Systems-level analysis of age-related macular degeneration reveals global biomarkers and phenotype-specific functional networks.

UNLABELLED: Please see related commentary: http://www.biomedcentral.com/1741-7015/10/21/abstract BACKGROUND: Age-related macular degeneration (AMD) is a leading cause of blindness that affects the central region of the retinal pigmented epithelium (RPE), choroid, and neural retina. Initially characterized by an accumulation of sub-RPE deposits, AMD leads to progressive retinal degeneration, and in advanced cases, irreversible vision loss. Although genetic analysis, animal models, and cell culture systems have yielded important insights into AMD, the molecular pathways underlying AMD's onset and progression remain poorly delineated. We sought to better understand the molecular underpinnings of this devastating disease by performing the first comparative transcriptome analysis of AMD and normal human donor eyes. METHODS: RPE-choroid and retina tissue samples were obtained from a common cohort of 31 normal, 26 AMD, and 11 potential pre-AMD human donor eyes. Transcriptome profiles were generated for macular and extramacular regions, and statistical and bioinformatic methods were employed to identify disease-associated gene signatures and functionally enriched protein association networks. Selected genes of high significance were validated using an independent donor cohort. RESULTS: We identified over 50 annotated genes enriched in cell-mediated immune responses that are globally over-expressed in RPE-choroid AMD phenotypes. Using a machine learning model and a second donor cohort, we show that the top 20 global genes are predictive of AMD clinical diagnosis. We also discovered functionally enriched gene sets in the RPE-choroid that delineate the advanced AMD phenotypes, neovascular AMD and geographic atrophy. Moreover, we identified a graded increase of transcript levels in the retina related to wound response, complement cascade, and neurogenesis that strongly correlates with decreased levels of phototransduction transcripts and increased AMD severity. Based on our findings, we assembled protein-protein interactomes that highlight functional networks likely to be involved in AMD pathogenesis. CONCLUSIONS: We discovered new global biomarkers and gene expression signatures of AMD. These results are consistent with a model whereby cell-based inflammatory responses represent a central feature of AMD etiology, and depending on genetics, environment, or stochastic factors, may give rise to the advanced AMD phenotypes characterized by angiogenesis and/or cell death. Genes regulating these immunological activities, along with numerous other genes identified here, represent promising new targets for AMD-directed therapeutics and diagnostics.

The pivotal role of the complement system in aging and age-related macular degeneration: hypothesis re-visited.

During the past ten years, dramatic advances have been made in unraveling the biological bases of age-related macular degeneration (AMD), the most common cause of irreversible blindness in western populations. In that timeframe, two distinct lines of evidence emerged which implicated chronic local inflammation and activation of the complement cascade in AMD pathogenesis. First, a number of complement system proteins, complement activators, and complement regulatory proteins were identified as molecular constituents of drusen, the hallmark extracellular deposits associated with early AMD. Subsequently, genetic studies revealed highly significant statistical associations between AMD and variants of several complement pathway-associated genes including: Complement factor H (CFH), complement factor H-related 1 and 3 (CFHR1 and CFHR3), complement factor B (CFB), complement component 2 (C2), and complement component 3 (C3). In this article, we revisit our original hypothesis that chronic local inflammatory and immune-mediated events at the level of Bruch's membrane play critical roles in drusen biogenesis and, by extension, in the pathobiology of AMD. Secondly, we report the results of a new screening for additional AMD-associated polymorphisms in a battery of 63 complement-related genes. Third, we identify and characterize the local complement system in the RPE-choroid complex - thus adding a new dimension of biological complexity to the role of the complement system in ocular aging and AMD. Finally, we evaluate the most salient, recent evidence that bears directly on the role of complement in AMD pathogenesis and progression. Collectively, these recent findings strongly re-affirm the importance of the complement system in AMD. They lay the groundwork for further studies that may lead to the identification of a transcriptional disease signature of AMD, and hasten the development of new therapeutic approaches that will restore the complement-modulating activity that appears to be compromised in genetically susceptible individuals.

Microduplication and triplication of 22q11.2: a highly variable syndrome.

22q11.2 microduplications of a 3-Mb region surrounded by low-copy repeats should be, theoretically, as frequent as the deletions of this region; however, few microduplications have been reported. We show that the phenotype of these patients with microduplications is extremely diverse, ranging from normal to behavioral abnormalities to multiple defects, only some of which are reminiscent of the 22q11.2 deletion syndrome. This diversity will make ascertainment difficult and will necessitate a rapid-screening method. We demonstrate the utility of four different screening methods. Although all the screening techniques give unique information, the efficiency of real-time polymerase chain reaction allowed the discovery of two 22q11.2 microduplications in a series of 275 females who tested negative for fragile X syndrome, thus widening the phenotypic diversity. Ascertainment of the fragile X-negative cohort was twice that of the cohort screened for the 22q11.2 deletion. We also report the first patient with a 22q11.2 triplication and show that this patient's mother carries a 22q11.2 microduplication. We strongly recommend that other family members of patients with 22q11.2 microduplications also be tested, since we found several phenotypically normal parents who were carriers of the chromosomal abnormality.

White-cap mutants and meiotic apoptosis in the basidiomycete Coprinus cinereus.

Among many white-cap mutants of Coprinus cinereus, four distinct classes have been identified cytologically. Mutants of one class progress through meiosis normally but fail to sporulate; the defect is post-meiotic and it triggers apoptosis in the tetrad stage. Mutants of the other three classes have defects in meiotic prophase and these are: (1) those that assemble synaptonemal complexes (SCs) normally; (2) those that assemble axial elements (AEs) but not SCs; and (3) those that assemble neither AEs nor SCs even though the chromosomes are condensed and also paired. All three meiotic mutant classes arrest at meiotic metaphase I and the arrest triggers meiosis-specific apoptosis showing characteristic chromatin condensation, DNA fragmentation as shown by the TUNEL assay, cytoplasmic shrinkage, and finally total DNA degradation. Apoptosis is very cell-type specific; it occurs only in the basidia while the neighboring somatic cells are perfectly healthy and the mushroom continues to develop and mature with very few basidiospores produced. The meiotic apoptosis in C. cinereus is under strict cell cycle control rather than at any time after defect; apoptosis is triggered only after entry to meiotic metaphase. It is intriguing to note that C. cinereus has two checkpoints for arrest and entry to apoptosis: one is meiotic at the metaphase I spindle checkpoint regardless of the time of defects, and one is post-meiotic at the tetrad stage. This is in striking contrast to multiple checkpoint arrests and entries to meiotic apoptosis found in the mouse.