Human platelet lysate as a potential clinical-translatable supplement to support the neurotrophic properties of human adipose-derived stem cells

Abstract

Background The autologous nerve graft, despite its donor site morbidity and unpredictable functional recovery, continues to be the gold standard in peripheral nerve repair. Rodent research studies have shown promising results with cell transplantation of human adipose-derived stem cells (hADSC) in a bioengineered conduit, as an alternative strategy for nerve regeneration. To achieve meaningful clinical translation, cell therapy must comply with biosafety. Cell extraction and expansion methods that use animal-derived products, including enzymatic adipose tissue dissociation and the use of fetal bovine serum (FBS) as a culture medium supplement, have the potential for transmission of zoonotic infectious and immunogenicity. Human-platelet-lysate (hPL) serum has been used in recent years in human cell expansion, showing reliability in clinical applications. Methods We investigated whether hADSC can be routinely isolated and cultured in a completely xenogeneic-free way (using hPL culture medium supplement and avoiding collagenase digestion) without altering their physiology and stem properties. Outcomes in terms of stem marker expression (CD105, CD90, CD73) and the osteocyte/adipocyte differentiation capacity were compared with classical collagenase digestion and FBS-supplemented hADSC expansion. Results We found no significant differences between the two examined extraction and culture protocols in terms of cluster differentiation (CD) marker expression and stem cell plasticity, while hADSC in hPL showed a significantly higher proliferation rate when compared with the usual FBS-added medium. Considering the important key growth factors (particularly brain-derived growth factor (BDNF)) present in hPL, we investigated a possible neurogenic commitment of hADSC when cultured with hPL. Interestingly, hADSC cultured in hPL showed a statistically higher secretion of neurotrophic factors BDNF, glial cell-derived growth factor (GDNF), and nerve-derived growth factor (NFG) than FBS-cultured cells. When cocultured in the presence of primary neurons, hADSC which had been grown under hPL supplementation, showed significantly enhanced neurotrophic properties. Conclusions The hPL-supplement medium could improve cell proliferation and neurotropism while maintaining stable cell properties, showing effectiveness in clinical translation and significant potential in peripheral nerve research.

Human Platelet Lysate Acts Synergistically With Laminin to Improve the Neurotrophic Effect of Human Adipose-Derived Stem Cells on Primary Neurons in vitro

“…The DRG explant outgrowth was the same, and no significant difference was found between serum-free and serum-added groups, independently of the ECM ( Appendix 1 ), confirming that the medium additives themselves did not influence DRG outgrowth directly. Applying hPL only during the expansion steps of hADSC was sufficient to obtain a significant neurotrophic effect on neurons, which was consistent with our previous results ( Palombella et al, 2020 ).…”

“…Interestingly, this effect was only present when hPL was applied as the additive medium during the hADSC expansion steps. Importantly, keeping the hPL supplementation ongoing during the direct co-culture did not seem to further improve neurite outgrowth as we evaluated previously (Palombella et al, 2020).…”

“…Recently, we developed a completely animal protein-free method to extract, culture, and maintain in vitro hADSC, guaranteeing patient biosafety and potential for clinical translation ( Palombella et al, 2020 ).…”

“…As a medium additive, hPL provides a safe, human-derived product, which has been shown to increase the proliferation of hADSC, preserve their stemness properties and differentiation potential, and even enhance their neural commitment ( Guiotto et al, 2020 ; Lischer et al, 2020 ; Palombella et al, 2020 ). For all these reasons, hADSC pre-expanded in hPL (hADSC hPL ) are the ideal candidate for cell-based therapies and can play a role in nerve tissue engineering encouraging regeneration.…”

“…Therefore, human adipose-derived stem cells (hADSC) expanded in hPL-supplemented medium (hADSC hPL ) can be seen as a viable cell population for therapy in nerve defects, with translational applicability and neurotrophic potential ( Palombella et al, 2020 ).…”