Glycative Stress Research
Online edition : ISSN 2188-3610
Print edition : ISSN 2188-3602
Received : October 9, 2020
Accepted : November 8, 2020
Published online : December 31, 2020
doi:10.24659/gsr.7. 4_283

Review article

Aging of the crystalline lens from the viewpoint of anti-glycation andanti-oxidation.
Eri Kubo
Department of Ophthalmology, Kanazawa Medical University, Ishikawa, Japan

Abstract

Ultraviolet rays, smoking, and drugs are considered to be risk factors for presbyopia and age-related cataracts. These
cause oxidative stress and glycative stress, therebyinducing disulfide bond (S-S bond) formation, followed by aggregation /denaturation and hardening of the nucleus of the lens protein, resulting in the formation of presbyopia and cataracts. Therefore,
antioxidant treatment and anti-glycation treatment can be expected to have a preventive / suppressive effect on cataracts andpresbyopia.
We have succeeded in delaying the progression of cataracts in the genetic cataract model, Shumiya Cataract Rat (SCR), by oral
administration of an antioxidant lutein (LU) and anti-glycation water chestnut (Trapa bispinosa Roxb.) extract (TBE), where
it was shown to increase the expression of the endogenous antioxidant protein peroxiredoxin 6 (Prdx6) and catalase in lensepithelial cells. Therefore, it can be effective in preventing cataracts to reduce not only the crystalline lens but also the wholebody of oxidative and glycative stress and increase the endogenous antioxidant protein by ingesting antioxidants and antiglycation supplements.

KEY WORDS: cataract, glycative stress, oxidative stress, lutein, water chestnut (Trapa bispinosa Roxb.)

1. Introduction

Cataracts are the number one cause of blindness inthe world and its prevalence is estimated to rise to 50 %by 2020 1). Cataracts are attributed to aging, ultraviolet(UV), radiation, inflammation, diabetes, smoking, steroids
intake, and highmyopia. Since the lens has a poor protein
turnover, the lens becomes opaque due to the accumulationof exogenous and endogenous oxidative stress and glycative
stress associated with aging 2, 3). Oxidative stress, i.e. free
radicals, reactive oxygen species (ROS), is known to cause
cell damage and protein denaturation in vivo. Therefore,
for the prevention of cataracts, the anti-cataract effects ofvarious supplements, phytochemicals, and functional foods
have been studied for the purpose of reducing these stresses
in the body and eyes 4). This paper outlines the results ofour research on the mechanism of age-related cataract
development associated with oxidative stress and glycative
stress, and on cataract prevention with lutein (LU) and water
chestnut (Trapa bispinosa Roxb.) extract (TBE).

2. Changes and functions of crystallin
protein in aging

About 90 % of the protein in the human lens is composedof α, β, and γ-crystallin, which plays a role in maintaining
the transparency of the lens and its refraction. In order tomaintain the transparency of the lens, it is necessary tocorrectly form the three-dimensional structure peculiar tothe crystallin protein. Therefore, a molecular chaperone ispresent to support correctly keeping the three-dimensional
structure. α- Crystallin has its chaperone activity, thus
suppressing the aggregation of partially denatured α, β, andγ-crystallin, as well as other proteins, and contributes to thetransparency of the lens 5). When the structure of α-crystallin
is changed by post-translational modification such asprotein oxidation, glycation, deamidation, and isomerization
due to the stresses, i.e. aging, UV exposure, its function
as a molecular chaperone is reduced. Furthermore, β and
γ-crystallin form large particles, abnormally aggregates,
and changes to insoluble proteins, causing light scattering

Corresponding to: Professor Eri Kubo, MD, PhD
Department of Ophthalmology, Kanazawa Medical University
1-1 Uchinada-machi, Kahoku-gun, Ishikawa 920-0293, Japan
TEL : +81-76 -286 -2211 (ext. 3412)
e-mail: kuboe@kanazawa-med.ac.jp

Glycative Stress Research 2020; 7 (4 ) : 283-286(c) Society for Glycative Stress Research_ 283 _

Aging of the Crystalline Lens, Anti-Gycation and Anti-Oxidationand swelling/ liquefaction of fibers in the lens, resulting
in the progression of cataracts. Since there is almost no
protein metabolism in the lens, it is necessary to maintainthe structure of α-crystallin protein normally, by keeping
the chaperone function, and suppress aggregate formation inorder to prevent cataracts.
Tryptophan, which is one of the constituent amino acidsof proteins, i.e. crystallin, has an absorption maximum
in the UVB region of 280 nm and is therefore affected by
UV rays. Tryptophan, by absorbing UVB, changes to be
kynurenine, which, under a further oxidized condition, thenbecomes a kynurenine derivative, 3-hydroxykynurenine.
This derivative absorbs UVA and releases its light energy
in the form of ROS, thus decomposing amino acids inthe protein, resulting in induction of structural changes
in the lens protein 6). Pyrenoxin, an anti-cataract eye drop
approved in Japan, has been reported to exert an effect ofmaintaining the transparency of the crystalline lens bysuppressing the photooxidation process from tryptophan
to N'-formylkynurenine and by competitively inhibiting
the cellular membrane dysfunction, which is induced byquinoid substances. The effectiveness of pyrenoxin remains
controversial. One report said that there is no scientificevidence for the anti-cataract effect of pyrenoxin 7), while
another report has shown that the use of pyrenoxin for earlycortical cataracts with an opacity area of 10 % or less had asignificant anti-cataract effect 8). It is necessary to select atherapeutic drug in consideration of the severity and type ofcataract.

3. Oxidative stress and cataracts
Oxidative stress such as free radicals and ROS, which
increase with aging, causes oxidation of the crystalline protein,
resulting in the formation of disulfide bonds (S-S bonds)
between the protein and glutathione (GSH), thus changing tooxidized glutathione. GSHs are maintained rich in the clear
lens, however with age, GSH levels become lower in the lens
nucleus compared to the outer cortical region. As a result, thelens nucleus is exposed to harmful conditions such as free
radicals.
This reduction in GSH promotes decreased lensantioxidant capacity, increased oxidative stress, formation of
S-S bonds between proteins, protein misfolding, aggregation,and insolubilization, leading to the initiation of cataracts 9, 10).
There exist proteins such as superoxide dismutase (SOD),
catalase, peroxiredoxin 6 (Prdx6), thioredoxin and glutathione
peroxidase (GPX), which are called antioxidant enzymes, inthe crystalline lens to eliminate oxidative stress. Vitamin E
and ascorbic acid are also present in the crystalline lens as
free radical scavenging substances. With aging, the decreasein these antioxidants and their activity elevates ROS
production and oxidative stress, and promotes the aggregationof lens proteins. Therefore, measures are required to prevent
cataracts as follows: one is the prevention of oxidative stressby avoiding UV exposure and by maintaining blood glucose;
another is the elimination and suppression of oxidative stressby the maintenance of endogenous antioxidant enzyme andby the administration of exogenous antioxidants.

4. Age-related cataracts and glycative stress
It has been reported that glycation of lens protein occurs
with aging 11). Lens proteins are prone to the formationof advanced glycation endproducts (AGEs), through theformation of carbonyl compounds, by reaction with reducing
sugars. AGEs usually accumulate in the crystalline lens withage, but the amount is more pronounced in cataract patients.
AGEs result in the formation of high molecular weightaggregation of crystalline in the lens, followed by inducing
lens opacity. AGEs are generally produced by reacting withsugars, but it is also known that kynurenine, a tryptophan
oxidation product of the lens, regulates AGE synthesis
by UV irradiation 11). Furthermore, it is reported that theoxidation of ascorbic acid in the lens caused by UV rays
induces AGE production, which becomes a photosensitizer,
resulting in inducing ROS production and oxidative stress in
the lens ( Fig. 1 ) 11).

5. Cataract prevention with lutein andwater chestnuts
LU and zeaxanthin (ZEA) contained in green leafy
vegetables are lipid-based antioxidants that protect the
crystalline lens by absorbing light in the outer layer of theretina and suppressing lipid peroxidation. Past large-scale
epidemiological studies have reported that LU intake reduces
the prevalence of nuclear cataracts 12, 13). In addition, astatistically significant preventive effect was shown on
nuclear cataracts in the LU and ZEA co-ingestion group in
a report by Cui et al., who meta-analyzed 7 cohort studies 14),
and a meta-analysis result from 14 cohort studies 15). LU andZEA cannot be produced in the body, therefore they must be
taken from diets and supplements. Foods rich in LU include
kale, paprika, spinach, parsley and other green-yellow
vegetables, however depending on eating habits, it may bedifficult to obtain a sufficient amount from the diet. AREDS2(Rep # 4) reported that in the group with the lowest dietary
LU and ZEA intake, the cataract surgery rate decreased dueto the intake of these supplements 16). Furthermore, the resultsof a meta-analysis have reported that low serum LU andZEA levels increase the risk of nuclear cataracts 17). It can besaid that LU and ZEA are supplements that are expected tohave a preventive effect on age-related cataracts.
In recent years, it has been reported that LU and TBE,
which is a peel extract of water chestnut (Trapa bispinosaRoxb.), has a delayed effect on sugar cataracts in a diabetic
rat model 18). TBE has been biochemically reported to haveanti-glycation effects, such as inhibition of AGE production19),
promotion of AGE cross-linking cleavage 19), and suppressionof α-crystallin glycation 20). Using SCR (Shumiya Cataract
Rat), which is a hereditary cataract model rat that develops
cataracts in about 66% due to mutation of lanosterol synthase,
we reported the effect of LU and TBE administration on
cataract prevention and the effect of inducing the expressionof antioxidant genes in the lens 21). The experiment consistsof two designs as follows: first, LU + TBE or Control(safflower oil) was administered to a 6-week-old cataractscheduled SCR (Cat +) and a non-cataract-free SCR (Cat −)for 4 weeks with an oral feeding needle; second, a 6-week-_ 284 _Glycative Stress Research

Aging, UV, DM, etc.
Water chestnuts
LuteinPrdx6
Catalase

Oxidative stress
Glycative stress(AGEs accumulation）

Age-related cataract
Fig. 1. Mechanism of cataract suppression by LU and TBE.

Due to aging, UV rays, DM, oxidative stress and glycative stress in the crystalline lens increase, and a cataract
develops. LU and TBE directly suppress these stresses; furthermore, they suppress oxidative stress in the lens by
inducing the endogenous antioxidant proteins Prdx6 and catalase. LU, lutein; TBE, water chestnuts (Trapa bispinosaRoxb.) extraxt; UV, ultraviolet; DM, diabetes mellitus; AGEs, advanced glycation endproducts.

old SCR (Cat +) was allowed to freely ingest a solid feed
containing LU, TBE, LU + TBE, and a standard solid feed(Control) for 3 weeks. As a result, lens opacity appeared in
SCR (Cat +) at 9-10 weeks of age, while the lens opacity inthe LU + TBE- administered group was significantly reduced
than that in the Control group and the TBE-only or LU-onlygroup. This result revealed that co-administration of LU +TBE delays the onset of cataracts in SCR. Furthermore, it
was demonstrated in vivo and in vitro that co-administrationof LU + TBE can induce the gene expression of antioxidant
enzymes as Prdx6 and catalase in lens epithelial cells. Since
the co-administration of LU + TBE showed stronger effects
than the single administration of LU or TBE in terms ofsuppressing cataract progression and inducing the expressionof antioxidant genes, it is speculated that the anti-cataract
effect may have been exhibited due to the synergistic actionof the anti-oxidation effect of LU and the anti-glycation
effect of TBE (Fig. 1). Therefore, LU + TBE can be expected
to be effective as an anti-cataract supplement, but it isnecessary to investigate the preventive effect of LU + TBE
on age-related cataracts in a large-scale epidemiologicalstudy in the future.

6. Other antioxidant supplements andcataract prevention
People often take vitamin C, vitamin A, vitamin E, or
β-carotene as antioxidant supplements because they reduce
oxidative stress as free radical scavengers. In 15 cohort
studies of vitamin C intake and cataract prevention effects,
8 studies showed that vitamin C intake reduced cataract
risk, while 7 studies found no significant risk reduction.
Taken together, these results showed a significant cataract
prevention effect in all types of cataracts, including thenucleus, cortex, and posterior subcapsule 22). However, there
are reports that high-dose (1,000 mg) vitamin C intake acts
as a prooxidant and increases the risk of cataracts 22), and thathigh-dose vitamin E and B increase the risk of cataracts 23, 24).
Attention is necessary to the excessive intake of the vitamin.
In other words, it suggests that the action of antioxidant
vitamins depends on the level of oxidative stress, and nonselective administration to individuals without excessiveoxidative stress has little effect or, conversely, risks.
In addition to vitamins, there are many food factors
with antioxidant capacity, and cataract prevention effects
have been reported at the animal experiment level. Animal
experiments and cell-culture studies have shown that cataracts
were suppressed by resveratrol, a type of polyphenol that has
antioxidant and anti-inflammatory effects 25), astaxanthin, atype of carotenoid 26), and curcumin, which has antioxidant
effects 27).

7. Conclusion
Cataracts are associated with aging and oxidative stressglycation not only in the crystalline lens but also throughout
the body. Antioxidant and anti-glycation supplements canprevent cataracts by improving the stressed condition in
the crystalline lens as well as the nutritional status of thewhole body. In the future, with the improvement of eating
habits as the first priority, it may be better to consider taking
supplements in the form of supplementing the shortage.

Conflict of interest
This study was funded in part by Santen Pharmaceutical
Co. Ltd (Osaka, Japan).

_ 285 _

Aging of the Crystalline Lens, Anti-Gycation and Anti-Oxidation

Reference
1) Mathew MC, Ervin AM, Tao J, et al. Antioxidant vitaminsupplementation for preventing and slowing the progressionof age-related cataract. Cochrane Database Cochrane
Database Syst Rev. 2012; 6(6): CD004567.
2) Spector A. Oxidative stress-induced cataract: Mechanismof action. FASEB J. 1995; 9: 1173-1182.
3) Spector A. Review: Oxidative stress and disease. J Ocul
Pharmacol Ther. 2000; 16: 193-201.
4) Chiu CJ, Taylor, A. Nutritional antioxidants and agerelated cataract and maculopathy. Exp Eye Res. 2007;
84: 229-245.
5) Bloemendal H, de Jong W, Jaenicke R, et al. Ageing andvision: Structure, stability and function of lens crystallins.
Prog Biophys Mol Biol. 2004; 86: 407- 485.
6) Mizdrak J, Hains PG, Truscott RJ, et al. Tryptophanderived ultraviolet filter compounds covalently bound tolens proteins are photosensitizers of oxidative damage.
Free Radic Biol Med. 2008; 44: 1108-1119.
7) Ibaraki N. Indications for drug therapy. Clinical Practice
Guidelines for Management of Cataract in Japan. J Jpn
Soc Cat Res. 2004; 16 (Supple.): 23-25. (in Japanese)
8) Kociecki J, Załecki K, Wasiewicz-Rager J, et al.
Evaluation of effectiveness of Catalin eyedrops inpatients with presenile and senile cataract. Klin Oczna.
2004; 106: 778-782. (in Polish)9) Lou MF. Redox regulation in lens. Prog Retin Eye Res.
2003; 22: 657-682.
10) Truscott, RJW. Age-related nuclear cataract-oxidation is
the key. Exp Eye Res. 2005; 80: 709-725.
11) Nagaraj RH, Linetsky M, Stitt AW. The pathogenic roleof Maillard reaction in the aging eye. Amino Acids. 2012;
42: 1205-1220.
12) Moeller SM, Voland R, Tinker L, et al. Associations
between age-related nuclear cataract and lutein andzeaxanthin in the diet and serum in the Carotenoids in the
Age-Related Eye Disease Study, an Ancillary Study of the
Women's Health Initiative. Arch Ophthalmol. 2008; 126:354-364.
13) Vu HT, Robman L, Hodge A, et al. Lutein and zeaxanthin
and the risk of cataract: The Melbourne visual impairment
project. Invest Ophthalmol Vis Sci. 2006; 47: 3783-3786.
14) Cui YH, Jing CX, Pan HW. Association of blood
antioxidants and vitamins with risk of age-related cataract:
A meta-analysis of observational studies. Am J Clin Nutr.
2013; 98: 778-786.
15) Sideri O, Tsaousis KT, Li HJ, et al. The potential roleof nutrition on lens pathology: A systematic review andmeta-analysis. Surv Ophthalmol. 2019; 64: 668-678.
16) The Age-Related Eye Disease Study 2 (AREDS2) ResearchGroup. Lutein/Zeaxanthin for the treatment of age-related
cataract AREDS2 randomized trial Report No. 4. JAMA
Ophthalmol. 2013; 131: 843-850.
17) Liu XH, Yu RB, Liu R, et al. Association between lutein
and zeaxanthin status and the risk of cataract: A metaanalysis. Nutrients. 2014; 6: 452- 465.
18) Kinoshita H, Sugawa T, Nanri R, et al. Trapa bispinosaRoxb. and lutein ameliorate cataract in type 1 diabetic
rats. J Clin Biochem Nutr. 2020; 66: 8-14.

19) Takeshita S, Yagi M, Uemura T, et al. Peel extract ofwater chestnut (Trapa bispinosa Roxb.) inhibits glycation,
degradesα-dicarbonyl compound, and breaks advanced
glycation end product crosslinks. Glycative Stress Res.
2015; 2: 72 -79.
20) Uemura T, Takeshita S, Yamada M. The effectiveness ofthe peel extract of water chestnut (Trapa bispinosa Roxb.)
in an α-crystallin glycation model with glyoxal. Glycative
Stress Res. 2017; 4: 104-108.
21) Ishida H, Shibata T, Kubo E, et al. Lutein plus water
chestnut (Trapa bispinosa Roxb.) extract inhibits thedevelopment of cataracts and induces antioxidant gene
expression in lens epithelial cells. Biomed Res Int. 2020;
2020: 9204620.
22) Rautiainen S, Lindblad BE, Morgenstern R, et al. Vitamin
C supplements and the risk of age-related cataract: A
population-based prospective cohort study in women. Am
J Clin Nutr. 2010; 91: 487-493.
23) Selin JZ, Lindblad BE, Bottai M, et al. High-dose
B-vitamin supplements and risk for age-related cataract:
A population-based prospective study of men and women.
Br J Nutr. 2017; 118: 154-160.
24) Ferrigno L, Aldiger R, Rosmini F, et al.: Italian-American
Cataract Study Group. Associations between plasma
levels of vitamins and cataract in the Italian-American
Clinical Trial of Nutritional Supplements and AgeRelated Cataract (CTNS): CTNS Report #2. Ophthalmic
Epidemiol. 2005; 12: 71-80.
25) Goutham G, Manikandan R, Beulaja M, et al. A focus onresveratrol and ocular problems, especially cataract: Fromchemistry to medical uses and clinical relevance. Biomed
Pharmacother. 2017; 86: 232-241.
26) Ishikawa S, Hashizume K, Nishigori H, et al. Effectof astaxanthin on cataract formation induced by
glucocorticoids in the chick embryo. Curr Eye Res. 2015;
40: 535-540.
27) Raman T, Ramar M, Arumugam M, et al. Cytoprotective
mechanism of action of curcumin against cataract.
Pharmacol Rep. 2016; 68: 561-569.

_ 286 _