DISCUSSION
Our hypothesis that the injection of leukocytes would cause an inflammatory response in rat tendons leading to catabolic outcomes was confirmed in the granulocyte group. It should be noted that prior to the catabolic outcome, there was a transient anabolic effect in the granulocyte group during the second week. Deterioration in morphology was observed in the tendons injected with granulocytes on day 7, which subsequently recovered in the following week. We found that injecting granulocytes into normal tendons resulted in an increase in inflammatory cellularity, when compared to monocytes and plasma injections.
Limitations inherent in this study are those similar to other in vivo studies. To begin with, the results of injections into rat tendons may not be translatable to human tendons. Despite this limitation, the rat is a common model for tendon research.31 Another limitation is that this study injected healthy Achilles tendons, rather than tendons with preexisting tendinopathy. In a naturally occurring tendinopathy, there may be other factors present that interact with PRP, and this model negates the contribution of these factors. Finally, while the immunohistochemistry (IHC) and morphological data are clear, the cellularity data are not clear in identifying the type of cells that were increased by granulocyte injection. However, the cells appeared rounded, resembling inflammatory infiltrate; a common cell type seen in tendons.2 While fibroblasts are also a common infiltrate during chronic tendinopathy, they are generally flat and appear on H&E as long spindle shaped cells. Thus, we believe the increased cellularity of the tendons after granulocyte injections is representative of an increase in inflammation. The increased cellularity could be due to the increased number of cells injected into the tendon; however, our conclusions are consistent with the increased inflammation previously reported linking leukocytes to tendon inflammation.20,22,32
In terms of morphology, we hypothesized that degenerative changes would be seen in the tendons that were injected with granulocytes due to the inflammatory action of these cells. As part of the granulocyte response, neutrophils release proteases and macrophages can stimulate collagen synthesis via fibroblasts, both causing change within the extracellular matrix.33,34 Indeed, we observed a significant change in tissue morphology in the granulocyte group over the course of 14 days. As the degenerative and regenerative effects of granulocytes take time to present, this is likely what we observed to occur between day 7 and 14 after treatment. These observations are also consistent with prior observations that leukocyte-rich PRP injections can be detrimental to tendon healing, but beneficial to tissue degeneration in the setting of chronic tendonitis.20
We hypothesized that the vascularity of the tendons would be similar in all preparations. This was based on previous studies demonstrating that the lack of platelets in the platelet-poor plasma fraction is sufficient to deplete VEGF, the angiogenic agent in PRP.35 In this study, there were no observable differences in vascularity of platelet-poor plasma, monocyte, and granulocyte injections. We attribute this to the lack of VEGF in any of these preparations. The aforementioned study also showed that the lack of platelets in injection was enough to prevent the angiogenic effect of this treatment.35
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