December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Establishment of an In Vitro Model of Human RPE/Bruch's Membrane
Author Affiliations & Notes
  • J ZhangJ Marshall
    GKT Department of Ophthalmology Rayne Inst St Thomas Hospital London United Kingdom
  • R Chibber
    London United Kingdom
  • J Hillenkamp
    London United Kingdom
  • J Marshall
    London United Kingdom
  • Footnotes
    Commercial Relationships   J. Zhang, None; R. Chibber , None; J. Hillenkamp, None; J. Marshall , None. Grant Identification: Study was supported by PPP Medical Trust
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 693. doi:
  • Views
  • Share
  • Tools
    • Alerts
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      J ZhangJ Marshall, R Chibber, J Hillenkamp, J Marshall; Establishment of an In Vitro Model of Human RPE/Bruch's Membrane . Invest. Ophthalmol. Vis. Sci. 2002;43(13):693.

      Download citation file:

      © ARVO (1962-2015); The Authors (2016-present)

  • Supplements

Abstract: : Purpose: Interactions between cell and its environment are known to play a major role in several cellular functions and maintenance of a stable differentiated phenotype. An important component of the extracellular environment is the extracellular matrix (ECM). Retinal pigment epithelium (RPE) is a layer of polarized epithelial cells interposed between the nerosensory retina and the Bruch's membrane/choriocapillaris. Bruch's membrane offers structural support and provides important barrier functions and selective filtration of solute molecules between the RPE and choriocapillaris. It is therefore important to provide these cells a similar in vivo environment to promote a development of their in vivo characteristics. The aim of this study was to develop an in vitro reconstituted model of RPE-Bruch's membrane complex, which recaptures the structure and function that is found in the living eye. Methods: Bruch's membrane preparations denuded of cells were prepared by incubating Bruch's/choriod preparations in Ca2+ and Mg2+ free HBSS for 4-6 weeks, and the preparations were washed with fresh HBSS once a week during incubating period. Human RPE cells were established and plated onto denuded human Bruch's membrane. The cells were grown to a confluent state and then maintained for an additional 4-6 weeks. Morphology of the RPE/Bruch's membrane complexes were examined by scanning and transmission electron microscopes. Formation of tight junctions and polarisation of the RPE cells on the complexes were determined by immunofluorescence microscopy. Results: Histological and electron microscopic examination demonstrated that the Bruch's -choroid preparations were free of cells, particularly in choroid, after 4-6 weeks HBSS treatment, indicating a relevant denuded preparation. It was also shown that the human RPE cells grown on these denuded Bruch's membranes form and maintain a health monolayer for entire culture period, and exhibit morphological polarization as seen in vivo. The RPE cells grown under this condition also developed a complete circumferential distribution of tight-junction proteins around each cell at the apical-lateral level. Na/K ATPase and Na/H exchangers were also found to be accumulated and polarized on the apical surface of these cells. Conclusion: For the first time, to our knowledge, we have demonstrated the ability to repopulate a denuded Bruch's membrane complex with a monolayer of RPE cells that exhibit many in vivo cell properties and characteristics. This provides a novel method of culturing cells on a collagen matrix and a useful model for the study of ageing and retina diseases.

Keywords: 567 retinal pigment epithelium • 333 Bruch's membrane • 403 extracellular matrix 

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.