Different pro-angiogenic potential of γ-irradiated PBMC-derived secretome and its subfractions

Abstract

Secretomes from various cell sources exert strong regenerative activities on numerous organs, including the skin. Although secretomes consist of many diverse components, a growing body of evidence suggests that small extracellular vesicles (EVs) account for their regenerative capacity. We previously demonstrated that the secretome of γ-irradiated peripheral blood mononuclear cells (PBMCs) exhibits wound healing capacity. Therefore, we sought to dissect the molecular composition of EVs present in the secretome and compared wound healing-related activities of these EVs to other subfractions of the secretome and the fully supplemented secretome (MNCaposec). Compared to EVs derived from non-irradiated PBMCs, γ-irradiation significantly increased the size and number and changed the composition of released EVs. Detailed characterization of the molecular components of EVs, i.e. miRNA, proteins, and lipids, derived from irradiated PBMCs revealed a strong association with regenerative processes. Reporter gene assays and aortic ring sprouting assays revealed diminished activity of the subfractions compared to MNCaposec. In addition, we showed that MNCaposec accelerated wound closure in a diabetic mouse model. Taken together, our results suggest that secretome-based wound healing represents a promising new therapeutic avenue, and strongly recommend using the complete secretome instead of purified subfractions, such as EVs, to exploit its full regenerative capacity.

The secretome of stressed peripheral blood mononuclear cells increases tissue survival in a rodent epigastric flap model

“…40,48 PBMCsec was categorized as a biological medicinal product (Directive 33 In most of the studied biological processes the integrity of the entire PBMC secretome was necessary to achieve full regenerative effects as sub-fractions or single cell types did not reach the same capacity. 32,33 Unlike previously described monocausal therapies, the effects of PBMCsec can therefore be attributed to its heterogeneity, resembling the physiological environment necessary for tissue regeneration. Previously described therapies with single growth factors significantly reduced postoperative flap necrosis at a similar rate to the described results.…”

“…31 Several mechanisms have already been characterized, indicating that the entire PBMCsec is required to exert its full action spectrum, as subfractions thereof did not reach the same beneficial effects. 32,33 The pleiotropic effects of the secretome components better resemble the physiologic environment of wound healing occurring in the body, thus resulting in a faster and better regeneration. PBMCsec was successfully applied in animal models of wound healing, tissue ischemia or inflammation.…”

Role for Lipids Secreted by Irradiated Peripheral Blood Mononuclear Cells in Inflammatory Resolution in Vitro

“…Briefly, PBMCs were isolated from heparinized blood using density gradient centrifugation via Ficoll-Paque PLUS (GE Healthcare Bio-Sciences AB, Uppsala, Sweden). The buffy coat containing the PBMCs was resuspended at 2.5 × 10 7 cells/mL in CellGro serum-free medium (CellGenix, Freiburg, Germany), irradiated with Cesium-137 (60 Gy), and cultivated for 24 h [55]. Supernatants were collected by centrifugation at 400× g for 9 min and passed through 0.2 µm filters.…”

“…Reconstitution was performed by serum-free media to receive a stock concentration equivalent to 2 × 10 7 cells/mL that was further diluted to a working concentration equivalent to or below 1 × 10 7 cells/mL. The lipid fraction was prepared based on a traditional chloroform/methanol extraction method [57] with minor modifications [55]. All experiments were performed with batches A000918399132 and A000918399129.…”

Quantitative Hybrid Cardiac [18F]FDG-PET-MRI Images for Assessment of Cardiac Repair by Preconditioned Cardiosphere-Derived Cells

“…Separate fractions, as well as secretomes of PBMC subpopulations, failed to replicate completely the effects regarding tissue repair, wound healing, and angiogenesis. 15 , 16 , 17 …”