Bone Marrow Derived Connective Tissue Progenitor Cell Responses on Microtextured Substrates with Controlled Mechanical Cues

Abstract

The relationship between mechanical and topographical features of tissue engineering scaffolds and the likely response of human adult stem cells was investigated by a simple, yet powerful in vitro model, based on varying substrate stiffness with the precise and reproducible patterning capabilities of micro fabrication techniques. Polydimethylsiloxane (PDMS) pre-polymer and cross-linker were combined at various weight ratios designated as PDMS-a to PDMS-e, corresponding to 5.7, 10.0, 14.3, 21.4, and 42.9 wt. % cross-linker, respectively. PDMS microtextures with 10 μm diameter and 6 μm height microposts were produced using soft lithography and correlated to preferential human bone marrow derived connective tissue progenitor cells (CTPs) behavior as a function of varying stiffness. To investigate cell proliferation and osteogenic differentiation, CTPs were cultured for 30 days on a topographical map of substrates that combines 3 different types of PDMS microtextures and smooth PDMS. Elastic modulus, which is directly related to stiffness, increased from 0.78 ± 0.25 MPa (PDMS-a) to 2.83 ± 0.26 MPa (PDMS-c), and decreased down to 1.66 ± 0.18 MPa (PDMS-e). The cell number and gene expression levels were proportional to the PDMS stiffness, and PDMS microtextures exhibited greater numbers of CTPs compared to smooth PDMS. Alkaline phosphatase expressed greater on post microtextures than smooth surfaces on early days. Regardless of surface topographies, however, cells on PDMS-b consistently expressed more osteocalcin compared on other substrates on day 30. These results indicate that CTP proliferation and early osteogenic differentiation are more likely to be affected by surface microtextures, while substrate stiffness is more likely to influence the late osteogenic differentiation.