Integrative genome-wide analysis reveals EIF3A as a key downstream regulator of translational repressor protein Musashi 2 (MSI2).

Musashi 2 (MSI2) is an RNA binding protein (RBP) that regulates asymmetric cell division and cell fate decisions in normal and cancer stem cells. MSI2 appears to repress translation by binding to 3' untranslated regions (3'UTRs) of mRNA, but the identity of functional targets remains unknown. Here, we used individual nucleotide resolution cross-linking and immunoprecipitation (iCLIP) to identify direct RNA binding partners of MSI2 and integrated these data with polysome profiling to obtain insights into MSI2 function. iCLIP revealed specific MSI2 binding to thousands of mRNAs largely in 3'UTRs, but translational differences were restricted to a small fraction of these transcripts, indicating that MSI2 regulation is not triggered by simple binding. Instead, the functional targets identified here were bound at higher density and contain more 'UAG' motifs compared to targets bound nonproductively. To further distinguish direct and indirect targets, MSI2 was acutely depleted. Surprisingly, only 50 transcripts were found to undergo translational induction on acute loss. Using complementary approaches, we determined eukaryotic translation initiation factor 3A (EIF3A) to be an immediate, direct target. We propose that MSI2 downregulation of EIF3A amplifies these effects on translation. Our results also underscore the challenges in defining functional targets of RBPs since mere binding does not imply a discernible functional interaction.

Random walker's view of networks whose growth it shapes.

We study a simple model in which the growth of a network is determined by the location of one or more random walkers. Depending on walker motility rate, the model generates a spectrum of structures situated between well-known limiting cases. We demonstrate that the average degree observed by a walker is a function of its motility rate. Modulating the extent to which the location of node attachment is determined by the walker as opposed to random selection is akin to scaling the speed of the walker and generates new limiting behavior. The model raises questions about energetic and computational resource requirements in a physical instantiation.

Using approximate Bayesian computation to quantify cell-cell adhesion parameters in a cell migratory process.

In this work, we implement approximate Bayesian computational methods to improve the design of a wound-healing assay used to quantify cell-cell interactions. This is important as cell-cell interactions, such as adhesion and repulsion, have been shown to play a role in cell migration. Initially, we demonstrate with a model of an unrealistic experiment that we are able to identify model parameters that describe agent motility and adhesion, given we choose appropriate summary statistics for our model data. Following this, we replace our model of an unrealistic experiment with a model representative of a practically realisable experiment. We demonstrate that, given the current (and commonly used) experimental set-up, our model parameters cannot be accurately identified using approximate Bayesian computation methods. We compare new experimental designs through simulation, and show more accurate identification of model parameters is possible by expanding the size of the domain upon which the experiment is performed, as opposed to increasing the number of experimental replicates. The results presented in this work, therefore, describe time and cost-saving alterations for a commonly performed experiment for identifying cell motility parameters. Moreover, this work will be of interest to those concerned with performing experiments that allow for the accurate identification of parameters governing cell migratory processes, especially cell migratory processes in which cell-cell adhesion or repulsion are known to play a significant role.

Variable species densities are induced by volume exclusion interactions upon domain growth.

In this work we study the effect of domain growth on spatial correlations in agent populations containing multiple species. This is important as heterogenous cell populations are ubiquitous during the embryonic development of many species. We have previously shown that the long-term behavior of an agent population depends on the way in which domain growth is implemented. We extend this work to show that, depending on the way in which domain growth is implemented, different species dominate in multispecies simulations. Continuum approximations of the lattice-based model that ignore spatial correlations cannot capture this behavior, while those that explicitly account for spatial correlations can. The results presented here show that the precise mechanism of domain growth can determine the long-term behavior of multispecies populations and, in certain circumstances, establish spatially varying species densities.

How domain growth is implemented determines the long-term behavior of a cell population through its effect on spatial correlations.

Domain growth plays an important role in many biological systems, and so the inclusion of domain growth in models of these biological systems is important to understanding how these systems function. In this work we present methods to include the effects of domain growth on the evolution of spatial correlations in a continuum approximation of a lattice-based model of cell motility and proliferation. We show that, depending on the way in which domain growth is implemented, different steady-state densities are predicted for an agent population. Furthermore, we demonstrate that the way in which domain growth is implemented can result in the evolution of the agent density depending on the size of the domain. Continuum approximations that ignore spatial correlations cannot capture these behaviors, while those that account for spatial correlations do. These results will be of interest to researchers in developmental biology, as they suggest that the nature of domain growth can determine the characteristics of cell populations.

Reconciling diverse mammalian pigmentation patterns with a fundamental mathematical model.

Bands of colour extending laterally from the dorsal to ventral trunk are a common feature of mouse chimeras. These stripes were originally taken as evidence of the directed dorsoventral migration of melanoblasts (the embryonic precursors of melanocytes) as they colonize the developing skin. Depigmented 'belly spots' in mice with mutations in the receptor tyrosine kinase Kit are thought to represent a failure of this colonization, either due to impaired migration or proliferation. Tracing of single melanoblast clones, however, has revealed a diffuse distribution with high levels of axial mixing--hard to reconcile with directed migration. Here we construct an agent-based stochastic model calibrated by experimental measurements to investigate the formation of diffuse clones, chimeric stripes and belly spots. Our observations indicate that melanoblast colonization likely proceeds through a process of undirected migration, proliferation and tissue expansion, and that reduced proliferation is the cause of the belly spots in Kit mutants.

Inference of cell-cell interactions from population density characteristics and cell trajectories on static and growing domains.

A key feature of cell migration is how cell movement is affected by cell-cell interactions. Furthermore, many cell migratory processes such as neural crest stem cell migration [Thomas and Erickson, 2008; McLennan et al., 2012] occur on growing domains or in the presence of a chemoattractant. Therefore, it is important to study interactions between migrating cells in the context of domain growth and directed motility. Here we compare discrete and continuum models describing the spatial and temporal evolution of a cell population for different types of cell-cell interactions on static and growing domains. We suggest that cell-cell interactions can be inferred from population density characteristics in the presence of motility bias, and these population density characteristics for different cell-cell interactions are conserved on both static and growing domains. We also study the expected displacement of a tagged cell, and show that different types of cell-cell interactions can give rise to cell trajectories with different characteristics. These characteristics are conserved in the presence of domain growth, however, they are diminished in the presence of motility bias. Our results are relevant for researchers who study the existence and role of cell-cell interactions in biological systems, so far as we suggest that different types of cell-cell interactions could be identified from cell density and trajectory data.

PIWI proteins and PIWI-interacting RNAs in the soma.

The discovery of millions of PIWI-interacting RNAs revealed a fascinating and unanticipated dimension of biology. The PIWI-piRNA pathway has been commonly perceived as germline-specific, even though the somatic function of PIWI proteins was documented when they were first discovered. Recent studies have begun to re-explore this pathway in somatic cells in diverse organisms, particularly lower eukaryotes. These studies have illustrated the multifaceted somatic functions of the pathway not only in transposon silencing but also in genome rearrangement and epigenetic programming, with biological roles in stem-cell function, whole-body regeneration, memory and possibly cancer.

The effect of occlusion on the semantics of projective spatial terms: a case study in grounding language in perception.

Although data-driven spatial template models provide a practical and cognitively motivated mechanism for characterizing spatial term meaning, the influence of perceptual rather than solely geometric and functional properties has yet to be systematically investigated. In the light of this, in this paper, we investigate the effects of the perceptual phenomenon of object occlusion on the semantics of projective terms. We did this by conducting a study to test whether object occlusion had a noticeable effect on the acceptance values assigned to projective terms with respect to a 2.5-dimensional visual stimulus. Based on the data collected, a regression model was constructed and presented. Subsequent analysis showed that the regression model that included the occlusion factor outperformed an adaptation of Regier & Carlson's well-regarded AVS model for that same spatial configuration.

A high omega-3 fatty acid diet reduces retinal lesions in a murine model of macular degeneration.

Age-related macular degeneration (AMD) is one of the leading cause of blindness among the elderly; however, current therapy options are limited. Epidemiological studies have shown that a diet that is high in omega-3 polyunsaturated (n-3) fatty acids can slow disease progression in patients with advanced AMD. In this study, we evaluated the effect of such a diet on the retinas of Ccl2(-/-)/Cx3cr1(-/-) mice, a model that develops AMD-like retinal lesions that include focal deep retinal lesions, abnormal retinal pigment epithelium, photoreceptor degeneration, and A2E accumulation. Ccl2(-/-)/Cx3cr1(-/-) mice that ingested a high n-3 fatty acid diet showed a slower progression of retinal lesions compared with the low n-3 fatty acids group. Some mice that were given high levels of n-3 fatty acids had lesion reversion. We found a shunted arachidonic acid metabolism that resulted in decreased pro-inflammatory derivatives (prostaglandin E(2) and leukotriene B(4)) and an increased anti-inflammatory derivative (prostaglandin D(2)). We also measured lower ocular TNF-alpha and IL-6 transcript levels in the mice fed a diet of high n-3 fatty acids. Our findings in these mice are in line with human studies of AMD risk reduction by long-chain n-3 fatty acids. This murine model provides a useful tool to evaluate therapies that might delay the development of AMD.

Peroxisome Proliferator-Activated Receptor Expression in Murine Models and Humans with Age-related Macular Degeneration.

Peroxisome proliferator-activated receptors (PPARs) play a role in oxidative stress and VEGF regulation, which are closely related to age-related macular degeneration (AMD). PPAR γ expression and its downstream molecules were examined in fat-1 mice (transgenic mice that convert n-6 to n-3 fatty acids), Ccl2(-/-)/Cx3cr1(-/-) mice (an AMD model), ARPE19 cells (a human retinal pigment epithelial cell line, RPE, a cell type with a critical role in AMD), and human eyes with and without AMD. PPAR α, ß, and γ, VEGF and receptors were determined by immunohistochemistry in the mice models, humans, and ARPE19 cells. Transcripts of PPARs, VEGF, MMP-9 and HO-1 were determined by RQ-PCR. PPARs were constitutively expressed in normal neuroretina and RPE of humans and mice. PPAR γ expression was increased in fat-1 and Ccl2(-/-)/Cx3cr1(-/-) mice. VEGF was decreased in fat-1 mice but increased in Ccl2(-/-)/Cx3cr1(-/-) mice. VEGF receptors were stable. VEGF, MMP9 and HO-1 transcript levels were increased in ARPE19 cells under H(2)O(2) - induced oxidative stress. Human AMD retinas exhibited higher PPAR γ. The findings of increased expression of PPAR γ and its downstream proteins (VEGF, MMP9, and HO-1) in H(2)O(2)-treated ARPE19 cells, Ccl2(-/-)/Cx3cr1(-/-) mice, and human AMD eyes, but decreased VEGF in fat-1 mice, suggest that PPAR γ may play a role in AMD.

The HtrA1 promoter polymorphism, smoking, and age-related macular degeneration in multiple case-control samples.

OBJECTIVE: To assess the association and combined effect on the risk of age-related macular degeneration (AMD) by the HtrA1 and complement factor H (CFH) polymorphisms, smoking, and serum cholesterol. DESIGN: Clinic-based and population-based case control study. PARTICIPANTS: A total of 805 AMD cases and 921 controls from The Eye Clinic of National Eye Institute, Age-Related Eye Diseases Study, Blue Mountain Eye Study Cohort, and Minnesota Lions Eye Bank. METHODS: DNA samples were genotyped for polymorphisms of rs11200638 in HtrA1 promoter and rs380390 in CFH. HtrA1 protein in ocular tissue was measured. Interactions of the HtrA1 risk allele with the CFH risk variant, smoking status, and cholesterol were assessed. MAIN OUTCOME MEASURES: AMD was evaluated by retinal specialists, and AMD subtypes (geographic atrophy and neovascularization) were determined. RESULTS: Strong associations of the HtrA1 risk allele (A) with AMD were present in all sample sets. A similar magnitude of association was observed for central geographic atrophy and neovascular AMD. The combination of the HtrA1 and CFH risk alleles increased AMD susceptibility, as did the combination of the HtrA1 risk allele with smoking. No combined effect of HtrA1 risk allele and cholesterol level was found. Enhanced expression of HtrA1 protein was detected in retina with AMD. CONCLUSIONS: Findings from multiple samples support an AMD genetic variant harbored within HtrA1. The risk of advanced AMD increased when the presence of risk alleles from HtrA1 was combined with either CFH risk alleles or history of smoking.

Ccl2/Cx3cr1-deficient mice: an animal model for age-related macular degeneration.

BACKGROUND/AIMS: Senescent Ccl2-/- mice develop cardinal features of human age-related macular degeneration (AMD). Loss-of-function single-nucleotide polymorphisms within CX3CR1 are associated with AMD. METHODS: We generated Ccl2-/-/Cx3cr1-/- [double-knockout (DKO)] mice and evaluated the eyes using fundoscopy routine histology, immunochemistry, biochemistry and proteomics. RESULTS: At 6 weeks old, all DKO mice developed AMD-like retinal lesions such as abnormal retinal pigment epithelium cells, drusen, photoreceptor atrophy and choroidal neovascularization, which progressed with age and reversed with high omega-3 long-chain polyunsaturated fatty acid diet. N-retinylidene-N-retinylethanolamine (A2E), a major lipofuscin fluorophore, illustrated by an emission peak at approximately 600 nm, was significantly higher in DKO retinal pigment epithelium. Decreased ERp29 was found in the retina of DKO mice. CONCLUSION: A broad spectrum of AMD pathologies with early onset and high penetrance in these mice implicate certain chemokines, A2E and endoplasmic reticulum proteins in AMD pathogenesis.

Immunological protein expression profile in Ccl2/Cx3cr1 deficient mice with lesions similar to age-related macular degeneration.

Age-related macular degeneration (AMD) is the leading cause of blindness in the United States. Ccl2 knock-out (KO) mice sporadically develop the cardinal features of AMD in their senescent stage. Humans bearing a loss of function variant or single nucleotide polymorphism (SNP) in CX3CR1 are at increased risk of developing AMD. We recently developed Ccl2(-/-)/Cx3cr1(-/-) mice, which consistently develop retinal degeneration with many AMD features. Since there is strong evidence for an immunological role in AMD pathogenesis, we examined ocular immune protein expression levels in Ccl2(-/-)/Cx3cr1(-/-), Ccl2(-/-), Cx3cr1(-/-), and age-matched wild-type (WT) mice. Immunohistochemistry revealed increased complement C3d in Bruch's membrane, retinal pigment epithelium (RPE), choroidal capillaries, and particularly drusen of the Ccl2(-/-)/Cx3cr1(-/-) mice relative to the WT controls. No change was detected in single KO mice. Real-time RT-PCR revealed a 2.5-fold increase in C3 expression in the Ccl2(-/-)/Cx3cr1(-/-). While the retinas of four month old WT and Ccl2(-/-) showed minimal immunoreactivity for markers of macrophages and microglia, infiltrates of these mononuclear phagocytic cells were detected in the Ccl2(-/-)/Cx3cr1(-/-)retinal lesions and a few foci in the Cx3cr1(-/-) retina. The Ccl2(-/-)/Cx3cr1(-/-) had reduced toll-like receptor 4 (TLR4) expression in the RPE. Following LPS injection, the Ccl2(-/-)/Cx3cr1(-/-) had significantly reduced endotoxin-induced uveitis scores and showed a diminished increase in Tlr4 mRNA expression. No changes in TLR4 expression were detected in either single KO. Autoantibodies against the retina and photoreceptors were also detected in the Ccl2(-/-)/Cx3cr1(-/-) serum. Real-time RT-PCR revealed significant increases in Ccl5 transcript in the Ccl2(-/-)/Cx3cr1(-/-) relative to the WT. These results suggest that innate immunity and possibly adaptive immunity play an important role in Ccl2(-/-)/Cx3cr1(-/-) retinal degeneration. Moreover, since human AMD patients show similar immunopathological profiles, these results support the Ccl2(-/-)/Cx3cr1(-/-) as a suitable model for human AMD.

The LOC387715 polymorphism, inflammatory markers, smoking, and age-related macular degeneration. A population-based case-control study.

OBJECTIVE: To assess combined effects on the risk of age-related macular degeneration (AMD) by the LOC387715 polymorphism, smoking, and inflammatory or hemostatic factors. DESIGN: Population-based case-control study. PARTICIPANTS: Two hundred seventy-eight AMD cases (224 early, 54 late) and 557 controls matched for age, gender, and smoking, drawn from the Blue Mountains Eye Study cohort. METHODS: Subjects were genotyped for the LOC387715 Ala69Ser polymorphism (rs# 10490924). Smoking was self-reported. Serum high-sensitivity C-reactive protein (CRP), interleukin 6 (IL-6), soluble intercellular adhesion molecule 1 (sICAM-1), fibrinogen, homocysteine, plasminogen activator inhibitor 1 (PAI-1), von Willebrand factor, and white cell count (WCC) were measured. Combined effects of this genetic variant plus any of these study factors on AMD risk were assessed using logistic regression models, adjusted for age and smoking. We defined interaction if the influence of 2 factors departed from the multiplicative scale, confirmed by a statistically significant interaction term. Otherwise, the combined effect was used. MAIN OUTCOME MEASURES: Age-related macular degeneration was graded using the Wisconsin grading system. RESULTS: Combined effects on the likelihood of early or late AMD were demonstrated for the LOC387715 Ala69Ser G/T and T/T genotypes with the markers high-sensitivity CRP (odds ratios [ORs], 1.2 for the highest tertile alone, 1.6 for G/T and T/T genotypes alone, and 2.2 for both G/T and T/T genotypes plus the highest tertile, compared with the G/G genotype with the 2 lower tertiles), IL-6 (corresponding ORs, 1.1, 1.6, and 2.2), sICAM-1 (ORs, 1.0, 1.5, and 2.3, respectively), and PAI-1 (ORs, 1.3, 1.7, and 2.3, respectively), but not with WCC, fibrinogen, homocysteine, and von Willebrand factor. Findings were similar for early and late AMD separately. Current smokers with G/T and T/T genotypes had strong combined effects on late AMD risk compared with those who never smoked or past smokers with the G/G genotype (ORs, 1.2 for current smokers alone, 1.8 for G/T and T/T genotypes alone, and 6.1 for current smokers plus G/T and T/T genotypes). CONCLUSIONS: We found no significant interaction but combined effects for the LOC387715 genotypes with 3 inflammatory markers and PAI-1 on the risk of early or late AMD, and with current smoking on the risk of late AMD.

Genetic markers and biomarkers for age-related macular degeneration.

Age-related macular degeneration (AMD) is the leading cause of visual impairment and blindness in the USA. Although the treatment of AMD has evolved to include laser photocoagulation, photodynamic therapy, surgical macular translocation and antiangiogenesis agents, treatment options for advanced AMD are limited. Furthermore, the dry form of AMD, albeit less devastating than the wet form, has even fewer viable treatment options. This review summarizes the various biomarkers of AMD and analyzes whether or not they may one day be exploited to determine risks of disease onset, measure progression of disease or even assess the effects of treatment of AMD. Potential biomarkers are important to identify since some might be utilized to reflect the disease state of a particular patient and to individualize therapy. Although studies have yielded promising results for nutrient and inflammatory biomarkers, these results have been inconsistent. At present, the best available markers of AMD risk are single nucleotide polymorphisms (SNPs). SNPs in complement factor H (CFH) and PLEKHA1/ARMS2/HtrA1 capture a substantial fraction of AMD risk and permit the identification of individuals at high risk of developing AMD.

Murine ccl2/cx3cr1 deficiency results in retinal lesions mimicking human age-related macular degeneration.

PURPOSE: Senescent Ccl2(-/-) mice are reported to develop cardinal features of human age-related macular degeneration (AMD). Loss-of-function single-nucleotide polymorphisms within CX3CR1 are also found to be associated with AMD. The authors generated Ccl2(-/-)/Cx3cr1(-/-) mice to establish a more characteristic and reproducible AMD model. METHODS: Single Ccl2- and Cx3cr1-deficient mice were crossbred to obtain Ccl2(-/-)/Cx3cr1(-/-) mice. Funduscopy, histopathology, retinal A2E quantification, proteomics, RT-PCR gene expression assay, immunochemistry, and Western blotting were used to examine the retina and to evaluate gene expression within the retinal tissue. RESULTS: By 6 weeks of age, all Ccl2(-/-)/Cx3cr1(-/-) mice developed AMD-like retinal lesions, including drusen, retinal pigment epithelium alteration, and photoreceptor degeneration. Furthermore, choroidal neovascularization occurred in 15% of the mice. These degenerative lesions progressed with age. A2E, a major lipofuscin fluorophore that accumulated during AMD progression, was significantly higher in the Ccl2(-/-)/Cx3cr1(-/-) retina than in the wild-type retina. Complement cofactor was higher in the Ccl2(-/-)/Cx3cr1(-/-) RPE. Proteomics data indicated that four proteins were differentially expressed in Ccl2(-/-)/Cx3cr1(-/-) retina compared with control. One of these proteins, ERp29, an endoplasmic reticulum protein, functions as an escort chaperone and in protein folding. CONCLUSIONS: The authors concluded that Ccl2(-/-)/Cx3cr1(-/-) mice develop a broad spectrum of AMD abnormalities with early onset and high penetrance. These observations implicate certain chemokines and endoplasmic reticulum proteins in AMD pathogenesis. Similar to the mechanism of neurodegeneration caused by dysfunction of endoplasmic reticulum proteins, decreased chaperoning may cause misfolded protein accumulation, leading to drusen formation and retinal degeneration.

The LOC387715 polymorphism and age-related macular degeneration: replication in three case-control samples.

PURPOSE: Age-related macular degeneration (AMD) is a multifactorial blinding disease in the elderly. LOC387715 harbors a single-nucleotide polymorphism that has an association with AMD. This study was conducted to confirm the association between LOC387715 and AMD and to refine estimates of the impact of this gene variation in using samples from three studies: an Australian population-based study and two U.S. clinic-based case-control studies. METHODS: Cases and controls were collected from a National Eye Institute (NEI) clinical protocol (n = 240), the Age-Related Eye Disease Study (AREDS; n = 488), and the Blue Mountains Eye Study (BMES; n = 851). After DNA extraction, subjects were genotyped for the LOC387715 Ala69Ser polymorphism (rs10490924). RESULTS: The combined NEI and AREDS samples yielded odds ratios (ORs) of 2.61 (95% CI 1.89-3.61, P = 1.42 x 10(-9)) and 8.59 (95% CI 4.49-16.5, P = 3.56 x 10(-13)) for the heterozygous and homozygous risk alleles, respectively. The corresponding odds ratios in the BMES sample were 1.69 (95% CI: 1.25-2.28, P = 0.0007) and 2.20 (95% CI: 1.05-4.62, P = 0.038) for the heterozygous and homozygous groups. Neither set of samples showed statistically significant interaction with smoking, although there appeared to be a trend of interaction between smoking and LOC387715 for risk of advanced AMD. CONCLUSIONS: Although these data from three case-control samples support an AMD genetic risk marker harbored within LOC387715, the nested case-control data from the population-based BMES samples showed lower estimates than from the clinic-based samples. This may be because the BMES samples consisted of largely early AMD cases while the clinic-based AMD samples consisted exclusively of advanced cases.

Human HtrA1 in the archived eyes with age-related macular degeneration.

PURPOSE: HtrA1 belongs to the high temperature requirement factor A family of serine proteases, which are involved in protein quality control and cell fate. A single-nucleotide polymorphism (SNP), rs11200638, in the promoter of HtrA1 at chromosome 10q26 is reported as a likely causal variant for age-related macular degeneration (AMD). The SNP is located in the regulatory region and increases production of HtrA1 protein. This study investigates HtrA1 expression and SNP genotypes in archived ocular slides with AMD. METHODS: Macular, nonretinal, and peripheral retinal cells were microdissected from archived slides from 57 eyes with AMD and 16 age-matched, non-AMD controls. HtrA1 rs11200638 SNP genotyping was performed using polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis. HtrA1 transcripts were measured using real-time reverse transcriptase-PCR. HtrA1 protein expression was evaluated using avidin-biotin complex immunohistochemistry. RESULTS: HtrA1 (G/A) SNP was successfully genotyped in 52 AMD cases and 13 non-AMD subjects. The frequencies of the risk allele (A) were 55 of 104 (52.9%) and 8 of 26 (30.8%) in AMD and control groups, respectively. HtrA1 mRNA was detected in normal peripheral and macular retinas, higher in the periphery than maculae. HtrA1 mRNA was much higher in the macula and a lot lower in the periphery of the AMD eyes as compared to control eyes. HtrA1 protein was expressed in normal retinal vascular endothelia and retinal pigment epithelia. Intense immunoreaction against HtrA1 was found in AMD lesions, slightly more in wet than dry AMD lesions. CONCLUSION: This study successfully analyzes HtrA1 SNP and transcript expression in microdissected cells from archived paraffin fixed slides. Up-regulation of HtrA1 is detected in the macular lesions of AMD eyes. The data further suggest that rs11200638 in HtrA1 promoter is associated with AMD development.