Prediction of Sphingosine 1-Phosphate-Stimulated Endothelial Cell Migration Rates Using Biochemical Measurements

Abstract

The ability to predict endothelial cell migration rates may aid in the design of biomaterials that endothelialize following implantation. However, the complexity of the signaling response to migration-promoting stimuli such as sphingosine 1-phosphate (S1P) makes such predictions quite challenging. A number of signaling pathways impact S1P-mediated cell migration, including the Akt and Src pathways, which both affect activation of the small GTPase Rac. Rac activation promotes the formation of lamellipodia, and thus should be intimately linked to cell migration rates. In immortalized endothelial cells, expression of proteins that inhibit Akt, Src, and Rac (PTEN, CSK, and β2-chimaerin, respectively) was decreased using RNA interference, resulting in increases in the basal level of activation of Akt, Src, and Rac. Cells were scrape-wounded and stimulated with 1 μM S1P. The timecourse of Akt, Src, and Rac activation was followed over 2 h in the perturbed cells, while migration into the scrape wound was measured over 6 h. Rac activation at 120 min post-stimulation was highly correlated with the mean migration rate of cells, but only in cells stimulated with S1P. Using partial least squares regression, the migration rate of cells into the scrape wound was found to be highly correlated with the magnitude of the early Akt peak (e.g., 5–15 min post-stimulation). These results demonstrated that biochemical measurements might be useful in predicting rates of endothelial cell migration.

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Abbreviations

CSK:

C-terminal Src kinase

HAEC-hT:

Human aortic endothelial cells immortalized with hTERT

hTERT:

Human telomerase reverse transcriptase

PLSR:

Partial least squares regression

PTEN:

Phosphatase and homolog deleted on chromosome ten

S1P:

Sphingosine 1-phosphate

VIP:

Variable importance to projection

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Acknowledgments

We gratefully acknowledge funding from NIH, HL085364 (DLE), CA085839 (GDL), and GM080673 (GDL). We thank Sheila Stewart for technical guidance with hTERT tranformations, and Doug Lauffenburger for use of microscopy equipment.

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Correspondence to Donald L. Elbert.

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Associate Editor Sriram Neelamegham oversaw the review of this article.

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Alford, S.K., Wang, Y., Feng, Y. et al. Prediction of Sphingosine 1-Phosphate-Stimulated Endothelial Cell Migration Rates Using Biochemical Measurements. Ann Biomed Eng 38, 2775–2790 (2010). https://doi.org/10.1007/s10439-010-0014-6

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Keywords

  • Endothelial cell
  • Migration
  • Modeling
  • Partial least squares regression
  • Biochemistry