Proliferation Of Lens Epithelial Cells Research Articles

A 3D in vitro model for assessing the influence of intraocular lens: Posterior lens capsule interactions on lens epithelial cell responses

Posterior Capsule Opacification (PCO), the most frequent complication of cataract surgery, is caused by the infiltration and proliferation of lens epithelial cells (LECs) at the interface between the intraocular lens (IOL) and posterior lens capsule (PLC). According to the “no space, no cells, no PCO” theory, high affinity (or adhesion force) between the IOL and PLC would decrease the IOL: PLC interface space, hinder LEC migration, and thus reduce PCO formation. To test this hypothesis, an in vitro hemisphere-shaped simulated PLC (sPLC) was made to mimic the human IOL: PLC physical interactions and to assess their influence on LEC responses. Three commercially available IOLs with different affinities/adhesion forces toward the sPLC, including Acrylic foldable IOL, Silicone IOL, and PMMA IOL, were used in this investigation. Using the system, the physical interactions between IOLs and sPLC were quantified by measuring the adhesion force and interface space using an adhesion force apparatus and Optical Coherence Tomography, respectively. Our data shows that high adhesion force and tight binding between IOL and sPLC contribute to a small interface space (or “no space”). By introducing LECs into the in vitro system, we found that, with small interface space, among all IOLs, acrylic foldable IOLs permitted the least extent of LEC infiltration, proliferation, and differentiation (or “no cells”). Further statistical analyses using clinical data revealed that weak LEC responses are associated with low clinical PCO incidence rates (or “no PCO”). The findings support that the in vitro system could simulate IOL: PLC interplays and predict IOLs’ PCO potential in support of the “no space, no cells, no PCO” hypothesis.

Role of C/EBP Homologous Protein (CHOP) and Nupr1 Interaction in Endoplasmic Reticulum Stress-Induced Apoptosis of Lens Epithelial Cells.

Our study mainly analyzed the mechanism of C/EBP homologous protein (CHOP) and its interacting protein Nupr1 on endoplasmic reticulum stress (ERS) induced lens epithelial cells (LEC) apoptosis. Cell proliferation was detected by CCK-8. Apoptosis was detected by flow cytometry and TUNEL. Nupr1 expressionwas detected byRT-qPCR. The expressions of CHOP, Nupr1, apoptosis-related protein, and ERS-related protein were detected by Western blot. DCFH-DA probe was used to detect cell ROS. The SOD, GSH-PX, and MDA contents were detected by the kit. Co-IP was used to detect the interaction between CHOP and Nupr1. The morphology of the lens was detected by HE staining. The result shows thatTunicamycin (TU) can induce endoplasmic reticulum stress and apoptosis in LEC in a concentration-dependent manner. TU induction leads to the occurrence of CHOP nuclear translocation. Overexpression of CHOP can further enhance the inhibitory effect of TU on LEC proliferation and the promotion of apoptosis, while knockdown of CHOP has the opposite effect. CHOP and Nupr1 are interacting proteins, and knockdown of Nupr1 or addition of Nupr1 inhibitor ZZW-115 can reverse the effects of TU and overexpression of CHOP, respectively. It has been observed in animal experiments that treatment with oe-CHOP can further aggravate the pathological lesions of the rat lens, while ZZW-115 can reverse the effect of oe-CHOP to a certain extent and improve the lesions of the rat lens. Overall,CHOP interacts with Nupr1 to regulate apoptosis caused by ERS and mediate cataract progression in rats, and this study provides a new potential therapeutic target for the treatment of cataract.

Zwitterionic modified and freeze-thaw reinforced foldable hydrogel as intraocular lens for posterior capsule opacification prevention

Posterior capsule opacification (PCO) is a predominant postoperative complication, often leading to visual impairment due to the aberrant proliferation and adhesion of lens epithelial cells (LECs) and protein precipitates subsequent to intraocular lens (IOL) implantation. To address this clinical issue, a foldable and antifouling sharp-edged IOL implant based on naturally-derived cellulose hydrogel is synthesized. The mechanical strength and transparency of the hydrogel is enhanced via repeated freeze-thaw (FT) cycles. The incorporated zwitterionic modifications can remarkably prevent the incidence of PCO by exhibiting proteins repulsion and cell anti-adhesion properties. The graft of dopamine onto both the haptic and the periphery of the posterior surface ensures the adhesion of the hydrogel to the posterior capsule and impedes the migration of LECs without compromising transparency. In in vivo study, the zwitterionic modified foldable hydrogel exhibits uveal and capsular biocompatibility synchronously with no signs of inflammatory response and prevent PCO formation, better than that of commercialized and PEG-modified IOL. With foldability, endurability, antifouling effect, and adhesive to posterior capsule, the reported hydrogel featuring heterogeneous surface design displays great potential to eradicate PCO and attain post-operative efficacy after cataract surgery.

Regulation of transforming growth factor β2-induced epithelial-mesenchymal transition and autophagy in lens epithelial cells by the miR-492/NPM1 axis.

A cataract is a clinically common blinding disease closely related to the ageing of the eye cells, which has become a major health killer in the elderly. Our research seeks to analyze the primary targets linked to the pathogenesis of cataracts during the ageing process. We performed bioinformatics analyses on the GSE101727 dataset to discover genes linked with ageing and cataracts. To explore the impacts of Nucleophosmin 1 (NPM1) on cell apoptosis, proliferation, as well as epithelial-mesenchymal transition (EMT) processes, in vitro tests such as western blotting, flow cytometry, and MTT were carried out. Additionally, the study incorporated transforming growth factor β2 (TGF-β2) to examine its function in cellular responses, chloroquine (CQ) to regulate autophagic flow, and H2O2 therapy to mimic oxidative stress. Our study discovered seven ageing-related genes, including NPM1, that had substantial relationships with cataracts. NPM1 overexpression was shown to boost cell proliferation and prevent apoptosis in SRA01/04 cells. Notably, NPM1 modulated the TGF-β signalling pathway, influencing cell proliferation and EMT processes. miR-429 was shown to be adversely regulating NPM1 and autophagy-related proteins, as demonstrated by changes in their expression in response to TGF-β2 treatment. Furthermore, NPM1 knockdown restored autophagy activity suppressed by miR-429 mimics, indicating a complex interaction of miR-429, NPM1, and TGF-β2 pathways in regulating autophagy and EMT. Lens epithelial cell proliferation and apoptosis were largely regulated by NPM1, as well as autophagy and EMT, which were significantly mediated by TGF-β2 and the miR-429/NPM1 axis. These results imply new possible targets for prognosis and therapy of cataracts.

Retraction Note: MiRNA-26b inhibits the proliferation, migration, and epithelial-mesenchymal transition of lens epithelial cells.

Retraction Note: MiRNA-26b inhibits the proliferation, migration, and epithelial-mesenchymal transition of lens epithelial cells.

Conditional Ablation of Spred1 and Spred2 in the Eye Lens Negatively Impacts Its Development and Growth.

The development and growth of the eye depends on normal lens morphogenesis and its growth. This growth, in turn, is dependent on coordinated proliferation of the lens epithelial cells and their subsequent differentiation into fiber cells. These cellular processes are tightly regulated to maintain the precise cellular structure and size of the lens, critical for its transparency and refractive properties. Growth factor-mediated MAPK signaling driven by ERK1/2 has been reported as essential for regulating cellular processes of the lens, with ERK1/2 signaling tightly regulated by endogenous antagonists, including members of the Sprouty and related Spred families. Our previous studies have demonstrated the importance of both these inhibitory molecules in lens and eye development. In this study, we build on these findings to highlight the importance of Spreds in regulating early lens morphogenesis by modulating ERK1/2-mediated lens epithelial cell proliferation and fiber differentiation. Conditional loss of both Spred1 and Spred2 in early lens morphogenesis results in elevated ERK1/2 phosphorylation, hyperproliferation of lens epithelia, and an associated increase in the rate of fiber differentiation. This results in transient microphakia and microphthalmia, which disappears, owing potentially to compensatory Sprouty expression. Our data support an important temporal role for Spreds in the early stages of lens morphogenesis and highlight how negative regulation of ERK1/2 signaling is critical for maintaining lens proliferation and fiber differentiation in situ throughout life.

Smurf1 Modulates Smad Signaling Pathway in Fibrotic Cataract Formation.

TGF-β/BMP signaling pathway plays a significant role in fibrotic cataract. Smurf1, a ubiquitin protein ligase, regulates the TGF-β/BMP signaling pathway through the ubiquitin-proteasome system (UPS). This study aims to investigate the role of Smurf1 in the progression of fibrotic cataract and its underlying mechanism. We used a mouse model of injury-induced anterior subcapsular cataract (ASC) and administered the Smurf1 inhibitor A01 for in vivo investigations. RNA sequencing was performed to examine global gene expression changes. Protein levels were assessed by Simple Western analysis. The volume of subcapsular opacity was determined using whole-mount immunofluorescence of lens anterior capsules. Lentivirus was utilized to establish cell lines with Smurf1 knockdown or overexpression in SRA01/04. Lens epithelial cell (LEC) proliferation was evaluated by CCK8 and EdU assays. Cell cycle profile was determined by flow cytometry. LEC migration was measured using Transwell and wound healing assays. The mRNA levels of genes associated with cell proliferation, migration, epithelial-mesenchymal transition (EMT), TGF-β/BMP pathway, and UPS were upregulated in mouse ASC model. Smurf1 mRNA and protein levels were upregulated in lens capsules of patients and mice with ASC. Anterior chamber injection of A01 inhibited ASC formation and EMT. In vitro, Smurf1 knockdown reduced proliferation, migration and TGF-β2-induced EMT of LECs, concomitant with the upregulation of Smad1, Smad5, and pSmad1/5. Conversely, overexpression of Smurf1 showed opposite phenotypes. Smurf1 regulates fibrotic cataract progression by influencing LEC proliferation, migration, and EMT through the modulation of the Smad signaling pathway, offering a novel target for the fibrotic cataract treatment.

Knockout of TGF-β receptor II by CRISPR/Cas9 delays mesenchymal transition of Lens epithelium and posterior capsule opacification

Posterior capsule opacification (PCO) is the most common postoperative complication of cataract surgery. Transforming growth factor-β (TGF-β) is related to epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) that is proven to induce PCO formation in clinical and experimental studies. In this study, CRISPR sequences targeting exon of TGF-βRII were knocked out with lentiviral transfection in LECs. Rabbits' PCO model was established and recombinant adeno-associated virus (AAV) for transferring the gRNA of TGF βRII were intravitreally injected. SgRNA inhibited TGF-βRII expression and human LECs proliferation. In TGF-βRII knockout group, LECs motility and migration were suppressed, N-cadherin and vimentin expressions were significantly decreased, whereas E-cadherin was increased. The animal model showed that TGF-βRII knockout in vivo was effective in suppressing PCO. The current study suggested that the CRISPR/Cas9 endonuclease system could suppress TGF-βRII secretion, which participates in the EMT procedure of LECs in vitro and PCO in vivo. These findings might provide a new gene-editing approach and insight into a novel therapeutic strategy for PCO.

JAM-C Is Important for Lens Epithelial Cell Proliferation and Lens Fiber Maturation in Murine Lens Development.

The underlying mechanism of congenital cataracts caused by deficiency or mutation of junctional adhesion molecule C (JAM-C) gene remains unclear. Our study aims to elucidate the abnormal developmental process in Jamc-/- lenses and reveal the genes related to lens development that JAM-C may regulate. Jamc knockout (Jamc-/-) mouse embryos and pups were generated for in vivo studies. Four key developmental stages from embryonic day (E) 12.5 to postnatal day (P) 0.5 were selected for the following experiments. Hematoxylin and eosin staining was used for histological analysis. The 5-bromo-2'-deoxyuridine (BrdU) incorporation assay and TUNEL staining were performed to label lens epithelial cell (LEC) proliferation and apoptosis, respectively. Immunofluorescence and Western blot were used to analyze the markers of lens epithelium, cell cycle exit, and lens fiber differentiation. JAM-C was expressed throughout the process of lens development. Deletion of Jamc resulted in decreased lens size and disorganized lens fibers, which arose from E16.5 and aggravated gradually. The LECs of Jamc-/- lenses showed decreased quantity and proliferation, accompanied with reduction of key transcription factor, FOXE3. The fibers in Jamc-/- lenses were disorganized. Moreover, Jamc-deficient lens fibers showed significantly altered distribution patterns of Cx46 and Cx50. The marker of fiber homeostasis, γ-crystallin, was also decreased in the inner cortex and core fibers of Jamc-/- lenses. Deletion of JAM-C exhibits malfunction of LEC proliferation and fiber maturation during murine lens development, which may be related to the downregulation of FOXE3 expression and abnormal localization patterns of Cx46 and Cx50.

Inhibition of miR-29a-3p Alleviates Apoptosis of Lens Epithelial Cells via Upregulation of CAND1

Purpose Accumulated evidence has shown that microRNAs (miRNAs) are closely related to the pathogenesis and progression of senile cataracts. Here we investigate the effect of miR-29a-3p in cataractogenesis and determined the potential molecular mechanism involved. Methods In this study, we constructed a selenite cataract model in rats and obtained the miRNAs related to cataracts by whole transcriptome sequencing. To investigate the effect and mechanism of miR-29a-3p on cataracts, we performed several in vivo and in vitro experiments, including CCK8 assay, flow cytometry, luciferase reporter assay, Edu assay, and western blot analysis. Result Sequencing data showed downregulation of miR-29a-3p in rats with selenite cataracts. Down-regulation of miR-29a-3p could promote lens epithelial cells (SRA01/04) proliferation and inhibit cell apoptosis, and miR-29a-3p silence could inhibit the development of cataracts. Additionally, CAND1 was a direct target gene for miR-29a-3p. Conclusion These data demonstrate that miR-29a-3p inhibits apoptosis of lens epithelial cells by regulating CAND1, which may be a potential target for senile cataracts.

Identification of Small Molecules for Prevention of Lens Epithelium-Derived Cataract Using Zebrafish.

Cataract is the leading cause of blindness worldwide. It can be treated by surgery, whereby the damaged crystalline lens is replaced by a synthetic lens. Although cataract surgery is highly effective, a relatively common complication named posterior capsular opacification (PCO) leads to secondary loss of vision. PCO is caused by abnormal proliferation and migration of residual lens epithelial cells (LECs) that were not removed during the surgery, which results in interruption to the passage of light. Despite technical improvements to the surgery, this complication has not been eradicated. Efforts are being made to identify drugs that can be applied post-surgery, to inhibit PCO development. Towards the goal of identifying such drugs, we used zebrafish embryos homozygous for a mutation in plod3 that develop a lens phenotype with characteristics of PCO. Using both biased and unbiased approaches, we identified small molecules that can block the lens phenotype of the mutants. Our findings confirm the relevance of zebrafish plod3 mutants' lens phenotype as a model for lens epithelium-derived cataract and add to our understanding of the molecular mechanisms that contribute to the development of this pathology. This understanding should help in the development of strategies for PCO prevention.

Mild Photothermal Therapy Prevents Posterior Capsule Opacification through Cytoskeletal Remodeling.

Cataract is the first leading cause of blindness in the world and posterior capsule opacification (PCO) is the most common long-term complication after surgery. The primary pathogenic processes contributing to PCO are the proliferation and migration of residual lens epithelial cells (LECs). This study aimed to explore the mild photothermal effect on LECs. Interestingly, this work finds that the mild photothermal effect significantly inhibited the proliferation and migration of LECs. The live cell fluorescence imaging reveals that the remodeling of the actin cytoskeleton and cell morphology attributed to the inhibition effect. Further mechanistic studies at molecular level suggest that the mild photothermal effect can regulate the phosphorylation of ERM, YAP, and Cofilin and thereby affect the proliferation and migration of LECs. In order to explore the potential clinical application of mild photothermal therapy for PCO prevention, PDA/PVA gel rings with photothermal effect is prepared by the repeated freeze-thaw method and conducted experiments in vivo, which achieved favorable PCO prevention effect. Overall, this study shows that the mild photothermal effect can regulate the proliferation and migration of LECs through cytoskeletal remodeling and the results of experiments in vivo demonstrate that mild photothermal effect is a promising approach for PCO prevention.

Arginase-1 promotes lens epithelial-to-mesenchymal transition in different models of anterior subcapsular cataract

BackgroundArginase-1 (ARG1) promotes collagen synthesis and cell proliferation. ARG1 is highly expressed in various tumour cells. The mechanisms of ARG1 in epithelial-to-mesenchymal transition (EMT)-associated cataracts were studied herein.MethodsC57BL/6 mice, a human lens epithelial cell line (HLEC-SRA01/04), and human lens capsule samples were used in this study. The right lens anterior capsule of the mouse eye was punctured through the central cornea with a 26-gauge hypodermic needle. Human lens epithelial cells (HLECs) were transfected with ARG1-targeted (siARG1) or negative control siRNA (siNC). For gene overexpression, HLECs were transfected with a plasmid bearing the ARG1 coding sequence or an empty vector. Medium containing 0.2% serum with or without transforming growth factor beta-2 (TGF-β2) was added for 6 or 24 h to detect mRNA or protein, respectively. The expression of related genes was measured by quantitative real-time polymerase chain reaction (RT–qPCR), western blotting, and immunohistochemical staining. Transwell assays and wound healing assays were used to determine cell migration. Cell proliferation, superoxide levels, nitric oxide (NO) levels, and arginase activity were estimated using Cell Counting Kit-8 assays, a superoxide assay kit, an NO assay kit, and an arginase activity kit.ResultsARG1, alpha-smooth muscle actin (α-SMA), fibronectin, and Ki67 expression increased after lens capsular injury, while zonula occludens-1 (ZO-1) expression decreased.Fibronectin and collagen type I alpha1 chain (collagen 1A1) expression increased, and cell migration increased significantly in ARG1-overexpressing HLECs compared with those transfected with an empty vector after TGF-β2 treatment. These effects were reversed by ARG1 knockdown.The arginase-related pathway plays an important role in EMT. mRNAs of enzymes of the arginase-related pathway were highly expressed after ARG1 overexpression. ARG1 knockdown suppressed these expression changes.Numidargistat (CB-1158) dihydrochloride (CB-1158), an ARG1 inhibitor, suppressed TGF-β2-induced anterior subcapsular cataract (ASC) by reducing the proliferation of lens epithelial cells (LECs) and decreasing fibronectin, α-SMA, collagen 1A1, and vimentin expression.Compared with that in nonanterior subcapsular cataract (non-ASC) patients, the expression of ARG1, collagen 1A1, vimentin, fibronectin, and Ki67 was markedly increased in ASC patients.ConclusionsARG1 can regulate EMT in EMT-associated cataracts. Based on the pathogenesis of ASC, these findings are expected to provide new therapeutic strategies for patients.Graphical abstract

Rotating the Intraocular Lens to Prevent Posterior Capsular Opacification in Cataract Surgeries.

Posterior capsule opacification (PCO) is a common postoperative complication of extracapsular cataract surgery, which is caused by the proliferation and migration of lens epithelial cells and can affect long-term visual outcomes significantly. The most effective treatment for PCO is neodymium-doped yttrium aluminum garnet (Nd:YAG) laser capsulotomy; however, this treatment is associated with posterior segment complication and can break the stability of capsular bag, affecting the position and function of trifocal or toric intraocular lenses (IOLs). Advances in surgical procedures, IOL design, and pharmacy have reduced the rate of PCO in recent years, concentrating on the inhibition of proliferative lens epithelial cells (LECs). This protocol aimed to clear LECs more thoroughly during phacoemulsification and IOL implantation. The first several steps, including clear corneal incision, continuous circular capsulorhexis, hydrodissection, hydrodelineation, and phacoemulsification, were completed as conventional procedures. After placing the IOL into the capsular bag, rotation of the IOL by at least 360° was performed using an irrigation/aspiration tip or a hook, with slight stress on the posterior capsule. Some residuals occurred in the originally transparent capsular bag after rotation of the IOLs. Then, these materials and the viscoelastic were cleared completely using an irrigation/aspiration system. A clear posterior capsule was observed after the surgery in patients undergoing this method. This method of rotating IOLs is a simple, effective, and safe way to prevent PCO by clearing residual LECs and can be carried out without extra tools or skills.

Characterization of a Novel Gja8 (Cx50) Mutation in a New Cataract Rat Model.

To describe a novel spontaneous cataract inbred strain isolated from large-scale breeding SD rats, identify the responsible gene mutation, and understand how this mutation affects lens function. Exome sequencing of 12 cataract-associated genes was performed in the affected and healthy relatives. Sequences of rat wild-type or mutant gap junction protein alpha 8 gene (Gja8) were transfected into cells. The expression level of protein was assayed by Western blot analysis. Subcellular localization of connexin 50 (Cx50) was analyzed in confocal fluorescent images. Wound-healing, 5-ethynyl-2'-deoxyuridine incorporation, and attachment assay were performed to characterize the cell migration, proliferation and adhesion. The abnormality was found to be inheritable in an autosomal semi-dominant pattern through different mating patterns. We found a G to T transversion at codon 655 in Gja8, leading to a substitution of valine by phenylalanine (p.V219F). Gja8V219F/+ heterozygotes expressed nuclear cataract while Gja8V219F/V219F homozygotes manifested microphthalmia in addition to cataract. Histology revealed fiber disorders and loss of organelle-free zone in the mutant lens. Cx50V219F altered its location in HeLa cells and inhibited the proliferation, migration and adhesion abilities of HLEB3 cells. The mutation also reduced the expression of focal adhesion kinase and its phosphorylation. The c.655G>T mutation (p.V219F) is a novel mutation in Gja8, inducing semi-dominant nuclear cataracts in a new spontaneous cataract rat model. The p.V219F mutation altered Cx50 distribution, inhibited lens epithelial cell proliferation, migration, and adhesion, and disrupted fiber cell differentiation. As a consequence, the nuclear cataract and small lens formed.

Let-7c-3p suppresses lens epithelial-mesenchymal transition by inhibiting cadherin-11 expression in fibrotic cataract.

Fibrotic cataract, including anterior subcapsular cataract (ASC) and posterior capsule opacification, always lead to visual impairment. Epithelial-mesenchymal transition (EMT) is a well-known event that causes phenotypic alterations in lens epithelial cells (LECs) during lens fibrosis. Accumulating studies have demonstrated that microRNAs are important regulators of EMT and fibrosis. However, the evidence explaining how microRNAs modulate the behavior and alter the cellular phenotypes of the lens epithelium in fibrotic cataract is insufficient. In this study, we found that hsa-let-7c-3p is downregulated in LECs in human ASC in vivo as well as in TGFβ2-induced EMT in vitro, indicating that hsa-let-7c-3p may participate in modulating the profibrotic processes in the lens. We then demonstrated that overexpression of hsa-let-7c-3p markedly suppressed human LEC proliferation and migration and attenuated TGFβ2-induced EMT and injury-induced ASC in a mouse model. In addition, hsa-let-7c-3p mediated lens fibrosis by directly targeting the CDH11 gene, which encodes cadherin-11 protein, an important mediator in the EMT signaling pathway. It decreased cadherin-11 protein expression at the posttranscriptional level but not at the transcriptional level by binding to a specific site in the 3-untranslated region (3'-UTR) of CDH11 mRNA. Moreover, blockade of cadherin-11 expression with a specific short hairpin RNA reversed TGFβ2-induced EMT in LECs in vitro. Collectively, these data demonstrated that hsa-let-7c-3p plays a clear role in attenuating ASC development and may be a novel candidate therapeutic for halting fibrosis and maintaining vision.

Spatiotemporal Localisation of Heparan Sulphate Proteoglycans throughout Mouse Lens Morphogenesis.

Heparan sulphate proteoglycans (HSPGs) consist of a core protein decorated with sulphated HS-glycosaminoglycan (GAG) chains. These negatively charged HS-GAG chains rely on the activity of PAPSS synthesising enzymes for their sulfation, which allows them to bind to and regulate the activity of many positively charged HS-binding proteins. HSPGs are found on the surfaces of cells and in the pericellular matrix, where they interact with various components of the cell microenvironment, including growth factors. By binding to and regulating ocular morphogens and growth factors, HSPGs are positioned to orchestrate growth factor-mediated signalling events that are essential for lens epithelial cell proliferation, migration, and lens fibre differentiation. Previous studies have shown that HS sulfation is essential for lens development. Moreover, each of the full-time HSPGs, differentiated by thirteen different core proteins, are differentially localised in a cell-type specific manner with regional differences in the postnatal rat lens. Here, the same thirteen HSPG-associated GAGs and core proteins as well as PAPSS2, are shown to be differentially regulated throughout murine lens development in a spatiotemporal manner. These findings suggest that HS-GAG sulfation is essential for growth factor-induced cellular processes during embryogenesis, and the unique and divergent localisation of different lens HSPG core proteins indicates that different HSPGs likely play specialized roles during lens induction and morphogenesis.

Low-dose ionizing radiation: Effects on the proliferation and migration of lens epithelial cells via activation of the Wnt/β-catenin pathway.

Eye lens opacification (cataract) induced by ionizing radiation is an important concern for radiation protection. Human lens epithelial cells (HLE-B3) were irradiated with γ-rays and radiation effects, including cell proliferation, cell migration, cell cycle distribution, and other changes related to the β-catenin pathway, were determined after 8-72h and 7 d. In an in vivo model, mice were irradiated; DNA damage (γH2AX foci) in the cell nucleus of the anterior capsule of the lens was detected within 1h, and radiation effects on the anterior and posterior lens capsules were observed after 3 months. Low-dose ionizing radiation promoted cell proliferation and migration. The expression levels of β-catenin, cyclin D1, and c-Myc were significantly increased in HLE-B3 cells after irradiation and β-catenin was translocated into the cell nucleus (activation of the Wnt/β-catenin pathway). In C57BL/6J mouse lens, even a very low irradiation dose (0.05Gy) induced the formation of γH2AX foci, 1h after irradiation. At 3 months, migratory cells were found in the posterior capsule; expression of β-catenin was increased and it was clustered at the nucleus in the epithelial cells of the lens anterior capsule. The Wnt/β-catenin signaling pathway may an important role in promoting abnormal proliferation and migration of lens epithelial cells after low-dose irradiation.

Prophylaxis of posterior capsule opacification through autophagy activation with indomethacin-eluting intraocular lens

Posterior capsule opacification (PCO) is the most common long-term postoperative complication of cataract surgery, leading to secondary vision loss. Optimized intraocular lens (IOL) structure and appropriate pharmacological intervention, which provides physical barriers and biological inhibition, respectively, can block the migration, proliferation, and epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) for PCO prophylaxis. Herein, a novel indomethacin-eluting IOL (INDOM-IOL) with an optimized sharper edge and a sustained drug release behavior was developed for PCO prevention. Indomethacin (INDOM), an ophthalmic non-steroidal anti-inflammatory drug (NSAID) used for postoperative ocular inflammation, was demonstrated to not only be able to suppress cell migration and down-regulate the expression of cyclooxygenase-2 (COX-2) and EMT markers, including alpha-smooth muscle actin (α-SMA) and cyclin D1, but also promote the autophagy activation in LECs. Additionally, autophagy was also verified to be a potential therapeutic target for the down-regulation of EMT in LECs. The novel IOL, serving as a drug delivery platform, could carry an adjustable dose of hydrophobic indomethacin with sustained drug release ability for more than 28 days. In the rabbit PCO model, the indomethacin-eluting IOL showed excellent anti-inflammatory and anti-PCO effects. In summary, indomethacin is an effective pharmacological intervention in PCO prophylaxis, and the novel IOL we developed prevented PCO in vivo under its sustained indomethacin release property, which provided a promising approach for PCO prophylaxis in clinical application.

BMP-4 and BMP-7 Inhibit EMT in a Model of Anterior Subcapsular Cataract in Part by Regulating the Notch Signaling Pathway.

The proliferation, migration, and epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) are believed to be the pathological mechanisms underlying anterior subcapsular cataract (ASC). Bone morphogenetic proteins (BMPs) inhibit transforming growth factor-beta (TGF-β)-induced fibrosis in the lens. Herein, we aimed to further clarify the roles of BMP-4/BMP-7 in the progression and the underlying mechanisms of fibrotic cataract. BMP-4/BMP-7, TGF-β2, jagged-1 peptide, or DAPT were applied in a mouse injury-induced ASC model and in the human LEC cell line SRA01/04. The volume of opacity was examined by a slit lamp and determined by lens anterior capsule whole-mount immunofluorescence. Global gene expression changes were assessed by RNA sequencing, and the levels of individual mRNAs were validated by real-time PCR. Protein expression was determined by the Simple Western sample dilution buffer. Cell proliferation was examined by CCK8 and EdU assays, and cell migration was measured by Transwell and wound healing assays. Anterior chamber injection of BMP-4/BMP-7 significantly suppressed subcapsular opacification formation. RNA sequencing of the mouse ASC model identified the Notch pathway as a potential mechanism involved in BMP-mediated inhibition of ASC. Consistently, BMP-4/BMP-7 selectively suppressed Notch1 and Notch3 and their downstream genes, including Hes and Hey. BMP-4/BMP-7 or DAPT suppressed cell proliferation by inducing G1 cell cycle arrest. BMP-4/BMP-7 also inhibited TGF-β2-induced cell migration and EMT by modulating the Notch pathway. BMP-4/BMP-7 attenuated ASC by inhibiting proliferation, migration, and EMT of LECs via modulation of the Notch pathway, thereby providing a new avenue for ASC treatment.