Abstract
Mesenchymal stem cells (MSCs) have been observed to have low engraftment rate after transplantation, yet their regenerative benefits remain long after their presence is lost. There is strong evidence that the therapeutic effects come from the cells’ paracrine mechanisms, which include secretion of free molecules (RNAs, proteins, etc.) and extracellular vesicles. In recent years the smallest extracellular vesicles, exosomes, have been of immense interest due to their delivery function of vital molecules to cells throughout the human body. There are many components of the exosomes that can be manipulated such as their internal components or their surface proteins; however, the production yield is typically very low when extracted from routine cell culture media. We proposed that the extracellular environment could be modified to increase the yield of exosomes from the cultured MSCs. In this study, we investigated MSCs derived from the chorionic villus of placenta tissue (PMSCs). Both the effect of oxygen tensions and extracellular matrices on the secretion of free proteins and exosomes were studied. Among free proteins, the secretory levels of brain-derived neurotrophic factor (BDNF) and hepatocyte growth factor (HGF) were examined. The BDNF levels decreased under the hypoxic conditions when compared to the normoxic conditions while the HGF levels increased overall at 3% O2. The exosome secretion had a negative effect under the hypoxic conditions compared to the 20% O2. For the extracellular matrices, the PMSCs were seeded onto the human amnion membrane (AM) and porcine small intestine submucosa (SIS) patches. The BDNF level was decreased to zero from the SIS while its level was decreased by 2.57-fold from the AM compared to the plastic surface. The HGF levels increased by 2.7-fold from the SIS and 3.13-fold from the AM compared to the plastic. The ECMs had negligible effect on the exosome secretion. We also proposed a possible mass-production method to producing exosome-mimicking nanovesicles with PMSCs and a mini-extruder. Our results indicated that the produced nanovesicles were similar to the native exosomes in terms of their size and the population was overall homogenous. Also, PMSCs could be modified so that the nanovesicles would reflect the modifications, such as the internal blocking of PMSCs, although much fine-tuning would be needed. Overall the PMSC culture conditions could be engineered to influence paracrine mechanisms and increase the production of exosomes.