Exercise and physical activity are effective ways to reduce the risk of cardiovascular diseases, such as heart attack and stroke, and can provide valuable benefits beyond those of medications. Improved cardiac performance, along with cardiac hypertrophy, is a major feature of endurance exercise, leading to a constellation of adaptations that affect the structure, electrical conduction, and function of the heart and contribute to appropriate increases in cardiac output (Khoury et al., 2019; Moreira et al., 2020). Studies in animal models of exercise-induced cardiac hypertrophy (here in response to treadmill exercise, voluntary wheel running, and swim training) have shown preserved or enhanced contractile function and relative cardiac hypertrophy (Konhilas et al., 2004; Radovits et al., 2013; Mi et al., 2019). The important differences that exist between these experimental methods may affect the interpretation of the results. There is a plethora of studies demonstrating that treadmill running is a preferred option in comparative studies of the effects of exercise training because of its ability to precisely control exercise intensity and volume (Kemi et al., 2004; Wang et al., 2010; Tang et al., 2011).

Physiological cardiac hypertrophy induced by endurance exercise is related to an increase in cardiac mass and individual cardiomyocyte growth in both length and width, with no interstitial or replacement fibrosis or cell damage; this is considered reversible and does not develop into heart failure (Ellison et al., 2012). In contrast, although pathological hypertrophy is initially induced as a compensatory response to the growth of the ventricle, this kind of hypertrophy progresses to ventricular chamber dilatation, with wall thinning through the loss of myocytes and contractile dysfunction, resulting in adverse cardiovascular events (Nakamura and Sadoshima, 2018). The impact of endurance exercise on heart health depends on the combination of intensity, time duration, frequency, and exercise type (Davos, 2019). However, less is known regarding cardiac adaptations to the different intensities of treadmill running in rats. Most studies have shown that low- and moderate-intensity exercise attenuates abnormal cardiac remodeling and myocardial dysfunction and improves functional capacity (Mi et al., 2019; Pagan et al., 2019). Additionally, clinical studies also support this recommendation by identifying the benefits that can be derived from low- and moderate-intensity exercise (Wasfy and Baggish, 2016). Exercise performed at a high intensity appears to convey greater cardioprotective benefits than exercise of a moderate intensity, which may be because of the increase in aerobic fitness (Swain and Franklin, 2006). However, recent data show that high-intensity exercise has an adverse effect on the heart, along with impairment of cardiomyocyte Ca²⁺ handling, mitochondrial respiration, and the activation of proapoptotic and profibrotic activity (Olah et al., 2015; Ljones et al., 2017). Given that intensity dependence for cardiac function remains controversial, it is imperative to better understand how different treadmill running intensities alter the phenotype of the murine heart.

Exploring the molecular mechanisms of cardiac hypertrophy is helpful to better understand the adaptability of the heart. Among the numerous signaling factors, the extracellular signal-regulated kinases 1/2 (ERK1/2) pathway has been suggested to promote cardiac hypertrophy (Yan et al., 2021). For instance, transgenic mice overexpressing an activated dual specificity mitogen-activated protein kinase kinase 1 (MEK1) mutant under the transcriptional control of the cardiac-specific α-myosin heavy chain promoter were found to induce cardiac hypertrophy in vivo, which constitutively activated ERK1/2 in the heart (Bueno et al., 2000). Furthermore, another study demonstrated that the ERK1/2 signaling pathway uniquely regulates the balance between eccentric and concentric growth of the heart (Kehat et al., 2011). Even though many studies conducted on pressure overload or mutagenesis have examined the role of the MEK1/2-ERK1/2 pathway, the regulatory mechanisms for exercise-induced cardiac hypertrophy remain unclear.

The present study was designed to develop a rat model of three treadmill running programs to test whether different types of intensity training can induce cardiac structural and functional changes. Our hypothesis was that different exercise loads would result in intensity-dependent myocardial hypertrophy, which might be associated with different functional consequences in the left ventricle (LV) in rats. Therefore, we aimed to provide a structural and functional characterization of the LV in exercise-induced hypertrophy in rats. Additionally, we investigated molecular alterations in exercise-induced cardiac hypertrophy.

In the present study, we provided an in vivo structural and functional comparison of exercise-induced hypertrophy in a rat model, depicting that changes in cardiac response differ based on exercise intensity and potential signal transduction. Here, our findings indicate that running leads to cardiac hypertrophy in an intensity-dependent manner. In contrast to LIR and MIR, 8 weeks of HIR-induced cardiac hypertrophy was characterized by potential cardiomyocyte injury, which increased the risk of pathological development. Furthermore, the ERK signaling pathway was mainly involved in the compensatory hypertrophic process of the myocardium in the early stage of exercise and was positively correlated with exercise load. However, long-term exercise may attenuate ERK signaling activation.

In the process of long-term adaptation to regular exercise, myocardial hypertrophy leads to an increased blood pumping ability to meet the increased metabolic needs of the whole body (Ellison et al., 2012). Cardiac hypertrophy is characterized by myocyte hypertrophy, structural rearrangement, and the accumulation of cardiac collagen (Krzesiak et al., 2017). Following exercise training in humans and animal models, heart mass typically increases (Wang et al., 2010; Davos, 2019). Macroscopically, we found that the increases in the size of the hearts in the running groups were more obvious after 4 and 8 weeks. Moreover, the HM/BM ratio, which is widely used to assess cardiac hypertrophy, has shown similar results. Although the ratio of heart mass to tibia length (HM/TL) would seem to be more reliable, data from other studies (Shioi et al., 2003; Tang et al., 2017) have indicated that there were no significant differences between HM/BM and HM/TL. Microscopically, our study showed that 1 week of HIR induced cardiac hypertrophy, while myocyte hypertrophy mirrored the macroscopic situation only after 4 and 8 weeks of running. Indeed, there is evidence that high-intensity treadmill models more easily alter cardiac phenotypes in rodents (Kemi et al., 2005; Liao et al., 2015). Furthermore, these cardiac dimensions are consistent with previous experimental results (Tang et al., 2011), which have suggested that myocardial hypertrophy occurs in tandem with an increase in exercise intensity and extension of exercise time. To induce a hypertrophic heart, a longer period of exercise may be required.

In terms of structural arrangement and cardiac collagen, we found that low-to-medium-intensity running led to proper cell morphological characteristics and decreased fibrosis, which likely contributed to overall myocardial tissue integrity and homeostasis for cardiac beneficial effects (Libonati et al., 2011; Pagan et al., 2019). HIR, however, may have a different impact on the heart. Although few studies have compared the effects of different exercise intensities on histological changes in rat hearts, Liao et al. (2015) have shown that a wide range of largely primary myocellular defects and myocardium ischemia were observed after high-intensity exercise. This is consistent with our finding of some counterproductive cardiac consequences induced by exercise. We also noted that collagen deposition in the myocardium also increased significantly. An elegant study demonstrated increased cardiac fibrosis after long-term intensive treadmill running in a rat model and represented potentially adverse cardiac remodeling (Benito et al., 2011). Data from reviews also suggest the development of ventricular fibrosis as assessed by magnetic resonance imaging (MRI) in endurance athletes (Malek and Bucciarelli-Ducci, 2020). As a result of the accumulation of these minor injuries, exercise may produce unfavorable results.

The mechanisms by which high-intensity exercise promotes cardiac pathologies are unknown. It is possible that long-term cardiac overload plays a role by promoting physiological remodeling in the early phases, but the heart, as the circulation center of energy and nutrition, is very sensitive to ischemia and hypoxia, which may eventually become maladaptive in the long term. A recent study supports the idea that balanced intense exercise training induces hypertrophy that is not associated with pathological remodeling (Olah et al., 2021).

Transthoracic echocardiography is accurate and widely used in the field to assess cardiac structure and function. The present study found that low-to-medium intensity running induced a significant increase in absolute LV diastolic diameter values and a decrease in LV relative wall thickness after 1 week. These results indicate a shift to eccentric LV remodeling. According to Morganroth’s dichotomous concept, regular aerobic sports, such as running, are accompanied by cardiac chamber dilation, which is referred to as eccentric hypertrophy (Morganroth et al., 1975; Ellison et al., 2012). However, in contrast, other studies have demonstrated that short-duration exercise appears to have little in the way of a negative impact on ventricular function (Oxborough et al., 2010). Our study also showed that FS and EF significantly decreased in the LIR and MIR groups, but not in the HIR group, in the short term. This disparity might be a consequence of myocyte hypertrophy in the HIR group. In short-term low-to-moderate-intensity training, cardiomyocyte contractility and maximal oxygen consumption (VO₂max) were like those of the control group (Kemi et al., 2004). Meanwhile, heart growth is related to an increase in chamber dilation. In order to maintain the normal ratio of the number of cardiomyocytes to blood capillaries, there may be a temporary decline in systolic function. Moreover, the impact of prolonged running on EF and FS was not significantly altered between the exercise groups and the SED group after weeks 4 and 8. The observed unchanged cardiac function is in line with recent findings on rat and athlete hearts (Oosthuyse et al., 2012; Asif et al., 2018). Although transthoracic echocardiography is accurate and widely used to evaluate cardiac structure and function in this field, MRI technology provides more accurate and reproducible measurements of cardiac function (Nikolaidou and Karamitsos, 2020; Russo et al., 2020).

cTnI is a highly sensitive and specific marker of myocardial injury and is suitable for early and late diagnoses (Mair et al., 1996). In the present study, no obvious myocardial injury was observed after treadmill running with varying intensities. In contrast to previous observations of animal models (Liao et al., 2015) and a large number of human exercise studies (Kosowski et al., 2019), the release of cTnI was markedly decreased after 4 weeks of running, and the same state also occurred after 8 weeks for the LIR and MIR groups, indicating that the leakage of cTnI decreased after exercise. The reason for this situation is that cardiac biomarker concentrations tend to return to baseline at 24–48 h postexercise (Lippi and Banfi, 2010). However, according to the current data, running for 8 weeks between the HIR and SED groups has no obvious changes, which may lead to potential myocardial injury. Our HE and PSR staining of the myocardium showed signs of sporadic cardiomyocyte damage and increased fibrosis. However, this probably represents a standard exercise intensity-dependent response rather than a pathological response.

It is generally accepted that ERK1/2 activation is essential for cardiac hypertrophy (Gallo et al., 2019). However, several partially contradictory studies have indicated that ERK1/2 can lead not only to maladaptive cardiac hypertrophy (Lorenz et al., 2009; Tomasovic et al., 2020), but also physiological hypertrophy (Bueno et al., 2000; Mutlak et al., 2018), and some studies have also suggested that it has no effect on cardiac hypertrophy (Purcell et al., 2007). In addition, Kehat et al. (2011) demonstrated that the ERK1/2 signaling pathway uniquely regulates the balance between eccentric and concentric growth of the heart. It has been theorized that the model used, circumstances, and upstream signals may affect the outcomes of these mouse studies on ERK1/2-mediated cardiac hypertrophy. In our rat model, we found that only HIR resulted in the activation of ERK1/2 after 1 week. The amount of phosphorylated ERK1/2 increased significantly by 134.3, 137.1, and 237.1% when the rats ran at low, medium, and high intensities, respectively, for 4 weeks. However, there were no significant changes in ERK1/2 activation between the running groups of varying intensities and the SED group at week 8. As reported, exercise induces the activation of multiple mitogen-active protein kinase (MAPK) pathways in the heart, an effect that gradually declines with the development of exercise-induced cardiac hypertrophy (Iemitsu et al., 2006). Interestingly, Hayashida et al. (2001) demonstrated that c-Jun N-terminal kinases (JNK) and p38-MAPK were similarly differentially regulated in the stages of the chronic hypertrophic process, while angiotensin (Ang) II-induced ERK activation was preserved. Therefore, the combined results from previous studies and our present evidence suggested that ERK1/2 represents a positive regulator in the progression of cardiac hypertrophy and is correlated with running intensity, which is, however, alleviated by increased running duration. Our research may provide valuable input regarding striking a balance between intensity and duration in terms of exercise-induced cardiac hypertrophy.

In summary, we confirmed that the effect of running on cardiac hypertrophy is intensity dependent. Compared with LIR and MIR, myocardial hypertrophy induced by HIR at 8 weeks was characterized by sporadic heart injuries, cumulative fibrosis, and increased myocardial enzymes, increasing the risk of pathological development. In addition, the ERK signaling pathway was mainly involved in the compensatory hypertrophy process of early exercise and was positively correlated with exercise load. However, long-term exercise may weaken the activation of the ERK signal.

The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.

The animal study was reviewed and approved by the Animal Ethics Committee of Fujian Medical University.

ZY and GN conceived research concept and drafted, revised, and edited the manuscript. ZY and NZ designed and performed the experiments, analyzed the data, and prepared the figures. NZ, JL, and TL contributed to the sample collection and sample storage. The authors read and approved the final manuscript. All authors contributed to the article and approved the submitted version.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.