Replenishing IRAK-M expression in retinal pigment epithelium attenuates outer retinal degeneration.

Chronic inflammation is a constitutive component of many age-related diseases, including age-related macular degeneration (AMD). Here, we identified interleukin-1 receptor-associated kinase M (IRAK-M) as a key immunoregulator in retinal pigment epithelium (RPE) that declines during the aging process. Rare genetic variants of IRAK3, which encodes IRAK-M, were associated with an increased likelihood of developing AMD. In human samples and mouse models, IRAK-M abundance in the RPE declined with advancing age or exposure to oxidative stress and was further reduced in AMD. Irak3-knockout mice exhibited an increased incidence of outer retinal degeneration at earlier ages, which was further exacerbated by oxidative stressors. The absence of IRAK-M led to a disruption in RPE cell homeostasis, characterized by compromised mitochondrial function, cellular senescence, and aberrant cytokine production. IRAK-M overexpression protected RPE cells against oxidative or immune stressors. Subretinal delivery of adeno-associated virus (AAV)-expressing human IRAK3 rescued light-induced outer retinal degeneration in wild-type mice and attenuated age-related spontaneous retinal degeneration in Irak3-knockout mice. Our data show that replenishment of IRAK-M in the RPE may redress dysregulated pro-inflammatory processes in AMD, suggesting a potential treatment for retinal degeneration.

Levels of the HtrA1 Protein in Serum and Vitreous Humor Are Independent of Genetic Risk for Age-Related Macular Degeneration at the 10q26 Locus.

Purpose: The purpose of this study was to determine if levels of the HtrA1 protein in serum or vitreous humor are influenced by genetic risk for age-related macular degeneration (AMD) at the 10q26 locus, age, sex, AMD status, and/or AMD disease severity, and, therefore, to determine the contribution of systemic and ocular HtrA1 to the AMD disease process. Methods: A custom-made sandwich ELISA assay (SCTM ELISA) for detection of the HtrA1 protein was designed and compared with three commercial assays (R&D Systems, MyBiosource 1 and MyBiosource 2) using 65 serum samples. Concentrations of HtrA1 were thereafter determined in serum and vitreous samples collected from 248 individuals and 145 human donor eyes, respectively. Results: The SCTM ELISA demonstrated high specificity, good recovery, and parallelism within its linear detection range and performed comparably to the R&D Systems assay. In contrast, we were unable to demonstrate the specificity of the two assays from MyBioSource using either recombinant or native HtrA1. Analyses of concentrations obtained using the validated SCTM assay revealed that genetic risk at the 10q26 locus, age, sex, or AMD status are not significantly associated with altered levels of the HtrA1 protein in serum or in vitreous humor (P > 0.05). Conclusions: HtrA1 levels in serum and vitreous do not reflect the risk for AMD associated with the 10q26 locus or disease status. Localized alteration in HTRA1 expression in the retinal pigment epithelium, rather than systemic changes in HtrA1, is the most likely driver of elevated risk for developing AMD among individuals with risk variants at the 10q26 locus.

Replenishing Age-Related Decline of IRAK-M Expression in Retinal Pigment Epithelium Attenuates Outer Retinal Degeneration.

Unchecked, chronic inflammation is a constitutive component of age-related diseases, including age-related macular degeneration (AMD). Here we identified interleukin-1 receptor-associated kinase (IRAK)-M as a key immunoregulator in retinal pigment epithelium (RPE) that declines with age. Rare genetic variants of IRAK-M increased the likelihood of AMD. IRAK-M expression in RPE declined with age or oxidative stress and was further reduced in AMD. IRAK-M-deficient mice exhibited increased incidence of outer retinal degeneration at earlier ages, which was further exacerbated by oxidative stressors. The absence of IRAK-M disrupted RPE cell homeostasis, including compromised mitochondrial function, cellular senescence, and aberrant cytokine production. IRAK-M overexpression protected RPE cells against oxidative or immune stressors. Subretinal delivery of AAV-expressing IRAK-M rescued light-induced outer retinal degeneration in wild-type mice and attenuated age-related spontaneous retinal degeneration in IRAK-M-deficient mice. Our data support that replenishment of IRAK-M expression may redress dysregulated pro-inflammatory processes in AMD, thereby treating degeneration.

Role of Erythropoietin Receptor Signaling in Macrophages or Choroidal Endothelial Cells in Choroidal Neovascularization.

Erythropoietin (EPO) has been proposed to reduce the progression of atrophic age-related macular degeneration (AMD) due to its potential role in neuroprotection. However, overactive EPO receptor (EPOR) signaling increased laser-induced choroidal neovascularization (CNV) and choroidal macrophage number in non-lasered mice, which raised the question of whether EPOR signaling increased CNV through the recruitment of macrophages to the choroid that released pro-angiogenic factors or through direct angiogenic effects on endothelial cells. In this study, we addressed the hypothesis that EPOR signaling increased CNV by direct effects on macrophages or endothelial cells. We used tamoxifen-inducible macrophage-specific or endothelial cell-specific EPOR knockout mice in the laser-induced CNV model, and cultured choroidal endothelial cells isolated from adult human donors. We found that macrophage-specific knockout of EPOR influenced laser-induced CNV in females only, whereas endothelial-specific knockout of EPOR reduced laser-induced CNV in male mice only. In cultured human choroidal endothelial cells, knockdown of EPOR reduced EPO-induced signal transducer and activator of transcription 3 (STAT3) activation. Taken together, our findings suggest that EPOR signaling in macrophages or choroidal endothelial cells regulates the development of CNV in a sex-dependent manner. Further studies regarding the role of EPO-induced signaling are required to assess EPO safety and to select or develop appropriate therapeutic approaches.

Is water quality in British rivers "better than at any time since the end of the Industrial Revolution"?

We explore the oft-repeated claim that river water quality in Great Britain is "better now than at any time since the Industrial Revolution". We review available data and ancillary evidence for seven different categories of water pollutants: (i) biochemical oxygen demand (BOD) and ammonia; (ii) heavy metals; (iii) sewage-associated organic pollutants (including hormone-like substances, personal care product and pharmaceutical compounds); (iv) macronutrients (nitrogen and phosphorus); (v) pesticides; (vi) acid deposition and (vii) other variables, including natural organic matter and pathogenic micro-organisms. With a few exceptions, observed data are scarce before 1970. However, we can speculate about some of the major water quality pressures which have existed before that. Point-source pollutants are likely to have increased with population growth, increased connection rates to sewerage and industrialisation, although the increased provision of wastewater treatment during the 20th century will have mitigated this to some extent. From 1940 to the 1990s, pressures from nutrients and pesticides associated with agricultural intensification have increased in many areas. In parallel, there was an increase in synthetic organic compounds with a "down-the-drain" disposal pathway. The 1990s saw general reductions in mean concentrations of metals, BOD and ammonia (driven by the EU Urban Waste Water Treatment Directive), a levelling out of nitrate concentrations (driven by the EU Nitrate Directive), a decrease in phosphate loads from both point-and diffuse-sources and some recovery from catchment acidification. The current picture is mixed: water quality in many rivers downstream of urban centres has improved in sanitary terms but not with respect to emerging contaminants, while river quality in catchments with intensive agriculture is likely to remain worse now than before the 1960s. Water quality is still unacceptably poor in some water bodies. This is often a consequence of multiple stressors which need to be better-identified and prioritised to enable continued recovery.

Protective chromosome 1q32 haplotypes mitigate risk for age-related macular degeneration associated with the CFH-CFHR5 and ARMS2/HTRA1 loci.

BACKGROUND: Single-variant associations with age-related macular degeneration (AMD), one of the most prevalent causes of irreversible vision loss worldwide, have been studied extensively. However, because of a lack of refinement of these associations, there remains considerable ambiguity regarding what constitutes genetic risk and/or protection for this disease, and how genetic combinations affect this risk. In this study, we consider the two most common and strongly AMD-associated loci, the CFH-CFHR5 region on chromosome 1q32 (Chr1 locus) and ARMS2/HTRA1 gene on chromosome 10q26  (Chr10 locus). RESULTS: By refining associations within the CFH-CFHR5 locus, we show that all genetic protection against the development of AMD in this region is described by the combination of the amino acid-altering variant CFH I62V (rs800292) and genetic deletion of CFHR3/1. Haplotypes based on CFH I62V, a CFHR3/1 deletion tagging SNP and the risk variant CFH Y402H are associated with either risk, protection or neutrality for AMD and capture more than 99% of control- and case-associated chromosomes. We find that genetic combinations of CFH-CFHR5 haplotypes (diplotypes) strongly influence AMD susceptibility and that individuals with risk/protective diplotypes are substantially protected against the development of disease. Finally, we demonstrate that AMD risk in the ARMS2/HTRA1 locus is also mitigated by combinations of CFH-CFHR5 haplotypes, with Chr10 risk variants essentially neutralized by protective CFH-CFHR5 haplotypes. CONCLUSIONS: Our study highlights the importance of considering protective CFH-CFHR5 haplotypes when assessing genetic susceptibility for AMD. It establishes a framework that describes the full spectrum of AMD susceptibility using an optimal set of single-nucleotide polymorphisms with known functional consequences. It also indicates that protective or preventive complement-directed therapies targeting AMD driven by CFH-CFHR5 risk haplotypes may also be effective when AMD is driven by ARMS2/HTRA1 risk variants.

Chromosome 10q26-driven age-related macular degeneration is associated with reduced levels of HTRA1 in human retinal pigment epithelium.

Genome-wide association studies have identified the chromosome 10q26 (Chr10) locus, which contains the age-related maculopathy susceptibility 2 (ARMS2) and high temperature requirement A serine peptidase 1 (HTRA1) genes, as the strongest genetic risk factor for age-related macular degeneration (AMD) [L.G. Fritsche et al., Annu. Rev. Genomics Hum. Genet. 15, 151-171, (2014)]. To date, it has been difficult to assign causality to any specific single nucleotide polymorphism (SNP), haplotype, or gene within this region because of high linkage disequilibrium among the disease-associated variants [J. Jakobsdottir et al. Am. J. Hum. Genet. 77, 389-407 (2005); A. Rivera et al. Hum. Mol. Genet. 14, 3227-3236 (2005)]. Here, we show that HTRA1 messenger RNA (mRNA) is reduced in retinal pigment epithelium (RPE) but not in neural retina or choroid tissues derived from human donors with homozygous risk at the 10q26 locus. This tissue-specific decrease is mediated by the presence of a noncoding, cis-regulatory element overlapping the ARMS2 intron, which contains a potential Lhx2 transcription factor binding site that is disrupted by risk variant rs36212733. HtrA1 protein increases with age in the RPE-Bruch's membrane (BM) interface in Chr10 nonrisk donors but fails to increase in donors with homozygous risk at the 10q26 locus. We propose that HtrA1, an extracellular chaperone and serine protease, functions to maintain the optimal integrity of the RPE-BM interface during the aging process and that reduced expression of HTRA1 mRNA and protein in Chr10 risk donors impairs this protective function, leading to increased risk of AMD pathogenesis. HtrA1 augmentation, not inhibition, in high-risk patients should be considered as a potential therapy for AMD.

Macular retinal thickness differs markedly in age-related macular degeneration driven by risk polymorphisms on chromosomes 1 and 10.

The two most common genetic contributors to age-related macular degeneration (AMD), a leading cause of irreversible vision loss worldwide, are variants associated with CFH-CFHR5 on chromosome 1 (Chr1) and ARMS2/HTRA1 on chromosome 10 (Chr10). We sought to determine if risk and protective variants associated with these two loci drive differences in macular retinal thickness prior and subsequent to the onset of clinically observable signs of AMD. We considered 299 individuals (547 eyes) homozygous for risk variants or haplotypes on Chr1 or Chr10 exclusively (Chr1-risk and Chr10-risk, respectively) or homozygous for a neutral haplotype (Chr1-neu), for the protective I62 tagged haplotype (Chr1-prot-I62) or for the protection conferring CFHR1/3 deletion haplotype (Chr1-prot-del) on Chr1 without any risk alleles on Chr10. Among eyes with no clinically observable signs of AMD, the deletion of CFHR1/3, which is strongly protective against this disease, is associated with significantly thicker retinas in the perifovea. When controlling for age, Chr10-risk eyes with early or intermediate AMD have thinner retinas as compared to eyes from the Chr1-risk group with similar disease severity. Our analysis indicates that this difference likely results from distinct biological and disease initiation and progression events associated with Chr1- and Chr10-directed AMD.

Viable bacterial colonization is highly limited in the human intestine in utero.

Mucosal immunity develops in the human fetal intestine by 11-14 weeks of gestation, yet whether viable microbes exist in utero and interact with the intestinal immune system is unknown. Bacteria-like morphology was identified in pockets of human fetal meconium at mid-gestation by scanning electron microscopy (n = 4), and a sparse bacterial signal was detected by 16S rRNA sequencing (n = 40 of 50) compared to environmental controls (n = 87). Eighteen taxa were enriched in fetal meconium, with Micrococcaceae (n = 9) and Lactobacillus (n = 6) the most abundant. Fetal intestines dominated by Micrococcaceae exhibited distinct patterns of T cell composition and epithelial transcription. Fetal Micrococcus luteus, isolated only in the presence of monocytes, grew on placental hormones, remained viable within antigen presenting cells, limited inflammation ex vivo and possessed genomic features linked with survival in the fetus. Thus, viable bacteria are highly limited in the fetal intestine at mid-gestation, although strains with immunomodulatory capacity are detected in subsets of specimens.

Intra-Target Microdosing - A Novel Drug Development Approach: Proof of Concept, Safety, and Feasibility Study in Humans.

Intra-Target Microdosing (ITM) is a novel drug development approach aimed at increasing the efficiency of first-in-human (FIH) testing of new molecular entities (NMEs). ITM combines intra-target drug delivery and "microdosing," the subpharmacological systemic exposure. We hypothesized that when the target tissue is small (about 1/100th of total body mass), ITM can lead to target therapeutic-level exposure with minimal (microdose) systemic exposure. Each of five healthy male volunteers received insulin microdose into the radial artery or full therapeutic dose intravenously in separate visits. Insulin and glucose levels were similar between systemic administration and ITM administration in the ipsilateral hand, and glucose levels demonstrated a reduction in the ipsilateral hand but not in the contralateral hand. Positron emission tomography (PET) imaging of 18 F-fluorodeoxyglucose (FDG) uptake demonstrated differences between the ipsilateral and contralateral arms. The procedures were safe and well-tolerated. Results are consistent with ITM proof-of-concept (POC) and demonstrate the ethical, regulatory, and logistical feasibility of the approach.

The Burden of the "False-Negatives" in Clinical Development: Analyses of Current and Alternative Scenarios and Corrective Measures.

The "false-negatives" of clinical development are the effective treatments wrongly determined ineffective. Statistical errors leading to "false-negatives" are larger than those leading to "false-positives," especially in typically underpowered early-phase trials. In addition, "false-negatives" are usually eliminated from further testing, thereby limiting the information available on them. We simulated the impact of early-phase power on economic productivity in three developmental scenarios. Scenario 1, representing the current status quo, assumed 50% statistical power at phase II and 90% at phase III. Scenario 2 assumed increased power (80%), and Scenario 3, increased stringency of alpha (1%) at phase II. Scenario 2 led, on average, to a 60.4% increase in productivity and 52.4% increase in profit. Scenario 3 had no meaningful advantages. Our results suggest that additional costs incurred by increasing the power of phase II studies are offset by the increase in productivity. We discuss the implications of our results and propose corrective measures.

Microdosing and Other Phase 0 Clinical Trials: Facilitating Translation in Drug Development.

A number of drivers and developments suggest that microdosing and other phase 0 applications will experience increased utilization in the near-to-medium future. Increasing costs of drug development and ethical concerns about the risks of exposing humans and animals to novel chemical entities are important drivers in favor of these approaches, and can be expected only to increase in their relevance. An increasing body of research supports the validity of extrapolation from the limited drug exposure of phase 0 approaches to the full, therapeutic exposure, with modeling and simulations capable of extrapolating even non-linear scenarios. An increasing number of applications and design options demonstrate the versatility and flexibility these approaches offer to drug developers including the study of PK, bioavailability, DDI, and mechanistic PD effects. PET microdosing allows study of target localization, PK and receptor binding and occupancy, while Intra-Target Microdosing (ITM) allows study of local therapeutic-level acute PD coupled with systemic microdose-level exposure. Applications in vulnerable populations and extreme environments are attractive due to the unique risks of pharmacotherapy and increasing unmet healthcare needs. All phase 0 approaches depend on the validity of extrapolation from the limited-exposure scenario to the full exposure of therapeutic intent, but in the final analysis the potential for controlled human data to reduce uncertainty about drug properties is bound to be a valuable addition to the drug development process.

Spatiotemporal variation and social determinants of suicide in China, 2006-2012: findings from a nationally representative mortality surveillance system.

BACKGROUND: Suicide in China has declined since the 1990s. However, there has been limited investigation of the potential spatiotemporal variation and social determinants of suicide during subsequent periods. METHOD: Annual suicide counts from 2006 to 2012 stratified by county, 5-year age group (⩾15 years) and gender were obtained from the Chinese Disease Surveillance Points system. Trends and geographic differentials were examined using multilevel negative binomial regression models to explore spatiotemporal variation in suicide, and the role of key sociodemographic factors associated with suicide. RESULTS: The suicide rate (per 100 000) in China decreased from 14.7 to 9.1, 2006-2012. Rates of suicide were higher in males than females and increased substantially with age. Suicide rates were higher in rural areas compared with urban areas; however, urban-rural disparities reduced over time with a faster decline for rural areas. Within both urban and rural areas, higher rates of suicide were evident in areas with lower socio-economic circumstances (SEC) [rate ratio (RR) 1.85, 95% confidence interval (CI) 1.31-2.62]. Suicide rates varied more than twofold (median RR 2.06) across counties, and were highest in central and southwest regions of China. A high proportion of the divorced population, especially for younger females, was associated with lower suicide rates (RR 0.60, 95% CI 0.46-0.79). CONCLUSIONS: Geographic variations for suicide should be taken into account in policy making, particularly for older males living in rural areas and urban areas with low SEC. Measures to reduce disparities in socio-economic level and alleviate family relation stress are current priorities.

Greener neighbourhoods, slimmer children? Evidence from 4423 participants aged 6 to 13 years in the Longitudinal Study of Australian children.

OBJECTIVES: There is a growing belief that green space (for example, parks) help prevent obesity. There is evidence of an inverse association between green space and childhood body mass index (BMI); however, the majority of these studies are cross-sectional. Longitudinal studies that track change in BMI across childhood in relation to levels of green space proximity would improve the quality of evidence available for decision making. METHODS: Objectively measured BMI was obtained every 2 years between 2006 and 2012 for 4423 participants initially aged 6-7 years in the Longitudinal Study of Australian Children (LSAC). The LSAC is a nationally representative study on a range of health and socio-demographic measures. Using Australian Bureau of Statistics mesh block data, which classify small scale land areas based on the main usage, each participant was assigned an objective measure of green space availability within their Statistical Area (level 2) of residence. Gender-stratified multilevel linear regression was used to estimate BMI growth curves across childhood in relation to green space availability. Family income, Australian Indigenous status, mothers' education and language spoken were used to adjust for socio-economic confounding. RESULTS: Age was found to be an effect modifier of associations between green space and BMI for boys (P=0.005) and girls (P=0.048). As children grew older, an inverse patterning of BMI by green space availability emerged. These findings held after adjustment for socio-economic circumstances for boys (P=0.009), though were less robust for girls after this adjustment (P=0.056). CONCLUSION: A beneficial effect of green space on BMI emerges as children grow older. However, there was little additional benefit after a modest amount of green space was met. Further research is needed to understand whether the drivers of this effect are from age-specific mechanisms, or whether the benefit of living in a greener neighbourhood is accumulated through childhood.

Neighborhood walkability, fear and risk of falling and response to walking promotion: The Easy Steps to Health 12-month randomized controlled trial.

In older adults the relationships between health, fall-related risk factors, perceived neighborhood walkability, walking behavior and intervention impacts are poorly understood. To determine whether: i) health and fall-related risk factors were associated with perceptions of neighborhood walkability; ii) perceived environmental attributes, and fall-related risk factors predicted change in walking behavior at 12 months; and iii) perceived environmental attributes and fall-related risk factors moderated the effect of a self-paced walking program on walking behavior. Randomized trial on walking and falls conducted between 2009 and 2012 involving 315 community-dwelling inactive adults ≥ 65 years living in Sydney, Australia. Measures were: mobility status, fall history, injurious fall and fear of falling (i.e., fall-related risk factors), health status, walking self-efficacy and 11 items from the neighborhood walkability scale and planned walking ≥ 150 min/week at 12 months. Participants with poorer mobility, fear of falling, and poor health perceived their surroundings as less walkable. Walking at 12 months was significantly greater in "less greenery" (AOR = 3.3, 95% CI: 1.11-9.98) and "high traffic" (AOR = 1.98, 95% CI: 1.00-3.91) neighborhoods. The intervention had greater effects in neighborhoods perceived to have poorer pedestrian infrastructure (p for interaction = 0.036). Low perceived walkability was shaped by health status and did not appear to be a barrier to walking behavior. There appears to be a greater impact of, and thus, need for, interventions to encourage walking in environments perceived not to have supportive walking infrastructure. Future studies on built environments and walking should gather information on fall-related risk factors to better understand how these characteristics interact.

Investigating 'place effects' on mental health: implications for population-based studies in psychiatry.

Background. Interest in features of our local environments that may promote better mental health and wellbeing continues to rise among decision makers. Our purpose was to highlight a selection of these challenges and some promising avenues for enhancing the quality of evidence. Method. An analysis of approximately 267, 000 people was used to test the local relative deprivation hypothesis, wherein the shortfall of a person's socioeconomic circumstances from their neighbours is said to impact negatively upon mental health. This case was used to anchor further discussion of challenges to identifying and interpreting genuine 'place effects' from spurious correlations. Results. A Median Odds Ratio of 1.29 computed via multilevel logistic regression showed that the odds of experiencing psychological distress (as measured by the Kessler score) varied by geographical area. Approximately 67% of this was attributed to a cross-classified measure of household income and neighbourhood deprivation. Compared to people on high incomes living in affluent neighbourhoods, the odds ratio of psychological distress for people on low incomes in affluent areas was 4.73 (95% confidence interval (95% CI) 4.39, 5.09), whereas that for people on low incomes in deprived areas was significantly higher at 5.83 (95% CI 5.41, 6.28). Conclusions. While no evidence was found to support local relative deprivation hypothesis, the pattern suggests that more affluent areas may contain features that are conducive to better mental health. Selection of bespoke geographical boundaries, use of directed acyclic graphs and more evaluations of natural experiments are likely to be important in taking the field of enquiry onwards.