Perfusion Culture Enhances Osteogenic Differentiation of Rat Mesenchymal Stem Cells in Collagen Sponge Reinforced with Poly(Glycolic Acid) Fiber

Abstract

The objective of this study was to obtain fundamental knowledge about in vitro culture systems to enhance the proliferation and differentiation of mesenchymal stem cells (MSCs) in collagen sponge reinforced by the incorporation of poly(glycolic acid) (PGA) fiber. A collagen solution with PGA fiber homogeneously localized at PGA:collagen weight ratios of 0.67, 1.25, 2.5, and 5 was freezedried, followed by cross-linking of combined dehydrothermal, glutaraldehyde, and ultraviolet treatment. Scanning electron microscopy revealed that collagen sponges exhibited homogeneous and interconnected pore structures with an average size of 180 microm, irrespective of PGA fiber incorporation. When rat MSCs were seeded into collagen sponge with or without PGA fiber incorporation, more attached cells were observed in collagen sponge incorporating PGA fiber than in collagen sponge without PGA fiber incorporation, irrespective of the PGA:collagen ratio. The proliferation and osteogenic differentiation of MSCs in PGA-reinforced sponge at a weight ratio of 5 were greatly influenced by the culture method and growth conditions. Alkaline phosphatase (ALP) activity and osteocalcin content of MSCs cultured in PGA-reinforced sponge by the perfusion method became maximum at a flow rate of 0.2 mL/min, although they increased with culture time period. It may be concluded that appropriate perfusion conditions enable MSCs to positively improve the extent of proliferation and differentiation.

Development of 3D in vitro platform technology to engineer mesenchymal stem cells

“…In comparison with conventional culture, the environment of cells maintained in the 3D culture scaffold is closer to that in vivo, which can affect the gene expression and behavior of cells. 32,33 The level of BMP-2 protein was enhanced by mixing plasmid DNA with dextran-spermine nanoparticles and incorporating the nanoparticles into the PGA/collagen nanofiber sheets. It is highly conceivable that the electrostatic interaction between the negatively charged cell membrane and the positively charged nanoparticles is a driving force in attaching nanoparticles to cells, which is an important initial step for gene transfection.…”

Nanoporous Titania Coating of Microwell Chips for Stem Cell Culture and Analysis

“…The collagen was prepared from porcine tendon by pepsin treatment in HCl (3 mg/ml, pH 3.0) and was supplied by Nitta Gelatin Inc., Osaka, Japan. Mesenchymal stem cells (MSC) were isolated from the femurs of 3-week-old male Wistar rats [16]. The bone marrow was flushed out using Dulbecco's modified Eagle's minimal essential medium (DMEM) supplemented with 15% fetal calf serum (FCS) and 50 U/ml penicillin and streptomycin.…”

A driving force for change: interstitial flow as a morphoregulator

“…Morphogen transport and gradient patterning are also believed to regulate the branching of developing lungs in the embryo [14], and it is expected that interstitial flows, caused by embryonic lung movements that simulate breathing, would influence morphogen distributions. Interstitial flow can also impart shear stress on the cell surface, which, as a mechanical stimulus, can itself drive embryonic cell differentiation and determine lineage fate [15,16] and could be responsible for shaping organs [17,18].…”