We performed a comprehensive assessment of muscle alterations as a result of ACL injury and rehabilitation through a combined approach involving muscle structural quantification using DTI-MRI and muscle morphology analysis using biopsied tissue. Alterations occurred in the vastus lateralis muscle of the injured limb compared with the control limb, and the alterations did not improve following subsequent surgery and rehabilitation. These results indicate that there are several pathophysiological responses of the muscle to an ACL tear as well as other alterations that become more apparent following reconstruction and rehabilitation of the ACL.
There were several notable alterations in the vastus lateralis muscle prior to surgery. One of the most important early alterations was the significant reduction in the PCSA as a result of ACL injury. Similar findings have been reported in research assessing the effects of disuse33. We examined the 3 individual components of the PCSA—volume, pennation angle, and fiber length—to determine which were driving this change (Table I). The fiber tract length did not change, which we speculate was because subjects did not maintain the limb in a shortened position. Previous work has shown that when subjects have undergone casting, fiber length decreases only if the limb is immobilized in a shortened position34. In contrast, we found that the pennation angle was significantly different. Although the pennation angle is not a major determinant of the PCSA, a reduction in this angle will limit the number of fibers within a given area and subsequently result in less force development. The observed reduction in the PCSA is most likely primarily attributable to a reduction in the volume of the vastus lateralis. The reduction in muscle volume is consistent with other studies, although the magnitude of the reduction in the present study is larger than in those studies35,36. Potential differences in subject characteristics and in the methods used to determine volume may explain some of the differences between studies.
There were several significant cellular alterations that occurred following the injury but prior to surgery. Although we did not find significant changes in the distribution of muscle fiber types before surgery, there was a significant reduction in the CSA of type-IIA muscle fibers. The selective atrophy of the type-IIA fibers confirms the speculations of previous authors6,16,26,27. Potentially, the selective reduction of type-IIA fiber CSA is due to a lack of input from the gamma motor neuron loop, which affects high-threshold motor neurons activating type-IIA fibers37,38. Additional work is needed to more directly test the link between sensory alterations due to an ACL tear and adaptations within the muscle.
Other cellular adaptations of the vastus lateralis muscle due to the ACL injury included greater extracellular matrix and fewer satellite cells prior to surgery. Little research has defined fibrotic changes in muscle as a result of an orthopaedic injury. For example, imaging studies in subjects who have had a hamstring strain have shown an increase in nonactive contractile components39,40. The end result of greater muscle extracellular matrix is less area occupied by active contractile components, effectively reducing force generation. Additionally, the reduction in satellite cell content in the injured muscle before surgery may impair the muscle’s ability to respond to subsequent rehabilitation. Satellite cell content is strongly correlated with strength gains in response to resistance training in the elderly, with the satellite cells providing myonuclei by fusing to the growing fibers41-43.
Muscle morphology did not improve significantly after surgery and rehabilitation. For example, the PCSA did not improve following reconstruction of the ACL and rehabilitation. Previous reports have found a reduction in muscle volume following surgery and rehabilitation but have not reported on PCSA35,36. We also found little change in muscle volume or pennation angle. Our results are in contrast to another report that used ultrasonography and found no difference in pennation angle at 2 years of follow-up22. Differences in the time to follow-up and in the technique used to assess pennation angle make a direct comparison between these studies difficult. However, these data provide further evidence of the need for rehabilitation strategies focused on earlier intervention, such as eccentric exercises, which are known to improve PCSA and the components contributing to it44-48.
We also report several other adaptations that occur after surgery and rehabilitation. There was a selective reduction in type-IIA muscle fiber frequency and increased abundance of type-IIA/X hybrid fibers. In addition, we found no significant improvement in the CSA of specific muscle fiber types. In the only previous study of fiber atrophy after ACL surgery that we are aware of, type-II muscle fiber CSA was reduced up to 1 year after surgery relative to that in the nonoperative limb15. However, that study focused only on fiber CSA, did not consider type-IIA/X hybrid fibers, and used outdated surgical and rehabilitation techniques15. The shift from type-IIA to type-IIA/X hybrid fibers has been shown in other populations to be indicative of a detrained state49-51. By contrast, subjects in the present study underwent a rehabilitation period, and at the time of the second biopsy they had begun to return to their previous activity levels. These results suggest that deficits in type-IIA fiber CSA and deleterious alterations in fiber type composition continue during current rehabilitation protocols.
The persistence of the increase in extracellular matrix and decrease in satellite cell content despite surgical reconstruction and rehabilitation demonstrate the need to intervene early following an ACL tear to prevent changes at the cellular level that may ultimately limit muscle adaptation during rehabilitation. Additionally, whether thickening of the muscle extracellular matrix can be reversed in humans through rehabilitation has yet to be determined. If irreversible, these alterations may necessitate the need to develop strategies to optimize the remaining muscle function. Emerging evidence suggests that satellite cells may play an important role in the regulation of extracellular matrix in muscle29. The lack of restoration of satellite cells following surgery may contribute to the protracted weakness, greater extracellular matrix, and reduced ability of muscle fibers to hypertrophy.
The between-limb differences in quadriceps strength were larger than in some previous reports but similar to the differences in others3,12,52-54. Differences in the mode of testing (isometric versus isokinetic), type of graft, timing of the testing, and age are all factors that may potentially explain these differences. Future studies in larger cohorts comparing the morphological features of those with the least and greatest between-limb differences are warranted given the observed differences.
The present study had several limitations. First, we were unable to control the time window between injury and surgery, although subjects had surgery within 2 months of the injury. Second, the sample size is small and includes subjects with either bone-patellar tendon-bone or hamstring grafts, but the level of complexity of the study and time commitments from subjects make large sample sizes for this work a challenge. Lastly, we were not able to image or take a biopsy from the uninjured limb following surgery, limiting our ability to identify contralateral changes in the examined variables.
In conclusion, we have shown that there are pathophysiological responses within muscle following an ACL injury, with reductions in muscle fiber volume and pennation angle resulting in reduced PCSA as well as greater extracellular matrix and reduced satellite cell frequency. These measures do not improve following surgery and rehabilitation. There was also a significant shift to a greater frequency of type-IIA/X muscle fibers following surgery and rehabilitation. Future strategies to address these clinical issues must be developed.
Disclaimer: The content of this article is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
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