Timing of antiretroviral therapy after diagnosis of cryptococcal meningitis.

BACKGROUND: Cryptococcal meningitis accounts for 20 to 25% of acquired immunodeficiency syndrome-related deaths in Africa. Antiretroviral therapy (ART) is essential for survival; however, the question of when ART should be initiated after diagnosis of cryptococcal meningitis remains unanswered. METHODS: We assessed survival at 26 weeks among 177 human immunodeficiency virus-infected adults in Uganda and South Africa who had cryptococcal meningitis and had not previously received ART. We randomly assigned study participants to undergo either earlier ART initiation (1 to 2 weeks after diagnosis) or deferred ART initiation (5 weeks after diagnosis). Participants received amphotericin B (0.7 to 1.0 mg per kilogram of body weight per day) and fluconazole (800 mg per day) for 14 days, followed by consolidation therapy with fluconazole. RESULTS: The 26-week mortality with earlier ART initiation was significantly higher than with deferred ART initiation (45% [40 of 88 patients] vs. 30% [27 of 89 patients]; hazard ratio for death, 1.73; 95% confidence interval [CI], 1.06 to 2.82; P=0.03). The excess deaths associated with earlier ART initiation occurred 2 to 5 weeks after diagnosis (P=0.007 for the comparison between groups); mortality was similar in the two groups thereafter. Among patients with few white cells in their cerebrospinal fluid (<5 per cubic millimeter) at randomization, mortality was particularly elevated with earlier ART as compared with deferred ART (hazard ratio, 3.87; 95% CI, 1.41 to 10.58; P=0.008). The incidence of recognized cryptococcal immune reconstitution inflammatory syndrome did not differ significantly between the earlier-ART group and the deferred-ART group (20% and 13%, respectively; P=0.32). All other clinical, immunologic, virologic, and microbiologic outcomes, as well as adverse events, were similar between the groups. CONCLUSIONS: Deferring ART for 5 weeks after the diagnosis of cryptococcal meningitis was associated with significantly improved survival, as compared with initiating ART at 1 to 2 weeks, especially among patients with a paucity of white cells in cerebrospinal fluid. (Funded by the National Institute of Allergy and Infectious Diseases and others; COAT ClinicalTrials.gov number, NCT01075152.).

Impact of ejection fraction on the clinical response to cardiac resynchronization therapy in mild heart failure.

BACKGROUND: Current guidelines recommend cardiac resynchronization therapy (CRT) in mild heart failure (HF) patients with QRS prolongation and ejection fraction (EF) ≤30%. To assess the effect of CRT in less severe systolic dysfunction, outcomes in the REsynchronization reVErses Remodeling in Systolic left vEntricular dysfunction (REVERSE) study were evaluated in which patients with left ventricular (LV) ejection fraction (LVEF) >30% were included. METHODS AND RESULTS: The results of patients with baseline EF >30% (n=177) and those with EF ≤30% (n=431), as determined by a blinded core laboratory, were compared. In the LVEF >30% subgroup, there was a trend for improvement in the clinical composite response with CRT ON versus CRT OFF (P=0.06) and significant reductions in LV end systolic volume index (-6.7 ± 21.1 versus 2.1 ± 17.6 mL/m(2); P=0.01) and LV mass (-20.6 ± 50.5 versus 5.0 ± 42.4 g; P=0.04) after 12 months. The time to death or first HF hospitalization was significantly prolonged with CRT (hazard ratio, 0.26; P=0.012). In the LVEF <30% subgroup, significant improvements in clinical composite response (P=0.02), reverse remodeling parameters, and time to death or first HF hospitalization (hazard ratio, 0.58; P=0.047) were observed. After adjusting for important covariates, the CRT ON assignment remained independently associated with improved time to death or first HF hospitalization (hazard ratio, 0.54; P=0.035), whereas there was no significant interaction with LVEF. CONCLUSIONS: Among subjects with mild HF, QRS prolongation, and LVEF >30%, CRT produced reverse remodeling and similar clinical benefit compared with subjects with more severe LV systolic dysfunction. CLINICAL TRIAL REGISTRATION- URL: http://www.clinicaltrials.gov. Unique identifier: NCT00271154.

Case-parent analysis of variation in pubertal hormone genes and pediatric osteosarcoma: a Children's Oncology Group (COG) study.

Osteosarcoma (OS) is a rare malignant bone tumor with an overall incidence rate of 4.6 cases per million children aged 0-19 years in the United States. While the etiology of OS is largely unknown, its distinctive age-incidence pattern suggests that growth and development is crucial in genesis. Prior studies have suggested that variants in genes in the estrogen metabolism (ESTR) and insulin-like growth factor/growth hormone (IGF/GH) pathways are associated with OS. We examined 798 single nucleotide polymorphisms (SNPs) in 42 genes from these pathways in a case-parent study (229 complete triads and 56 dyads) using buccal cell samples. Relative risks (RR) and 95% confidence intervals (CI) associated with transmitting one or two copies of the variant were estimated using log-linear models. After Bonferroni correction, 1 SNP within the ESTR pathway (rs1415270: RR = 0.50 and 8.37 for 1 and 2 vs. 0 copies, respectively; p = 0.010), and two SNPs in the IGF/GH pathway (rs1003737: RR = 0.91 and 0.0001 for 1 and 2 vs. 0 copies, respectively; p <0.0001 and rs2575352: RR = 2.62 and 0.22 for 1 and 2 vs. 0 copies; p < 0.0001) were significantly associated with OS incidence. These results confirm previous findings that variation in the estrogen metabolism and bone growth pathways influence OS risk and further support a biologically and epidemiologically plausible role in OS development.

New methods for finding common insertion sites and co-occurring common insertion sites in transposon- and virus-based genetic screens.

Insertional mutagenesis screens in mice are used to identify individual genes that drive tumor formation. In these screens, candidate cancer genes are identified if their genomic location is proximal to a common insertion site (CIS) defined by high rates of transposon or retroviral insertions in a given genomic window. In this article, we describe a new method for defining CISs based on a Poisson distribution, the Poisson Regression Insertion Model, and show that this new method is an improvement over previously described methods. We also describe a modification of the method that can identify pairs and higher orders of co-occurring common insertion sites. We apply these methods to two data sets, one generated in a transposon-based screen for gastrointestinal tract cancer genes and another based on the set of retroviral insertions in the Retroviral Tagged Cancer Gene Database. We show that the new methods identify more relevant candidate genes and candidate gene pairs than found using previous methods. Identification of the biologically relevant set of mutations that occur in a single cell and cause tumor progression will aid in the rational design of single and combinatorial therapies in the upcoming age of personalized cancer therapy.

Sarcomas in TP53 germline mutation carriers: a review of the IARC TP53 database.

BACKGROUND: Sarcoma is the index diagnosis of Li-Fraumeni syndrome (LFS), a familial predisposition to cancer that also includes brain cancer, breast cancer, and adrenal cortical carcinoma. Germline mutations in the TP53 gene are detected in approximately 80% of families that fulfill LFS criteria and in 15% to 25% of families that fulfill criteria for Li-Fraumeni-like syndrome (LFS), a group of related syndromes with broader clinical criteria. METHODS: The authors of this report used the International Agency for Research on Cancer TP53 database to analyze the types, age at onset and mutation patterns of sarcoma in TP53 mutation carriers. Those data were compared with sarcoma types in the general population of Caucasians using data from the National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) Program. RESULTS: Overall, sarcomas represented 25% of tumors in TP53 mutation carriers, and 95.6% occurred before age 50 years compared with 38.3% before age 50 years in the SEER data set. Sarcomas were more likely to be rhabdomyosarcoma in carriers aged <5 years (odds ratio [OR], 11.6; 95% confidence interval [CI], 6.1-21.9) and osteosarcoma in carriers at any age (aged <20 years: OR, 1.41; 95% CI, 1.02-1.94; age >20 years: OR, 4.61; 95% CI, 2.72-7.83). Early sarcoma (at age <20 years) was associated with missense mutations in exons encoding the DNA-binding domain of p53 protein. Conversely, p53 null mutations (frameshift, splice sites, nonsense) and mutations outside the DNA-binding domain were associated with leiomyosarcoma (OR, 10.1; 95% CI, 3.4-29.9), a type of sarcoma that occurred after age 20 years. CONCLUSIONS: The current results further demonstrated genotype-phenotype correlations and age-dependent variations in sarcoma types in carriers of germline TP53 mutations.

Statistical Approaches to Assess the Effects of Disease on Neurocognitive Function Over Time.

INTRODUCTION: Assessment of the effects of disease on neurocognitive outcomes in children over time presents several challenges. These challenges are particularly pronounced when conducting studies in low-income countries, where standardization and validation is required for tests developed originally in high-income countries. We present a statistical methodology to assess multiple neurocognitive outcomes over time. We address the standardization and adjustment for age in neurocognitive testing, present a statistical methodology for development of a global neurocognitive score, and assess changes in individual and global neurocognitive scores over time in a cohort of children with cerebral malaria. METHODS: Ugandan children with cerebral malaria (CM, N = 44), uncomplicated malaria (UM, N = 54) and community controls (N = 89) were assessed by cognitive tests of working memory, executive attention and tactile learning at 0, 3, 6 and 24 months after recruitment. Tests were previously developed and validated for the local area. Test scores were adjusted for age, and a global score was developed based on the controls that combined the assessments of impairment in each neurocognitive domain. Global normalized Z-scores were computed for each of the three study groups. Model-based tests compare the Z-scores between groups. RESULTS: We found that continuous Z-scores gave more powerful conclusions than previous analyses of the dataset. For example, at all four time points, children with CM had significantly lower global Z-scores than controls and children with UM. Our methods also provide more detailed descriptions of longitudinal trends. For example, the Z-scores of children with CM improved from initial testing to 3 months, but remained at approximately the same level below those of controls or children with UM from 3 to 24 months. Our methods for combining scores are more powerful than tests of individual cognitive domains, as testing of the individual domains revealed differences at only some but not all time points.

Malaria with neurological involvement in Ugandan children: effect on cognitive ability, academic achievement and behaviour.

BACKGROUND: Malaria is a leading cause of ill health and neuro-disability in children in sub-Saharan Africa. Impaired cognition is a common outcome of malaria with neurological involvement. There is also a possibility that academic achievement may be affected by malaria with neurological involvement given the association between cognitive ability and academic achievement. This study investigated the effect of malaria with neurological involvement on cognitive ability, behaviour and academic achievement. METHODS: This prospective case-control study was carried out in Kampala City, Uganda between February 2008 and October 2010. Sixty-two children with a history of malaria with neurological involvement were followed up and given assessments for cognitive ability (working memory, reasoning, learning, visual spatial skills and attention), behaviour (internalizing and externalizing problems) and academic achievement (arithmetic, spelling and reading) three months after the illness. Sixty-one community controls recruited from the homes or neighbouring families of the cases were also given the same assessments. Tests scores of the two groups were compared using analysis of covariance with age, sex, level of education, nutritional status and quality of the home environment as covariates. This study was approved by the relevant ethical bodies and informed consent sought from the caregivers. RESULTS: Children in the malaria group had more behavioural problems than the community controls for internalizing problems (estimated mean difference = -3.71, 95% confidence interval (CI), = -6.34 to -1.08, p = 0.007). There was marginal evidence of lower attention scores (0.40, CI = -0.05 to 0.86, p = 0.09). However, excluding one child from the analyses who was unable to perform the tests affected the attention scores to borderline significance (0.32, CI, = 0.01 to 0.62, p = 0.05). No significant differences were observed in other cognitive abilities or in academic achievement scores. CONCLUSION: Malaria with neurological involvement affects behaviour, with a minimal effect on attention but no detectable effect on academic achievement at three months post discharge. This study provides evidence that development of cognitive deficits after malaria with neurological involvement could be gradual with less effect observed in the short term compared to the long term.

Transmission of HLA-DP variants from parents to children with B-cell precursor acute lymphoblastic leukemia: log-linear analysis using the case-parent design.

Childhood B-cell precursor acute lymphoblastic leukemia (BCP ALL) is usually initiated in utero and is thought to progress to overt leukemia under the influence of delayed exposure to a common infection. Based on the hypothesis that polymorphic HLA-DP variants can restrict T-cell responses to infection, we previously compared DP supertype frequencies in BCP ALL patients with that of unrelated newborn controls. We reported that the DP2 supertype was associated with susceptibility, whereas DP1 was associated with protection. However, the association of genetic variants in children with early-onset diseases such as ALL may be a proxy for parental effects. Here we examine whether maternal DP1 and DP2 are associated with BCP ALL by fitting log-linear models in a combined series of family triads (both parents and case child) and dyads (1 parent and case child; n = 571) in comparison with similar models in non-BCP leukemia (n = 198). We report no evidence of maternal DP1 or DP2 associations with BCP ALL, but we did identify suggestive evidence of maternal undertransmission of the infrequent supertypes DP11 and DP15. Although these results require confirmation, they suggest that DP11 and DP15 may be protective or that there is transmission ratio distortion of these supertypes in BCP ALL.

Proportion statistics to detect differentially expressed genes: a comparison with log-ratio statistics.

BACKGROUND: In genetic transcription research, gene expression is typically reported in a test sample relative to a reference sample. Laboratory assays that measure gene expression levels, from Q-RT-PCR to microarrays to RNA-Seq experiments, will compare two samples to the same genetic sequence of interest. Standard practice is to use the log(2)-ratio as the measure of relative expression. There are drawbacks to using this measurement, including unstable ratios when the denominator is small. This paper suggests an alternative estimate based on a proportion that is just as simple to calculate, just as intuitive, with the added benefit of greater numerical stability. RESULTS: Analysis of two groups of mice measured with 16 cDNA microarrays found similar results between the previously used methods and our proposed methods. In a study of liver and kidney samples measured with RNA-Seq, we found that proportion statistics could detect additional differentially expressed genes usually classified as missing by ratio statistics. Additionally, simulations demonstrated that one of our proposed proportion-based test statistics was robust to deviations from distributional assumptions where all other methods examined were not. CONCLUSIONS: To measure relative expression between two samples, the proportion estimates that we propose yield equivalent results to the log(2)-ratio under most circumstances and better results than the log(2)-ratio when expression values are close to zero.

Antiretroviral therapy down-regulates innate antiviral response genes in patients with AIDS in sub-saharan Africa.

OBJECTIVE: HIV pathogenesis is characterized by destructive imbalances between virus-mediated immune damage, antiviral immune responses, and immune activation. We characterized the effects of successful antiretroviral therapy (ART) to identify the breadth and patterns of HIV-associated gene expression. METHODS: In a prospective observational, longitudinal cohort study of 10 ART-naive Ugandans with AIDS (median 30 CD4/µL), we measured mRNA gene profiles in peripheral blood using Affymetrix U133_Plus2.0 microarrays at 0, 2, 4, 8, and 24 weeks after ART initiation. RESULTS: We identified 160 mRNA transcripts that were consistently down-regulated and 48 that were up-regulated after ART at each point over 24 weeks based on linear regression modeling (adjusted P < 0.05), Of these 208 transcripts, approximately half represent heretofore unrecognized ART-responsive genes and one-third have no known function. The down-regulated genes with known function encoded mediators of innate antiviral responses, including antiviral restriction factors, pattern recognition receptors, and interferon response proteins, and mediators of immune activation, cellular proliferation, and apoptosis. CONCLUSIONS: By using ART to block the viral stimulus, we identified transcripts involved in innate antiviral immunity, including antiviral restriction factors and pattern recognition receptors, that were not previously known to be induced by HIV infection.

Use of signal quality measurements to gain efficiency in the analysis of cDNA microarray data.

This research provides a new way to measure error in microarray data in order to improve gene expression analysis. Microarray data contains many sources of error. In order to glean information about mRNA expression levels, the true signal must first be segregated from noise. This research focuses on the variation that can be captured at the spot level in cDNA microarray images. Variation at other levels, due to differences at the array, dye, and block levels, can be corrected for by a variety of existing normalization procedures. Two signal quality estimates that capture the reliability of each spot printed on a microarray are described. A parametric estimate of within-spot variance, referred to here as sigma(2)(spot), assumes that pixels follow a normal distribution and are spatially correlated. A non-parametric estimate of error, called the mean square prediction error (MSPE), assumes that spots of high quality possess pixels that are similar to their neighbors. This paper will provide a framework to use either spot quality measure in downstream analysis, specifically as weights in regression models. Using these spot quality estimates as weights can result in greater efficiency, in a statistical sense, when modeling microarray data.

Signal quality measurements for cDNA microarray data.

Concerns about the reliability of expression data from microarrays inspire ongoing research into measurement error in these experiments. Error arises at both the technical level within the laboratory and the experimental level. In this paper, we will focus on estimating the spot-specific error, as there are few currently available models. This paper outlines two different approaches to quantify the reliability of spot-specific intensity estimates. In both cases, the spatial correlation between pixels and its impact on spot quality is accounted for. The first method is a straightforward parametric estimate of within-spot variance that assumes a Gaussian distribution and accounts for spatial correlation via an overdispersion factor. The second method employs a nonparametric quality estimate referred to throughout as the mean square prediction error (MSPE). The MSPE first smoothes a pixel region and then measures the difference between actual pixel values and the smoother. Both methods herein are compared for real and simulated data to assess numerical characteristics and the ability to describe poor spot quality. We conclude that both approaches capture noise in the microarray platform and highlight situations where one method or the other is superior.

TLR9 polymorphisms are associated with altered IFN-gamma levels in children with cerebral malaria.

Toll-like receptor (TLR) polymorphisms have been associated with disease severity in malaria infection, but mechanisms for this association have not been characterized. The TLR2, 4, and 9 single nucleotide polymorphism (SNP) frequencies and serum interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) levels were assessed in Ugandan children with cerebral malaria (CM, N = 65) and uncomplicated malaria (UM, N = 52). The TLR9 C allele at -1237 and G allele at 1174 were strongly linked, and among children with CM, those with the C allele at -1237 or the G allele at 1174 had higher levels of IFN-gamma than those without these alleles (P = 0.03 and 0.008, respectively). The TLR9 SNPs were not associated with altered IFN-gamma levels in children with UM or altered TNF-alpha levels in either group. We present the first human data that TLR SNPs are associated with altered cytokine production in parasitic infection.

Clinical features and serum biomarkers in HIV immune reconstitution inflammatory syndrome after cryptococcal meningitis: a prospective cohort study.

BACKGROUND: Although antiretroviral therapy (ART) improves survival in persons with cryptococcal meningitis (CM) and AIDS, ART frequently elicits HIV immune reconstitution inflammatory syndrome (IRIS), an exaggerated and frequently deadly inflammatory reaction that complicates recovery from immunodeficiency. The pathogenesis of IRIS is poorly understood and prediction of IRIS is not possible. METHODS AND FINDINGS: We prospectively followed 101 ART-naïve Ugandans with AIDS and recent CM for one year after initiating ART, and used Luminex multiplex assays to compare serum cytokine levels in participants who did or did not develop IRIS. IRIS occurred in 45% of participants with recent CM on ART, including 30% with central nervous system (CNS) manifestations. The median time to CM-IRIS was 8.8 wk on ART. Overall mortality on ART was 36% with IRIS and 21% without IRIS. CM-IRIS was independently associated with death (HR = 2.3, 95% CI 1.1-5.1, p = 0.04). Patients experiencing subsequent CM-IRIS had 4-fold higher median serum cryptococcal antigen (CRAG) levels pre-ART (p = 0.006). Higher pre-ART levels of interleukin (IL)-4 and IL-17 as well as lower tumor necrosis factor (TNF)-α, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and vascular endothelial growth factor (VEGF) predicted future IRIS in multivariate analyses (area under the curve [AUC] = 0.82). An algorithm based on seven pre-ART serum biomarkers was a robust tool for stratifying high (83%), moderate (48%), and low risk (23%) for IRIS in the cohort. After ART was initiated, increasing levels of C-reactive protein (CRP), D-dimer, IL-6, IL-7, IL-13, G-CSF, or IL-1RA were associated with increasing hazard of IRIS by time-to-event analysis (each p≤0.001). At the time of IRIS onset, multiple proinflammatory cytokine responses were present, including CRP and IL-6. Mortality was predicted by pre-ART increasing IL-17, decreasing GM-CSF, and CRP level >32 mg/l (highest quartile). Pre-ART CRP level >32 mg/l alone was associated with future death (OR = 8.3, 95% CI 2.7-25.6, p<0.001). CONCLUSIONS: Pre-ART increases in Th(17) and Th(2) responses (e.g., IL-17, IL-4) and lack of proinflammatory cytokine responses (e.g., TNF-α, G-CSF, GM-CSF, VEGF) predispose individuals to subsequent IRIS, perhaps as biomarkers of immune dysfunction and poor initial clearance of CRAG. Although requiring validation, these biomarkers might be an objective tool to stratify the risk of CM-IRIS and death, and could be used clinically to guide when to start ART or use prophylactic interventions.

A transposon-based genetic screen in mice identifies genes altered in colorectal cancer.

Human colorectal cancers (CRCs) display a large number of genetic and epigenetic alterations, some of which are causally involved in tumorigenesis (drivers) and others that have little functional impact (passengers). To help distinguish between these two classes of alterations, we used a transposon-based genetic screen in mice to identify candidate genes for CRC. Mice harboring mutagenic Sleeping Beauty (SB) transposons were crossed with mice expressing SB transposase in gastrointestinal tract epithelium. Most of the offspring developed intestinal lesions, including intraepithelial neoplasia, adenomas, and adenocarcinomas. Analysis of over 16,000 transposon insertions identified 77 candidate CRC genes, 60 of which are mutated and/or dysregulated in human CRC and thus are most likely to drive tumorigenesis. These genes include APC, PTEN, and SMAD4. The screen also identified 17 candidate genes that had not previously been implicated in CRC, including POLI, PTPRK, and RSPO2.

Donors with group B KIR haplotypes improve relapse-free survival after unrelated hematopoietic cell transplantation for acute myelogenous leukemia.

Survival for patients with acute myeloid leukemia (AML) is limited by treatment-related mortality (TRM) and relapse after unrelated donor (URD) hematopoietic cell transplantation (HCT). Natural killer (NK)-cell alloreactivity, determined by donor killer-cell immunoglobulin-like receptors (KIRs) and recipient HLA, correlates with successful HCT for AML. Hypothesizing that donor KIR genotype (A/A: 2 A KIR haplotypes; B/x: at least 1 B haplotype) would affect outcomes, we genotyped donors and recipients from 209 HLA-matched and 239 mismatched T-replete URD transplantations for AML. Three-year overall survival was significantly higher after transplantation from a KIR B/x donor (31% [95% CI: 26-36] vs 20% [95% CI: 13-27]; P = .007). Multivariate analysis demonstrated a 30% improvement in the relative risk of relapse-free survival with B/x donors compared with A/A donors (RR: 0.70 [95% CI: 0.55-0.88]; P = .002). B/x donors were associated with a higher incidence of chronic graft-versus-host disease (GVHD; RR: 1.51 [95% CI: 1.01-2.18]; P = .03), but not of acute GVHD, relapse, or TRM. This analysis demonstrates that unrelated donors with KIR B haplotypes confer significant survival benefit to patients undergoing T-replete HCT for AML. KIR genotyping of prospective donors, in addition to HLA typing, should be performed to identify HLA-matched donors with B KIR haplotypes.

Sex and age dimorphism of myocardial gene expression in nonischemic human heart failure.

BACKGROUND: We report the first comprehensive analysis of gene expression differences by sex and age in left ventricular samples from 102 patients with dilated cardiomyopathy. METHODS AND RESULTS: Gene expression data (HG-U133A gene chip, Affymetrix) were analyzed from 30 females and 72 males from 3 separate centers. More than 1800 genes displayed sexual dimorphism in the heart (adjusted P value <0.05). A significant number of these genes were highly represented in gene ontology pathways involved in ion transport and G-protein-coupled receptor signaling. Localization of these genes revealed enrichment on both the sex chromosomes as well as chromosomes 3, 4, and 14. The second goal of this study was to determine the effect of age on gene expression. Within the female cohort, >140 genes were differentially expressed in the <55 years age group compared with the >55 years age group. These genes were highly represented in gene ontology pathways involved in DNA damage. In contrast, zero genes in the male cohort <55 years met statistical significance when compared with the >55 years age group. CONCLUSIONS: Gene expression in dilated cardiomyopathy displayed evidence of sexual dimorphism similar to other somatic tissues and age dimorphism within the female cohort.

Using linkage and association to identify and model genetic effects: summary of GAW15 Group 4.

Group 4 at Genetic Analysis Workshop 15 focused on methods that exploited both linkage and association information to map disease loci. All contributions considered the dichotomous trait of rheumatoid arthritis, using either affected sibpairs and/or unrelated controls. While one contribution investigated linkage and association approaches separately in genome-wide analyses, the remaining others focused on joint linkage and association methods in specific genomic regions. The latter contributions proposed new methods and/or examined existing methods that addressed whether one or more polymorphisms partially or fully explained a linkage signal, particularly the methods proposed by Li et al. that are implemented in the computer program Linkage and Association Modeling in Pedigrees (LAMP). Using simulated SNP data under linkage peaks, several contributions found that existing family-based association approaches such as those of Martin et al. and Lake et al. had power similar to LAMP and to several methods proposed by the contributors for testing that a single nucleotide polymorphism partially explains a linkage peak. In evaluating methods for identifying if a polymorphism or a set of polymorphisms fully accounted for a linkage signal, several contributions found that it was important to understand that these methods may be subject to low power in some situations and thus, a non-significant result was not necessarily indicative of the polymorphism(s) being fully responsible for the linkage signal. Finally, modeling the disease using association evidence conditional on linkage may improve understanding of the etiology of disease.

A subpopulation of human peripheral blood NK cells that lacks inhibitory receptors for self-MHC is developmentally immature.

How receptor acquisition correlates with the functional maturation of natural killer (NK) cells is poorly understood. We used quantitative real-time polymerase chain reaction (PCR) assays to compare NKG2 and killer immunoglobulin-like receptor (KIR) gene expression in NK cells from allogeneic transplant recipients and their donors. Marked differences were observed in the NK subsets of recipients who had 8-fold more CD56(bright) cells, diminished KIR expression (except 2DL4), and increased NKG2A. In normal blood not all CD56(dim) cells express KIR, and a novel subpopulation of cells committed to the NK-cell lineage was defined. These cells, which comprise 19.4% +/- 2.8% of the CD56(dim) NK population in healthy donors, express the activating NKG2D and NKG2E receptors but no KIR or NKG2A. Although the CD56(dim) NKG2A(-) KIR(-) NK cells lack "at least one" inhibitory receptor for autologous MHC class I, they are not fully responsive, but rather functionally immature cells with poor cytotoxicity and IFN-gamma production. Upon culture with IL-15 and a stromal cell line, CD56(dim) and CD56(bright) KIR(-) NK cells proliferate, express KIR, and develop cytotoxicity and cytokine-producing potential. These findings have implications for the function of NK cells reconstituting after transplantation and support a model for in vivo development in which CD56(bright) cells precede CD56(dim) cells.