Figure 5. Lens Ab extraction efficiency in sodium dodecyl sulfate and formic acid. See text for details. doi:10.1371/journal.pone.0010659.g005 fiber cell cytoplasm and upregulated in AD brain [33]. We hypothesized that accumulation of amyloidogenic Ab peptides in the lens fiber cell would potentiate cytosolic lens protein aggregation and increase Rayleigh light scattering. To investigate this hypothesis, we incubated synthetic Ab with human lens protein extract. Addition of Ab to human lens protein extract generated birefringent congophilic aggregates consistent with amyloid (Fig. 7A,B). Double immunogold electron microscopic analysis revealed electron-dense aggregates that co-immunostained for Ab and aB-crystallin (Fig. 7C). These hetero-oligomeric aggregates demonstrated comparable size, morphology, and immunoreactivity relative to the cytosolic protein aggregates identified in postmortem DS lens specimens. Significantly, the dimensions of these Ab- containing protein aggregates (,5pone.0010645 pone.0010646 200 nm in longest axial cross-section) describe classical Rayleigh light scattering centers. Therefore, we sought to determine whether Ab potentiated lens protein aggregation as assessed by quasi-elastic light scattering (QLS) analysis. Addition of synthetic human Ab to human lens protein extract resulted in dose- and time-dependent increases in backscatterd light intensity that was not observed in control lens protein extracts without added human Ab (Fig. 7D). These findings are consistent with Ab-potentiated lens protein aggregation. Discussion In this report, we identify the origin and pathogenic mechanism of cataractogenesis in DS, establish the distinctive lens phenotype Ab in Lens in Down Syndrome in DS as a genetic cataract, and define the molecular correspondence of DS-associated pathology in the lens and brain. Taken together with our previous findings of supranuclear Ab lens pathology in late-onset sporadic AD [28], these results also establish the pathogenic relationship linking Ab pathology in the lens and brain in both DS and AD. AD neuropathology is an invariant feature of DS in subjects over the age of 30 [18,19] but may emerge as early as the first decade [18,34]. A critical factor contributing to the association of DS and AD Ab pathology is the locus of the amyloid precursor protein gene (APP, 21q21) on the long arm of chromosome 21 [16,35,36]. Chromosome 21 triplication leads to increased dosage of the APP gene, accelerated cerebral Ab accumulation, progressive AD neuropathology, and age-dependent cognitive sequelae [18,37,38,39,40]. With the exception of a single reported case of an individual with an atypical DS karyotype involving a specific chromosome 21 micro-deletion in the APP gene locus [41], all subjects with Down syndrome (including the rare cases involving Robertsonian translocation, partial duplications, or trisomic mosaicism) demonstrate triplication of the APP gene locus (21q21), overexpress amyloid precursor protein (APP) in affected somatic cells, accumulate b-amyloid peptides (Ab) in the brain, and reveal evidence of early-onset Alzheimer’s disease (AD) neuropathology [18,19,34,40,42]. In this report, we hypothesize and confirm that this genotype- phenotype concordance extends to Ab-linked molecular pathol- ogy in the supranucleus and deep cortex of the lens. The characteristic cataract phenotype associated with Ab lens pathology has been identified in only two clinical disorders, Alzheimer’s disease [28] and as reported here, Down syndrome. This distinctive supranuclear cataract phenotype has not been reported in normal individuals, and is not observed in other non- AD neurodegenerative diseases, nor in normal aged controls [28]. The classical cerulean ‘‘blue dot’’ cataracts characteristic of DS initially emerge at the equatorial periphery of the lens, and over time, encompass progressively larger areas of the supranucleus and deep cortex (Fig. 8). While lens pathology observed in both infantile and mature DS lenses reveal opaque regions that are clinically described as supranuclear, the ages of the corresponding fiber cells comprising these regions are different (Fig 8). In this respect, it is important to note that the lens pathology in DS lenses does not implicate involvement of lens fibers cells in the fetal nucleus. Distinctive lens pathology is recognized as a characteristic early- onset ocular phenotype in subjects with DS that may be clinically detectable early in life, and in some cases, at birth [20,22,23,24,26,43,44]. By contrast, DS neuropathology and cognitive sequelae typically emerge in the third decade of life and age-dependently progress thereafter [18,19,37,38,39,40]. In the present study, early-onset lens pathology is histochemically documented in illustrative cases of two young adults (21 and 22 years of age) with DS. Lenses obtained from these donors revealed marked Ab amyloid lens pathology that was evident not only in the supranuclear subregion, but also throughout the anterior and equatorial cortex and epithelium. Ab biochemical analysis of DS and normal control lenses indicates that increased Ab expression in the lenses of people with DS may be evident very early in life. In one case included in this study, we detected abnormally high tissue levels of Ab in the lens of a DS subject at 2 years of age. Although visual impairment may not become clinically apparent due to the peripheral distribution of these lenticular lesions relative to the iris, it is certainly the case that Ab molecular pathology in the lens may be amongst the earliest expressed age-dependent phenotypes in Down syndrome. PLoS ONE | www.plosone.org 8 May 2010 | Volume 5 | Issue 5 | e10659