The role of Red cell characteristics, Angiogenesis, Viscosity and Oxygenation in the pathophysiology of Sickle cell related retinopathy (RAVOS study)

Sickle cell disease (SCD) is characterized by chronic hemolysis and recurrent
microvascular occlusion, resulting in chronic inflammation, increased
endothelial adhesiveness, local ischemia and organ damage. This process can
affect almost every organ, which can lead to significant morbidity and
mortality. Interestingly, a large clinical variability is noticed between
individuals with SCD which is currently not well understood. SCD is
predominantly seen in individuals of African, Middle Eastern, Mediterranean or
Asian descent, but is most prevalent in sub-Saharan countries. Worldwide, more
than 300.000 individuals are born with SCD every year. While the prevalence is
significantly lower in Western Europe, the numbers are rising due to the
increase of migration, making SCD an increasingly important health problem in
Western Europe.

In this project, we focus on sickle cell related retinal damage. The retina is
very sensitive to ischemic damage, resulting in sickle cell retinopathy (SCR).
SCR can be divided in non-proliferative retinopathy (NPSCR) and proliferative
retinopathy (PSCR). The latter is characterized by the development of
neovascularization progressing to scarring, which can lead to vitreous
hemorrhage or retinal detachment. These complications can drastically impair
the visual acuity. Therefore, regular ophthalmologic screening is recommended
by performing a dilated eye examination every one or two years, starting from
the age of ten years. However, the frequency of individual screenings is not
well defined, since evidence is lacking. However, there is a remarkable
difference in the prevalence of, in particular PSCR, between patients with the
HbSS and HbSC genotype. Patients with the combined heterozygosity (HbSC) do
more frequently have PSCR despite the fact that the clinical presentation and
frequency of other sickle cell-related complications in HbSC patients is milder
as compared to patients with homozygous SCD (HbSS).

SCR results from a combination of ischemic vascular disease and secondary
angiogenesis, but it is unknown which factors play a significant role in the
pathogenesis. Studies on this subject are still in an early phase. The aim of
this study is to get more insight in the pathogenesis of (P)SCR.

The aim of this study is to get more insight in the pathogenesis of (P)SCR by
comparing: (1) Red cell characteristics (deformability and point of sickling in
hypoxia measured by the Oxygenscan), (2) Angiogenesis by determining several
biomarkers of angiogenetic activity in plasma samples, (3) Whole blood
viscosity and (4) Oxygenation by the assessment of vessel density in the retina
by the angio-OCT and by the assessment of the oxygen saturation in the retinal
vessels with the Oxymap scan between patients with and without (P)SCR in both
genotypes.

The Oxygenscan is a new method to determine ex vivo the deformability of red
blood cells as a function of the partial pressure of oxygen. OCT angiography is
a non-invasive routinely used device at the department of Ophthalmology to
assess vessel density and macular thinning as markers of impaired oxygenation.
The Oxymap scan is a non-invasive device, which will be used to assess the
oxygen saturation in retinal arterioles and venules.

In our specialized teaching hospital, more than 300 adult patients with SCD
have been systematically screened on all forms of organ damage including
retinopathy. For this study, 80 adult patients with SCD with the HbSS or HbSC
genotype will be selected for inclusion. For both genotype groups, 20 patients
without SCR and 20 patients with PSCR will be included. When these numbers
cannot be achieved, patients with non-proliferative SCR will be included to
achieve 40 patients per genotype. Patients will receive a complete
ophthalmologic examination (best corrected visual acuity, slit lamp
examination, dilated fundus examination and OCT angiography) to determine their
current status of retinopathy. Blood samples will be drawn for the analysis of
the red blood cell characteristics including adhesiveness, deformability and
point of sickling by Oxygenscan (as described above). Information on other SCD
characteristics (e.g. patient demographics and sickle cell related organ
damage) will be retrieved from the operational clinical database. To determine
the role of angiogenesis, plasma levels of VEGF (vascular endothelial growth
factor), CTGF (connective tissue growth factor), CD105 (endoglin) and
angiopoietin-2 will be assessed in these blood samples using ELISA (enzyme
linked immunosorbent assay) kits. To determine the role of whole blood
viscosity, we will use a cone/plate viscometer (Brookfield DVII+ with CPE40
spindle, Brookfield Engineering Labs, Natick, MA, USA).

Included patients will not experience direct advantages of their participation.
However, their participation will increase knowledge on risk factors of (P)SCR
and might therefore improve health care for SCD patients. We aim to collect the
blood samples during planned venepuncture for routine care (thus: no extra
risks for the patient). Furthermore, we request patients to fast 8 hours prior
to the venepuncture (or, if fasting is not feasible, to consume a low-fat
breakfast). As we also plan the venepunctures in the morning to avoid
unnecessary long fasting, we deem the risk for the patients low.