**12. Apo ε4 allele status and AMD**

370 Lipoproteins – Role in Health and Diseases

Much of the debris that accumulates in BrM in the form of BlinDs is composed of lipoproteins and lipoprotein particles.[14] It has been found that almost 60% of the total cholesterol within these lipoproteins is esterified cholesterol.[96] Furthermore, the esterified cholesterol within BrM was enriched between 16 and 40-fold compared to plasma. If these extracellular lipid deposits had been derived from plasma, more than 90% of the phospholipid would be phosphatidylcholine, whereas in actual fact, these lipoproteins are comprised of less than 50% phosphatidylcholine.[96] Indeed, the composition of drusen, which are essentially large BlinDs, has been shown to include esterified and unesterified cholesterol, and multiple apolipoproteins, including apolipoproteins B, A-I, C-I, C-II, and E, appearing with frequencies ranging from 100% (ApoE) to approximately 60% (A-I).[88;91;97;98] Interestingly, ApoC-III, although abundant in plasma, is present in fewer drusen (16.6%) than ApoC-I (93.1%), which is not present in plasma in large quantities, indicating either a specific retention of plasma-derived apolipoproteins within drusen, or an intraocular source for these apolipoproteins.[93] It is now understood that the majority of lipoproteins in BrM have undergone intracellular processing within the RPE prior to secretion as neutral lipids, mainly esterified cholesterol.[99;100] The RPE origin has been definitively shown by two groups using metabolic labelling and immunoprecipitation in ratderived and human-derived RPE cell lines that were shown to secrete full-length ApoB.[101;102] This evidence is further strengthened by the finding of microsomal triglyceride transfer protein within native human RPE, indicating that the RPE is capable of secreting lipoprotein particles.[102] The pattern of lipid deposition in BrM with age, in which debris appears firstly in the elastic layer and then fills in towards the RPE, is also

The hydrophobic nature of the age-related thickening of BrM has been implicated in the aetiopathogenesis of AMD. In the case of Apo E, it is noteworthy that ApoE4 presents a positive charge relative to both ApoE2 and ApoE3. ApoE4 possesses arginine at residue 112 of the amino acid sequence, whereas ApoE3 possesses cysteine at this position, and in the case of ApoE2, the most frequent variant has cysteine instead of the normally occurring arginine at residue 158. Thus, ApoE3 presents a neutral charge, and ApoE2 a negative charge, relative to ApoE4.[53] Souied *et al.* suggested that this difference in charges between the ApoE isoforms

It appears that Müller cells are the most prominent biosynthetic sources of ApoE in the neural retina, and RPE cells are the most prominent sources in the RPE/choroid.[91] However, it remains unclear whether the concentration of ApoE in the cytoplasm of some RPE cells, especially those in close proximity to drusen, is the result of biosynthesis or selective accumulation. It has been shown that, in both the central and peripheral nervous systems, ApoE expression by astrocytes is up-regulated in response to neuronal injury and neuro-degenerative disease.[84;105;106] Indeed, there is evidence for ApoE up-regulation by Müller cells in degenerating human retina, where increased ApoE immuno-reactivity is found in the sub-retinal space of detached retinas[107] and in the Müller cells of retinas affected by glaucoma or AMD.[108] Furthermore, the relatively high levels of ApoE mRNA detected in the retina, especially in the eyes of older donors and in an individual with

may also contribute to differences in the clearance of debris through BrM.[104]

consistent with this lipid being primarily of RPE origin.[103]

*ApoE* gene status is believed to be a determinant of AMD risk.[88;104;109-111] The *ApoE* gene has three separate alleles: Apo ε2, Apo ε3 and Apo ε4, resulting in six common phenotypes: three homozygous (ε3ε3, ε2ε2, ε4ε4) and three heterozygous (ε2ε3, ε2ε4, ε3ε4) phenotypes. The ε4 allele has been found to be associated with a reduced risk of AMD, whereas the ε2 allele has been associated with an increased risk of developing this disease.[88;104;109-113]

Due to the lack of cysteine residues at positions 112 and 158, preventing the formation of disulphide bridges with ApoA-II or other peptide components, the Apo ε4 allele has an inability to form dimers. It has been suggested that this inability of the Apo ε4 allele to form dimers, when compared with the Apo ε2 and Apo ε3 alleles, favours easier transport of lipids through BrM because of the smaller sized lipid particles, thus protecting against a loss of permeability of BrM.[104]

In the same way, it is possible that the neurosensory retina and the RPE respond to conditions of high oxidative injury by up-regulation of ApoE synthesis and/or accumulation, with implications for selective capture and stabilisation of L and Z in the retina.[91] It has been demonstrated that there is selective binding of certain receptors within the CNS to HDL particles enriched with ApoE, and that there is a lack of binding of these receptors to HDL particles deficient in ApoE.[114] Should this selectivity of the uptake mechanism be dependent on the ApoE polymorphism of the transporting lipoproteins, and given that the Apo ε4 allele is putatively protective for AMD, it is tempting to hypothesise that retinal capture of L and Z may be related to apolipoprotein profile. In other words, the apolipoprotein composition as well as the lipoprotein profile, may play an important role in the transport and delivery of L and Z, and their subsequent accumulation and stabilisation within the retina.[115] Therefore, it is possible that the putative protective effect of the Apo ε4 allele against AMD is attributable, at least in part, to the role its phenotypic expression (ApoE4) plays in the transport and delivery of the macular carotenoids to the retina, and to their stabilisation within the retina. Furthermore, recent research has shown an association between possession of at least one Apo ε4 allele and higher levels of MP across the macula, which is consistent with the view that apolipoprotein profile influences the transport and/or retinal capture of the macular carotenoids.[74]

#### **13. Conclusion**

In conclusion, the role that lipoproteins and apolipoproteins play in the ageing eye and in the aetiopathogenesis of AMD is complex and, as yet, incompletely understood. Lipoproteins and apolipoproteins play an important role in the delivery of potentially protective nutrients from the digestive tract to the eye. The local ocular metabolic activity,

centred on the RPE and BrM, involves an exchange of nutrients from the choroidal circulatory system via BrM to the RPE and retina, with a reverse process whereby waste products are removed from the retina by the RPE through BrM in association with locally produced lipoproteins and apolipoproteins (particularly ApoB and ApoE). Unfortunately, over time it appears that these lipoproteins and apolipoproteins can accumulate between the RPE and BrM, and within BrM, leading to degradation in the metabolic efficiency between these two structures and the choroidal circulation. This deposition has been described as a 'lipid wall' and precedes the development of AMD.[93;94] Methods to detect and arrest or delay this process before it becomes clinically apparent and visually consequential to the patient have yet to be developed. Recent advances in our understanding of the lipoprotein and apolipoprotein molecular biology of the ageing and AMD-affected eye will help to direct future treatment strategies.[100]

#### **Author details**

Edward Loane

*Department of Ophthalmology, Mater Misericordiae University Hospital, Dublin, Ireland* 

#### **14. References**


[12] van der Schaft TL, de Bruijn WC, Mooy CM, de Jong PT. Basal laminar deposit in the aging peripheral human retina. Graefes Arch Clin Exp Ophthalmol 1993 Aug;231(8):470-5.

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**Author details** 

**14. References** 

Edward Loane

centred on the RPE and BrM, involves an exchange of nutrients from the choroidal circulatory system via BrM to the RPE and retina, with a reverse process whereby waste products are removed from the retina by the RPE through BrM in association with locally produced lipoproteins and apolipoproteins (particularly ApoB and ApoE). Unfortunately, over time it appears that these lipoproteins and apolipoproteins can accumulate between the RPE and BrM, and within BrM, leading to degradation in the metabolic efficiency between these two structures and the choroidal circulation. This deposition has been described as a 'lipid wall' and precedes the development of AMD.[93;94] Methods to detect and arrest or delay this process before it becomes clinically apparent and visually consequential to the patient have yet to be developed. Recent advances in our understanding of the lipoprotein and apolipoprotein molecular biology of the ageing and AMD-affected eye will help to

*Department of Ophthalmology, Mater Misericordiae University Hospital, Dublin, Ireland* 

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**Section 5** 

**Lipid Oxidation and Anti-Oxidants** 

380 Lipoproteins – Role in Health and Diseases

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