A rat model for retinal vascular occlusion by human sickle erythrocytes

G. A. Lutty, A. Phelan, D. S. McLeod, C. Merges, S. M. Suzuka, M. E. Fabry, R. L. Nagel

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Abstract

Purpose. To determine if human sickled erythrocytes (SRBCs) are retained in the retinal vasculature and what type of SRBC is retained using a rat model. Methods. RBCs from patients homozygous for hemoglobin S (SS) or heterozygous (SC) were separated on Percoll-Larex continuous gradients, labeled with fluorescein isothiocyanate (FITC), and delivered via the left ventricle to anesthetized, ventilated rats. Blood gas levels were altered by changing inspired gas and monitored via a femoral arterial catheter. After 5 min, animals were exsanguinated and retinas processed by our ADPase flatmount technique. Results. Higher number of high density SS cells (SS4) were retained in the rat retinal vasculature than normal density SS cells (SS2). The number of SS4 cells retained was inversely dependent on the arterial oxygen tension, whereas the number of SS2 cells retained was independent of the rat's PaO2. Most SS4 cells were retained in peripheral capillaries (average lumenal diameter 5.36 μm) and occupied the full lumenal diameter of vessels in most cases. In contrast to SS donors, very few RBCs from SC donors (normal or high density) were retained in the normal retinal vasculature and the number did not increase significantly with hypoxia. Preliminary data suggests that stimulation of endothelium with lipopolysaccharide, which is known to stimulate release of cytokines, increases retention of SC RBCs. Conclusions. This model demonstrates that high density SS cells, which include irreversibly sickled cells, are retained in normal retinal vessels of the rat and that the number retained is oxygen dependent. Preferential retention of high density cells and blockage of the full diameter of the vessel suggests hemoglobin polymer-mediated, mechanical obstruction and not adhesion is responsible for retention of SS4 cells in the normal retinal vasculature. In contrast, very few SC cells of any density are retained unless the endothelium is stimulated. Therefore, adhesion of sickle erythrocytes may not be a feature of retinal vaso-occlusion in the steady state, but endothelial activation may be an important modulating phenomena.

Original languageEnglish (US)
JournalInvestigative Ophthalmology and Visual Science
Volume37
Issue number3
StatePublished - Feb 15 1996

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Retinal Vessels
Cell Count
Erythrocytes
Sickle Hemoglobin
Endothelium
Gases
Tissue Donors
Apyrase
Oxygen
Thigh
Fluorescein
Heart Ventricles
Lipopolysaccharides
Retina
Arterial Pressure
Catheters
Cytokines

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Lutty, G. A., Phelan, A., McLeod, D. S., Merges, C., Suzuka, S. M., Fabry, M. E., & Nagel, R. L. (1996). A rat model for retinal vascular occlusion by human sickle erythrocytes. Investigative Ophthalmology and Visual Science, 37(3).

A rat model for retinal vascular occlusion by human sickle erythrocytes. / Lutty, G. A.; Phelan, A.; McLeod, D. S.; Merges, C.; Suzuka, S. M.; Fabry, M. E.; Nagel, R. L.

In: Investigative Ophthalmology and Visual Science, Vol. 37, No. 3, 15.02.1996.

Research output: Contribution to journalArticle

Lutty, GA, Phelan, A, McLeod, DS, Merges, C, Suzuka, SM, Fabry, ME & Nagel, RL 1996, 'A rat model for retinal vascular occlusion by human sickle erythrocytes', Investigative Ophthalmology and Visual Science, vol. 37, no. 3.
Lutty GA, Phelan A, McLeod DS, Merges C, Suzuka SM, Fabry ME et al. A rat model for retinal vascular occlusion by human sickle erythrocytes. Investigative Ophthalmology and Visual Science. 1996 Feb 15;37(3).
Lutty, G. A. ; Phelan, A. ; McLeod, D. S. ; Merges, C. ; Suzuka, S. M. ; Fabry, M. E. ; Nagel, R. L. / A rat model for retinal vascular occlusion by human sickle erythrocytes. In: Investigative Ophthalmology and Visual Science. 1996 ; Vol. 37, No. 3.
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abstract = "Purpose. To determine if human sickled erythrocytes (SRBCs) are retained in the retinal vasculature and what type of SRBC is retained using a rat model. Methods. RBCs from patients homozygous for hemoglobin S (SS) or heterozygous (SC) were separated on Percoll-Larex continuous gradients, labeled with fluorescein isothiocyanate (FITC), and delivered via the left ventricle to anesthetized, ventilated rats. Blood gas levels were altered by changing inspired gas and monitored via a femoral arterial catheter. After 5 min, animals were exsanguinated and retinas processed by our ADPase flatmount technique. Results. Higher number of high density SS cells (SS4) were retained in the rat retinal vasculature than normal density SS cells (SS2). The number of SS4 cells retained was inversely dependent on the arterial oxygen tension, whereas the number of SS2 cells retained was independent of the rat's PaO2. Most SS4 cells were retained in peripheral capillaries (average lumenal diameter 5.36 μm) and occupied the full lumenal diameter of vessels in most cases. In contrast to SS donors, very few RBCs from SC donors (normal or high density) were retained in the normal retinal vasculature and the number did not increase significantly with hypoxia. Preliminary data suggests that stimulation of endothelium with lipopolysaccharide, which is known to stimulate release of cytokines, increases retention of SC RBCs. Conclusions. This model demonstrates that high density SS cells, which include irreversibly sickled cells, are retained in normal retinal vessels of the rat and that the number retained is oxygen dependent. Preferential retention of high density cells and blockage of the full diameter of the vessel suggests hemoglobin polymer-mediated, mechanical obstruction and not adhesion is responsible for retention of SS4 cells in the normal retinal vasculature. In contrast, very few SC cells of any density are retained unless the endothelium is stimulated. Therefore, adhesion of sickle erythrocytes may not be a feature of retinal vaso-occlusion in the steady state, but endothelial activation may be an important modulating phenomena.",
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AU - Lutty, G. A.

AU - Phelan, A.

AU - McLeod, D. S.

AU - Merges, C.

AU - Suzuka, S. M.

AU - Fabry, M. E.

AU - Nagel, R. L.

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N2 - Purpose. To determine if human sickled erythrocytes (SRBCs) are retained in the retinal vasculature and what type of SRBC is retained using a rat model. Methods. RBCs from patients homozygous for hemoglobin S (SS) or heterozygous (SC) were separated on Percoll-Larex continuous gradients, labeled with fluorescein isothiocyanate (FITC), and delivered via the left ventricle to anesthetized, ventilated rats. Blood gas levels were altered by changing inspired gas and monitored via a femoral arterial catheter. After 5 min, animals were exsanguinated and retinas processed by our ADPase flatmount technique. Results. Higher number of high density SS cells (SS4) were retained in the rat retinal vasculature than normal density SS cells (SS2). The number of SS4 cells retained was inversely dependent on the arterial oxygen tension, whereas the number of SS2 cells retained was independent of the rat's PaO2. Most SS4 cells were retained in peripheral capillaries (average lumenal diameter 5.36 μm) and occupied the full lumenal diameter of vessels in most cases. In contrast to SS donors, very few RBCs from SC donors (normal or high density) were retained in the normal retinal vasculature and the number did not increase significantly with hypoxia. Preliminary data suggests that stimulation of endothelium with lipopolysaccharide, which is known to stimulate release of cytokines, increases retention of SC RBCs. Conclusions. This model demonstrates that high density SS cells, which include irreversibly sickled cells, are retained in normal retinal vessels of the rat and that the number retained is oxygen dependent. Preferential retention of high density cells and blockage of the full diameter of the vessel suggests hemoglobin polymer-mediated, mechanical obstruction and not adhesion is responsible for retention of SS4 cells in the normal retinal vasculature. In contrast, very few SC cells of any density are retained unless the endothelium is stimulated. Therefore, adhesion of sickle erythrocytes may not be a feature of retinal vaso-occlusion in the steady state, but endothelial activation may be an important modulating phenomena.

AB - Purpose. To determine if human sickled erythrocytes (SRBCs) are retained in the retinal vasculature and what type of SRBC is retained using a rat model. Methods. RBCs from patients homozygous for hemoglobin S (SS) or heterozygous (SC) were separated on Percoll-Larex continuous gradients, labeled with fluorescein isothiocyanate (FITC), and delivered via the left ventricle to anesthetized, ventilated rats. Blood gas levels were altered by changing inspired gas and monitored via a femoral arterial catheter. After 5 min, animals were exsanguinated and retinas processed by our ADPase flatmount technique. Results. Higher number of high density SS cells (SS4) were retained in the rat retinal vasculature than normal density SS cells (SS2). The number of SS4 cells retained was inversely dependent on the arterial oxygen tension, whereas the number of SS2 cells retained was independent of the rat's PaO2. Most SS4 cells were retained in peripheral capillaries (average lumenal diameter 5.36 μm) and occupied the full lumenal diameter of vessels in most cases. In contrast to SS donors, very few RBCs from SC donors (normal or high density) were retained in the normal retinal vasculature and the number did not increase significantly with hypoxia. Preliminary data suggests that stimulation of endothelium with lipopolysaccharide, which is known to stimulate release of cytokines, increases retention of SC RBCs. Conclusions. This model demonstrates that high density SS cells, which include irreversibly sickled cells, are retained in normal retinal vessels of the rat and that the number retained is oxygen dependent. Preferential retention of high density cells and blockage of the full diameter of the vessel suggests hemoglobin polymer-mediated, mechanical obstruction and not adhesion is responsible for retention of SS4 cells in the normal retinal vasculature. In contrast, very few SC cells of any density are retained unless the endothelium is stimulated. Therefore, adhesion of sickle erythrocytes may not be a feature of retinal vaso-occlusion in the steady state, but endothelial activation may be an important modulating phenomena.

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