Using the OCTA technique, this study compares the differences in structural, and vascular density parameters between eyes treated with Conbercept or Ranibizumab, with spontaneously regressed ROP lesions and those of age-matched, full-term healthy children. Besides, we are also the first to compare the long-term effects of two anti-vascular endothelial growth factor drugs on retinal structure and retinal microcirculation: We found a significant reduction in FAZ area in children with ROP. Anti-VEGF treatment significantly reduced CFT, and Conbercept showed a better performance in comparison to Ranibizumab. In addition, FAZ area was negatively correlated with foveal VD and CFT.
With regard to visual function, we did not find a correlation between foveal structural parameters and visual acuity using linear regression analysis. Currently, studies on factors associated with long-term visual outcome have competing results. Villegas et al. noted similar conclusion that none of the structural factors was significantly correlated to BCVA16. Balasubramanian et al. found that worse BCVA was associated with a smaller FAZ in a cross-sectional study of 32 eyes of preterm children17, while chen et al. found superficial VD and inner retinal thickness were associated with suboptimal visual acuity18. Stoica et al. discovered a better VA was correlated with higher BW and GA at birth, as well as with a thinner CFT19. Embryogenesis demonstrates that the development of the fovea externa is relatively independent from foveal pit20, thus despite the structural differences in FAZ and CFT in ROP eyes, the development of visual acuity may not be as significantly affected, which supported our observation.
Blood vessels in the human optic disc region first appear around week 14-1521, and the vascular plexus encircling the central avascular zone begins to form around fetal week 25. The central area of the macula has never been vascularized,22, 23. In contrast, foveal retinal tissue continues to mature for several years after birth, and Dubis et al. has found that morphologic development of the fovea may be completed by around 17 months24. Therefore, changes in the local microenvironment during this time period can affect the development of both the macula tissure and FAZ region. VEGF plays an important role in the growth of retinal vessels towards the fovea section. Local hypoxia generated from ganglion cells differentiation drives angioblast migration at the vascular front by promoting the local synthesis of VEGF within astrocyte precursor cells, in this way, capillaries expand into the previously hypoxic retina following the template of astrocytes, thus mitigating the local hypoxia21.At the same time, the fovea center expresses more antiangiogenic molecules than the perifoveal and peripheral retinal areas, mainly pigment epithelium-derived factor (PEDF), brain natriuretic peptide, etc. The macular pigment is also involved in this process20. The balance between these factors is the key to maintain FAZ25. Therefore, during the formation of FAZ and fovea pit, the increase in the concentration of VEGF and anti-angiogenic molecules mentioned above in eyes with ROP might break this balance, resulting in the decrease or even disappearance of FAZ, which explains the significant reduction in FAZ area with ROP. In the present study, there was no significant difference in the FAZ area between group A, B, C. Therefore, it can be demonstrated that the types of drug might have little effect on the area of FAZ. VEGF-A is the most potent stimulator of pathological angiogenesis among the VEGF family members. Placenta growth factor (PlGF) is a pleiotropic cytokine, analogous to vascular endothelial growth factor (VEGF), could stimulate angiogenesis by binding to its membrane-bound receptor, Fms-like tyrosine kinase 1 (Flt-1). Ranibizumab is a recombinant, humanised monoclonal antibody that binds to VEGF-A with a high affinity to making it inert26, whilst Conbercept is a 143-kDa designed fusion protein that binds to all VEGF-A isoforms as well as the related VEGFR-1 ligands, VEGF-B and PlGF. Considering the difference in affinity for mentioned molecules, the latter binds to VEGF 50 times better than Ranibizumab. With respect to peripheral vascular structural anomalies verified on FFA, there is no significant difference between ROP neonates who received intravitreal Conbercept (IVC) or Ranibizumab (IVR)27. However, Cheng et al. reported that IVC yield less recurrence and longer treatment intervals compared with IVR28, proving the higher effect of the Conbercept in short-term observation. Our results demonstrated that there was no difference in the long-term FAZ area changes between different drug choices. One possible explanation is that the occurrence of FAZ reduction has already occurred before the injection of anti-VEGF agents, and the formation of FAZ differences stems mainly from the pathological mechanism of ROP itself rather that treatment chosing.
Besides the area of FAZ, we also found that the mean VD-SCP (foveal) and VD-DCP (foveal) in the group A, B, C were higher than those in the group D. Although there was no significant difference between the four groups, the relative trend existed. Takagi et al. found that the vessel density at the fovea in the ROP group was significantly higher than in the control group (35.7 vs 30.1%, p < 0.01)29. Chen et al. also discovered increased foveal VD (of SCP and DCP) in IVB-treated preterm children in comparison to healthy controls18. Asli Vural et al. found smaller FAZ, and increased central foveal VD in all eyes with ROP regardless of treatment options30. In the present study, we speculated that the difference in significance came from the relatively small sample size. However, the trend of quantitative parameters remained consistent with previous studies mentioned above. Furthermore, we also found that the FAZ area and foveal VD (at the level of both the SCP and DCP) are strongly negatively correlated. We suspect that there are 2 possible reasons for this phenomenon: first, increase of VEGF concentration might lead to the compensatory elevation of retinal blood vessel density in the foveal section; second, the physiological displacement of inner retina might be incomplete due to the smaller avascular area in ROP. We also found that the VD-SCP (parafoveal) of group A and B was significantly smaller than that of group C, indicated that concentration of VEGF may simultaneously affect the vascular density of parafoveal section.
With regard to choroidal VD, Rezar-Dreindl et al. conducted a cross-sectional study of 15 children (30 eyes) with a history of ROP, which showed no statistically significant differences in choroidal VD between the ROP and control groups.31. Our study obtained similar results: choriocapillary VD was not related to treatment or drug choice, indicating that choriocapillaris might not be significantly affected in the disease course. However, different conclusions have been made regarding the vascularization of other levels of choroidal tissue. Lavric et al. has found that children with ROP had a much reduced choroidal vascularity index (defined as the proportion of luminal areas to the total sub-foveal choroidal area), which indicated compromised choroidal vascularity32. A smaller choroidal vascularity index might demonstrate a reduction of oxygen delivery to the outer retina, and thinner choroidal thickness was also associated with worse VA33.Until now, studies between choroid and ROP are still relatively sparse, and further researches are needed to determine the role of the choroid in ROP development.
The CFT of ROP children was higher than that of age matched healthy children in our study. What’s more, our study confirmed that a smaller FAZ was significantly correlated with a thicker CFT, which was consistent with the findings from Asli Vural et al, who also reported that FAZ was negatively correlated with CFT (r=-0.27, p = 0.03)30. Bowl et al. has also found that the mean values of the CFT were highest in the spontaneously regressed ROP group, intermediate in the premature children without ROP group, and lowest in the age-matched term-born group34. FAZ is prerequisite for formation of a fovea pit. The foveal pit is formed by the centrifugal migration of inner retinal neurons away from the fovea and the centripetal migration of cone cell nuclei towards the center35. Decrease in the migration ability of retinal neurons in the inner layer of the fovea is the main cause of the increase in retinal thickness36. Vascularized tissue is less elastic than avascular retinal tissue, thus it's reasonable to believe that the retina on FAZ section is more elastic and malleable than the vascularized retina around it under the action of intraocular pressure37. The existence of capillaries will mechanically interfere with the centrifugal displacement of GCL and INL cells, hence a smaller FAZ will limit foveal pit widening35, which increases the foveal thickness subsequently36. Our study has a further discovery that the CFT of group B was lower than group C and higher than group A and D, which indicates that anti-VEGF agents can effectively reduce the CFT of ROP eyes, and the effect of Conbercept is even better. We speculated that the formation of CFT may not only be affected by FAZ. Conbercept may affect the CFT through other pathways, although more researches is needed to verify this conclusion.
Our study still has some limitations. First, we did not enroll children who had received laser treatment and had a history of prematurity but no ROP. Prematurity itself or laser treatment may have contributed to the retinal vascular and structural changes mentioned above. Second, all children in our study were Chinese, and retinal vascular or structural parameters may vary based on ethnicity. Finally, the sample size of this study is relatively small and needs to be expanded for better validation.
In conclusion, we used OCT and OCTA technologies to investigate the foveal structural and microvascular characteristics of ROP children (treated with Conbercept, Ranibizumab, and spontaneously regressed) and age-matched children. OCTA, as a non-invasive test, can help us to understand more clearly the macular microstructural and microvascular abnormalities in children with ROP, such as decreased FAZ, and increased CFT, as well as the correlations between OCTA parameters and demographic data. We also have proved that Conbercept have a better performance in reducing CFT than Ranibizumab. However, further studies are needed to gain more insight into the developmental processes of retinal vessels and structures in ROP, as well as to identify the pros and cons of various treatment options.