DNMT1 driven by mouse amniotic fluid mesenchymal stem cell exosomes improved corneal cryoinjury via inducing microRNA-33 promoter DNA hypermethylation modification in corneal epithelium cells.

Severe corneal cryoinjury can cause permanent corneal swelling and bullous keratopathy, one of the main reason for loss of sight. Mouse amniotic fluid mesenchymal stem cells (mAF-MSCs) can repair corneal damage caused by freezing; however, whether the exosomes derived from mAF-MSCs have the same repair effect is unknown. In this study, the mAF-MSC-exosomes were transplanted into the eyeballs of corneal cryoinjured mice. Histopathological examination showed that the mAF-MSC-exosomes improved the corneal structure and status of corneal epithelial cells in corneal cryoinjured mice. RRBS-sequencing showed that compared with the control group, four genes (Rpl13-ps6, miR-33, Hymai, and Plagl1), underwent DNA hypermethylation modification after mAF-MSC-exosomes treatment. The result of FISH indicated that miR-33-3p hybridization signals were enhanced in corneal epithelial cells from mice treated with mAF-MSC-exosomes. Semi-quantitative PCR and western blotting indicated that mAF-MSC-exosomes contained high levels of DNMT1 mRNA and protein. Additionally, luciferase report assays indicated that miR-33-3p overexpression in NIH-3T3 mouse embryonic fibroblast cells inhibited the activity of luciferase carrying a sequence from the 3' untranslated region of Bcl6. Moreover, BCL6 mRNA and protein levels in corneal tissues from mice treated with mAF-MSC-exosomes were higher than those in the control group. Therefore, our results suggested that mAF-MSC-exosomes could repair corneal cryoinjury by releasing DNMT1, which induced hypermethylation of the miR-33 promoter in corneal epithelial cells. Consequent downregulated miR-33 transcription upregulated Bcl6 expression, ultimately achieving the repair of corneal cryoinjury in mice.

Prevalence of dry eye disease among Chinese high school students during the COVID-19 outbreak.

PURPOSE: The study aimed to investigate the prevalence of dry eye disease (DED) and relevant risk factors among Chinese high school students during the COVID-19 outbreak. METHODS: A cross-sectional study was conducted from November to December 2020, and 4825 high school students from nine high schools in Shanghai were recruited. All students completed ocular surface disease index (OSDI) and perceived stress scale (PSS) questionnaires and answered other questions designed to ascertain information on the risk factors related to DED. DED was diagnosed when OSDI scores were greater than or equal to 13. The prevalence of symptomatic DED was determined. A T-test, Kruskal-Wallis test, Chi-square test, and logistic regression analysis were used to examine the possible risk factors. RESULTS: The prevalence of symptomatic DED among Chinese high school students was 70.5%. In univariate analysis, higher PSS scores (P<0.001), prolonged video display terminal (VDT) use (P<0.001), wearing contact lenses (P=0.001), poor sleep quality (P<0.001), and being female (P<0.001) were significantly correlated with dry eyes. In multivariate logistic regression analysis, higher PSS scores (P<0.001, OR=1.20), prolonged VDT use (P<0.001, OR=1.07), poor sleep quality (P<0.001, OR=1.84), and being female (P=0.001, OR=1.25) were significant risk factors associated with DED. CONCLUSIONS: Due to the epidemic, most Chinese high school students are in a high-risk environment in which they are more likely to suffer from DED, such as long online courses and heavy stress from school. Relevant preventive measures that may have a positive impact on public health and quality of life for high school students should be brought to the forefront.

Amniotic fluid mesenchymal stem cells repair mouse corneal cold injury by promoting mRNA N4-acetylcytidine modification and ETV4/JUN/CCND2 signal axis activation.

Severe corneal injury is one of the main causes of loss of visual function. Mesenchymal stem cells (MSCs) have the ability to repair damaged cells in vivo. The present study aimed to explore whether MSCs could function as a cell therapy tool to replace traditional methods to treat corneal injury. CD44 + /CD105 + mesenchymal stem cells isolated from mouse amniotic fluid (mAF-MSCs) were injected into mice after cryoinjury to induce corneal endothelial cell injury. Histopathological assays indicated that mAF-MSCs could promote the growth of corneal epithelial cells, reduce keratitis, and repair the corneal damage caused by low temperature. cDNA microarray analysis revealed that the mAF-MSCs affected the expression patterns of mRNAs related to cell proliferation and differentiation pathways in the mice after transplantation. The results of quantitative real-time PCR and western blotting revealed that NAT12, NAT10, and the ETV4/JUN/CCND2 signaling axis were elevated significantly in the mAF-MSC-transplantation group, compared with those in the phosphate-buffered saline-treated groups. High performance liquid chromatography-mass spectroscopy results revealed that mAF-MSCs could promote mRNA N4-acetylcytidine (ac4C) modification and high expression of N-acetyltransferase in the eyeballs. RNA immunoprecipitation-PCR results showed that a specific product comprising Vegfa, Klf4, Ccnd2, Jun, and Etv4 mRNA specific coding region sites could be amplified using PCR from complexes formed in mAF-MSC-transplanted samples cross-linked with anti-ac4C antibodies. Thus, mouse amniotic fluid MSCs could repair the mouse corneal cold injury by promoting the ETV4/JUN/CCND2 signal axis activation and improving its stability by stimulating N4-acetylcytidine modification of their mRNAs.

Effect of Rapamycin Microspheres in Sjögren Syndrome Dry Eye: Preparation and Outcomes.

Purpose: To prepare a drug delivery system of rapamycin poly-3-hydroxybutyrate-co-3-hydroxyvalerate microspheres (RPM) and analyze its effect on the eyes of a Sjögren's syndrome mouse model.Methods: RPM was generated using a solvent evaporation method, and observed under light and electron microscopy. No obesity diabetes (NOD) mice with Sjögren's syndrome were randomized into normal saline, empty microspheres, and RPM groups, with healthy Kunming mice as the controls. Dry eye signs were assessed using a slit lamp. Tear secretion, corneal fluorescein, keratoconjunctival rose bengal, and conjunctival impression cytology were recorded.Results: RPM was smooth and spherical, about 20-50 µm in diameter, without agglomeration or adhesion. Mice receiving RPM secreted more tears. And they had a normal corneal histology, exhibiting reduced corneal fluorescein, rose bengal staining, and conjunctival impression cytology.Conclusion: RPM improved the signs of dry eyes, improved tear secretion, decreased corneal endothelial cell injury, and improved histological damage of the cornea in NOD mice.

Induction of reprogramming of human amniotic epithelial cells into iPS cells by overexpression of Yap, Oct4, and Sox2 through the activation of the Hippo-Yap pathway.

The present study has reported a novel method for producing induced pluripotent stem (iPS) cells. Primary human amniotic epithelial cells (HuAECs) were isolated from the amniotic membranes of pregnant women who received Cesarean sections. These cells were infected with retroviruses carrying octamer-binding transcription factor 4 (Oct4), (sex determining region Y)-box 2 (Sox2) and Yes-associated protein (Yap) (OSY). Following in vitro culture for ~14 days, epithelial-like HuAECs exhibited several iPS clone-like cell colonies (OSY-iPS). These cell clones presented positive alkaline phosphatase features and expressed high levels of embryonic stem cell-like markers (Nanog homeobox, Sox2, Oct4, reduced expression protein 1, and SSES3/4). Additionally, epigenetic analysis results indicated that the methylation of CpG islands on endogenous Oct4 and Sox2 promoters was reduced in OSY-iPS cells. Furthermore, the majority of the histone H3 at lysine 9 sites that interacted with the Oct4 and Sox2 promoters were acetylated, suggesting that the transcription activities of the above two transcription factors significantly increased. In vivo and in vitro induced differentiation experiments demonstrated that OSY-iPS could develop into embryoid bodies in vitro, and express numerous cellular markers in the three germ layers. Furthermore, OSY-iPS could form teratomas in immunodeficient mice. The pathological detection results suggest that these teratomas contain numerous types of cells from the three germ layers. However, the results from the quantitative polymerase chain reaction and western blot analyses suggest that the Hippo-Yap signaling pathway was significantly activated in OSY-iPS cells. In conclusion, a novel method for iPS induction was established in the present study. HuAECs were successfully induced to reprogram iPS cells through the introduction of OSY to activate the Hippo-Yap signaling pathway.

Vision Health-Related Quality of Life in Chinese Glaucoma Patients.

This cross-sectional study evaluated VRQOL in Chinese glaucoma patients and the potential factors influencing VRQOL. The VRQOL was assessed using the Chinese-version low vision quality of life questionnaire. Visual field loss was classified by the Hodapp-Parrish-Anderson method. The correlations of VRQOL to the best corrected visual acuity and the VF loss were investigated. The potential impact factors to VRQOL of glaucoma patients were screened by single factor analysis and were further analyzed by multiple regression analysis. There were significant differences in VRQOL scores between mild VF loss group and moderate VF loss group, moderate VF loss group and severe VF loss group, and mild VF loss group and severe VF loss group according to the better eye. In multiple linear regression, the binocular weighted average BCVA significantly affected the VRQOL scores. Binocular MD was the second influencing factor. In logistic regression, binocular severe VF loss and stroke were significantly associated with abnormal VRQOL. Education was the next influencing factor. This study showed that visual acuity correlated linearly with VRQOL, and VF loss might reach a certain level, correlating with abnormal VRQOL scores. Stroke was significantly associated with abnormal VRQOL.

Low temperature induces cryoinjury in mouse corneal endothelial cells by stimulating the Stk11-p53 signal pathway.

Cryoinjury, or injury caused by extremely low temperatures, can occur in corneal endothelial cells (CECs) and lead to visual impairment. However, the mechanism of cryoinjury in CECs is not clear. The Stk11­p53 signaling pathway regulates the proliferation and division of cells. Activity of the Stk11­p53 signaling pathway arrests the cell cycle at the G0/G1 phase and induces apoptosis. In this study, a mouse model of cryoinjury in CECs was used. Following injury, significant mouse CEC death and shedding were observed. In addition, the mRNA and protein levels of core factors from the Stk11­p53 signaling pathway (Stk11, p21 and p53) were elevated and Caspase­3 was activated following cryoinjury. In addition, chromatin immunoprecipitation revealed that Stk11 catalyzed p53 serine 15 phosphorylation, and the Stk11­p53 complex bound to the p21 promoter and stimulated gene transcription. Thus, the results of the present study suggest that cryoinjury leads to the damage and apoptosis of mouse CECs by activation of the Stk11­p53 signaling pathway, phosphorylation of p53 serine 15 and p21 gene transcription.

The impact of unilateral or bilateral cataract surgery on visual acuity and life quality of elderly patients.

In the current study, the CLVQOL was used to assess VRQOL before unilateral or bilateral cataract surgery and at the end of the follow-up period in order to determine the greater beneficial mode of surgery for patients, if one of the two surgical methods is more beneficial over the other. The patients were classified as receiving unilateral (group A) and bilateral cataract surgery (group B). There were no significant differences between groups A and B before the operation in terms of life quality scores, binocular weighted average LogMAR BCVA, age, educational level, gender, systematic and ocular comorbidities, and the complications of the operation. It was shown that visual acuity improved more significantly with bilateral cataract surgery than with unilateral surgery in elderly patients with a high preoperative disease burden in Shanghai city. However, the improvement in life quality was not different in patients receiving either bilateral or unilateral cataract surgery.