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Effects of Intermittent and Incremental Cyclic Stretch on ERK Signaling and Collagen Production in Engineered Tissue


Intermittent cyclic stretching and incrementally increasing strain amplitude cyclic stretching were explored to overcome the reported adaptation of fibroblasts in response to constant amplitude cyclic stretching, with the goals of accelerating collagen production and understanding the underlying cell signaling. The effects of constant amplitude, intermittent, and incremental cyclic stretching regimens were investigated for dermal fibroblasts entrapped in a fibrin gel by monitoring the extracellular signal-regulated kinase (ERK1/2) and p38 pathways, collagen transcription, and finally the deposited collagen protein. Activation of ERK1/2, which has been shown to be necessary for stretch-induced collagen transcription, was maximal at 15 min and decayed by 1 h. ERK1/2 was reactivated by an additional onset of stretching or by an increment in the strain amplitude 6 h after the initial stimulus, which was approximately the lifetime of activated p38, a known ERK1/2 inhibitor. While both intermittent and incremental regimens reactivated ERK1/2, only incremental stretching increased collagen production compared to samples stretched with constant amplitude, resulting in a 37% increase in collagen per cell after 2 weeks. This suggests that a regimen with small, frequent increments in strain amplitude is optimal for this system and should be used in bioreactors for engineered tissues requiring high collagen content.

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Analysis of variance


Collagen Iα1/collagen IIIα1




Extracellular signal-regulated kinase 1/2


Mitogen-activated protein kinase


Phosphate buffered saline


Sodium dodecyl sulfate polyacrylamide gel electrophoresis


Tris-buffered saline


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The authors thank Naomi Ferguson, Sandra Johnson, Jay Reimer, Dr. Colleen Witzenburg, and Dr. M. Cristine Charlesworth for technical assistance and Kiley Schmidt for providing illustrations. Nanoimmunoassay was performed in the Mayo Clinic Proteomics Core. This study was funded by a National Science Foundation Graduate Research Fellowship (to J.B.S) and National Institutes of Health/National Heart, Lung, and Blood Institute award HL107572 (to R.T.T.).

Conflict of interest

Jillian B. Schmidt, Kelley Chen, and Robert T. Tranquillo declare that they have no conflicts of interest.

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No human or animal studies were carried out by the authors for this article.

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Correspondence to Robert T. Tranquillo.

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Associate Editor Michael R. King oversaw the review of this article.

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Supplemental Fig. 1 Chemiluminescence intensity vs. isoelectric point for static (blue) and 5% continuously stretched (green) samples at 15 min. Peak identification was validated with HeLa control lysates and isoform specific antibodies. Supplementary material 1 (TIFF 1910 kb)


Supplemental Fig. 2 Representative Western blots for phosphorylated and total p38 in static and continuously stretched (5%) samples at 15 min, 1, 3, and 6 h. Supplementary material 2 (TIFF 4576 kb)

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Schmidt, J.B., Chen, K. & Tranquillo, R.T. Effects of Intermittent and Incremental Cyclic Stretch on ERK Signaling and Collagen Production in Engineered Tissue. Cel. Mol. Bioeng. 9, 55–64 (2016).

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  • Fibrin
  • Fibroblast
  • Mitogen activated protein kinase
  • p38
  • Mechanical conditioning