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Pilot Production of Mesenchymal Stem/Stromal Freeze-Dried Secretome for Cell-Free Regenerative Nanomedicine: A Validated GMP-Compliant Process

2018; Multidisciplinary Digital Publishing Institute; Volume: 7; Issue: 11 Linguagem: Inglês

10.3390/cells7110190

2073-4409

Elia Bari, Sara Perteghella, Dario Di Silvestre, Marzio Sorlini, Laura Catenacci, Milena Sorrenti, Giorgio Marrubini, Rossana Rossi, Giuseppe Tripodo, Pierluigi Mauri, Mario Marazzi, Maria Luisa Torre,

Mesenchymal stem cell research

In this paper, a pilot production process for mesenchymal stem/stromal freeze-dried secretome was performed in a validated good manufacturing practice (GMP)-compliant cell factory. Secretome was purified from culture supernatants by ultrafiltration, added to cryoprotectant, lyophilized and characterized. We obtained a freeze-dried, "ready-off-the-shelf" and free soluble powder containing extracellular vesicles and proteins. In the freeze-dried product, a not-aggregated population of extracellular vesicles was detected by nanoparticle tracking analysis; Fourier transform infrared spectra showed the simultaneous presence of protein and lipids, while differential scanning calorimetry demonstrated that lyophilization process successfully occurred. A proteomic characterization allowed the identification of proteins involved in immune response, response to stress, cytoskeleton and metabolism. Moreover, the product was not cytotoxic up to concentrations of 25 mg/mL (on human fibroblasts, chondrocytes and nucleus pulposus cells by MTT assay) and was blood compatible up to 150 mg/mL. Finally, at concentrations between 5 and 50 mg/mL, freeze-dried secretome showed to in vitro counteract the oxidative stress damage induced by H₂O₂ on nucleus pulposus cells by MTT assay.