to lipid hydroperoxide (HP), and subsequently undergo cleavage to aldehydes, some of which react with TBA (1,11).TBA and HP concentrations were significantly increased in cardiac tissue of both trained groups.It is evident that both training programs had a higher workload than normal, which leads to an imbalance of the oxidant/antioxidant systems.These observations were demonstrated with antioxidant enzymes (Table II).The SOD activity was significantly decreased in response to the training (T group).Nevertheless, within the same group no changes were observed in GSH-Px activities.It is interesting to note that when rats trained with load (TL) they showed the lowest SOD and GSH-Px activities.SOD removes superoxide radical by reducing them to hydrogen peroxide.GSH-Px catalyses the conversion of hydrogen peroxide to water (1).Therefore, our results support the idea that the alterations in antioxidant enzymes were not sufficient to inhibit the oxidative stress and the lipoperoxidation in cardiac tissue of both trained rats (T and TL).
On the other hand, Table II shows similar TBA values in T and TL animals, while HP concentration was markedly lower in T than in the TL group.These observations indicated that over-training, as occurred in the TL group, accelerated the breakdown of HP and increased aldehyde production.These observations are substantially important in explaining the adverse alterations on serum lipids (1,4).Lipid hydroperoxides give rise to a large variety of products, many of which react with TBA and with serum lipids (1).Oxidation of LDL from hydroperoxide products may have substantial importance on dyslipidemic profile.Zadeh et al., (36) showed that LDLcholesterol is the major carrier of lipid hydroperoxide in plasma.Oxidative modification of LDL is the key step in the sequence of events leading to atherosclerosis (4).Since serum TBA and LDL-cholesterol concentrations were increased in TL animals, we can assume that increased lipoperoxidation was associated to LDLcholesterol.The HDL-cholesterol is extensively degraded due oxidative processes (3).The reduced SOD and GSH-Px activities in the serum were the result of load training.While no alterations were observed in HP serum concentrations, the increased TBA in the serum of TL animals indicated that they were largely affected by training intensity and suggested that substances that react with thiobarbituric acid were liberated to the blood from the tissues.These observations explained why the T rats had decreased myocardial HP levels and no differences were observed in myocardial TBA levels between T and TL animals.
The most significant risk indicators for cardiovascular alterations, which are considered to be parameters of oxidative stress (1), are increased serum cholesterol, triacylglycerol, LDL-cholesterol (36) and decreased HDL-cholesterol (31).Various theories have suggested that cardiovascular damage was the result of an oxidative stress process (31,36).Loadless training had beneficial effects on serum lipids (Table III).The beneficial effects of loadless training were demonstrated in relation to LDL and HDL-cholesterol.The T animals showed decreased LDL and increased HDLcholesterol.These effects were not observed when the training was made with load.Note that the TL animals showed decreased HDL and increased LDLcholesterol (Table III).
In conclusion, the effects of exercise on lipid profile and markers of oxidative stress were dependent on the training program performed.Loadless training programs induced beneficial effects on the serum lipid profile, while load training induced dyslipidemic profiles that were associated with serum oxidative stress.The training program with overload 2% body weight was deleterious relative to loadless training program, increasing myocardial oxidative stress.