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The present paper is related to the field of cell culture medium supplements. In particular, it refers to (i) an heparin-free human Platelet Lysate devoid of serum or plasma components (v-PL) and (ii) an heparin-free human serum devoid of platelet lysate components (Pl-s), and to their use as single components or in combination in primary or cell line cultures. Human Mesenchymal Stem Cells (MSC) primary cultures were obtained from adipose, bonemarrow and umbilical cord. The cumulative cell doubling number in the presence of the different culture medium supplements was determined at different culture times. In general, MSC expanded in the presence of Pl-s alone showed a low or no proliferation in comparison to cells grown with the combination of Pl-s and v-PL. Confluent, growth arrested cells, either human MSC or human articular chondrocytes, treated with v-PL resumed proliferation, whereas control cultures, not supplemented with v-PL, remained quiescent and did not proliferate. Interestingly, signal transduction pathways distinctive of proliferation were activated also in cells treated with v-PL in the absence of serum, when cell proliferation did not occur, indicating that v-PL could induce the cell re-entry in the cell cycle (cell commitment), but the presence of serum proteins was an indispensable requirement for cell proliferation to happen. Indeed, Pl-s alone supported cell growth in constitutively activated cell lines (U-937, HeLa, HaCaT, V-79) regardless the co-presence of v- PL. Plasma and plasma-derived serum were equally able to sustain cell proliferation although, for cells cultured in adhesion, the Pl-s was more efficient than the plasma from which it was derived. Moreover, cell expanded in the presence of the new additives maintained their differentiation potential and did not show alterations in their karyotype. In a different work we showed how Platelet Lysate (PL) derived from Platelet Rich Plasma (PRP) was capable to support growth and isolation of MSC (Mesenchymal Stromal Cell). However, the molecular mechanisms involved in these processes were still unknown. We focused our attention on a family of highly conserved proteins involved in different cellular mechanisms such as cell growth, proliferation and apoptosis, MYC's family. In particular, MYC is a transcription factor that activates or inhibits the transcription of several genes by interacting with other proteins. The three proteins of C-MYC gene have the same c-terminal part, differing in the N-terminal region due to an alternative translation at the start site. The three forms are C-MYC1, C-MYC2 and CMYCS, differently expressed during cell growth.

Chronic skin ulcers, consequence of diabetes and other pathological conditions, heavily compromise the patient life quality and represent a high and constantly growing cost for National Health Services. Autologous platelet-rich plasma (PRP), has been proposed to treat these lesions. The absence of guidelines for the PRP production and the need of a fresh preparation for each treatment lead us to develop a protocol for the production of an allogenic PRP-based bioactive membrane (BAM), standardized for platelet concentration and growth factor release. This work compares BAMs obtained starting from two different platelet concentrations. There was no direct correlation between the amount of growth factors released by BAM in vitro and the initial platelet count. However, different release kinetics were noticed for different growth factors, suggesting that they were differently retained by the two BAMs. The angiogenic potential of both BAMs was determined by Luminex Angiogenesis Assay. The biological activity of the factors released by the two BAMs was confirmed by cell proliferation and migration. A diabetic mouse chronic ulcer model was used to define the best PRP therapeutic dose in vivo. Both BAMs induced wound healing by increasing the thickness of the regenerated epidermis and the vessel number. However, a too high platelet concentration resulted in a slowdown of the membrane resorption that interfered with the skin healing. Overall, the results indicate that the BAMs could represent a natural and effective wound healing tool for the treatment of skin ulcers. Copyright © 2016 John WileySons, Ltd.