Chitosan‐based scaffold counteracts hypertrophic and fibrotic markers in chondrogenic differentiated mesenchymal stromal cells

Abstract

Cartilage tissue engineering remains problematic because no systems are able to induce signals that contribute to native cartilage structure formation. Therefore, we tested the potentiality of gelatin‐polyethylene glycol scaffolds containing three different concentrations of chitosan (CH; 0%, 8%, and 16%) on chondrogenic differentiation of human platelet lysate‐expanded human bone marrow mesenchymal stromal cells (hBM‐MSCs). Typical chondrogenic (SOX9, collagen type 2, and aggrecan), hypertrophic (collagen type 10), and fibrotic (collagen type 1) markers were evaluated at gene and protein level at Days 1, 28, and 48. We demonstrated that 16% CH scaffold had the highest percentage of relaxation with the fastest relaxation rate. In particular, 16% CH scaffold, combined with chondrogenic factor TGFβ3, was more efficient in inducing hBM‐MSCs chondrogenic differentiation compared with 0% or 8% scaffolds. Collagen type 2, SOX9, and aggrecan showed the same expression in all scaffolds, whereas collagen types 10 and 1 markers were efficiently down‐modulated only in 16% CH. We demonstrated that using human platelet lysate chronically during hBM‐MSCs chondrogenic differentiation, the chondrogenic, hypertrophic, and fibrotic markers were significantly decreased. Our data demonstrate that only a high concentration of CH, combined with TGFβ3, creates an environment capable of guiding in vitro hBM‐MSCs towards a phenotypically stable chondrogenesis.

A Blood Bank Standardized Production of Human Platelet Lysate for Mesenchymal Stromal Cell Expansion: Proteomic Characterization and Biological Effects

“…Therefore, "hPL should be considered a product category and not a single product" in which it is important to distinguish "essential" from "non-essential" content based on the intended application (Henschler et al, 2019). We have previously reported that our hPL4c A18 batch does not induce hMSC chondrogenic differentiation when used alone, but it exerts an inhibitory effect when used in combination with a specific chondrogenic differentiation medium containing TGFβ3 (Manferdini et al, 2019). It has been reported that the composition of hPL can induce hMSC to release cytokines affecting their proliferative and differentiative potential (Mathews et al, 2014;Karlsen and Brinchmann, 2019).…”

Chitosan-Hydrogel Polymeric Scaffold Acts as an Independent Primary Inducer of Osteogenic Differentiation in Human Mesenchymal Stromal Cells

“…Similar results were obtained by aggrecan transcript quantification in AT-hMSC cultured in OM, even if with low statistical significance. The negative impact of hPL on chondrogenic differentiation of hMSCs cultured in 3D Hydrogel-chitosan scaffolds was previously described [34] and it is probably due to the presence of cell receptor-binding competitors within hPL. On the other hand, the quantified adipogenic marker presents the opposite behavior to the different stimuli: statistically significant differences were observed only in AT-hMSCs, as expected, while no differences came to light by FABP4 transcript quantification in BM-hMSCs.…”

“…Chitosan is a biomimetic, biodegradable, biocompatible, non-immunogenic, and versatile polymer presenting appropriate mechanical strength and anti-bacterial properties [ 16 ]. It is also able to support growth factor activity [ 32 , 34 ]. For these reasons, chitosan is used in tissue engineering, production of artificial organs, biotechnologies, and for the regeneration of damaged tissues [ 41 , 42 , 43 ].…”

“…The complete structural and mechanical characterization of the chitosan-based scaffold has been previously reported [32,34].…”

Impedance-Based Monitoring of Mesenchymal Stromal Cell Three-Dimensional Proliferation Using Aerosol Jet Printed Sensors: A Tissue Engineering Application

“…In particular, gelatin and chitosan have been extensively used for tissue engineering applications, since they are biocompatible, biodegradable, non-antigenic, non-toxic and present antimicrobial activity [6,7]. Starting from this evidence and aiming to regenerate bone, recent literature has provided examples of optimized combinations of these materials to realize hybrid hydrogel scaffolds, presenting a structure that mimics native ECM constituents and reporting morphological and mechanical properties suitable to support cell growth, osteo-differentiation and mineralization [8][9][10][11].…”