Results did not support our first hypothesis that changes with altered loading would begin at the proximal insertion into bone and progress along the tendon length with time. After 1 week of altered loading, changes were seen in composition only and the changes found were seen at multiple locations along the tendon. However, after 4 weeks of increased loading, organizational changes were seen only in the intra-articular space, which then progressed to the proximal intratubercular groove by 8 weeks. It is possible that organizational and histological parameters do not change at the proximal insertion into bone due to its normal state being a disorganized one with a prevalent rounded cell phenotype. Changes therefore would be seen first next to this region, as was seen at 4 weeks with increased angular deviation in the intra-articular space with increased loading. The increased compressive loading in this region that normally sees primarily tensile loading, resulted in collagen organization much more like the proximal insertion into bone than the distal portion of the tendon. Additional analysis of the collagen organization data showed that with increased loading, organization was not different between the proximal insertion into bone, intra-articular space and proximal intratubercular groove. Organization did not increase in this group until the distal intratubercular groove. Conversely, in both the SI only and SI+DEC groups, tendon organization improved significantly between the proximal insertion into bone and intra-articular space and remained constant between in the intra-articular space and proximal and distal intratubercular grooves.
Our second hypothesis that increased loading would result in further detrimental changes was supported as increased angular deviation was seen compared to detachment alone at both 4 and 8 weeks. However, there were no differences between increased loading and detachment alone at any time point in cellularity and cell shape. This may be due to the fact that both detachment alone and detachment followed by increased loading have altered pathology, and therefore, it was difficult to detect a difference in a semi-quantitative parameter such as histological grading. For instance, both groups were a grade 2 or 3 in both parameters at all locations, and therefore statistical differences in these semi-quantitative parameters were not seen between the two as were seen in a quantitative parameter such as angular deviation. After 4 weeks, increased loading did result in increased aggrecan and collagen I production at the proximal insertion into bone compared to detachment alone, but by 8 weeks there were no compositional differences between these groups.
Results also support our third hypothesis that decreased loading would result in improved tendon properties, or properties more like an uninjured tendon than one in the presence of a rotator cuff tear. After 1 week, changes with decreased loading were seen only in composition where increased production of several ECM components such as collagens I, III and XII were seen. However, 8 weeks following detachments, increased organization was found compared to both SI only and SI+INC in the intra-articular space and proximal and distal intratubercular grooves. Decreased loading also resulted in decreased cellularity in the intra-articular space and proximal intratubercular groove as well as a more elongated cell phenotype along the entire tendon away from the proximal insertion into bone. Decreases in expression of aggrecan and biglycan were also seen at this time point. Aggrecan is an ECM component usually seen in cartilage or regions of compressive loading and biglycan is a proteoglycan indicative of an injury response. The decreased expression of these components with decreased loading in combination with the increased organization and more elongated cell shape indicates that with decreased loading, the structure of the tendon more closely resembles its normal structure and composition.11
Interestingly, changes were not seen in histological grading or organization with altered loading 1 week following detachments. Additionally, compositional changes seen at this time point were not consistent in location and often detrimental changes were seen with decreased loading, which is not supported at later time points. Tendons in all groups were in the presence of rotator cuff tears, and these results indicate that after a short period of time, altered loading does not seem to have an effect. After 4 weeks, decreased loading did not yet show an improvement compared to detachment alone but did not exhibit detrimental compositional changes as seen after 1 week. These results were not surprising, as both improved and detrimental changes in mechanical parameters were seen with decreased loading at this time point in our earlier studies.14
It was not until 8 weeks following detachments that improved tendon properties were seen with decreased loading. However, improvements in tendon mechanics were not seen at this time point.14
It is possible that the composition and organization more like uninjured tendons found with decreased loading in the current study may lead to improved tendon mechanics at later time points, as was seen in other studies investigating the effect of decreased loading by immobilization.3,17
Organizational changes in the biceps tendon with a supraspinatus+infraspinatus detachment were seen first in the intra-articular space when compared to a sham surgery.11
In this study, organizational changes with increased loading were also seen first in the intra-articular space, further indicating that this may be the site where pathological changes originate. Results of this study also support the theory of increased loading as the mechanism responsible for biceps tendon pathology in the presence of rotator cuff tears. In this study, further changes were seen in organization with increased loading at the same location changes were first seen with detachment alone. In addition, decreasing the load on the tendon resulted in dramatically improved tendon organization and cell morphology compared to those in the presence of cuff tears alone or cuff tears followed by increased loading, indicating that with removal of load, the tendon’s organization is more like an uninjured tendon. The results regarding decreased loading indicate that this is a promising area of future study and that with rotator cuff repair, it is possible that biceps tendon pathology may be recoverable. For example, studies could be done to investigate the effect of delayed decreased loading, to see if a positive effect is still seen in tendons that see a period of normal loading following cuff tendon detachment before the initiation of decreased load.
This study is not without limitation. Quantification of the amount of load added to or taken away from the biceps tendon in our increased and decreased loading scenarios was not possible. However, preliminary studies were conducted to confirm that the methods used to induce altered loading did indeed increase and decrease the load on the tendon. Additionally, in patients with rotator cuff tears, changes to the biceps tendon take place after being exposed to an abnormal mechanical environment for a long period of time. The changes that occur in this study happen much faster, and the method of increased loading by detaching the short head of the biceps tendon would not occur clinically. It is also possible that additional biological changes may be taking place that were not detected with the immunohistochemical staining performed in this study. A group of targets were selected that, according to previous studies, would be expected to show differences across both time and the length of the tendon and have been extensively used for rotator cuff tendons in the rat model. Future work with this model may include additional biological assays such as PCR or staining for targets involved in collagen turnover.