Disrupted potassium ion homeostasis in ciliary muscle in negative lens-induced myopia in Guinea pigs.

Myopia is a main cause of preventable or treatable visual impairment, it has become a major public health issue due to its increasingly high prevalence worldwide. Currently, it is confirmed that the development of myopia is associated with the disorders of accommodation. As a dominant factor for accommodation, ciliary muscle contraction/relaxation can regulate the physiological state of the lens and play a crucial role in the development of myopia. To investigate the relationship between myopia and ciliary muscle, the guinea pigs were randomly divided into a normal control (NC) group and a negative lens-induced myopia (LIM) group, and the animals in each group were further randomly assigned into 2-week (n = 18) and 4-week (n = 21) subgroups in accordance with the duration of myopic induction of 2 and 4 weeks, respectively. In the present study, right eyes of the animals in LIM group were covered with -6.0 D lenses to induce myopia. Next, we performed the haematoxylin and eosin (H&E) staining to observe the pathological change of ciliary muscle, determined the contents of adenosine triphosphate (ATP) and lactate acid (LA), and measured the Na+/K+-ATPase expression and activity in ciliary muscles in both NC and LIM groups. Moreover, we also analyzed the potassium ion (K+) flux in ciliary muscles from 4-week NC and LIM guinea pigs. As a result, we found that the arrangements of ciliary muscles in LIM guinea pigs were broken, dissolved or disorganized; the content of ATP decreased, whereas the content of LA increased in ciliary muscles from LIM guinea pigs. Monitoring of K+ flux in ciliary muscles from LIM guinea pigs demonstrated myopia-triggered K+ influx. Moreover, we also noted a decreased expression of Na+/K+-ATPase (Atp1a1) at both mRNA and protein levels and reduced activity in ciliary muscles from LIM guinea pigs. Overall, our results will facilitate the understanding of the mechanism associated with inhibitory Na+/K+-ATPase in lens-induced myopia and which consequently lead to the disorder of microenvironment within ciliary muscles from LIM guinea pigs, paving the way for a promising adjuvant approach in treating myopia in clinical practice.

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.

Inhibitory effect of miR-138-5p on choroidal fibrosis in lens-induced myopia guinea pigs via suppressing the HIF-1α signaling pathway.

Myopia is one of the most common eye diseases in children and adolescents worldwide. Currently, there is no effective treatment in clinical practice. Ocular tissue fibrosis is involved in the development of myopia and this study aimed to investigate the effect of miR-138-5p on choroidal fibrosis in myopic guinea pigs via regulating the HIF-1α signaling pathway. First, guinea pigs were randomly divided into a normal control (NC) group, a lens-induced myopia (LIM) group, a LIM + miR-138-5p-carried Lentivirus treatment (LV) group, and a LIM + miR-138-5p-Vector treatment (VECTOR) group. All animals were induced experimental myopia with a -6.0 diopter lens except those in the NC group. Meanwhile, animals in the LV group were supplemented with 5 µl of miR-138-5p-carried Lentivirus, while those in the VECTOR group were only supplemented with the same volume of miR-138-5p-Vector. After myopia induction for 2 and 4 weeks, the refractive status and other ocular parameters of the guinea pigs were measured. Further, the expression of hypoxia-inducible factor (HIF)-1α, transforming growth factor (TGF)-ß, collagen I, hydroxyproline (HYP), interleukin 1 beta (IL-1ß), tumor necrosis factor alpha (TNF-α), and a-smooth muscle actin (α-SMA) in choroidal tissues was investigated. Results showed that the refraction and axial length of the experimental myopic guinea pigs increased, and choroid fibrosis aggravated after experimental myopic induction. miR-138-5p can efficiently decrease the refraction and ocular length, and ameliorate the choroidal fibrosis of the experimental myopic guinea pigs via downregulating the fibrosis-related TGF-ß1, collagen I, HYP, IL-1ß, TNF-α, and α-SMA expression through inhibiting the HIF-1α signaling pathway. Our results provide new insight into controlling myopic development using microRNAs in clinical practice.

Enhanced Apoptosis in Choroidal Tissues in Lens-Induced Myopia Guinea Pigs by Activating the RASA1 Signaling Pathway.

Purpose: This study aimed to explore the role of the RAS p21 protein activator 1 (RASA1) signaling pathway in apoptosis in choroid tissues from guinea pigs with negative lens-induced myopia (LIM). Methods: Biometric measurements were performed to examine refractive status, ocular parameters, and choroidal thickness (ChT) after myopia induction. The choroidal morphology was observed by hematoxylin and eosin (H&E) staining and TUNEL assay. The expression of the RASA1 signaling pathway at the mRNA and protein levels in choroidal tissues was measured by real-time quantitative PCR (qPCR) and western blot assays. Results: Compared with the normal control (NC) group, the ocular length of the guinea pigs in LIM increased remarkably, as did the myopic refraction. ChT decreased after myopia induction. H&E staining showed that the thickness and laxity of the choroidal tissues in LIM were strikingly reduced. The number of apoptotic cells in the LIM eyes was increased. Moreover, qPCR and western blot assays showed that the expression levels of both RASA1 and BCL-2-associated agonist of cell death (BAD) were higher in the LIM group than in the NC group, whereas the expression level of B-cell lymphoma 2 (BCL-2) was decreased after 2 weeks of experimental myopia. However, the trend of RASA1, BAD, and BCL-2 expression was reversed after 4 weeks of experimental myopia compared with levels after 2 weeks of experimental myopia. Conclusions: Results showed that the RASA1 signaling pathway is activated in choroid tissues in myopic guinea pigs. Activated RASA1 signaling induces high BAD expression and low BCL-2 expression, which in turn promotes apoptosis and ultimately causes ChT thinning in myopic guinea pigs.

Quantitative proteomic analysis of rat retina with experimental autoimmune uveitis based on tandem mass tag (TMT) peptide labeling coupled with LC-MS/MS.

Uveitis is a recurrent, inflammatory eye disease that occurs in the retina, iris, ciliary body and choroid. Currently, the detailed mechanism is still unclear. Proteomics can offer a powerful set of tools for the direct high-throughput study and a key contribution to the understanding of protein functions. This approach can also allow us to compare the protein profiling of the cells in healthy and diseased states that can be used to identify proteins associated with disease development and progression. In the present study, we first established an autoimmune uveitis (EAU) rat model. On day 12 after immunization, we isolated the rat retinas from both normal and EAU animals to collect total proteins. Using tandem mass tag (TMT) peptide labeling coupled with LC-MS/MS quantitative proteomics technique, we identified the differentially expressed proteins in EAU rat retinas, performed bioinformatics analyses, validated the expression of the COX1, NADH1, C3, and C9 proteins, and determined the adenosine triphosphate (ATP) levels. The results indicated that there were 190 upregulated and 103 downregulated proteins in EAU rat retinas. Bioinformatics analysis revealed the differentially expressed proteins were mainly involved in acute inflammatory response, visual perception and eye photoreceptor cell differentiation that were mainly related to complement and coagulation cascades, phagosome, PI3K-Akt signaling, and metabolic pathways. In conclusion, based on the TMT-based quantitative proteomics technique, the differentially expressed proteins in EAU rat retinas were mainly associated with complement and coagulation cascades and metabolic pathways. Our findings will facilitate the understanding of the pathogenesis of uveitis and will be useful for subsequent studies.