Background

Many interventions aiming to increase children’s physical activity have been developed and implemented in a variety of settings, and these interventions have previously been reviewed; however the focus of these reviews tends to be on the intervention effects on physical activity outcomes without consideration of the reasons and pathways leading to intervention success or otherwise.

To systematically review the efficacy of physical activity interventions targeting 5-12 year old children on potential mediators and, where possible, to calculate the size of the intervention effect on the potential mediator.

Methods

A systematic search identified intervention studies that reported outcomes on potential mediators of physical activity among 5-12 year old children. Original research articles published between 1985 and April 2012 were reviewed.

Results

Eighteen potential mediators were identified from 31 studies. Positive effects on cognitive/psychological potential mediators were reported in 15 out of 31 studies. Positive effects on social environmental potential mediators were reported in three out of seven studies, and no effects on the physical environment were reported. Although no studies were identified that performed a mediating analysis, 33 positive intervention effects were found on targeted potential mediators (with effect sizes ranging from small to large) and 73% of the time a positive effect on the physical activity outcome was reported.

Conclusions

Many studies have reported null intervention effects on potential mediators of children’s physical activity; however, it is important that intervention studies statistically examine the mediating effects of interventions so the most effective strategies can be implemented in future programs.

Background

Despite clear evidence regarding how social determinants of health and structural inequities shape health, Aboriginal women’s birth outcomes are not adequately understood as arising from the historical, economic and social circumstances of their lives. The purpose of this study was to understand rural Aboriginal women’s experiences of maternity care and factors shaping those experiences.

Methods

Aboriginal women from the Nuxalk, Haida and 'Namgis First Nations and academics from the University of British Columbia in nursing, medicine and counselling psychology used ethnographic methods within a participatory action research framework. We interviewed over 100 women, and involved additional community members through interviews and community meetings. Data were analyzed within each community and across communities.

Results

Most participants described distressing experiences during pregnancy and birthing as they grappled with diminishing local maternity care choices, racism and challenging economic circumstances. Rural Aboriginal women’s birthing experiences are shaped by the intersections among rural circumstances, the effects of historical and ongoing colonization, and concurrent efforts toward self-determination and more vibrant cultures and communities.

Conclusion

Women’s experiences and birth outcomes could be significantly improved if health care providers learned about and accounted for Aboriginal people’s varied encounters with historical and ongoing colonization that unequivocally shapes health and health care. Practitioners who better understand Aboriginal women’s birth outcomes in context can better care in every interaction, particularly by enhancing women’s power, choice, and control over their experiences. Efforts to improve maternity care that account for the social and historical production of health inequities are crucial.

Alterations in the mitochondrial genome have been chronicled in most solid tumors, including breast cancer. The intent of this paper is to compare and document somatic mitochondrial D-loop mutations in paired samples of ductal carcinoma in situ (DCIS) and invasive breast cancer (IBC) indicating a potential breast ductal epithelial cancerization field effect. Paired samples of these histopathologies were laser-captured microdissected (LCM) from biopsy, lumpectomy, and mastectomy tissues. Blood samples were collected as germplasm control references. For each patient, hypervariable region 1 (HV1) in the D-loop portion of the mitochondrial genome (mtGenome) was sequenced for all 3 clinical samples. Specific parallel somatic heteroplasmic alterations between these histopathologies, particularly at sites 16189, 16223, 16224, 16270, and 16291, suggest the presence of an epithelial, mitochondrial cancerization field effect. These results indicate that further characterization of the mutational pathway of DCIS and IBC may help establish the invasive potential of DCIS. Moreover, this paper indicates that biofluids with low cellularity, such as nipple aspirate fluid and/or ductal lavage, warrant further investigation as early and minimally invasive detection mediums of a cancerization field effect within breast tissue.

Human embryonic stem cells can differentiate into CD34+ hematopoietic progenitors by co-culture on murine feeders such as OP9 and S17. These CD34+ progenitors can be further differentiated into several cells of the hematopoietic lineage including macrophages. However, co-culture on murine feeders is time consuming and involves extensive manipulations. Furthermore, CD45 expression is low on hematopoietic cultures derived from stromal co-cultures. In this study we describe a novel and highly efficient system of generating differentiated macrophages from hematopoietic progenitors generated from embryoid body cultures of human embryonic stem cells. The hematopoietic progenitors generated from these embryoid bodies express higher numbers of CD45+ cells and are able to differentiate to macrophages when cultured in presence of cytokines. Using this system we were able to generate higher yields of CD14+ macrophages compared to traditional stromal cell culture methods. The embryoid body derived macrophages are phagocytic, respond to Toll-like receptor stimulation and express phenotypic markers of mature macrophages. Importantly, the embryoid body system generates hematopoietic progenitors suitable for clinical use by eliminating the need for murine feeder cells. Furthermore, this system is amenable to genetic manipulation and may thus be used to study important mechanisms of macrophage differentiation and function.

Background

Obstetrical interventions, including caesarean sections, are increasing in Canada. Canadian women’s psychological states, fatigue, and sleep have not been examined prospectively for contributions to obstetric interventions and adverse neonatal outcomes.

Context and purpose of the study: The prospective study was conducted in British Columbia (BC), Canada with 650 low-risk pregnant women. Of those women, 624 were included in this study. Women were recruited through providers’ offices, media, posters, and pregnancy fairs. We examined associations between pregnant women’s fatigue, sleep deprivation, and psychological states (anxiety and childbirth fear) and women’s exposure to obstetrical interventions and adverse neonatal outcomes (preterm, admission to NICU, low APGARS, and low birth weight).

Methods

Data from our cross-sectional survey were linked, using women’s personal health numbers, to birth outcomes from the Perinatal Services BC database. After stratifying for parity, we used Pearson’s Chi-square to examine associations between psychological states, fatigue, sleep deprivation and maternal characteristics. We used hierarchical logistic regression modeling to test 9 hypotheses comparing women with high and low childbirth fear and anxiety on likelihood of having epidural anaesthetic, a caesarean section (stratified for parity), assisted vaginal delivery, and adverse neonatal outcomes and women with and without sleep deprivation and high levels of fatigue on likelihood of giving birth by caesarean section, while controlling for maternal, obstetrical (e.g., infant macrosomia), and psychological variables.

Results

Significantly higher proportions of multiparas, reporting difficult and upsetting labours and births, expectations of childbirth interventions, and health stressors, reported high levels of childbirth fear. Women who reported antenatal relationship, housing, financial, and health stressors and multiparas reporting low family incomes were significantly more likely to report high anxiety levels. The hypothesis that high childbirth fear significantly increased the risk of using epidural anaesthesia was supported.

Conclusions

Controlling for some psychological states and sleep quality while examining other contributors to outcomes decreases the likelihood of linking childbirth fear anxiety, sleep deprivation, and fatigue to increased odds of caesarean section. Ameliorating women’s childbirth fear to reduce their exposure to epidural anaesthesia can occur through developing effective interventions. These include helping multiparous women process previous experiences of difficult and upsetting labour and birth.

The impact of human activity on the selection for antibiotic resistance in the environment is largely unknown, although considerable amounts of antibiotics are introduced through domestic wastewater and farm animal waste. Selection for resistance may occur by exposure to antibiotic residues or by co-selection for mobile genetic elements (MGEs) which carry genes of varying activity. Class 1 integrons are genetic elements that carry antibiotic and quaternary ammonium compound (QAC) resistance genes that confer resistance to detergents and biocides. This study aimed to investigate the prevalence and diversity of class 1 integron and integron-associated QAC resistance genes in bacteria associated with industrial waste, sewage sludge and pig slurry. We show that prevalence of class 1 integrons is higher in bacteria exposed to detergents and/or antibiotic residues, specifically in sewage sludge and pig slurry compared with agricultural soils to which these waste products are amended. We also show that QAC resistance genes are more prevalent in the presence of detergents. Studies of class 1 integron prevalence in sewage sludge amended soil showed measurable differences compared with controls. Insertion sequence elements were discovered in integrons from QAC contaminated sediment, acting as powerful promoters likely to upregulate cassette gene expression. On the basis of this data, >1 × 1019 bacteria carrying class 1 integrons enter the United Kingdom environment by disposal of sewage sludge each year.

It is important to recognise that migraine is a ‘biological’ and not a ‘psychological’ entity. However, psychological factors can be involved in migraine in 4 different ways:- 1) Migraines can be triggered by psychological stressors; 2) Severe migraine can itself be a cause of significant psychological stress which can, in turn, exacerbate the problem; 3) Even if psychological stress is not significantly involved in the genesis of the headache, pain management techniques can help people cope with their pain more effectively; 4) Longitudinal data demonstrate a complex bidirectional association between mood disorders and migraine. Treatment of a co-existing mood disorder, for example with cognitive behavioural techniques, may therefore reduce the impact of migraine. It would thus appear logical to view medical and psychological approaches as potentially synergistic rather than mutually exclusive. Functional imaging indicates that cognition, emotions, and pain experiences change the way the brain processes pain inputs. This may provide a physiological rationale for psychological interventions in pain management. As most studies of psychological management of migraine have been relatively small and the approach often varies between clinicians, the magnitude of benefit, optimum method of delivery, and the length of intervention are uncertain.

The design of adaptation strategies that promote urban health and well-being in the face of climate change requires an understanding of the feedback interactions that take place between the dynamical state of a city, the health of its people, and the state of the planet. Complexity, contingency and uncertainty combine to impede the growth of such systemic understandings. In this paper we suggest that the collaborative development of conceptual models can help a group to identify potential leverage points for effective adaptation. We describe a three-step procedure that leads from the development of a high-level system template, through the selection of a problem space that contains one or more of the group’s adaptive challenges, to a specific conceptual model of a sub-system of importance to the group. This procedure is illustrated by a case study of urban dwellers’ maladaptive dependence on private motor vehicles. We conclude that a system dynamics approach, revolving around the collaborative construction of a set of conceptual models, can help communities to improve their adaptive capacity, and so better meet the challenge of maintaining, and even improving, urban health in the face of climate change.

Background

Physical activity (PA) is associated with positive cardio-metabolic health and emerging evidence suggests sedentary behavior (SB) may be detrimental to children's health independent of PA. The primary aim of the Transform-Us! study is to determine whether an 18-month, behavioral and environmental intervention in the school and family settings results in higher levels of PA and lower rates of SB among 8-9 year old children compared with usual practice (post-intervention and 12-months follow-up). The secondary aims are to determine the independent and combined effects of PA and SB on children's cardio-metabolic health risk factors; identify the factors that mediate the success of the intervention; and determine whether the intervention is cost-effective.

Methods/design

A four-arm cluster-randomized controlled trial (RCT) with a 2 × 2 factorial design, with schools as the unit of randomization. Twenty schools will be allocated to one of four intervention groups, sedentary behavior (SB-I), physical activity (PA-I), combined SB and PA (SB+PA-I) or current practice control (C), which will be evaluated among approximately 600 children aged 8-9 years in school year 3 living in Melbourne, Australia. All children in year 3 at intervention schools in 2010 (8-9 years) will receive the intervention over an 18-month period with a maintenance 'booster' delivered in 2012 and children at all schools will be invited to participate in the evaluation assessments. To maximize the sample and to capture new students arriving at intervention and control schools, recruitment will be on-going up to the post-intervention time point. Primary outcomes are time spent sitting and in PA assessed via accelerometers and inclinometers and survey.

Discussion

To our knowledge, Transform-Us! is the first RCT to examine the effectiveness of intervention strategies for reducing children's overall sedentary time, promoting PA and optimizing health outcomes. The integration of consistent strategies and messages to children from teachers and parents in both school and family settings is a critical component of this study, and if shown to be effective, may have a significant impact on educational policies as well as on pedagogical and parenting practices.

Trial registration

ACTRN12609000715279; Current Controlled Trials ISRCTN83725066

Background

The transcription factor MYC is a critical regulator of diverse cellular processes, including both replication and apoptosis. Differences in MYC-regulated gene expression responsible for such opposing outcomes in vivo remain obscure. To address this we have examined time-dependent changes in global gene expression in two transgenic mouse models in which MYC activation, in either skin suprabasal keratinocytes or pancreatic islet β-cells, promotes tissue expansion or involution, respectively.

Results

Consistent with observed phenotypes, expression of cell cycle genes is increased in both models (albeit enriched in β-cells), as are those involved in cell growth and metabolism, while expression of genes involved in cell differentiation is down-regulated. However, in β-cells, which unlike suprabasal keratinocytes undergo prominent apoptosis from 24 hours, there is up-regulation of genes associated with DNA-damage response and intrinsic apoptotic pathways, including Atr, Arf, Bax and Cycs. In striking contrast, this is not the case for suprabasal keratinocytes, where pro-apoptotic genes such as Noxa are down-regulated and key anti-apoptotic pathways (such as Igf1-Akt) and those promoting angiogenesis are up-regulated. Moreover, dramatic up-regulation of steroid hormone-regulated Kallikrein serine protease family members in suprabasal keratinocytes alone could further enhance local Igf1 actions, such as through proteolysis of Igf1 binding proteins.

Conclusions

Activation of MYC causes cell growth, loss of differentiation and cell cycle entry in both β-cells and suprabasal keratinocytes in vivo. Apoptosis, which is confined to β-cells, may involve a combination of a DNA-damage response and downstream activation of pro-apoptotic signalling pathways, including Cdc2a and p19Arf/p53, and downstream targets. Conversely, avoidance of apoptosis in suprabasal keratinocytes may result primarily from the activation of key anti-apoptotic signalling pathways, particularly Igf1-Akt, and induction of an angiogenic response, though intrinsic resistance to induction of p19Arf by MYC in suprabasal keratinocytes may contribute.

Selumetinib (AZD6244, ARRY-142886) is a selective, non–ATP-competitive inhibitor of mitogen-activated protein/extracellular signal–regulated kinase kinase (MEK)-1/2. The range of antitumor activity seen preclinically and in patients highlights the importance of identifying determinants of response to this drug. In large tumor cell panels of diverse lineage, we show that MEK inhibitor response does not have an absolute correlation with mutational or phospho-protein markers of BRAF/MEK, RAS, or phosphoinositide 3-kinase (PI3K) activity. We aimed to enhance predictivity by measuring pathway output through coregulated gene networks displaying differential mRNA expression exclusive to resistant cell subsets and correlated to mutational or dynamic pathway activity. We discovered an 18-gene signature enabling measurement of MEK functional output independent of tumor genotype. Where the MEK pathway is activated but the cells remain resistant to selumetinib, we identified a 13-gene signature that implicates the existence of compensatory signaling from RAS effectors other than PI3K. The ability of these signatures to stratify samples according to functional activation of MEK and/or selumetinib sensitivity was shown in multiple independent melanoma, colon, breast, and lung tumor cell lines and in xenograft models. Furthermore, we were able to measure these signatures in fixed archival melanoma tumor samples using a single RT-qPCR–based test and found intergene correlations and associations with genetic markers of pathway activity to be preserved. These signatures offer useful tools for the study of MEK biology and clinical application of MEK inhibitors, and the novel approaches taken may benefit other targeted therapies.

Cellular signalling pathways are critical in regulating the balance between latency and lytic replication of herpesviruses. Here, we investigated the effect of the phosphatidylinositol 3-kinase (PI3K)-Akt pathway on replication of two gamma-2 herpesviruses, murine gammaherpesvirus-68 (MHV-68) and human herpesvirus-8/Kaposi's sarcoma-associated herpesvirus (HHV-8/KSHV). We found that de novo infection of MHV-68 induced PI3K-dependent Akt activation and the lytic replication of MHV-68 was enhanced by inhibiting the PI3K-Akt pathway with both chemical inhibitors and RNA interference technology. Inhibiting the activity of Akt using Akt inhibitor VIII also facilitated the reactivation of KSHV from latency. Both lytic replication and latency depend on the activity of viral transactivator RTA and we further show that the activity of RTA is increased by reducing Akt1 expression. The data suggest that the PI3K-Akt pathway suppresses the activity of RTA and thereby contributes to the maintenance of viral latency and promotes tumorigenesis.

Lavender Foal Syndrome (LFS) is a lethal inherited disease of horses with a suspected autosomal recessive mode of inheritance. LFS has been primarily diagnosed in a subgroup of the Arabian breed, the Egyptian Arabian horse. The condition is characterized by multiple neurological abnormalities and a dilute coat color. Candidate genes based on comparative phenotypes in mice and humans include the ras-associated protein RAB27a (RAB27A) and myosin Va (MYO5A). Here we report mapping of the locus responsible for LFS using a small set of 36 horses segregating for LFS. These horses were genotyped using a newly available single nucleotide polymorphism (SNP) chip containing 56,402 discriminatory elements. The whole genome scan identified an associated region containing these two functional candidate genes. Exon sequencing of the MYO5A gene from an affected foal revealed a single base deletion in exon 30 that changes the reading frame and introduces a premature stop codon. A PCR–based Restriction Fragment Length Polymorphism (PCR–RFLP) assay was designed and used to investigate the frequency of the mutant gene. All affected horses tested were homozygous for this mutation. Heterozygous carriers were detected in high frequency in families segregating for this trait, and the frequency of carriers in unrelated Egyptian Arabians was 10.3%. The mapping and discovery of the LFS mutation represents the first successful use of whole-genome SNP scanning in the horse for any trait. The RFLP assay can be used to assist breeders in avoiding carrier-to-carrier matings and thus in preventing the birth of affected foals.

Author Summary

Genetic disorders affect many domesticated species, including the horse. In this study we have focused on Lavender Foal Syndrome, a seizure disorder that leads to suffering and death in foals soon after birth. A recessively inherited disorder, its occurrence is often unpredictable and difficult for horse breeders to avoid without a diagnostic test for carrier status. The recent completion of the horse genome sequence has provided new tools for mapping traits with unprecedented resolution and power. We have applied one such tool, the Equine SNP50 genotyping chip, to a small sample set from horses affected with Lavender Foal Syndrome. A single genetic location associated with the disorder was rapidly identified using this approach. Subsequent sequencing of functional candidate genes in this location revealed a single base deletion that likely causes Lavender Foal Syndrome. From a practical standpoint, this discovery and the development of a diagnostic test for the LFS allele provides a valuable new tool for breeders seeking to avoid the disease in their foal crop. However, this work also illustrates the utility of whole-genome association studies in the horse.

Quiescent T lymphocytes containing latent human immunodeficiency virus (HIV) provide a long-lived viral reservoir. This reservoir may be the source of active infection that is reinitiated following the cessation of antiretroviral therapy. Therefore, it is important to understand the mechanisms involved in latent infection to develop new strategies to eliminate the latent HIV reservoir. We have previously demonstrated that latently infected quiescent lymphocytes can be generated during thymopoiesis in vivo in the SCID-hu mouse system. However, there is still a pressing need for an in vitro model of HIV latency in primary human cells. Here, we present a novel in vitro model that recapitulates key aspects of dormant HIV infection. Using an enhanced green fluorescent protein-luciferase fusion protein-containing reporter virus, we have generated a stable infection in primary human CD4+ CD8+ thymocytes in the absence of viral gene expression. T-cell activation induces a >200-fold induction of reporter activity. The induced reporter activity originates from a fully reverse-transcribed and integrated genome. We further demonstrate that this model can be useful to study long terminal repeat regulation, as previously characterized NF-κB response element mutations decrease the activation of viral gene expression. This model can therefore be used to study intricate molecular aspects of activation-inducible HIV infection in primary cells.

The herpesvirus life cycle has two distinct phases: latency and lytic replication. The balance between these two phases is critical for viral pathogenesis. It is believed that cellular signals regulate the switch from latency to lytic replication. To systematically evaluate the cellular signals regulating this reactivation process in Kaposi sarcoma–associated herpesvirus, the effects of 26,000 full-length cDNA expression constructs on viral reactivation were individually assessed in primary effusion lymphoma–derived cells that harbor the latent virus. A group of diverse cellular signaling proteins were identified and validated in their effect of inducing viral lytic gene expression from the latent viral genome. The results suggest that multiple cellular signaling pathways can reactivate the virus in a genetically homogeneous cell population. Further analysis revealed that the Raf/MEK/ERK/Ets-1 pathway mediates Ras-induced reactivation. The same pathway also mediates spontaneous reactivation, which sets the first example to our knowledge of a specific cellular pathway being studied in the spontaneous reactivation process. Our study provides a functional genomic approach to systematically identify the cellular signals regulating the herpesvirus life cycle, thus facilitating better understanding of a fundamental issue in virology and identifying novel therapeutic targets.

Author Summary

Kaposi sarcoma is a cancer that commonly occurs in AIDS patients. The tumor-associated virus, Kaposi sarcoma–associated herpesvirus, has two distinct phases in its life cycle: inactive latency and active lytic replication. The balance between these two phases is critical for viral pathogenesis. Cellular signals play a role in the switch from latency to lytic replication, termed reactivation. To systematically evaluate the cellular signals regulating this reactivation process in Kaposi sarcoma–associated herpesvirus, a genome-wide cDNA library screen was conducted. Twenty-six thousand mammalian genes were individually expressed in cells that harbor the latent virus, and their effect on reactivation was assessed through a sensitive reporter system. A group of diverse cellular signaling proteins were identified and validated. Further analysis revealed that the activation of the cellular Raf/MEK/ERK/Ets-1 pathway is shared by multiple upstream inducers to trigger reactivation. This work provides a functional genomic approach to systematically identify the cellular signals regulating the herpesvirus life cycle, thus facilitating better understanding of a fundamental issue in virology and identifying novel therapeutic targets.

The host cell activation state impacts the nature of human immunodeficiency virus infection. Activated cells facilitate productive infections; quiescent cells enable the virus to enter a latent state, the major obstacle to viral clearance. We wanted to understand how these differences affected viral gene expression. In quiescent cells activated prior to infection, viral RNA was seen 12 h postinfection; when cells were stimulated postinfection, viral RNA was not seen until 36 h postinfection. Up-regulation of viral RNA in latently infected cells occurred within 2 h poststimulation. This hierarchy also held true for viral protein production. These results may explain the rapid reemergence of viremia following termination of therapy.

Antigenic variation inherent in human immunodeficiency virus type 1 (HIV-1) virions that successfully instigate new infections transferred by sex has not been well defined. Yet this is the viral “challenge” which any vaccine-induced immunity must deal with. Closely timed comparisons of the virus circulating in the “donor” and that which initiates new infection are difficult to carry out rigorously, as suitable samples are very hard to get in the face of ethical hurdles. Here we investigate HIV-1 variation in four homosexual couples where we sampled blood from both parties within several weeks of the estimated transmission event. We analyzed variation within highly immunogenic HIV-1 internal proteins encoding epitopes recognized by cytotoxic Tlymphocytes (CTLs). These responses are believed to be crucial as a means of containing viral replication. In the donors we detected virions capable of evading host CTL recognition at several linked epitopes of distinct HLA class I restriction. When a donor transmitted escape variants to a recipient with whom he had HLA class I molecules in common, the recipient's CTL response to those epitopes was prevented, thus impeding adequate viral control. In addition, we show that even when HLA class I alleles are disparate in the transmitting couple, a single polymorphism can abolish CTL recognition of an overlapping epitope of distinct restriction and so confer immune escape properties to the recipient's seroconversion virus. In donors who are themselves controlling an early, acute infection, the precise timing of onward transmission is a crucial determinant of the viral variants available to compose the inoculum.

Reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV) lytic replication is mediated by the viral RTA transcription factor, but little is known about the physiological processes controlling its expression or activity. Links between autonomic nervous system activity and AIDS-associated Kaposi's sarcoma led us to examine the potential influence of catecholamine neurotransmitters. Physiological concentrations of epinephrine and norepinephrine efficiently reactivated lytic replication of KSHV in latently infected primary effusion lymphoma cells via β-adrenergic activation of the cellular cyclic AMP/protein kinase A (PKA) signaling pathway. Effects were blocked by PKA antagonists and mimicked by pharmacological and physiological PKA activators (prostaglandin E2 and histamine) or overexpression of the PKA catalytic subunit. PKA up-regulated RTA gene expression, enhanced activity of the RTA promoter, and posttranslationally enhanced RTA's trans-activating capacity for its own promoter and heterologous lytic promoters (e.g., the viral PAN gene). Mutation of predicted phosphorylation targets at RTA serines 525 and 526 inhibited PKA-mediated enhancement of RTA trans-activating capacity. Given the high catecholamine levels at sites of KSHV latency such as the vasculature and lymphoid organs, these data suggest that β-adrenergic control of RTA might constitute a significant physiological regulator of KSHV lytic replication. These findings also suggest novel therapeutic strategies for controlling the activity of this oncogenic gammaherpesvirus in vivo.

The role of cytotoxic T-lymphocyte (CTL) escape in rapidly progressive infant human immunodeficiency virus type 1 (HIV-1) infection is undefined. The data presented here demonstrate that infant HIV-1-specific CTL can select for viral escape variants very early in life. These variants, furthermore, may be selected specifically in the infant, despite the same CTL specificity being present in the mother. Additionally, pediatric CTL activity may be compromised both by the transmission of maternal escape variants and by mother-to-child transmission of escape variants that originally arose in the father. The unique acquisition of these CTL escape forms may help to explain the severe nature of some pediatric HIV infections.

Murine gammaherpesvirus 68 (MHV-68), Kaposi's sarcoma-associated herpesvirus (HHV-8), and Epstein-Barr virus (EBV) are all members of the gammaherpesvirus family, characterized by their ability to establish latency in lymphocytes. The RTA protein, conserved in all gammaherpesviruses, is known to play a critical role in reactivation from latency. Here we report that HHV-8 RTA, not EBV RTA, was able to induce MHV-68 lytic viral proteins and DNA replication and processing and produce viable MHV-68 virions from latently infected cells at levels similar to those for MHV-68 RTA. HHV-8 RTA was also able to activate two MHV-68 lytic promoters, whereas EBV RTA was not. In order to define the domains of RTA responsible for their functional differences in viral promoter activation and initiation of the MHV-68 lytic cycle, chimeric RTA proteins were constructed by exchanging the N-terminal and C-terminal domains of the RTA proteins. Our data suggest that the species specificity of MHV-68 RTA resides in the N-terminal DNA binding domain.

HIV-specific CD4+ T helper lymphocytes are preferred targets for infection. Although complete interruption of combination antiretroviral therapy (ART) can form part of therapeutic manipulations, there is grave concern that the resumption of viral replication might destroy, perhaps irreversibly, these T helper populations. High viremia blocks the proliferation capacity of HIV-specific helper cells. However, cytokine production assays imply that some antigen-specific effector function is retained. Despite this careful work, it remains unclear whether the return of HIV-1 replication physically destroys HIV-1–specific T helper cells in the peripheral blood. Difficulties in producing stable peptide-MHC class II complexes and the very low frequencies of antigen-specific CD4+ T cells have delayed the application of this powerful technique. Here we employ HLA class II tetramers and validate a sensitive, quantitative cell-enrichment technique to detect HIV-1 T helper cells. We studied patients with early-stage HIV infection who were given a short, fixed course of ART as part of a clinical study. We did not find significant deletion of these cells from the peripheral circulation when ART was stopped and unfettered HIV replication returned. The turnover of these virus-specific cells increased and they adopted an effector phenotype when viremia returned.

Kaposi's sarcoma-associated herpesvirus and murine gammaherpesvirus-68 (MHV-68) establish latent infections and are associated with various types of malignancies. They are members of the gamma-2 herpesvirus subfamily and encode a replication and transcriptional activator, RTA, which is necessary and sufficient to disrupt latency and initiate the viral lytic cycle in vitro. We have constructed a recombinant MHV-68 virus that overexpresses RTA. This virus has faster replication kinetics in vitro and in vivo, is deficient in establishing latency, exhibits a reduction in the development of a mononucleosis-like disease in mice, and can protect mice against challenge by wild-type MHV-68. The present study, by using MHV-68 as an in vivo model system, demonstrated that RTA plays a critical role in the control of viral latency and suggests that latency is a determinant of viral pathogenesis in vivo.

Objective To examine the access to the renal transplant waiting list and renal transplantation in Scotland.

Design Cohort study.

Setting Renal and transplant units in Scotland.

Participants 4523 adults starting renal replacement therapy in Scotland between 1 January 1989 and 31 December 1999.

Main outcome measures Impact of age, sex, social deprivation, primary renal disease, renal or transplant unit, and geography on access to the waiting list and renal transplantation.

Results 1736 of 4523 (38.4%) patients were placed on the waiting list for renal transplantation and 1095 (24.2%) underwent transplantation up to 31 December 2000, the end of the study period. Patients were less likely to be placed on the list if they were female, older, had diabetes, were in a high deprivation category, and were treated in a renal unit in a hospital with no transplant unit. Patients living furthest away from the transplant centre were listed more quickly. The only factors governing access to transplantation once on the list were age, primary renal disease, and year of listing. A significant centre effect was found in access to the waiting list and renal transplantation.

Conclusions A major disparity exists in access to the renal transplant waiting list and renal transplantation in Scotland. Comorbidity may be an important factor.

Murine gammaherpesvirus 68 (MHV-68 [also referred to as γHV68]) is phylogenetically related to Kaposi's sarcoma-associated herpesvirus (KSHV [also referred to as HHV-8]) and Epstein-Barr virus (EBV). However, unlike KSHV or EBV, MHV-68 readily infects fibroblast and epithelial cell lines derived from several mammalian species, providing a system to study productive and latent infections as well as reactivation of gammaherpesviruses in vivo and in vitro. To carry out rapid genome-wide analysis of MHV-68 gene expression, we made DNA arrays containing nearly all of the known and predicted open reading frames (ORFs) of the virus. RNA obtained from an MHV-68 latently infected cell line, from cells lytically infected with MHV-68 in culture, and from the lung tissue of infected mice was used to probe the MHV-68 arrays. Using a tightly latent B-cell line (S11E), the MHV-68 latent transcription program was quantitatively described. Using BHK-21 cells and infected mice, we demonstrated that latent genes are transcribed during lytic replication and are relatively independent of de novo protein synthesis. We determined that the transcription profiles at the peak of lytic gene expression are similar in cultured fibroblast and in the lung of infected mice. Finally, the MHV-68 DNA arrays were used to examine the gene expression profile of a recombinant virus that overexpresses replication and transcription activator (RTA), C-RTA/MHV-68, during lytic replication in cell culture. The recombinant virus replicates faster then the parental strain and the DNA arrays revealed that nearly every MHV-68 ORF examined was activated by RTA overexpression. Examination of the gene expression patterns of C-RTA/MHV-68 over a time course led to the finding that the M3 promoter is RTA responsive in the absence of other viral factors.

Nasopharyngeal carcinoma, Kaposi's sarcoma, and B-cell lymphomas are human malignancies associated with gammaherpesvirus infections. Members of this virus family are characterized by their ability to establish latent infections in lymphocytes. The latent viral genome expresses very few gene products. The infected cells are therefore poorly recognized by the host immune system, allowing the virus to persist for long periods of time. We sought to identify the cell-specific factors that allow these viruses to redirect their life cycle from productive replication to latency. We find that the cellular transcription factor NF-κB can regulate this process. Epithelial cells and fibroblasts support active (lytic) gammaherpesvirus replication and have low NF-κB activity. However, overexpression of NF-κB in these cells inhibits the replication of the gammaherpesvirus murine herpesvirus 68 (MHV68). In addition, overexpression of NF-κB inhibits the activation of lytic promoters from MHV68 and human gammaherpesviruses Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV). In lymphocytes latently infected with KSHV or EBV, the level of NF-κB activity is high, and treatment of these cells with an NF-κB inhibitor leads to lytic protein synthesis consistent with virus reactivation. These results suggest that high levels of NF-κB can inhibit gammaherpesvirus lytic replication and may therefore contribute to the establishment and maintenance of viral latency in lymphocytes. They also suggest that NF-κB may be a novel target for the disruption of virus latency and therefore the treatment of gammaherpesvirus-related malignancies.