Purpose: Tissue regeneration can be improved by local application of autologous bone marrow derived progenitor cells (BMSC) and platelet rich plasma (PRP). However, there is a lack of standardized application procedures for clinical use. Therefore, a technique in accordance with the guidelines for advanced therapies medical products of the European Medicine Agency was developed and established.
Methods: In detail, a process for the isolation and formulation of autologous bone marrow cells (BMC) and PRP in a clinical setting was validated. To investigate the influence of storage time and temperature on gel formation and gel stability, different concentrations of BMC were stored with and without additional platelets, thrombin and fibrinogen and analyzed over a period of 28 days. In addition, cell vitality using a live-dead staining and migration ability of human mesenchymal stem cells (hMSC) in the gel clot was investigated.
Results: For an optimized stable gel clot, human BMC and PRP should be combined with 10% to 20% fibrinogen (9 mg/mL to 18 mg/mL) and 5% to 20% thrombin (25 I.E. to 100 I.E.). Both freshly prepared and stored cells for 1 to 7 days had a stable consistence over 28 days at 37°C. Different platelet concentrations did not influence gel clot formation. The ratio of living cells did not decrease significantly over the observation period of 5 days in the live-dead staining.
Conclusions: The study identified an optimal gel texture for local application of BMC and PRP. Seeded hMSC could migrate therein and were able to survive to initiate a healing cascade.