Association Between Age-related Macular Degeneration And Apolipoprotein E

Senile macular degeneration (SMD) is also called Age-relatedmacular degeneration, AMD. Age related macular degeneration (AMD)is the leading cause of irreversible vision loss in the elderly populationin Western Europe and USA. In our country, with the increasing oldpopulation and the coming of senile society, its morbidity increasedyear after year. However, The etiology of AMD remains unknown andthe effects of therapy are not ideal. According to the epidemiologicalstudies, the risk factors for AMD include age , positive family historyof AMD, smoking , race, low level of antioxidants such as vitaminsand zinc in diet and plasma has been demonstrated to be related toAMD. Other possible risk factor include female gender, lightening ofiris colour, underlying cardiovascular disease, light exposure etc.among this risk factor, age and genetic factor plays crucial roles inprogression of AMD. Recently, several families and MonozygoticTwins studies have convincingly demonstrated the importance ofgenetics in AMD. AMD may be a autosomal dominant disease, justlike glaucoma, the AMD inheritance in their progenies not simplyaccording to Mendel law, it more likely polygenic inherited disease.However, the pathogenesis of genetic factors involved AMD need tofurther study.Apolipoprotein E (APOE), the major apolipoprotein of the CNSand an important regulator of cholesterol and lipid transport, appears tobe associated with neurodegeneration. Some research revealed that theexpression of APOE was increased in the extracellular accumulationsof retinal pigment epithelium (RPE) and Bruch's membrane, suggestthe function disorder of APOE was involved in the pathogenesis ofAMD. Some studies reported that the APOE polymorphism showed asignificant association with the risk for AMD in USA and Europe.Since the APOE polymorphism has race specificity, we study theAssociation between Age-related Macular Degeneration andApolipoprotein E in Chinese. Further study the genetic mechanism ofAMD will promote the gene therapy for AMD.Part1 Drusen proteome analysis for Age-relatedMacular DegenerationObjectiveDevelop an approach to the etiology of age-related maculardegeneration by using Drusen proteome analysis.Mathods4 eyes from 2 corpses were thawed overnight at 4°C, the anteriorsegment was removed with a circumferential cut behind the limbus,the vitreous was poured from the posterior globe, the optic nerve wascut, and the retina was removed. Drusen appear as bump-likeelevations of the RPE when viewed at magnifications of 20–40diameters and the macular region of the RPE is identifiable by itsheavier pigmentation. The RPE was removed from the interior surfaceof the globe under a dissecting microscope with repetitive brushingand aspiration in the presence of PBS. At magnifications between 25and 40 diameters, drusen were removed with fine forceps. Many of thedrusen isolated were sorted into separate groupings, and transferred tostorage tubes with a capillary pipette for further analysis. Isolateddrusen samples were identified by LC MS/MS analyses.ResultsWe have identified 40 proteins by LC MS/MS analyses ofindividual tryptic digests of isolated drusen preparations, among theseproteins, 20 were unknown proteins. Averaged 10 proteins per drusenpreparation were identified, and the most common proteins were foundin drusen preparations. Tissue metalloproteinase inhibitor 3 (TIMP3),clusterin, vitronectin and serum albumin appear to be common drusenproteins, detectable in up to 60% of drusen preparations. Crystallinswere detected more frequently in AMD donor drusen;however,analysis of additional AMD donors is required to establish thesignificance of this observation.ConclusionsThe following general conclusions regarding the isolationprocedure can be made:1.Most drusen remain on Bruch's membrane after brushing toremove the RPE.2.The abundance of drusen varies per donor and accordingly, theamount of protein recovered is variable and in the low microgramrange.3.Not every drusen preparation has yielded protein identificationsby LC MS/MS. Our observations during drusen isolation are consistentwith previous funduscopic reports indicating(1) That most elderly eyes have drusen, particularly in theperiphery;(2) When a large population of drusen are found in one eye, theother eye also contains abundant drusen;(3) Multiple drusen types (hard and soft) can be present in a singleeye.(4) AMD is a multifactorial and polygenic factors involveddisease. 4. This approach can be a useful method for etiologyresearch and diagnosis of AMD.Part2 Association between Age-related MacularDegeneration and Apolipoprotein EObjectiveTo understand the physiological and pathological characters ofAPOE and it's Association with Age-related Macular Degeneration.MethodsGenomic DNA was isolated from 2ml venous blood leukocytesusing a standard phenol-chloroform extraction procedure. Polymerasechain reaction (PCR) amplification was undertaken with the primers5'-AGGGCGCTGATGGACGAGAC-3'and 5'-AGGCGCTCGCGGATGGCGC-3' to produce a 360 bp product covered amino acid position112 and 158. Aliquots of amplicons were confirmed by agarose gelelectrophoresis, and then the product was ligated into a T vectorbefore sequencing. Sequence analysis was performed respectivelytwice to confirm mismatch.ResultsOur results suggest that the APOE polymorphism has somerelationship with AMD. The APOE e4 allele was associated with adecreased risk and the e2 allele was associated with a slightlyincreased risk of AMD.ConclusionsOur findings support an association of APOE with AMD.Specifically, they implicate the APOE2 genotype as a susceptibilitygene that confers an increased risk for AMD. Further investigate thegenetic mechanism of AMD will benefit for the prevention and therapyof AMD. Gene therapy will be the promising approach for AMD. Butthere is a lot of things need us to further investigate for gene therapy.