Decellularised extracellular matrices promote mesenchymal stem cell expansion and support osteogenlc differentiation

摘要

Aims: Mesenchymal stem cell (MSCs) therapies require extensive ex vivo cell expansion to obtain sufficient cell numbers. However, MSC expansion is limited to few passages (～5) after which cells senescence, spontaneously differentiate, and/or lose phenotype. Recent studies have shown that decellularized extracellular matrix (dECM) from fetal bone marrow-derived MSCs can maintain the phenotype of adult bone marrow-derived MSCs during extensive ex vivo expansion. However, the source material is very limited and encumbered by legal and ethical concerns. Placental MSCs share some properties of fetal MSC and are abundant and readily obtainable. The general hypothesis is that dECM derived from placental stem cells will provide a superior platform for the growth of MSCs and maintain of their sternness during prolonged culture. Methods: Placental MSC cultures were decellularised using PBS containing 0.5% Triton X-100 and 20mM NH4OH. Decellularised ECM (dECM) composition was assessed. Proliferation and differentiation properties of two types of MSC, decidua basalis MSCs (DMSCs) and chorionic villous MSCs (CMSCs), grown on the dECM-coated surface were compared with standard culture surfaces including artificial matrices (i.e. fibronectin) and tissue culture plastic (TCP). Results: Decellularization was confirmed by light microscopy and the absence of DAPI nuclear signal. DMSCs and CMSCs grown on dECM showed significantly increased cell numbers compared with controls (DMSCs: dECM; 1550 ± 370.7, fibronectin; 526.7 ± 32.45, TCP; 591.7 ± 89.5: CMSCs: ECM; 1091 ± 160.2, fibronectin; 576.7 ± 35.86, TCP; 538.3 ± 4.4, all figures are mean ± SEM, 1x103 cells). Osteogenic differentiation was quantified by hydroxyapatite/mineral deposition. After 14 days induction period, both DMSCs and CMSCs cultured on dECM deposited significantly more mineral compared to controls. Conclusions: These data indicate that dECM supports the growth of at least 2 different types of MSC; CMSCs and DMSCs, and also increases cell proliferation and enhances osteogenic differentiation. dECM substrates have the potential to support extensive expansion of MSCs, while maintaining sternness. This is an essential prerequisite for the preparation of MSCs for human stem cell therapies.