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.

Bone Marrow-Derived Cell Concentrates Have Limited Effects on Osteochondral Reconstructions in the Mini Pig

“…The medium was changed every 3 days. In a previous study with the same culture media, 23 the fibroblast nature of these cells has been demonstrated using antibodies for collagen I and III and by the up-regulation of tenascin-C. Fibroblast colonies were Giemsa stained and counted using an …”

Indirect co-culture with tenocytes promotes proliferation and mRNA expression of tendon/ligament related genes in rat bone marrow mesenchymal stem cells

“…JuncosaMelvin et al further showed that mechanical stimulation of MSCs-collagen sponges constructs significantly improved tendon repair in rabbit model (Juncosa-Melvin et al 2006). In vitro, MSCs have been successfully induced into tenocytes by exposure to growth factors BMP-12 (Violini et al 2009) or low-dose FGF-2 (Hankemeier et al 2005). Meanwhile, GDF-5 has also been reported to increase mRNA expression of Col I and SCX in MSCs which are believed to be the markers of tendon/ligament differentiation (Farng et al 2008).…”

Regenerative Medicine for Tendon Regeneration and Repair: The Role of Bioscaffolds and Mechanical Loading

“…(Yao L et al 2006) (Cao D et al 2006) Anyway the use of tenocytes have some drawbacks such as limited availability of donor sites for cell harvest, the requirement for lengthy in vitro culture to expand the number of cells, and donor-site morbidity limit the practicality of this technique. (Hankemeier S et al 2005) (Awad HA et al 2000) Stem cells may represent the ideal source for tendon engineering.There are 2 types of stem cells: embryonic stem cells and adult stem cells, embryonic cells are totipotent, but their practical use may be limited because of ethical issues and concerns regarding cell regulation. Adult stem cells, also known as mesenchymal stem cells, show excellent regenerative capacity, the ability to proliferate rapidly in culture, and the ability to differentiate into a wide variety of cell types.…”

“…(Hankemeier S et al 2005) ) (Abrahamsson SO et al 1991) (Rickert M et al 2001) (Wang XT 2004) (Hsu C & Chang J 2004). The results of in vitro studies have shown that growth factors can promote cell proliferation and protein synthesis (Fu SC et al 2003) (Jann HW et all 1999)Injured tendons treated with growth factors show improved mechanical properties.…”