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Users' guides to the medical literature: XXIV. How to use an article on the clinical manifestations of disease. Evidence-Based Medicine Working Group.

Clinicians rely on knowledge about the clinical manifestations of disease to make clinical diagnoses. Before using research on the frequency of clinical features found in patients with a disease, clinicians should appraise the evidence for its validity, results, and applicability. For validity, 4 issues are important-how the diagnoses were verified, how the study sample relates to all patients with the disease, how the clinical findings were sought, and how the clinical findings were characterized. Ideally, investigators will verify the presence of disease in study patients using credible criteria that are independent of the clinical manifestations under study. Also, ideally the study patients will represent the full spectrum of the disease, undergo a thorough and consistent search for clinical findings, and these findings will be well characterized in nature and timing. The main results of these studies are expressed as the number and percentages of patients with each manifestation. Confidence intervals can describe the precision of these frequencies. Most clinical findings occur with only intermediate frequency, and since these frequencies are equivalent to diagnostic sensitivities, this means that the absence of a single finding is rarely powerful enough to exclude the disease. Before acting on the evidence, clinicians should consider whether it applies to their own patients and whether it has been superseded by new developments. Detailed knowledge of the clinical manifestations of disease should increase clinicians' ability to raise diagnostic hypotheses, select differential diagnoses, and verify final diagnoses. JAMA. 2000;284:869-875

Diagnosis↗

Secular trend in body size among college athletes.

Height and weight were compared across five birth decades (1850-1899) among 1,121 Harvard athletes who were lettermen in various sports. There were considerable differences in the magnitude of the secular trend among the sport categories (crew, baseball, football, track, ice hockey, and two or more sports). Comparing the 1890-1899 and 1860-1869 birth-cohort samples, football lettermen were 2.6 inches (6.6 cm) taller (p less than 0.001) and 20 pounds (9.1 kg) heavier (p less than 0.001). Crew lettermen were 2.6 inches taller (p less than 0.001) and 8.5 pounds (3.9 kg) heavier (p less than 0.05). For lettermen in other sports, changes in mean height and weight were smaller in magnitude. Differential selection for body size may explain the differences in the magnitude of the secular trend when analyzed by specific sport.

Body Height↗

Partial correlation of distance matrices in studies of population structure.

Anthropological studies of human population structure commonly compare various monogenic and polygenic (metric) distance matrices to distance matrices obtained from measures of geographical dispersion, linguistic differences, and migration patterns in an attempt to infer something about the effects of evolutionary factors (drift and differential selection, in particular). It is, though, commonly recognized that geography, language, and migration patterns may be intercorrelated due to the common effects of historical and social processes. Previous attempts to deal with the problems of assessing relative effects among such sets of intercorrelated factors using partial correlations have resulted in coefficients that are either not well defined or have no known sampling distribution or both. Here, we outline a general approach to partialling distance matrices that results in well-defined coefficients and valid significance testing procedures. Application of the matrix partialling methods to a variety of distance matrices obtained for a sample of eight ethnolinguistic groups from the Harvard Solomon Islands Expedition (Friedlaender et al., 1986) reveals a close association between language dissimilarity and dermatoglyphics controlling for geography, thus reinforcing earlier suggestions that dermatoglyphics, properly used, reflect historical relationships of groups in this region better than do anthropometry, odontometrics, or small batteries of blood polymorphisms.

Anthropology↗

Lineage pathway of human brain progenitor cells identified by JC virus susceptibility.

Multipotential human central nervous system progenitor cells, isolated from human fetal brain tissue by selective growth conditions, were cultured as undifferentiated, attached cell layers. Selective differentiation yielded highly purified populations of neurons or astrocytes. This report describes the novel use of this cell culture model to study cell type-specific recognition of a human neurotropic virus, JC virus. Infection by either JC virions or a plasmid encoding the JC genome demonstrated susceptibility in astrocytes and, to a lesser degree, progenitor cells, whereas neurons remained nonpermissive. JC virus susceptibility correlated with significantly higher expression of the NFI-X transcription factor in astrocytes than in neurons. Furthermore, transfection of an NFI-X expression vector into progenitor-derived neuronal cells before infection resulted in viral protein production. These results indicate that susceptibility to JC virus infection occurs at the molecular level and also suggest that differential recognition of the viral promoter sequences can predict lineage pathways of multipotential progenitor cells in the human central nervous system.

Brain↗

Regulation of myeloid cell function and major histocompatibility complex class II expression by tumor necrosis factor.

OBJECTIVE: Tumor necrosis factor (TNF)-neutralizing agents are the most successful means of ameliorating systemic autoimmune inflammation. Neutralization of TNF, however, is often associated with the development of autoantibodies, particularly to nuclear antigens, and the mechanisms of this are unknown. We undertook this study to analyze the effect of TNF and its neutralization on the expression of major histocompatibility complex class II molecules and on the function of antigen-presenting myeloid cells in rheumatoid arthritis (RA). METHODS: Monocytes were isolated from the peripheral blood of RA patients before and after anti-TNF monoclonal antibody (mAb) treatment and from the peripheral blood of controls by negative selection, differentiated in vitro to macrophages, and analyzed by flow cytometry for HLA-DR expression. T cell responses to activation by myeloid cells were assessed in proliferation assays, and messenger RNA (mRNA) levels of the class II transactivator (CIITA) were determined by semiquantitative reverse transcriptase-polymerase chain reaction. RESULTS: HLA-DR expression was significantly reduced on myeloid cells from RA patients with active disease, but was increased to normal levels after anti-TNF mAb treatment. Concordantly, in vitro application of TNF to monocytes from healthy individuals reduced their ability to up-regulate HLA-DR during differentiation to macrophages and, importantly, inhibited their ability to stimulate T cells in mixed lymphocyte reactions. Molecular analysis revealed that the effect of TNF on HLA-DR expression was mediated via suppression of the transcription factor CIITA. CONCLUSION: The data indicate that TNF decreases HLA-DR expression by reducing CIITA mRNA levels in myeloid cells, functionally resulting in a decreased capacity of myeloid cells to stimulate T cells. Concordantly, ameliorating disease activity in chronic inflammatory diseases by neutralizing TNF restores expression of HLA-DR on myeloid cells as well as the ability of myeloid cells to stimulate T cells. Thus, anti-TNF treatment might lead to augmented T cell activation by myeloid cells, thereby promoting immune responses to (auto)antigens and the development of antinuclear antibodies that are frequently associated with anti-TNF therapy.

Adult↗

Guanine 6-O-methylation pattern within the dioxin responsive element of the CYP1A1 enhancer shows two critical guanines for AhR/ARNT binding.

The core-recognition motif for TCDD-liganded AhR/ARNT complex of the dioxin-responsive element (DRE) contains four guanine residues, three on the antisense (5'-T(T)/(A)GCGTG-3') and one on the sense (5'-CACGC(A)/(T)A-3') strand. It has been reported that, in methylation-protection and methylation-interference assays, the TCDD-liganded AhR/ARNT contacts all four guanine residues. On the other hand, it is known that some anticancer drugs, and various environmental and workplace chemicals, including strongly human carcinogenic nitrosamines, lead to the highly miscoding 6-O-methylation of guanine. In the present study, we have investigated whether specific methylation of guanine at the 6-O-position interferes with the binding of TCDD-liganded AhR/ARNT complex to its recognition motif in the CYP1A1 enhancer, and how individual 6-O-methylated guanines contribute to this interference. We found that only two 6-O-methylguanine residues are critical: the closest to the 5'-end within the three-nucleotide sequence (5'-GTG-3'), identical to a half-site E-box element, on the antisense strand, and the only guanine on the sense strand. In contrast, the 6-O-methylguanine closest to the 5'-end (well) and the one closest to the 3'-end (to a lesser extent), both on the antisense strand, were still able to bind the TCDD-liganded AhR/ARNT complex. This shows that the 6-O-methylation of guanine in the core sequences of CYP1A1 enhancer interferes with the binding of the ligand-activated AhR/ARNT complex in a differentially selective manner; it fully impedes binding of this complex to DRE (the prerequisite of most of the toxic effects of TCDD) only when one of the two particular guanines is methylated at the 6-O-position.

Animals↗

Unsupervised immunophenotypic profiling of chronic lymphocytic leukemia.

BACKGROUND: Proteomics and functional genomics have revolutionized approaches to disease classification. Like proteomics, flow cytometry (FCM) assesses concurrent expression of many proteins, with the advantage of using intact cells that may be differentially selected during analysis. However, FCM has generally been used for incremental marker validation or construction of predictive models based on known patterns, rather than as a tool for unsupervised class discovery. We undertook a retrospective analysis of clinical FCM data to assess the feasibility of a cell-based proteomic approach to FCM by unsupervised cluster analysis. METHODS: Multicolor FCM data on peripheral blood (PB) and bone marrow (BM) lymphocytes from 140 consecutive patients with B-cell chronic lymphoproliferative disorders (LPDs), including 81 chronic lymphocytic leukemia (CLLs), were studied. Expression was normalized for CD19 totals, and recorded for 10 additional B-cell markers. Data were subjected to hierarchical cluster analysis using complete linkage by Pearson's correlation. Analysis of CLL in PB samples (n = 63) discovered three major clusters. One cluster (14 patients) was skewed toward "atypical" CLL and was characterized by high CD20, CD22, FMC7, and light chain, and low CD23. The remaining two clusters consisted almost entirely (48/49) of cases recorded as typical BCLL. The smaller "typical" BCLL cluster differed from the larger cluster by high CD38 (P = 0.001), low CD20 (P = 0.001), and low CD23 (P = 0.016). These two typical BCLL clusters showed a trend toward a difference in survival (P = 0.1090). Statistically significant cluster stability was demonstrated by expanding the dataset to include BM samples, and by using a method of random sampling with replacement. CONCLUSIONS: This study supports the concept that unsupervised immunophenotypic profiling of FCM data can yield reproducible subtypes of lymphoma/chronic leukemia. Expanded studies are warranted in the use of FCM as an unsupervised class discovery tool, akin to other proteomic methods, rather than as a validation tool.

ADP-ribosyl Cyclase 1↗

Genome-wide identification of female-enriched genes in zebrafish.

Characteristic differences in morphology, physiology, and behavior between a male and female are correlated to the differential selection of sex-dependent transcriptomes. By using a cDNA array carrying approximately 9,000 zebrafish unique genes, we identified a group of genes whose expression are enriched in the female fish. A subset of these genes have been confirmed and further grouped as egg-enriched genes, as both somatic- and egg-enriched genes or as somatic-enriched genes by means of RNA gel blot hybridization. Most importantly, a significant proportion of these genes are either functionally unknown or are novel genes. Thus, future studies of this group of genes will help us greatly to understand more about sex-determination and sex-related physiology and behavior. In addition, comparison of zebrafish female-enriched genes with that in Drosophila, we found that only germline genes are shared between vertebrate and invertebrate, suggesting that the process of oogenesis is highly conserved during the evolution.

Animals↗

Genetic (Genomic) and Morphological Evidence Suggest That the Korean Endemic Fat Minnow (Rhynchocypris kumgangensis) and the Deogyu Population Represent Distinct Species.

Geographically disconnected populations of freshwater fishes often show ecological and genetic divergence in response to opposing selection differentials in different habitat environments, sometimes leading to the formation of ecotypes and even speciation. Nevertheless, evidence for the speciation through allopatric processes in freshwater fish systems remains scarce. Kumkang fat minnow (Rhynchocypris kumgangensis), a Korean endemic coldwater fish, has recently been suggested to diverge into a separate species (Rhynchocypris deogyuensis). However, the level of their genetic and ecological divergence remains largely unknown. We analyzed population genetic structure of R. kumgangensis together with R. deogyuensis using mitochondrial DNA (mtDNA) and eight newly developed microsatellite markers. Genetic divergence at mitogenome level was further assessed between the presumed two species. Moreover, we examined morphology between two groups by analyzing morphometric traits and also conducted geometric morphometrics on body shape. We found distinct population structure between R. kumgangensis and R. deogyuensis at both mtDNA and microsatellites. One remnant population of R. deogyuensis harbored only a single haplotype and showed a very high level of inbreeding. Comparative analysis of the mitogenomes showed approximately 2.9% divergence between R. kumgangensis and R. deogyuensis, supporting the species-level divergence. The analyses of both morphometric traits and geometric morphometrics indicated significant morphological divergence between the two species. The observed low values of length-weight relationship and condition factor for R. deogyuensis are likely to be attributed to effects of the elevated level of inbreeding and depleted genetic diversity. Overall, our combined genetic/genomic and morphological analyses suggest the considerable divergence between R. kumgangensis and R. deogyuensis, supporting the hypothesis that they are distinct species. The mechanisms underpinning how they speciate still need to be studied further in detail.

Deogyu fat minnow↗

c-fos is induced in the hippocampus during consolidation of sexual imprinting in the zebra finch (Taeniopygia guttata).

c-fos was used to mark regions of enhanced neuronal activity during sexual imprinting, an early learning process by which information about the prospective sexual partner is acquired and consolidated. In the present study, we demonstrate that the hippocampus, already known for its specialized spatial memory capacities in navigating pigeons and in food-storing birds, depicts a selective differential c-fos induction in a situation shown to lead to sexual imprinting, that is, exposing previously isolated male birds to a female for 1 h. c-fos induction is lateralized, the left hippocampus showing more c-fos activity than the right. Our results would indicate a role for the hippocampus in the consolidation process of imprinting, probably in the transfer of information to the other telencephalic areas that show alterations in synaptic connectivity as a result of consolidation of sexual imprinting.

Animals↗

dbQSNP: a database of SNPs in human promoter regions with allele frequency information determined by single-strand conformation polymorphism-based methods.

We present a database, dbQSNP (http://qsnp.gen.kyushu-u.ac.jp/), that provides sequence and allele frequency information for single-nucleotide polymorphisms (SNPs) located in the promoter regions of human genes, which were defined by the 5' ends of full-length cDNA clones. We searched for the SNPs in these regions by sequencing or single-strand conformation polymorphism (SSCP) analysis. The allele frequencies of the identified SNPs in two ethnic groups were quantified by SSCP analyses of pooled DNA samples. The accuracy of our estimation is supported by strong correlations between the frequencies in our data and those in other databases for the same ethnic groups. The frequencies vary considerably between the two ethnic groups studied, suggesting the need for population-based collections and allele frequency determination of SNPs, in, e.g., association studies of diseases. We show profiles of SNP densities that are characteristic of transcription start site regions. A fraction of the SNPs revealed a significantly different allele frequency between the groups, suggesting differential selection of the genes involved.

Alleles↗

Alpha-lipoic acid induces p27Kip-dependent cell cycle arrest in non-transformed cell lines and apoptosis in tumor cell lines.

alpha-Lipoic acid is a naturally-occurring co-factor found in a number of multi-enzyme complexes regulating metabolism. We report here that alpha-lipoic acid induces hyperacetylation of histones in vivo and has differential effects on the growth and viability of normal versus transformed cell lines. The human tumor cell lines FaDu and Jurkat, as well as a Ki-v-Ras-transformed Balb/c-3T3 murine mesenchymal cell line, all initiated apoptosis following exposure to alpha-lipoic acid. In contrast, treatment of non-transformed cell lines with alpha-lipoic acid resulted only in reversible cell cycle arrest in G0/G1. Treatment with butyrate, another short-chain fatty acid, induced a G0/G1 arrest in both transformed and non-transformed cell lines. alpha-Lipoic acid caused a post-translational elevation in the levels of the cyclin-dependent kinase inhibitor p27Kip1. Studies using p27Kip1-deficient MEF cells demonstrated that p27Kip1 was required for the alpha-lipoic acid-mediated cell cycle arrest. The mechanism of apoptosis was independent of Fas-mediated signaling, as alpha-lipoic acid-treated Jurkat cell mutants deficient in Fas or FADD retained sensitivity to apoptosis. The differential selectivity of the pro-apoptotic effects of alpha-lipoic acid for transformed cells supports its potential use in the treatment of neoplastic disorders.

3T3 Cells↗

Blockade of interleukin-6 receptor suppresses reactive astrogliosis and ameliorates functional recovery in experimental spinal cord injury.

Endogenous neural stem/progenitor cells (NSPCs) have recently been shown to differentiate exclusively into astrocytes, the cells that are involved in glial scar formation after spinal cord injury (SCI). The microenvironment of the spinal cord, especially the inflammatory cytokines that dramatically increase in the acute phase at the injury site, is considered to be an important cause of inhibitory mechanism of neuronal differentiation following SCI. Interleukin-6 (IL-6), which has been demonstrated to induce NSPCs to undergo astrocytic differentiation selectively through the JAK/STAT pathway in vitro, has also been demonstrated to play a critical role as a proinflammatory cytokine and to be associated with secondary tissue damage in SCI. In this study, we assessed the efficacy of rat anti-mouse IL-6 receptor monoclonal antibody (MR16-1) in the treatment of acute SCI in mice. Immediately after contusive SCI with a modified NYU impactor, mice were intraperitoneally injected with a single dose of MR16-1 (100 microg/g body weight), the lesions were assessed histologically, and the functional recovery was evaluated. MR16-1 not only suppressed the astrocytic diffentiation-promoting effect of IL-6 signaling in vitro but inhibited the development of astrogliosis after SCI in vivo. MR16-1 also decreased the number of invading inflammatory cells and the severity of connective tissue scar formation. In addition, we observed significant functional recovery in the mice treated with MR16-1 compared with control mice. These findings suggest that neutralization of IL-6 signaling in the acute phase of SCI represents an attractive option for the treatment of SCI.

Analysis of Variance↗

Proteomic analysis of pulmonary sclerosing hemangioma.

Sclerosing hemangioma (SH) is a rare benign pulmonary tumor derived from the primitive respiratory epithelium. However, the pathogenesis of SH has not yet been clear. Surfactant protein, thyroid transcription factor-1, epithelial membrane antigen, cytokeratin, and vimentin have been identified in SH by immunohistochemistry and electron microscopy. To identify proteins specifically regulated in SH, 2-D PAGE was performed using SH and paired normal tissues. Ten selected differentially expressed protein spots were identified by PMF, MALDI-TOF-MS, and database searching. Apolipoprotein A-1, antizyme inhibitor, heat shock 27-kDa protein 1, and antioxidant proteins, such as peroxiredoxin II (Prx II) and GST, were identified among the down-regulated proteins in SH. Western blot and immunohistochemistry confirmed reduced expressions of Prx II and GST in SH versus normal lung tissue. This study is the first report on the reduced expressions of Prx II and GST in SH.

Adult↗

Accounting for correlation and compliance in cluster randomized trials.

This paper discusses causal inference with survival data from cluster randomized trials. It is argued that cluster randomization carries the potential for post-randomization exposures which involve differentially selective compliance between treatment arms, even for an all or nothing exposure at the individual level. Structural models can be employed to account for post-randomization exposures, but should not ignore clustering. We show how marginal modelling and random effects models allow to adapt structural estimators to account for clustering. Our findings are illustrated with data from a vitamin A trial for the prevention of infant mortality in the rural plains of Nepal.

Cluster Analysis↗

Estimating incidence of HIV infection in childbearing age African women using serial prevalence data from antenatal clinics.

Ades and Medley provided the first flexible method for estimating age- and time-specific HIV incidence using HIV prevalence data collected among pregnant women and adjusting for the effect of differential selection between infected and uninfected women. This paper extends the approach proposed by these authors. We used a parametric model that allows the relative inclusion rate to depend on both age, calendar time, and duration of HIV infection. We developed a two dimensional penalized log-likelihood approach for estimating time- and age-specific incidence using a binomial likelihood function and a quadratic roughness penalty which allows smoothing over both age and time. Identifiability of the model parameters and effect of sample size are studied through simulations. The method is illustrated using prenatal HIV testing data recorded from 1995 to 2002 in Abidjan, Côte d'Ivoire, to estimate the HIV annual incidence rate among women aged 12-40 year old, from the beginning of the epidemic to 2002. We show that estimated incidence rates are highly dependent on hypotheses made to model the relative inclusion rate. Despite this dependency, the application of the method leads to new and accurate findings on HIV incidence qualitative features in Abidjan. We highlight the relevance of such a method in monitoring the dynamics of HIV epidemic in Africa which is essential for planning vaccine trials and future treatment needs, and for assessment of prevention policy.

Adolescent↗

Effect of ischemia on the fraction of ryanodine-sensitive cardiac sarcoplasmic reticulum.

The effect of 15 min of global, normothermic ischemia on cardiac sarcoplasmic reticulum (SR) was investigated using the Ca2+ uptake rate and 3H-ryanodine binding of ventricular homogenates and isolated SR vesicles. Ischemia did not affect ryanodine binding in the homogenate, while it increased it in the isolated SR vesicles. Although ischemia decreased the homogenate oxalate-supported Ca2+ uptake rate, measured in the presence of high ryanodine to close the ryanodine-sensitive efflux pathway (+RY), its decrease of the Ca2+ uptake rate, measured in the absence of ryanodine (-RY), was more marked. This finding was also observed in the isolated SR. Although inhibition of the Ca-ATPase and its coupled Ca2+ uptake by thapsigargin proportionately decreased SR Ca2+ uptake -RY and +RY, ischemia decreased the Ca2+ uptake -RY proportionately more. This result suggested that there was a greater fraction of Ca2+ uptake activity in ryanodine-sensitive vesicles after ischemia. However, ischemia also reduced the yield of SR activity in the isolated SR fraction and the results could potentially be due to differential selection of ryanodine-sensitive and ryanodine-insensitive SR in the isolation procedure. We directly tested the hypothesis that ischemia changes the fraction of Ca2+ uptake activity in the ryanodine-sensitive vesicles by estimating the Ca-oxalate capacity measured +RY and -RY. Ischemia decreased the capacity -RY much more than +RY in the homogenate, indicating that more of the SR volume and Ca2+ uptake activity was in the ryanodine-sensitive vesicles after ischemia.

Animals↗

Genetic variability in sensitivity to population density affects the dynamics of simple ecological models.

Many 1-dimensional discrete time ecological models contain a sensitivity parameter that does not affect the dynamic complexity of these models. We show that genetic variability in this parameter can have a strong effect on population dynamics. We incorporate ecological dynamics in two different population genetic models with one locus and two alleles. The first is the classical model of a randomly mating population in Hardy-Weinberg equilibrium, and the second is a model of differential selection in males and females. In populations in Hardy-Weinberg equilibrium, variability in the sensitivity parameter can be maintained by overdominance. In this case, the dynamics of the polymorphic population tend to be much simpler than those of monomorphic populations. In the model with different selection in males and females, polymorphisms can be maintained in various ways, e.g., by opposing directional selection in males and females. Polymorphism in the sensitivity parameter tends to simplify population dynamics in the model with different selection in males and females as well. A number of interesting dynamic effects can be observed, e.g., multiple attractors with complicated basins of attraction. Then the final state of the system after a successful invasion by mutant alleles may depend on the mutation rate and on the distribution of mutational steps. In addition, there are situations in which genetic variability destabilizes a stable population dynamic equilibrium in the monomorphic model. There is an analogy between genetic variability and variability imposed by the environment. If differences in sensitivity are caused by the environment, dynamic effects similar to those in the genetic models can be observed. In addition, source-sink structures that are known to occur in spatially structured models can be seen in the genetic model if one of the genotypes is inviable. The results suggest that combining ecological and population genetic models can lead to a number of new insights. More work is needed, e.g., with fertility models, in which fitnesses are not assigned to individuals, but to mating pairs.

Alleles↗