This study aimed to analyse the types and characteristics of rehabilitation specialists’ experience in administering virtual reality (VR) programs using Q methodology as well as provide basic data regarding the introduction of VR programs in rehabilitation therapy. Thirty-three statements were derived based on a literature review and an in-depth interview with rehabilitation specialists. Q-sorting was performed by a Q-sample of 22 rehabilitation specialists with experience administering VR programs. Data were analysed using the QUANL Program. Rehabilitation specialists’ experience with administering VR programs were classified into three types: type 1 was labelled ‘the need to develop VR programs customised for disability level and type’, type 2 was labelled ‘emphasis on experts’ role of paying attention continuously and their experience’, and type 3 was labelled ‘the need to develop safety equipment by disability type’. The study’s findings demonstrate that there are a variety of rehabilitation specialists’ perceptions on their experience administering VR programs. This study is of significance because it suggests ways to improve VR programs for disabled people, with the consultation of rehabilitation specialists. In addition, rehabilitation specialists’ perceptions on VR programs have not been explored before.  An important technology within the fourth industrial revolution is virtual reality (VR). This refers to a human-computer interface in which a specific environment and situation is created to enable users to experience it as if they were interacting with a real environment . RecentlVR has been utilised in fields such as healthcare, education, rehabilitation, industries, and games ,4,5,6,7.The aim for the disabled person is social security practice, with the objectives of ‘independent living’ and ‘normalisation’. Preparation of the disabled person for independence, such as rehabilitation training, is an essential component for living in a community. Through rehabilitation training, the disabled person can have jobs and form social networks via interpersonal interaction at work, which then leads to opportunities for social participation.To live independently, the disabled person can receive a variety of rehabilitation training appropriate to their circumstances in rehabilitation hospitals and centres, as well as an education. Through rehabilitation training to cope with society, the disabled person can prepare themselves for a social life and communicate with others in daily life . DespiteRehabilitation is not a service provided only for the individuals with physical disabilities or organ failure, but a critical element in effective health management for those with acute or chronic pain and functional disorders. As such, rehabilitation should be available to a wider population. Hence, in a broad sense, rehabilitation is a health management strategy and is more effective if it is accompanied with a personalised management strategy and performed with essential assistive devices ,22,23,24It is rehabilitation specialists whose roles consist of championing programs appropriate for disabled individuals and supporting them to improve quality of life, functional rehabilitation, and social participation, aside from physical rehabilitation ,25. RehaAlthough VR programs are presently administered to disabled people and shown to be effective, research has not been conducted to investigate rehabilitation specialists’ (who have an important role in the field of rehabilitation therapy) perception of experience with VR program administration. As a core factor explaining the perception  of experience with VR program administration, the structure of subjectivity and understanding in rehabilitation specialists is necessary for creating appropriate programs for disabled individuals.Q methodology is a useful method to uncover individuals’ characteristics and subjectivity and identify difference among types of individuals. This methodology has been evaluated and found to be useful in classifying, analysing, and gaining an understanding of the structure of subjectivity, with a purpose of hypothesis generation rather than hypothesis test and generalisation ,30. Q meAccordingly, the current study aimed to typify the perception of rehabilitation specialists working at the Korea National Rehabilitation Center (KNRC), regarding their experience of administrating VR programs and to identify the characteristics of each type by using Q methodology. This was to suggest strategies to advance rehabilitation therapy by considering the specialists’ opinions on the utilisation of VR programs and contribute to the improvement of VR program quality.(1)What are the types of subjectivity in rehabilitation specialists’ experience with VR program administration?(2)What are the characteristics of each type of subjectivity in rehabilitation specialists’ experience with VR program administration?Specific research questions were as follows:The purpose of the current study was to identify the types and characteristics of rehabilitation specialists’ subjective perception of VR programs by using Q methodology. Q methodology is a method to uncover and explain individuals’ subjective tendencies and values. It is a simultaneously qualitative and quantitative method to analyse individuals’ operant or self-referent subjectivity  and can Based on the features of Q methodology, this study’s researchers determined that it was the most appropriate method for the current study, the aims of which were to explore a variety of subjective perceptions of experience with VR program administration and identify the types of perception by interpreting diverse intrinsic meanings. Particularly, Q methodology was appropriate in this study because an objective of the methodology is to examine individuals’ subjectivity in viewing a specific subject-matter by studying their implicit beliefs, values, and attitudes .Study participants were rehabilitation specialists with experience administering augmented VR programs  as well as immersive VR programs  10-times or more. Both programs required head mount displays (HMD) for program participation.In the study, according to the Q methodology procedure, a Q-population was constructed and from this population came a Q-sample and then a P-sample to sort the Q-sample . The stuA Q-population refers to Q statements related to the study topic. Q statements are collected primarily through in-depth interviews with experts on the topic, ordinary people, as well as a literature review . To consTo select the Q-sample, 74 statements in the Q-population were iteratively read. Redundant statements were revised and integrated. To ensure the validity of the statements, a Q-methodology expert, a rehabilitation specialist experienced in augmented and immersive VR program administration, and a hands-on worker in rehabilitation with experience in VR program administration were consulted. Of the statements, the 33 which were determined to better reflect the experience were selected for the Q-sample. The 33 statements are shown in P-sample refers to the sample of a small number of participants who know the study topic and perform a Q-sort task . In geneQ-sorting refers to a task the P-sample performs to sort Q-statements using a Q-sort distribution grid . In thisSpecifically, participants (P-sample) were instructed to sort the 33 statements (Q-sample) to the most agreed, the most disagreed, or neutral, based on their subjective judgements and then, fill in the blanks in the Q-sample distribution grid with the statements from the farthest left (−4) or the farthest right (+4) toward the centre (0).Q-sort data obtained from P-sample were converted into scores using a 9-point scale. The coding was performed by assigning 1 point to most disagreed items (−4), 9 points to most agreed items (+4), and 5 points to neutral items (0). The distribution chart of Q classification depends on the number of statements, and if the number of statements is 40 or less, a 9-point scale is generally used . Coded dThe results of principal component analysis using the PC-QUANL Program were as follows. Rehabilitation specialists’ perceptions of experience in VR program administration were sorted into three types as shown in Regarding correlation between types, that is, a measure of the degree of similarity of the types, the correlation coefficient was 0.566 between types 1 and 2, 0.627 between types 1 and 3, and 0.470 between types 2 and 3 .Twelve participants were classified to type 1. They had 13 years of experience, on average. Their perception of experience with VR program administration was as shown in Type 1 agreed the most to the following statements: ‘I think that programs for disabled individuals should be appropriate to their levels’ (z = 1.77), ‘I think that VR programs should be designed to be specific to each disability type’ (z = 1.66), ‘I think that VR programs reflecting the needs of disabled individuals should be developed’ (z = 1.61), and ‘I think that VR programs with a variety of sports contents specifically targeting disabled individuals should be developed’ (z = 1.22).P15 who showed the highest factor weight in type 1, 1.5406, was a physical therapist with 15 years of experience. P15 stated that ‘because the level of disability is different in different individuals, I think that like all other rehabilitation programs, diverse programs reflecting a wide range of disability level should be provided to increase program effect’. P11 (factor weight = 1.2831), an occupational therapist with 11 years of experience, stated that ‘programs designed by considering disability type should be provided to better motivate participants and increase treatment effect. Thus, I think that we need customised programs with the characteristics of each disability type to be taken into account’.Rehabilitation therapists in type 1 perceived that VR programs should reflect disability type, need, and level of disabled individuals and so, this type was labelled ‘the need to develop VR programs customised for disability level and type’.Five participants were classified to type 2. They had 9 years of experience, on average. Their perception of experience with VR program administration is as shown in Type 2 agreed the most to the following statements: ‘I think that sensitivity of experts is essential because patients’ attitude toward program participation varies depending on their condition’ (z = 1.60), ‘I think that therapists with a lot of experience with disabled individuals should provide VR programs’ (z = 1.44), and ‘I think that to motivate disabled individuals to participate in VR programs, instructors should continue to pay attention and communicate with them’ (z = 1.36).P1 who showed the highest factor weight in type 2, 1.7204, was an occupational therapist with 8 years of experience. P1 stated that ‘I think the therapist’s experience is most critical in program application’. P12 (factor weight = 1.1602), a rehabilitation sports instructor with 3 years of experience, said that ‘I think that to achieve the goal of disabled individuals’ participation in sports and exercise, VR programs participation should be consistent and continuous. Hence, I think it the most essential for rehabilitation specialists is to continue to pay attention to induce participants’ interest and select programs designed to achieve the goal’. P12 added that ‘exercise performance and participation enthusiasm vary day by day, depending on the patients’ condition and so, therapists should be sensitive to patient condition and induce active participation’.As shown above, rehabilitation specialists in type 2 were characterised by their emphasis on the expertise and roles of therapists in providing VR programs to disabled individuals and so, type 2 was labelled ‘emphasis on experts’ role in paying attention continuously and their experience’.Five participants were classified to type 3. They had 9 years of experience, on average. Their perception of experience in VR program administration is shown in Type 3 rehabilitation specialists agreed the most to the following statements: ‘I think that VR programs should be designed to be specific to each disability type’ (z = 2.18), ‘I think that equipment easy for disabled individuals to wear should be developed’ (z = 1.38), ‘I think that it is mandatory to provide safety equipment when disabled individuals participate in VR programs’ (z = 1.31), and ‘I think that it will be more effective if equipment specialised for each disability type is provided’ (z = 1.28). P20 who showed the highest factor weight in type 3, 1.0349, and was a rehabilitation sports instructor who stated that ‘safety problem occurs during VR programs for various reasons. I think that there should be a safety system to predict all such situations’ and that ‘after having tried VR program applications, regarding program applicability by disability type, I think that users and instructors both can be satisfied if equipment is developed for each and every disability type’. P5 (factor weight = 0.6823), a rehabilitation sports instructor with 8 years of experience, stated that ‘I predict that the participation rate will be low and disabled individuals will not even try, because I think wearing the equipment causes a hassle or is dangerous’.As shown above, rehabilitation specialists in type 3 had the opinion that the effect of VR programs would be greater if safety equipment and the equipment essential for each disability type were developed and utilised during the programs. In this regard, type 3 was labelled ‘the need to develop safety equipment by disability type’.This study was conducted to explore rehabilitation specialists’ subjective perception of experience in VR program administration using Q methodology and identified the following three types: ‘the need to develop disability-customised programs’, ‘emphasis on experts’ role’, and ‘the need to develop safety equipment’. Based on the findings, major characteristics of rehabilitation specialists’ perception of experience with VR program administration will be discussed pertaining to each research question.First, type 1, labelled ‘the need to develop VR programs customised for disability level and type’, was characterised by the opinions that programs should be provided appropriately for disability type and level and that VR programs should be developed by reflecting the needs of people with disabilities. In addition, rehabilitation specialists in type 1 thought that to increase the program effect, VR programs should reflect participants’ functional level because disability level differs in different individuals and that participants will be motivated to participate in programs in which the type of disability is taken into consideration. In other words, type 1 rehabilitation specialists perceived the need for VR programs to reflect the type and level of disability and the needs of disabled individuals. This finding is in line with a previous study’s findings that the development of programs customised for disability level, the characteristics of disabled individuals, and meeting the demand of the time should be included in VR . AdditioType 1 rehabilitation specialists said that even though programs should be provided based on the consideration of various factors, including participants’ functional level and disability type, current programs administered to the disabled were not specialised for them and so, it was not possible to provide programs customised for disabled individuals. Accordingly, to leverage the strengths of VR programs while decreasing risk and ultimately, to contribute to the development of rehabilitation and social return of people with disabilities via VR programs, the needs of disabled individuals should be identified via usability assessment ) and needs-based VR programs developed. This is consistent with previous studies showing high levels of satisfaction as a solution for rehabilitation and treatment of people with brain lesion and securing positive and effective data for patients through continuous clinical trials and usability evaluation . In addiSecond, type 2, labelled ‘emphasis on experts’ role in paying attention continuously and their experience’, was of opinion that rehabilitation specialists should be sensitive because disabled individuals’ attitude toward program participation varies according to their condition and that VR programs should be administered by highly experienced therapists. In addition, type 2 thought that rehabilitation specialists’ experience was important in VR program application and participation in VR programs required consistency and continuity, accordingly the specialists should continue to pay attention to their patients. In other words, rehabilitation specialists in type 2 seemed to think that the expertise of therapists was essential for administering VR programs to disabled individuals and their role was critical. This finding is consistent with previous study findings that the levels of rehabilitation specialists and service were directly linked with each other . In addiType 2 rehabilitation specialists stated that because the therapists’ experience would impact disabled individuals participating in VR programs, highly experienced and adequately educated therapists should administer the programs. Hence, to improve program quality, rehabilitation specialists should be educated to increase expertise in VR program administration. Furthermore, requirements to nationally register and certify practitioners in order to be qualified and administer programs to people with disabilities should be considered. This is consistent with the findings that the role of competent supervisors and experts plays a decisive role in the rehabilitation of the people with disabilities  and showThird, type 3, labelled ‘the need to develop safety equipment by disability type’, had the opinion that it would be necessary to develop equipment for VR programs for disabled individuals to wear and to introduce equipment customised by disability type. Additionally, type 3 rehabilitation specialists thought that a safety system should be in place to predict safety problems in VR program applications and that users and therapists both would be satisfied if safety equipment was developed by disability type. That is, they perceived that the effect of VR programs would be greater if safety problems that might occur during VR programs could be prevented, and safety equipment customised for each disability type be available. This finding is in line with previous studies findings that users may refuse to wear VR devices or have difficulty wearing them . FurtherRehabilitation specialists in type 3 stated that currently, for various reasons, safety problems occurred during the administration of VR programs and so, both disabled individuals and rehabilitation specialists would be satisfied if safety systems were to be introduced for risk prevention and equipment specialised for each disability type was available. It will be possible to create safer VR programs and specialised for a wide range of disability type, if rehabilitation specialists administering VR programs discuss with VR hardware experts and software experts when developing VR programs.As discussed above, the existence of rehabilitation specialists’ various perceptions demonstrates the importance for the specialists to self-manage, in order to effectively administer VR programs to people with disabilities. Further, it suggests that it is important to examine the extent of using a checklist and that continuous and professional management at organisation level is necessary ,58,59. TIn this study, types, and characteristics of rehabilitation specialists’ subjective perception of experience with VR program administration were analysed by using Q methodology to explore the experience of administering VR programs and provide basic data needed for quality improvement and follow-up research regarding VR programs in rehabilitation therapy.Below, the limitations of the study and suggestions for follow-up studies are discussed.First, the demographic background of the rehabilitation specialists was not sufficiently discussed. Rehabilitation specialists’ subjectivity with respect to the experience with VR program administration may vary according to the area of expertise, level of experience, and the number of times of having administered VR programs. Second, subjective perception types and characteristics of rehabilitation specialists’ experience in VR program administration were identified by using Q methodology and so, the study findings are limited in generalisability, because unlike R methodology, Q methodology assesses subjectivity based on a small sample of participants randomly selected by the researcher . A thirdBased on the current study, it is suggested that follow-up research should be performed by varying rehabilitation specialists’ background, such as the number of times of having administered and provided VR programs and the level of experience. Additionally, it is believed that a dissemination of each of the perception types derived in this study will help rehabilitation specialists provide disabled individuals with high quality VR programs and diverse VR programs can be developed to increase accessibility.This study is significant because the structure of rehabilitation specialists’ subjective perception of experience in program administration was classified and examined with focus on VR programs. The study’s findings suggested ways to improve VR programs and laid a foundation for practical directions in program administration. The study’s findings demonstrate that there are various types of perceptions amongst rehabilitation specialists regarding VR program administration and that diverse needs exist. The researchers made suggestions for responding to each perception type and hope that the findings will be useful in providing basic data to identify disabled individuals’ needs and improve their access to VR programs to advance their rehabilitation therapy.


P = 0.002, both), fellow eyes , and controls . There was no significant difference between the acute CSC eyes, fellow eyes, controls, and 1-year follow-up eyes. As subfoveal choroidal thickness decreased and choroidal vascularity (CVI) increased, the perifoveal density and number of HRF was measured higher with a significant correlation in univariate regression analysis of the acute and resolved CSC eyes . The authors hypothesized that stromal edema induced by choroidal congestion and hyperpermeability has the greatest influence on HRF measurement, possibly affected by inflammatory cells and materials extravasation.The purpose of this study is to evaluate choroidal hyperreflective foci (HRF) changes in central serous chorioretinopathy (CSC) on en-face optical coherence tomography (OCT). Retrospective analysis of 42 patients with unilateral CSC  and 42 age- and sex-matched controls. With 4.5 × 4.5 mm macular scans, structural en-face OCT choriocapillaris (CC) slabs were used to calculate the density and number of HRF in acute CSC eyes with serous retinal detachment (SRD), resolved CSC eyes without SRD, unaffected fellow eyes, control eyes, and 1-year follow-up eyes. Based on the 2-disc diameter (3000 μm), the en-face OCT scan was divided into foveal and perifoveal lesion and analyzed to consider the impact of SRF in HRF measurement. Regression analyses were performed on the several factors with HRF number and density in the acute and resolved CSC eyes. The perifoveal density and number of CC HRF was significantly lower in the resolved CSC eyes when compared to the acute CSC eyes ( In the aggravated case of recurrent episodes, prominent retinal pigment epithelium (RPE) alteration could be observed and lead to choroidal neovascularization (CNV) with persistent SRD3. Spaide4 introduced the starling resistor theory, which can be explained that the structure of noncompliant outershell like brain or eye, that excessively increased blood inflow cannot enlarge vessels anymore, rather constrict the blood flow, leading to maintain suitable volume in the restricted space. In the choroid, vortex vein might perform the role of starling resistor as dilated vortex veins were frequently observed in several CSC studies5. The constriction of resisting point on vortex vein would contribute to choroidal congestion and increased hydrostatic pressure, causing choroidal hyperpermeability. These choroid physics could raise the question of whether the acute CSC sign; SRD, PED, enlarged choroidal vessel and stroma edema, is a series of severe clinical features, or is it a product of a defense mechanism resulting from the operation of a compensatory mechanism to maintain a constant ocular volume. In addition, there could be another question of whether CSC pathophysiology is explained by the movement of serous fluid from increased hydrostatic pressure and permeability, or whether the inflammation is involved in the disease process and progression.Central serous chorioretinopathy (CSC) is a vision-threatening disease characterized by increased choroidal blood flow and hyperpermeability, causing serous neurosensory retinal detachment (SRD), pigment epithelial detachment (PED), or thickened choroid11. In particular, in the study of choroidal HRF in CSC, hypotheses emerged that this would be an exudate of a lipid component or stromal edema12. In other studies of degenerative retinal diseases and normal eyes, possible candidates of HRF were microglial cells or migrated RPE cells, containing lipids or byproducts of photoreceptors11.Hyperreflective foci (HRF) is a characteristic feature that appears in optical coherence tomography (OCT) and en-face OCT, and has already been introduced in several studiesIn the present study, to help understand the CSC pathophysiology, we measured continuously the density and number of choriocapillaris (CC) HRF of acute CSC patients using en-face OCT, and obtained clues about HRF identity and choroidal vascular dynamics.This retrospective observational study was conducted in the Department of Ophthalmology and Visual Science at Seoul St Mary’s Hospital and adhered to the tenets of the Declaration of Helsinki. All protocols were approved by the Institutional Review Board (IRB) of Seoul St. Mary’s Hospital, The Catholic University of Korea (KC22RISI0493). Owing to the retrospective nature and anonymized data of this study, written informed consent procedures were exempted based on the provisions of the IRB of Seoul St. Mary’s Hospital.Forty-two patients who were diagnosed with monocular CSC at our clinic were selected to participate in our study; their corresponding fellow eyes and sex-, age-matched control eyes were also studied. All participants were selected between March 2019 and September 2020 at Seoul St. Mary's Hospital in Korea. A retrospective review of their medical records was performed. The exclusion criteria were as follows: (1) refractive errors of more than spherical equivalent of 6 diopters of either myopia or hyperopia; (2) eyes with a history of any ocular trauma, laser treatment, or intraocular surgery; (3) eyes with a history of intravitreal injections for other ocular diseases; (4) other systemic diseases that could affect the retina except hypertension; (5) presence of other retinal diseases, including glaucoma, age-related macular degeneration, diabetic retinopathy, neurodegenerative disease, or other retinal disease affecting macula; (6) media opacity that could affect image quality; and (7) any history of uveitis.Demographic data, medical history, and ophthalmologic history were recorded at the initial visit. All subjects underwent an ocular examination, including slit-lamp microscopy, dilated fundus examination, OCT, and OCTA. OCT/OCTA imaging was performed using the Topcon DRI Triton SS-OCT device using a 1050-nm wavelength light source, and a scanning speed of 100,000 A-scans/s. The OCT scan protocol was three-dimensional (3D) scan, which consisted of a number of horizontal line scans covering 12 mm × 9 mm rectangular area. 128 B-scans were scanned in total and each B-scan was consisted of 512 A-scans. The OCTA protocol was also 3D scan, covering macular 4.5 mm × 4.5 mm area (320 pixel × 320 pixel) within the height of 4 mm. Acute CSC was diagnosed with OCT scan images when the patient revealed subretinal fluid (SRF) with neurosensory retinal detachment and choroidal thickening without flat irregular PED. The CSC subjects with SRD, not exceeding 3000 μm diameter in the OCT B-scan, were included in the present study. Fluorescein angiography (FAG), indocyanine green angiography (ICG) were performed to confirm typical acute CSC characteristics of hyperfluorescent leaking and choroidal vessel hyperpermeability without any evidence of CNV. Resolutions of SRD within 6 months were confirmed in all subjects regardless of therapeutic intravitreal injections. The presence of visual symptoms in the past day was included into the exclusion criteria to diagnose the first episode of acute CSC. Scans of unaffected fellow eyes and sex- and age-matched normal controls were also used to be compared with those of the acute and resolved CSC eyes. All en-face OCT images and medical records were reviewed by two experienced independent retinal specialists (Y-H.P. and B-E.H.). Two graders made an agreement about whether CSC patients meet inclusion criteria for the analysis of HRFs.13, which were calculated from the OCTARA segmentation algorithm built in the Topcon imageNET software  were applied to the images 15 that calculated all particles of continuous white pixels in the range of 314–2000 μm, the density (%), count (n) of HRF were measured separately within the foveal and the perifoveal lesion, based on the 2-disc diameter (3000 μm)  bigger than 2000 μm to minimalize excessively measured errors due to including scar-like RPE fibrotic areas in the HRF calculation. Consider the diameter of the CC and the size of the intercapillary distance16, hyperreflective areas larger than 2000 μm were likely to be scar lesion than cells or inflammatory materials. Oh et al.6 and Piri et al.7 also defined choroidal HRF as round or oval hyperreflective areas with 10–50 μm in diameter. The CC OCTA images were also binarized with the phansalkar threshold method (15 window radius), also using the “Analyze particle” to calculate all threshold areas bigger than or equal to 1 pixel of flow void, the percentage (%) of CC flow void area were also measured17.All en-face OCT images in this study met an image quality of 60 or higher with no line artifacts or noise. Figure 18. We measured CT manually at the foveal center using digital calipers provided by the SS-OCT software. Using the protocol described by Agrawal et al.19, OCT b-scan images were binarized into black and white pixels with a Niblack auto local threshold. Then, the ratio of luminal area (vessel component) to total choroidal area  was defined as the choroidal vascularity index (CVI). Two experienced independent observers measured the SFCT and CVI, and the average values were used in the analyses to avoid inter-observer variation.Choroidal thickness (CT) was calculated using the automatic built-in software within the SS-OCT device. Subfoveal CT was determined by calculating the distance from the outer border of the RPE to the inner edge of the suprachoroidal spaceP-value. The intraclass correlation coefficient (ICC) analysis was performed to calculate intergrader reliability. Regression analyses were performed on the several possible relative factors with HRF density and number in the acute CSC eyes and resolved CSC eyes.Statistical analysis was performed using the Statistical Package for the Social Sciences for Windows . The mean differences between acute CSC eyes with SRD, resolved acute CSC eye, corresponding fellow eyes, normal control eyes, and 1-year follow-up eyes (n = 20) were assessed using the one-way ANOVA test, followed by post-hoc paired t-test for the HRF density & number. Bonferonni corrections for multiple comparison testing were applied to the significant Owing to the retrospective nature of image analysis and anonymized data of this study, written informed consent procedures were exempted. All protocols were approved by the Institutional Review Board (IRB) of Seoul St. Mary’s Hospital, The Catholic University of Korea (KC22RISI0493).P < 0.0001), which were evaluated as an excellent agreement between two graders20. The variation of HRF density (%) and count (n) were statistically significant  on the comparison of acute CSC eyes with SRD, resolved acute CSC eyes, unaffected fellow eyes, normal control eyes, and 1-year follow-up eyes. On the post hoc analysis, the density and number of perifoveal HRF was lower significantly in the resolved CSC eyes when compared to acute CSC eyes, fellow eyes, controls . The number of perifoveal HRF was significantly higher in 1-year follow-up eyes than in resolved CSC eyes. There was no significant difference between the acute CSC eyes, fellow eyes, controls, and 1-year follow-up eyes  are in Table P < 0.05). . Table , Fig. 3.11; (2) stromal localized cell proliferation like microglial cell, melanocyte, fibroblast6; (3) exudates or macrophage/monocyte extravasation from increased hydrostatic pressure or inflammation21; (4) chronic fibrotic response8. The present study dealt with acute and resolved CSC eyes, the case of (4) might not be a high-probability candidate.Referring to previous papers, choroidal HRF candidates were summarized as follow; (1) migration of RPE cell with lipid components from retina (photoreceptor)6, the number of HRF was higher in the group of lower CVI in the eyes of normal population. It was suggested that the components in the stroma would be HRFs, which were microglial cells, melanocytes, and fibrocytes. In the acute or resolved CSC eyes of the present study, enlarged hydrostatic pressure and choroidal hyperpermeability could contribute to stromal edema. The significant reduction of density and number of HRF at the perifoveal lesion in the resolved CSC eyes might be considered as the dilution of the localized stromal cells, or other inflammatory materials due to sustained stromal edema in the recovery process of CSC. During the SRF resolution, the time difference between the decrease in hydrostatic pressure of congested vessel and the reduction in stromal edema might prevent sudden changes of choroidal volume, which could aggravate the imbalance of autonomic modulation in CSC.In the study of Oh and colleagues22, but the absolute amount of stromal edema would be the largest in CSC, especially in the inner choroid, due to the choroidal hyperpermeability, which could contribute to the HRF reduction.In the acute stage with SRD, we speculated that the increased choroidal blood inflow was combined with choroidal congestion, which would be induced by the starling resistor, maximized choroidal hyperpermeability, accelerating the inflammatory cells or hyperreflective materials extravasation. However, the movement of fluid also considerably aggravated stromal edema, probably not leading to a significantly measured higher in the density and number of HRF in the acute stage, when compared to those of fellow eyes and controls. Sonoda et al. demonstrated that inner choroidal hyperreflective lesion would mean stroma, which was made up of vascular walls, melanocytes, fibrocytes, or neural tissues. They reported that the stromal portion in the inner choroid was significantly larger in CSC eyes than that of fellow eyes and normal control eyes. As the ratio of stroma to the total choroid was relatively lower in CSC23. As microglia reside in the retina, and tissue-resident macrophage or blood-driven monocyte do the similar roles in the choroid24, the alteration of HRF in CC layer from the hemodynamic changes of CSC might be due to the blood-driven inflammatory cell reactions. Regarding lipid exudates, Shinojima et al.9 introduced that hyperreflective areas in OCT corresponded ICG-hypofluorescent areas in CSC patients, suggesting that ICG block was due to accumulation of lipid material, or possibly lipid exudates. Sirakaya et al.25 presented elevated monocyte to high-density lipoprotein ratio in CSC eyes, which meant increased inflammatory and oxidative stress would be remarkably involved in the CSC pathophysiology. However, if the extravasation of lipid exudates or inflammatory cells do more crucial role than the stromal edema from increased hydrostatic pressure with hyperpermeability in the CSC process, it could not be supported by the result that the higher the choroidal thickness, the less HRF was measured in the regression analysis of our study. Similar results were also found in other HRF studies for the negative correlation between choroidal thickness and HRF12. In this regard, it was hypothesized that as the large vessel area and the stromal area increases simultaneously, it could be considered that the movement of fluid or solute is much more dominant than the exudation of inflammatory cells or lipid materials.In addition to the dilution effect of the stromal edema on HRF, the authors also focused on lipid or proteinous exudates, and related blood-driven inflammatory cell reactions for the changes of HRF in the present study. In the studies of retinal hyperreflective foci in diabetic retinopathy, HRFs were considered as an inflammation OCT biomarker, especially predicting microglial cell activation and aggregation19. The positive correlation between HRF and CVI in the regression analysis was in line with the hypothesis that HRF would come from inflammatory extravasation due to choroidal vein congestion and induced-hyperpermeability, the detrimental effect of which was resolved by photodynamic therapy remarkably as shown in the report of Kinoshita et al26. In this respect, the significant correlation result from the perivascular HRF number and CC flow voids of acute CSC  also suggested that HRF increased with hyperpermeability of choroidal vessel despite of the dilution effect of stromal edema. We authors reported the reversible CC flow reduction in acute CSC eye with a novel compensation method17. Several studies also demonstrated CC flow delay or hypoperfusion in CSC with choroidal hyperpermeability28. All of these studies supported the significant relevance between choroidal congestion, choroidal hyperpermeability and CC flow reduction. Nevertheless, further research will be needed because the quantitative measurement of CC flow in acute CSC is still challenging and inaccurate.CVI was known as the parameter for the proportion of choroidal large vessel in Haller’s layer, especially vein7 observed the HRF in choroids of Stargardt disease, hypothesizing the HRF might be lipofucin materials from outer retina and RPE lesion. In the study of retinal degenerative disease29, retinitis pigmentosa, the thicker the choroid, the more HRFs. The opposite results would be because the disease character was different. The HRF of the RP study would be more likely to be lipofucin-related RPE cell migration. In the present study, damaged detached neurosensory retina might induce dysfunction of RPE cell, causing RPE cell migration and lipofucin accumulation in the CC level. Although this hypothesis was not in the line with the result of regression analysis, the result of significant recovery of perifoveal HRF number of 1-year follow-up data would make it possible to speculate that the RPE-lipofucin related mechanism do a partial role in the HRF alteration with the gradual resolution of stromal edema. Through a long-term observational study of the study subjects, the researchers will further clarify the identity of HRFs, focusing on a gradual increasing tendency in chronic process. Additional research, like postmortem or animal studies, will also be needed to clarify this matter.Piri et el.8. This might be because there was no quantitative restriction on the size when measuring HRF, so the fibrotic changes of RPE from repeated SRD attacks could also appear hyperreflective. It would not be of much help in predicting changes in choroid vessel dynamics when fibrotic changes of RPE were involved in the HRF calculations. If chronic CSC was involved in our HRF study, the results would be uncertain because HRFs would more likely be RPE cell migration with photoreceptor damages from recurrent SRD, resembling degenerative disease, which Arrigo et al.21 reported similarly as a high risk factor for macula complications.In the Ruiz-Medrano et al. HRF study, CSC affected eyes, especially chronic ones, tended to have more HRF numbers and hyperreflective fibrous wall thickeningSince we planned to focus on the perifoveal lesion that is not affected by SRF during the research planning stage, we delt with the alteration of perifoveal HRFs in the preceding discussions. In fact, even assuming that the effect of SRF on HRF measurements is minimal, the only significant result of lower HRF number in the foveal lesion in the acute CSC would also present the similar the hypothesis we suggested that the excessive stromal edema would occur more largely than hyperreflective inflammatory cell or material extravasation.5, who presented the geographical correspondence of dilated vortex veins and delayed CC flows. If a wider range of area was analyzed with ICG angiography, more valuable evidences would be presented to explain the change in HRF with choroidal fluid dynamics. Second, a comprehensive analysis for the correlation between the en-face choroidal microvasculature HRF and the whole choroidal HRF on B-scan images was not properly conducted in the present study. The reason for analyzing CC slab in this study was that exudates, inflammatory cell extravasation, and hydrostatic fluid shift with the altered permeability would most clearly appear in the CC layer14. We additionally calculated the density and number of HRF in the entire choroid by applying the same method on OCT b-scan image. However, any significant correlations between the en-face OCT perifoveal CC HRF and the whole choroidal HRF on b-scan image was not observed (Table 10. For this reason, the reference point will be different from the threshold value set in the macula scan of CC. The difference between the axial and lateral resolution also have contributed the irrelevance when calculated in the same automatic way. These technical limitations require the effective way to standardize the difference between the two different image scans and should be discussed through further research. The third limitation was the suitability of mean + 2SD for the image binarization threshold and the range of 314–2000 μm for HRF size. Another method of comparing RPE reflectivity when defining HRF has also been introduced29. It was not apparent to figure out the better automatic method to distinguish whether it is HRF or not in terms of stromal cells, inflammatory cell, or exudates, when only using OCT/OCTA scan and image J tool. Fourth, foveal HRFs were not accurately measured when acute SRD attack occurred. Although we aimed to avoid signal changes from SRF and focused on the perifoveal changes, foveal HRF measurement during acute CSC attack was needed through a more robust method to elucidate CSC pathophysiology more clearly. Fifth, in order to know the exact identity of HRF, these imaging studies will have to be supported by cadaver researches or animal experiments. The unavoidable limitations of image quality in the clinical settings, and the absence of long-term follow up images were other shortcomings in the current study.The present study would be the prior research, giving opportunity to consider the fluid dynamics and inflammatory process in CSC with the alteration of HRF. However, we acknowledge that the exact histopathological correlate to HRF in CSC was not established and all discussions above rely heavily on hypotheses or conjectures. There were some other limitations to this study. The first was that analysis was only performed in just 4.5 × 4.5 mm sized macular area. Regional variation of vasculature in CSC was found in the report of Kishi et al.ed Table . In the In conclusion, although several possibilities remain open for the clear origin of HRF in en-face OCT, we identified the serial changes of HRF in the acute CSC eyes, revealing significantly association with choroidal vascular dynamics. The novel result might give the clinician to get closer to CSC pathophysiology and to consider HRF as parameter for choroidal hyperpermeability and inflammation in eyes with CSC.Supplementary Information.


To describe the development and outcome of mid-phase pinpoint hyperfluorescent spots (MPHS) on fundus fluorescein angiography (FFA) in acute (< 7-day symptom onset) central retinal artery occlusion (CRAO) patients.This retrospective study included acute CRAO patients who underwent multimodal imaging utilizing optical coherence tomography (OCT) and FFA between June 2017 and January 2023. The correlation of FFA images with the OCT images in various stages and severity of CRAO were studied.Twenty-three FFA studies on 23 patients with acute CRAO were included. In 11 (48%) cases, an important finding noted on FFA was the presence of single or multiple MPHS with adjacent minimal late vascular leakage. Of these 11 cases, eight (73%) were males and median age was 41 years (inter quartile range: 33–55 years). Visual acuity ranged from ‘light perception’ to 6/18, and these patients presented anytime on the same day to seven days after symptom onset. On OCT, three eyes had severe CRAO, seven eyes had moderate CRAO, and one eye had mild CRAO. MPHS were primarily observed at the posterior pole and more frequently observed in moderate CRAO severity. During follow-up, the MPHS and retinal vessel staining on FFA disappeared as the CRAO showed signs of resolution.MPHS at the posterior pole on FFA in acute CRAO patients could indicate a more severe occlusion and poor visual outcomes, even after treatment. This finding is most likely caused by red blood cell aggregation or rouleaux formation.Not applicable.The online version contains supplementary material available at 10.1186/s40942-023-00478-5. A clinical diagnosis of CRAO is based on typical patient symptoms such as sudden onset painless vision loss, fundus findings such as extensive retinal whitening and cherry red spot, retinal vessel attenuation, and preservation of optic nerve head perfusion, and OCT characteristics such as inner retinal layer opacification and thickening . In routIn our limited experience with FFA in acute CRAO cases, we observed in the middle phase of the angiogram, pinpoint hyperfluorescent spots primarily at the posterior pole, along with subtle late vascular leakage. There is limited literature on these hyperfluorescent pinpoint spots. To learn more about hyperfluorescent spots on FFA and its association with the severity of CRAO and visual prognosis, we aimed to examine the FFA findings in patients with acute CRAO and a symptom onset duration of less than 7 days. In addition, this study seeks to determine the etiopathogenesis of this imaging finding observed on FFA in acute CRAO.In this study, we retrospectively reviewed the medical records of patients who were clinically diagnosed with acute CRAO with a symptom onset duration of less than 7 days. Acute occlusion of the central retinal artery was defined as a recent history of sudden, painless vision loss accompanied by retinal thickening and whitening, retinal vessel attenuation, and a cherry-red spot in the macula. Patients who visited the retina clinic between June 2017 and January 2023 were included in the study. In addition, only patients who had undergone an invasive fundus fluorescein angiography and OCT imaging were included in the subsequent analysis. In a case of acute CRAO, a FFA was performed at the discretion of the clinician in order to document the arm-retina time and thus estimate the location and severity of occlusion and final visual outcome. In addition, it was performed to rule out partial or total ophthalmic artery occlusion. The clinician considered repeat FFA primarily when the OCT findings worsened or to examine the changes in the previous FFA findings. The local Institutional Review Board/Ethics Committee approved the research because it adhered to the Declaration of Helsinki’s principles. Prior to the procedures, separate informed patient consent was obtained for invasive retinal imaging with FFA.Demographics, associated systemic history, corrected distance visual acuity and time interval between onset of symptoms and presentation to clinic were all recorded during the presentation and follow-up visits. Clinical features were documented using the Optos Optomap Daytona Panoramic 200Tx  or Topcon TRC-50Dx  machines, and OCT was performed using the spectral domain Spectralis . The cSLO-based Spectralis machine was also used for FFA imaging. All patients had a complete hemogram, random blood sugar for diabetes, blood pressure check-up, lipid profile, serum homocysteine levels, carotid doppler ultrasound, cardiac evaluation, and stroke work-up.Macular volumetric assessments were performed using 512 A-scans per line with a 30° scanning area and 25-line horizontal raster scans centered on the fovea. In addition, each eye obtained a 12-line foveal-centered radial scan. The presence of inner retinal layer hyperreflectivity, inner retinal layer thickening and loss of individual inner retinal layer stratification, presence of cystoid macular edema, and neurosensory detachment were all observed on OCT. The severity of acute CRAO was classified based on OCT findings into mild, moderate, and severe occlusion [www.graphpad.com. Only statistical tests related to the analysis of non-parametric data were used in this study. Quantitative data between the 2 groups of cases were analysed using the Mann-Whitney U test. Chi-square test was used to compare the categorical data between 2 groups. P values < 0.05 were considered statistically significant.All data were analysed using GraphPad Prism version 9.5.0 (730) for Windows, GraphPad Software, San Diego, California USA, This retrospective study identified 48 eyes in 48 cases of acute CRAO with a symptom onset duration of less than seven days. In 23 (48%) of the 48 cases with acute CRAO, fluorescein angiography information was available. Eleven (48%) of the 23 eyes with acute CRAO demonstrated the presence of single or multiple, pinpoint hyperfluorescent spots in the mid-phase of the angiogram. We shall henceforth refer to them as mid-phase pinpoint hyperfluorescent spots or MPHS. Table Further, we analysed in detail 15 FFA image sets from 11 cases of acute CRAO with MPHS. Only these 11 acute CRAO patients with MPHS were included in subsequent demographic and ocular data reporting. Eight (73%) males and three 27%) females participated in the study. In 8 (73%) cases, the right eye was affected, and in 3 (27%) cases, the left eye was affected. The median age of the patients was 41 years, and the interquartile range was 33 to 55 years. 82% (n = 9) of patients with CRAO had at least one associated systemic condition, such as diabetes mellitus, valvular heart disease, hypertension, internal carotid artery occlusion, stroke, or hyperhomocysteinemia. Visual acuity ranged from ‘light perception’ to 6/18, and patients presented to the clinic anytime from the same day to seven days after the onset of symptoms. No patient had a significant refractive error, and the anterior segment of all 11 patients was unremarkable. In these cases, FFA was performed on the same day or within two days of presentation to the retina clinic. All the 11 eyes with MPHS showed adjacent minimal late vascular leakage on FFA. Based on OCT classification, three eyes had severe CRAO, seven eyes had moderate CRAO, and one eye had mild CRAO. One patient (case 4) presented with cystoid macular edema and neurosensory detachment, while another patient (case 3) exhibited only a small subfoveal pocket of subretinal fluid. Table 7% femaleFFA was repeated in three cases  during subsequent follow-up visits. The purpose of repeating the angiographies was to compare the FFA image changes with the findings from the previous visit and to correlate them with the OCT image changes as well. Within two weeks, the second round of angiographies were performed on all three patients. In two of the three cases, the MPHS seen on the initial angiographies had disappeared (case 4 and 10) Fig. . At the This study describes the presence of a peculiar FFA finding of MPHS in the eyes of patients with acute CRAO. To the best of our knowledge, no previous literature reports have documented or described this finding. The typical characteristics of these MPHS are their location predominantly at the posterior pole, their prevalence in moderate to severe grades of acute CRAO, and their emergence during the middle phase of the fluorescein angiogram with late staining and subtle adjacent perivascular leakage. These MPHS persist until the severity of occlusion exist and disappear in the resolving stages of CRAO. There are no significant relationships between the presence of MPHS and age of presentation, symptom duration, CRAO severity or arm-retina time delay.In the case of an ophthalmic emergency, such as acute CRAO, the primary goal of treatment is to quickly relieve the obstruction and restore retinal perfusion. As a result of its invasive nature and limited ability to influence the treatment decision, FFA is a relatively uncommon imaging technique in clinical practice. Consequently, retinal imaging findings with FFA in acute CRAO are rarely described in the scientific literature. This could be the primary reason why MPHS on FFA in acute CRAO cases have yet to be described in the literature. Gong et al. identified three types of CRAO patterns on FFA, including poor perfusion, exudation, and mixed patterns, based on the arm-retina time and retinal vascular leakage . These fThe occurrence of multiple hyperfluorescent spots at the posterior pole beginning in the middle phase of the angiogram and extending until the late phase of the acute stages was an interesting and significant finding in the current series. The exact nature of these MPHS is unknown. One may consider them to be true emboli (fibrin-platelet or cholesterol) released from the thrombus responsible for the obstruction of the retinal artery. This may be unlikely, however, in the absence of non-perfusion distal to these hyperfluorescent spots. Red blood cell (RBC) aggregates in the retinal circulation is a possible alternative explanation for these hyperfluorescent spots. In the aftermath of an acute occlusion of the central retinal artery, blood flow distal to the occlusion becomes stagnant or diminished. Blood flow is affected more often as occlusion grades increase. The surface properties of RBCs and fibrinogen concentrations could also lead to RBC clustering . This leThe aggregation of RBCs is a reversible phenomenon. The MPHS in our current series of cases disappeared in subsequent FFA studies as the CRAO showed signs of resolution. This supports our contention that the MPHS seen on FFA are most likely RBC aggregates similar to the ‘box-carring’ of larger retinal arteries seen on clinical examination and FFA. The MPHS could reveal the severity of the occlusion and the visual prognosis. The presence of the MPHS, however, had no effect on the treatment decision.Single or multiple hyperfluorescent spots at the posterior pole are seen in a number of choroidal diseases, including central serous chorioretinopathy, Vogt-Koyanagi Harada syndrome, sympathetic ophthalmia, posterior scleritis, and occasionally due to drusen staining in age-related macular degeneration . Aside fThis study has several limitations. In this study, only a small proportion of cases initially underwent FFA. Second, only patients who had undergone invasive FFA were included in the analysis, which introduces the possibility of selection bias. As the FFA was recommended at the discretion of the clinician and the study design was retrospective, this could not have been avoided. Third, the timing of the FFA was inconsistent in all instances, which could be attributed to its retrospective nature, which is one of its major limitations. Fourth, the FFA result was not correlated with the treatment modality or treatment outcomes; therefore, it does not contribute to the treatment decision. On the other hand, research on this topic is limited. This study describes a novel imaging finding on FFA in acute CRAO cases and offers a possible explanation for its presence. The discovery could be an indirect predictor of visual prognosis in cases of acute CRAO.To summarize, MPHS at the posterior pole on FFA in acute CRAO cases could imply greater occlusion severity and, as a result, poor visual outcomes even after treatment. The most likely pathogenesis for this finding is RBC aggregation. Future research would be needed to determine whether FFA is truly a necessary investigation for acute CRAO.Below is the link to the electronic supplementary material.Supplementary Material 1


Our aim was to investigate the incidence and risk factors for aortic regurgitation (AR) requiring unplanned surgery after transcatheter closure of ventricular septal defect (VSD) in children.Medical records of 876 children with VSD who underwent transcatheter closure from July 2009 to September 2018 in our hospital were retrospectively reviewed. Groups with and without new-onset or increasing AR requiring unplanned surgery were compared. Univariate and multivariate analyses were used to identify the possible risk factors. Smoothing plot and threshold effect analysis were carried out to find the relationship between possible factors and risk of new-onset or increasing AR.p < 0.001), larger ratio between diameter to body surface area , intracristal VSD , and shorter distance from the upper edge of defect to the aortic valve (or the sub-aortic rim)  were risk factors for new-onset or increasing AR requiring unplanned surgery. And, low risk of AR after muscular VSD transcatheter closure was found. An L-shaped nonlinear relationship between the sub-aortic rim and the risk of new-onset or increasing AR was observed, and the risk of new-onset or increasing AR with the sub-aortic rim up to the turning point (2 mm) . With a median time of 7.3 years' follow-up, no new-onset or increasing AR has been found for children who initially did not have unplanned surgery.A total of 29 children (3.3%) underwent unplanned surgery after transcatheter closure owing to new-onset or increasing AR, including 6 children with new-onset AR and 23 children with increasing AR. Multivariate regression analysis revealed that preoperative mild AR  could increase the risk of new-onset or increasing AR requiring unplanned surgery after transcatheter closure of VSD. Ventricular septal defect (VSD) is one of the most common congenital heart diseases, and surgical closure has been recommended as the standard closure strategy . In receMedical records of children who underwent transcatheter VSD closure in Children's Hospital of Chongqing Medical University were retrospectively reviewed from July 2009 to September 2018. Fig. In our study, based on the recommendations for valvular regurgitation evaluation, AR requiring unplanned surgery was identified as a new-onset or increasing AR detected by both transthoracic color Doppler echocardiography  and ascending aorta angiography  during the procedure. Once a new-onset or increasing AR was detected, the procedure of transcatheter closure was interrupted, the occluder was retrieved, and an unplanned surgery would be performed , 10.The legal guardian of each child has been fully informed of the potential risks of transcatheter closure and signed an informed consent form before the procedure. This study was approved by the Ethics Committee of Children's Hospital of Chongqing Medical University.We collected the patient's information from the hospital's electronic medical record system, (1) demographic information: gender, height, weight, age; (2) preoperative data: VSD diameter, VSD type, with or without aortic valve prolapses, with or without AR, with or without arrhythmia; (3) intraoperative data: VSD diameter, VSD type, pulmonary arterial systolic pressure (PASP), occluder size, occluder type, and delivery sheath size.Based upon standard domestic guidelines, the antegrade catheterization procedure was performed in accordance with established standard operation protocols, as previously published , 12, 13.Devices with four subtypes  employed in our study were the modified double-disk VSD occluders made by LifeTech Scientific , Shanghai Shape Memory Alloy , Starway Medical  . The colAspirin  was administrated to all children for 6 months. Follow-up was performed at 1, 3, 6, and 12 months at least after the procedure. Physical examination, electrocardiogram, and TTE were included as follow-up contents.2 or Fisher's exact tests. Continuous variables were expressed as mean ± standard deviations (SD), and the Kolmogorov-Smirnov test was used for the normality test. Independent t tests were used for normally distributed data, while the Mann-Whitney U tests were used for data with a non-normal distribution. Univariate and multivariate logistic regression analyses were used to determine risk factors for AR requiring unplanned surgery after transcatheter VSD closure. Factors significant in the univariate analysis were subsequently included in the multivariate analysis. Smoothing plot and threshold effect analysis were performed to find the relationship between possible factors and risk of new-onset or increasing AR. p < 0.05 was considered as statistically significant. All tests were performed by IBM SPSS 22.0 , R (http://www.R-project.org), and EmpowerStats software .Categorical variables were expressed as counts and percentages and were compared by χA follow-up duration up to 12.4 years and a median time of 7.3 years were conducted. No new-onset or increasing AR, or any unplanned surgery related to cardiology has been found for those children in the non-AR group during our follow-up.2. Comprehensive baseline and procedural data comparing AR group and non-AR group are listed in Table In our center, 1,379 children underwent transcatheter VSD closure were reviewed, and 876 children met inclusion and exclusion criteria were selected. A total of 29 out of 876 children (3.3%) underwent unplanned surgery due to new-onset or increasing AR detected by both echocardiography and angiography, including 6 children with new-onset AR and 23 children with increasing AR, all of which were done at a median time of 4 days (IQR 3–6 days) after transcatheter closure. In the AR group, 51.7% were men, the mean (SD) age was 38.82 (20.54) months, the mean (SD) weight was 14.34 (4.45) kg, and the mean (SD) body surface area (BSA) was 0.62 (0.12) mp < 0.05). Moreover, compared to the patients with perimembranous VSD (pmVSD) (20/841 = 2.4%) and intracristal VSD (9/24 = 37.5%), patients with muscular VSD (0/11 = 0%) had a lower incidence of new-onset or increasing AR requiring unplanned surgery after transcatheter closure.Compared to the non-AR group, patients in the AR group had a higher percentage of leaflet prolapse (69.0 vs. 12.3%), a higher percentage of preoperative mild AR (79.3 vs. 2.4%), a higher percentage of intracristal VSD (31.0 vs. 1.8%), a higher percentage of asymmetric occluder (51.7 vs. 9.6%), a higher ratio of defect diameter (angio)/BSA (7.5 ± 4.0 vs. 5.5 ± 3.0), shorter distance from the upper edge of the defect to the aortic valve (or the sub-aortic rim) (0.9 ± 1.0 vs. 3.5 ± 1.0), higher PASP (34.5 ± 11.7 vs. 30.1 ± 8.7), larger occluder size (7.4 ± 1.9 vs. 6.3 ± 1.5), and larger delivery sheath size (6.8 ± 0.8 vs. 6.3 ± 0.7) than patients did not undergo unplanned surgery , higher ratio of defect diameter (angio)/BSA , intracristal VSD , and the sub-aortic rim  were identified as independent and significant risk factors.Univariate and multivariate logistic regression analyses were performed between groups to identify risk factors for new-onset or increasing AR requiring unplanned surgery after transcatheter VSD closure . When the distance exceeded 2 mm, the distance was not associated with the risk of new-onset or increasing AR .Then, an L-shaped nonlinear relationship between the sub-aortic rim and the risk of new-onset or increasing AR was observed by the smoothing plot , which may result from the leaflet distortion such as instrumental injury, leaflet propped by the occluder, leaflet clamped by the occluder, filamentous hyperemia at the root of the leaflet. Additionally, the ratio of defect diameter (angio)/BSA, the sub-aortic rim, preoperative AR, intracristal VSD were identified as risk factors for AR requiring unplanned surgery.Transcatheter closure in children with preoperative mild AR may further aggravate the damage of aortic valve function or even AR . HoweverThe higher defect diameter/BSA, shorter sub-aortic rim, and intracristal VSD interpret the relative relationship between the defect and the aortic valve from different perspectives. The difference of relative location of VSD determined the focus of preoperative evaluation and closure strategies to a great extent. Well-known, compared to perimembranous VSD and muscular VSD, intracristal VSD was closer to the aortic valve anatomically. Because of the lack of sub-aortic muscle of the ventricular septum, and contact closely between the occluder and aortic valve, the risks of secondary aortic valve injury increased after occluder placing . NotablyThe sub-aortic rim was the intuitive description of the proximity of the VSD. The shorter the distance was, the more likely the occluder interfered with the aortic valve after the double-disk occluder was deployed. The general view of Chinese medical experts is positive in performing procedures in children with a sub-aortic rim >2 mm . ImportaOccluder type was considered to be different in the comparison of general characteristics. Adjusted multivariate analysis did not suggest that occluder-related factors were the possible risk factors for AR requiring unplanned surgery, surprisingly, while according to our understanding and experience from other studies occluder-related factors should be associated with the occurrence of AR , 27, 28.n = 876) in a Chinese pediatric tertiary cardiology specialty. Preoperative mild AR, intracristal VSD, higher ratio between defect diameter (angio)/BSA and shorter sub-aortic rim were found as risk factors for new-onset or increasing AR requiring unplanned surgery after accepting VSD transcatheter closure (n = 29) for those children, and an L-shaped nonlinear relationship between the sub-aortic rim and the risk of new-onset or increasing AR was observed. Some limitations should be considered. This was a single-center retrospective study, lacking more comprehensive perioperative variables and detailed long-term follow-up. Although cardiologists in our center strictly abided by the China Expert Consensus on VSD, the development of technology in the time span and different cardiologists may introduce confounding bias. Pulmonary to systemic blood flow ratio (Qp/Qs) was not presented. It was very unfortunate that we did not find pictures to further present the situation of the aortic valves for these five children with special descriptions of valve conditions. The advantages/disadvantages of an eccentric occluder for AR should be discussed further. And, the long-term implications of VSD occluder devices remained uncertain given that they are relatively new.This study enrolled a large sample of children with VSD /BSA and shorter sub-aortic rim  were identified as risk factors for new-onset or increasing AR requiring unplanned surgery. And, no new-onset or increasing AR has been found in children who did not undergo unplanned surgery after a median follow-up of 7.3 years.The study protocol was reviewed and approved by the Ethics Committee of Children's Hospital of Chongqing Medical University, approval number (202010). Written informed consent from parents/legal guardians for all participants aged under 18 was obtained for the publication of any potentially identifiable images or data included in this article.The authors declare that they have no conflicts of interest.This study was supported by Chongqing Science and Technology Commission Social Innovation and Social Security Special Project (CSTC 2018 jscx-msybX0041).Kaijun Zhang, Penghui Yang, Mi Li, and Ping Xiang: concept and design. Kaijung Zhang, Penghui Yang, Dan Yin, and Ping Xiang: data collection, analysis, and interpretation. Kaijung Zhang and Penghui Yang: drafting of the manuscript. Mi Li, Tiewei Lv, and Ping Xiang: administrative support, supervision, and critical revision of the manuscript. Xiaohua Liang: statistical support. Min Zheng: cardiac ultrasound support. All authors contributed to the article and approved the submitted version.The original contributions presented in this study are included in the article; further inquiries can be directed to the corresponding author.


OLIG2, CHI3L1, TIMP3, TNFAIP2, and TNFAIP6 transcripts were significantly overexpressed in U87-ST8Sia3 compared to U87-MG. RT-qPCR confirmed these results and demonstrated that neuraminidase decreased gene expression in GBM CSC lines. Moreover, neuraminidase drastically reduced ganglioside expression in GBM CSC lines. Neuraminidase, by its pleiotropic action, is an attractive local treatment against GBM.Glioblastoma (GBM) contains cancer stem cells (CSC) that are resistant to treatment. GBM CSC expresses glycolipids recognized by the A2B5 antibody. A2B5, induced by the enzyme ST8 alpha-N-acetyl-neuraminide alpha-2,8-sialyl transferase 3 (ST8Sia3), plays a crucial role in the proliferation, migration, clonogenicity and tumorigenesis of GBM CSC. Our aim was to characterize the resulting effects of neuraminidase that removes A2B5 in order to target GBM CSC. To this end, we set up a GBM organotypic slice model; quantified A2B5 expression by flow cytometry in U87-MG, U87-ST8Sia3 and GBM CSC lines, treated or not by neuraminidase; performed RNAseq and DNA methylation profiling; and analyzed the ganglioside expression by liquid chromatography–mass spectrometry in these cell lines, treated or not with neuraminidase. Results demonstrated that neuraminidase decreased A2B5 expression, tumor size and regrowth after surgical removal in the organotypic slice model but did not induce a distinct transcriptomic or epigenetic signature in GBM CSC lines. RNAseq analysis revealed that  IDH wildtype, is the most common malignant primary tumor of the central nervous system x- ions of the different ganglioside molecular species. This fragment was used for precursor ion scanning, whereby the [M-xH]x- ions of gangliosides were specifically detected . For quantification, data were acquired using monitoring reaction monitoring (MRM). The precursor and product ion pairs for the MRM analysis were selected based on the precursor ion scanning, but some species were not considered in SRM because they stood below the quantification limit . The proportion of each molecular species of a specific ganglioside class was calculated as the ratio of its peak area to the sum of all detected peak areas in this class, every ganglioside class being considered separately. For more information about the developed analytical methods, see Masson et al. [The total lipids were extracted and purified according to Khoury et al. . Liquid n et al. .p < 0.05. Significances: *** p < 0.001, ** p < 0.01, * p < 0.05.The non-parametric Mann–Whitney test was conducted using the GraphPad 5.0 statistical software to analyze flow cytometry, tumor areas and differential gene expression results. All statistical tests were two-sided, and the threshold for statistical significance was In this paper, we have reported that neuraminidase administration in an organotypic slice model reduces tumor size as well as regrowth after removal but is not associated with a transcriptomic or epigenetic signature, although a strong decrease in gene expression involved in stemness, invasion and proliferation occurred. Moreover, neuraminidase administration also induces major changes in cell surface ganglioside expression and therefore sialylation that might influence the interactions between GBM tumor cells and the microenvironment. Although very promising, numerous experiments must be carried out before including neuraminidase in the GBM therapeutic arsenal, especially as local treatment, likely in combination with other drugs.These include, among others, the measurement of the long-term action of neuraminidase, stabilization of the enzyme, and combination with other drugs such as temozolomide. In addition, given the role of sialic acids in both innate and adaptative immune responses, dedicated experiments must be carried out to analyze the role of neuraminidase in these processes.


Androgenic insensitivity syndrome is the most common cause of disorders of sexual differentiation in 46,XY individuals. It results from alterations in the androgen receptor gene, leading to a frame of hormonal resistance, which may present clinically under 3 phenotypes: complete (CAIS), partial (PAIS) or mild (MAIS). The androgen receptor gene has 8 exons and 3 domains, and allelic variants in this gene occur in all domains and exons, regardless of phenotype, providing a poor genotype – phenotype correlation in this syndrome. Typically, laboratory diagnosis is made through elevated levels of LH and testosterone, with little or no virilization. Treatment depends on the phenotype and social sex of the individual. Open issues in the management of androgen insensitivity syndromes includes decisions on sex assignment, timing of gonadectomy, fertility, physcological outcomes and genetic counseling. AR) are found in most individuals with CAIS but in less individuals with PAIS (Androgen Insensitivity Syndrome (AIS) is an X-linked genetic disease and it is the most common cause of disorders of sex development (DSD) in 46,XY individuals . The pheith PAIS .AIS was first described by Morris, in 1953, with the clinical description of 82 female patients with testes but female phenotype and for this reason Morris named the syndrome as testicular feminization . Later, PAIS should be considered in all individuals with atypical genitalia at birth regardless of the degree of external genitalia virilization and MAIS is a possible diagnosis in males with persistent gynecomastia and or infertility .Role of Androgens in Male Fetal Development: androgens are key elements for appropriate internal and external male sex differentiation. After normal testes development, the Leydig cells produce testosterone, which promotes Wolffian duct differentiation into epididymes, vasa deferentia and seminal vesicles . The conAR gene is located at chromosome Xq11-12, is encoded by eight exons and codifies a 919 aminoacids protein  (residues 1-555), a DNA-binding domain (DBD) (556-623 residues), a hinge domain (624-665 residues) and a C-terminal ligand-binding domain (LBD) (666–919 residues)  . The NTDCAIS prevalence in 46,XY males is estimated from 1 in 20.400 to 1 in 99.100 . Except Final height in CAIS is above normal mean female height, probably due to the action of the growth-controlling gene (GCY) located at the Y chromosome . InteresDifferential diagnosis of CAIS includes complete gonadal dysgenesis, Mayer-Rokitanski-Kuster-Hauser syndrome and Mullerian ducts anomalies . BiosyntAR residual function and ranges from proximal hypospadias to micropenis . This condition is due to the hyperexpansion of the CAG repeats (> 38), present in R exon 1 . These pIn AIS the endocrine profile is consistent with androgen resistance characterized by elevated or normal basal serum testosterone levels associated with high serum LH levels . ElevateAR residual function among the mutated receptors between CAIS and PAIS phenotypes, no difference are observed in hormonal levels  can be helpful in order to detect deletions, insertions and duplications in the AR gene (AR mutations described in a website database associated with AIS and prostate cancer (http://www.mcgill.ca/androgendb) and around 600 of them were described in AIS . Defects in the NTD domain are more frequent in CAIS's patients and variants in exons 5 and 6 (that encode LBD) are more frequent in PAIS's patients  have been described but are very rare in AIS  deletion and deletion of entire e in AIS .AR allelic variants resulting in somatic mosaicism are rarely described in AIS  are implicated with this phenomenon. Other possibilities are variations in the level of 5α-reductase type 2 activity resulting in different DHT availability, and the presence of germ-line AR allelic variants at a post zygote stage conferring somatic mosaicism  can be used, in order to increase the penile size and to promote virilization . Maximumin situ – CIS – or Intrabular germ cell neoplasia unclassified – IGCNU). In 2016, the World Health Organization suggested to change the nomenclature of this initial germinative neoplastic lesion from CIS or IGCNU to germ cell neoplasia in situ (GCNIS) (Disorders of sex development are recognized as a risk factor for type II germ cell tumors (GCTs). These tumors are classified as seminomatous and non-seminomatous types . The sem (GCNIS) . GCNIS a (GCNIS) .In CAIS, the risk of GCTs is considered low and related to age . The estPAIS individuals may maintain their germ cells because of the presence of residual androgen receptor responsiveness, differently of CAIS . TherefoIn patients who maintained the gonads, a careful monitoring including ultrasonography (US) or MRI has been suggested . Due to A normal androgen receptor is necessary for normal male reproduction, because testosterone and FSH, are essential factors for male spermatogenesis. Therefore, mutations in the androgen receptor gene have been searched in order to identify possible causes for male infertility. As previously described, infertility may be the only clinical manifestation of undervirilization in MAIS phenotype ,51.The strategy to obtain fertility in AIS individuals has not been defined yet . In CAISAR variant allelics, were found in male individuals with preserved fertility  causing improvement in sperm count. The gonadotropin concentrations remained unaffected and intracytoplasmic sperm injection with a single sperm directly into an egg resulted in proved fertility  can be an indicative of fertility success, but should be tested in further studies as well as the use of aromatase inhibitors and clomiphene citrate to obtain fertility in these patientsPsychological support is essential for AIS individuals and their parents, in general . DialoguThe gender identity, gender role and sexual orientation show a female pattern in CAIS individuals. In PAIS patients, in general, gender identity aligned with both sex of rearing male or female .Gender change is very rarely described in CAIS and there are just four cases of gender change in individuals with CAIS . TherefoAlthough there is no inconsistency in gender identity, male PAIS individuals show disappointment with undervirilization signs. The absence or paucity of facial and body hair, the high-pitched voice compromised their self-perception of manhood . In femaAR defect. It has an X-linked inheritance and affects 50% of the male offspring. In CAIS, the diagnosis can be done intrauterus, at birth, childhood or after puberty. In PAIS, the diagnosis is usually at birth due to the atypical external genitalia. In MAIS, the diagnosis should be considered in cases of pubertal gynecomastia and male infertility. AR defects are found along AR gene in all AIS phenotypes. Non-synonymous point mutations are the commonest AR defects reported in AIS. Molecular diagnosis is achieved in almost all patients with CAIS and in a lower frequency in PAIS individuals. AIS is characterized by elevated serum LH and testosterone. In CAIS, there is a low risk of GCTs before puberty and postponing surgery to after puberty may allow the development of spontaneous puberty. In PAIS there is a risk of GCTs in 15% of the patients, and bilateral gonadectomy is recommended at childhood in all individuals raised in the female social sex. For males with PAIS, the testis should be placed in the scrotum and regularly monitored. Fertility was described in one PAIS individuals, and therapeutic strategy for successful fertility could be experienced in PAIS and MAIS individuals. In AIS, gender identity usually follows the sex of rearing, but quality of sexual life, sexual functioning and quality of life can be slightly compromised and are important issues for keeping patients in psychological care.AIS is the most common molecular diagnosis in newborns with 46,XY DSD and results of an


This article has been corrected at the request of the authors to correct the affiliation for Mohammed Khashab from Orthopedic Surgery, King Abdulaziz Medical City, Jeddah, SAU to:1. Orthopedics, King Abdulaziz Medical City, Jeddah, Saudi Arabia2. Research Office, King Abdullah International Medical Research Center, Jeddah, Saudi Arabia.3. Orthopedics, College of Medicine, King Saud bin Abdulaziz University for Health Sciences, Jeddah, Saudi ArabiaThe authors deeply regret that these errors were not identified and addressed prior to publication.