November 1997
Volume 38, Issue 12
Free
Articles  |   November 1997
A new method for studying the selective adherence of blood lymphocytes to the microvasculature of human retina.
Author Affiliations
  • T A Hill
    Department of Immunology, United Medical and Dental Schools, London, United Kingdom.
  • M R Stanford
    Department of Immunology, United Medical and Dental Schools, London, United Kingdom.
  • E M Graham
    Department of Immunology, United Medical and Dental Schools, London, United Kingdom.
  • D C Dumonde
    Department of Immunology, United Medical and Dental Schools, London, United Kingdom.
  • K A Brown
    Department of Immunology, United Medical and Dental Schools, London, United Kingdom.
Investigative Ophthalmology & Visual Science November 1997, Vol.38, 2608-2618. doi:
  • Views
  • PDF
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      T A Hill, M R Stanford, E M Graham, D C Dumonde, K A Brown; A new method for studying the selective adherence of blood lymphocytes to the microvasculature of human retina.. Invest. Ophthalmol. Vis. Sci. 1997;38(12):2608-2618.

      Download citation file:


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

      ×
  • Supplements
Abstract

PURPOSE: To develop a sensitive and reproducible technique for measuring the adherence of blood lymphocytes to vessel walls exposed in sections of human retina and for examining the role of lymphocyte and vascular adhesion molecules in these events. METHODS: Cryostat sections of human retina were overlaid with blood lymphocytes from healthy subjects, and experimental conditions were sought by which preferential attachment of the cells occurred to blood vessel walls in the retinal sections. Adherent lymphocytes were identified by staining with methyl green-thionine, and transected blood vessels were identified by their structure and by staining of basement membranes with periodic acid-Schiff. The adherence of enriched preparations of CD4+ (T-helper) and CD8+ (T-cytotoxic) lymphocytes, of interleukin-2 (IL-2)-activated cells, and of lymphocytes from patients with ocular Behçet's disease was examined. The distribution of adhesion molecules on retinal vessel walls was determined by immunohistochemistry, and the contribution of leukocyte integrins to lymphocyte binding was studied by blocking experiments with monoclonal antibodies. RESULTS: The optimal selectivity of blood lymphocyte attachment to retinal vessel walls occurred when purified lymphocytes were suspended in culture medium with 10% fetal calf serum and overlaid onto retinal sections for 30 minutes at 23 degrees C with gentle agitation. Under these conditions, 92% of the lymphocytes that adhered to the section were confined to the retinal microvasculature, and CD4+ T cells were more adherent than CD8+ T cells (P < 0.01). Prior exposure of normal lymphocytes to IL-2 enhanced their binding to retinal blood vessels, and lymphocytes from patients with Behçet's disease showed supranormal vascular adherence (P < 0.005). Many transected vessels stained positively for CD31; PECAM (mean 62%), CD54; ICAM-1 (mean 73%), CD62E; E-selectin (mean 35%), CD62P; P-selectin (mean 61%), and CD106; VCAM-1 (mean 42%). However, these vascular adhesion molecules occupied < 20% of the area of the blood vessel walls. Lymphocyte adhesion to the retinal vessels was more dependent on CD29 (the common chain of the beta 1 integrins) expression than either CD11a/CD18 or CD49d. CONCLUSIONS: This technique allows measurements to be made of lymphocyte adherence to vascular and nonvascular structures of retina ex vivo. Extension of this approach to the study of leukocyte adherence to sections of pathologic retina may be of clinical and experimental applicability in understanding mechanisms of retinal inflammation.

×
×

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.

×