Defects in the functions of LOXL1 are one of the major contributors to abnormal deposits of PXEM in ocular tissues. LOXL1 essentially maintains the homeostasis of fibrillar ECM via regulating the generation, maintenance and repair of the elastic connective tissue (49). LOXL1 essentially acts as a framework element ensuring spatially defined elastin deposition. Particularly, LOXL1 is involved in the crosslinking of elastin and collagen through its pro-peptide, which binds to both fibulin-5 and tropoelastin to target elastic microfibrils at elastogenesis sites (49). It has been demonstrated that LOXL1 is a vital factor in preventing age-related elastic fibre damage and loss of elasticity (50). Furthermore, LOXL1 expression is essential for standard IOP control, while deficiency causes modified conventional outflow physiology and ECM repair and homeostasis (51). Specifically, the fibrillar material deposits found in PEXS patients contain elastin and tropoelastin, suggesting the link between defects of LOXL1 and the pathogenesis of PEXS. Changes in LOXL1 activation can result in an excessive aggregation of elastic fibre fragments into PEXS eyes.

Further, LOXL1 deficiency was found in the eye, and its deficiency increases susceptibility to optic nerve damage (52). Since the dysregulation in elastic fibre production and crosslinking is hypothesized to be the major contributor to the pathogenesis of PEXS, LOXL1 expression and polymorphisms have also been linked to the pathogenesis of this syndrome. Besides this, OXL1-AS1, a long non-coding RNA (lncRNA) synthesised from the LOXL1 gene, also has been linked to the PEXS. The nuclear LOXL1-AS1 selectively bind to the mRNA processing protein, the heterogeneous nuclear ribonucleoprotein-L (hnRNPL). Both have a vital role in regulating total gene expression in eye cells. Interestingly, SNPs regulating the expression of LOXL1-AS1 have been found in the patients of PEXS, suggesting the vital role of the LOXL1 gene in the pathogenesis of PEXS (53).

Another critical protein involved in the ECM remodelling and the pathogenesis of PEXS is tumor growth factor-β1 (TGF-β1), a fibrosis-associated growth factor found in high levels, specifically in fibrotic diseases and experimental fibrosis models (54). Increased TGF-β1 levels were noted in the AH of PEXS patients, and it has been associated with the production of several elastic fibrillary elements, like fibrillin-1, that comprise the PEXM (55). Notably, TGF-b1 is one of the most vital factors that triggers the expression of both LOXL1 and fibrillin-1, which is the critical element of PEXS fibrils. Additionally, these factors also seemed to activate the construction of a specific elastic microfibrillar network into PEXS-like fibrils, suggesting the contribution of TGF-β1 in the PEXM deposition (56). Further, TGF-β1 expression is correlated with decreased degradation of ECM via regulation of the activities of matrix metalloproteases (MMPs) and their tissue inhibitors (TIMPs). Of note, TGF-β1 reduces the expression of MMP1 and MMP3 while increasing the expression of MMP2, TIMP1, and TIMP3, leading to reduced degradation of the newly synthesised matrix material. In patients with PEXS, inactive forms of MMP-2, MMP-3, and the active forms of TIMP1 and TIMP2 are higher than other MMPs. The aberrant expression of these tissue remodelling enzymes leads to insufficient degradation of excess matrix material leading to the accumulation of PEXM (57).

Studies suggest that TGF-β1 activation causes downregulation of clusterin (CLU), a molecular chaperone essential for folding denatured and misfolded proteins in the AH during the PEXM generation (58,59). CLU is a glycoprotein component of biological fluids and is found at higher levels in ocular cells. CLU isoforms act as an extracellular chaperone that reduces abnormal aggregation of proteins by favouring their unfolded state for proper refolding. Notably, the expression levels of CLU have been correlated with both physiological and pathological processes, including regulation of lipid transport, apoptosis, cell-cell and cell-matrix interactions, and OS. Interestingly, elevated levels of CLU are found in PEXM deposits, which are colocalised with exfoliation fibrils and LOXL1 (59,60). Furthermore, the expression of CLU decreases in PEX individuals, which could be responsible for reduced chaperone function and deposition of PEXM in the anterior segment of the eye (61).

Studies have suggested that LOXL1 activity is tightly regulated by fibulin-5 (FBLN5), an extracellular scaffold protein. FBLN5 plays a crucial role in the activation of LOXL1, thereby controlling the deposition of elastin in the ECM. Notably, FBLN5 activates LOXL1 via binding to the N-terminus of LOXL1. Studies have suggested that two polymorphisms that have been found in the noncoding part of the FBLN5 gene could be a risk factor for PEXS (37). Further, studies have shown that low mRNA and protein levels of FBLN5 in the lens of PEXS patients promote deposition of PEXM by affecting ECM dynamics. Remarkably, FBLN5 deficiency and the loss of interaction between FBLN5 and LOXL1 could cause accumulation of the inactive form of LOXL1, leading to the pathogenesis of PEXS.

OS is defined as excess reactive oxygen species (ROS) production in cells, mainly due to the imbalance in the generation and clearance of free radicals and reactive metabolites. The presence of active oxygen radicals in biological materials was first established in 1954 by Gerschman et al (62). Specifically, the toxic nature of oxygen was related to its partially reduced forms (63). Two years later, a hypothesis that oxygen radicals are produced as by-products of biological reactions were responsible for mutations, cellular damage, cancer, and ageing (64). The discovery of the enzyme peroxidase dismutase was the beginning of a new era for exploring the effects of ROS on living organisations (65). In the subsequent decade, extensive investigations revealed that ROS is capable of causing oxidative damage in DNA, lipids, proteins, and other cellular targets (66). Nowadays, it has become clear that living organisms have adapted to moderately increased levels of ROS and have also developed mechanisms for using them in numerous physiological functions. Free radicals are now products of normal cellular metabolism and play a dual role: either beneficial to cells and organisms or harmful, depending on the amount generated at a particular time (67).

In biological systems, OS typically occurs when ROS are overproduced or the antioxidant defense mechanisms are insufficient. The delicate balance between ROS's beneficial and harmful effects is critical to living organisms and is maintained by ‘redox regulation’. The redox regulation maintains homeostasis and protects living organisms from OS (68). Importantly, OS is essentially a disorder in the redox regulation (69), OS plays a significant role in biology and has been implicated in numerous pathophysiological processes (70). Depending on OS, a wide range of disorders may occur involving cellular dysregulation or altered processes such as inflammatory responses dysfunction, accelerated ageing, abnormal proliferation, carcinogenesis and even cell death (71).

Recently we have reviewed the critical role of ECM in pathogenesis and treatment and particularly the roles of ECM effectors and biochemical pathways involved in the development and the progression of the PEXS (Fig. 1) (8). Here we further focused on the emerging roles of OS in PEXS. The eye is a highly metabolic organ devouring large amounts of energy. OS can affect the eye due to its anatomical and functional features. Specifically, the structural characteristics of the anterior eye segment tissues render them susceptible to a number of risk factors that can lead to an oxidative status (20). Notably, the eye is one of the organs constantly exposed to environmental factors that induce ROS production. Its anterior part and mainly the cornea is directly exposed to harmful atmospheric oxygen, toxins, radiation, physical abrasion, air pollution, artificial light, cigarette smoke, fumes and gases from household cleaning products, toxic chemicals and some drugs (72,73). Further, the solar UVR consists of UVA (315-400 nm), UVB (280-315 nm) and UVC (100-280 nm) is the primary source of ROS in the eye. The cornea is exposed directly to UVR and absorbs all UVC, 80% UVB and 34% UVA. Besides this, AH absorbs some of the UVB, the lens absorbs 66% of UVA and 20% of UVB, and the retina absorbs only a minimal percentage of UVA (<1%), but no UVB or UVC (72). Absorption of UVR by ocular tissues, especially UVC and UVB, eventually leads to photochemical production of ROS [e.g. singlet oxygen (¹O₂), superoxide (O₂^•-), hydroxyl radical (OH^•), peroxyl radical (ROO^•)] (73,74) causing UVR-induced molecular modifications (e.g. chain breaking, pyrimidine and thymine dimers and protein crosslinks) associated with pathological ophthalmic conditions such as cataract, glaucoma and age-related macular degeneration (AMD) (75,76).

In addition to UVR, some chemotherapeutic, phototoxic or even herbal origin drugs and diagnostic dyes can induce the generation of ROS in the eye and thus cause early cataracts or transient vision loss (77). For example, a drug widely used in photodynamic tumour therapy, such as γ-cyclodextrin bicapped C₆₀ [(γCyD)₂/C₆₀, CDF0], can effectively produce ¹O₂ (78). Apart from its anatomical characteristics, the eye can also be affected by OS by virtue of its physical and metabolic characteristics. Notably, the mitochondria are a significant endogenous source of ROS in the eye, as it is a metabolically active organ that consumes large amounts of O₂. Additionally, the eye's transparent structures, such as the cornea, AH, lens, vitreous and retina, allow continuous photochemical production of ROS (79). The biomolecular effectors of OS are summarised in Table I.

The lens is particularly vulnerable to OS due to its continuous exposure to solar UVR and oxidants throughout its life. Since the lens has mostly fibrous cells, it does not regenerate after damage. Moreover, the reduced levels of antioxidant molecules in the lens nucleus and the absence of protein turnover lead to an impaired ability to repair. Thus, damages accumulate over time (80). Except for solar UVR, other sources of OS in the lens include smoke and oxidants from AH or those produced by the lens cells themselves. The lens acts as a filter and absorbs over 60% of UVA and 20% of UVB, thus preventing much of the radiation to reach the retina (73). As a result, the photooxidation of the thiol groups of the lens's crystallins forms disulfide bridges between the molecules, leading to protein aggregation and cataract formation (81) ROS in the lens [e.g., O₂^•-, hydrogen peroxide (H₂O₂), OH^•] can also be created endogenously through cellular metabolism in different cell compartments, such as mitochondria, peroxisomes and cytoplasm.

For example, O₂^•-can be produced by the typical electron transport system and the activity of cytochrome P450. The nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex with Rac GTPases, associated with activating plasma membrane receptors by external signals like growth factors, can also produce O₂^•-. In addition, growth factor receptors can be activated by the UVR and produce ROS. Intracellular H₂O₂ can be derived from superoxide dismutase (SOD) activity or created from ascorbate and O₂ in the presence of Fe⁺³ or even generated from the AH. Finally, the reaction of H₂O₂ with metal ions (M+) can lead to OH^• H O₂^•- via the Fenton reaction (82). However, the endogenous production of ROS in the lens does not necessarily lead to OS since low concentrations of H₂O₂ could play a role as a signal transduction factor in the differentiation of lens epithelial cells and also be a significant regulatory molecule of numerous vital enzymes, such as phosphatases and kinases (83). Under OS conditions, the selective oxidation of specific amino acids in the lens results in aggregation and degradation of proteins, reduction of water solubility, crosslinking, charge changes, etc. (84,85) can significantly contribute to cataract development (86).

Additionally, ROS causes increased oxidation of amino acid residues of methionine, cysteine, tryptophan and phenylalanine in the lens, as evidenced by the formation of protein disulfides kynurenine, o-Tyr and Met-SO (87,88). Specifically, the associated ageing increase in Met-SO formation is related to the loss of a number of protein activities that affect various functions of the lens (89,90). To protect against oxidation, the lens has evolved as an anaerobic system containing high concentrations of ascorbic acid and glutathione (GSH), an endogenous antioxidant molecule part of the antioxidant defence system (91). However, these defensive antioxidant molecules decrease with age, with GSH declining significantly in the lens nucleus. Subsequently, ascorbic acid is increasingly oxidised, leading to the accumulation of crystalline-bound advanced glycation end products (AGEs) that contribute to the cataractogenesis (92).

The metabolically active lens epithelial cells are primarily responsible for their defence against OS (93). UVA is the main responsible for ROS production in the lens epithelial cells. About 70% of UVA passes through the cornea towards lens epithelial cells reacting with NADPH and nicotinamide adenine dinucleotide (NADH), which are present in high concentrations in these cells producing ROS (e.g., ¹O₂, O₂^•-, H₂O₂) (94). Exposure of lens epithelial cells to UVA results in increased lipid peroxidation, decreased antioxidant enzymes e.g. catalase (CAT), reduced viability and cell death (95). UVB is estimated to be only 3% of the total UVR reaching the lens and can trigger apoptotic mechanisms in the lens epithelial cells and inactivate enzymes (96,97).

In addition to UV-induced photooxidation, oxidative damage to the macromolecules of lens epithelial cells is also caused by elevated levels of cellular oxidants produced by exposure to toxic chemicals and failure of antioxidant defences (97). Moreover, lens epithelial cells can be affected by the high levels of H₂O₂ in the AH (98). H₂O₂ and peroxynitrite (ONOO^-) are thought to be the essential oxidants of acute or chronic exposure of lens epithelial cells (97) and their elevated levels can cause oxidation-dependent inactivation of crucial enzymes such as CAT, proteasome, and arylamine N-acetyltransferases (NATs) (97,99). Oxidative damage to lens epithelial cells can also cause osmotic swelling of the lens and loss of its transparency (100).

ROS production in AH is mainly due to UVR and inflammatory processes in adjacent structures, e.g., surgery (101,102). The AH contains ascorbic acid, proteins, certain amino acids (e.g. tyrosine, phenylalanine, cysteine, tryptophan) and uric acid, which act as filters absorbing the majority of UVR, leaving only a tiny part of it reaching the structures of the eye (103). Despite the critical role of endogenous UV filters like ascorbic acid, their photooxidation can produce potent oxidising molecules, such as ¹O₂ and H₂O₂ (104). The increased concentration of H₂O₂ in the AH has been found to reduce GSH metabolism (105). It may cause damage to the corneal endothelium, the lens and the radial body, particularly the trabecular meshwork. In vitro studies have associated increased H₂O₂ levels with reduced AH drainage, resulting in glaucoma (79).

In contrast to the findings mentioned earlier, there is a view that OS does not cause permanent damage to the AH. Any alterations of vital components are considered to be part of its antioxidant defence and are relatively reversible after restoring optimal conditions. However, long-term exposure to OS results in loss of antioxidants and tissue damage (89).

OS plays a crucial role in the pathogenesis of several eye diseases, including PEXS. Disturbances in the delicate balance between ROS and antioxidant defense mechanisms of the eye may contribute to the development of PEXS. Evidence suggests that high malondialdehyde (MDA) levels, which are the end-product of polyunsaturated fatty acid peroxidation reaction and a marker of free radical-mediated lipid peroxidation, are found in the patients of PEXS (106-108). Similarly, high levels of thiobarbituric acid reactive substances (TBARS), the major breakdown products of lipid peroxides, were significantly higher in the AH samples collected from primary open-angle glaucoma patients (109). Besides, increased levels of AGEs are also observed in the AH and serum of PEXS patients (110,111).

Additionally, these specific oxidation and glycation products could trigger the glaucoma formation associated with PEXS (112). Specifically, these end products can induce ROS generation, thus resulting in the deterioration of trabecular meshwork cells. Therefore, correcting the OS-induced damage using antioxidants could be considered a therapeutic strategy to prevent glaucoma in PEXS patients (113). According to these findings, OS and its end products might be a vital factor in PEXS pathogenesis. Further, the levels of superoxide dismutase 2 (SOD2), aldehyde dehydrogenase 1a1 (ALDH1A1), and microsomal glutathione transferase 1 (MGST1) which are part of the essential antioxidant defense system, are high in the anterior lens capsule of PEXS patients (114). However, antioxidant enzymes, SOD and CAT were significantly lower in PEXS patients (110). Similarly, the levels of GSH decrease in the lens epithelial cells of PXES patients (115,116). The antioxidant system defence failure could result in inadequate OS response and pseudoexfoliation development.

OS elevates the levels of free radicals, TGF-b1 and other growth factors in the eye. It is a critical factor in developing fibrosis in the PEXS-affected eyes (117). Also, OS disrupts the balance between MMPs and TIMPs, leading to dysregulated ECM in the eye of PEXS patients (118). TIMPs imbalance has been implicated in various abnormal fibroblastic disorders (119), including the development of PEXM and PEXG (120). Studies have found that TGF-b1 may up-regulate OS and have a synergic external role in the PEXS development (121,122). Especially, TGF-b1-mediated upregulation of LOXL1 could promote fibrosis in the eye of PEXS patients. Further, studies have indicated the synergy between TGF-β1 and OS in the activation of the LOXL1 (20,122). Therefore it could be suggested that the abnormal ECM deposition can be triggered by OS, and TGF-b1 under the high-risk LOXL1 haplotype may contribute to the PEXS aggregates in the ocular tissues (20).

OS is also known to modify glutamine synthase and thus influence glutamate/glutamine metabolism, leading to increased neurotoxic concentrations of glutamate (123). Further, OS can also damage the mitochondria present in the cells of optic nerves resulting in a reduced energy supply. Considering the critical role of mitochondrial disfunction in glaucoma evolution, therapies targeting mitochondria with specific antioxidants may improve the survival of retina ganglion cells to protect them from glaucomatous degeneration (124). Indirectly, OS can also cause vascular changes resulting in impaired blood flow to the optic nerve, injury to trabecular meshwork that decreases AH outflow and elevated IOP and glial cell dysfunction (110,125,126). Overall these studies point to the regulation of OS to regulate the pathogenesis of PEXS.