This research was conducted following the Declaration of Helsinki.
This retrospective study was approved by the Institutional Review Board of Fukushima Medical University (No. 2020-091) and waived the individual consent for this analysis.
In this retrospective comparative study, we reviewed the medical records of 862 Japanese patients diagnosed with unilateral PCV between June 2017 and August 2020 at Fukushima Medical University Hospital, Fukushima, Japan.
The inclusion criterium was diagnosed with unilateral PCV, which evaluated by three ophthalmologists (T.S., Y.S., and A.K.) based on the presence of branching vascular networks, polypoidal lesions, and dilatated aneurysmal lesions on ICGA (TRC 50DX, Topcon, Tokyo, Japan) according to the published criteria.1,2 Two of the participants were allergic to the ICGA dye, so we used indirect ophthalmoscopy to detect orange polypoidal structures for diagnosis. Fifty-seven fellow eyes of the patients diagnosed with unilateral retinal vein occlusion were selected as an age-matched normal control group. All RVO patients received FA for clinical evaluation and were confirmed no abnormality on FA in the contralateral eyes of RVO. All participants in both groups had no evidence of any retinal pathological change in the enrolled eyes.
The exclusion criteria for all eyes were as follows: 1. The eyes with any combined retinal diseases or intraocular diseases, such as macula hole, central serous chorioretinopathy, uveitis, and glaucoma. 2. The patients with any systemic diseases that affect CC circulation: uncontrolled hypertension, diabetes, gout, and neoplasm. 3. The refractive error is greater than -6 diopters. 4. Any observed drusen, RPE abnormality, or segmentation errors on the B-scan of SS-OCT images.
In which 771 patients met the exclusion criteria 1- 3 has been excluded. Then we reviewed the rest 91 patients’ SS-OCT images. Among them, 39 patients met the exclusion criterium 4 and has been excluded.
All the patients who did not meet the exclusion criteria has been enrolled: 52 fellow eyes of 52 patients were PCV fellow eyes group (PCVF). According to the results of the choroidal vascular hyperpermeability (CVH) status, the PCVF was divided into two groups: PCV fellow eyes with CVH (CVH[+]) and PCV fellow eyes without CVH (CVH[-]). And 57 Japanese patients diagnosed with unilateral retinal vein occlusion (RVO) as controls.
All included patients received comprehensive ophthalmic examinations, including indirect ophthalmoscopy, slit-lamp biomicroscopy, color, and red-free fundus photography, SS-OCTA, and ICGA at the first visit. RVO patients did not receive the ICGA for ethical reasons.
The choroidal vascular hyperpermeability (CVH) was defined as hyper fluorescence seen in the mid-phase (10 minutes after dye injected) of the ICGA images, the correspondent area of the OCTA images. (Fig. 1)
The CVH was evaluated by two masked ophthalmologists (H.W. and A.K.). The 5-minute images were as the reference and scored the ICGA images around 10-minute. The scores were defined as follows: All participants of the control group were scored 0. Scored 1 (PCV-): There was no distinguishable CVH in the area. Scored 2 (PCV[+]): There was obvious and strong CVH in the area. If the results were inconsistent, another experienced ophthalmologist (T.S.) will make the judgment.
The subfoveal choroidal thickness (SFCT) was measured by masked ophthalmologists (K.I. and H.S.) as the vertical distance between the RPE and the choroidoscleral border at the center of the fovea on the B-scan of SS-OCT through ImageJ software, version 1.52p (National Institutes of Health, Bethesda, Maryland, USA. https://imagej.nih.gov/ij/index.html).
To analyze the FV, we conducted the binarization of the CC slab and grouped the FV according to our previous research.10 (Fig. 2)
Briefly, we imported the CC slab bounded from 31 to 40μm beneath the RPE reference11 and the correspondent en-face structural image to MATLAB R2019a. The compensation algorithm was used to eliminate the influence of the shadowing effect on the angiography images.12 Then we imported the compensated images into ImageJ and processed the “Phansalkar local threshold” (radius= 5) for binarization.13 After binarization, we processed "Analyze Particles" and “watershed irregular” 14 to remove the noise.
Since the center of the macula of some subjects was displacement from the center of the image (-210~ +455 μm), we performed manual correction to ensure that all images contain the same 5×5mm2 area centered at the fovea. We applied "Analyze Particles" to summarize the FV in each interval.
Statistical calculations were performed by IBM SPSS V.26 for Windows (IBM Co., Armonk, New York, USA). The P-value lower than 0.05 was defined as the statistical significance. The Kolmogorov-Smirnov test was performed to detect the normality of distribution. Because the K-S test of FV number showed significance in most intervals, all variables were reported as median and quartile deviation. The independent samples median test adjusted by the Bonferroni correction has calculated the differences between groups. The independent samples Mann-Whitney U test was conducted to investigate the distribution differences. The Spearman's rank correlation coefficient was performed to evaluate the correlation between the FV and the CVH. The Youden index was used to determine the best cut-off point for the receiver operating characteristic (ROC) curve between the CVH[+] and the controls.
The size of flow void analysis
We compared the number and the distribution of the FV between PCVF and controls in the followed intervals to confirm the difference between the fellow eyes of PCV and normal eyes.
The primary outcome was the number of the flow void in different intervals among PCVF and controls.
We calculated the total area of FV sizes from 400μm2 to 25000μm2 for comparison (FVarea).
The number of the FV was divided into 7 groups.
FVall: The FV sizes from 400μm2 to 25000μm2 (from 16pixels to 1000pixels). FV500: The FV sizes from 400μm2 to 500μm2 (from 16pixels to 20pixels). FV625: The FV sizes from 525μm2 to 625μm2 (from 21pixels to 25pixels). FV750: The FV sizes from 650μm2 to 750μm2 (from 26pixels to 30pixels). FV875: The FV sizes from 775μm2 to 875μm2 (from 31pixels to 35pixels). FV1000: The FV sizes from 900μm2 to 1000μm2 (from 36pixels to 40pixels). FV1125: The FV sizes from 1025μm2 to 1125μm2 (from 41pixels to 45pixels).
The relationship between choroidal vascular hyperpermeability and the flow void
To investigate the relationship between FV and CVH status, we performed Spearman's rank correlation analysis among the FV and CVH status in all intervals.
Then we compared the number of FV between CVH[+], CVH[-] and controls in each interval to confirm the influence of CVH.
Receiver operating characteristic curve analysis
We used binary logistic regression analysis for CVH[+] and controls depending on the results of FV and CVH analysis.
The FV smaller than 750μm2 were grouped every 25μm2 (1 pixel) and subjected to binary logistic regression analysis after standardization.
We performed the receiver operating characteristic curve analysis to find a cut-off value for maximizing sensitivity and specificity to discriminate two groups.