The Mechanosensitive Piezo1 Channel Mediates Mechanochemical Transmission in Myopic Eyes

Abstract

Purpose: To identify the expression of the mechanosensitive ion channel Piezo1 in the retina of guinea pigs with form deprivation myopia (FDM) and to investigate mechanisms by which Piezo1 channels might regulate myopia.

Method: Sixty 3-week-old guinea pigs were divided into four groups randomly: normal control, FDM, FDM + vehicle control (DMSO), and FDM + Piezo1 inhibitor (GsMTx4). Measurements of spherical equivalent (SE) and axial length (AL) of the guinea pig were taken using retinoscopy and A-scan ultrasound examination, respectively. Location of Piezo1 protein was determined using immunohistochemistry. The histological structure and thickness changes of the guinea pig retina were observed by hematoxylin and eosin. Expression of Piezo1 in the retina was detected using quantitative RT-PCR and Western blot. Reactive oxygen species (ROS) levels in the retina were measured using flow cytometry.

Result: After 4 weeks of form deprivation, the FDM group exhibited a significantly increased myopic degree and axial length compared with the normal control group (all P < 0.001), and had higher expression levels of Piezo1 and ROS than the normal control group (P < 0.001 and P = 0.002, respectively). Piezo1 protein expression was down-regulated in guinea pigs given GsMTx4 compared with the DMSO group (P = 0.037). Additionally, the GsMTx4 group showed lower myopic degree (P < 0.001) and lower ROS levels (P = 0.019) compared with the DMSO group.

Conclusions: The Piezo1 channel may be activated in the retinas of FDM guinea pigs and be involved in the development of myopia by regulating intraocular ROS levels.

The prevalence of myopia is increasing alarmingly worldwide, especially in East Asian populations.^1,2 The rapid progression of myopia dramatically increases the risk of high myopia, which is closely associated with potential blinding complications and poses a long-term burden on the economy and public health.³ However, the etiology and pathogenesis of myopia have not been elucidated. 

The sclera is the primary load-bearing connective tissue of the eye, providing stable mechanical support for delicate intraocular structures, such as the retina.⁴ However, during the progression of myopia, the thickness of the sclera decreases significantly as the axial length (AL) increases, which greatly decreases the scleral support for the retina.⁵ As a result, the retina is significantly stretched, especially at the posterior pole of the eye, owing to excessive lengthening of the AL and decreased scleral support.⁶ This process may involve the effects of mechanical force and mechanotransmission.⁷ Mechanotransmission depends on mechanosensitive ion channel, which induce specific regulatory responses by depolarizing cell membranes and triggering Ca²⁺ inward flow to mediate multiple physiological activities or disease development. Previous studies have found that various mechanosensitive ion channel, such as Piezo1, transient receptor potential vanilloid 4 channel (TRPV4), transient receptor potential canonical 1 channel, and TWIK-related arachidonic acid-sensitive K⁺ channel, are present in the retina.^8–11 Mechanical stretching activates these mechanosensitive ion channels and affects cellular function by regulating the calcium homeostasis and excitability of the cells. Piezo1 is a mechanically gated cation channel consisting of a central ion channel and a three-lobed propeller-like structure that converts mechanical stimuli into electrochemical signals. It plays an important role in regulating mechanical force-mediated cellular biological behavior. Compared with other mechanosensitive ion channels, the Piezo1 channel is more sensitive to certain mechanical stimuli.¹² It has been reported that elevated IOP up-regulates Piezo1 expression in the mouse retina and that activation of the Piezo1 channel inhibits neurite outgrowth in retinal ganglion cells (RGCs).¹³ It is worth noting that the IOP of guinea pigs did not change significantly after 4 weeks of form deprivation.¹⁴ This finding is also consistent with reports examining the relationship between IOP and myopia development in Chinese children, where the IOP was largely unrelated to myopia development in school-aged children.¹⁵ However, even if myopia does not result in a significant increase in IOP, the weakened scleral biomechanics caused by myopia may still make the retina more susceptible to the stretching (swelling) of the IOP. These findings suggest that the Piezo1 channel may play an important role in retinal perception of mechanical stimuli. 

In addition, the Piezo1 channel has previously been demonstrated to be involved in converting mechanical stimuli into reactive oxygen species (ROS) signals. Numerous studies have shown that mechanical stretch promotes increased ROS production.^16–19 ROS is one of the major oxidative systems in the oxidative stress reaction, which is closely associated with the development of myopia.^20–22 On the one hand, ROS can cause retinal damage, resulting in photoreceptor dysfunction and affecting refractive development²³; RGC axons are vulnerable to oxidative stress injury caused by ROS because of their high lipid content and high mitochondrial density.²⁴ Intrinsically, photosensitive RGCs are the main type of RGCs. It was found that the ablation of intrinsically photosensitive RGCs in the mouse retina induced myopia, indicating that RGCs are crucial to the development of myopia.²⁵ Therefore, it is speculated that the Piezo1 channel mediates the production of ROS induced by retinal stretch during the development of myopia. However, whether Piezo1 senses myopia-induced retinal stretching stimuli and how it is involved in myopia remains unclear. 

Therefore, we propose to construct a model of form deprivation myopia (FDM) to investigate the changes of Piezo1 expression, and the effects of the inhibition of Piezo1 channel on ROS production in the retinas of FDM guinea pigs. 

Materials and Methods

Animals and Grouping

Sixty healthy 3-week-old male pigmented guinea pigs (Cavia porcellus, British short-hair stock, tricolor strain), weighing 150 to 180 g, were provided by Chongqing Tengxin Biotechnology Co. The Experimental Animal Ethics Committee approved the study (NSMC 2022037), and all animal experiments followed the ARVO statement on the use of animals in ophthalmic and vision research. All guinea pigs had free access to food and water and were regularly fed vegetables and fruits. A daily 12-hour light/12-hour dark cycle (starting at 8:00 am) with approximately 500 lux. 

A random number table method was used to divide sixty guinea pigs into four groups: the normal control (NC; n = 15), FDM (n = 15), FDM + vehicle control (DMSO; n = 15) and FDM + Piezo1 inhibitor (GsMTx4, grammostola spatulata mechanotoxin 4; n = 15). The NC group's guinea pigs had both eyes left untreated, whereas the other three groups' right eyes were covered with translucent balloons to establish the FDM model and the uncovered left eye was assigned as the self-control (SC) group. GsMTx4, a mechanosensitive channel-blocking peptide, has been shown to potentially inhibit the activity of the Piezo1 channel by down-regulating the expression of Piezo1.²⁶ After the start of form-deprivation, 10% DMSO (Sigma, St Louis, MO, USA) and 10 µΜ GsMTx4^27,28 (MCE, Zelienople, PA, USA) solutions were injected into the vitreous cavity of the covered eyes of the DMSO group and GsMTx4 group, respectively, at 5 µL²⁹ every 3 days. Oxyfloxacin ophthalmic ointment (Santen, Osaka, Japan) is used twice daily for infection prevention. 

After 4 weeks, guinea pigs were anesthetized by inhaling excessive sevoflurane until the corneal and nociceptive reflexes, then the eyes were quickly enucleated. The eyes for hematoxylin and eosin staining and immunohistochemistry were placed in 4% paraformaldehyde (Solarbio, Beijing, China) and fixed for 48 hours at 4°C. The cornea, lens, vitreous body, retinal pigment epithelium, choroid, and sclera were rapidly removed from the eyes. The remaining retinal tissue used in the quantitative RT-PCR, Western blot and flow cytometry experiments was stored at −80°C. 

Biometric Measurements

The refraction and AL were measured at five time points (0, 1, 2, 3, and 4 weeks) in guinea pigs. Tropicamide eye drops (Santen) were administered three times at 5-minute intervals to paralyze the ciliary muscles, and the pupillary light reflex was checked using a penlight 30 minutes after the last drop. If the pupils could not contract promptly, it indicated that the cycloplegia was effective. Otherwise, additional eye drops were administered until the ciliary muscle was sufficiently paralyzed. The spherical equivalent (SE) (SE = spherical power + 1/2 cylindrical power) was then measured using a retinoscopy (66 Vision Technology Co., Suzhou, China). It should be noted that, during the measurement process, the guinea pig's line of sight should be fixed, and the retinoscopy lens should be aligned with the pupil. Then, the lens power is adjust to neutralization and the final power is recorded. 

A-scan ultrasound imaging (Quantel Medical, Auvergne, France) was used to measure the AL. Before the measurement, the surface of both eyes was anesthetized using oxybutynin hydrochloride eye drops (Santen). The measurement began when the probe touched the surface of the cornea without eliciting a blink reflex. If a blink reflex was observed, additional oxybutynin hydrochloride eye drops were administered to ensure adequate corneal anesthesia. During the measurement, the A-scan probe was held vertically to the cornea and gently placed at the center of the pupil to avoid depressing the cornea. Each eye was measured at least five times, and the average of the three data with the best repeatability was taken for further analysis. 

Hematoxylin and Eosin Staining

Paraffin sections of 5 µm thick were prepared. Staining was carried out according to the standard operating instructions of the hematoxylin and eosin staining kit (Biosharp, Hefei city, Anhui Province, China). The sections were dewaxed and hydrated, followed by hematoxylin for 3 minutes. Next, the paraffin sections were treated with 1% hydrochloric acid alcohol for 2 seconds. Then, eosin staining was performed for 1 minutes. After that, the sections were dehydrated and transparent. Finally, the sections were sealed with neutral resin for observation under a light microscope (Leica, Paris, France). 

Immunohistochemical Staining

Paraffin sections of 3 µm thick were prepared and stained according to the standard operating instructions of the Immunohistochemistry Kit (SA1027, Boster, Wuhan, China). The histological sections were dewaxed and hydrated, followed by antigen repairing for 20 minutes. The sections incubated with 3% hydrogen peroxide–methanol solution protected from light at 37°C for 15 minutes. The primary antibody used was Piezo1 (1:50, M1005-2; Huabio, Hangzhou, China) was then added overnight at 4°C. Anti-mouse secondary antibody (SA1027, Boster) was incubated at room temperature for 1 hour. Immunoreactivity was subsequently tested with the DAB reagent (ZSGB-BIO). Then the sections were restained with hematoxylin and dehydrated transparent. Finally, the sections were sealed with neutral resin and observed under a light microscope (Leica). 

Quantitative RT-PCR

Total RNA was extracted from retina of guinea pigs using Trizol (Beyotime, Jiangsu, China) and reverse transcribed to cDNA using PrimeScript RT Master Mix (Takara Bio, Shiga, Japan) according to the manufacturer's protocol. Using SYBR Premix Ex Taq II (Takara), amplify the designed primers on a Roche Light Cycler 480 instrument. The amplification was performed under the following cycling conditions: 95°C for 30 seconds, 40 cycles at 95°C for 5 seconds, 60°C for 30 seconds. and 97°C for 1 second. At the end of the amplification, the experimental results are analyzed according to the 2^−△△Ct method. The primer sequences in this study are as follows: Piezo1 forward primer 5'-GTGCGTGTGAATGCGCTCT-3' and reverse primer 5′- CAGGAGCCGGATCGAGTG-3', GAPDH forward primer 5′- GGTATTCCTTCTTCCCGTGC-3′ and reverse primer 5′-CCAAATCCGTTCACTCCGA-3′. Primer amplification efficiency of 2 is the optimum and 1.9 to 2.1 is preferable, according to the criteria of Roche Light Cycler 480. The GAPDH and Piezo1 primers used in this experiment had efficiencies of 1.97 and 2.006, respectively, and the efficiency met the standard. 

Western Blot Analysis

RIPA lysis buffer (Solarbio) was added to the retina and well cut up with scissors, followed by further lysis with ultrasound (Qsonica, Newtown, CT, USA). The supernatant was collected after centrifugation at 12,000×g for 15 minutes, and protein concentration was determined by the BCA kit (Beyotime). The samples were mixed with a 5× protein loading buffer (Beyotime) and then placed at 100°C for 15 minutes; the protein samples were loaded on 8% SDS-PAGE gel and transferred to polyvinylidene difluoride membranes. Then, the membranes were blocked with 5% nonfat milk at room temperature for 1.5 hours and incubated with the primary antibody (anti-Piezo1, 1:500, M1005-2, Huabio; anti-GAPDH, 1:5000, ET1601-4, Huabio) at 4°C overnight. After that, the membranes were incubated with a secondary antibody (1:10,000; Boster) for 1 hour at room temperature. Finally, the bands were exposed using the ECL developer solution (Biosharp), and the optical density was quantified using ImageJ software. 

Flow Cytometry

ROS levels were detected by ROS fluorescent probe (ThermoFisher Scientific, Waltham, MA, USA). Retina was mixed with a small amount of saline before being homogenized. Saline solution is added, mixed by blowing, and then strained through nylon mesh with a mesh size of 100 into a test tube. Saline washing was performed three times, followed by brief, low-speed centrifuging each time. The cell suspension was temporarily kept in a refrigerator at 4°C after the cell clumps were separated by filtration through 300 mesh nylon mesh. In a flow tube, place a 100-µL cell sample. ROS probe (500 µL) was added, then mixed well and incubated at 37°C for 20 minutes without light. After 20 minutes, add 1 mL PBS to the cells, mix well, and centrifuge at 350×g for 5 minutes, discarding the supernatant. Then 400 µL PBS was added and gently vortexed. Finally, flow cytometry was used to measure ROS levels. 

Statistical Analysis

The data are presented as the mean ± SD and analyzed by SPSS 21.0 software (SPSS, Inc, Armonk, NY, USA). The statistical graphs were made using GraphPad Prism 9 (GraphPad, San Diego, CA, USA). Paired t tests were used to assess the significance between the left and right eyes within groups; unpaired t tests were used to assess the significance between the remaining groups. A one-way ANOVA followed by the Bonferroni multiple comparisons was applied to analyze multiple groups. A P value of less than 0.05 was considered statistically significant. 

Results

Changes in SE and AL

Before myopia induction, all groups of guinea pigs were moderately hyperopic, and the differences in SE and AL were not statistically significant (P = 0.999 and P = 0.703, respectively). After myopic induction for 1 week, the FDM group began to show myopic drift. When inducted for 4 weeks, SE and AL of the form-deprived eyes increased significantly in the FDM group compared with the NC and SC groups (all P < 0.001). After 4 weeks of induction, the differences in SE and AL between the FDM group and DMSO group were not statistically significant (all P > 0.05). At the end of form deprivation, the GsMTx4 group had less myopic degree and shorter AL compared with the DMSO and FDM group (all P < 0.001) (Table, Fig. 1). 

Table.

 

Changes of SE and AL in Different Groups (mean ± SD)

 

Figure 1.

 

The changes of SE (A) and AL (B). NC (n = 15); SC (n = 15); FDM (n = 15); DMSO (n = 15), vehicle control group; GsMTx4 (n = 15), Piezo1 inhibitor group. The differences between all groups at each time spot were analyzed with one-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001.

The Morphology of the Retina

After 4 weeks of form deprivation, the retina and ganglion cell layer of guinea pigs in FDM group were thinner than those in NC and SC groups, the difference was statistically significant (all P < 0.05). In addition, compared with the control group, the structure arrangement of the inner and outer nuclear layers of the retina in FDM group was more disordered (Fig. 2). 

Figure 2.

 

Hematoxylin and eosin staining and quantitative analysis of total retinal thickness and ganglion cell layer thickness. The thickness was measured 1mm around the optic nerve. n = 4. Scale bar = 50 µm. (A) hematoxylin and eosin staining of the whole eye and retina of guinea pigs. (B) the statistical result of total retinal thickness. (C) the statistical result of ganglion cell layer thickness. GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; ns, no significance; OPL, outer plexiform layer; ONL, outer nuclear layer; IS/OS, inner and outer segments of the photoreceptor. *P < 0.05, **P < 0.01.

Distribution and Expression of Piezo1

Immunohistochemical staining showed the degree of expression of Piezo1 protein in the NC guinea pig eyes (Fig. 3). In the retinal tissues, Piezo1 protein mainly distributed in the ganglion cell layer, the inner nuclear layer, the outer plexiform layer, and the inner segment of the photoreceptors, where Piezo1 was strongly expressed in the ganglion cell layer (Fig. 3A). Negative control of Piezo1 immunostaining did not show a nonspecific reaction in the retina of guinea pig (Fig. 3B). 

Figure 3.

 

Expression of Piezo1 protein in guinea pig eyes. DAB staining shows the expression of Piezo1 protein. The NC group of guinea pigs was used in this experiment. Scale bar = 50 µm. (A) The expression of Piezo1 in the cross-section and retina of guinea pig eye. (B) Negative control of Piezo1 immunostaining. GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer; IS/OS, inner and outer segments of the photoreceptor.

The mRNA Level of Piezo1

After 4 weeks, the expression of retina Piezo1 gene was examined in each group. The relative expression of Piezo1 mRNA was significantly up-regulated in the FDM group compared with the NC group (all P < 0.001) and the SC group (all P < 0.001). The expression of Piezo1 mRNA was down-regulated after applying Piezo1 channel inhibitors, with statistically significant differences between the GsMTx4 and the DMSO groups (P = 0.025) (Fig. 4). 

Figure 4.

 

The expression of Piezo1 mRNA in retina of guinea pigs. n = 3. DMSO, vehicle control group; GsMTx4, Piezo1 inhibitor; ns, no significance. *P < 0.05, ***P < 0.001.

The Protein Level of Piezo1

The expression of Piezo1 protein in the guinea pig retina were further investigated. After 4 weeks of form deprivation, Piezo1 expression was significantly up-regulated in the FDM group compared with the NC group (P < 0.001) and the SC group (P = 0.004) (Fig. 5A). After the pharmacological intervention, the expression of Piezo1 protein was down-regulated in the GsMTx4 group compared with the DMSO group (P = 0.037); however, the expression of Piezo1 protein in the GsMTx4 group was still higher than the NC group (P = 0.001) (Fig. 5B). 

Figure 5.

 

The expression of Piezo1 protein in retina of guinea pigs. n = 3. (A) Expression level of Piezo1 protein in the NC group, SC group, and FDM group. (B) Expression level of Piezo1 protein in the NC group, DMSO group and GsMTx4 group. DMSO, vehicle control group; GsMTx4, Piezo1 inhibitor group; ns, no significance. *P < 0.05, **P < 0.01, ***P < 0.001.

The Level of ROS in the Retina

Using flow cytometry, we assessed the ROS levels in retina of guinea pigs. The results showed that ROS levels were significantly higher in the FDM group (12.50 ± 1.38%) than in the NC group (6.49 ± 0.42%) (P = 0.002) after 4 weeks of form deprivation. There was no statistically significant difference in the level of ROS between the DMSO group and the FDM group (P = 0.399). After inhibition of Piezo1 channel activity, ROS levels were reduced in the GsMTx4 group (8.31 ± 0.78%) compared with the DMSO group (11.50 ± 1.22%) (P = 0.019). In addition, the GsMTx4 group had higher levels of ROS than the NC group (P = 0.024) (Fig. 6). 

Figure 6.

 

The ROS levels in retina of guinea pigs. n = 3. Flow cytometry gating is shown in a forward (FSC-A) vs side scattering (SSC-A) plot in (A1–D1). The use of the DCFH-DA probe to detect ROS in each group (A2–D2). (A1–A2) the NC group; (B1–B2) the FDM group; (C1–C2) the DMSO group; (D1–D2) the GsMTx4 group. (E) The statistical results of ROS levels between the groups. DMSO, vehicle control group; GsMTx4, Piezo1 inhibitor group; ns, no significance. *P < 0.05, **P < 0.01.

Discussion

Mechanical force can trigger various biological signals involved in physiological functions, so when the transmission of mechanical force is abnormal, it may have adverse effects on the body.³⁰ Mechanosensitive ion channels play an important role in the perception and transmission of mechanical forces. Piezo1, one of mechanosensitive ion channels, effectively converts the mechanical stimulation of the retina into its bioelectrical and chemical signals.^27,31 The present study provides the first evidence that the expression of Piezo1 protein is up-regulated in the guinea pig retina following induction of myopia by form deprivation, suggesting that Piezo1 may be involved in the development of myopia. In addition, we showed that Piezo1 may influence ROS levels in the retina to some extent. 

As the AL increases, the retina is subjected to continuous mechanical stretching and thinning.³² Studies have shown that mechanical stretching on the retina can induce abnormal states and lead to RGC damage.^33,34 Lin et al.³⁵ recently showed that myopia affects the remodeling and rearrangement of retinal vascular, retinal astrocytes, and RGCs in marmosets, changing the RGC axon distribution and decreasing axonal survival. In our study, we showed that form deprivation caused excessive AL elongation and a refractive condition toward myopia in guinea pigs. Hematoxylin and eosin staining also showed that the total retinal thickness and RGC layer of guinea pigs in FDM group were thinner. Previous research discovered that mouse RGC contain the mechanosensitive ion channel TRPV4, and that TRPV4 agonists caused dose-dependent death of RGC.^8,9 In this study, Piezo1 protein expressed in the ganglion cell layer, inner nuclear layer, outer plexiform layer, and inner segment of the photoreceptors of guinea pig retinas. Moreover, Piezo1 was up-regulated in the FDM group compared with the control group at both mRNA and protein levels. Because FDM had no significant effect on IOP in guinea pigs,¹⁴ it can be excluded that Piezo1 up-regulation is the result of IOP elevation. As a result, mechanical straining of the retina may promote activation of Piezo1 channels, which could lead to apoptosis and altered cell function. In fact, it has been demonstrated that increased mechanical stress up-regulates Piezo1 expression in mouse retina and suppresses RGC neurite growth.²⁷ Therefore, it is possible that increased Piezo1 channel function is associated with myopia-induced RGC damage. To further clarify the effect of Piezo1 on myopia, we decreased the activity of Piezo1 by vitreous cavity injection of GsMTx4 (an inhibitor of Piezo1).³⁶ The results show that, when compared with the vehicle control group, the expression of Piezo1 was down-regulated and the AL was decreased in the GsMTx4 group, implying that GsMTx4 suppressed the function of Piezo1 to some extent and may inhibit the development of myopia. GsMTx4 is known to protect mouse RGC by decreasing mechanical stimulus-induced abnormalities in RGC axonal guidance and promoting synaptic growth.^27,37 Given that RGC can influence refractive development, this could be the reason why GsMTx4 delays the development of myopia.²⁵ 

The ROS system coexists with antioxidant systems in the human body to maintain the balance of redox reactions. Owing to the downregulation of antioxidants, oxidative stress caused by the inability of the body to eliminate excess ROS is an important factor in the pathogenesis of many ocular diseases, including myopia.²³ Previous research has shown that oxidative stress causes photoreceptors and other retinal nerve cells to deteriorate and die.²² With the progression of myopia, choroidal atrophy takes place, which causes retinal hypoxia and increased ROS generation, aggravating oxidative stress.³⁸ Unsaturated fatty acids make up a substantial portion of the retina, and lipid peroxides are the result of free radicals or ROS reacting with these fatty acids. In the subretinal fluid of individuals with retinal detachment, clinical research discovered a link between the levels of lipid peroxide and the myopic degree.³⁹ These studies all demonstrate how oxidative stress affects myopia. According to the results of the current study, FDM increased the level of ROS in retina of guinea pigs, which suggests that the tissue damage and apoptosis brought on by oxidative stress may be a cause for the development of myopia. 

It was interestingly demonstrated that the Piezo1 signaling pathway may affect the production of ROS. Previous research has showed that mechanical stretching enhances the formation of ROS in a variety of cells.^16–19 Prosser et al.⁴⁰ demonstrated that stretch induction significantly increased ROS fluorescence intensity in cardiac myocytes. Jiang et al.¹⁹ discovered that Piezo1 functions in cardiac myocytes as a mechanosensor and can transform mechanical stimuli into ROS signals instantly. In Piezo1 knockout cardiac myocytes, an agonist of the Piezo1 channel was unable to increase the amount of ROS, indicating that mechanical stretching can boost ROS generation through opening the Piezo1 channel. Our findings demonstrate that ROS levels in retina of the GsMTx4 group were lower than those of the vehicle control group, suggesting that suppressing the function of Piezo1 pathway may inhibit ROS production. However, the ROS levels were still lower in the NC group than in the GsMTx4 group, indicating that the Piezo1 channel may be responsible for regulating only partial ROS production in the eye. Taken together, we demonstrate that ROS in the retina may also be regulated by Piezo1 channels in vivo. However, it has been confirmed that myopia causes decreased choroidal perfusion, which directly results in retinal hypoxia and increased ROS, so the ROS in the retina detected in this study probably originated from the choroid.²¹ Therefore, additional in vitro studies in the retina to show the impact of Piezo1 on ROS production are required to demonstrate the existence of the Piezo1–ROS pathway in the retina. 

Conclusions

Our work investigated for the first time the connection between the Piezo1 channel and myopia, and discovered that Piezo1 expression was increased in retina of FDM guinea pigs. Additionally, the progression of myopia was slowed by the suppression of the function of Piezo1 channel, which also decreased the level of ROS in retina of FDM guinea pigs to some extent. These results suggest that the Piezo1 channel in the retina of FDM guinea pigs may be activated and regulate the level of intraocular ROS. These findings help to further investigate how biomechanical changes in the eye are involved in the development of myopia. 

Acknowledgments

Supported by the Project of Science & Technology from Department of Sichuan Province (No. 23NSFSC1940), the Strategic Cooperation of City and College (No. 22SXFWDF0003) and Affiliated Hospital of North Sichuan Medical College (No. 2023ZD010). 

Author Contributions: WZ performed the experiments, analyzed the data, and wrote the manuscript; CL, ZG, and QT interpreted the results. XX and HZ provided technical and material support. XL designed the study and reviewed and revised the paper. All authors read and approved the final paper. 

Ethics: This research was approved by the Animal Care and Ethics Committee at the North Sichuan Medical College (NSMC 2022037), and all treatment and care of animals were performed according to the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. 

Data Availability Statement: Related research datasets of this study are available from the first authors upon reasonable request. 

Disclosure: W. Zhong, None; C. Lan, None; Z. Gu, None; Q. Tan, None; X. Xiang, None; H. Zhou, None; X. Liao, None