ool for clinical use to avoid animal derived additives. Bieback and colleagues showed that only HPL supplemented medium, in con trast to human sera and platelet enriched plasma, is able to increase the growth rate of MSCs cultured in FBS supplemented medium. The high levels and num bers of platelet derived growth factors contained in HPL are likely to be responsible for the accelerated growth rate of MSCs that could act in synergy with each other. Many growth factors have been found in platelet con centrates. Among these, TGF b, PDGF, and FGF2 have been shown to be critical for both differen tiation and proliferation of MSCs. In addition, we assessed the cytokine secretion pro file of MSCs expanded in regular medium and in three HPL supplemented media. We used a multiplex bead based immunoassay array to simultaneously measure concentrations of seven cyto kines in MSC supernatants in comparison with corresponding media.
All MSC supernatants contained IL 6, IL 8, and VEGF at variable levels according to the different expansion culture conditions, whereas FGF2, G CSF, and GM CSF remained undetectable in supernatants in all culture conditions of MSCs. The presence of HPL increased IL 6 and IL 8 concentra tions, in MLN9708 solubility particular, at 10%. Variations of RANTES expression cannot be evaluated, because of the high level of this chemokine in HPL supplemented media, whereas consistent baseline levels were found in super natants of MSCs cultured in regular medium. In fact, the expression cytokine profile in MSCs cultured in regular medium is now well established. We previously reported by mRNA array that MSCs express a number of tyrosine kinase receptor cytokines, including VEGF, Ang 1, PDGFA, FGF1 and 2, EGF, IGF 1, and HGF, but also a number of CC and CXC chemokines, including RANTES, MCP 1, GRO alpha and beta, IL 8, and SDF 1.
Similar results were found in other studies at the protein level by large scale analysis. Although constitutive expression of the hema topoietic growth factors G CSF and GM CSF, has also been reported in other URB597 studies, these growth factors were not found in the present study. { This may reflect technical differences with lower sensitivity of the Bio Plex technique, as indicated by the lack of detection of cytokines in HPL containing media. Otherwise, we demonstrated here that secretion of both the pro inflammatory and hematopoietic cytokine IL 6 and the chemokine IL 8 was increased in MSCs previously expanded in HPL supplemented medium, as described elsewhere for IL 6. This indicates that HPL con tains strong inductors of such cytokines that could minimize the anti inflammatory activity of MSCs. Conclusions MSCs can be expanded in media supplemented with HPL that can totally replace FBS as well as FGF2. HPL represents an important t