Modulation of proliferation and differentiation of human bone marrow stromal cells by fibroblast growth factor 2: potential implications for tissue engineering of tendons and ligaments

Abstract

Bone marrow stromal cells (BMSCs) play a central role in the repair and regeneration of mesenchymal tissues. For tissue engineering of ligaments and tendons, both stimulation of cell proliferation and differentiation with increased expression of essential extracellular matrix proteins and cytoskeletal elements are desirable. This study analyzes the effect of low-dose (3 ng/mL) fibroblast growth factor 2 (FGF-2) and high-dose FGF-2 (30 ng/mL) on proliferation (bromodeoxyuridine content, spectrophotometry), differentiation (transcription of collagen I, collagen III, fibronectin, elastin, alpha-smooth muscle actin, and vimentin, reverse transcription-polymerase chain reaction, and cell density and apoptosis (annexin V, fluorescence-activated cell sorting) of human BMSCs, and compares the results with those of a control group without FGF-2. Low-dose FGF-2 triggered a biphasic BMSC response: on day 7, cell proliferation reached its maximum and was significantly higher compared with the other groups. On days 14 or 28, collagen I, collagen III, fibronectin, and alpha- smooth muscle actin mRNA expression was significantly enhanced in the presence of low-dose FGF-2. In contrast, high-dose FGF-2 did not stimulate differentiation or proliferation. Vimentin mRNA was expressed only in cultures with low-dose and high-dose FGF-2 after 14 and 28 days. Cell density was significantly higher in cultures with low-dose FGF-2 compared with the group with high-dose FGF-2 on days 7, 14, and 28. The apoptosis rate remained stable, at a rather high level, in all groups. Microscopic investigation of the cell cultures with low-dose FGF-2 showed more homogeneous, dense, fibroblast-like, spindle-shaped cells with long cell processes compared with cultures with high-dose, or no FGF-2. Low-dose FGF-2 may be useful for tissue engineering of ligaments and tendons by increasing BMSC proliferation and stimulating mRNA expression of specific extracellular matrix proteins and cytoskeletal elements.