Strain-time cell-death threshold for skeletal muscle in a tissue-engineered model system for deep tissue injury

Amit Gefen, Bastiaan van Nierop, Dan L. Bader, Cees W. Oomens

Research output: Contribution to journalArticlepeer-review


Deep tissue injury (DTI) is a severe pressure ulcer that results from sustained deformation of muscle tissue overlying bony prominences. In order to understand the etiology of DTI, it is essential to determine the tolerance of muscle cells to large mechanical strains. In this study, a new experimental method of determining the time-dependent critical compressive strains for necrotic cell death (Ezzc (t)) in a planar tissue-engineered construct under static loading was developed. A half-spherical indentor is used to induce a non-uniform, concentric distribution of strains in the construct, and Ezzc (t) is calculated from the radius of the damage region in the construct versus time. The method was employed to obtain Ezzc (t) for bio-artificial muscles (BAMs) cultured from C2C12 murine cells, as a model system for DTI. Specifically, propidium iodine was used to fluorescently stain the development of necrosis in BAMs subjected to strains up to 80%. Two groups of BAMs were tested at an extracellular pH of 7.4 (n=10) and pH 6.5 (n=5). The lowest strain levels causing cell death in the BAMs were determined every 15 min, during 285-min-long trials, from confocal microscopy fluorescent images of the size of the damage regions. The experimental Ezzc (t) data fitted a decreasing single-step sigmoid of the Boltzmann type. Analysis of the parameters of this sigmoid function indicated a 95% likelihood that cells could tolerate engineering strains below 65% for 1 h, whereas the cells could endure strains below 40% over a 285 min trial period. The decrease in endurance of the cells to compressive strains occurred between 1-3 h post-loading. The method developed in this paper is generic and suitable for studying Ezzc(t) in virtually any planar tissue-engineered construct. The specific Ezzc (t) curve obtained herein is necessary for extrapolating biological damage from muscle-strain data in biomechanical studies of pressure ulcers and DTI.

Original languageEnglish
Pages (from-to)2003-2012
Number of pages10
JournalJournal of Biomechanics
Issue number9
StatePublished - 2008
Externally publishedYes


  • Acidosis
  • Deformation damage
  • Etiology
  • In vitro model
  • Pressure ulcer


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