*3.2.2. Antioxidant function*

Of the 25 dietary carotenoids found in human tissues and blood, the selectively high rate of absorption and accumulation of lutein and zeaxanthin in human retina remained unclear until the discovery of specific macular carotenoid-binding proteins. Bernstein et al. have demonstrated that tubulin, a hydrosoluble protein, could bind to both lutein and zeaxanthin, and may be involved in the high distribution of MPs in the retina, but presented relatively low binding affinity and specificity. As a result, the research team continued to identify carotenoid-binding proteins with higher affinity and specificity. Subsequently, glutathione S-transferase P1 (GSTP1) was identified to bind zeaxanthin in the macula specifically compared to GSTM1 and GSTA1, the members of GST protein family [24]. GSTP1 was further confirmed to prevent lipid membrane from oxidation. In 2011, steroidogenic acute regulatory domain protein 3 (StARD3), one of lipid transfer-related protein family, was discovered as the lutein-binding protein [25]. Further studies need to be carried out to reveal more functions of StARD3. Generally, GSTP1 and StARD3 selectively bind zeaxanthin and lutein, respectively, leading to the high concentration and stabilization in human retina. In addition, the retinoid transporters including inter-photoreceptor retinoid-binding protein (IRBP) and retinol binding protein 4 (RBP4) are believed to be involved in the transport of MPs from circulation to

**Figure 2.** The human eye. (A) A schematic diagram demonstrating the anatomy of the human eye [23]. (B) A schematic

image of optical coherence tomography (OCT) showing the vertical section of the center of the retina.

The peak value of MPs absorption is about 460 nm, which lies in the range of wave length of blue light (450–495 nm). Therefore, MPs can absorb 40–90% of incident high-energy, visible blue light depending on the concentration. The absorption offers protection from light-induced damages and reduction of light scatter in the retina. Junqhans A et al. [27] have investigated the efficacy of various carotenoids as the blue light filter using unilamellar liposomes with a

retina [26].

*3.2.1. Blue light filter*

176 Progress in Carotenoid Research

**3.2. Lutein and visual functions**

A free radical is defined as a molecule, atom, or ion containing an unpaired electron. Because of the unpaired electron in the outer shell, the free radical is chemically highly reactive and unstable. Therefore, the free radical will react with other substances, even with themselves to reach steady state. Free radicals generated from oxygen are called reactive oxygen species (ROS), including superoxide anion (O2 —), perhydroxyl radical (also known as hydroperoxyl radical, HO2 ), and hydroxyl radical (OH) [29]. Superoxide can be inverted into only hydrogen peroxide (H2 O2 ) and H2 O2 together with singlet oxygen (non-radical compound) by enzymatic and non-enzymatic reactions, respectively [30]. Singlet oxygen, perhydroxyl radical, and hydroxyl radical are oxidants causing oxidation of protein, DNA as well as lipid peroxidation in cell membrane lipid bilayer, resulting in damages to the integrity of biological membrane and subsequently cell necrosis [29]. In physiological condition, production and detoxification of ROS are balanced in the body. However, when the balance is disrupted, no matter the increase in ROS generation or reduction of endogenous antioxidants, damages in the body occur. Thus it has been defined as "oxidative stress".

The retina is constantly exposed to ROS due to its high consumption of oxygen, conversion of light photons into electrochemical signals, and a number of mitochondria in rods. Massive blood supplies to the choroid in the retina make it the highest oxygen uptake tissue in the human body. Continuous exposure to the light photons, especially the blue light, triggers photo-oxidative reactions and damages DNA in RPE cells. Mitochondria, which are believed to be the major site for the generation of ROS, are rich in the inner segments of rod cells. It has been estimated that about 5% activated oxygen electrons in mitochondria could leak out as they go through the complicated electron transport chain, forming superoxide radicals [31]. Furthermore, a high content of polyunsaturated fatty acids in the outer segments of rods makes it more prone to peroxidation. In general, retina exhibits high susceptibility to ROS, resulting in irreversible oxidative damages.

Depending on the unique structure of MPs, one of the major biological functions of MPs in the retina is the prevention from oxidative damages via either physical quenching of singlet oxygen or chemical scavenging of free radicals. In the process of quenching of non-radical compound, such as singlet oxygen, the energy of singlet oxygen is transferred to the molecules of MPs, leading to excited triplet state of MPs and ground state of oxygen. Subsequently, the MPs in the triplet state dissipate the energy and return into the ground state. Since it is a physical mechanism, the structure of MPs is not changed, thus can be reused in the quenching cycles. It has been estimated that among carotenoids, lutein can react with singlet oxygen more strongly [32]. In contrast to physical mechanism, scavenging of ROS is achieved through chemical reactions in two ways. First, ROS accepts the missing electrons from MPs in which electrons are available in the polyene chain, thus cannot induce oxidation of lipid, protein and DNA in cells. Second, lutein and zeaxanthin insert themselves into the cell membrane to pair the single electron in ROS, making the lipid bilayers more rigid. Lutein was found to insert into the biological membranes in perpendicular and parallel orientation, while zeaxanthin follows the perpendicular orientation in the lipid membrane [28]. It is the transmembrane alignment of MPs that reduce the susceptibility of lipid bilayers to oxidative injury and maintain the integrity and rigidity of biological membranes [33].

chloride)-induced chemical hypoxia [39]. Moreover, in the treatment of CoCl2

**4. Lutein and AMD**

to almost 3 million [42].

also correlated with AMD.

vision loss.

**4.1. AMD**

[41]. In general, lutein could protect retinal neurons from hypoxia-induced injury.

lutein not only improved cell viability and enhanced cell survival but also inhibited the formation of autophagosome [40]. In the rat model of retinal detachment, lutein preserved cells in ONL and rhodopsin expression, indicating its neuroprotective and anti-apoptotic effects

AMD is the leading cause of visual impairment in people 65 years and above in developed countries. It is a slowly progressive disease that affects the central retina or macula. As estimated by the United Nations, approximately 20–25 million people are affected by AMD across the world, and the prevalence inevitably rises with the increasing of aged populations. By the end of 2020, it is expected that only in the USA, the number of AMD patients will reach

AMD is a complicated, multifactorial ocular disease, and the exact etiology still remains unclear. However, a number of risk factors are thought to be related to the pathogenesis of AMD. Of all those factors, age is the most obvious risk factor. Both the incidence and prevalence of AMD increase with age. Many investigators revealed that the family members of AMD patients were more prone to develop this disease, demonstrating the genetic factors in the genesis of AMD. Furthermore, the incidence in Caucasians is higher than that in other ethnic populations. There is no apparent sex preference in AMD patients, although some studies have indicated that women may be more susceptible [43]. In addition to the unmodified factors mentioned above, several other factors that can be modified are also involved in the pathogenesis of AMD. Smoking is considered as a frequent environmental risk factor, which is proved to double the AMD risk through increasing the oxidative stress in the macula [44]. Excessive exposure to the sunlight can lead to lipid peroxidation on cell membranes. Hypertension, overweight or obesity, poor nutrition status, and cardiovascular diseases are

AMD is classified into a non-exudative or atrophic (dry) form, accounting for 90% of AMD, and an exudative (wet) form, accounting for only 10% of AMD. The atrophic form is characterized by the accumulation of drusen under the macula formed by photo-oxidation of lipids plus proteins, and progressive degeneration of RPE cells in the macula, affecting central vision to varying degrees. The exudative form is associated with choroidal neovascularization (CNV) in the submacular area and subsequent retinal hemorrhage, leading to severe central

The most destructive type of AMD is the exudative or wet form because of the sudden loss of vision. Therapies for the wet form of AMD mainly focus on halting the progression of CNV, of which intravitreal injection of anti-vascular endothelial growth factor (anti-VEGF) drugs have been widely adopted by ophthalmologists as a standard treatment due to the up-regulation of

on Müller cells,

179

Lutein and the Aging Eye

http://dx.doi.org/10.5772/intechopen.79604

ROS is directly or indirectly involved in the most pathological processes observed in the retina, including inflammation, neuron degeneration, angiogenesis, or cell apoptosis. In the process of inflammation, excessive generation of ROS has been found to simulate many pro-inflammatory pathways. Moreover, oxidative injury is also associated with certain downstream signaling pathways in inflammation. Our research team has evaluated the anti-inflammatory effects of lutein in mouse model of ischemia/reperfusion and demonstrated that several proinflammatory factors, including nuclear factor-kappa B (NF-κB), interleukin 1β(IL-1β), and cyclooxygenase-2 (Cox-2), from Müller cells were significantly decreased in lutein-treated group when compared with control group, suggesting protection effects of lutein in retinal ischemia/reperfusion damage was achieved by its anti-inflammatory property [34]. Similarly, supplementation of lutein and zeaxanthin decreased NF-κB activity, while increased levels of erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1), which are the key factors to initiate phase II antioxidant protection to eliminate oxidative stress, in rats fed with high fat diet [35].

## *3.2.3. Other functions*

In addition to the functions mentioned above, lutein also plays an important role in maintaining other visual performance. A huge number of studies have shown that lutein and/or zeaxanthin, or in combination with other antioxidants have improved visual acuity and contrast sensitivity in healthy, young adults, in subjects with AMD at early and/or advanced stage, and in people with diabetes. High levels of MPs have been reported to decrease the influence of bright lights via quick recover from bright lights and improvement of ability to see in glare conditions. Daily uptake of lutein (20 mg/day) for a year increased visual contrast and glare sensitivity in healthy Chinese drivers, thus benefiting driving or other vision-related tasks performed at night [17]. Furthermore, MPs are able to speed conversion of photic impulses into electrical impulses in retina as well as the transmission to the visual cortex in the brain by keeping neurons in healthy state [36].

Furthermore, lutein has been shown to have neuroprotective effects in the retina. We have reported that in mouse model of ischemia/reperfusion, lutein decreased the expression of nitrotyrosine, and nuclear poly(ADP-ribose) (PAR) in GCL and INL, which are the markers for oxidative stress; thus exhibited protection effects on cell loss and cell apoptosis in inner retinal neurons [37]. Similar results were observed in the cerebral ischemia/reperfusion injury [38]. We further used the in vitro model of oxidative stress and hypoxia to evaluate the neuroprotective function of lutein in retinal ganglion cells. Our data revealed that lutein could protect ganglion cells from either H2 O2 -induced oxidative stress or CoCl2 (cobalt chloride)-induced chemical hypoxia [39]. Moreover, in the treatment of CoCl2 on Müller cells, lutein not only improved cell viability and enhanced cell survival but also inhibited the formation of autophagosome [40]. In the rat model of retinal detachment, lutein preserved cells in ONL and rhodopsin expression, indicating its neuroprotective and anti-apoptotic effects [41]. In general, lutein could protect retinal neurons from hypoxia-induced injury.
