Pediatrics Faculty Papers

Permanent URI for this collectionhttps://digital.lib.washington.edu/handle/1773/11650

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CODEHOP-mediated PCR - A powerful technique for the identification and characterization of viral genomes
(2005) Rose, Timothy M.
Consensus-Degenerate Hybrid Oligonucleotide Primer (CODEHOP) PCR primers derived from amino acid sequence motifs which are highly conserved between members of a protein family have proven to be highly effective in the identification and characterization of distantly related family members. Here, the use of the CODEHOP strategy to identify novel viruses and obtain sequence information for phylogenetic characterization, gene structure determination and genome analysis is reviewed. While this review describes techniques for the identification of members of the herpesvirus family of DNA viruses, the same methodology and approach is applicable to other virus families.
Development of a real-time QPCR assay for the detection of RV2 lineage-specific rhadinoviruses in macaques and baboons
(2005) Bruce, A Gregory; Bakke, Angela M.; Thouless, Margaret E.; Rose, Timothy M.
Background: Two distinct lineages of rhadinoviruses related to Kaposi's sarcoma-associated herpesvirus (KSHV/HHV8) have been identified in macaques and other Old World non-human primates. We have developed a real-time quantitative PCR (QPCR) assay using a TaqMan probe to differentially detect and quantitate members of the rhadinovirus-2 (RV2) lineage. PCR primers were derived from sequences within ORF 60 and the adjacent ORF 59/60 intergenic region which were highly conserved between the macaque RV2 rhadinoviruses, rhesus rhadinovirus (RRV) and Macaca nemestrina rhadinovirus-2 (MneRV2). These primers showed little similarity to the corresponding sequences of the macaque RV1 rhadinoviruses, retroperitoneal fibromatosis herpesvirus Macaca nemestrina (RFHVMn) and Macaca mulatta (RFHVMm). To determine viral loads per cell, an additional TaqMan QPCR assay was developed to detect the single copy cellular oncostatin M gene. Results: We show that the RV2 QPCR assay is linear from less than 2 to more than 300,000 copies using MneRV2 DNA, and is non-reactive with RFHVMn DNA up to 1 billion DNA templates per reaction. RV2 loads ranging from 6 to 2,300 viral genome equivalent copies per 106 cells were detected in PBMC from randomly sampled macaques from the Washington National Primate Research Center. Screening tissue from other primate species, including another macaque, Macaca fascicularis, and a baboon, Papio cynocephalus, revealed the presence of novel rhadinoviruses, MfaRV2 and PcyRV2, respectively. Sequence comparison and phylogenetic analysis confirmed their inclusion within the RV2 lineage of KSHV-like rhadinoviruses. Conclusions: We describe a QPCR assay which provides a quick and sensitive method for quantitating rhadinoviruses belonging to the RV2 lineage of KSHV-like rhadinoviruses found in a variety of macaque species commonly used for biomedical research. While this assay broadly detects different RV2 rhadinovirus species, it is unreactive with RV1 rhadinovirus species which commonly co-infect the same primate hosts. We also show that this QPCR assay can be used to identify novel RV2 rhadinoviruses in different primate species.
Hospital care for children and young adults in the last year of life: a population-based study
(2003) Feudtner, Chris; DiGiuseppe, David L.; Neff, Jeff M.
Background: To help design population-based pediatric palliative care services, we sought to describe the hospital care received in the last year of life by children and young adults who died. We also determined the proportion with complex chronic conditions (CCCs) and tested whether the use of hospital services increased as the date of death drew nearer. Methods: For all deaths occurring under 25 years of age from 1990 to 1996 in Washington State, USA, we linked death certificate information to hospital utilization records and analyzed the timing and duration of hospitalizations and the nature of hospital procedures during the year prior to death. Results: Of the 8 893 deaths, 25 % had CCCs. Among infants with CCCs, 84 % were hospitalized at the time of death and 50 % had been mechanically ventilated during their terminal admission. Among the 458 CCC neonates dying under a week of age, 92% of all days of life were spent in the hospital; among the 172 CCC neonates dying during the second to fourth weeks of life, 85 % of all days of life were spent hospitalized; among the 286 CCC infants dying during the second to twelfth month of life, 41 % of all days of life were spent hospitalized. Among children and young adults with CCCs, 55 % were hospitalized at the time of death, and 19 % had been mechanically ventilated during their terminal admission. For these older patients, the median number of days spent in the hospital during the year preceding death was 18, yet less than a third of this group was hospitalized at any point in time until the last week of their lives. The rate of hospital use increased as death drew near. For subjects who had received hospital care, 44 % had governmental insurance as the source of primary payment. Conclusions: Infants who died spent a substantial proportion of their lives in hospitals, whereas children and adolescents who died from CCCs predominantly lived outside of the hospital during the last year of life. To serve these patients, pediatric palliative and end-of-life care will have to be provided in an integrated, coordinated manner both in hospitals and home communities.
Immigrant families' perceptions on walking to school and school breakfast: a focus group study
(2007) Greves, H. Mollie; Lozano, Paula; Liu, Lenna; Busby, Katie; Cole, Jen; Johnston, Brian
Background: Immigrant children face an increased risk of being overweight. Little is known about how immigrant families perceive school programs that may help prevent obesity, such as walking to school and school breakfast. Methods: Six focus groups (n = 53) were conducted with immigrant parents of school-aged children, two each in three languages: Vietnamese, Spanish, and Somali. A facilitator and translator conducted the focus groups using a script and question guide. Written notes and audio transcripts were recorded in each group. Transcripts were coded for themes by two researchers and findings classified according to an ecological model. Results: Participants in each ethnic group held positive beliefs about the benefits of walking and eating breakfast. Barriers to walking to school included fear of children's safety due to stranger abductions, distrust of neighbors, and traffic, and feasibility barriers due to distance to schools, parent work constraints, and large families with multiple children. Barriers to school breakfast participation included concerns children would not eat due to lack of appealing/appropriate foods and missing breakfast due to late bus arrival or lack of reminders. Although some parents acknowledged concerns about child and adult obesity overall, obesity concerns did not seem personally relevant. Conclusion: Immigrant parents supported the ideals of walking to school and eating breakfast, but identified barriers to participation in school programs across domains of the ecological model, including community, institution, and built environment factors. Schools and communities serving immigrant families may need to address these barriers in order to engage parents and children in walking and breakfast programs.
Report card on school snack food policies among the United States' largest school districts in 2004�2005: Room for improvement
(2006) Greves, H. Mollie; Rivara, Frederick P.
Background: Federal nutritional guidelines apply to school foods provided through the national school lunch and breakfast programs, but few federal regulations apply to other foods and drinks sold in schools (labeled "competitive foods"), which are often high in calories, fat and sugar. Competitive food policies among school districts are increasingly viewed as an important modifiable factor in the school nutrition environment, particularly to address rising rates of childhood overweight. Congress passed legislation in 2004 requiring all school districts to develop a Wellness Policy that includes nutrition guidelines for competitive foods starting in 2006-2007. In addition, the Institute of Medicine (IOM) recently published recommendations for schools to address childhood obesity. Methods: Representatives of school districts with the largest student enrollment in each state and D.C. (N = 51) were interviewed in October-November 2004 about each school district's nutrition policies on "competitive foods." District policies were examined and compared to the Institute of Medicine's recommendations for schools to address childhood obesity. Information about state competitive food policies was accessed via the Internet, and through state and district contacts. Results: The 51 districts accounted for 5.9 million students, representing 11% of US students. Nineteen of the 51 districts (39%) had competitive food policies beyond state or federal requirements. The majority of these district policies (79%) were adopted since 2002. School district policies varied in scope and requirements. Ten districts (53%) set different standards by grade level. Most district policies had criteria for food and beverage content (74%) and prohibited the sale of soda in all schools (63%); fewer policies restricted portion size of foods (53%) or beverages (47%). Restrictions more often applied to vending machines (95%), cafeteria � la carte (79%), and student stores (79%) than fundraising activities (47%). Most of the policies did not address more comprehensive approaches to the school nutrition environment, such as nutrition education (32%) or advertising to students (26%), nor did they include guidelines on physical education (11%). In addition, few policies addressed monitoring (32%) or consequences for non-compliance (11%). No policy restricted foods sold for after-school fundraising or required monitoring physical health indicators (e.g. BMI). Conclusion: When compared to the Institute of Medicine's recommendations for schools' role in preventing obesity, none of the nutrition policies among each state's largest school district had addressed all the recommendations by 2004�2005. Nutritionists, nurses, pediatricians, parents, and others concerned about child health have an unprecedented opportunity to help shape and implement more comprehensive school district nutrition policies as part of the Congressional requirement for a "Wellness Policy" by 2006-2007.
Television viewing, computer use, obesity, and adiposity in US preschool children
(2007) Mendoza, Jason A.; Zimmerman, Fred J.; Christakis, Dimitri A.
Background: There is limited evidence in preschool children linking media use, such as television/video viewing and computer use, to obesity and adiposity. We tested three hypotheses in preschool children: 1) that watching > 2 hours of TV/videos daily is associated with obesity and adiposity, 2) that computer use is associated with obesity and adiposity, and 3) that > 2 hours of media use daily is associated with obesity and adiposity. Methods: We conducted a cross-sectional study using nationally representative data on children, aged 2�5 years from the National Health and Nutrition Examination Survey, 1999-2002. Our main outcome measures were 1) weight status: normal versus overweight or at risk for overweight, and 2) adiposity: the sum of subscapular and triceps skinfolds (mm). Our main exposures were TV/video viewing (= 2 or > 2 hours/day), computer use (users versus non-users), and media use (= 2 or > 2 hours/day). We used multivariate Poisson and linear regression analyses, adjusting for demographic covariates, to test the independent association between TV/video viewing, computer use, or overall media use and a child's weight status or adiposity. Results: Watching > 2 hours/day of TV/videos was associated with being overweight or at risk for overweight (Prevalence ratio = 1.34, 95% CI [1.07, 1.66]; n =1340) and with higher skinfold thicknesses ([Beta] = 1.08, 95% CI [0.19, 1.96]; n = 1337). Computer use > 0 hours/day was associated with higher skinfold thicknesses ([Beta] = 0.56, 95% CI [0.04, 1.07]; n = 1339). Media use had borderline significance with higher skinfold thicknesses ([Beta] = 0.85, 95% CI [-0.04, 1.75], P=0.06; n = 1334) Conclusion: Watching > 2 hours/day of TV/videos in US preschool-age children was associated with a higher risk of being overweight or at risk for overweight and higher adiposity-findings in support of national guidelines to limit preschool children's media use. Computer use was also related to higher adiposity in preschool children, but not weight status. Intervention studies to limit preschool children's media use are warranted.
CTLA4Ig-mediated blockade of T-cell costimulation in patients with psoriasis vulgaris
(1999) Abrams, Judith R.; Lebwohl. Mark G.; Guzzo, Cynthia A.; Jegasothy. Brian V.; Goldfarb, Michael T.; Goffe, Bernard S.; Menter, Alan; Lowe, Nicholas J.; Krueger, Gerald; Brown, Michael J.; Weiner, Russell S.; Birkhofer, Martin J.; Warner, Garvin L.; Berry, Karen K.; Linsley, Peter S.; Krueger, James G.; Ochs, Hans D.; Kelley, Susan L.; Kang, Sewon
Engagement of the B7 family of molecules on antigen-presenting cells with their T cell–associated ligands, CD28 and CD152 (cytotoxic T lymphocyte–associated antigen-4 [CTLA-4]), provides a pivotal costimulatory signal in T-cell activation. We investigated the role of the CD28/CD152 pathway in psoriasis in a 26-week, phase I, open-label dose-escalation study. The importance of this pathway in the generation of humoral immune responses to T cell–dependent neoantigens, bacteriophage φX174 and keyhole limpet hemocyanin, was also evaluated. Forty-three patients with stable psoriasis vulgaris received 4 infusions of the soluble chimeric protein CTLA4Ig (BMS-188667). Forty-six percent of all study patients achieved a 50% or greater sustained improvement in clinical disease activity, with progressively greater effects observed in the highest-dosing cohorts. Improvement in these patients was associated with quantitative reduction in epidermal hyperplasia, which correlated with quantitative reduction in skin-infiltrating T cells. No markedly increased rate of intralesional T-cell apoptosis was identified, suggesting that the decreased number of lesional T cells was probably likely attributable to an inhibition of T-cell proliferation, T-cell recruitment, and/or apoptosis of antigen-specific T cells at extralesional sites. Altered antibody responses to T cell–dependent neoantigens were observed, but immunologic tolerance to these antigens was not demonstrated. This study illustrates the importance of the CD28/CD152 pathway in the pathogenesis of psoriasis and suggests a potential therapeutic use for this novel immunomodulatory approach in an array of T cell–mediated diseases.

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