The pachychoroid spectrum disease including PNV is a recently proposed disease entity characterized by pachyvessels1,3 Our results showed that IL-4 was significantly associated with the presence of pachyvessels in the choroid and the disease spectrum of PNV. More specifically, the remodeling of the pachyvessels was associated with higher levels of expression of IL-4, which was partly associated with the formation of polypoidal lesions. These findings suggested that IL-4 is significantly involved in the vascular remodeling of the choroidal vessels and may play different roles in the PNV disease spectrum.
In the disease spectrum of PNV, IL-4 was specifically associated with the choroidal thickness, and the choroidal thickness was associated with the single nucleotide polymorphism of IL-4-590 (rs2243250), which is a well-recognized allele associated with asthma and rhinitis.14,15
Earlier, the molecular pathology of nAMD was examined by analyzing the aqueous humor of the patients. Several studies found a significant association of inflammatory cytokines with nAMD. For example, VEGF16,17, CCL25,6,18,19, CCL319, CCL419, IL-1α 20, IL-67,8,9, IL-87,9,10, IL-1520, IL-36β21, TGF-β18, CXCL9 (C-X-C Motif Chemokine Ligand 9)18, CXCL1020, CXCL1222, angiopoietin-210, HGF (hepatocyte growth factor)10, TIMP1 (tissue inhibitor of metallopeptidase 1)10, FGF1 (fibroblast growth factor 1)21, and angiogenin21 were shown to be significantly associated with nAMD.
In contrast, there have been few reports for PNV. Earlier studies showed that it was associated with elevated levels of VEGF6, CCL2 (C-C motif chemokine 2)6,22, IL-8, angiopoietin-2, PGF (placental growth factor), CXCL12 (C-X-C Motif Chemokine Ligand 12), and CXCL13 (C-X-C Motif Chemokine Ligand 13)22. Thus, eyes with PNV have overlapping profile of cytokine elevation with nAMD, however, there remains unclear whether PNV have specific cytokine profile.
PNV is characterized by pachyvessels and by vascular hyperpermeability and vasodilation of the outer choroidal vessels. Thus, for example, the vascular hyperpermeability of the pachyvessels causes choroidal thickening and subretinal exudative changes.10 However, previous reports on PNV assessed its association with the disease category, and no detailed analyses for the disease characteristics have been performed. Thus, it remains unclear how the observed cytokines may be associated with or contribute to the formation of pachyvessels or vascular remodeling.
The critical disease-associated cytokines for PNV appears to be VEGF based on previous information on nAMD. However, in PNV, the contribution of the VEGF in the aqueous is inconsistent depending on the study. Kato et al reported an elevation of VEGF in PNV eyes,6 however VEGF was reported to be down-regulated in eyes with PNV with drusen.22
Inoda et al reported an elevation of CRP, CXCL12, CXCL13, IL-8, angiopoietin-2, PIGF, and CCL2 in eyes with PNV.22 An important clinical characteristic of PNV was an increase in the choroidal thickness, however an assessment of their association with choroidal thickness was not performed. In our analyses, an elevation of IL-8 and CCL2 was not observed in PNV patients.
Because CCL2 is well recognized to be associated with nAMD, we also examined the association of CCL2 with choroidal thickness by structural equation modeling analyses. We found a significant and negative effect of CCL2 on choroidal thickness (coefficient: -0.18, P = 0.007). However, the inclusion of CCL2 into the structural equation modeling analysis did not appreciably change the structure of outcomes of the structural equation model. Thus, the role of CCL2 is contentious and presumably context dependent.
For the increased vascular permeability and vasodilatability, T helper 2 (Th2) lymphocytes and related cytokines including IL-4, IL-5, IL-6, IL-9, IL-13, and IL-17E (IL-25) play significant roles. Of these, IL-4 decreases the endothelial barrier function which causes the vascular hyperpermeability.23,24 Importantly, the IL-4-mediated hyperpermeability of the vascular endothelial cells has been shown to be a major mechanism of the inflammatory edema.25
Mechanistically, IL-4 impairs the barrier function of the vascular endothelium by forming gaps between the endothelial cells through the rearrangement of the cytoskeleton.26 Another mechanism of IL-4 mediated barrier dysfunction has been recently reported. IL-4 stimulates the formation of actin stress fibers by a non-canonical Wnt ligand, Wnt5A.27 This eventually causes a remodeling of the cytoskeleton leading to an impairment of the barrier function.27 Thus, we propose that inflammatory networks featuring IL-4 contributes to vascular hyperpermeability leading to the development of the pachyvessels.
Another characteristic of pachyvessels is the vasodilation of the outer choroidal vessels. Vasodilation has also been reported to be associated with IL-4. IL-4 promotes both the lumen formation13,28 and the endothelial cell cycle.29 In addition, IL-4 stimulates the VCAM-1 promoter via STAT630 and exerts proangiogenic effects on the VCAM-1/alpha4 integrin pathway.28 Alternatively, vasodilation itself induces chronic ischemia and the hypoxic conditions of the choriocapillaris2 leading to IL-4 induction.31
Considering the pathogenic mechanism of PNV as a venous overload choroidopathy, PNV is homologous to diseases with chronic venous congestions.3 A wide variety of venous disorders throughout the body including varicose veins32, pregnancy33,34, pelvic congestion syndrome35, and gastrointestinal varices associated with cirrhosis and portal hypertension36 are characterized as vasculopathy due to venous overload. The venous overload promotes capillary dilation and increases the permeability, inflammatory cell infiltration, activation of matrix metalloproteinases (MMPs)37, and induction of cell adhesion molecules (CAMs).38
Importantly, IL-4 has been shown to be associated with the development of vasodilatability, hypervascular permeability, and tissue vascular remodeling which is exemplified in the placenta during pregnancy39 and the regeneration of cardiomyocytes.40
Analysis of the aqueous humor showed that IL-4 was also associated with polypoidal lesions.11 In addition, elevated IL-4 levels may induce polypoidal formation or choroidal neovascularization.13 Polypoidal lesions are often accompanied by pachyvessels.41 Because large polypoidal lesions can be regarded as neovascular tangles or branching neovascular networks rather than actual polypoidal lesions or aneurysmal dilatations in OCT images,42 polypoidal lesions may reflect angiogenic lesions.41,43 Our results showed that IL-4 was significantly associated with the choroidal thickness and the presence of polypoidal lesions in eyes with PNV. This is consistent with previous reports that showed that the development of polypoidal lesions was closely associated with development of pachyvessels.44 Collectively, the PNV diseases appear to be associated with a vascular remodeling including the development of polypoidal lesions. In addition, the pachyvessels may also promote choroidal neovascularization.45
There are some limitations in this study. Our study was a retrospective analysis and may not reflect direct causal relationships. For example, there is a possibility that the contribution of IL-4 may be indirect effects by unobserved factors or molecules. However, our previous analyses using IL-4 deficient mice supported the significant and direct roles of IL-4 in vascular remodeling which supports our conclusions.13
To understand the roles of IL-4 in greater detail, clinical trials of blocking IL-4 in PNV patients are required. Nevertheless, our study provides a strong basis for future therapy blocking the IL-4 pathway to treat patients with PNV spectrum of diseases.
In conclusion, our findings indicate that IL-4 represents a crucial mediator in the development of PNV spectrum of disease as well as the vascular remodeling in PNV patients.