Acute and Chronic Effects of Endurance Running on Inflammatory Markers: A Systematic Review

In order to understand the effect of endurance running on inflammation, it is necessary to quantify the extent to which acute and chronic running affects inflammatory mediators. The aim of this study was to summarize the literature on the effects of endurance running on inflammation mediators. Electronic searches were conducted on PubMED and Science Direct with no limits of date and language of publication. Randomized controlled trials (RCTs) and non-randomized controlled trials (NRCTs) investigating the acute and chronic effects of running on inflammation markers in runners were reviewed by two researchers for eligibility. The modified Downs and Black checklist for the assesssments of the methodological quality of studies was subsequently used. Fifty-one studies were finally included. There were no studies with elite athletes. Only two studies were chronic interventions. Results revealed that acute and chronic endurance running may affect anti- and pro-inflammatory markers but methodological differences between studies do not allow comparisons or generalization of the results. The information provided in this systematic review would help practitioners for better designing further studies while providing reference values for a better understanding of inflammatory responses after different running events. Further longitudinal studies are needed to identify the influence of training load parameters on inflammatory markers in runners of different levels and training background.

In order to understand the effect of endurance running on inflammation, it is necessary to quantify the extent to which acute and chronic running affects inflammatory mediators. The aim of this study was to summarize the literature on the effects of endurance running on inflammation mediators. Electronic searches were conducted on PubMED and Science Direct with no limits of date and language of publication. Randomized controlled trials (RCTs) and non-randomized controlled trials (NRCTs) investigating the acute and chronic effects of running on inflammation markers in runners were reviewed by two researchers for eligibility. The modified Downs and Black checklist for the assesssments of the methodological quality of studies was subsequently used. Fifty-one studies were finally included. There were no studies with elite athletes. Only two studies were chronic interventions. Results revealed that acute and chronic endurance running may affect anti-and pro-inflammatory markers but methodological differences between studies do not allow comparisons or generalization of the results. The information provided in this systematic review would help practitioners for better designing further studies while providing reference values for a better understanding of inflammatory responses after different running events. Further longitudinal studies are needed to identify the influence of training load parameters on inflammatory markers in runners of different levels and training background.

INTRODUCTION
Running is an important natural ability of our species that has contributed to our survival and body adaptations (Bramble and Lieberman, 2004). In the Paleolithic Era, survival was dependent on hunting and gathering, and therefore it has been suggested that the ancient physical activity pattern included mostly prolonged, low-intensity physical activities, including endurance running, interspersed with high-intensity bursts of activity (O'Keefe et al., 2010;Boullosa et al., 2013). Nowadays, endurance running is probably the most popular sport worlwide and it is practiced for recreational, health and competitive purposes (Chiampas and Goyal, 2015).
There is a close link between endurance running and the activity of the immune system. The importance of this relationship has led to important investigations over the last decades. Previously, Peters and Bateman (1983) identified an increased prevalence of upper respiratory tract infection (URTI) in 150 runners following a 56.0 km ultramarathon. Subsequently, specialized literature has suggested that even highly trained individuals, when subjected to frequent strenuous exercise, could develop a proinflammatory condition that favors the onset of a number of health problems, including damage to myocardial cells and connective tissues, overload of the atria and right ventricle, coronary artery disease (CAD), and coronary artery calcification among others (Peters and Bateman, 1983;Febbraio and Pedersen, 2005;Petersen and Pedersen, 2005;Zaldivar et al., 2006;Mohlenkamp et al., 2007;Hubble et al., 2009;Nieman, 2009;Meeusen et al., 2013;O'Keefe and Lavie, 2013;Taylor et al., 2014). However, little is known about whether the development of these chronic pathologies is the result of an excess of training volume, intensity, or both, associated with an insufficient recovery, which often promotes an increased susceptibility to infections and subsequent reduction in performance (Smith, 2004;Zaldivar et al., 2006;Hubble et al., 2009). Furthermore, systematic and non-systematic inflammation after running might be related with functional overreaching (Steinacker et al., 2004). In contrast, it has been suggested that periodised training with adequate recovery may be associated with positive adaptations including an adequate balance between pro-inflammatory and anti-inflammatory responses (Febbraio and Pedersen, 2005;Petersen and Pedersen, 2005;Zaldivar et al., 2006).
A growing body of evidence highlights the importance of studying inflammation promoted by endurance running as a factor which is linked to the physiopathology of a number of cardiovascular diseases (Mohlenkamp et al., 2007). It has also been suggested a link between myocardial damage and small thrombotic or even atherosclerotic emboli following a marathon, or after a quick session of exercise, accompanied by a transient monocytosis (about 2 h) (Walsh et al., 2011). The tissue factor is known as the key initiator of coagulation, and is highly dependent on vascular injury and mediators of inflammation such as tumor necrosis factor alpha (TNF-α), which has been reported to increase during and predominantly after marathon running (O'Brien, 2012;Gill et al., 2015b).
Contrary to other endurance sports, eccentric muscle contractions play a key role in running exercises, leading up to the occurrence of different levels of damage in muscle, connective and bone tissues (Suzuki et al., 2003;Jarvinen et al., 2013). The repair of these tissues involves the presence of inflammatory cells into the damaged site, which stimulates the release of proinflammatory cytokines such as TNF-α and interleukin-1 beta (IL-1β), thus triggering inflammation (Nieman et al., 1989(Nieman et al., , 1990. However, little is known about the impact of this chronic cycle tissue damage and repair in runners. On the other hand, it is also important to emphasize that signaling promoted by repeated muscle contractions as in running, stimulates the production of anti-inflammatory mediators by myocytes, especially interleukin-6 (IL-6), which acts as an inhibitor of pro-inflammatory cytokines such as TNF-α by stimulating the production of its soluble receptor antagonists (Pedersen, 2013). In addition, IL-6 also stimulates the production of interleukin-10 (IL-10) and interleukin-1 receptor antagonist (IL-1ra), generating an anti-inflammatory environment which may counterbalance the pro-inflammatory responses associated to repetitive eccentric actions (Pedersen and Febbraio, 2012).
Despite the growing body of evidence regarding the effects of endurance running on inflammation, the link between transient acute responses and chronic adaptations needs to be addressed (Gleeson, 2007). This information would be important to shed light on the possible role of the inflammatory milieu in the pathophysiology of a number of diseases, especially the cardiovascular ones. Thus, the aim of this systematic review was to investigate the effects of different doses (i.e., training and competitive loads) of endurance running on the acute and chronic inflammatory responses, and the immune effects of this practice on runners of different levels and training backgrounds.
Once the abstracts were reviewed, the complete versions of the papers that met the criteria were obtained. In addition, the reference lists of the papers that fulfilled the inclusion criteria were analyzed for identification of additional studies. The exclusion of studies with irrelevant content and duplicates was carried out after the title, abstract and full-text were read.

Definition of Terms
An "athlete" was defined according to the Medical Subject Headings (MeSH) and was considered to be an individual who has developed skills, physical fitness and strength, or who has participated in sports running (MeSH, 2015). We have considered the definition proposed by Stirling and Kerr (2006) that defines a "recreational athlete" as being an individual who plays on a sports team at an amateur level, works out 1-4 times a week, does not train and compete nationally or internationally, and does not train for the same activity for more than 8 h per week. Novice runners were those individuals who had not been running on a regular basis in the previous 12 months (10 km total in all training sessions in the previous 12 months), and recreational runners were considered as individuals running a mean of 24.94 km/week (Videbaek et al., 2015).
The following thesaurus terms registered in the database from MeSH were also used: "running, " "aerobic exercise, " "inflammation, " and "cytokines." These terms were associated with the free terms "recreational runners, " "novice runners, " "marathon runners, " and "ultramarathon." Science Direct (Elsevier) 1. "marathon runners" OR "novice runners" AND "cytokines"; 94 4 Total: 94 Other sources (reference lists of the papers that fulfilled the inclusion criteria were analyzed for the identification of additional studies) Total: 19 6 *Truncation or wildcard.

Inclusion and Exclusion Criteria
The inclusion criteria were as follows: randomized controlled trials (RCTs) and non-randomized controlled trials (NRCTs); studies investigating the acute and chronic effects of running on markers of inflammation in marathon runners, recreational runners and novice runners; the terms runners, marathon runners, recreational runners and novice runners should be cited in the paper; only healthy participants; only full-text article citations with no restriction on languages; with individuals aged over 19 as the World Health Organisation (WHO) defines adolescence as the period in human growth and development that occurs between childhood and adulthood, from ages 10-19 (WHO, 2015) 1 . Meeting abstracts, unpublished data, observational studies, review articles, studies using walking and jogging as independent variables, and studies on the effects of any kind of supplements on running, diet restrictions, use of devices (e.g., equipment, compression garments), comparisons between running and other sports, and effects of environmental conditions (ex. dry and hot) were excluded.

Quality Assessment
The quality and assessment of all eligible articles was evaluated using a modified version of the Downs and Black checklist (Downs and Black, 1998). Disagreements between authors were discussed and subsequently solved. This modified version consists of 27 objective questions (Downs and Black, 1998

Research Strategy
Results of the research strategy are presented in Figure 1. Initially, 60 studies were selected, with 51 studies being finally included according to the inclusion/exclusion criteria. Nine studies were excluded as follows: one study was excluded due to the use of heat stress, three because the subjects were adolescents, one following the reading of the full paper, two because of comparison with other sporting activities, and one because of medication use; in addition, one paper was not available in fulltext version (Saravia et al., 2010). A total of 49 studies verified the acute effect of running on inflammation and two studies focused on the chronic effects.

Methodological Quality Assessment
Quality assessment of the studies according to the modified Downs and Black scale is summarized in Table 2. One important finding was that characteristics of the patients' included were not cleary described in 32 studies. Important adverse events and description of patients' characteristics lost to follow-up was not reported in 34 and 32 studies, respectively. None of the studies were randomized controlled trials and power was provided in 4 studies ( Table 2).

Characteristics of the Studies and Summary of Outcome Measures
An overview of the studies' characteristics is provided in Table 3 with sample size, age, sex, and exercise protocols. A summary of outcome measures in selected studies is presented on Table 4.
The 51 studies included resulted in a total of 1,421 subjects, of whom 163 were female and 1,234 males; 24 subjects were not identified by sex in one study (Neidhart et al., 2000). All trials provided age ranges for the subjects and the mean age was 39.16 years.
The common protocols adopted included marathon in 17 studies, ultra-marathon in 22, half-marathon in three studies, different distance protocols (42.195, 21.1, 12, 10 km and treadmill) in seven studies, and chronic training only in two studies (see Table 3). For other questions 1 = yes, 0 = no, and 0* = unable to determine.

DISCUSSION
The aim of this systematic review was to analyze studies that verified the effects of different endurance running exercises on acute and chronic inflammatory responses in runners of different training background. The present systematic review allows an initial understanding of this issue. It seems that acute and chronic endurance running may affect anti-and proinflammatory markers. However, important differences between studies in terms of methods as well as in runners' charactersitics do not allow appropriate comparison or generalization of results.

Inflammatory Markers
The timing in data collection could be considered an important limiting factor for adequate comparisons, given the heterogeneity observed across the reported acute (i.e., 5-20 min) (Fallon et al., 2001;Kim et al., 2007;Vaisberg et al., 2013;Grabs et al., 2015) and delayed responses (24 h to 8 days) (Uchakin et al., 2003;Siegel et al., 2007;Hewing et al., 2015). This is not a simple issue given the different kinetics and biological availability of the molecules considered as IL-6 and CRP (Kasprowicz et al., 2013;Reihmane et al., 2013). Of note, some inflammatory markers (e.g., monocyte chemoattractant protein-1, granulocyte colonystimulating factor) (Suzuki et al., 2003) have not been included in the current review. Further studies should elaborate appropriate study designs that consider both the appropriateness of the inflammatory markers selected as well as their kinetics.
Another important limitation for generalization of the results refers to the heterogeneity of running exercises (e.g., distance, intensity) used for evaluation of acute inflammatory responses. Furthermore, ambient characteristics (e.g., altitude, temperature) and race profile (e.g., uphill and downhill running) which have been suggested to influence muscle contraction and physiological responses (Vernillo et al., 2017), have not been always reported (Millet et al., 2011;Saugy et al., 2013). These aspects should be controlled in further studies for isolating the relative effect of every specific factor on inflammation.

Body Fatness and Inflammation
Given the relationship between adipose tissue and inflammation (Pedersen and Febbraio, 2012), attention should be paid to overweight and obese runners. For instance, a higher level of CRP 24 h following a marathon has been observed in obese nonelite runners when compared to lean elite runners (Nickel et al., 2012). Furthermore, obese non-elite runners when compared to lean elite and lean non-elite runners demonstrated a higher level of IL-6 and a lower level of IL-10 serum levels at baseline (Nickel et al., 2012). However, all groups presented an increase for serum IL-10 and TNF-α, and a decrease for serum IL-6 levels, immediately post-marathon (Nickel et al., 2012). It must be considered that increments in IL-10 induced by exercise may be responsible for the elevation in IL1-ra which exerts an antiinflammatory action by antagonizing IL-1 and IL-1β (Dinarello, 2000;Moldoveanu et al., 2001;Petersen and Pedersen, 2005;Pedersen, 2011). Nevertheless, it could be suggested that, in overweighted individuals, a higher pro-inflammatory status at baseline and post-marathon could be observed, with unknow consequences for health in the long term.

Inflammation and Cardiovascular Health
One relevant issue refers to the link between inflammation and cardiovascular health. Interestingly, the exercise-induced increase of IL-6 after the marathon in 20 lean male runners was associated with a lower prevalence of arrhythmias during and after the marathon race (Grabs et al., 2015). When produced by muscle contraction, IL-6 stimulates the synthesis of other anti-inflammatory cytokines such as IL-1ra and IL-10, thus providing an inhibitory effect on pro-inflammatory cytokines such as IL-1β and TNF-α (Pedersen and Febbraio, 2012;Pedersen, 2013). However, CRP, a strong predictor of cardiovascular events, is an acute phase protein synthesized in the liver by the stimulation of IL-6 (Ridker et al., 2002). Chronic endurance training may decrease CRP values, especially when accompanied by a loss in fat mass, therefore promoting further reduction of risk for cardiovascular events (Fallon et al., 2001;Tomaszewski et al., 2003;Walsh et al., 2011;Grabs et al., 2015;Kim et al., 2015). Of note, CRP may be more susceptible to chronically decrease in individuals presenting higher baseline levels (Barnett et al., 2005). Therefore, caution should be taken when evaluating the anti-and pro-inflammatory effects of running in individuals with different characteristics regarding cardiovascular risk factors (e.g., body composition) (Moldoveanu et al., 2000;Petersen and Pedersen, 2005;Walsh et al., 2011).

Studies' Characteristics
Most studies included in this systematic review were acute interventions (49 studies). However acute changes in inflammatory markers might not be related with anti-and pro-inflammatory outcomes during chronic aerobic training interventions. For instance, there were divergent responses for CRP changes in chronic interventions (Mattusch et al., 2000;Auersperger et al., 2012). Thus, while Mattusch et al. (2000) observed a reduction in CRP levels, Auersperger et al. (2012) did not observe any change. Therefore, further studies must consider this important limitation, while providing training load charactersitics as volume, intensity, and frequency of training sessions. An important question to be answered refers to the minimal training load required for runners of different levels when preparing different competitive distances, while analyzing the impact of these factors on inflammatory markers. Additionally, there is a prevalence of male runners on literature therefore more studies with female runners are needed.

CONCLUSION
In summary, our results revealed that acute and chronic endurance running may affect anti-and pro-inflammatory markes but methodological differences between studies do not allow comparisons or generalization of the results. Only two studies were chronic interventions. There are no studies with elite athletes. Thus, RCTs are urgently needed to identify the appropriate dose of endurance running (volume, intensity, and frequency) required to elicit improvements in inflammatory markers in runners of different levels and training background. External (e.g., ambient characteristics, race profile) and internal factors (e.g., fitness level, training experience) to runners should be considered in further studies for a better understanding of the relationship between running and the mediators of inflammation. The information provided in this systematic review would help practitioners for better designing further studies while providing reference values for a better understanding of inflammatory responses after different running events.