Nanotopography Driven Mesenchymal Stem Cell Differentiation and Proliferation

Abstract

Plastic-adherent, fibroblast-like populations of bone marrow cells, termed multipotent mesenchymal stromal/stem cells (MSCs), constitute a tissue specific stem cell population that is responsible for tissue homeostasis and repair of bone, cartilage and adipose tissue. Previous work with MSCs had shown that they are capable of differentiation into specific lineages in response to the physical properties of the extracellular matrix such as matrix elasticity and cell geometry. Remarkably MSCs have been demonstrated to differentiate into bone-forming osteoblast cells by modifying the substrate nanotopography. In this work, the role of nanotopography on the long-term response of MSCs is explored using biodegradeable polycaprolactone (PCL) nanopillar and nanofiber surfaces seeded with rat MSCs and cultured in normal growth media for four weeks. We found that after four weeks in culture under normal expansion media conditions, MSCs cultured on nanofibers exhibit better adherence, significantly increased proliferation, and maintain increasingly dense fibroblast-like morphologies. In contrast, MSCs seeded on nanopillar surfaces display lowered adherence, reduced proliferation, and adopt highly elongated cellular morphologies. Cell shape and area quantification reveals that MSCs cultured on smooth substrates adopt uniformly spread morphologies covering large surface areas, while MSCs cultured on nanopillars have significantly larger length-to-width ratios and significantly reduced coverage areas. Immunofluorescent staining of MSCs on PCL nanopillars reveals the presence of two bone marker proteins, osteopontin and osteocalcin, providing evidence for differentiation into osteoblast-like cells. Unlike the nanopillar topography, MSCs cultured on nanofiber and smooth PCL surfaces did not appear to undergo surface induced osteogenesis. We put forward a mechanotransductive model to interpret these and other's findings.