Objective : To investigate the effect of laminarin(LAM) on nonproliferative diabetes retinopathy by high throughput sequencing(RNA-seq). Methods : The diabetes model was established by intraperitoneal injection of streptozotocin(STZ), and the effect of LAM on diabetic mice was observed.C57BL/6 mice were randomly divided into three groups: Control group, Model group, and LAM group, with 8 mice in each group. After 8 weeks of modeling, the LAM group received a 4-week intraperitoneal injection of LAM treatment. Changes in blood glucose and body weight of the three groups of mice were recorded, HE staining was performed to examine retinal lesions, and RNA-seq was used to identify differentially expressed genes(DEGs) in diabetic retinopathy(DR) under the action of STZ and LAM. Results : STZ successfully established the model of DR, and LAM reduced the blood sugar in diabetic mice to a certain extent and improved the pathological morphology of retinal structural looseness in diabetic mice. After RNA-seq analysis of DEGs, it was found that there were a total of 214 DEGs in the retina of the Model group mice compared to the Control group. Enrichment analysis revealed that DR could exacerbate the lesions through the PI3K Akt signaling pathway. There were a total of 42 DEGs in the retina of the Model group and LAM group mice, and enrichment showed that LAM improved the lesions through the neutrophil extracellular trap pathway. Early growth response factor 1(Egr1), FBJ osteosarcoma oncogene(Fos), nuclear receptor subfamily 4A member 1(Nr4a1), and salt-induced kinase 1(Sik1) were regulated by STZ, and LAM significantly regulated their expression, which might be closely related to LAM′s treatment of diabetic retinopathy. Conclusion DEGs can exacerbate the severity of diabetic retinopathyviathe PI3K-Akt signaling pathway. LAM can mitigate diabetic retinopathyviathe neutrophil extracellular trap pathway. Egr1, Fos, Nr4a1, and Sik1 are key genes involved in LAM treatment of STZ-induced DR.

Objective:To screen differentially expressed genes in the retinas of mice with experimental autoimmune uveitis (EAU) through transcriptome sequencing.Methods:Twenty 8-week-old SPF grade male C57BL/6J mice were randomly divided into EAU group and control group by random number table method, with 10 mice in each group.In the EAU group, an emulsion of interphotoreceptor retinoid-binding protein 1-20 and complete Freund adjuvant was injected subcutaneously into the inner side of the hind limbs and intravenously into the tail vein of the mice, and pertussis toxin was injected intraperitoneally to establish the EAU model.In the control group, mice were subcutaneously injected with the corresponding volume of phosphate buffer solution.On the 21st day after model establishment, the anterior segment inflammation of the mouse eyes was examined using a slit-lamp microscope.Retinal blood vessels were observed by fundus photography and clinical inflammation was scored.Hematoxylin-eosin staining was used to observe the morphological changes in the retinal tissues of the two groups.Retinal tissues were collected from two groups of mice.RNA was extracted and cDNA libraries were constructed for transcriptome sequencing.The clusterProfiler package in R programming language was used to screen for differentially expressed genes in EAU, and to conduct GO and KEGG enrichment analyses.The expression of core differentially expressed genes in the retinal tissues was then verified using real-time fluorescent quantitative PCR.The breeding and use of experimental animals were in compliance with the relevant regulations of the Animal Ethics Committee of Anhui Medical University.The study protocol was approved by the Ethics Committee of Anhui Medical University (No.LLSC20221208).Results:On the 21st day after model establishment, the clinical score of the EAU group was 2.83±0.94, significantly higher than 1.89±0.93 of the control group, and the difference was statistically significant ( t=2.290, P<0.05). The retinal blood vessels of mice in the EAU group showed tortuosity and dilation.Histopathological examination revealed a large number of inflammatory cell infiltrations in the retina, indicating successful establishment of the EAU mouse model.Transcriptome sequencing of the mouse retinal tissues identified a total of 32 473 genes, and 154 differentially expressed genes were screened out.Among them, 79 genes were up-regulated with Gm38574, Tmem81, Ptpn7 being top three and 75 genes were down-regulated with Gm19802, Fga, Cyp3a11 being top three.GO enrichment analysis showed that the differentially expressed genes were mostly related to immune response and cytokine action.KEGG pathway enrichment analysis showed that the differentially expressed genes were mainly enriched in the Toll-like pathway and cytokine signaling pathway.Compared with the control group, the relative expression levels of Gm38574, Tmem81, and Ptpn7 in the retinal tissues of the EAU group were up-regulated, while the expression levels of Gm19802, Fga, and Cyp3a11 were down-regulated, and the differences were statistically significant ( t=9.755, 5.358, 6.289, 4.312, 6.577, 6.118; all P<0.05). Conclusions:There are significant differences in the key genes Gm38574, Tmem81, Ptpn7, Gm19802, Fga, and Cyp3a11 in the retinal tissues of EAU mice, mainly corelated with the cytokine interaction pathway, Toll-like receptor signaling pathway.

Objective:To screen differentially expressed genes in the retinas of mice with experimental autoimmune uveitis (EAU) through transcriptome sequencing.Methods:Twenty 8-week-old SPF grade male C57BL/6J mice were randomly divided into EAU group and control group by random number table method, with 10 mice in each group.In the EAU group, an emulsion of interphotoreceptor retinoid-binding protein 1-20 and complete Freund adjuvant was injected subcutaneously into the inner side of the hind limbs and intravenously into the tail vein of the mice, and pertussis toxin was injected intraperitoneally to establish the EAU model.In the control group, mice were subcutaneously injected with the corresponding volume of phosphate buffer solution.On the 21st day after model establishment, the anterior segment inflammation of the mouse eyes was examined using a slit-lamp microscope.Retinal blood vessels were observed by fundus photography and clinical inflammation was scored.Hematoxylin-eosin staining was used to observe the morphological changes in the retinal tissues of the two groups.Retinal tissues were collected from two groups of mice.RNA was extracted and cDNA libraries were constructed for transcriptome sequencing.The clusterProfiler package in R programming language was used to screen for differentially expressed genes in EAU, and to conduct GO and KEGG enrichment analyses.The expression of core differentially expressed genes in the retinal tissues was then verified using real-time fluorescent quantitative PCR.The breeding and use of experimental animals were in compliance with the relevant regulations of the Animal Ethics Committee of Anhui Medical University.The study protocol was approved by the Ethics Committee of Anhui Medical University (No.LLSC20221208).Results:On the 21st day after model establishment, the clinical score of the EAU group was 2.83±0.94, significantly higher than 1.89±0.93 of the control group, and the difference was statistically significant ( t=2.290, P<0.05). The retinal blood vessels of mice in the EAU group showed tortuosity and dilation.Histopathological examination revealed a large number of inflammatory cell infiltrations in the retina, indicating successful establishment of the EAU mouse model.Transcriptome sequencing of the mouse retinal tissues identified a total of 32 473 genes, and 154 differentially expressed genes were screened out.Among them, 79 genes were up-regulated with Gm38574, Tmem81, Ptpn7 being top three and 75 genes were down-regulated with Gm19802, Fga, Cyp3a11 being top three.GO enrichment analysis showed that the differentially expressed genes were mostly related to immune response and cytokine action.KEGG pathway enrichment analysis showed that the differentially expressed genes were mainly enriched in the Toll-like pathway and cytokine signaling pathway.Compared with the control group, the relative expression levels of Gm38574, Tmem81, and Ptpn7 in the retinal tissues of the EAU group were up-regulated, while the expression levels of Gm19802, Fga, and Cyp3a11 were down-regulated, and the differences were statistically significant ( t=9.755, 5.358, 6.289, 4.312, 6.577, 6.118; all P<0.05). Conclusions:There are significant differences in the key genes Gm38574, Tmem81, Ptpn7, Gm19802, Fga, and Cyp3a11 in the retinal tissues of EAU mice, mainly corelated with the cytokine interaction pathway, Toll-like receptor signaling pathway.

Objective:To investigate the relationship between three-dimensional choroidal vascularity index (3D CVI) and the severity of diabetic retinopathy (DR) using swept-source optical coherence tomography angiography (SS-OCTA).Methods:A cross-sectional study was conducted.A total of 139 eyes of 139 subjects, including 115 eyes with diabetes mellitus and 24 control eyes without diabetes, were enrolled in the Second People's Hospital of Hefei from March to December 2022.DR was graded according to the standard seven-field ETDRS color fundus photographs.Eyes with diabetes mellitus were divided into non-DR (NDR) group (34 eyes), non-proliferative DR (NPDR) group (42 eyes), NPDR with diabetic macular edema (DME) group (21 eyes) and PDR group (18 eyes).3D CVI in central fovea 1 mm (C1) and parafoveal 3 mm (C3), choroidal vascular volume (CVV), and choroidal thickness were measured by SS-OCTA in the area of 3 mm×3 mm centered on the fovea using the built-in automated quantification software.Parafoveal region was divided into superior, inferior, temporal and nasal quadrants, and 3D CVI of the different quadrants was detected.3D CVI was defined as the ratio of CVV to total choroidal volume.The monocular data were analyzed to compare 3D CVI among the five groups, and multiple linear regression analysis was used to evaluate the influencing factors.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of the Second People's Hospital of Hefei, Hefei Hospital Affiliated to Anhui Medical University (No.2022064).All subjects were aware of the study purpose and agreed to participate the study.Results:There were significant differences in 3D CVI-C1 and 3D CVI-C3 among all groups ( F=3.103, 3.036, both at P<0.05).In PDR group, 3D CVI-C1 was lower than in non-DR group, and 3D CVI-C3 was lower than in control group and non-DR group, with statistically significant differences (all at P<0.05).There were significant differences in 3D CVI in the inferior and nasal quadrants among all groups ( F=2.714, 4.020, both at P<0.05).In PDR group, 3D CVI in the inferior quadrant was lower than that in non-DR group, and 3D CVI in nasal quadrant in PDR group was lower than that in control group, non-DR group, NPDR group and NPDR with DME group, with statistically significant differences (all at P<0.05).Multiple linear regression showed that after controlling for age, course of disease and glycosylated hemoglobin, the severity of DR was the influencing factor of 3D CVI in fovea and parafovea.PDR eyes had the greatest impact on 3D CVI in fovea and parafovea.Compared with non-DR eyes, there was a -0.019(95% CI: -0.031--0.007, P=0.003) decrease in central foveal 3D CVI and a -0.019(95% CI: -0.030--0.008, P=0.001) decrease in parafoveal 3D CVI in PDR eyes, followed by a 0.014(95% CI: -0.027-0.000, P=0.044) decrease in central foveal 3D CVI in NPDR with DME eyes. Conclusions:Macular foveal 3D CVI correlates with DR severity, and a decrease in 3D CVI of large vessels in the macular choroid may be a sensitive indicator of DR exacerbation.

Objective: To explore the damaging effect of the IP3-protein kinase C pathway on human retinal vascular endothelial cells under high glucose conditions. Methods: Human retinal vascular endothelial cells in the logarithmic growth phase were taken and divided into the experimental group(10 mmol/L glucose and 20 mmol/L glucose) and the control group(5 mmol/L glucose). The morphological changes of cells were observed under different concentrations of glucose culture medium. The apoptosis of human retinal vascular endothelial cells was detected by flow cytometry. Real-time fluorescent quantitative reverse transcription polymerase chain reaction was used to detect the RNA expression level of PKC in the IP3-protein kinase C(PKC) pathway, and Western blot was used to detect the protein expression level of PKC in cells. Results: Under high glucose conditions, the volume of human retinal vascular endothelial cells increased, the degree of extension decreased, and the apoptosis rate increased. Compared with the control group, the apoptosis rate of human retinal vascular endothelial cells in the 20 mmol/L glucose group increased(P<0.05); the IP3 level of human retinal vascular endothelial cells in the 20 mmol/L glucose group(587.9±15.2) ng/ml was lower than that of the control group(738.9±1.0) ng/ml(P<0.05). Under high glucose treatment, the expression levels of PKC mRNA and protein increased(P<0.05). Conclusion The IP3-PKC pathway may be involved in the damage process of human retinal vascular endothelial cells under high glucose conditions, and thus may play a role in diabetic retinopathy.

Objective To identify possible associated genetic variants and characterise the clinical presentation of isolated ectopia lentis (IEL).Methods Forty-eight members with 5 generations of an IEL family were enrolled in this study.Peripheral blood samples of all members were collected, and clinical manifestations were observed through physical examination and routine ophthalmological examination.Whole-exome sequencing (WES) was per-formed for two patients to identify disease-causing variants.The target variants were verified by Sanger sequencing in family members and 200 normal controls.Then, candidate variants were verified using Sanger sequencing in family members and 200 healthy controls.SIFT, PolyPhen and MutationTester were used to predict the protein function.Results A total of 13 IEL patients in this family which inherited in an autosomal dominant pattern.The mean age at disease onset was 51.5 years.The main clinical phenotype of this ICE was characterised by ectopia lentis which anterior inclinated to the anterior chamber.As the anterior chamber became shallow, and the angle of the chamber became narrow, and eventually resulted in the secondary glaucoma.A heterozygous missense variant in the fibrillin gene-1 (FBN1) gene (c.3463G>A) was identified by WES, which was present in all patients but was absent in 200 healthy controls.SIFT, PolyPhen and MutationTester predicted that the variant affected protein function.Conclusion This IEL family is characterized by secondary glaucoma as the first symptom which is caused by ectopia lens with inclination.The c.3463G>A of FBN1 gene may be the pathogenic mutation leading to IEL in this family.

Objective To identify possible associated genetic variants and characterise the clinical presentation of isolated ectopia lentis (IEL).Methods Forty-eight members with 5 generations of an IEL family were enrolled in this study.Peripheral blood samples of all members were collected, and clinical manifestations were observed through physical examination and routine ophthalmological examination.Whole-exome sequencing (WES) was per-formed for two patients to identify disease-causing variants.The target variants were verified by Sanger sequencing in family members and 200 normal controls.Then, candidate variants were verified using Sanger sequencing in family members and 200 healthy controls.SIFT, PolyPhen and MutationTester were used to predict the protein function.Results A total of 13 IEL patients in this family which inherited in an autosomal dominant pattern.The mean age at disease onset was 51.5 years.The main clinical phenotype of this ICE was characterised by ectopia lentis which anterior inclinated to the anterior chamber.As the anterior chamber became shallow, and the angle of the chamber became narrow, and eventually resulted in the secondary glaucoma.A heterozygous missense variant in the fibrillin gene-1 (FBN1) gene (c.3463G>A) was identified by WES, which was present in all patients but was absent in 200 healthy controls.SIFT, PolyPhen and MutationTester predicted that the variant affected protein function.Conclusion This IEL family is characterized by secondary glaucoma as the first symptom which is caused by ectopia lens with inclination.The c.3463G>A of FBN1 gene may be the pathogenic mutation leading to IEL in this family.

Objective To identify possible associated genetic variants and characterise the clinical presentation of isolated ectopia lentis (IEL).Methods Forty-eight members with 5 generations of an IEL family were enrolled in this study.Peripheral blood samples of all members were collected, and clinical manifestations were observed through physical examination and routine ophthalmological examination.Whole-exome sequencing (WES) was per-formed for two patients to identify disease-causing variants.The target variants were verified by Sanger sequencing in family members and 200 normal controls.Then, candidate variants were verified using Sanger sequencing in family members and 200 healthy controls.SIFT, PolyPhen and MutationTester were used to predict the protein function.Results A total of 13 IEL patients in this family which inherited in an autosomal dominant pattern.The mean age at disease onset was 51.5 years.The main clinical phenotype of this ICE was characterised by ectopia lentis which anterior inclinated to the anterior chamber.As the anterior chamber became shallow, and the angle of the chamber became narrow, and eventually resulted in the secondary glaucoma.A heterozygous missense variant in the fibrillin gene-1 (FBN1) gene (c.3463G>A) was identified by WES, which was present in all patients but was absent in 200 healthy controls.SIFT, PolyPhen and MutationTester predicted that the variant affected protein function.Conclusion This IEL family is characterized by secondary glaucoma as the first symptom which is caused by ectopia lens with inclination.The c.3463G>A of FBN1 gene may be the pathogenic mutation leading to IEL in this family.

Objective To identify possible associated genetic variants and characterise the clinical presentation of isolated ectopia lentis (IEL).Methods Forty-eight members with 5 generations of an IEL family were enrolled in this study.Peripheral blood samples of all members were collected, and clinical manifestations were observed through physical examination and routine ophthalmological examination.Whole-exome sequencing (WES) was per-formed for two patients to identify disease-causing variants.The target variants were verified by Sanger sequencing in family members and 200 normal controls.Then, candidate variants were verified using Sanger sequencing in family members and 200 healthy controls.SIFT, PolyPhen and MutationTester were used to predict the protein function.Results A total of 13 IEL patients in this family which inherited in an autosomal dominant pattern.The mean age at disease onset was 51.5 years.The main clinical phenotype of this ICE was characterised by ectopia lentis which anterior inclinated to the anterior chamber.As the anterior chamber became shallow, and the angle of the chamber became narrow, and eventually resulted in the secondary glaucoma.A heterozygous missense variant in the fibrillin gene-1 (FBN1) gene (c.3463G>A) was identified by WES, which was present in all patients but was absent in 200 healthy controls.SIFT, PolyPhen and MutationTester predicted that the variant affected protein function.Conclusion This IEL family is characterized by secondary glaucoma as the first symptom which is caused by ectopia lens with inclination.The c.3463G>A of FBN1 gene may be the pathogenic mutation leading to IEL in this family.

Objective To identify possible associated genetic variants and characterise the clinical presentation of isolated ectopia lentis (IEL).Methods Forty-eight members with 5 generations of an IEL family were enrolled in this study.Peripheral blood samples of all members were collected, and clinical manifestations were observed through physical examination and routine ophthalmological examination.Whole-exome sequencing (WES) was per-formed for two patients to identify disease-causing variants.The target variants were verified by Sanger sequencing in family members and 200 normal controls.Then, candidate variants were verified using Sanger sequencing in family members and 200 healthy controls.SIFT, PolyPhen and MutationTester were used to predict the protein function.Results A total of 13 IEL patients in this family which inherited in an autosomal dominant pattern.The mean age at disease onset was 51.5 years.The main clinical phenotype of this ICE was characterised by ectopia lentis which anterior inclinated to the anterior chamber.As the anterior chamber became shallow, and the angle of the chamber became narrow, and eventually resulted in the secondary glaucoma.A heterozygous missense variant in the fibrillin gene-1 (FBN1) gene (c.3463G>A) was identified by WES, which was present in all patients but was absent in 200 healthy controls.SIFT, PolyPhen and MutationTester predicted that the variant affected protein function.Conclusion This IEL family is characterized by secondary glaucoma as the first symptom which is caused by ectopia lens with inclination.The c.3463G>A of FBN1 gene may be the pathogenic mutation leading to IEL in this family.