Extensive reduction in choroidal thickness after photodynamic therapy in eyes with central serous chorioretinopathy

We examined the effect of reduced fluence (rf)-photodynamic therapy (PDT) of the macular area on the wide-field choroidal thickness in 20 eyes with central serous chorioretinopathy (CSC) and 20 age- and sex-matched control eyes. The choroidal thickness at the posterior pole was measured before and after rf-PDT, using a grid with inner and outer rings, each divided into superotemporal, inferotemporal, superonasal, and inferonasal quadrants, respectively, making up a total of nine subfields including the central 3 mm ring. Before treatment, all eyes showed wide-field choroidal thickening from the dilated vortex vein ampulla to the fovea, along the course of the vein. After rf-PDT of the macular area, the choroidal thickness significantly decreased, not only in the irradiated macular area but also outside the arcade vessels in all quadrants (p < 0.001 for all inner subfields; p = 0.035 and p = 0.024 for the outer superonasal and inferonasal subfields, respectively; p < 0.001 and p = 0.004 for the outer superotemporal and inferotemporal subfields, respectively). For control eyes, the choroidal thickness did not differ between the initial visit and follow-up 1.2 ± 0.7 months after the initial visit (p > 0.05 for all subfields). These findings provide new insights into the pathogenesis of CSC and explain the reasons for the effectiveness of rf-PDT for this condition.

vortex veins at the scleral outlet contributes to the choroidal overload, resulting in the formation of SRF at the posterior pole 16 .
Most of the SRF from CSC is spontaneously absorbed. However, persistent SRF may cause poor visual prognosis 17,18 . Currently, photodynamic therapy (PDT) is the treatment of choice for chronic or recurrent CSC 19 . Studies have shown that half-dose (hd) and reduced-fluence (rf)-PDT are both effective and safe treatments for CSC [20][21][22][23] . A key benefit of rf-PDT is its ability to control the disease while minimizing risks. Some treatments can harm the choriocapillaris, a layer of tiny blood vessels in the eye, which may cause the retina to become thinner and potentially lead to vision loss. This risk is especially concerning for CSC patients, who generally have good visual acuity that they want to preserve. As such, rf-PDT, which mitigates these risks, is a highly favored treatment choice for this group of patients.
Several studies have reported changes in the CSC-associated choroidal structures after PDT [24][25][26] . Maruko et al. 26 showed reduced subfoveal choroidal thickness and choroidal vascular hyperpermeability in CSC after PDT. Presently, PDT is considered to reduce the recurrence of SRF in CSC more effectively than does focal photocoagulation 27 .
However, the effect of PDT, when applied to the macular area on the extensive choroid has not been fully elucidated. Most recently, a study using WF SS-OCT showed that the choroidal thickening extended to the periphery, including vortex vein ampulla 12 . The present study examined the effect of rf-PDT on the WF choroidal thickness in CSC. The current findings may be helpful to elucidate the mechanism of rf-PDT for CSC and improve our understanding of the pathophysiology of CSC.

Results
This study included 20 eyes of 20 patients (14 men, 6 women) with unilateral CSC treated with rf-PDT, as well as 20 eyes of 20 patients (12 men, 8 women) with unilateral CSC that was untreated for a duration of 1.2 ± 0.7 months (Fig. 1).
The choroidal thickness map, captured using WF SS-OCT, covered the vortex vein ampulla. All 40 eyes exhibited some signs of dilated vortex veins and thickening of the choroid from the vortex vein ampulla to the fovea along the course of the vein at the initial visit (Figs. 2 and 3). Choroidal thickness map analysis using enhanced depth imaging in wide-field swept-source optical coherence tomography (A) Infrared scanning laser ophthalmoscopy images with a measurement grid that was developed for assessing changes in the wide-field (WF) choroidal thickness. The grid consisted of nine subfields divided by three circles with diameters of 3, 9, and 18 mm, respectively, and four lines. The circumferential and zonal areas enclosed by these circles were divided into superotemporal, inferotemporal, superonasal, and inferonasal subfields, considering the arrangement of the vortex veins.  Table 1 delineates the baseline characteristics of patients experiencing visual disturbances due to SRF. The mean pre-rf-PDT SRF height at the fovea and the subfoveal choroidal thickness were 151 ± 101 µm and 352 ± 101 µm, respectively, in the rf-PDT group and 140 ± 78 µm and 372 ± 100 µm, respectively, in the control group. There were no significant between-group differences at the initial visit (p = 0.70 for foveal SRF height and p = 0.25 for subfoveal choroidal thickness; Table 1).
Rf-PDT was performed for all affected eyes in the rf-PDT group. The mean diameter of the spot size for rf-PDT before treatment was 5390 ± 1159 µm. No systemic or ocular adverse events were observed following rf-PDT. After rf-PDT, the SRF height and subfoveal choroidal thickness in the rf-PDT group reduced to 33 ± 60 µm and 239 ± 56 µm, respectively. In contrast, in the control group, the SRF height decreased to 80 ± 65 µm; however, the subfoveal choroidal thickness remained unchanged at 370 ± 87 µm after an observational period of 1.2 ± 0.7 months. The difference in the period was due to the fact that the natural history of CSC was monitored at 1-to 2-month intervals, whereas follow-up after treatment was conducted within 1 month. Both the SRF height and subfoveal choroidal thickness in the rf-PDT group were significantly decreased compared with those in the control group at follow-up (p = 0.01 for foveal SRF height, p < 0.001 for subfoveal choroidal thickness; Table 2). Table 3 summarizes the comparisons of choroidal thickness in the nine subfields at baseline and at the followup time point in both groups. Relative to that before rf-PDT, the choroidal thickness after rf-PDT showed a significant decrease in not only all inner subfields but also all outer subfields at the posterior pole, including the vicinity of the vortex vein ampulla (p < 0.001 for all inner subfields; p = 0.035 and p = 0.024 for the outer superonasal and inferonasal subfields, respectively; p < 0.001 and p = 0.004 for the outer superotemporal and inferotemporal subfields, respectively). Contrarily, no significant difference was observed in the choroidal thickness across any subfields in the control eyes between the initial visit and the follow-up point.
Our analysis revealed a significant positive correlation between the rate of reduction in choroidal thickness in the irradiated area and that in the non-irradiated area (correlation coefficient, 0.765; p < 0.001, Supplemental Figure).

Discussion/Conclusion
Accumulating evidence has revealed that both choroidal thickening 6,7,16 21 suggested the efficacy of hd-PDT for acute CSC. Thereafter, various investigators reported the efficacy of hd/rf-PDT for chronic or recurrent CSC 23,29 . Using OCT or ICGA, several studies have shown post-PDT changes in the choroidal structures in eyes with CSC 21,24,25,30 . Using ICGA, Chan et al. 21,24 showed a reduction in the diameter of the macular choroidal vessels as well as the choroidal vascular leakage in CSC following PDT. Maruko et al. 31 showed that subfoveal choroidal thickness and choroidal vascular hyperpermeability were reduced in CSC after hd-PDT. Izumi et al. 25 showed a marked reduction in the thickness of the large choroidal vessel layer, but not of the choriocapillaris-medium choroidal vessel layer, after hd-PDT. However, previous studies focused on the examination of the choroidal changes within the macular area where the PDT laser was irradiated. This study compared the choroidal thickness at the posterior pole before and after rf-PDT in eyes with CSC using EDI of WF SS-OCT. Previous research by Sugano et al. 32 reported that changes in the subfoveal choroidal thickness mirrored those in the choroidal thickness outside the laser-irradiated area after PDT in eyes with polypoidal choroidal vasculopathy. Consistent with the findings of that study, our study found a positive correlation between choroidal thickness changes in the irradiated area and those in the nonirradiated areas (Supplemental Figure). However, our study was different in that it focused on eyes with CSC, and two-dimensional evaluation of the region up to the vicinity of the vortex vein was possible.
Histopathologic evidence demonstrates that PDT selectively occludes the choriocapillaris in targeted areas 33 . With its lower irradiation energy, rf-PDT is presumed to induce even fewer morphological and functional changes within the choriocapillaris 34 . This leads us to propose that the effectiveness of rf-PDT arises from a moderated reduction in choriocapillaris blood flow in the irradiated region, resulting in a similar moderated decrease in the blood flow of the short posterior ciliary artery, given its constant pressure and increased resistance.
Recent studies utilizing WF choroidal thickness maps have shown choroidal thickening from the vortex vein ampulla along the dilated vortex veins in eyes with CSC 12 . This suggests that the pathogenesis of CSC may be associated with impaired drainage of the vortex veins at the sclera. In CSC, it is believed that overload or congestion in the choroid leads to increased permeability at the choriocapillaris, which results in the formation of SRF at the posterior pole 10,16 . In the present study, we observed that rf-PDT resulted in a substantial reduction in choroidal thickening, not only in the irradiated macular area but also outside the vascular arcade (Table 3, Figs. 2, 3).
Considering the substantial blood flow in the macular choriocapillaris 9,35 and its upstream position in the choroidal circulation 36 , it is plausible that it could influence the outflow into the vortex veins across all quadrants. Accordingly, we suggest that rf-PDT reduces SRF in CSC by selectively curtailing blood flow in the Table 2. Longitudinal changes in morphologic and functional parameters in the control group and the reduced-fluence photodynamic therapy group. Data are shown as the mean ± standard deviation unless otherwise indicated. n.a. not applicable, rf-PDT reduced-fluence photodynamic therapy, logMAR logarithm of the minimum angle of resolution.  Table 3. Comparisons of swept-source optical coherence tomography-examined wide-field choroidal thicknesses between baseline and follow-up in the control group and the reduced-fluence photodynamic therapy group. Data are shown as the mean ± standard deviation unless otherwise indicated. rf-PDT reducedfluence photodynamic therapy.  33,34 and by decreasing the volume of blood entering the vortex veins. This action could alleviate vortex vein congestion and reduce SRF at the posterior pole. This study has several limitations. First, the number of patients included in the study was limited. Second, the follow-up period was short although the duration was comparable to a previous report 31 . Third, this study did not determine the precise cause of the observed decrease in the peripheral choroidal thickness. This reduction could be attributed to suppression of the increased permeability of choroidal vessels by PDT. Nevertheless, considering that the reduction in the choroidal thickness was much more extensive than the PDT-applied area, we speculate that this effect might primarily be due to the alleviation of overload at the vortex venous ampulla. Future research is planned to prospectively investigate the actual factors contributing to this extensive reduction in choroidal thickness after rf-PDT. Fourth, because this was a retrospective study, the observational period in the control group was longer than that in the rf-PDT group. Finally, WF SS-OCT images may have been overestimated at the periphery because the measurements were made obliquely to the choroidal interface at the periphery. However, as we compared the same subfields of the same eye before and after treatment, possible distortions in the measurement of choroidal thickness at the periphery did not seem to be significant enough to overturn our conclusions.
In conclusion, eyes with CSC showed WF choroidal thickening from the ampulla of dilated vortex vein to the fovea along the vascular course. However, rf-PDT achieved an extensive reduction in the choroidal thickness not only in the irradiated macular area but also outside the arcade vessels in all quadrants. These findings provide new insights into the pathogenesis of CSC and explain the reasons for the effectiveness of rf-PDT for this condition. This can pave way for the development of best-practice guidelines for the treatment of CSC.

Methods
Patients and methods. This observational study received approval from the Institutional Review Board of Kyoto University Graduate School of Medicine, Kyoto, Japan and adhered to the principles of the Declaration of Helsinki. Written informed consent was obtained from all participants before the study procedures and examinations were initiated.
Patients with treatment-naïve unilateral CSC were enrolled at Kyoto University Hospital between December 2021 and July 2022 (ClinicalTrials.gov identifier, NCT02081339). The diagnosis of CSC was based on the presence of SRF at the posterior pole, hyperfluorescent leakage areas on fluorescein angiography (FA) with RPE damage, and choroidal vascular hyperpermeability appearing as hyperfluorescent in the middle-phase ICGA. Exclusion criteria included previous treatment for CSC, corticosteroid use, and intraocular surgery other than cataract surgery as described previously 28 . Patients with other choroidal neovascularization based on OCT and FA/ICGA results that may affect foveal structures, as well as those with poor-quality OCT images were also excluded. Finally, 20 eyes of 20 consecutive patients were included in the study. In addition, we retrospectively analyzed the chart records of 20 consecutive patients adjusted for age and sex as a control group. The same analytical process applied to the eyes with CSC treated with rf-PDT was employed for the control group.
Each patient underwent a comprehensive ophthalmic examination, including refraction measurement, decimal best-corrected visual acuity (BCVA) measurement, measurement of intraocular pressure and axial length, slit-lamp biomicroscopy, color fundus photography, FA, ICGA, fundus autofluorescence photography, and SS-OCT. Using the Early Treatment Diabetic Retinopathy Study (ETDRS) grid, we defined the subfoveal choroidal thickness as the averaged choroidal thickness in the central grid (1 mm in diameter). Objective refraction was measured with an autorefractor, and the axial length was measured with an interferometer (IOL Master 700; Carl Zeiss Meditec, Dublin, CA, USA). The spherical equivalent was calculated as the sum of the spherical power and half of the cylindrical power. Color fundus photography was performed using a fundus camera system. FA and ICGA were performed using a confocal scanning laser ophthalmoscope as previously described 12 . reduced-fluence photodynamic therapy. In this study, all 20 patients in the rf-PDT group underwent treatment with a standard dose of verteporfin (6 mg/m 2 ) that was intravenously administered over a span of 10 min. Laser treatment was performed exactly 15 min after the drug infusion, with a reduced fluence of 25 J/ cm 2 and an exposure duration of 83 s. The laser was delivered to cover both, areas with the leak point based on FA and the choroidal vascular hyperpermeability on ICGA 29 that had been observed before treatment. In this study, all PDT spots were within a circle with an approximate diameter of 7 mm on the temporal side of the disc. After treatment, patients were instructed to avoid exposure to sunlight for 48 h. rf-PDT was recommended for CSC diagnosed by multimodal imaging, with reference to the PLACE trial 37 . Specifically, rf-PDT was used for eyes with SRF that was observed to affect the fovea on OCT, as well as those with focal or diffuse hyperfluorescent leakage areas, an RPE window defect observed on FA, hyperfluorescent changes on ICGA, and RPE abnormality on fundus autofluorescence.
In this study, patients with CSC who met the criteria for the PLACE study were indicated for rf-PDT. However, in clinical settings, some patients are unable to undergo treatment or refuse treatment for various reasons. Therefore, for each individual case, the final decision was made based on discussions with macular specialists after considering individual patient factors such as BCVA, symptom duration, number of recurrences, the ophthalmic status determined by multimodal imaging data, and consent for treatment.

Evaluations of choroidal thickness by EDI of WF SS-OCT.
We examined the choroidal structure using SS-OCT (Xephilio OCT-S1, Canon Medical Systems, Otawara, Japan) with 1010-1110-nm near-infrared light (scanning laser ophthalmoscope, 780 nm) and a scanning speed of 100,000 A-scans/s. No supplementary equipment or lenses were required for image acquisition. www.nature.com/scientificreports/ To measure changes in choroidal thickness on WF, the scanning area was captured at 20 mm (vertical, 128 B-scans) and 23 mm (horizontal, 1024 pixels) with a 5.3-mm scan depth (1396 pixels) for 3D volumetric data using EDI in SS-OCT. The choroidal thickness was defined as the vertical distance from the Bruch's membrane to the choroidoscleral border which was evaluated by the built-in software supported by artificial intelligence. When automatic machine segmentation of the inner and outer edges of the choroid was performed inappropriately, the segmentation errors were corrected manually as needed.
In this study, the choroidal thickness was measured using a grid consisting of three circles of 3, 9, and 18 mm diameter, respectively (Fig. 1) 12 . The inner and outer rings were defined as the fields enclosed between the 3-and 9-mm circles, and between the 9-and 18-mm circles, respectively. Each ring was divided into four subfields, superotemporal, inferotemporal, superonasal, and inferonasal quadrants, respectively. Measurements were performed over a total of nine subfields together with the central 3-mm ring (Fig. 1). We measured the choroidal thickness in each subfield both before and after rf-PDT during the first visit to the outpatient clinic. For the control group, measurements were taken during the initial visit and the subsequent follow-up visit without treatment.
Where choroidal thickness was measured, we corrected for axial length-related magnification using the measurements of axial length in the modified Littmann formula (Bennett procedure) 12,38 .
In the rf-PDT group, we explored the correlation between the reduction in choroidal thickness in areas irradiated with rf-PDT and that in non-irradiated areas.
Statistical analysis. Statistical analyses were performed using JMP 16 software (SAS Institute Inc., Cary, NC, USA). All values were presented as mean ± standard deviation. The difference in sex distribution between the rf-PDT group and the control group was evaluated using the chi-square test. Other comparisons between the two groups were performed using the Mann-Whitney U test. The mean foveal thickness, SRF height at the fovea, and subfoveal choroidal thickness for each sector were compared between different observational points using the Wilcoxon signed-rank test. The correlation between the reduction in choroidal thickness in areas irradiated with rf-PDT and the reduction in choroidal thickness in non-irradiated areas was evaluated by the Spearman rank test. To quantify the visual changes, a Landolt chart was converted to the logarithm of the minimum angle of resolution (logMAR) BCVA for analysis. Statistical significance was set at p < 0.05.

Data availability
The datasets generated during and/or analyzed during the current study are available from the first author [Naomi Nishigori] on reasonable request.