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Technical Briefs

A High Throughput System for Long Term Application of Intermittent Cyclic Hydrostatic Pressure on Cells in Culture

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Markus Rottmar, Sabine Ackerknecht, Peter Wick

Laboratory for Materials-Biology Interactions, Empa, Swiss Federal Laboratories for Materials Science and Technology, Lerchenfeldstrasse 5, CH-9014 St. Gallen, Switzerland

Katharina Maniura-Weber

Laboratory for Materials-Biology Interactions, Empa, Swiss Federal Laboratories for Materials Science and Technology, Lerchenfeldstrasse 5, CH-9014 St. Gallen, Switzerlandkatharina.maniura@empa.ch

J Biomech Eng 133(2), 024502 (Jan 31, 2011) (5 pages) doi:10.1115/1.4003313 History: Received March 24, 2010; Revised July 15, 2010; Posted December 22, 2010; Published January 31, 2011; Online January 31, 2011

The process of bone remodeling is governed by mechanical stresses and strains. Studies on the effects of mechanical stimulation on cell response are often difficult to compare as the nature of the stimuli and differences in parameters applied vary greatly. Experimental systems for the investigation of mechanical stimuli are mostly limited in throughput or flexibility and often the sum of several stimuli is applied. In this work, a flexible system that allows the investigation of cell response to isolated intermittent cyclic hydrostatic pressure (icHP) on a high throughput level is shown. Human bone derived cells were cultivated with or without mechanical stimulus in the presence or absence of chemical cues triggering osteogenesis for 7–10 days. Cell proliferation and osteogenic differentiation were evaluated by cell counting and immunohistochemical staining for bone alkaline phosphatase as well as collagen 1, respectively. In either medium, both cell proliferation and level of differentiation were increased when the cultures were mechanically stimulated. These initial results therefore qualify the present system for studies on the effects of isolated icHP on cell fate and encourage further investigations on the details behind the observed effects.

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Copyright © 2011 by American Society of Mechanical Engineers
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Figures

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Figure 1

An experimental system for the application of dynamic pressure. (a) Image of the system placed in a 37°C regulated climatic chamber. (b) Schematic of one of four chambers each harboring space for two cell culture plates (7) are driven by a crankshaft (3). An unbalance (8) above the chambers with the corresponding mechanics (4) converts the rotation into an up-and-down movement of a stamp (5) on a flexible membrane (2). The reduction in the volume within the incubating chambers is building up dynamic pressure stimulation. The system is continuously flooded (6) with humidified 5% CO2/95% air and cyclic pressure is computer controlled and monitored (1).

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Figure 2

Pressure and frequency range of the system. Optimal pressure and frequency range (left of trend line) determined by measuring the pressure difference with stamp amplitudes of 0.5 mm (square), 2 mm (diamond), and 3.9 mm (triangle) over the frequency range of the system.

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Figure 3

Representative pressure and frequency profiles. (a) Pressure profile with maximum (upper lines), minimum (lower lines), and average (middle lines) pressure and (b) frequency profile of a 24 h time frame with 30 min of stimulation followed by a 7 h 30 min break (gaps in graph).

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Figure 4

Cell proliferation of stimulated and unstimulated cells. Numbers of stimulated (+, filled bars) relative to unstimulated (−, structured bars) cells cultivated in expansion (black) or osteogenic (gray) medium. Significance p<0.05.

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Figure 5

Osteogenic differentiation of stimulated and unstimulated HBCs. HBCs stimulated (+) or unstimulated (−) for 10 days in osteogenic (O) or expansion (E) medium. Cells stained for ALP (upper row), collagen 1 (lower row), and the nuclei. (Scale bar 50 μm.)

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