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• Leucocyte Rich and Poor Platelet Concentrates and Tenocyte Proliferation
  Marko Bodor, MD, Ryan Dregalla, PhD and Yvette Uribe, BA
  Published on: 3 December 2017

• Published on: 3 December 2017

  Leucocyte Rich and Poor Platelet Concentrates and Tenocyte Proliferation

  • Marko Bodor, MD, Interventional Spine and Sports Medicine Physician University of California San Francisco, University of California Davis, Bodor Clinic
  • Other Contributors:
    • Ryan Dregalla, PhD, Regenerative Science Research & Development
    • Yvette Uribe, BA, Clinical Researcher

  We read with interest the article by Parrish et al, “Normal platelet function in platelet concentrates requires non-platelet cells: a comparative in vitro evaluation of leucocyte-rich (type 1a) and leucocyte-poor (type 3b) platelet concentrates.”(1)

  Parrish et al define PRP as a preparation with a platelet concentration of at least 5x over baseline, yet the LP-PRP they prepared (Arthrex Autologous Conditioned Plasma) was significantly lower at 2x over baseline, while the LR-PRP (Mitek Sports Medicine PEAK PRP) was significantly higher at 8x over baseline. We might reasonably expect that the ratio of growth factors between their LR-PRP and their LP-PRP to be approximately 8x/2x or 4:1, and this was indeed the case as seen in their Figure 4.

  Subsequently, the authors grew tenocytes (tendon cells) exposed to serum derived from LR-PRP and LP-PRP preparations. Given that their LR-PRP was approximately 4 times richer in growth factors than their LP-PRP, we might reasonably expect that the 2.5% solution of serum derived from their LR-PRP have approximately the same effect as the 10% solution of serum derived from their LP-PRP. However, their 10% LP-PRP solution actually resulted in higher growth of tenocytes (2656 light units) than their 2.5% LR-PRP solution (1001 light units), as seen in their Table 5, but not discussed by the authors. The fact that their 10% LR-PRP-derived serum caused tenocytes to grow to confluence while their 10% LP-PRP-derived serum did...

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  We read with interest the article by Parrish et al, “Normal platelet function in platelet concentrates requires non-platelet cells: a comparative in vitro evaluation of leucocyte-rich (type 1a) and leucocyte-poor (type 3b) platelet concentrates.”(1)

  Parrish et al define PRP as a preparation with a platelet concentration of at least 5x over baseline, yet the LP-PRP they prepared (Arthrex Autologous Conditioned Plasma) was significantly lower at 2x over baseline, while the LR-PRP (Mitek Sports Medicine PEAK PRP) was significantly higher at 8x over baseline. We might reasonably expect that the ratio of growth factors between their LR-PRP and their LP-PRP to be approximately 8x/2x or 4:1, and this was indeed the case as seen in their Figure 4.

  Subsequently, the authors grew tenocytes (tendon cells) exposed to serum derived from LR-PRP and LP-PRP preparations. Given that their LR-PRP was approximately 4 times richer in growth factors than their LP-PRP, we might reasonably expect that the 2.5% solution of serum derived from their LR-PRP have approximately the same effect as the 10% solution of serum derived from their LP-PRP. However, their 10% LP-PRP solution actually resulted in higher growth of tenocytes (2656 light units) than their 2.5% LR-PRP solution (1001 light units), as seen in their Table 5, but not discussed by the authors. The fact that their 10% LR-PRP-derived serum caused tenocytes to grow to confluence while their 10% LP-PRP-derived serum did not is irrelevant given that the latter was obtained from 4-fold fewer platelets.

  One of the purported new findings of the study was that LP-PRP was “deficient” in terms of coagulation, clot retraction and platelet growth factor release. This statement was based on an average 20% lower release of growth factors relative to whole blood following activation with CaCl2, as seen in their Figure 5. In addition, the authors stated “Exogenous thrombin addition rescues coagulation and clot retraction deficits and improves platelet growth factor release in LP-PRP.” These statements are not supported by their research because coagulation and clot retraction assays were not conducted in this study, and use of the word “rescues” is misleading.

  Platelet concentrations vary broadly (150-450K/µL) in normal individuals (2) with normal coagulation and clot retraction function. Whether an average 20% lower immediate release of growth and coagulation factors induced by CaCl2 would translate into an actual in-vitro or in-vivo coagulation or clot retraction deficit is not clear and was not assessed by this study. Furthermore, platelet growth factor release is temporal and multiphasic in nature and there is a known differential between preparation methods (3-5). Hence, whatever differences were seen at 80 minutes, the time point when serum was collected from the preparations, may not have been present at later time points.

  Another purported new finding, reflected by the title of the article, was that non-platelet cellular components in platelet concentrates are important for proper platelet function, including thrombin generation and clot retraction. Again, this was not supported by their research because platelet function, thrombin generation and clot retraction studies were not done and the statement appears to hinge on growth factors released within 80 minutes of the preparations.

  In conclusion, the presented findings must be considered as artifacts of the specific materials, methods and time points selected. If an implicit goal of the article was to demonstrate the superiority of LR-PRP over LP-PRP, then the platelet concentrations should have been similar and platelet rich plasma used instead of serum. Nevertheless, correcting for platelet concentration LP-PRP serum appears to have been at least twice as effective at inducing tenocyte proliferation. Furthermore, even if LR-PRP and LP-PRP were produced with similar levels of growth factors, LR-PRP has pro-inflammatory and catabolic factors attributable to leucocytes, a fact not acknowledged by the authors. (6,7)

  Acknowledgement: We would like to thank the Napa Medical Research Foundation, a 501(c)3 non-profit foundation, for their encouragment and support.

  References

  1 Parrish, W. R. et al. Normal platelet function in platelet concentrates requires non-platelet cells: a comparative in vitro evaluation of leucocyte-rich (type 1a) and leucocyte-poor (type 3b) platelet concentrates. BMJ Open Sport Exerc Med 2, e000071, doi:10.1136/bmjsem-2015-000071 (2016).
  2 Brown, L. M. et al. A normal platelet count may not be enough: the impact of admission platelet count on mortality and transfusion in severely injured trauma patients. J Trauma 71, S337-342, doi:10.1097/TA.0b013e318227f67c (2011).
  3 Schar, M. O., Diaz-Romero, J., Kohl, S., Zumstein, M. A. & Nesic, D. Platelet-rich concentrates differentially release growth factors and induce cell migration in vitro. Clin Orthop Relat Res 473, 1635-1643, doi:10.1007/s11999-015-4192-2 (2015).
  4 McCarrel, T. & Fortier, L. Temporal growth factor release from platelet-rich plasma, trehalose lyophilized platelets, and bone marrow aspirate and their effect on tendon and ligament gene expression. J Orthop Res 27, 1033-1042, doi:10.1002/jor.20853 (2009).
  5 David, M. D. E. et al. Do the Fibrin Architecture and Leukocyte Content Influence the Growth Factor Release of Platelet Concentrates? An Evidence-based Answer Comparing a Pure Platelet-Rich Plasma (P-PRP) Gel and a Leukocyte- and Platelet-Rich Fibrin (L-PRF). Current Pharmaceutical Biotechnology 13, 1145-1152, doi:http://dx.doi.org/10.2174/138920112800624382 (2012).
  6 Sundman, E. A., Cole, B. J. & Fortier, L. A. Growth factor and catabolic cytokine concentrations are influenced by the cellular composition of platelet-rich plasma. Am J Sports Med 39, 2135-2140, doi:10.1177/0363546511417792 (2011).
  7 Anitua, E., Zalduendo, M., Troya, M., Padilla, S. & Orive, G. Leukocyte inclusion within a platelet rich plasma-derived fibrin scaffold stimulates a more pro-inflammatory environment and alters fibrin properties. PLoS One 10, e0121713, doi:10.1371/journal.pone.0121713 (2015).

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  Conflict of Interest:
  None declared.

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