Laurent Jeannin

Synthetic peptide matrices as support for stem cells culture

Background Nowadays, stem cells draw great interest in regenerative medicine [1]. However, due to low occurrence in tissues, their in vitro expansion is required to gain access to therapeutic applications [2]. Furthermore, the existing culture systems often include xeno-derived components, present upscaling limitations [3,4] and do not necessary address important considerations, such as those related to biosafety, availability and homogeneity of (biological) raw material and process reproducibility. Moreover, the regulatory guidelines focus on the importance of defined environments, minimal handling and continuity between development and industrialization scales [5,6]. In this context, new scalable solutions a reactively considered in order to achieve production of stem cells at higher densities [7]. Among these, synthetic peptides are of particular interest [4] as potential alternatives to xeno-derived culture supports, able to mimic the physico-biochemical properties of natural cell matrices. In this way our work aims at assessing new xeno-free culture matrices based on short, synthetic and bioactive peptides and demonstrating the scalability, reproducibility and performance of such peptides-based matrices for human stem cells culture. This report is focused on a part of the work performed to study the stability of synthetic peptide coatings on polymer surfaces and the capability of such coatings to favour stem cells adhesion and growth.

New peptide-based and animal-free coatings for animal cell culture in bioreactors

Background Anchorage dependent cells require an appropriate extracellular matrix for their survival, migration, proliferation, phenotyping and/or differentiation [1-3]. These cells interact with extracellular matrix proteins, primarily through integrins, which induces focal adhesion contacts assembly and activation of signalling pathways that regulate diverse cellular processes [4]. Culture supports usually include biochemical components allowing such cells to adhere and to reconstitute an extracellular environment close to that found in vivo. Currently, this artificial environment is achieved by extracellular matrix constituents deposition, adsorption or grafting; among them collagens, fibronectin, laminin, artificial lamina propria... [5]. However, such animal proteins used in cell culture may induce pro-inflammatory stress, be unstable against proteolysis or loose activity after adsorption [6,7]. Synthetic microenvironments should be more suitable for clinical purposes: (i) improved control of physicochemical and mechanical properties, (ii) limited risks of immunogenicity, (iii) increased biosafety (animal free) and (iv) facilitated scale-up [1]. In this framework, research has recently focused on synthetic peptides or peptidomimetics that can mimic the extracellular matrix. Such molecules can be immobilized as recognition motifs on the surface of culture supports with a greater stability and easier surfaces characterization [5]. Self-assembling peptide hydrogels could mimic the chemical and mechanical aspects of the natural extracellular matrix [8,9] by undergoing large deformations, as in mammalian tissues. They have an inherent biocompatibility and should be able to direct cell behaviour [10]. They also can be functionalized with various biologically active ligands constituting good candidates to a new range of smart biomaterials, able to ensure adhesion of different cell types [11-13]. The range of biomimetic peptides that direct cell adhesion and are recognized by integrins is large. Recognition sequences derived from different extracellular matrix proteins include RGD [1], which are specific to different cell lines [1,5,6]. In this context, this work aims at designing animal-free, chemically defined and industrially scalable coatings for animal cell culture, as an alternative to collagen, fibronectin or Matrigel for laboratory and industrial large scale applications. These are based on self-assembling short peptides bearing adhesion bioactive sequences like RGDderived or other adhesion sequences developed to coat polystyrene or polyethylene terephthalate surfaces. Adhesion sequences should be recognized by cells, which should favour their anchorage and spreading.