Gene expression-based comparison of the human secretory neuroepithelia of the brain choroid plexus and the ocular ciliary body:potential implications for glaucoma

BACKGROUND: The neuroepithelia of the choroid plexus (CP) in the brain and the ciliary body (CB) of the eye have common embryological origins and share similar micro-structure and functions. The CP epithelium (CPE) and the non-pigmented epithelium (NPE) of the CB produce the cerebrospinal fluid (CSF) and the aqueous humor (AH) respectively. Production and outflow of the CSF determine the intracranial pressure (ICP); production and outflow of the AH determine the intraocular pressure (IOP). Together, the IOP and ICP determine the translaminar pressure on the optic disc which may be involved in the pathophysiology of primary open angle glaucoma (POAG). The aim of this study was to compare the molecular machinery of the secretory neuroepithelia of the CP and CB (CPE versus NPE) and to determine their potential role in POAG. METHODS: We compared the transcriptomes and functional annotations of healthy human CPE and NPE. Microarray and bioinformatic studies were performed using an Agilent platform and the Ingenuity Knowledge Database (IPA). RESULTS: Based on gene expression profiles, we found many similar functions for the CPE and NPE including molecular transport, neurological disease processes, and immunological functions. With commonly-used selection criteria (fold-change > 2.5, p-value < 0.05), 14% of the genes were expressed significantly differently between CPE and NPE. When we used stricter selection criteria (fold-change > 5, p-value < 0.001), still 4.5% of the genes were expressed differently, which yielded specific functions for the CPE (ciliary movement and angiogenesis/hematopoiesis) and for the NPE (neurodevelopmental properties). Apart from a few exceptions (e.g. SLC12A2, SLC4A4, SLC4A10, KCNA5, and SCN4B), all ion transport protein coding genes involved in CSF and AH production had similar expression profiles in CPE and NPE. Three POAG disease genes were expressed significantly higher in the CPE than the NPE, namely CDH1, CDKN2B and SIX1. CONCLUSIONS: The transcriptomes of the CPE and NPE were less similar than we previously anticipated. High expression of CSF/AH production genes and candidate POAG disease genes in the CPE and NPE suggest that both might be involved in POAG

Gene Expression and Functional Annotation of the Human Ciliary Body Epithelia

Purpose: The ciliary body (CB) of the human eye consists of the non-pigmented (NPE) and pigmented (PE) neuro-epithelia. We investigated the gene expression of NPE and PE, to shed light on the molecular mechanisms underlying the most important functions of the CB. We also developed molecular signatures for the NPE and PE and studied possible new clues for glaucoma. Methods: We isolated NPE and PE cells from seven healthy human donor eyes using laser dissection microscopy. Next, we performed RNA isolation, amplification, labeling and hybridization against 44×k Agilent microarrays. For microarray conformations, we used a literature study, RT-PCRs, and immunohistochemical stainings. We analyzed the gene expression data with R and with the knowledge database Ingenuity. Results: The gene expression profiles and functional annotations of the NPE and PE were highly similar. We found that the most important functionalities of the NPE and PE were related to developmental processes, neural nature of the tissue, endocrine and metabolic signaling, and immunological functions. In total 1576 genes differed statistically significantly between NPE and PE. From these genes, at least 3 were cell-specific for the NPE and 143 for the PE. Finally, we observed high expression in the (N)PE of 35 genes previously implicated in molecular mechanisms related to glaucoma. Conclusion: Our gene expression analysis suggested that the NPE and PE of the CB were quite similar. Nonetheless, cell-type specific differences were found. The molecular machineries of the human NPE and PE are involved in a range of neuro-endocrinological, developmental and immunological functions, and perhaps glaucoma

The Mass Concentrations of Serum Troponin T and Creatine Kinase-MB are Elevated before Creatine Kinase and Creatine Kinase-MB Activities in Acute Myocardial Infarction

Peer Reviewe

Developmental defects and male sterility in mice lacking the ubiquitin-like DNA repair gene mHR23B.

mHR23B encodes one of the two mammalian homologs of Saccharomyces cerevisiae RAD23, a ubiquitin-like fusion protein involved in nucleotide excision repair (NER). Part of mHR23B is complexed with the XPC protein, and this heterodimer functions as the main damage detector and initiator of global genome NER. While XPC defects exist in humans and mice, mutations for mHR23A and mHR23B are not known. Here, we present a mouse model for mHR23B. Unlike XPC-deficient cells, mHR23B(-/-) mouse embryonic fibroblasts are not UV sensitive and retain the repair characteristics of wild-type cells. In agreement with the results of in vitro repair studies, this indicates that mHR23A can functionally replace mHR23B in NER. Unexpectedly, mHR23B(-/-) mice show impaired embryonic development and a high rate (90%) of intrauterine or neonatal death. Surviving animals display a variety of abnormalities, including retarded growth, facial dysmorphology, and male sterility. Such abnormalities are not observed in XPC and other NER-deficient mouse mutants and point to a separate function of mHR23B in development. This function may involve regulation of protein stability via the ubiquitin/proteasome pathway and is not or only in part compensated for by mHR23A

Dietary magnesium, not calcium, prevents vascular calcification in a mouse model for pseudoxanthoma elasticum

Pseudoxanthoma elasticum (PXE) is a heritable disorder characterized by ectopic calcification of connective tissue in skin, Bruch’s membrane of the eye, and walls of blood vessels. PXE is caused by mutations in the ABCC6 gene, but the exact etiology is still unknown. While observations on patients suggest that high calcium intake worsens the clinical symptoms, the patient organization PXE International has published the dietary advice to increase calcium intake in combination with increased magnesium intake. To obtain more data on this controversial issue, we examined the effect of dietary calcium and magnesium in the Abcc6−/− mouse, a PXE mouse model which mimics the clinical features of PXE. Abcc6−/− mice were placed on specific diets for 3, 7, and 12 months. Disease severity was measured by quantifying calcification of blood vessels in the kidney. Raising the calcium content in the diet from 0.5% to 2% did not change disease severity. In contrast, simultaneous increase of both calcium (from 0.5% to 2.0%) and magnesium (from 0.05% to 0.2%) slowed down the calcification significantly. Our present findings that increase in dietary magnesium reduces vascular calcification in a mouse model for PXE should stimulate further studies to establish a dietary intervention for PXE

Impaired Genome Maintenance Suppresses the Growth Hormone–Insulin-Like Growth Factor 1 Axis in Mice with Cockayne Syndrome

Cockayne syndrome (CS) is a photosensitive, DNA repair disorder associated with progeria that is caused by a defect in the transcription-coupled repair subpathway of nucleotide excision repair (NER). Here, complete inactivation of NER in Csb(m/m)/Xpa(−/−) mutants causes a phenotype that reliably mimics the human progeroid CS syndrome. Newborn Csb(m/m)/Xpa(−/−) mice display attenuated growth, progressive neurological dysfunction, retinal degeneration, cachexia, kyphosis, and die before weaning. Mouse liver transcriptome analysis and several physiological endpoints revealed systemic suppression of the growth hormone/insulin-like growth factor 1 (GH/IGF1) somatotroph axis and oxidative metabolism, increased antioxidant responses, and hypoglycemia together with hepatic glycogen and fat accumulation. Broad genome-wide parallels between Csb(m/m)/Xpa(−/−) and naturally aged mouse liver transcriptomes suggested that these changes are intrinsic to natural ageing and the DNA repair–deficient mice. Importantly, wild-type mice exposed to a low dose of chronic genotoxic stress recapitulated this response, thereby pointing to a novel link between genome instability and the age-related decline of the somatotroph axis

Vitamin K supplementation increases vitamin K tissue levels but fails to counteract ectopic calcification in a mouse model for pseudoxanthoma elasticum

Pseudoxanthoma elasticum (PXE) is an autosomal recessive disorder in which calcification of connective tissue leads to pathology in skin, eye and blood vessels. PXE is caused by mutations in ABCC6. High expression of this transporter in the basolateral hepatocyte membrane suggests that it secretes an as-yet elusive factor into the circulation which prevents ectopic calcification. Utilizing our Abcc6−/− mouse model for PXE, we tested the hypothesis that this factor is vitamin K (precursor) (Borst et al. 2008, Cell Cycle). For 3 months, Abcc6−/− and wild-type mice were put on diets containing either the minimum dose of vitamin K required for normal blood coagulation or a dose that was 100 times higher. Vitamin K was supplied as menaquinone-7 (MK-7). Ectopic calcification was monitored in vivo by monthly micro-CT scans of the snout, as the PXE mouse model develops a characteristic connective tissue mineralization at the base of the whiskers. In addition, calcification of kidney arteries was measured by histology. Results show that supplemental MK-7 had no effect on ectopic calcification in Abcc6−/− mice. MK-7 supplementation increased vitamin K levels (in skin, heart and brain) in wild-type and in Abcc6−/− mice. Vitamin K tissue levels did not depend on Abcc6 genotype. In conclusion, dietary MK-7 supplementation increased vitamin K tissue levels in the PXE mouse model but failed to counteract ectopic calcification. Hence, we obtained no support for the hypothesis that Abcc6 transports vitamin K and that PXE can be cured by increasing tissue levels of vitamin K

The role of cardiovascular magnetic resonance imaging and computed tomography angiography in suspected non-ST-elevation myocardial infarction patients:Design and rationale of the CARdiovascular Magnetic rEsoNance imaging and computed Tomography Angiography (CARMENTA) trial

BackgroundAlthough high-sensitivity cardiac troponin (hs-cTn) substantially improves the early detection of myocardial injury, it lacks specificity for acute myocardial infarction (MI). In suspected non–ST-elevation MI, invasive coronary angiography (ICA) remains necessary to distinguish between acute MI and noncoronary myocardial disease (eg, myocarditis), unnecessarily subjecting the latter to ICA and associated complications. This trial investigates whether implementing cardiovascular magnetic resonance (CMR) or computed tomography angiography (CTA) early in the diagnostic process may help to differentiate between coronary and noncoronary myocardial disease, thereby preventing unnecessary ICA.Study DesignIn this prospective, single-center, randomized controlled clinical trial, 321 consecutive patients with acute chest pain, elevated hs-cTnT, and nondiagnostic electrocardiogram are randomized to 1 of 3 strategies: (1) CMR, or (2) CTA early in the diagnostic process, or (3) routine clinical management. In the 2 investigational arms of the study, results of CMR or CTA will guide further clinical management. It is expected that noncoronary myocardial disease is detected more frequently after early noninvasive imaging as compared with routine clinical management, and unnecessary ICA will be prevented. The primary end point is the total number of patients undergoing ICA during initial admission. Secondary end points are 30-day and 1-year clinical outcome (major adverse cardiac events and major procedure-related complications), time to final diagnosis, quality of life, and cost-effectiveness.ConclusionThe CARMENTA trial investigates whether implementing CTA or CMR early in the diagnostic process in suspected non–ST-elevation MI based on elevated hs-cTnT can prevent unnecessary ICA as compared with routine clinical management, with no detrimental effect on clinical outcome

Mitochondrial DNA D-loop variants correlate with a primary open-angle glaucoma subgroup

IntroductionPrimary open-angle glaucoma (POAG) is a characteristic optic neuropathy, caused by degeneration of the optic nerve-forming neurons, the retinal ganglion cells (RGCs). High intraocular pressure (IOP) and aging have been identified as major risk factors; yet the POAG pathophysiology is not fully understood. Since RGCs have high energy requirements, mitochondrial dysfunction may put the survivability of RGCs at risk. We explored in buffy coat DNA whether mtDNA variants and their distribution throughout the mtDNA could be risk factors for POAG.MethodsThe mtDNA was sequenced from age- and sex-matched study groups, being high tension glaucoma (HTG, n=71), normal tension glaucoma patients (NTG, n=33), ocular hypertensive subjects (OH, n=7), and cataract controls (without glaucoma; n=30), all without remarkable comorbidities.ResultsNo association was found between the number of mtDNA variants in genes encoding proteins, tRNAs, rRNAs, and in non-coding regions in the different study groups. Next, variants that controls shared with the other groups were discarded. A significantly higher number of exclusive variants was observed in the D-loop region for the HTG group (~1.23 variants/subject), in contrast to controls (~0.35 variants/subject). In the D-loop, specifically in the 7S DNA sub-region within the Hypervariable region 1 (HV1), we found that 42% of the HTG and 27% of the NTG subjects presented variants, while this was only 14% for the controls and OH subjects. As we have previously reported a reduction in mtDNA copy number in HTG, we analysed if specific D-loop variants could explain this. While the majority of glaucoma patients with the exclusive D-loop variants m.72T&gt;C, m.16163 A&gt;G, m.16186C&gt;T, m.16298T&gt;C, and m.16390G&gt;A presented a mtDNA copy number below controls median, no significant association between these variants and low copy number was found and their possible negative role in mtDNA replication remains uncertain. Approximately 38% of the HTG patients with reduced copy number did not carry any exclusive D-loop or other mtDNA variants, which indicates that variants in nuclear-encoded mitochondrial genes, environmental factors, or aging might be involved in those cases.ConclusionIn conclusion, we found that variants in the D-loop region may be a risk factor in a subgroup of POAG, possibly by affecting mtDNA replication