The Effect of a Single Freeze-Thaw Cycle on Matrix Metalloproteinases in Different Human Platelet-Rich Plasma Formulations

Kaitlyn E Whitney; Grant J Dornan; Jillian King; Jorge Chahla; Thos A Evans; Marc J Philippon; Robert F LaPrade; Johnny Huard

doi:10.3390/biomedicines9101403

The Effect of a Single Freeze-Thaw Cycle on Matrix Metalloproteinases in Different Human Platelet-Rich Plasma Formulations

Biomedicines. 2021 Oct 6;9(10):1403. doi: 10.3390/biomedicines9101403.

Authors

Kaitlyn E Whitney¹, Grant J Dornan¹, Jillian King¹, Jorge Chahla², Thos A Evans³, Marc J Philippon³, Robert F LaPrade⁴, Johnny Huard¹

Affiliations

¹ Steadman Philippon Research Institute, Vail, CO 81657, USA.
² Rush University Medical Center, Midwest Orthopaedics at Rush, Chicago, IL 60612, USA.
³ The Steadman Clinic, Vail, CO 81657, USA.
⁴ Twin Cities Orthopaedics, Edina, MN 55435, USA.

Abstract

Storing platelet-rich plasma (PRP) for future use is a compelling approach, presuming the retention of biological properties is maintained. However, certain factors in PRP preparations have deleterious effects for the treatment of certain musculoskeletal conditions. The purpose of this study was to measure and compare matrix metalloproteinase protein (MMP) concentrations between fresh and freeze-thawed leukocyte-rich PRP (LR-PRP) inactivated (LR-I) and activated (LR-A) preparations, and leukocyte-poor PRP (LP-PRP) inactivated (LP-I) and activated (LP-A) preparations. A volume of 60 mL of whole blood was drawn from 19 healthy donors. LP-I and LR-I samples were processed using a manual extraction and centrifugation methodology. LP-A and LR-A products were activated with 10% CaCl₂ and recombinant thrombin. Blood fractions were either immediately assayed and analyzed or stored at -80 °C for 24, 72 and 160 h. Multiplex immunoassay was used to measure MMP-1, MMP-2, MMP-3, MMP-9, MMP-10, and MMP-12. MMP-1 concentrations increased in LR-A (p < 0.05) and MMP-9 significantly increased in LR-I (p < 0.05), while MMP-2 significantly decreased in LR-I (p < 0.05) and MMP-3 concentrations significantly decreased in LR-A (p < 0.05). MMP-12 concentrations also significantly decreased in LR-I (p < 0.05) from baseline concentrations. There were no significant differences between LP-A and LP-I preparations and MMP concentrations. MMP-10 concentrations in all PRP samples compared to each freezing time point were also not significantly different. MMPs regulate components of the extracellular matrix (ECM) in the remodeling phase of musculoskeletal injury. In this study, we observed a significant increase and decrease in MMP concentrations in response to a single freeze-thaw cycle in inactivated PRP and activated PRP preparations. This evidence contributes to the growing body of literature on the optimization of PRP preparation and storage strategies prior to delivery. Our findings suggest that specific PRP preparations after a single freeze-thaw may be more advantageous for certain musculoskeletal applications based on the presence of MMP concentrations.

Keywords: freeze–thaw; leukocyte-poor PRP (LP-PRP); leukocyte-rich PRP (LR-PRP); matrix metalloproteinases (MMPs); platelet-rich plasma (PRP).