The results of this study demonstrate that prolonged performance of repetitive/cyclic movement triggers significantly elevated pro-inflammatory cytokines expression in the associated ligaments regardless of the load magnitude. Furthermore, repetitive exposure to high loads triggers significantly higher expression in several (IL-6, IL-8 and TGFβ1), but not all, cytokines than exposure to low loads. Elevated cytokines expression indicates that significant tissue degradation, consistent with an acute inflammation, occurred post-loading. Low cyclic loads, therefore, should not be simply dismissed as a no-risk activity.
Cytokines are soluble proteins that respond to ligament injury by binding to their receptors and initiating a cellular response [35
]. Interleukin-1 (IL-1) is a strong pro-inflammatory cytokine that works both directly and indirectly. IL-1 induces B-cell differentiation and acute phase proteins [8
]. It also drives extracellular matrix destruction by increasing degradative enzymes such as matrix metalloproteinases [44
]. In addition, IL-1 stimulates fibroblasts to express and secrete other inflammatory cytokines including IL-6 [9
]. The action of IL-6 is typically pro-inflammatory during the acute phase by inducing the production of acute phase proteins (some distinct from those produced by IL-1 and TNFα) and can also promote B- and T-cell functions and stimulate the secretion of immunoglobulin [9
]. IL-8 is considered a chemokine because it attracts neutrophils to the site of inflammation [23
], for example, in inflammatory joint disease [46
]. The action of TNFα is similar to that of IL-1, but is thought to be less potent in most tissues. The remaining cytokine measured in this study, TGFβ, is a multifunctional cytokine that acts in normal physiology and pathology. It may act in potentially contradictory ways depending on tissue and pathology [21
]. In inflammation, TGFβ can recruit B and T cells, but can also lower the immune response and promote extracellular matrix production.
Anterior flexion executed to a pre-determined angle should manifest with an identical strain (elongation %) regardless of the load applied. Why, then, a significantly higher pro-inflammatory cytokines expression is exhibited following flexion under high loads? It is well established that viscoelastic tissues (such as ligaments, facet capsules, dorsolumbar fascia, etc., in this case) are highly sensitive to the load application rate. In the group subjected to low loads, the peak load of 20 N was reached within 2 s. The load application rate, in this case, was 10 N/s. Conversely, in the group subjected to high loads, the peak load of 60 N was also reached within 2 s. Thus, the load application rate was 30 N/s, or three times (3×) the rate of the low-load group. Viscoelastic tissues can adapt to slow application of load with larger strain and minimal development of micro-fractures within its collagen fibers. Fast application of loads, however, results in the inability of the tissue to strain fast enough to accommodate the load rate, development of high tension [28
] and substantial development of micro-ruptures within its collagen fibers [7
]. Overall, high rates of loading result in substantially larger damage within the collagen fibers and the expected larger inflammatory response.
Indeed, the creep recorded at the end of the first 10 min of cyclic loading was 48.9% for the low-load group, but was 34.4% for the high-load group, demonstrating the ability of the tissue to strain more in the low-load group and the inability to strain fast enough in the high-load group. Similarly, at the end of the six loading/rest periods, the final creep in the low-load group was 85.9%, but only 62.1% in the high-load group.
The final, or residual, creep at the end of the 7-h rest period is most illuminating. It was 24.8% for the low-load group, but a much higher 30.2% in the high-load group. Similar results were obtained throughout our long investigation of cyclic loading [2
], and the residual creep at the end of 7-h recovery was always higher in the high-load condition. Finally, our study assessing the direct impact of increased frequency of the cyclic load at the same load [18
] confirm the increased impact of frequency on the elicited neuromuscular disorder, leaving the assessment of the change in cytokines expression to the next study on our agenda. Preliminary data confirm that significant increase in cytokines is observed at high-frequency loading.
During the cyclic loading periods, the lumbar viscoelastic tissues of the group subjected to high load continuously incurs micro-damage to more and more collagen fibers. At the end of the loading periods, large numbers of fibers are damaged and are not functional. Therefore, at the end of the rest period, the viscoelastic tissues exhibit the recovery of the remaining undamaged, functional collagen fibers together with the laxity induced by the dysfunctional, damaged fibers. The overall higher residual creep, therefore, represents the larger damage inflicted by the high-load rate. Since significantly higher pro-inflammatory cytokines expression elicits inflammatory tissue degradation, more pronounced degradation is expected to be observed in the group exposed to higher load.
Although a significant increase in cytokines expression was observed in the group subjected to low loads, the epidemiology, biomechanics and neuromuscular findings do not support the existence of a repetitive disorder. The epidemiology does not describe a pronounced increase in reports of disability associated with repetitive activity at low loads [10
]. The biomechanical and neurophysiological evidence also show minimal spasms during loading, absence of muscular spasms/hyperexcitability post-loading and fair recovery of lumbar laxity [11
]. With the lack of supporting evidence for a disorder, one is left to conclude that the cytokines expression level recorded in the low-load group did not exceed a certain physiological threshold required to trigger a disorder directly or indirectly.
Significantly greater expression of IL-6, IL-8 and TGFβ1 were observed in the ligaments harvested from the group exposed to high load relative to the group exposed to low loads. The remaining cytokines, IL-1β and TNFα, were not significantly different in the high-load group. The specific cytokines that registered elevated expression in the high-load group may lead us to gain insight into the threshold discussed above. Indeed, a recent report points out that upregulation of IL-6 and IL-8 are each positively correlated with pain intensity in humans with an acute inflammation [47
]. The authors went onto conclude that increased IL-6 and IL-8 expression contributed to the development of acute inflammation and inflammatory pain. Considering their conclusion in the context of our data, the significantly lower levels of these two cytokines in the low-load group are probably below the threshold of triggering an acute inflammation and the associated pain. Another possibility is that a mild acute inflammation was present in the low-load group, but the pain level was not sufficient to trigger a neuromuscular disorder. A third possibility is that regardless of the presence or level of the acute inflammation, the pain level may be the trigger of the neuromuscular disorder. Unfortunately, we could not assess pain in anesthetized animals. It is obvious that more research is required to reveal the details of this important and fascinating issue.
The significant increase in the expression of all five pro-inflammatory cytokines tested in the ligaments from the group exposed to low loads indicates that repetitive activity under such conditions cannot be designated as “no risk.” Possibly, with higher number of repetitions [24
] and with less intermittent rest [11
], low load can become problematic as the amount of damage to the collagen fibers may exceed a certain threshold. It may be more appropriate to designate such category as “low risk”.
Inflammation of damaged viscoelastic tissue such as tendons and ligament is biphasic. Once the damage in the tissue is detected, pro-inflammatory substances and cells invade the affected tissue and mediate the removal of the damaged molecules, passing them to the circulatory system [14
]. Once this first phase is complete, a second phase is initiated during which a reconstruction of the damaged collagen matrix is performed. Therefore, a minor acute inflammation due to repetitive physical activity can be spontaneously resolved if sufficient rest is allowed. Often, 24–72 h are sufficient to spontaneously resolve most repetitive exercise-related damage [14
]. Indeed, a model based on animal work [37
] shows that creep/laxity in ligaments together with a neuromuscular disorder will not fully recover within 7–8 h of rest post-cyclic exercise, as it leaves a large residual laxity [6
] probably due to the lost tension from the damaged collagen fibers [7
]. Measurements performed in humans confirm this model and show that 24-h post-exercise rest resolves the residual laxity/creep of the tissue [4
]. Most likely, the second phase of the inflammation had been completed by then, restoring the lost viscoelastic properties of the damaged fibers. The major factor associated with a successful healing by the inflammation seems to be rest. Therefore, any moderate repetitive/cyclic exercise can benefit from at least 24 h of rest, and 48 h may be required for exercise under high loads. Understanding of the healing process provides justification for taking a 48-h rest between exercise sessions of the same joints. For example, exercise of a specific joint performed at 3:00 p.m. of 1 day should not be repeated the next day and preferably addressed again the second day, at about the same time. Such rest practice is already common in the athletic circles, but is validated here based on the inflammatory process.
Based on the literature reviewed above, we can propose a biphasic recovery theory of viscoelastic tissues exposed to repetitive exercise. The theory is summarized in Fig. and shows the initial displacement, Do, associated with load application as well as the exponentially increasing laxity during the exercise period reaching a maximum of DL at the end of loading. DL can be expressed as creep when calculated as a percentage of Do. As the post-exercise rest begins, Phase I of the inflammation is initiated, during which pro-inflammatory cytokines begin to be accumulated in the ligaments and direct the removal of the damaged molecules into the circulatory system. The duration of Phase I is not documented yet and may depend on the amount of damage present in the tissue, which is further associated with the load magnitude, frequency and duration of the physical activity. So far, the longest duration of Phase I available in the literature is 7–8 h. At the end of Phase I, a residual laxity, R1, is present, and is at an asymptotic saturation of its exponential decay, indicating that further significant recovery is not expected in this phase. As mentioned above, the residual laxity, R1, is due to the lost viscoelastic properties of the damaged collagen fibers. As Phase II begins, a new exponentially decaying equation describes the gain in the viscoelastic tension in the ligament as the reconstruction/repair of the damaged fibers is underway. This phase also reaches asymptotic saturation near baseline, signifying a full recovery to normal operating functionality. While literature fully supports and validates the different components, Phase II is a highly educated speculation and in need of experimental validation.
Fig. 4 A schematic of ligament elongation during loading and the biphasic recovery to baseline over the following rest. Note the large residual laxity at the end of the first phase and the second exponential decrease to baseline at the end of the second phase (more ...)
The balance of pro- and anti-inflammatory mediators determines the final tissue response. The limitation of most studies on this topic is that one cannot quantify all possible mediators. Furthermore, some cytokines, such as TGFb for example, exist as either pro- or anti-inflammatory mediators depending on the context and the circumstances (tissue type, injury status, repair status, etc.). Nevertheless, a significant elevated level of the major pro-inflammatory cytokines and especially of the pain-inducing IL-8 suggest a significant tissue injury and inflammatory response.
In conclusion, low-load activities such as aerobics, for example, should not be exempt from such work/rest protocols if overuse/cumulative disorder are to be prevented. Two types of rest, therefore, are available as a preventive measure for pro-inflammatory activity: an inter-session rest during the performance of exercise as an attenuation measure [33
] and an inter-day rest as a preventive measure from propagation of the acute phase into chronic inflammation. Repeating exercise of the same ligament/tendon on a daily basis for prolonged periods (weeks/months) can further aggravate a minor inflammation and convert it into a chronic inflammation: an irreversible and degenerative disease called overuse or cumulative disorder.