1. Introduction
Refractive error significantly contributes to visual impairment. Myopia is the common cause of considerable pressure on healthcare systems and economies worldwide [
1]. Approximately 153 million individuals over the age of 5 experience varying degrees of distant vision impairment; of these, eight million individuals are blind as a result of untreated refractive errors [
2]. Myopia is a prevalent yet insufficiently managed ocular condition. While glasses, contact lenses, and refractive surgery can effectively correct most cases of myopia, roughly 33% of visual impairment cases are still caused by uncorrected refractive errors [
3]. High myopia is a key factor that leads to serious cataracts, macular choroidal degeneration, retinal detachment, and glaucoma. Patients with myopia of −6 D or less have a 3.2% annual risk of retinal detachment. The incidence of macular choroidal neovascularization in this group is 9-fold greater [
4,
5]. Myopia has been observed in the global initiative aimed at eliminating preventable blindness [
6].
Transforming growth factor β (TGF-β) is a multifunctional cytokine with essential function in modulating extracellular matrix synthesis, cellular development, inflammation, and apoptosis [
7]. TGF-β serves a crucial role in controlling the synthesis of matrix metalloproteinases, collagen, and proteoglycan in the extracellular matrix [
8]. A previous study demonstrated the presence of TGF-β isoforms within the mammalian sclera [
9]. Another study has connected heightened TGF-β2 function in the sclera to the progression of myopia generated by the use of negative lenses in guinea pigs [
10]. Clinical investigations indicate that patients with high myopia have elevated amounts of TGF-βs in the vitreous humor [
11]. Animal experiments have demonstrated that an increase in TGF-βs inside the choroid is associated with the elongation of the eye [
12]. TGF-β2 increases matrix metalloproteinase 2 (MMP2) and decreases collagen I expression, promoting myopia [
13]. TGF-β1 regulates the expression of MMP2 in fibroblasts by activating nuclear factor κB (NF-κB) [
14], and MMP2 has the ability to break down collagen types I, III, and V, leading to the restructuring of the sclera [
14].
The initiation of the complement system within the human organism is carefully managed to minimize overstimulation and the associated negative effects of inflammation. It is maintained at a steady and low turnover rate and plays an important function in preserving ocular immune privilege. Various eye illnesses, including age-related macular degeneration, autoimmune uveitis, diabetic retinopathy, and glaucoma, exhibit abnormal regulation of the complement system [
15]. The deregulation of the complement system has been associated with the development of myopia. Patients with pathologic myopia (−8 D to −25 D) showed significantly higher levels of C3 (
p = 0.004) and CH50 (
p < 0.001) [
16]. Myopic guinea pigs who received defocused negative lenses had considerably increased amounts of C1q, C3, and C5b-9 in their sclera [
17]. Activation of the complement system may promote extracellular matrix remodeling and contribute to myopia development [
17]. A meta-analysis was performed on eight different transcriptome databases to evaluate the link between lens-induced or form-deprivation myopia in chicks. The study found that the activation of the complement system is one of the biochemical pathways implicated in the advancement of myopia [
18]. It is determined that the eyes are highly susceptible to disruption of the complement system [
18]. Hence, abnormal activation of the complement system significantly contributes to the development of myopia. CD55 is a glycoprotein that is attached to the cell membrane by glycosylphosphatidylinositol. CD55 inhibits the activation of C3 and C5 by preventing the synthesis of C3 and C5 convertases and allowing their degradation. CD55 inhibits the activation of the complement system [
19,
20,
21].
Inflammasomes serve a crucial role in innate immunity by stimulating the development and synthesis of cytokines that promote inflammation, IL-1β and IL-18 [
22]. Studies highlight a significant crosstalk between the complement system and inflammasome activation [
23,
24]. For instance, complement activation products like C3a and C5a can act as signaling molecules, enhancing inflammasome activation [
23,
24]. C5a has been shown to bind to its receptor, C5aR, promoting NLRP3 inflammasome assembly and activation [
23,
24]. Inflammasome-derived cytokines can influence complement activation by activating NF-κB [
25,
26], creating a feedback loop that amplifies the immune response.
Based on the provided information, TGF-β, the complement system, and the inflammasome all involve molecules that possess the ability to facilitate the development of myopia. Our intention is to assess the possible correlation between these three factors in the development of myopia. By elucidating these pathways, we can identify potential therapeutic targets to modulate scleral remodeling and inflammation, potentially leading to new treatments for myopia.
2. Materials and Methods
2.1. Vector Production, Purification and Injection
CD55 AAV vector (Rat) (CMV), AAV Blank control vector (CMV), CD55 AAV siRNA pooled vector (Rat), and scrambled AAV siRNA control plasmids were obtained from Applied Biological Materials Inc. (Richmond, BC, Canada). An AAV-DJ helper free packaging system (Cell Biolabs, San Diego, CA, USA) was utilized to generate AAV virus in AAV-100 cells (Cell Biolabs, San Diego, CA, USA). AAV viruses were recovered using ViralBingTM AAV purification kit (Cell Biolabs, San Diego, CA, USA) and titrated by a QuickTiterTM AAV quantitation kit (Cell Biolabs, San Diego, CA, USA).
The rats were administered Zoletil at a dose of 5 mg/0.1 kg body weight for anesthesia and were then positioned on a heating pad to regulate their body temperature. A small opening was created in the sclera near the limbus using a 30-gauge disposable needle. Then, a 33-gauge blunt-tip needle connected to a Hamilton syringe was introduced through the scleral aperture into the vitreous area to provide intravitreal injections. The needle was left in the vitreous cavity for approximately 2–3 s to guarantee a complete injection, following which it was removed. Each eye was administered 1 µL of vector with a titer of 1 × 1012 GC/mL.
2.2. Myopia Animal Models
Three-week-old male Brown Norway rats were acquired from the National Laboratory Animal Center in Taipei, Taiwan, and randomly allocated to experiment groups; 210 rats were utilized in this investigation and were housed on a 12 h light/12 h dark cycle. Ten rats were assigned to each group for every experiment. All procedures were approved by the Institutional Animal Care and Use Committee of China Medical University (CMUIACUC-2019-173 approved on 29 December 2018). All operations followed the rules for the use of animals in ophthalmic and vision research.
2.2.1. Form-Deprivation Myopia Animal Model
In the monocular form-derived myopia study, fusion of the right eyelid caused myopia to develop. The concentration of TGF-β2 was measured by enucleating the eyes on days 0, 7, 14, and 21 and collecting the vitreous humor, retina, and sclera.
2.2.2. TGF-β-Induced Myopia Animal Model
PeproTech Inc., Cranbury, NJ, USA, provided recombinant TGF-β1 (100-21), 2 (100-35B), and 3 (100-36E), which were dissolved in balanced salt solution (BSS). The diluted solutions were kept at −20 °C until use. The recombinant TGF-βs were injected subconjunctivally on the first, seventh, and fourteenth days. The rats were administered Zoletil (5 mg/0.1 kg) to induce anesthesia. The eye’s surface was sterilized using iodophor and subsequently rinsed extensively with sterile BSS. In the myopic eyes, a total of 1.5 ng of recombinant TGF-βs was injected, while the control eyes were injected with BSS.
The axial lengths (AL) and refractive errors (RE) of the eyes were measured on days 0 and 21 of each study.
2.3. Ocular Biometry Assessment
A portable streak retinoscope was used to examine each eye’s refractive status. Tropicamide was administered to cause pupil dilatation. The animals were given Zoletil (5 mg/0.1 kg) to induce anesthesia. Ocular refraction was measured before and after the experiment. The initial experiment used five different degrees of lens. During the second cycle, the refractive condition was measured within a ±0.5-degree range using pupil luminescence measurement. The PacScan 300 Plus instrument from New Hyde Park, NY, USA, was employed to measure the axial lengths using A-scan ultrasonography. A mean value was determined from ten different measurements. After the study, the animals were euthanized via CO2 asphyxiation. The eyes were enucleated and either paraffin-embedded for immunofluorescence labeling or protein-extracted for Western blotting.
2.4. Determination the Level of Transforming Growth Factor β2 (TGF-β2)
Using RIPA buffer supplemented with a combination of protease and phosphoprotease inhibitors (Roche, West Sussex, UK), vitreous humor, retina, and sclera tissues were homogenized; 50 μg total protein was subjected to treatment with Sample Activation 1 (DY010; R&D systems, Minneapolis, MN, USA). The TGF-β2 concentration in the tissue lysate was determined by a Mouse/Rat/Canine/Porcine TGF-β2 Quantikine ELISA Kit (MB200; R&D systems, Minneapolis, MN, USA).
2.5. Immunofluorescence Staining
The eyes were submerged in a solution of 4% paraformaldehyde (Sigma-Aldrich Corp., St. Louis, MO, USA) in PBS overnight before being embedded in paraffin. Tissue blocks were chopped into 3 μm slices and placed on glass slides. The sections were treated to remove paraffin and then prepared for antigen retrieval by incubating them in a buffer solution (Epitope Retrieval Solution, Leica Biosystems, Buffalo Grove, IL, USA) for 20 min. They were subsequently submerged in a 3% hydrogen peroxide solution for 30 min (Polymer Detection System, Leica Biosystems, Buffalo Grove, IL, USA). After that, the sections were blocked with a solution containing 5% normal goat serum in PBS for 30 min at room temperature. Finally, they were incubated overnight at 4 °C with the specific primary antibodies against CD55 (sc-9156, RRID:AB_2075970, Santa Cruz, TX, USA), transforming growth factor β (TGF-β) (ab66043, RRID:AB_1143428, Abcam, Cambridge, UK), C3 (GTX72994, RRID:AB_374787, GeneTex, Irvine, CA, USA), collagen type I (collagen 1) (GTX20292, RRID:AB_384293, GeneTex, Irvine, CA, USA), C5 (GTX33052, GeneTex, Irvine, CA, USA), C5b-9, IL-1β (ab9722, RRID: AB_308765, Abcam), matrix metalloproteinase-2 (MMP-2) (ab37150, RRID:AB_881512, Abcam, Cambridge, UK), TNF-α (BS1857, RRID:AB_1662107, Bioworld, Dublin, OH, USA), NFκB (ab16502, RRID:AB_443394, Abcam) and NLRP3 (13158, RRID:AB_2798134, Cell Signaling Technology, Danvers, MA, USA). The slides were incubated with goat anti-Rabbit IgG (GTX213110-04 RRID:AB_2887579, Genetex) conjugated to biotin (Alexa Fluor 488 or 546) or Cy™3 (11-165-003 RRID:AB_2338000, Jackson ImmunoResearch Inc., West Grove, PA, USA) for 1 h at room temperature the next day. A confocal spectral microscope, equipped with a white light laser, was utilized as imaging equipment to take retinal pictures from the posterior segment of the eyes at magnifications of 20× or 63× (Leica TCS SP8 X).
2.6. Cell Culture
The human retinal pigment epithelial cells (ARPE-19) were acquired from the Bioresource Collection and Research Center in HsinChu, Taiwan (BCRC; BCRC-60,383). The cells were grown in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) at 37 °C and 5% CO2. The medium was changed every three to four days. For TGF-β treatment, the cells were distributed evenly in 6-well plates with a density of 1 × 105 cells per well with the medium changed every other day for 21 days. They were then exposed to TGF-β1, 2, or 3 at a concentration of 10 ng/mL, or a combination of TGF-β2 at 10 ng/mL and SB431542 at a concentration of 5 μM (Sigma-Aldrich), for the specified durations. Cell lysates were obtained to perform real-time quantitative PCR (qPCR) to quantitate gene expression.
2.7. Real-Time Quantitative PCR (qPCR)
The total RNA was extracted using the Qiagen RNeasy Mini Kit. To generate cDNA, 5 μg of RNA was reverse-transcribed using the Superscript First Strand Synthesis kit (Invitrogen, Carlsbad, CA, USA). For qPCR analysis, we utilized the Universal Probes Library system (Roche). The specific primer pair and probe number utilized are documented in
Supplementary Table S1. The transcript levels were normalized to the expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in all of the samples.
2.8. Western Blot Analysis
Retina tissues were collected after animals were sacrificed and homogenized using RIPA buffer, which was supplemented with a mixture of protease and phosphoprotease inhibitors (Roche, West Sussex, UK). The cell lysates were also lysed using radioimmunoprecipitation assay (RIPA) buffer. The protein concentrations were measured with the Bradford protein assay (Bio-Rad, Hercules, CA, USA). The extracts (15 μg) were separated by SDS-PAGE and transferred to PVDF membranes (0.45 μm; Millipore, Billerica, MA, USA). The membranes were blocked for an hour in PBST with 5% nonfat milk, and then primary antibodies were added, including p-NFκB (3033, RRID:AB_331284, Cell Signaling Technology, Danvers, MA, USA), Smad2/3 (3102, RRID:AB_10698742, Cell Signaling Technology, Danvers, MA, USA), NFκB (ab16502, RRID:AB_443394, Abcam), p-Smad2(Ser456/467)/Smad3(Ser423/425) (9510, RRID:AB_2193178, Cell Signaling Technology, Danvers, MA, USA), IL-1β (ab9722, RRID: AB_308765, Abcam), NLRP3 (13158, RRID:AB_2798134, Cell Signaling Technology, Danvers, MA, USA), and β-actin (4970, RRID:AB_2223172, Cell Signaling Technology, Danvers, MA, USA), overnight at 4 °C. After three PBST washes, the membranes were treated for one hour at room temperature with anti-mouse or anti-rabbit antibodies coupled with horseradish peroxidase. Proteins of interest were identified using an enhanced chemiluminescence assay kit, following the instructions provided by the manufacturer. Chemiluminescence was used with an ImageQuant LAS4000 mini (GE Healthcare, Little Chalfont, UK) equipment to view the reaction. The NIH in Bethesda, MD, USA, provided ImageJ 1.49 software, which was used to quantify the immunoblots.
2.9. Transepithelial Electrical Resistance
We followed the protocol established by Markert et al. [
27] with modification. ARPE-19 cells (1 × 10
5 cells) were seeded onto VTN-N-coated transwell inserts (0.25 μg/mL; ThermoFisher Scientific) placed in 12-well culture plates. The culture medium was replaced every other day for four weeks. Transepithelial electrical resistance (TEER) was measured using an EVOM2 epithelial voltohmmeter (World Precision Instruments, Sarasota, FL, USA). After four weeks, the mean TEER was 72.25 ± 3.32 Ω·cm
2. Only wells with TEER values around 70 Ω·cm
2 were used for subsequent experiments.
2.10. Statistical Analysis
ANOVA was used to statistically analyze the differences between the experimental groups with the GraphPad Prism program (Version 9, San Diego, CA, USA). Tukey’s multiple comparison tests are suitable to compare treatment groups in pairs. p values less than 0.05 were used to determine statistical significance for the means of the data.
4. Discussion
The study investigates the roles of transforming growth factor-β (TGF-β), the complement system, and inflammasome activation in the pathogenesis of myopia, proposing a significant interplay among these factors that potentially drives disease progression.
The present investigation revealed an elevation in the expression of C3 and C5 caused by TGF-βs, along with the suppression of CD55. Inhibiting TGF-β signaling prevented the elevation of C3 and C5 as well as the down-regulation of CD55. We also found an activation of Smad2 and NF-κB by the treatment of TGF-βs. Increasing NF-κB has been noted in promoting myopia development [
28,
29,
30]. While direct evidence for the TGF-β/Smad2 signaling pathway in promoting C3 and C5 expression is limited, the regulatory capabilities of the TGF-β/Smad pathway suggest a plausible role. TGF-β activates TRAF6 by Smad2, which in turn activates TAK1 (TGF-β-activated kinase 1), leading to the phosphorylation and activation of the IKK (IκB kinase) complex and subsequent NF-κB activation [
30]. We found an increase in TAK1 and NF-κB activation in the retina of TGF-β2-treated mice, while the activation was inhibited by CD55 administration (
Supplementary Figure S7). TGF-β can activate NF-κB independently of the canonical SMAD pathway. This involves the recruitment and activation of other signaling molecules, such as MAP kinases and PI3K/AKT, which can further activate NF-κB [
31]. NF-κB is known to promote the expression of C3 and C5 [
25]. The TGF-β/Smad pathway is also known to inhibit expression of certain genes, such as c-Myc [
32]. However, how the TGF-β/Smad pathway inhibits CD55 expression is not known. There are three potential mechanisms that the TGF-β/Smad pathway may use to inhibit gene expression. First, Smad2 may bind to the promoters of target genes, directly repressing their transcription. Second, Smad2 may interact with co-repressors and other transcription factors to inhibit gene expression directly. Thirdly, Smad2 may alter chromatin structure, making the promoter regions of certain genes less accessible to the transcriptional machinery. More detailed experiments should be performed to provide molecular mechanisms of either promoting or inhibiting complement protein expression by TGF-β2.
In interpreting the effects of TGF-β2 in our experimental myopia model, it is important to acknowledge that TGF-β is secreted in a latent form and requires extracellular activation to exert its biological functions. This activation process, involving proteolytic cleavage, integrin engagement, or conformational changes in the latent complex, is essential for enabling TGF-β to interact with its receptors. Although we administered recombinant TGF-β2, which is in its active form, we did not directly assess whether latent TGF-β2 requires activation in vivo under physiological or pathological conditions. Importantly, MMP2, which we observed to be upregulated following TGF-β2 treatment, is one of the major proteases capable of activating latent TGF-β. MMP2 cleaves the latency-associated peptide (LAP) from TGF-β, thereby converting it into its bioactive form [
33]. This suggests a possible amplification mechanism in our system: the administration of active TGF-β2 may induce MMP2 expression, which in turn promotes further activation of latent TGF-β already present in ocular tissues, especially within the sclera or retinal pigment epithelium. Such positive feedback could sustain or escalate TGF-β signaling, thus contributing to the observed upregulation of complement components and inflammasome activation. This possibility further supports the relevance of MMP2 as both a downstream effector and an upstream enhancer of TGF-β signaling, consistent with its role in scleral remodeling and myopia development. We acknowledge that the dynamics of latent TGF-β activation, particularly in a physiological context, were not fully examined in this study. Future work should include the evaluation of latent versus active TGF-β pools, as well as endogenous activators such as MMP2, integrins (e.g., αvβ6), or thrombospondin-1, to more precisely model the endogenous regulation of TGF-β activity in the progression of myopia.
TGF-β2 is a large protein (~25 kDa); it cannot cross the blood–retina barrier. However, previous studies have demonstrated that subconjunctival injection enables diffusion of proteins and macromolecules into the choroidal circulation via scleral and episcleral vasculature [
34]. This provides a plausible route through which TGF-β2 may reach RPE and choroidal targets, consistent with our proposed mechanism. Furthermore, TGF-β is known to possess auto-inductive properties, whereby exogenous TGF-β stimulates endogenous TGF-β expression in a feed-forward loop [
35]. This auto-induction could account for the elevated levels of TGF-β2 we observed in the apical compartment following basolateral exposure. We performed a directional secretion assay using ARPE-19 cells cultured on Transwell inserts. This setup mimics the in vivo polarity of RPE cells, forming tight junctions to distinguish apical and basolateral compartments. When active TGF-β2 was applied to the basolateral chamber, we observed a marked increase in TGF-β2 concentration in the apical medium after 24 h. Apically applied TGF-β2 also increased the TGF-β2 concentration in the basolateral medium (
Supplementary Figure S8). This directional release supports the physiological relevance of basolateral TGF-β exposure, as would occur via diffusion from the choroidal side. These findings reinforce the concept that subconjunctivally administered TGF-β2, while not directly entering the retina, can stimulate intraocular signaling cascades via the choroid–RPE axis, leading to increased local TGF-β expression and downstream pathological remodeling relevant to myopia progression.
Beyond its role in myopia, TGF-β is involved in a variety of ocular disorders. Its broad impact on cellular processes such as extracellular matrix (ECM) remodeling, differentiation, apoptosis, and proliferation makes it a critical factor in the pathogenesis of several eye conditions. One notable condition where TGF-β is implicated is cataract formation, particularly in high myopia. After cataract surgery, capsular contraction syndrome (CCS) has been linked to greater levels of TGF-β2 in the aqueous humor of myopic eyes. This syndrome is characterized by the shrinkage and opacification of the lens capsule, leading to impaired vision and potential intraocular lens decentration. TGF-β2 promotes the transformation of lens epithelial cells into myofibroblasts, contributing to capsular fibrosis and contraction [
36]. In the context of retinal diseases, TGF-β is involved in the progression of proliferative vitreoretinopathy (PVR), a condition that can follow retinal detachment. TGF-β2, in particular, stimulates the proliferation and migration of RPE cells and fibroblasts into the vitreous cavity, leading to the formation of fibrotic membranes that contract and cause recurrent retinal detachments. The signaling pathways activated by TGF-β2 in PVR include the Smad-dependent pathway, which regulates the transcription of genes involved in fibrosis and ECM production [
37]. Age-related macular degeneration (AMD) is another condition where TGF-β signaling plays a significant role. In the early stages of AMD, TGF-β helps maintain the homeostasis of the RPE and choroidal tissues. However, in the advanced stages, dysregulated TGF-β signaling can contribute to the formation of choroidal neovascularization, a hallmark of wet AMD. TGF-β promotes angiogenesis through the induction of vascular endothelial growth factor and other pro-angiogenic factors. This neovascularization leads to severe visual impairment if left untreated [
38,
39]. TGF-β is also involved in the pathogenesis of glaucoma, particularly in the remodeling of the trabecular meshwork (TM) and the optic nerve head. Elevated TGF-β2 levels in the aqueous humor of glaucoma patients induce ECM deposition in the TM, increasing outflow resistance and intraocular pressure. This ECM remodeling is mediated by TGF-β-induced activation of connective tissue growth factor and subsequent fibrotic changes [
40]. In summary, TGF-β is a pivotal cytokine in the pathogenesis of various eye diseases, influencing processes from ECM remodeling to cell proliferation and differentiation. Its role in fibrosis and angiogenesis underscores the potential of TGF-β as a therapeutic target. Inhibiting TGF-β signaling pathways could offer new treatment strategies for managing conditions like CCS, PVR, AMD, and glaucoma, thereby preserving vision and improving patient outcomes.
However, the regulatory activity of TGF-β should be noted. TGF-β2 plays a central role in maintaining immune privilege in the eye, primarily through its regulation of regulatory T cells (Tregs) [
41]. TGF-β2 is the dominant isoform of TGF-β in the ocular environment, particularly in the aqueous humor, where it contributes to the immunosuppressive milieu necessary to prevent destructive inflammation that could compromise vision. This cytokine is critical in the induction and maintenance of Tregs, which are essential for immune tolerance and preventing autoimmune responses [
42]. TGF-β2 induces the expression of Foxp3 in CD4+ T cells, converting them into Tregs capable of suppressing effector T cells and maintaining immune homeostasis [
41]. In the eye, RPE cells and other ocular tissues produce TGF-β2, which not only acts locally to suppress immune responses but also promotes the differentiation of Tregs in both the anterior and posterior segments of the eye. These Tregs then suppress bystander T cells through mechanisms that include direct cell-to-cell contact and the secretion of inhibitory cytokines like IL-10 [
41]. The protective role of TGF-β2 in ocular immune privilege is further demonstrated in experimental models where the administration of recombinant TGF-β2 can prevent or reduce the severity of autoimmune uveitis by enhancing the population of Tregs in the affected tissues [
38]. Therefore, TGF-β2 not only supports immune privilege by directly suppressing inflammation but also by fostering a population of Tregs that perpetuates this immunosuppressive state, highlighting its importance in ocular health and disease.
Accordingly, TGF-β2 functions as positive and negative regulator in the eye. The homeostasis of TGF-β2 is important to maintain normal visual function. Therefore, simply promoting or inhibiting TGF-β2 or its signaling pathway may cause detrimental effects on visual function. It is difficult to determine the most suitable concentration or signaling responses for TGF-β2 for any individual due to the multiple functions of TGF-β2. Blocking the key pathways altered by aberrant TGF-β2 signaling would be a better choice.
The interplay between TGF-β and inflammasome activation, particularly through the NLRP3 pathway, forms a self-perpetuating vicious cycle that significantly contributes to pathological conditions like fibrosis [
43]. This process begins with TGF-β enhancing the expression of NLRP3 and pro-IL-1β through the activation of TAK1-NF-κB signaling. Subsequently, TGF-β also increases intracellular ROS levels via the Smad-NOX4 axis, which serves as a secondary signal for the activation of the NLRP3 inflammasome [
44]. Once activated, the NLRP3 inflammasome promotes the cleavage of pro-IL-1β into its active form, IL-1β. This IL-1β further stimulates the secretion of TGF-β, creating an autocrine loop that sustains and amplifies both TGF-β production and NLRP3 inflammasome activation [
44]. This loop can drive chronic inflammation and fibrosis, as seen in hepatic stellate cells, where the continuous production of TGF-β perpetuates the fibrotic response. Inhibiting key components of this cycle, such as TAK1 or IL-1R, has been shown to disrupt this pathological feedback, highlighting potential therapeutic strategies for conditions driven by this TGF-β–inflammasome axis. We and others have indicated that inflammasome activation promotes myopia development [
45,
46].
Accordingly, TGF-β2 can activate the inflammasome directly, which does not involve the complement system. CD55 is primarily known for its role in regulating the complement system by preventing the formation of the membrane MAC. However, it has also been implicated in modulating various signaling pathways, including c-Jun N-terminal kinase, Janus kinase/signal transducers and activators of transcription, mitogen-activated protein kinase/NF-κB, and lymphocyte-specific protein tyrosine kinase pathways [
47]. Interfering with these pathways, especially the NF-κB signaling pathway, may alter the TGF-β–inflammasome axis.
This study highlights the role of TGF-β2 in promoting myopia through the activation of complement components C3 and C5 and suppression of CD55, leading to enhanced inflammasome activity. The overexpression of CD55 effectively counteracts these effects, suggesting that targeting the complement system, particularly CD55, could be a novel therapeutic strategy for myopia. The findings underscore the complex interplay between TGF-β2, the complement system, and inflammasome activation in myopia development, providing a foundation for future research aimed at developing targeted treatments for this condition.