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Mitochondrial (mt) DNA haplogroups, defined by specific single nucleotide polymorphism (SNP) patterns, represent populations of diverse geographic origins and have been associated with increased risk or protection of many diseases. The H haplogroup is the most common European haplogroup while the K haplogroup is highly associated with the Ashkenazi Jewish population. Transmitochondrial cybrids (cell lines with identical nuclei, but mtDNA from either H (n = 8) or K (n = 8) subjects) were analyzed by the Seahorse flux analyzer, quantitative polymerase chain reaction (Q-PCR) and immunohistochemistry (IHC). Cybrids were treated with amyloid-β peptides and cell viabilities were measured. Other cybrids were demethylated with 5-aza-2′-deoxycytidine (5-aza-dC) and expression levels for APOE and NFkB2 were measured. Results show K cybrids have (a) significantly lower mtDNA copy numbers, (b) higher expression levels for MT-DNA encoded genes critical for oxidative phosphorylation, (c) lower Spare Respiratory Capacity, (d) increased expression of inhibitors of the complement pathway and important inflammasome-related genes; and (e) significantly higher levels of APOE transcription that were independent of methylation status. After exposure to amyloid-β1–42 peptides (active form), H haplogroup cybrids demonstrated decreased cell viability compared to those treated with amyloid-β42–1 (inactive form) (p < 0.0001), while this was not observed in the K cybrids (p = 0.2). K cybrids had significantly higher total global methylation levels and differences in expression levels for two acetylation genes and four methylation genes. Demethylation with 5-aza-dC altered expression levels for NFkB2, while APOE transcription patterns were unchanged. Our findings support the hypothesis that mtDNA-nuclear retrograde signaling may mediate expression levels of APOE, a key factor in many age-related diseases. Future studies will focus on identification of the mitochondrial-nuclear retrograde signaling mechanism(s) contributing to these mtDNA-mediated differences.

Background: Cisplatin, a powerful antitumor agent, causes formation of DNA adducts, and activation of apoptotic pathways. Presently, cisplatin resistance develops in up to 70% of patients but the underlying molecular mechanism(s) are unclear and there are no markers to determine which patients will become resistant. Mitochondria play a significant role not only in energy metabolism but also retrograde signaling (mitochondria to nucleus) that modulates inflammation, complement, and apoptosis pathways. Maternally inherited mitochondrial (mt) DNA can be classified into haplogroups representing different ethnic populations that have diverse susceptibilities to diseases and medications. Methods: Transmitochondrial cybrids, where all cell lines possess identical nuclear genomes but either the H (Southern European) or J (Northern European) mtDNA haplogroups, were treated with cisplatin and analyzed for differential responses related to viability, oxidative stress, and expression levels of genes associated with cancer, cisplatin-induced nephrotoxicity and resistance, apoptosis and signaling pathways. Results: The cisplatin-treated-J cybrids showed greater loss of cell viability along with lower levels of reactive oxygen species and mitochondrial membrane potential compared to cisplatin-treated-H cybrids. After cisplatin treatment, J cybrids showed increased gene expression of BAX, CASP3, and CYP51A, but lower levels of SFRP1 compared to untreated-J cybrids. The cisplatin-treated-H cybrids had elevated expression of CDKN1A/P21, which has a role in cisplatin toxicity, compared to untreated-H cybrids. The cisplatin-treated H had higher transcription levels of ABCC1, DHRS2/HEP27, and EFEMP1 compared to cisplatin-treated-J cybrids. Conclusions: Cybrid cell lines that contain identical nuclei but either H mtDNA mitochondria or J mtDNA mitochondria respond differently to cisplatin treatments suggesting involvement of the retrograde signaling (from mitochondria to nucleus) in the drug-induced cell death. Varying toxicities and transcription levels of the H vs. J cybrids after cisplatin treatment support the hypothesis that mtDNA variants play a role in the expression of genes affecting resistance and side effects of cisplatin.

Mitochondrial (mt) DNA haplogroups, defined by specific single nucleotide polymorphism (SNPs) patterns, represent populations of diverse geographic origins and may play a role in disparate disease susceptibilities found in different ethnic/racial populations. The most common European haplogroup is H, while the K haplogroup is highly associated with Ashkenazi Jewish populations. Studies using transmitochondrial cybrids (cell lines with identical nuclei but mitochondria from either H or K haplogroup subjects) demonstrated significant molecular and biological differences but mechanisms for these disparities are unclear. In this study, we hypothesized that there is differential retrograde signaling occurring between the Stimulator of Interferon Genes (STING) pathway and H versus K mtDNA haplogroups. Results showed that K cybrids exhibit increased levels of cytoplasmic mtDNA fragments. After STING Knock-Down, H cybrids had lower expression levels for EGFR, BRCA1, DNMT3A, DNMT3B, HDAC1, and IFNα genes, but upregulated DNMT3A compared to control H cybrids. The STING-KD K cybrids showed downregulation of EGFR, DNMT3A, HDAC1, HCAD9, CFH, and CHI, along with upregulation of DNMT1 and IL-6 compared to control K cybrids. Since all cybrids have identical nuclei, the STING DNA sensor system interacts differently with K haplogroup mtDNA compared to H mtDNA for genes related to cancer (EGFR, BRCA1), methylation (DNMT1, DNMT3A, DNMT3B), acetylation (HDAC1, HDCA9), complement (CFH, CHI) and inflammation (IFNα, IL-6). In summary, in non-pathologic conditions, (a) STING is an important retrograde signaling mechanism(s) and (b) cybrids possessing Ashkenazi Jewish mtDNA (K haplogroup) interact with the STING complex differently compared to H cybrids which affects various disease-related pathways.

The geographic origins of populations can be identified by their maternally inherited mitochondrial DNA (mtDNA) haplogroups. This study compared human cybrids (cytoplasmic hybrids), which are cell lines with identical nuclei but mitochondria from different individuals with mtDNA from either the H haplogroup or L haplogroup backgrounds. The most common European haplogroup is H while individuals of maternal African origin are of the L haplogroup. Despite lower mtDNA copy numbers, L cybrids had higher expression levels for nine mtDNA-encoded respiratory complex genes, decreased ATP (adenosine triphosphate) turnover rates and lower levels of reactive oxygen species production, parameters which are consistent with more efficient oxidative phosphorylation. Surprisingly, GeneChip arrays showed that the L and H cybrids had major differences in expression of genes of the canonical complement system (5 genes), dermatan/chondroitin sulfate biosynthesis (5 genes) and CCR3 (chemokine, CC motif, receptor 3) signaling (9 genes). Quantitative nuclear gene expression studies confirmed that L cybrids had (a) lower expression levels of complement pathway and innate immunity genes and (b) increased levels of inflammation-related signaling genes, which are critical in human diseases. Our data support the hypothesis that mtDNA haplogroups representing populations from different geographic origins may play a role in differential susceptibilities to diseases.

A new approach for obtaining metasurfaces realized by using frequency selective surfaces with reduced periodicity printed on a new class of high dielectric constant materials is presented. The proposed engineered ad-hoc materials and the innovative electromagnetic design provide an improvement of the overall device in terms of compactness and angular stability, as shown by the use of our novel band vs. incidence angles chart. © 2009 Wiley Periodicals, Inc. Microwave Opt Technol Lett 51: 2753–2758, 2009; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/mop.24731

In a volcano monitoring application, various geophysical and geochemical sensors generate continuous high-fidelity data, and there is a compelling need for real-time raw data for volcano eruption prediction research. It requires the network to support network synchronized sampling, online configurable sensing and situation awareness, which pose significant challenges on sensing component design. Ideally, the resource usages shall be driven by the environment and node situations, and the data quality is optimized under resource constraints. In this paper, we present our smart sensing component design, including hybrid time synchronization, configurable sensing, and situation awareness. Both design details and evaluation results are presented to show their efficiency. Although the presented design is for a volcano monitoring application, its design philosophy and framework can also apply to other similar applications and platforms.

Mitochondrial (mt) DNA can be classified into haplogroups representing different geographic and/or racial origins of populations. The H haplogroup is protective against age-related macular degeneration (AMD), while the J haplogroup is high risk for AMD. In the present study, we performed comparison analyses of human retinal cell cybrids, which possess identical nuclei, but mtDNA from subjects with either the H or J haplogroups, and demonstrate differences in total global methylation, and expression patterns for two genes related to acetylation and five genes related to methylation. Analyses revealed that untreated-H and -J cybrids have different expression levels for nuclear genes (CFH, EFEMP1, VEGFA and NFkB2). However, expression levels for these genes become equivalent after treatment with a methylation inhibitor, 5-aza-2'-deoxycytidine. Moreover, sequencing of the entire mtDNA suggests that differences in epigenetic status found in cybrids are likely due to single nucleotide polymorphisms (SNPs) within the haplogroup profiles rather than rare variants or private SNPs. In conclusion, our findings indicate that mtDNA variants can mediate methylation profiles and transcription for inflammation, angiogenesis and various signaling pathways, which are important in several common diseases.

The yeast Saccharomyces cerevisiae has a finite replicative life span. Yeasts possess two prohibitins, Phb1p and Phb2p, in similarity to mammalian cells. These proteins are located in the inner mitochondrial membrane, where they are involved in the processing of newly-synthesized membrane proteins. We demonstrate that the elimination of one or both of the prohibitin genes in yeast markedly diminished the replicative life span of cells that lack fully-functional mitochondria, while having no effect on cells with functioning mitochondria. This deleterious effect was suppressed by the deletion of the RAS2 gene. The expression of PHB1 and PHB2 declined gradually up to 5-fold during the life span. Cells in which PHB1 was deleted in conjunction with the absence of a mitochondrial genome displayed remarkable changes in mitochondrial morphology, distribution, and inheritance. This loss of mitochondrial integrity was not seen in cells devoid of PHB1 but possessing an intact mitochondrial genome. In a subset of the cells, the changes in mitochondrial integrity were associated with increased production of reactive oxygen species, which co-localized with the altered mitochondria. The mitochondrial deficits described above were all suppressed by deletion of RAS2. Our data, together with published information, are interpreted to provide a unified view of the role of the prohibitins in yeast aging. This model posits that the key initiating event is a decline in mitochondrial function, which leads to progressive oxidative damage that is exacerbated in the absence of the prohibitins. This aggravation of the initial damage is ameliorated by the suppression of the production of mitochondrial proteins in the absence of Ras2p signaling of mitochondrial biogenesis.

Human Molecular Genetics, 2014, Vol. 23, No. 13 doi:10.1093/hmg/ddu065 Advance Access published on February 28, 2014 Inherited mitochondrial DNA variants can affect complement, inflammation and apoptosis pathways: insights into mitochondrial –nuclear interactions M. Cristina Kenney 1,2, ∗ , Marilyn Chwa 1 , Shari R. Atilano 1 , Payam Falatoonzadeh 1 , Claudio Ramirez 1 , Deepika Malik 1 , Mohamed Tarek 1 , Javier Ca´ceres-del-Carpio 1 , Anthony B. Nesburn 1,4 , David S. Boyer 5 , Baruch D. Kuppermann 1 , Marquis Vawter 3 , S. Michal Jazwinski 6 , Michael Miceli 6 , Douglas C. Wallace 7 and Nitin Udar 1 Gavin Herbert Eye Institute, 2 Department of Pathology and Laboratory Medicine, 3 Functional Genomics Laboratory, Department of Psychiatry and Human Behavior, University of California Irvine, Irvine, CA, USA, 4 Cedars-Sinai Medical Center, Los Angeles, CA, USA, 5 Retina-Vitreous Associates Medical Group, Beverly Hills, CA, USA, 6 Tulane Center for Aging, Tulane University, New Orleans, LA, USA and 7 Children’s Hospital of Philadelphia, Center for Mitochondrial and Epigenomic Medicine, Philadelphia, PA, USA Received December 17, 2013; Revised February 3, 2014; Accepted February 10, 2014 Age-related macular degeneration (AMD) is the leading cause of vision loss in developed countries. While linked to genetic polymorphisms in the complement pathway, there are many individuals with high risk alleles that do not develop AMD, suggesting that other ‘modifiers’ may be involved. Mitochondrial (mt) haplogroups, defined by accu- mulations of specific mtDNA single nucleotide polymorphisms (SNPs) which represent population origins, may be one such modifier. J haplogroup has been associated with high risk for AMD while the H haplogroup is protective. It has been difficult to assign biological consequences for haplogroups so we created human ARPE-19 cybrids (cyto- plasmic hybrids), which have identical nuclei but mitochondria of either J or H haplogroups, to investigate their effects upon bioenergetics and molecular pathways. J cybrids have altered bioenergetic profiles compared with H cybrids. Q-PCR analyses show significantly lower expression levels for seven respiratory complex genes encoded by mtDNA. J and H cybrids have significantly altered expression of eight nuclear genes of the alternative complement, inflammation and apoptosis pathways. Sequencing of the entire mtDNA was carried out for all the cybrids to identify haplogroup and non-haplogroup defining SNPs. mtDNA can mediate cellular bioenergetics and expression levels of nuclear genes related to complement, inflammation and apoptosis. Sequencing data sug- gest that observed effects are not due to rare mtDNA variants but rather the combination of SNPs representing the J versus H haplogroups. These findings represent a paradigm shift in our concepts of mt–nuclear interactions. INTRODUCTION Mitochondria (mt) are unique organelles that have their own DNA. Human mtDNA is maternally inherited and forms a circle of double-stranded DNA with 16 569 nucleotide pairs. The non-coding mt-Dloop (also called the control region) con- tains 1121 nucleotides important for replication and transcrip- tion. The coding region of mtDNA encodes for 37 genes, including 13 protein subunits essential for oxidative phosphorylation (OXPHOS), 2 ribosomal RNAs and 22 transfer RNAs (1 – 3). The mtDNA can be categorized into haplogroups that are defined by a group of single nucleotide polymorphism (SNP) variants that have accumulated over tens of thousands of years and correspond to various geographic populations. While some studies have shown that certain mtDNA hap- logroups can be either protective or increase risk for human dis- eases (4–10), others report no haplogroup association with diseases (11– 13). However, many of these studies identified To whom correspondence should be addressed at: Gavin Herbert Eye Institute, Discovery Center for Eye Research, University of California Irvine, Hewitt Hall, Room 2028, 843 Health Science Road, Irvine, CA 92697, USA. Tel: +1 9498247603; Fax: +1 9498249626; Email: mkenney@uci.edu # The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Objectives: In this study we evaluated the association between chronic prostatitis syndrome (CPS), varicocele and hemorrhoids as manifestations of a pelvic venous disease.

Purpose. Due to a possible link in humans between atherosclerosis, a high-fat diet, and the development of age-related retinal degenerations, we investigated retinal changes with age and diet in high-fat-fed C57BL/6 mice, a mouse model of human atherosclerosis. Methods. We fed C57BL/6J mice either a normal chow diet or an atherogenic diet containing 15% total fat for either 15, 30 or 45 weeks. We sacrificed the animals and examined the eyes using fluorescence microscopy, light microscopy and transmission electron microscopy. Results. At 15 weeks, in high-fat-fed mice, there was an increase in the number of autofluorescent granules in the retinal pigment epithelium (RPE) compared to chow-fed mice. By light microscopy, we noted no remarkable differences in the inner retinal layers between mice fed either diet for 15 or 30 weeks. In contrast, we noted major changes in transmission electron micrographs from the 30 week high-fat group. In the RPE, these included: (1) an increase in the number and size of autop...

We have previously shown that an experimental, low-zinc environment decreased catalase activity in cultured human fetal retinal pigment epithelial (RPE) cells. The purpose of this study was to investigate the effect of zinc supplementation on catalase expression in cultured human fetal RPE cells.Confluent fetal RPE cells incubated in Coon's modified Ham's F12 (CMF-12) were treated (18 h) with zinc chloride (ZnCl2) (15, 30, or 100 microM) to assess changes in catalase enzyme activity or for 6 h to assess the induction of catalase mRNA by Northern analysis and in situ hybridization. RPE cells were also treated with 30 microM ZnCl2 for 2, 6, 24, 48 and 72 h to assess the time course of changes in catalase enzyme activity, changes in mRNA levels and status of the Sp1 transcription factor.Catalase activity was increased above control by the addition of 15, 30 and 100 microM ZnCl2. Catalase gene expression was induced by 30 microM zinc in 6 h, but decreased to non-treated control levels by 24 h. The transcription factor Sp1 was also activated by zinc treatment (30 microM) which peaked at 2 h and declined to non-treated control levels by 24 h. Catalase enzyme activity peaked at 24 h and decreased to control levels by 72 h.Our results demonstrate that zinc treatment of RPE cells increases catalase expression and activates the transcription factor Sp1. The results suggest zinc may play a role in the transcriptional regulation of catalase in RPE cells.

Mitochondrial DNA Variants Mediate Energy Production and Expression Levels for CFH, C3 and EFEMP1 Genes: Implications for Age-Related Macular Degeneration M. Cristina Kenney 1 *, Marilyn Chwa 1 , Shari R. Atilano 1 , Janelle M. Pavlis 1 , Payam Falatoonzadeh 1 , Claudio Ramirez 1 , Deepika Malik 1 , Tiffany Hsu 1 , Grace Woo 1 , Kyaw Soe 1 , Anthony B. Nesburn 1,2 , David S. Boyer 3 , Baruch D. Kuppermann 1 , S. Michal Jazwinski 4 , Michael V. Miceli 4 , Douglas C. Wallace 5 , Nitin Udar 1 1 Gavin Herbert Eye Institute, University of California Irvine, Irvine, California, United States of America, 2 Cedars-Sinai Medical Center, Los Angeles, California, United States of America, 3 Retina-Vitreous Associates Medical Group, Beverly Hills, California, United States of America, 4 Tulane Center for Aging and Department of Medicine, Tulane University, New Orleans, Louisiana, United States of America, 5 Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania, United States of America Abstract Background: Mitochondrial dysfunction is associated with the development and progression of age-related macular degeneration (AMD). Recent studies using populations from the United States and Australia have demonstrated that AMD is associated with mitochondrial (mt) DNA haplogroups (as defined by combinations of mtDNA polymorphisms) that represent Northern European Caucasians. The aim of this study was to use the cytoplasmic hybrid (cybrid) model to investigate the molecular and biological functional consequences that occur when comparing the mtDNA H haplogroup (protective for AMD) versus J haplogroup (high risk for AMD). Methodology/Principal Findings: Cybrids were created by introducing mitochondria from individuals with either H or J haplogroups into a human retinal epithelial cell line (ARPE-19) that was devoid of mitochondrial DNA (Rho0). In cybrid lines, all of the cells carry the same nuclear genes but vary in mtDNA content. The J cybrids had significantly lower levels of ATP and reactive oxygen/nitrogen species production, but increased lactate levels and rates of growth. Q-PCR analyses showed J cybrids had decreased expressions for CFH, C3, and EFEMP1 genes, high risk genes for AMD, and higher expression for MYO7A, a gene associated with retinal degeneration in Usher type IB syndrome. The H and J cybrids also have comparatively altered expression of nuclear genes involved in pathways for cell signaling, inflammation, and metabolism. Conclusion/Significance: Our findings demonstrate that mtDNA haplogroup variants mediate not only energy production and cell growth, but also cell signaling for major molecular pathways. These data support the hypothesis that mtDNA variants play important roles in numerous cellular functions and disease processes, including AMD. Citation: Kenney MC, Chwa M, Atilano SR, Pavlis JM, Falatoonzadeh P, et al. (2013) Mitochondrial DNA Variants Mediate Energy Production and Expression Levels for CFH, C3 and EFEMP1 Genes: Implications for Age-Related Macular Degeneration. PLoS ONE 8(1): e54339. doi:10.1371/journal.pone.0054339 Editor: Walter Lukiw, Louisiana State University Health Sciences Center, United States of America Received October 1, 2012; Accepted December 10, 2012; Published January 24, 2013 Copyright: s 2013 Kenney et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Funding provided by The Discovery Eye Foundation www.discoveryeye.org, Lincy Foundation lincyinstitute.unlv.edu/lincy.html, Beckman Macular Research Initiative www.beckmanmacular.org, The Henry Guenther Foundation, Polly and Michael Smith Foundation, Research to Prevent Blindness Foundation www.rpbusa.org, and the National Institute on Aging (AG006168). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: mkenney@uci.edu specific SNP variants that have accumulated over tens of thousands of years and correspond to different geographic populations of the world. The H haplogroup is the most common European haplogroup, while the J haplogroup originates from the Northern European region and is defined by SNP variants that are associated with heat production as an adaptation to colder climates [4]. The mtDNA plays an important role in aging and diseases [4– 6]. Specific haplogroups are associated with a variety of eye diseases including age-related macular degeneration (AMD) [7– 10], diabetic retinopathy [11], pseudoexfoliation glaucoma [12,13], primary open-angle glaucoma [14], keratoconus [15], multiple sclerosis-related optic neuritis [16,17], and Leber Introduction Mitochondria provide critical cellular energy using the tricar- boxylic acid (TCA) cycle, oxidative phosphorylation (OXPHOS), and beta-oxidation of fatty acids for metabolism, cell division, production of reactive oxygen species (ROS), and apoptosis. Human mitochondrial (mt) DNA forms a circle of double stranded DNA with 16,569 nucleotide pairs. The non-coding mtDNA Dloop contains 1121 nucleotides and is important for replication and transcription. The coding region of mtDNA encodes for 37 genes including 13 protein subunits essential for OXPHOS, 2 ribosomal RNAs, and 22 transfer RNAs [1–3]. The mtDNA can be categorized into haplogroups that are defined by a set of PLOS ONE | www.plosone.org January 2013 | Volume 8 | Issue 1 | e54339

We previously reported an age-related decline of catalase activity and metallothionein (MT) in human retinal pigment epithelium (RPE) isolated from donor eyes. MT content and catalase activity also were shown to decrease in RPE cells cultured in zinc-deficient medium. We now sought to determine whether zinc content and superoxide dismutase activity change with age and signs of macular degenerative disease in isolated human RPE. Eyes from 57 donors were graded with respect to disease by the presence or absence of visible drusen and macular pigment changes. RPE was collected from macular and peripheral regions. Tissues from 39 donors were separated into soluble and pigment granule fractions and zinc content determined by flame atomic absorption spectroscopy. RPE from 18 separate donors was used for SOD activity analysis. Total RPE zinc was 9% lower in eyes from donors >70 years of age compared to that from donors age 70 and showed a greater decline in the soluble fraction of macular RPE in eyes with signs of macular disease. Total SOD activity increased significantly (P < .05) with donor age. We conclude that zinc content decreased dramatically in the soluble fraction of macular RPE with age and signs of macular degeneration. Total SOD activity correlated negatively with changes in zinc content. It remains to be determined whether the age-related decline in macular zinc is causal with respect to previously observed decreases in cellular MT and catalase. © 1996 Wiley-Liss, Inc.

The temporal evolution of the spectrum emitted by a two level atom irradiated by a laser pulse is obtained by means of the Gabor transform of the acceleration of the electron. This transform is useful in revealing fine structures of the spectrum which are otherwise invisible.

The goal of this study was to determine the effect of extracellular matrix components on the phagocytic function of the retinal pigment epithelial (RPE) cell. Cultured human fetal RPE cells were established in culture and plated on three commercially-prepared substrates: collagen IV, fibronectin and laminin and on three native matrices: bovine corneal endothelial cell matrix (BCEM) denuded bovine Bruch's membrane and denuded human Bruch's membrane. Cultured cells were allowed to become confluent and maintained for an additional two weeks before uptake of fluorescent bovine retinal outer segments (ROS) was measured by flow cytometry. Morphology by phase contrast microscopy and melanization was also determined as measures of differentiation. The results showed that morphology, melanization and ROS uptake by cells on collagen IV, laminin and fibronectin were not different from control cells plated on tissue culture plastic. However, ROS uptake by cells plated on BCEM was significantly less than that of cells cultured on plastic and melanization was greater. ROS uptake by cells plated on both types of Bruch's membrane was also significantly less than control cells. Treatment of cells plated on tissue culture plastic with 44 mM NaHCO3, which increased melanization, also reduced ROS uptake. We conclude that native matrices seem to contain components that significantly depress ROS uptake in culture. The inhibition is not mimicked by collagen, laminin or fibronectin coated wells. The ECM may play a significant role in controlling phagocytosis of ROS either by determining morphology, increasing differentiation or by directly influencing intracellular metabolism, and thus serve as another level of control for this RPE function which may not occur in cells plated on tissue culture plastic. These results may also have implications for the effects of aging or disease in which there are changes in the ECM.

An investigation was undertaken to determine the toxicity of an intravitreal injection of a novel peptide drug, Shiva-1, in rabbits. The drug, a synthetic peptide modeled after lytic peptides secreted by certain insects, has antiproliferative and antibacterial properties. Initial in vitro experiments showed that the drug, at a concentration of 100 microM, was toxic to both Y-79 retinoblastoma cells and human retinal pigment epithelial cells. A wide range of doses (6-1200 micrograms) was injected into the rabbit vitreous in an attempt to determine the maximum tolerated dose. Retinal toxicity was evaluated clinically, by electroretinography, and by light microscopy. Some localized toxicity was evident at 200 micrograms; all doses of 240 micrograms and above were toxic. While the drug appears to exhibit a narrow range between effective and toxic doses, the results suggest that this and other peptides of similar design merit further investigation for the treatment of proliferative and infectious diseases of the eye.

This study investigates the mechanism by which insulin stimulates phagocytosis in cultured human retinal pigment epithelium, using a flow cytometric assay of retinal outer segment uptake. RPE cells were isolated from adult human donors and grown in culture. Retinal outer segments were isolated from fresh bovine eyes and covalently labeled with the fluorescent dye carboxy-SNAFL-2. Retinal outer segment binding and uptake was quantified by direct visualization and the increase in cellular fluorescence measured using a flow cytometer and the methods compared. Uptake measured by flow cytometry was further characterized by concentration, time, temperature and serum stimulation, and shown to be comparable to other published methods. Uptake of retinal outer segments was acutely stimulated by insulin in the absence or presence of serum with half maximal stimulation occurring between 0.1 and 1.0 microgram ml-1. Theophylline, forskolin and cholera toxin all reduced retinal outer segment uptake by RPE cells, but had no effect upon insulin stimulation. Measurements of cAMP showed that insulin did not change intracellular cAMP concentration compared to controls in the absence or presence of added retinal outer segments. One hundred nanomolar okadaic acid also inhibited retinal outer segment uptake but not insulin-stimulated uptake, nor did 100 microM genistein have an effect upon insulin-stimulated uptake. Preincubation of RPE cells with 25 microM ZnCl2 overnight stimulated retinal outer segment uptake and appeared to inhibit the insulin stimulation. Preincubation of the cells in 25 mM glucose overnight also increased the uptake of retinal outer segments over control and reduced the effect of insulin. We conclude that insulin stimulates retinal outer segment phagocytosis by an as yet unknown process which may involve specific tyrosine phosphatases.

To investigate the nature of the oxidative event that occurs during phagocytosis of retinal outer segments (ROS) by cultured human retinal pigment epithelial (RPE) cells, cells were incubated with isolated bovine ROS labeled with either the fluorescence probe carboxy-SNAFL-2 or the nonfluorescent, oxidizable probe 2',7'-dichlorodihydrofluorescein (H2DCF). The increase in fluorescence following phagocytosis was measured by a flow cytometer. Other measurements included: oxygen consumption using a Clark-type oxygen electrode, extracellular superoxide release by superoxide dismutase inhibitable lucigenin chemiluminescence, intracellular hydrogen peroxide (H2O2) production, and the effect of catalase inhibition on cellular thiobarbituric acid-reactive substances (TBARS) caused by phagocytosis. The activities of the enzymes NADPH oxidase and palmitoyl-CoA oxidase were also measured. H2DCF attached to bovine ROS was oxidized during phagocytosis with a time course suggesting oxidation subsequent to ROS uptake. Measurements of oxygen consumption showed a time-dependent increase of 10%, 4 h after ROS feeding, attributable to a doubling of the cyanide-resistant oxygen consumption. Intracellular H2O2 production also doubled 4 h after ROS phagocytosis. ROS uptake by RPE cells produced no significant extracellular superoxide, while extracellular superoxide production was readily demonstrated in a control macrophage cell line. Enzyme activity measurements showed that incubation of RPE cells with ROS doubled catalase activity without affecting superoxide dismutase or glutathione peroxidase activities. Inhibition of catalase during ROS uptake increased TBARS by 66%. Other enzyme activity measurements showed that human RPE cells possess both NADPH oxidase and palmitoyl-CoA oxidase activities. We conclude that ROS phagocytosis subjects RPE cells to an oxidative event on the same order of magnitude as measured in a macrophage. The event is not an extracellular macrophage-type respiratory burst and may be due to intracellular H2O2 resulting from an NADPH oxidase in the phagosome or from beta-oxidation of ROS lipids in peroxisomes. Irrespective of case, the enzyme catalase appears to be essential in protecting the RPE cell against reactive oxygen species produced during phagocytosis.

The harmonic spectrum emitted by a two-level atom driven by a resonant laser field is calculated. The spectrum is seen to depend on the shape of the pulse. In the case of an asymmetric pulse profile, harmonic emission is present also at a relatively low intensity field.

Grids, clouds and cloud-like infrastructures are capable of supporting a broad range of data-intensive applications. There are interesting and unique performance issues that appear as the volume of data and degree of distribution increases. New scalable data-placement and management techniques, as well as novel approaches to determine the relative placement of data and computational workload, are required. We develop and study a genome sequence matching application that is simple to control and deploy, yet serves as a prototype of a data-intensive application. The application uses a SAGA-based implementation of the All-Pairs pattern. This paper aims to understand some of the factors that influence the performance of this application and the interplay of those factors. We also demonstrate how the SAGA approach can enable data-intensive applications to be extensible and interoperable over a range of infrastructure. This capability enables us to compare and contrast two different approaches for executing distributed data-intensive applications—simple application-level data-placement heuristics versus distributed file systems.

MapReduce has emerged as an important data-parallel programming model for data-intensive computing – for Clouds and Grids. However most if not all implementations of MapReduce are coupled to a specific infrastructure. SAGA is a high-level programming interface which provides the ability to create distributed applications in an infrastructure independent way. In this paper, we show how MapReduce has been implemented using SAGA and demonstrate its interoperability across different distributed platforms – Grids, Cloud-like infrastructure and Clouds. We discuss the advantages of programmatically developing MapReduce using SAGA, by demonstrating that the SAGA-based implementation is infrastructure independent whilst still providing control over the deployment, distribution and runtime decomposition. The ability to control the distribution and placement of the computation units (workers) is critical in order to implement the ability to move computational work to the data. This is required to keep data network transfer low and in the case of commercial Clouds the monetary cost of computing the solution low. Using data-sets of size up to 10GB, and upto 10 workers, we provide detailed performance analysis of the SAGA-MapReduce implementation, and show how controllingthe distribution of computation and the payload per worker helps enhance performance.

A new approach for obtaining metasurfaces realized by employing Frequency Selective Surfaces (FSSs) with small periodicity printed on a new class of high dielectric constant materials is presented. The synergic use of ad-hoc engineered materials and innovative electromagnetic design provides an improvement of the overall device in terms of size reduction and radiative properties. Zr 0.8 Sn 0.2 TiO 4 (ZST) exhibits a high dielectric constant, low dielectric losses, good thermal stability and good high frequency characteristics. Consequently, it is one of the most promising materials for these specific purposes. In particular, in this work the microstructure and the microwave dielectric properties of Zr 0.8 Sn 0.2 TiO 4 (ZST) perovskite with NiO addition, prepared with conventional solid-state route, are investigated. The addition of NiO (0.2 wt.%) as doping can efficiently promote the densification and reduce dielectric losses of ZST ceramics. Estimates of the physical and electric characteristics, with particular attention to losses, will be presented through measurements within different range of frequencies. Up to now, ZST materials have been successfully applied in memory devices, filters in cellular base stations, miniaturization of microwave devices, duplexers and general wireless applications [1].

1 Abstract Previously the cost and extremely limited capabilities of sensors prohibited Quality of Service (QoS) implementations in wireless sensor networks. With advances in technology, sensors are becoming significantly less expensive and the increases in computational and storage capabilities are opening the door for new, sophisticated algorithms to be implemented. Newer sensor network applications require higher data rates with more stringent priority requirements. We introduce a dynamic scheduling algorithm to improve bandwidth for high priority data in sensor networks, called Tiny-DWFQ. Our Tiny-Dynamic Weighted Fair Queuing scheduling algorithm allows for dynamic QoS for prioritized communications by continually adjusting the treatment of communication packages according to their priorities and the current level of network congestion. For performance evaluation, we tested Tiny-DWFQ, Tiny-WFQ (traditional WFQ algorithm implemented in TinyOS), and FIFO queues on an Imote2-based wireless sensor network and report their throughput and packet loss. Our results show that Tiny-DWFQ performs better in all test cases.

Summary Yeast cells become older with each division, but their daughters are born young. Mutational analysis shows that maintenance of this age asymmetry requires segregation of a complement of active mitochondria to daughters and that this process breaks down in older mother cells. This decline has implications for stem cell aging in higher organisms. PEX6, a peroxisome biogenesis gene, has been isolated as a multicopy suppressor of an atp2 age asymmetry mutant. Suppression depended on the presence of particular amino acid residues in Atp2p, and required adenosine triphosphate (ATP) binding and/or ATP hydrolysis activity of Pex6p. Extra copies of PEX6 corrected the deficit in Atp2p in mitochondria in the mutant by improving its import kinetics, resulting in near normal mitochondrial inheritance by daughter cells. The novel function of Pex6p described here may provide insights into peroxisomal and mitochondrial disorders and into metabolic diseases in general.

Purpose To measure changes in nuclear gene expression resulting from mitochondrial dysfunction in retinal pigment epithelial cells. Methods ARPE-19 retinal pigment epithelial cells were depleted of their mitochondrial (mt)DNA by passaging in a low concentration of ethidium bromide. Loss of mitochondrial DNA was determined by uridine auxotrophy and quantitative real-time polymerase chain reaction of isolated DNA. Loss of mitochondrial membrane potential was estimated by uptake of JC-1. Changes in nuclear gene expression were determined by quantitative real-time reverse transcription-polymerase chain reaction of isolated total RNA from ethidium-bromide-treated and untreated cells. Morphologic and phenotypic changes were determined by phase-contrast microscopy, sensitivity to the oxidant tert-butyl hydroperoxide (tBH), and invasion assay. Results ARPE-19 cells became auxotrophic for growth on uridine after eight passages in 50 ng/mL ethidium bromide. Quantitative PCR revealed almost complete loss of mitochondrial DNA (rho(0) cells). Uptake of JC-1 was reduced in the rho(0) cells, indicating reduction of mitochondrial membrane potential. Quantitative RT-PCR measured increased expression of genes coding for drusen components, lipid transport, extracellular matrix components, and responses to inflammation in the rho(0) cells. The rho(0) cells also exhibited an increased sensitivity to killing by tBH and increased migration and invasion through solubulized basement membrane-coated tissue culture inserts. Conclusions ARPE-19 cells respond to loss of mitochondrial function by changes in nuclear gene expression that resemble changes observed in age-related macular degeneration. The results lead to the hypothesis that loss of mitochondrial function with age and resultant changes in nuclear gene expression may explain some of the changes in the macula that are associated with the known clinical manifestations of age-related macular degeneration.

The yeast Saccharomyces cerevisiae has a finite replicative life span. Yeasts possess two prohibitins, Phb1p and Phb2p, in similarity to mammalian cells. These proteins are located in the inner mitochondrial membrane, where they are involved in the processing of newly-synthesized membrane proteins. We demonstrate that the elimination of one or both of the prohibitin genes in yeast markedly diminished the replicative life span of cells that lack fully-functional mitochondria, while having no effect on cells with functioning mitochondria. This deleterious effect was suppressed by the deletion of the RAS2 gene. The expression of PHB1 and PHB2 declined gradually up to 5-fold during the life span. Cells in which PHB1 was deleted in conjunction with the absence of a mitochondrial genome displayed remarkable changes in mitochondrial morphology, distribution, and inheritance. This loss of mitochondrial integrity was not seen in cells devoid of PHB1 but possessing an intact mitochondrial genome. In a subset of the cells, the changes in mitochondrial integrity were associated with increased production of reactive oxygen species, which co-localized with the altered mitochondria. The mitochondrial deficits described above were all suppressed by deletion of RAS2. Our data, together with published information, are interpreted to provide a unified view of the role of the prohibitins in yeast aging. This model posits that the key initiating event is a decline in mitochondrial function, which leads to progressive oxidative damage that is exacerbated in the absence of the prohibitins. This aggravation of the initial damage is ameliorated by the suppression of the production of mitochondrial proteins in the absence of Ras2p signaling of mitochondrial biogenesis.

To characterise the noise behaviour of transistors, different noise equivalent circuits are used. In these circuits the noise properties are modelled as noise voltage and current generators in order to design microwave devices for low noise applications. In this paper we present a stochastic approach to study the noise modelling of bipolar microwave transistors. Starting from experimental on-wafer measurements of the scattering and noise parameters of these bipolar devices, we obtain the corresponding Giacoletto model with the noise internal sources. We write down then the stochastic differential equations of the Giacoletto model for a short-circuited output in order to study the noise properties of these devices. The probability distribution of the output current is obtained. We give the analytical temporal behaviour of the second moment of the output current, assuming particular given correlation functions between the internal noise sources. From the power spectrum of the output current we obtain the behaviour of the noise figure as a function of the operating frequency. Our theoretical results are in good agreement with experimental ones. The present approach allows us to study the statistical properties of the output current for different statistics of the internal noise sources of the bipolar transistors (BJT and HBT).

To determine the relative expression of metallothionein isoforms and their differential induction by oxidative stress in cultured RPE cells and to localize the isoforms in the human chorioretinal complex.Total RNA was isolated from cultured human retinal pigment epithelial cells using TRI-Reagent. An "anchor-oligo-dT primer" was used for the synthesis of cDNA, reverse transcribed using avian reverse transcriptase and subsequently subjected to PCR analysis using oligonucleotides specific for metallothionein (MT) I, MT II, and MT III. The selected transcripts were then used to assess the expression of the above elements in fixed tissue sections by in situ hybridization. Cultured RPE cells were allowed to phagocytose bovine photoreceptor outer segments (ROS) or were treated with H(2)O(2) for 6 hours and then analyzed by RT-PCR or in situ hybridization to ascertain the effect of oxidative stress on metallothionein mRNA isoform expression.Relative density analysis of amplified products demonstrate the presence of MT I, MT II and MT III in RPE cells, with an apparent relative expression MT IIMT IMT III [corrected]. Expression of MT I and MT II mRNA was increased by both phagocytosis and hydrogen peroxide, however MT III was not induced by either stress. In situ hybridization corroborated the findings of the RT-PCR analysis and showed that MTs were mainly localized in the RPE and the photoreceptor layer of the retina.The localization of MT and the response of MT to oxidative stress are consistent with a role for MT as an antioxidant in the RPE and retina. Studies are ongoing to determine the specific mechanisms of action of these antioxidants in RPE cells.

In this study we have gathered 86 ALTE infants' characteristics about physiological, familiar, remote and near pathological case history, modality of apnoea episode's presentation, environmental and behavioural conditions of babies and we have compared these ones with main risk factors for SIDS to find possible similarities or differences. Many of the already known differences or similarities between ALTE and SIDS have been confirmed by our data and new ones have been observed. So we can say that the association between ALTE and SIDS remains contrasting because beside undiscussed factors of association there are other ones which differentiate the two Syndromes.

The aim of this research is to value, with a questionnary for the families, epidemiologic aspects in 20 children died for SIDS. The risk factors we have considered regard: ambiental condition, pregnancy, neonatal and post-neonatal period, and modality of death. The result is that sleeping position, child's age and month in which the death happens, are still the most significant risk factors for SIDS.

To investigate if the use of a contrast agent (Levovist) improves the specificity of US in the diagnosis of prostate carcinoma, having the results of prostate biopsy as the gold standard.Thirty patients with physical findings suspicious for prostate carcinoma and PSA ranging 5 ng/mL to 15 ng/mL were examined with transrectal US (TRUS) integrated with the color Doppler mode and contrast agent administration (4 g injected at 4 mL/min: 300 mL concentration).Based on bioptic and surgical results, 14 prostate carcinomas were found, all of them1.2 cm. Conventional US recognized the suspected nodule in 11 cases, with 78% sensitivity and 93% specificity. Color Doppler showed color signals in 8/14 cases, which were peripheral to the lesion or intranodular, but did not add any important finding to those of gray-scale US. In contrast, contrast enhanced studies showed 13/14 carcinomas, which improved sensitivity significantly (92%). Particularly, 11/14 lesions had a typical avascular pattern within the strongly enhanced peripheral gland, while 2 small lesions only exhibited intranodular vessels.We compared our results with the Microscopic Angiogenesis Grading System (MAGS) index and found it exceeded 30 in the 11 avascular lesions value indicates microneoangiogenesis. In the other two cases a value30 was correlated with a different type of tumor vascularization typical of macroangiogenesis.

Feeding, deglutition, and swallowing are the most natural means of alimentation. These means provide nutrition and hydration to the body. Management of dysphagia has become a widespread clinical practice in acute care, rehabilitation, and long-term skilled nursing facilities from rehabilitation and nursing perspectives. Caring for chronic dysphagic and feeding-impaired patients on nursing units is a central concern of this article. The purpose of this descriptive article is to report on a practical meal supervision program that may be used on nursing units to facilitate the continuum of dysphagia care after skilled therapeutic intervention. This program stresses the importance of: (a) identifying patient help level and corresponding nursing care; (b) centralized, supervised dining; (c) organized seating assignments; and (d) an effective, integrated communication system. This article addresses staff training and continuous quality improvement monitoring, and discusses potential investigations concerning compliance.

We present a new approach to study the complete stochastic properties of fluctuations of the output current of microwave transistors. We obtain the π-hybrid model of bipolar microwave transistors with the noise internal sources starting from experimental on-wafer measurements of the scattering and noise parameters. We derive the stochastic differential equations of the Giacoletto model for different loads and source admittances. We give the analytical temporal behavior of the second moment of the output current, assuming particular given correlation functions between the internal noise sources.

To determine the effect of moderate zinc deficiency on antioxidant defenses and measures of oxidative stress in the retina and retinal pigment epithelium (RPE) of Brown Norway Rats.Twenty-four rats were housed individually and divided into three groups of 8 rats each. Group 1 was fed ad libitum a semipurified control diet formulated to contain 50 parts per million [ppm] total zinc; group 2 was fed ad libitum an identical diet but containing 5 ppm total zinc; and group 3 was pair-fed the control diet but restricted in amount to that consumed by group 2. Food intake was measured daily and the rats weighed weekly. After 6 weeks, the rats were killed and the following measurements were made: serum zinc, serum alkaline phosphatase, retinal zinc, RPE-choroid zinc, RPE-choroid catalase, liver metallothionein (MT), retinal MT, RPE-choroid MT, retinal catalase, and retinal thiobarbituric reactive substances (TBARS).The following showed statistically significant differences between groups 2 and 3, respectively: serum Zn (1216 micro/l versus 1555 microg/l, Por = 0.01), serum alkaline phosphatase (3.75 U/mg versus 5.10 U/mg, Por = 0.05), liver MT (4.3 microg/mg protein versus 16.7 microg/mg, Por = 0.0001), RPE-choroid MT (1.3 microg/mg protein versus 2.2 microg/mg, Por = 0.02), retinal MT (0.85 microg/mg protein versus 2.8 microg/mg, Por = 0.05), and retinal TBARS (6.2 nM/mg protein versus 2.2 nM/mg, Por = 0.05).The results show that retinal MT and RPE MT concentrations are very sensitive to intake of dietary zinc. The increase in retinal TBARS in group 2 indicates that moderate zinc deficiency increases oxidative stress to the retina. The results also suggest that MT is protective against lipid peroxidation of retinal membranes.

This study was undertaken to determine whether bioavailable zinc can influence the effects of oxidative stress on cultured human retinal pigment epithelial (RPE) cells. RPE cells were maintained for 7 d in culture medium containing 14 microM total zinc, or in medium containing 0.55 microM total zinc. After 1 week, MTT assays were performed to determine the relative cytotoxicity of H2O2 or paraquat on RPE cells. Conjugated dienes and thiobarbituric acid reactive substances (TBARS) were measured in RPE cells treated with 0, 0.5 mM H2O2, 10 microM FeSO4 + 0.5 mM H2O2 or 10 microM FeSO4 + xanthine/xanthine oxidase for 24 h or paraquat for 7 d. Oxidized proteins were determined by the formation of carbonyl residues. The antioxidants metallothionein, catalase, superoxide dismutase, and glutathione peroxidase were also measured. The MTT assays showed that zinc protected cultured RPE from the toxicity of H2O2 and paraquat. RPE cells in 0.55 microM zinc medium contained higher levels of TBARS, conjugated dienes and protein carbonyls due to the oxidative stresses, compared to cells in 14 microM zinc. Catalase and MT content were reduced in cells cultured in 0.55 microM zinc medium and were reduced additionally when treated with above stresses. Superoxide dismutase activity increased in 0.55 microM zinc medium in response to these stresses. Our results show RPE cells cultured in zinc-reduced medium are more susceptible to oxidative insult.

Numerous reports have shown metallothionein protects tissue from reactive oxygen intermediates, considered to be a factor in aging. Because zinc stimulates MT induction and because loss of metal increases proteolysis of the apoprotein, zinc may have a protective role and may indirectly protect against aging. Our laboratory has shown age-related decreases in catalase, zinc and metallothionein in human retinal pigment epithelium. This study was undertaken to investigate the role of bioavailable zinc on MT content in RPE cells subjected to an oxidative stress.

To examine the internalization of liposomes containing various phospholipids by human retinal pigment epithelial (RPE) cells in culture. The internalization was compared to that exhibited by macrophages and fibroblast cells.The uptake into cells was monitored by using a non-degradable, radioactive cholesterol-hexadecyl ether derivative and a pH-sensitive, water-soluble fluorescent dye. Variables tested to determine their effect on uptake included time, phospholipid concentration, the presence or absence of serum in medium and the presence or absence of cholesterol in the liposome bilayer. The most avidly ingested liposome was tested to determine its ability to deliver a liposome-dependent drug.Liposome uptake was time and concentration dependent. Uptake for RPE was maximal in serum-free media, while the opposite was true for the other cells tested. Cell survival following exposure to fluoroorotic acid-containing liposomes correlated with the radioactive cholesterol-hexadecyl ether and fluorescent dye uptake data.No single liposome carrier or set of conditions tested appeared to be optimum for the delivery of a cytotoxic agent to these three cell types. This bears consideration when designing strategies for the prevention and treatment of intraocular proliferative diseases such as proliferative vitreoretinopathy (PVR).

To examine whether the vitronectin (VN) in serum is responsible for the serum stimulation of phagocytosis in the rod outer segment (ROS) by cultured retinal pigment epithelial (RPE) cells.Vitronectin was removed from fetal bovine serum by heparin-agarose affinity chromatography. Concentrations in normal and depleted serum were determined by enzyme-linked immunosorbent assay, using a polyclonal antibody against bovine VN and commercially prepared human VN as a standard. A monoclonal antibody against human alpha v beta 5 was used in localization and in blocking experiments. Rod outer segment phagocytosis was measured using a flow cytometric assay.Affinity chromatography removed 95% of the VN from serum as determined by enzyme-linked immunosorbent assay. Vitronectin-depleted serum did not stimulate ROS phagocytosis by RPE cells. Commercially prepared VN added to serum-free medium stimulated ROS phagocytosis in a dose-dependent manner. Pretreatment of RPE cells with an antibody against alpha v beta 5, an integrin receptor for VN, had no effect on phagocytosis in the absence of serum but completely blocked the serum stimulation of ROS phagocytosis. Antibody against alpha v beta 5 demonstrated a variable labeling pattern on the cultured RPE cell surface with morphologically distinct cell clusters exhibiting less labeling. Those cell clusters exhibiting less receptor labeling also showed less uptake of fluorescent-labeled ROS.Vitronectin is the component responsible for serum stimulation of ROS uptake, and this uptake appears to be mediated by an alpha v beta 5 integrin. Although clearly important in vitro, a role for VN in ROS uptake by RPE cells in situ remains to be determined.