Scleral remodeling during myopia development in mice eyes: a potential role of thrombospondin-1.

BACKGROUND: Scleral extracellular matrix (ECM) remodeling plays a crucial role in the development of myopia, particularly in ocular axial elongation. Thrombospondin-1 (THBS1), also known as TSP-1, is a significant cellular protein involved in matrix remodeling in various tissues. However, the specific role of THBS1 in myopia development remains unclear. METHOD: We employed the HumanNet database to predict genes related to myopic sclera remodeling, followed by screening and visualization of the predicted genes using bioinformatics tools. To investigate the potential target gene Thbs1, we utilized lens-induced myopia models in male C57BL/6J mice and performed Western blot analysis to detect the expression level of scleral THBS1 during myopia development. Additionally, we evaluated the effects of scleral THBS1 knockdown on myopia development through AAV sub-Tenon's injection. The refractive status and axial length were measured using a refractometer and SD-OCT system. RESULTS: During lens-induced myopia, THBS1 protein expression in the sclera was downregulated, particularly in the early stages of myopia induction. Moreover, the mice in the THBS1 knockdown group exhibited alterations in myopia development in both refraction and axial length changed compared to the control group. Western blotting analysis confirmed the effectiveness of AAV-mediated knockdown, demonstrating a decrease in COLA1 expression and an increase in MMP9 levels in the sclera. CONCLUSION: Our findings indicate that sclera THBS1 levels decreased during myopia development and subsequent THBS1 knockdown showed a decrease in scleral COLA1 expression. Taken together, these results suggest that THBS1 plays a role in maintaining the homeostasis of scleral extracellular matrix, and the reduction of THBS1 may promote the remodeling process and then affect ocular axial elongation during myopia progression.

The role of transforming growth factor beta in myopia development.

Myopia is widely recognized as an epidemic. Studies have found a link between Transforming Growth Factor-beta (TGF-ß) and myopia, but the specific molecular mechanisms are not fully understood. In this study, a monocular model in tree shrews (Tupaia belangeri) was established to verify the molecular mechanism of TGF-ß in myopia. The results indicated that there were significant changes in TGF-ßs during the treatment of myopia, which could enhance the refractive ability and axial length of the eye. Immunohistochemical staining, real-time fluorescent quantitative PCR, and immunoblotting results showed a significant upregulation of MMP2 and NF-κB levels, and a significant downregulation of COL-I expression in the TGF-ß treated eyes, suggesting that NF-κB and MMP2 are involved in the signaling pathways of TGF-ßs induced myopia and axial elongation. Moreover, the expression levels of IL-6, IL-8, MCP-1, IL-1ß, TNF-α, TAK1, and NF-κB in the retina were all significantly elevated. This indicates that TGF-ß stimulates the inflammatory response of retinal pigment epithelial cells through the TAK1-NF-κB signaling pathway. In conclusion, this study suggests that TGF-ß promotes the progression of myopia by enhancing intraocular inflammation.

Augmentation of scleral glycolysis promotes myopia through histone lactylation.

Myopia is characterized of maladaptive increases in scleral fibroblast-to-myofibroblast transdifferentiation (FMT). Scleral hypoxia is a significant factor contributing to myopia, but how hypoxia induces myopia is poorly understood. Here, we showed that myopia in mice and guinea pigs was associated with hypoxia-induced increases in key glycolytic enzymes expression and lactate levels in the sclera. Promotion of scleral glycolysis or lactate production induced FMT and myopia; conversely, suppression of glycolysis or lactate production eliminated or inhibited FMT and myopia. Mechanistically, increasing scleral glycolysis-lactate levels promoted FMT and myopia via H3K18la, and this promoted Notch1 expression. Genetic analyses identified a significant enrichment of two genes encoding glycolytic enzymes, ENO2 and TPI1. Moreover, increasing sugar intake in guinea pigs not only induced myopia but also enhanced the response to myopia induction via the scleral glycolysis-lactate-histone lactylation pathway. Collectively, we suggest that scleral glycolysis contributes to myopia by promoting FMT via lactate-induced histone lactylation.

Metabolomic profiling of ocular tissues in rabbit myopia: Uncovering differential metabolites and pathways.

To investigate the metabolic difference among tissue layers of the rabbits' eye during the development of myopia using metabolomic techniques and explore any metabolic links or cascades within the ocular wall. Ultra Performance Liquid Chromatography - Mass Spectrometry (UPLC-MS) was utilized for untargeted metabolite screening (UMS) to identify the significant differential metabolites produced between myopia (MY) and control (CT) (horizontal). Subsequently, we compared those key metabolites among tissues (Sclera, Choroid, Retina) of MY for distribution and variation (longitudinal). A total of 6285 metabolites were detected in the three tissues. The differential metabolites were screened and the metabolic pathways of these metabolites in each myopic tissue were labeled, including tryptophan and its metabolites, pyruvate, taurine, caffeine metabolites, as well as neurotransmitters like glutamate and dopamine. Our study suggests that multiple metabolic pathways or different metabolites under the same pathway, might act on different parts of the eyeball and contribute to the occurrence and development of myopia by affecting the energy supply to the ocular tissues, preventing antioxidant stress, affecting scleral collagen synthesis, and regulating various neurotransmitters mutually.

Diurnal retinal and choroidal gene expression patterns support a role for circadian biology in myopia pathogenesis.

The prevalence of myopia (nearsightedness) is increasing to alarming levels, but its etiology remains poorly understood. Because both laboratory and clinical findings suggest an etiologic role for circadian rhythms in myopia development, we assayed gene expression by RNA-Seq in retina and choroid at the onset of unilateral experimental myopia in chick, isolating tissues every 4 h during a single 24-h period from myopic and contralateral control eyes. Occluded versus open eye gene expression differences varied considerably over the 24-h sampling period, with some occurring at multiple times of day but with others showing differences at only a single investigated timepoint. Some of the genes identified in retina or choroid of chick myopia were previously identified as candidate genes for common human myopia. Like differentially expressed genes, pathways identified by Gene Set Enrichment Analysis also varied dramatically by sampling time. Considered with other laboratory data, human genetic and epidemiology data, these findings further implicate circadian events in myopia pathogenesis. The present results emphasize a need to include time of day in mechanistic studies of myopia and to assess circadian biology directly in trying to understand better the origin of myopia and to develop more effective therapies.

Intraocular cetuximab: Safety and effect on axial elongation in young Guinea pigs with lens-induced myopization.

This study aimed to examine the intraocular tolerability of the epidermal growth factor receptor antibody cetuximab, when applied intravitreally, and its effect on axial elongation. Guinea pigs aged 2-3 weeks were subjected to bilateral plano glasses and bilateral lens-induced myopization (LIM) as a single procedure for group I (n = 8) and group II (n = 8), respectively. In the animals of group III (n = 8), group IV (n = 8), and group V (n = 8), the right eyes of the animals, in addition to LIM, received four weekly intravitreal injections of cetuximab (Erbitux®) in doses of 6.25 µg, 12.5 µg, and 25 µg, respectively. As controls, the left eyes, in addition to LIM, received corresponding intraocular injections of phosphate-buffered saline. The animals underwent regular ophthalmoscopic examinations and biometry for axial length measurements. With increasing doses of cetuximab, the inter-eye difference in axial elongation (at study end, left eyes minus right eyes) were significantly the smallest in group I (0.00 ± 0.02 mm) and group II (-0.01 ± 0.02 mm), they were larger in group III (0.04 ± 0.04 mm) and group IV (0.10 ± 0.03 mm), and they were the largest in group V (0.11 ± 0.01 mm). The inter-eye difference in axial elongation enlarged (P < 0.001) with the number of injections applied. Retinal thickness at the posterior pole (right eyes) was significantly thicker in group V than in group II (P < 0.01). The density of apoptotic cells (visualized by TUNEL-staining) did not vary significantly between any of the groups (all P > 0.05). The results suggest that intravitreal injections of cetuximab in young guinea pigs with LIM resulted in a reduction in axial elongation in a dose-dependent and number of treatment-dependent manner. Intraocular toxic effects, such as intraocular inflammation, retinal thinning, or an increased density of apoptotic cells in the retina, were not observed in association with the intravitreally applied cetuximab.

Exploration of the pathophysiology of high myopia via proteomic profiling of human corneal stromal lenticules.

This study aimed to investigate the underlying pathophysiology of high myopia by analyzing the proteome of human corneal stromal lenticule samples obtained through small incision lenticule extraction (SMILE). A total of thirty-two patients who underwent SMILE were included in the study. Label-free quantitative proteomic analysis was performed on corneal stromal lenticule samples, equally representing high myopia (n = 10) and low myopia (n = 10) groups. The identified and profiled lenticule proteomes were analyzed using in silico tools to explore biological characteristics of differentially expressed proteins (DEPs). Additionally, LASSO regression and random forest model were employed to identify key proteins associated with the pathophysiology of high myopia. The DEPs were found to be closely linked to immune activation, extracellular matrix, and cell adhesion-related pathways according to gene ontology analysis. Specifically, decreased expression of COL1A1 and increased expression of CDH11 were associated with the pathogenesis of high myopia and validated by western blotting (n = 6) and quantitative real time polymerase chain reaction (n = 6). Overall, this study provides evidence that COL1A1 and CDH11 may contribute to the pathophysiology of high myopia based on comparative proteomic profiling of human corneal stromal lenticules obtained through SMILE.

Investigation of Mitochondrial Homeostasis Changes in Lens Epithelium of High-Myopic Cataract.

PURPOSE: High myopia is demonstrated as a pathogenic factor for nuclear cataract. The main mechanism of high-myopia cataracts (HMC) is oxidative damage, which causes mitochondrial homeostasis imbalance. This study aimed to explore the mitochondrial homeostasis alterations in lens epithelial cells (LECs) of HMC. METHODS: The lens epithelium tissues of 20 patients with HMC and 20 control subjects with age-related cataracts (ARC) were collected. The real-time quantitative PCR and western blot assays were performed for gene expressions. Immunofluorescence (IF) assays were performed for mitochondrial marker TOM20, DNA damage marker 15A3, and autophagosome marker LC3. Transmission electron microscopy (TEM) was used to observe the changes in mitochondria morphology. Mitochondrial ROS, and mitochondrial membrane potential were detected by MitoSOX fluorescence, and JC-1 MitoMP staining, respectively. Rat lenses cultured in vitro were pretreated with CCCP and H2O2 (10 and 400 µM) for 24 h. RESULTS: The copy number of mtDNA was decreased in HMC patients compared to the ARC patients. Increased mitochondrial-oriented oxidative stress response was detected in LECs of HMC compared to that of ARC. Altered expressions of mitochondrial homeostasis and mitophagy markers, including TFAM, PGC1α, MFN1, MFN2, Drp1, PINK1, Parkin and LC3, were found in HMC patients. Reciprocally, no significant differences in the expression of BNIP3 and FUNDC1 were found between HMC and ARC patients. Importantly, TEM revealed that the obvious mitochondrial fission and mitophagy phenomena occur in the LECs of HMC patients compared to the ARC patients. Moreover, CCCP aggreated the mitoROS production and depolarized mitochondrial membrane potential in the H2O2-treated human lens epithelial cells line (SRA01/04); Most important, rat lens organ culture experiments indicated a significant increase in H2O2-induced lens opacity following mitochondrial uncoupling CCCP treatment. CONCLUSION: This study has identified for the first time the abnormal mitochondrial homeostasis in HMC, and provide a new perspective on the potential mechanisms of HMC, which occurs earlier and at a higher incidence rate than ARC.

Targeting scleral remodeling and myopia development in form deprivation myopia through inhibition of EFEMP1 expression.

The role of extracellular matrix (ECM) remodeling in the axial elongation associated with myopia has not been fully elucidated, although it is considered a significant factor. EFEMP1, a regulator of ECM, has been associated with various pathological conditions. This study aimed to examine the involvement of EFEMP1 in scleral remodeling during form deprivation myopia. The results indicate a progressive increase in EFEMP1 expression following prolonged form deprivation treatment, followed by a subsequent decrease upon recovery. To gain a deeper understanding of the mechanism of EFEMP1, we conducted transcriptome sequencing on primary scleral fibroblasts that were subjected to lentivirus-mediated overexpression of EFEMP1. Validation was performed using lentivirus-induced overexpression and shRNA targeting EFEMP1 in combination with LY294002, a PI3K inhibitor. Our findings suggest that EFEMP1 may be involved in the development of FDM by regulating the expression of the PI3K/AKT/MMP2 axis. The AAV-mediated injection of shEFEMP1 under Tenon's capsule in guinea pigs was observed to effectively delay the progression of myopia and posterior scleral remodeling. In contrast, the AAV-mediated overexpression of EFEMP1 exacerbated the development of myopia and resulted in further thinning of collagen fibers in the posterior sclera. In summary, adjusting EFEMP1 concentrations could potentially serve as a viable approach to prevent and treat myopia by influencing the remodeling process of the posterior sclera.

Insights into myopic choroidal neovascularization based on quantitative proteomics analysis of the aqueous humor.

BACKGROUND: Previous studies on the biomarkers of pathologic myopia choroidal neovascularization (pmCNV) development merely detected limited types of proteins and provide a meagre illustration of the underlying pathways. Hence, a landscape of protein changes in the aqueous humor (AH) of pmCNV patients is lacking. Here, to explore the potential mechanisms and biomarkers of pmCNV, we analyzed the clinical data and protein profile among atrophic (A) lesions, tractional lesions (T) and neovascular (N) lesions in myopic patients based on the ATN grading system for myopic maculopathy (MM). RESULTS: After investigating demographic data of our patients, a correlation was found between A and N lesions (R = 0.5753, P < 0.0001). Accordingly, groups were divided into patients without MM, patients with myopic atrophic maculopathy (MAM), and patients with pmCNV (N2a lesion). In proteomics analysis, the increased protein level of GFAP and complement-associated molecules in AH samples of the 3 groups also indicated that MAM and pmCNV shared similar characteristics. The GO enrichment and KEGG pathway analysis were performed, which mapped that differential expressed proteins mainly engaged in JAK-STAT pathway between the pmCNV group and two controls. Furthermore, we identified several potential biomarkers for pmCNV, including FCN3, GFAP, EGFR, SFRP3, PPP2R1A, SLIT2, and CD248. CONCLUSIONS: Atrophic lesions under pathologic myopic conditions demonstrated similarities to neovascularization development. Potential biomarkers including GFAP were associated with the pathogenesis of pmCNV. In summary, our study provides new insights for further research on pmCNV development.

Intraperitoneal Injection of MCC950 Inhibits the Progression of Myopia in Form-Deprivation Myopic Mice.

Myopia, one of the most prevalent ocular diseases worldwide, is projected to affect nearly half of the global population by 2050. The main cause of myopia in most patients is axial myopia, which primarily occurs due to the elongation of the eyeball, driven by changes in the extracellular matrix (ECM) of scleral cells. Previous studies have shown that NLRP3, an important inflammatory mediator, plays a critical role in regulating the expression of MMP-2 in the sclera. This, in turn, leads to a decrease in the expression of Collagen-1, a major component of the scleral ECM, triggering the remodeling of the scleral ECM. This study aimed to investigate the effect of MCC950, an inhibitor of NLRP3, on the progression of myopia using a mouse form-deprivation myopia (FDM) model. The FDM mouse model was constructed by subjecting three-week-old C57BL/6J mice to form-deprivation. The mice were divided into experimental (n = 10/group; FDM2M, FDM4M, FDM2W, and FDM4W) and control groups (n = 5/group). The experimental groups were further categorized based on the duration of form deprivation (2 and 4 weeks, labeled as 2 and 4, respectively) and the type of injection received (MCC950 or saline, labeled as M and W, respectively). MCC950 was injected at a concentration of 50 mg/mL, with a dose of 10 mg per kilogram of body weight. Meanwhile, the saline group received the same volume of saline. Refraction and axial length measurements were performed for each eye. The expression levels of NLRP3, caspase-1, IL-1ß, IL-18, MMP-2, and Collagen-1 in the sclera were assessed using immunohistochemistry and Western blotting. The intraperitoneal injection of MCC950 did not significantly affect refraction or axial length in normal mice (p > 0.05). However, in FDM mice, MCC950 attenuated the elongation of the axial length and resulted in a smaller shift towards myopia compared to the saline group (FDM4M vs. FDM4W, p = 0.03 and p < 0.05, respectively). MCC950 decreased MMP-2 expression (p < 0.05) but increased Collagen-1 expression (p < 0.05) in the experimental eyes when compared to the saline group. Within the MCC950 group, the expression of MMP-2 was increased in the experimental eyes at 4 weeks (p < 0.05), while that of Collagen-1 was decreased (p < 0.05), which is consistent with changes in refractive error. Immunohistochemical analysis yielded similar results (p < 0.05). MCC950 also reduced the expression levels of NLRP3 (p = 0.03), caspase-1 (p < 0.05), IL-1ß (p < 0.05), and IL-18 (p < 0.05) in the experimental eyes compared to the saline group. Within the MCC950 group, the expression levels of NLRP3 and caspase-1 were comparable between the experimental and control eyes (p > 0.05), whereas IL-18 expression was higher in experimental eyes (p < 0.05). IL-1ß expression was higher in the experimental eyes only at week 4 (p < 0.05). The intraperitoneal injection of MCC950 can inhibit the progression of myopia in FDM mice, possibly by regulating collagen remodeling in the sclera through the NLRP3-MMP-2 signaling pathway. Therefore, MCC950 holds promise as a potential therapeutic agent for controlling the progression of myopia.

Opsin 5 mediates violet light-induced early growth response-1 expression in the mouse retina.

Myopia is an abnormal vision condition characterized by difficulties in seeing distant objects. Myopia has become a public health issue not only in Asian countries but also in Western countries. Previously, we found that violet light (VL, 360-400 nm wavelength) exposure effectively suppressed myopia progression in experimental chick and mice models of myopia. The inhibitory effects of VL on myopia progression are reduced in retina-specific opsin 5 (Opn5) knockout (KO) mice. Furthermore, VL exposure upregulated early growth response-1 (Egr-1) expression in the chorioretinal tissues of chicks. However, the expression of EGR-1 and role of OPN5 in mice following VL exposure remain unclear. In this study, we examined whether VL exposure-induced EGR-1 upregulation depends on Opn5 expression in the mouse retina. EGR-1 mRNA and protein expressions increased in the mouse retina and mouse retinal 661W cells following VL exposure. These increases were consistently reduced in retina specific Opn5 conditional KO mice and Opn5 KO 661W cells. Our results suggest that OPN5 mediates VL-induced EGR-1 upregulation in mice. These molecular targets could be considered for the prevention and treatment of myopia.

Suppressive effect of nitric oxide synthase (NOS) inhibitor L-NMMA acetate on choroidal fibrosis in experimental myopic guinea pigs through the nitric oxide signaling pathway.

Myopia is one of the most prevalent eye diseases that seriously threaten the eyesight of children and adolescents worldwide. However, the pathogenesis is still unclear, and effective drugs are still scarce. In the present study, the guinea pigs were randomly divided into a normal control (NC) group, a lens-induced myopia (LIM) group, a NOS inhibitor (L-NMMA) injection group, and a NOS inhibitor solvent phosphate-buffered saline (PBS) group and the animals received relevant treatments. After 2- and 4-week different treatments, we noted that the refraction and choroidal thickness in the LIM group decreased compared with the NC group, whereas the ocular axial length increased significantly, and the choroid showed a fibrotic trend. The expression of NOS1, NOS3, TGF-ß1, COLI, and α-SMA at gene and protein levels was increased significantly in the choroid (all P < 0.05). After intravitreal injection of NOS inhibitor L-NMMA, we found that compared with the LIM group, the refraction and the choroidal thickness significantly increased, whereas the axial length reduced significantly, accompanied by an increase of choroidal thickness and an improvement of choroidal fibrosis. The expression levels of choroidal NOS1, NOS3, TGF-ß, COLI, and α-SMA were significantly reduced (all P < 0.05). In conclusion, the trend of choroidal fibrosis in LIM guinea pigs is positively correlated with the increase in axial length. The NOS inhibitor L-NMMA can alleviate the process of choroidal fibrosis in myopic guinea pigs by inhibiting NO signaling pathway.

A common cause for nystagmus in different congenital stationary night blindness mouse models.

In Nyxnob mice, a model for congenital nystagmus associated with congenital stationary night blindness (CSNB), synchronous oscillating retinal ganglion cells (RGCs) lead to oscillatory eye movements, i.e. nystagmus. Given the specific expression of mGluR6 and Cav 1.4 in the photoreceptor to bipolar cell synapses, as well as their clinical association with CSNB, we hypothesize that Grm6nob3 and Cav 1.4-KO mutants show, like the Nyxnob mouse, oscillations in both their RGC activity and eye movements. Using multi-electrode array recordings of RGCs and measurements of the eye movements, we demonstrate that Grm6nob3 and Cav 1.4-KO mice also show oscillations of their RGCs as well as a nystagmus. Interestingly, the preferred frequencies of RGC activity as well as the eye movement oscillations of the Grm6nob3 , Cav 1.4-KO and Nyxnob mice differ among mutants, but the neuronal activity and eye movement behaviour within a strain remain aligned in the same frequency domain. Model simulations indicate that mutations affecting the photoreceptor-bipolar cell synapse can form a common cause of the nystagmus of CSNB by driving oscillations in RGCs via AII amacrine cells. KEY POINTS: In Nyxnob mice, a model for congenital nystagmus associated with congenital stationary night blindness (CSNB), their oscillatory eye movements (i.e. nystagmus) are caused by synchronous oscillating retinal ganglion cells. Here we show that the same mechanism applies for two other CSNB mouse models - Grm6nob3 and Cav 1.4-KO mice. We propose that the retinal ganglion cell oscillations originate in the AII amacrine cells. Model simulations show that by only changing the input to ON-bipolar cells, all phenotypical differences between the various genetic mouse models can be reproduced.

Integrative Transcriptome and Proteome Analyses Elucidate the Mechanism of Lens-Induced Myopia in Mice.

Purpose: The purpose of this study was to investigate the underlying molecular mechanism of lens-induced myopia (LIM) through transcriptome and proteome analyses with a modified mouse myopia model. Methods: Four-week-old C57BL/6J mice were treated with a homemade newly designed -25 diopter (D) lens mounting by a 3D printing pen before right eyes for 4 weeks. Refraction (RE) and axial dimensions were measured every 2 weeks. Retinas were analyzed by RNA-sequencing and data-independent acquisition liquid chromatography tandem mass spectrometry. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation, and STRING databases were used to identify significantly affected pathways in transcriptomic and proteomic data sets. Western blot was used to detect the expression of specific proteins. Results: The modified model was accessible and efficient. Mice displayed a significant myopic shift (approximately 8 D) following 4 weeks' of lens treatment. Through transcriptomics and proteomics analysis, we elucidated 175 differently expressed genes (DEGs) and 646 differentially expressed proteins (DEPs) between binoculus. The transcriptomic and proteomic data showed a low correlation. Going over the mRNA protein matches, insulin like growth factor 2 mRNA binding protein 1 (Igf2bp1) was found to be a convincing biomarker of LIM, which was confirmed by Western blot. RNA-seq and proteome profiling confirmed that these two "omics" data sets complemented one another in KEGG pathways annovation. Among these, metabolic and human diseases pathways were considered to be correlated with the LIM forming process. Conclusions: The newly constructed LIM model provides a useful tool for future myopia research. Combining transcriptomic and proteomic analysis may potentially brighten the prospects of novel therapeutic targets for patients with myopia.

The Role of Retinal Dopamine D1 Receptors in Ocular Growth and Myopia Development in Mice.

Dopamine is a key neurotransmitter in the signaling cascade controlling ocular refractive development, but the exact role and site of action of dopamine D1 receptors (D1Rs) involved in myopia remains unclear. Here, we determine whether retinal D1Rs exclusively mediate the effects of endogenous dopamine and systemically delivered D1R agonist or antagonist in the mouse form deprivation myopia (FDM) model. Male C57BL/6 mice subjected to unilateral FDM or unobstructed vision were divided into the following four groups: one noninjected and three groups that received intraperitoneal injections of a vehicle, D1R agonist SKF38393 (18 and 59 nmol/g), or D1R antagonist SCH39166 (0.1 and 1 nmol/g). The effects of these drugs on FDM were further assessed in Drd1-knock-out (Drd1-KO), retina-specific conditional Drd1-KO (Drd1-CKO) mice, and corresponding wild-type littermates. In the visually unobstructed group, neither SKF38393 nor SCH39166 affected normal refractive development, whereas myopia development was attenuated by SKF38393 and enhanced by SCH39166 injections. In Drd1-KO or Drd1-CKO mice, however, these drugs had no effect on FDM development, suggesting that activation of retinal D1Rs is pertinent to myopia suppression by the D1R agonist. Interestingly, the development of myopia was unchanged by either Drd1-KO or Drd1-CKO, and neither SKF38393 nor SCH39166 injections, nor Drd1-KO, affected the retinal or vitreal dopamine and the dopamine metabolite DOPAC levels. Effects on axial length were less marked than effects on refraction. Therefore, activation of D1Rs, specifically retinal D1Rs, inhibits myopia development in mice. These results also suggest that multiple dopamine D1R mechanisms play roles in emmetropization and myopia development.SIGNIFICANCE STATEMENT While dopamine is recognized as a "stop" signal that inhibits myopia development (myopization), the location of the dopamine D1 receptors (D1Rs) that mediate this action remains to be addressed. Answers to this key question are critical for understanding how dopaminergic systems regulate ocular growth and refraction. We report here the results of our study showing that D1Rs are essential for controlling ocular growth and myopia development in mice, and for identifying the retina as the site of action for dopaminergic control via D1Rs. These findings highlight the importance of intrinsic retinal dopaminergic mechanisms for the regulation of ocular growth and suggest specific avenues for exploring the retinal mechanisms involved in the dopaminergic control of emmetropization and myopization.

The role of the serotonergic system in atropine's anti-myopic effects.

The muscarinic cholinergic antagonist atropine is the most widely used pharmacological treatment for the visual disorder myopia (short-sightedness), the leading cause of low-vision worldwide. This study sought to better define the mechanism by which atropine inhibits myopic growth. Although classified as a muscarinic-cholinergic antagonist, atropine has been found to bind and modulate the activity of several non-cholinergic systems (e.g., serotonin). Thus, this study investigated whether the serotonergic system could underly atropine's anti-myopic effects. Using a chick model of myopia, we report that atropine's growth-inhibitory effects can be attenuated by pharmacological stimulation of the serotonin system. This may suggest that atropine can slow the development of myopia through inhibiting serotonergic receptor activity. We also observed that pharmacological antagonism of serotonergic receptors inhibits the development of experimental myopia in a dose-dependent manner, further demonstrating that modulation of serotonergic receptor activity can alter ocular growth rates. Finally, we found that neither experimental myopia, nor atropine treatment, induced a significant change in retinal serotonergic output (i.e., synthesis, transport, release and catabolism). This may suggest that, although myopic growth can be inhibited through modulation of serotonergic receptor activity (by atropine or serotonergic antagonists), this does not require a change in serotonin levels. These findings regarding a serotonergic mechanism for atropine may have significant ramifications for the treatment of human myopia. This includes assessing the use of atropine in patients who are also undergoing treatment to upregulate serotonergic signaling (e.g., serotonergic anti-depressants).

Correlation analysis of aqueous humor metabolomics with myopic axial length and choroidal parameters.

BACKGROUND: To explore differential metabolites in the aqueous humor of patients with different axial lengths and their correlations with axial length and choroidal parameters. METHODS: In this study, we included 12 patients with axial lengths less than 24 mm, 11 patients with axial lengths between 24 and 26 mm, and 11 patients with axial lengths greater than 26 mm. We collected their aqueous humor samples during cataract surgery for liquid chromatography-mass spectrometry metabolomic analysis. Simultaneously, we collected relevant clinical parameters such as axial length, subfoveal choroidal thickness, and choroidal vascular index. Correlations between clinical data, differential metabolites, and clinical indicators were analyzed. In addition, we plotted receiver operating characteristic curves. RESULTS: The results showed that axial length was significantly negatively correlated with choroidal thickness (r=-0.7446, P < 0.0001), and that several differential metabolites were significantly correlated with certain clinical parameters. After analyzing receiver operating characteristic curves, 5-methoxytryptophol and cerulenin were found to have excellent discriminative power, demonstrating their potential as biomarkers. In the enrichment analysis, we found that the differential metabolites among each group were involved in several special pathways (Taurine and Hypotaurine Metabolism, Vitamin B6 Metabolism, Pantothenate, and coenzyme A Biosynthesis), suggesting that abnormalities in these metabolic pathways may play a role in the process of axial myopia. CONCLUSIONS: Our study identified alterations in certain metabolic pathways in different axial lengths. At the same time, we found several metabolites with significant correlation with clinical indicators, among which 5-methoxytryptophol and cerulenin were associated with axial myopia. CLINICAL TRIAL REGISTRATION: Registration date:11/04/2022. TRIAL REGISTRATION NUMBER: ChiCTR2200058575. TRIAL REGISTRY: The First Affiliated Hospital of the Zhejiang University School of Medicine.

Molecular Basis of Transglutaminase-2 and Muscarinic Cholinergic Receptors in Experimental Myopia: A Target for Myopia Treatment.

Myopia, a prevalent refractive error disorder worldwide, is characterized by the elongation of the eye, leading to visual abnormalities. Understanding the genetic factors involved in myopia is crucial for developing therapeutic and preventive measures. Unfortunately, only a limited number of genes with well-defined functionality have been associated with myopia. In this study, we found that the homozygous TGM2-deleted gene in mice protected against the development of myopia by slowing down the elongation of the eye. The effectiveness of gene knockdown was confirmed by achieving a 60 percent reduction in TGM-2 transcript levels through the use of TGM-2-specific small interfering RNA (siRNA) in human scleral fibroblasts (SFs). Furthermore, treating normal mouse SFs with various transglutaminase inhibitors led to the down-regulation of TGM-2 expression, with the most significant reduction observed with specific TGM-2 inhibitors. Additionally, the study found that the pharmacological blockade of muscarinic receptors also slowed the progression of myopia in mice, and this effect was accompanied by a decrease in TGM-2 enzyme expression. Specifically, mice with homozygous mAChR5, mAChR1, and/or mAChR4 and knockout mice exhibited higher levels of TGM-2 mRNA compared to mice with homozygous mAChR2 and three knockout mice (fold changes of 5.8, 2.9, 2.4, -2.2, and -4.7, respectively; p < 0.05). These findings strongly suggest that both TGM-2 and muscarinic receptors play central roles in the development of myopia, and blocking these factors could potentially be useful in interfering with the progression of this condition. In conclusion, targeting TGM-2 may have a beneficial effect regarding myopia, and this may also be at least partially be the mechanism of anti-muscarinic drugs in myopia. Further studies should investigate the interaction between TGM-2 and muscarinic receptors, as well as the changes in other extracellular matrix genes associated with growth during the development of myopia.

Effects of dopamine D2 receptor antagonists on retinal pigment epithelial/choroid complex metabolism in form-deprived myopic guinea pigs.

The retinal pigment epithelial (RPE)/choroid complex regulates myopia development, but the precise pathogenesis of myopia remains unclear. We aimed to investigate the changes in RPE/choroid complex metabolism in a form deprivation myopia model after dopamine D2 receptor (D2R) modulation. Guinea pigs were randomly divided into normal (NC), form deprivation myopia (FDM), and FDM treated with dopamine D2R antagonist groups. Differential metabolites were screened using SIMCA-P software and MetaboAnalyst metabolomics analysis tool. Functions of differential metabolites were analyzed using KEGG enrichment pathways. Relative to the NC group, 38 differential metabolites were identified, comprising 29 increased metabolites (including nicotinic acid, cytosine, and glutamate) and 9 decreased metabolites, of which proline exhibited the largest decrease. Pathway analysis revealed regulation of arginine/proline and aspartate/glutamate metabolism. Intravitreal D2R antagonist injection increased proline concentrations and activated arginine/proline and purine metabolism pathways. In sum, D2R antagonists alleviated the myopia trend of refractive biological parameters in form deprivation myopic guinea pigs, suggesting the involvement of dopamine D2R signaling in myopia pathogenesis. The RPE/choroid may provide glutamate to the retina by activating proline metabolism via metabolic coupling with the retina. Dopamine D2R antagonism may modulate proline/arginine metabolic pathways in the RPE/choroid and regulate metabolism, information presentation, and myopia.