Development of a Platform for Studying 3D Astrocyte Mechanobiology: Compression of Astrocytes in Collagen Gels


Glaucoma is a common optic neuropathy characterized by retinal ganglion cell death. Elevated intraocular pressure (IOP), a key risk factor for glaucoma, leads to significant biomechanical deformation of optic nerve head (ONH) cells and tissues. ONH astrocytes respond to this deformation by transforming to a reactive, proliferative phenotype, which has been implicated in the progression of glaucomatous vision loss. However, little is known about the mechanisms of this transformation. In this study, we developed a 3D collagen gel culture system to mimic features of ONH deformation due to elevated IOP. Compressive loading of astrocyte-seeded collagen gels led to cell alignment perpendicular to the direction of strain, and increased astrocyte activation, as assayed by GFAP, vimentin, and s100β levels, as well as MMP activity. This proof-of-concept study shows that this system has potential for studying mechanisms of astrocyte mechanobiology as related to the pathogenesis of glaucoma. Further work is needed to establish the possible interplay of mechanical stimulation, matrix properties, and hypoxia on the observed response of astrocytes.

This is a preview of subscription content, access via your institution.

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5


  1. 1.

    Albon, J., W. S. Karwatowski, N. Avery, D. L. Easty, and V. C. Duance. Changes in the collagenous matrix of the aging human lamina cribrosa. Br. J. Ophthalmol. 79:368–375, 1995.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Anderson, D. R. Collaborative normal tension glaucoma study. Curr. Opin. Ophthalmol. 14:86–90, 2003.

    Article  PubMed  Google Scholar 

  3. 3.

    Anderson, D. R., and A. Hendrickson. Effect of intraocular pressure on rapid axoplasmic transport in monkey optic nerve. Investig. Ophthalmol. Vis. Sci. 13:771–783, 1974.

    CAS  Google Scholar 

  4. 4.

    Balasubramanian, S., J. A. Packard, J. B. Leach, and E. M. Powell. Three-dimensional environment sustains morphological heterogeneity and promotes phenotypic progression during astrocyte development. Tissue Eng. Part A 22:885–898, 2016.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  5. 5.

    Bell, B. J., E. Nauman, and S. L. Voytik-Harbin. Multiscale strain analysis of tissue equivalents using a custom-designed biaxial testing device. Biophys. J. 102:1303–1312, 2012.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Biran, R., M. D. Noble, and P. A. Tresco. Directed nerve outgrowth is enhanced by engineered glial substrates. Exp. Neurol. 184:141–152, 2003.

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Bott, K., Z. Upton, K. Schrobback, M. Ehrbar, J. A. Hubbell, M. P. Lutolf, and S. C. Rizzi. The effect of matrix characteristics on fibroblast proliferation in 3D gels. Biomaterials 31:8454–8464, 2010.

    CAS  Article  PubMed  Google Scholar 

  8. 8.

    Burgoyne, C. F., J. C. Downs, A. J. Bellezza, J.-K. F. Suh, and R. T. Hart. The optic nerve head as a biomechanical structure: a new paradigm for understanding the role of IOP-related stress and strain in the pathophysiology of glaucomatous optic nerve head damage. Prog. Retin. Eye Res. 24:39–73, 2005.

    Article  PubMed  Google Scholar 

  9. 9.

    Chen, B., and M. O. Platt. Multiple× zymography captures stage-specific activity profiles of cathepsins K, L, and S in human breast, lung, and cervical cancer. J. Transl. Med. 9:109, 2011.

    Article  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Chong, R. S., and K. R. Martin. Glial cell interactions and glaucoma. Curr. Opin. Ophthalmol. 26:73–77, 2015.

    Article  PubMed  PubMed Central  Google Scholar 

  11. 11.

    Coudrillier, B., D. Geraldes, N. Vo, R. Atwood, C. Reinhard, I. Campbell, Y. Raji, J. Albon, R. Abel, and R. Ethier. Phase-contrast micro-computed tomography measurements of the intraocular pressure-induced deformation of the porcine lamina cribrosa. IEEE Trans. Med. Imaging 35:988–999, 2015.

    Article  PubMed  Google Scholar 

  12. 12.

    De Hoz, R., B. Rojas, A. I. Ramírez, J. J. Salazar, B. I. Gallego, A. Triviño, and J. M. Ramírez. Retinal macroglial responses in health and disease. Biomed. Res. Int. 2016:2954721, 2016.

    PubMed  PubMed Central  Google Scholar 

  13. 13.

    Downs, J. C., C. F. Burgoyne, W. P. Seigfreid, J. F. Reynaud, N. G. Strouthidis, and V. Sallee. 24-hour IOP telemetry in the nonhuman primate: implant system performance and initial characterization of IOP at multiple timescales. Investig. Ophthalmol. Vis. Sci. 52:7365–7375, 2011.

    Article  Google Scholar 

  14. 14.

    Downs, J. C., M. D. Roberts, and C. F. Burgoyne. Mechanical environment of the optic nerve head in glaucoma. Optom. Vis. Sci. 85:425–435, 2008.

    Article  PubMed  PubMed Central  Google Scholar 

  15. 15.

    East, E., J. P. Golding, and J. B. Phillips. A versatile 3D culture model facilitates monitoring of astrocytes undergoing reactive gliosis. J. Tissue Eng. Regen. Med. 3:634–646, 2009.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Ellis, E. F., J. S. McKinney, K. A. Willoughby, S. Liang, and J. T. Povlishock. A new model for rapid stretch-induced injury of cells in culture: characterization of the model using astrocytes. J. Neurotrauma 12:325–339, 1995.

    CAS  Article  PubMed  Google Scholar 

  17. 17.

    Exler, R. E., X. Guo, D. Chan, I. Livne-Bar, N. Vicic, J. G. Flanagan, and J. M. Sivak. Biomechanical insult switches PEA-15 activity to uncouple its anti-apoptotic function and promote erk mediated tissue remodeling. Exp. Cell Res. 340:283–294, 2016.

    CAS  Article  PubMed  Google Scholar 

  18. 18.

    Gillette, B. M., J. A. Jensen, M. Wang, J. Tchao, and S. K. Sia. Dynamic hydrogels: switching of 3D microenvironments using two-component naturally derived extracellular matrices. Adv. Mater. Weinh. 22:686–691, 2010.

    CAS  Article  Google Scholar 

  19. 19.

    Heijl, A., M. C. Leske, B. Bengtsson, L. Hyman, B. Bengtsson, M. Hussein, and Early Manifest Glaucoma Trial Group. Reduction of intraocular pressure and glaucoma progression: results from the early manifest glaucoma trial. Arch. Ophthalmol. 120:1268–1279, 2002.

    Article  PubMed  Google Scholar 

  20. 20.

    Hernandez, M. R. The optic nerve head in glaucoma: role of astrocytes in tissue remodeling. Prog. Retin. Eye Res. 19:297–321, 2000.

    CAS  Article  PubMed  Google Scholar 

  21. 21.

    Hernandez, M. R., H. Miao, and T. Lukas. Astrocytes in glaucomatous optic neuropathy. In: Glaucoma: An Open Window to Neurodegeneration and Neuroprotection, Amsterdam: Elsevier, 2008, pp. 353–373.

    Google Scholar 

  22. 22.

    Jaworski, J., and C. M. Klapperich. Fibroblast remodeling activity at two- and three-dimensional collagen-glycosaminoglycan interfaces. Biomaterials 27:4212–4220, 2006.

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Johnson, E. C., and J. C. Morrison. Friend or foe? Resolving the impact of glial responses in glaucoma. J. Glaucoma 18:341–353, 2009.

    Article  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Lam, D. S. C., J. K. H. Chua, C. C. Y. Tham, and J. S. M. Lai. Efficacy and safety of immediate anterior chamber paracentesis in the treatment of acute primary angle-closure glaucoma: a pilot study. Ophthalmology 109:64–70, 2002.

    Article  PubMed  Google Scholar 

  25. 25.

    Lye-Barthel, M., D. Sun, and T. C. Jakobs. Morphology of astrocytes in a glaucomatous optic nerve. Invest. Ophthalmol. Vis. Sci. 54:909–917, 2013.

    Article  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Marshak, D. R. S100 beta as a neurotrophic factor. Prog. Brain Res. 86:169–181, 1990.

    CAS  Article  PubMed  Google Scholar 

  27. 27.

    Morgan, J. E. Optic nerve head structure in glaucoma: astrocytes as mediators of axonal damage. Eye (Lond) 14(Pt 3B):437–444, 2000.

    Article  Google Scholar 

  28. 28.

    Morrison, J. C., W. O. Cepurna, S. Tehrani, T. E. Choe, H. Jayaram, D. C. Lozano, B. Fortune, and E. C. Johnson. A period of controlled elevation of IOP (CEI) produces the specific gene expression responses and focal injury pattern of experimental rat glaucoma. Invest. Ophthalmol. Vis. Sci. 57:6700–6711, 2016.

    Article  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Munemasa, Y., and Y. Kitaoka. Molecular mechanisms of retinal ganglion cell degeneration in glaucoma and future prospects for cell body and axonal protection. Front. Cell. Neurosci. 6:60, 2012.

    PubMed  Google Scholar 

  30. 30.

    Neufeld, A. H., and B. Liu. Glaucomatous optic neuropathy: when glia misbehave. Neuroscientist 9:485–495, 2003.

    CAS  Article  PubMed  Google Scholar 

  31. 31.

    Ng, C. P., B. Hinz, and M. A. Swartz. Interstitial fluid flow induces myofibroblast differentiation and collagen alignment in vitro. J. Cell Sci. 118:4731–4739, 2005.

    CAS  Article  PubMed  Google Scholar 

  32. 32.

    Nguyen, A. H., Y. Wang, D. E. White, M. O. Platt, and T. C. McDevitt. MMP-mediated mesenchymal morphogenesis of pluripotent stem cell aggregates stimulated by gelatin methacrylate microparticle incorporation. Biomaterials 76:66–75, 2016.

    CAS  Article  PubMed  Google Scholar 

  33. 33.

    Orban, J. M., L. B. Wilson, J. A. Kofroth, M. S. El-Kurdi, T. M. Maul, and D. A. Vorp. Crosslinking of collagen gels by transglutaminase. J. Biomed. Mater. Res. A 68:756–762, 2004.

    Article  PubMed  Google Scholar 

  34. 34.

    Placone, A. L., P. M. McGuiggan, D. E. Bergles, H. Guerrero-Cazares, A. Quiñones-Hinojosa, and P. C. Searson. Human astrocytes develop physiological morphology and remain quiescent in a novel 3D matrix. Biomaterials 42:134–143, 2015.

    CAS  Article  PubMed  Google Scholar 

  35. 35.

    Quigley, H. A., and E. M. Addicks. Chronic experimental glaucoma in primates. II. Effect of extended intraocular pressure elevation on optic nerve head and axonal transport. Investig. Ophthalmol. Vis. Sci. 19:137–152, 1980.

    CAS  Google Scholar 

  36. 36.

    Radany, E. H., M. Brenner, F. Besnard, V. Bigornia, J. M. Bishop, and C. F. Deschepper. Directed establishment of rat brain cell lines with the phenotypic characteristics of type 1 astrocytes. Proc. Natl. Acad. Sci. USA 89:6467–6471, 1992.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Ridet, J. L., S. K. Malhotra, A. Privat, and F. H. Gage. Reactive astrocytes: cellular and molecular cues to biological function. Trends Neurosci. 20:570–577, 1997.

    CAS  Article  PubMed  Google Scholar 

  38. 38.

    Rogers, R. S., M. Dharsee, S. Ackloo, J. M. Sivak, and J. G. Flanagan. Proteomics analyses of human optic nerve head astrocytes following biomechanical strain. Mol. Cell. Proteom. 11:M111.012302, 2012.

    Article  Google Scholar 

  39. 39.

    Schneider, M., and R. Fuchshofer. The role of astrocytes in optic nerve head fibrosis in glaucoma. Exp. Eye Res. 142:49–55, 2016.

    CAS  Article  PubMed  Google Scholar 

  40. 40.

    Seliktar, D., R. A. Black, R. P. Vito, and R. M. Nerem. Dynamic mechanical conditioning of collagen-gel blood vessel constructs induces remodeling in vitro. Ann. Biomed. Eng. 28:351–362, 2000.

    CAS  Article  PubMed  Google Scholar 

  41. 41.

    Sigal, I. A., J. G. Flanagan, I. Tertinegg, and C. R. Ethier. Predicted extension, compression and shearing of optic nerve head tissues. Exp. Eye Res. 85:312–322, 2007.

    CAS  Article  PubMed  Google Scholar 

  42. 42.

    Simon, D. D., C. O. Horgan, and J. D. Humphrey. Mechanical restrictions on biological responses by adherent cells within collagen gels. J. Mech. Behav. Biomed. Mater. 14:216–226, 2012.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Sofroniew, M. V., and H. V. Vinters. Astrocytes: biology and pathology. Acta Neuropathol. 119:7–35, 2010.

    Article  PubMed  Google Scholar 

  44. 44.

    Son, J. L., I. Soto, E. Oglesby, T. Lopez-Roca, M. E. Pease, H. A. Quigley, and N. Marsh-Armstrong. Glaucomatous optic nerve injury involves early astrocyte reactivity and late oligodendrocyte loss. Glia 58:780–789, 2010.

    PubMed  Google Scholar 

  45. 45.

    Sundararaghavan, H. G., G. A. Monteiro, N. A. Lapin, Y. J. Chabal, J. R. Miksan, and D. I. Shreiber. Genipin-induced changes in collagen gels: correlation of mechanical properties to fluorescence. J. Biomed. Mater. Res. A 87:308–320, 2008.

    Article  PubMed  Google Scholar 

  46. 46.

    Sung, K. E., X. Su, E. Berthier, C. Pehlke, A. Friedl, and D. J. Beebe. Understanding the impact of 2D and 3D fibroblast cultures on in vitro breast cancer models. PLoS ONE 8:e76373, 2013.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Tehrani, S., L. Davis, W. O. Cepurna, T. E. Choe, D. C. Lozano, A. Monfared, L. Cooper, J. Cheng, E. C. Johnson, and J. C. Morrison. Astrocyte structural and molecular response to elevated intraocular pressure occurs rapidly and precedes axonal tubulin rearrangement within the optic nerve head in a rat model. PLoS ONE 11:e0167364, 2016.

    Article  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Tehrani, S., E. C. Johnson, W. O. Cepurna, and J. C. Morrison. Astrocyte processes label for filamentous actin and reorient early within the optic nerve head in a rat glaucoma model. Investig. Ophthalmol. Vis. Sci. 55:6945–6952, 2014.

    CAS  Article  Google Scholar 

  49. 49.

    Tham, Y.-C., X. Li, T. Y. Wong, H. A. Quigley, T. Aung, and C.-Y. Cheng. Global prevalence of glaucoma and projections of glaucoma burden through 2040: a systematic review and meta-analysis. Ophthalmology 121:2081–2090, 2014.

    Article  PubMed  Google Scholar 

Download references


This work was supported by the Georgia Research Alliance, the Irish Research Council through the ELEVATE: Irish Research Council International Career Development Fellowship—co-funded by Marie Curie Actions, the National Space Biomedical Research Institute through NCC 9-58, and the National Science Foundation Graduate Research Fellowship under Grant No. DGE-1148903.

Author information



Corresponding author

Correspondence to C. Ross Ethier.

Additional information

Associate Editor Joel D. Stitzel oversaw the review of this article.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 1976 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Mulvihill, J.J.E., Raykin, J., Snider, E.J. et al. Development of a Platform for Studying 3D Astrocyte Mechanobiology: Compression of Astrocytes in Collagen Gels. Ann Biomed Eng 46, 365–374 (2018).

Download citation


  • Glaucoma
  • Astrocyte mechanobiology
  • Optic nerve head
  • 3D culture model
  • Mechanical conditioning