Physical activity is associated with reduced serum concentration of inflammatory markers. Previous studies have shown that the type and the degree of exercises increases oxidative stress, which, in turn causes the generation of reactive oxygen species (ROS), that influences inflammatory markers including C-reactive protein (CRP) (Kasapis and Thompson, 2005; Majka et al., 2009) and genetic factors (Shen and Ordovas, 2009). Athletes have also been reported to have better endothelial function than sedentary controls (Franzoni et al., 2004), although, prolonged, brisk exercise is also reported to transiently raise markers of endothelial activation, such as ICAM-1 and E-selectin (Bartzeliotou et al., 2007). In the light of these studies we have hypothesed that intensive training alters the markers of inflammation, endothelial functions and autoimmunity in adolescent females.
Reactive oxygen species (ROS) are generated during exercise from several sources including leakage from the electron transport chain. These ROS may cause cellular damage that may subsequently lead to lipid oxidation, protein denaturation or DNA damage (Droge, 2002). However, exercise is also reported to enhance the endogenous antioxidant defences (Rousseau et al., 2006). For example, repeated episodes of aerobic exercise has been reported to induce the expression of antioxidant enzymes (Ji, 2002), and several studies have shown a relationship between extent of physical activity and plasma antioxidant concentrations (Dekany et al., 2006). Furthermore Pialoux and his colleagues have reported that the intensity of exercise and exposure to hypoxia may have a cumulative effect on oxidative stress (Pialoux et al., 2006) and that though antioxidant defences are enhanced in elite athletes this does not allow them sufficient buffering to counterbalance the over-production of free radicals during exercise (Pialoux et al., 2009). The potential damaging effects of reactive oxygen species can be partially reversed by molecular chaperones, such as heat shock protein 27 (Hsp27) that may be involved in a process of protein renaturation (Ferns et al., 2006). However, during this process the chaperone molecules appear to be altered in a way that may render them antigenic (Wick et al., 2004).
The principle aim of this current study was to determine weather intensive exercise alters markers of inflammation, endothelial activation and autoimmunity by comparing adolescent female gymnasts, engaged in high physical activity with sedentary girls, matched for associations between age and associations between them. Using this young age group of subjects removed the potential confounding effects of existing vascular disease. The term intensive training/exercise refers to a physical activity related to the average number of hours per day of gymnastic training which incorporates impact exercise including evaluating jumping, tumbling and strength work. The specific objectives were to assess their serum levels of CRP, sICAM-1 and Hsp27 antigen and antibodies, and to determine their association with age, adiposity and dietary intakes of macro- and micro-nutrients.
Subjects were recruited as part of a three-year longitudenal investigation of the effects of exercise on peak bone mass (PBM) development as previously reported (Nurmi-Lawton et al., 2004). Twenty-five female gymnasts, 8-17 years of age were originally recruited from five athletics clubs in the South of England. The gymnasts were eligible to join the study if they trained > 10 h/week and regularly took part in competitions (at club regional level). Nineteen female healthy normally active controls were randomly recruited through the database of local General Practices (GPs) in Surrey, South England. Controls were involved in normal activities (including walking to school and PE classes) for on average 5.6 [+ or -] 2.6 h/week (determined by a checklist), but not in sports requiring all year training at competition level. In addition, anthropometric, physical activity and dietary intake were estimated as previously described (Nurmi-Lawton et al., 2004). Anthropometric data were determined by a nutritionist. None of the subjects had evidence of acute infection, or inflammation at the time of recruitment, or at the time of blood sample collection. Each gymnast was matched to a non-active control, initially by age and then subsequently by pubertal age once Tanner staging had been completed and analysed (Nurmi-Lawton et al 2004). The baseline data were collected between the months of October and December of the year for both gymnasts and controls. We were not able to collect information on the timing of the menstruation phase cycle during the collection of blood samples.
Ethical approval was obtained from the South-West Surrey local research ethics committee, Guildford, Surrey, and from University of Surrey ethics committee.
The dietary intake of gymnasts and controls was recorded for 7 days at baseline using estimated food diaries, which have been shown to have an acceptable relative validity (Bingham et al., 1994). Instructions, including how to estimate portion size, were given both verbally and in writing to each subject, or their parents by a registered dietician. Gymnasts were asked to complete the diary on a non-competition week, during the athletic training season and controls on a non-holiday week, without changing their usual dietary habits. The diaries were analyzed using Diet5 for Windows computer package (Robert Gordon University, Aberdeen, UK), which is based on McCance and Widdowson Food Composition Tables (Mark and Paul, 1992). This allowed an estimation of dietary macronutrient (protein, carbohydrate and fat) and micronutrients (including fat and water soluble antioxidants)
Assessment of menarche
Pubertal status was assessed at each measurement occasion, using self-assessment of secondary, sexual characteristics. Staging of secondary, sex characteristics according to Tanner TM (Tanner, 1962) has provided a means of assessing sexual development, but requires the individual to undress and then be physically examined. It is very difficult to collect such personal data in non-medical settings. Self assessment of the level of sexual maturation by adolescents has been found to be both valid and reliable, and in excellent agreement with physical examination (Morris and Udry, 1980), enabling accurate assessment of an individual's own developmental stage according to standard photographs by Tanner (Marshall and Tanner, 1969). In the present study, the use of adolescent self-staging was deemed most appropriate.
Physical activity assessment
The hours of training, changes in training regimen of each gymnast, and the type and amount of exercise undertaken by controls, were assessed using a questionnaire. The Blair score was calculated as described previously (Blair et al., 1985), which gives an indication of the individual's level of physical activity:
Non-fasted blood samples were collected from both the controls and gymnasts. Due to the recruitment procedure required for the gymnasts, that is, via gymnastic clubs that only met late afternoon/early evening, blood samples after a 12 hour fast were not possible. Control subjects predominantly attended the University of Surrey Clinical Investigation Unit (CIU) in the afternoon/early evening. Collections of samples for the gymnast group were carried out before training and out of competition week. Collections of samples for the controls were carried out during the school holiday. In all cases, they were collected around the time that the dietary estimates were obtained. Blood samples were centrifuged at 3000g at room temperature to obtain serum for analysis of markers of inflammation, endothelial activation, autoimmunity, antioxidants and trace elements. Serum samples were frozen at -80 [degrees]C until they were assayed.
Determination of ICAM-1 and hsCRP by ELISAs
Serum soluble intercellular adhesion molecule 1 (sICAM-1) and high sensitivity C-reactive protein (hsCRP) concentrations were measured in duplicate in all subjects using enzyme-linked immunosorbent assay (ELISA) kit purchased from [Quantikine (R & D Systems Europe, Ltd. U.K)], or an in-house ELISA respectively. hsCRP in-house ELISA: The...