Effects of static or dynamic mechanical stresses on osteoblast phenotype expression in three-dimensional contractile collagen gels

Abstract

Studies performed at tissular (three-dimensional, 3-D) or cellular (two-dimensional, 2-D) levels showed that the loading pattern plays a crucial role in the osteoblastic physiology. In this study, we attempted to investigate the response of a 3-D osteoblastic culture submitted to either no external stress or static or dynamic stresses. Rat osteosarcoma cells (ROS 17/2.8) were embedded within collagen type I lattices and studied for 3 weeks. Entrapment and proliferation of cells within the hydrated collagen gel resulted in the generation of contractile forces, which led to contraction of the collagen gel. We used this ability to evaluate the influence of three modes of mechanical stresses on the cell proliferation and differentiation: (1) the freely retracted gels (FRG) were floating in the medium, (2) the tense gels (TG) were stretched statically and isometrically, with contraction prevented in the longitudinal axis, and (3) the dynamic gels (DG) were floating gels submitted to periodic stresses (50 or 25 rpm frequency). Gels showed maximum contraction at day 12 in 50 rpm DG, followed by 25 rpm DG, then FRG (88%, 81%, 70%, respectively) and at day 16 in TG (33%). The proliferation rate was greater in TG than in FRG (ϩ52%) but remained low in both DGs. Gel dimensions were related to the collagen concentration and on a minor extent to cell number. Cells in DG appeared rounder and larger than in other conditions. In TG, cells were elongated and oriented primarily along the tension axis. Scanning electron microscopy (SEM) showed that tension exerted by cells in TG led to reorientation of collagen fibers which, in turn, determined the spatial orientation and morphology of the cells. Transmission electron microscopy (TEM) performed at maximum proliferation showed a vast majority of cells with a distended well-developed RER filled with granular material and numerous mitochondria. Alkaline phosphatase activity peaked close to the proliferation peak in FRG, whereas in TG, a biphasic curve was observed with a small peak at day 4 and the main peak at day 16. In DG, this activity was lower than in the two other conditions. A similar time course was observed for alkaline phosphatase gene expression as assessed by Northern blots. Regardless of the conditions, osteocalcin level showed a triphasic pattern: a first increase at day 2, followed by a decrease from day 4 to 14, and a second increase above initial values at day 18. Microanalysis-x indicated that mineralization occurred after 14 days and TEM showed crystals within the matrix. We showed that static and dynamic mechanical stresses, in concert with 3-D collagen matrices, played a significant role on the phenotypic modulation of osteoblast-like cells. This experimental model provided a tool to investigate the significance and the mechanisms of mechanical activity of the 3-D cultured osteoblast-like cells. J. Cell. Biochem. 76:217-230, 1999. 1999 Key words: ROS 17/2.8; type I collagen; ultrastructure; proliferation; alkaline phosphatase; osteocalcin; mineralization...

Phenolic Hydroxy Groups Incorporated for the Peroxidase‐Catalyzed Gelation of a Carboxymethylcellulose Support: Cellular Adhesion and Proliferation

“…[21] In addition, the mechanical strength of hydrogels has been reported to affect the growth and function of enclosed mammalian cells. [22,23] Figure 3 shows stress/strain profiles of CMC-Ph gels prepared from 1.0 and 2.0 wt.-% solutions of CMC-Ph8 and CMC-Ph15. The stress/strain profiles of the gels were similar before rupture of the CMCPh15 gel.…”

Influences of hyaluronan on type II collagen fibrillogenesis in vitro

“…Recent studies have showed that collagen fibril morphology is critically important in determining the tensile strength of the associated tissues and, thus, provides a scaffold for the cell adhesion [5,6]. Some experimental data showed that the collagen morphological structures could affect the adhering, growth and differentiation of a variety of cells in cell culture applications [7,8].…”

Chondrocyte intracellular calcium, cytoskeletal organization, and gene expression responses to dynamic osmotic loading

“…While static loading conditions facilitate experimental investigation, the natural physiological environment is dynamic and changes with time. In fact, studies have demonstrated that many cell types respond differently depending on whether a given mechanical stimulus (e.g., stretch, flow) is applied in a static or dynamic manner (1,4,12,14,36,40,44,46,58). For instance, while static deformation causes a reduction in chondrocyte biosynthetic activities, dynamic deformational loading results in an upregulation of chondrocyte biosynthetic activity (6,49).…”