Identification of regulatory T cell molecules associated with severity of multiple sclerosis.

BACKGROUND: Regulatory CD4+ T cells (Tregs) exhibit functional alterations in patients with multiple sclerosis (MS). Transforming growth factor (TGF)-ß is a key regulator of Treg development and function. OBJECTIVE: The objective of this study is to determine whether the expression of functionally relevant TGF-ß-regulated molecules is altered in Tregs from patients with MS. METHODS: Expression of nine Treg markers was analyzed by multi-color flow cytometry in CD4+ T cells and Treg subpopulations of 31 untreated MS patients and age- and sex-matched healthy donors (HDs). Correlations between Treg marker expression and clinical variables were sought. RESULTS: Expression of the transcription factor Helios, which defines thymic-derived Tregs, was decreased in this Treg subpopulation. The frequency of peripherally generated Tregs was increased in patients with MS, particularly in patients with progressive MS. Low frequencies of thymic-derived Tregs were associated with magnetic resonance imaging (MRI) lesion-burden and a high relapse rate. Four surface markers associated with TGF-ß signaling (ABCA1, BTLA, DNAM-1, and GARP) were differentially expressed on Tregs from patients with MS and HDs. Expression levels of CD73, CD103, ABCA1, and PAR2 showed strong correlations with disease severity. CONCLUSION: We have identified novel markers abnormally expressed on Tregs from patients with MS that could detect patients with severe disease.

Comparison of clinical and cognitive characteristics of a Mexican adult clinical population with and without ADHD / Comparación de las características clínicas y cognitivas de una población clínica adulta mexicana con y sin TDAH

Abstract Introduction Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder affecting approximately 5% of the world population, with symptoms that may persist into adulthood. Despite the findings on the clinical course of this disorder, information regarding comorbidity patterns, psychosocial and executive functioning in adult life in those with and without ADHD in Latin American samples is scarce. Objective The aim of this study is to compare the comorbidity pattern, psychosocial, and executive functioning of adults with and without ADHD from a clinical sample. Method One hundred and fifty-one patients between 20 and 45 years, with screened positively on ASRS-V1.1, were invited to continue an evaluation process as part of clinical research program (PROMETEO): 1) K-SADS-PL Mx interview, 2) MINI-Plus interview, ASRS-V1-1 18 item version, BRIEF self-reported questionnaire, SCQA-ADHD, and 3) Individual case review by clinical expert in ADHD. Results Individuals in the ADHD group had a higher average of comorbid disorders (2.5 SD 1.1 vs. 1.3 SD 1.0 respectively, F = .439; t = -6.621; df = 149; p < .001), more likelihood of procrastinating (OR = 6.5; 95% CI[2.6, 16.2]; z = 4.0) and were more likely to present difficulties in both the behavior regulation index (OR = 104.9; 95% CI[31.8, 345.7]; z = 7.65) and the metacognitive index (OR = 94.79; 95% CI[29.10, 308.76]; z = 7.56) compared to the non-ADHD group, regardless of gender. Discussion and conclusions Our results indicate that the ADHD adult group presented with more comorbidity, and worse psychosocial and executive functioning than non-ADHD adults.

Resumen Introducción El trastorno por déficit de atención con hiperactividad (TDAH) es un trastorno del neurodesarrollo que afecta aproximadamente al 5% de la población mundial, persistiendo hasta la adultez. A pesar de los hallazgos acerca del curso clínico de este trastorno, la información es escasa con respecto a los patrones de comorbilidad, funcionamiento psicosocial y ejecutivo en la vida adulta entre aquellos con y sin TDAH en muestras latinoamericanas. Objetivo Comparar el patrón de comorbilidad, el funcionamiento psicosocial y ejecutivo de adultos con y sin TDAH de una muestra clínica. Método Ciento cincuenta y un pacientes entre 20 y 45 años, quienes inicialmente presentaron un tamizaje positivo del ASRS-V1.1, fueron evaluados dentro de un programa de investigación clínica (PROMETEO) con los siguientes instrumentos: 1) la entrevista K-SADS-PL-Mx, 2) la entrevista MINI-Plus, la version de 18 items del ASRS-V1-1, y los cuestionarios autoaplicados BRIEF y SCQA-ADHD y 3) Revisión de cada caso por un clínico experto en el diagnóstico de TDAH. Resultados El grupo de TDAH comparado con aquel sin TDAH presentó un mayor promedio de trastornos comórbidos (2.5 DE 1.1 vs 1.3 DE 1.0 respectivamente, F = .439; t = -6.621; gl = 149; p < .001), mayor probabilidad de procrastinar (OR = 6.5; 95% IC[2.6, 16.2]; z = 4.0), y mayor probabilidad de presentar dificultades tanto en el índice de regulación de la conducta (OR = 104.9; 95% IC[31.8, 345.7]; z = 7.65) como en el índice metacognitivo (OR = 94.79; 95% IC[29.10, 308.76]; z = 7.56) independientemente del sexo. Discusión y conclusión Nuestros resultados señalan que los adultos con TDAH presentan mayor comorbilidad y peor funcionamiento psicosocial y ejecutivo que los adultos sin TDAH.

High frequency of Taenia pisiformis metacestodes and high sex-associated susceptibility to cysticercosis in naturally infected wild rabbits.

Sexual dimorphism is a well-documented phenomenon observed at all levels of the animal kingdom, with the inclusion of both sexes in clinical trials and basic research becoming mandatory. Regarding parasitosis, in several animal species, the signs and virulence of the disease may change depending on the sex of the affected animal. In the cestodiasis caused by Taenia solium and Taenia crassiceps, females are more susceptible to experimental infection than males. Cysticercosis by Taenia pisiformis in rabbits has acquired relevance due to its economic impact, namely affecting welfare and production. In America, specifically in Mexico, there are no formal reports on the infection with T. pisiformis metacestodes in populations of wild rabbits, despite being the country with more endemic species (about 15 species), among them, the volcanoes rabbits or the endangered teporingo (Romerolagus diazi). In this study, 31 wild rabbits were obtained by hunters of some regions of Morelos state during several hunting seasons, and sex, physiological stage, and number of metacestodes were recorded. A high frequency of infection by T. pisiformis metacestodes (67.7%) was found. Also, a higher susceptibility to this infection was observed in does (80% infected) compared to bucks (40%), finding 84.2% of metacestodes (235 metacestodes) in does and 15.8% of metacestodes (44 metacestodes) in bucks. The percentage of infection was higher in lactating compared with pregnant and non-pregnant does, with metacestodes lodging mainly in the uterus. Increasing our knowledge regarding parasitic infections can help us better understand transmission circles as well as the parasite-host interaction of these increasingly at risk rabbit species.

No association of IL2, IL4, IL6, TNF, and IFNG gene polymorphisms was found with Taenia solium human infection or neurocysticercosis severity in a family-based study.

Neurocysticercosis (NC) is caused by the establishment of the metacestode stage of Taenia solium in the human central nervous system. A great heterogeneity in the susceptibility to the infection and to the disease has been reported. While the factors involved in this heterogeneity are not completely understood, clearly different immune-inflammatory profiles have been associated to each condition. This study evaluated the association of cytokine single nucleotide polymorphisms (SNPs) with susceptibility to infection and disease severity in NC patients. Blood samples from 92 NC cases and their parents (trios) were genotyped for SNPs in five cytokines relevant for the immune response: IL4 (-589C/T), IL6 (-174C/G), IFNG (+874T/A), TNF (-238G/A), and IL2 (-330G/T). Specific DNA fragments were amplified by the polymerase chain reaction, using the 5'-nuclease Taqman assay on a 7500 platform, allowing the detection of the polymorphism genotypes. No association between the polymorphisms evaluated neither with susceptibility to infection nor with disease severity was found, although previous studies reported variations in the levels of these cytokines among different NC clinical pictures. These results, nevertheless, add new elements to our understanding of the complex pathogenic mechanisms involved in susceptibility to infection by T. solium cysticerci and the severity of the ensuing disease.

Shared resource laboratories: impact of new design criteria to consolidate flow cytometry diagnostic service.

INTRODUCTION: The Shared Resource Laboratory (SRL) model recently described for research activities would also appear to be highly suitable for diagnostic services. Using modern SRL design criteria and benchmarks, the aim of our study was to verify whether the consolidation of a diagnostic cytofluorimetric activity could improve the overall service. METHODS: Outcome indicators such as impact on analytical quality, clinical satisfaction, team work involvement, and economic performance were evaluated in the planning and setting up of a new central laboratory. Comparison with preconsolidation status allowed us to investigate possible indicators of improvement. RESULTS: A total of 30 140 cytofluorimetric analyses performed before and after consolidation at the Central Laboratory in Pievesestina in north-central Italy were evaluated. The overall score of the clinical satisfaction questionnaire (range, between 1 and 5) increased from 4.3 to 4.9. Full-time equivalent (FTE) operators were reduced from 9 to 4.5 and cytofluorimeters from 6 to 2; economic indicator analyses showed a 17.75% reduction in unitary test costs. CONCLUSIONS: The adoption of new benchmarks and design criteria increased the quality of cytofluorimetric analysis, thus improving specialized diagnostic services and promoting the shared resource clinical laboratory.

Isolation of stem/progenitor cells from normal lung tissue of adult humans.

OBJECTIVES: This study aimed to isolate and characterize stem/progenitor cells, starting from normal airway epithelia, obtained from human adults. MATERIALS AND METHODS: Cultures of multicellular spheroids were obtained from human lung tissue specimens after mechanical and enzymatic digestion. Tissue-specific markers were detected on their cells by immunohistochemical and immunofluorescent techniques. Ultrastructural morphology of the spheroids (termed as bronchospheres) was evaluated by electron microscopy, gene expression analysis was performed by reverse transcription-polymerase chain reaction, and gene down-regulation was analysed by an RNA interference technique. RESULTS: Bronchospheres were found to be composed of cells with high expression of stem cell regulatory genes, which was not or was only weakly detectable in original tissues. Morphological analysis showed that bronchospheres were composed of mixed phenotype cells with type II alveolar and Clara cell features, highlighting their airway resident cell origin. In addition to displaying specific pulmonary and epithelial commitment, bronchospheres showed mesenchymal features. Silencing of the Slug gene, known to play a pivotal role in epithelial-mesenchymal transition processes and which was highly expressed in bronchospheres but not in original tissue, led bronchospheres to gain a differentiated bronchial/alveolar phenotype and to lose the stemness gene expression pattern. CONCLUSIONS: Ours is the first study to describe ex vivo expansion of stem/progenitor cells resident in human lung epithelia, and our results suggest that the epithelial-mesenchymal transition process, still active in a subset of airway cells, may regulate transit of stem/progenitor cells towards epithelial differentiation.

Role of p53 codon 72 arginine allele in cell survival in vitro and in the clinical outcome of patients with advanced breast cancer.

BACKGROUND: The p53 codon 72 polymorphism, which results in either an arginine or proline residue, plays a different role in vitro and in vivo in cell survival and drug resistance. We verified, in vitro, the impact of the arginine allele on cell survival under normoxia and hypoxia, and investigated in vivo the role of p53 codon 72 arginine homozygosity in the clinical outcome of advanced breast cancer patients. METHODS: Tumors at advanced stages grow in vivo in a hypoxic environment, and we mimicked such conditions in vitro using p53 null breast cancer cells transfected with either the arginine or proline allele. We also analyzed in vivo the p53 codon 72 genotype status of advanced breast cancer patients. RESULTS: In vitro transfection of the arginine allele induced higher cell death under normoxia, whereas cell death was greater in proline-transfected cells under hypoxia. The arginine allele upregulated BCRP-I, a hypoxia response gene, which increases drug resistance. Metastatic breast cancer patients homozygous for arginine had a significantly shorter time to progression and overall survival than those with heterozygous arginine/proline tumors. CONCLUSION: We provide a molecular explanation for the association of the arginine allele with tumor aggressiveness and treatment resistance in advanced breast cancer.

Pro-apoptotic activity of cyclopentenone in cancer cells.

Studies on cyclopentenone prostaglandins (CPPGs), clavulones and other cyclopentenones have shown that these compounds have a significant anticancer activity mediated by their cyclopentenone (CP) chemical moiety. In this study the cytotoxicity against cancer cells of the model compound cyclopent-2-en-1-one (2CP) was investigated. Being a highly water soluble small molecule, 2CP could be an ideal candidate to overcome pharmacological issues related to drug delivery and penetration. Its cytotoxic activity was tested on various melanoma and lung cancer cells. Interestingly, 2CP was both cytotoxic and pro-apoptotic, more pronounced on melanoma cells, at concentrations in the sub-micromolar range. On melanoma cells its mechanism of action was mediated by the mitochondria and the activation of caspase 3.

Risk of chronic kidney disease in hypertensive patients with other metabolic conditions.

Using a retrospective cohort design and electronic medical records, we examined chronic kidney disease (CKD) risk over a 6-year period among hypertensive patients in relation to the presence of diabetes, hyperlipidaemia and/or high body mass index. After adjusting for age, sex, smoking status and baseline glomerular filtration rate (GFR), hypertensive patients without other metabolic risk factors had a relative risk of CKD (versus normotensive patients) of 2.0 (95% CI 1.8-2.2); hypertensive patients with other metabolic conditions had adjusted relative risks ranging from 2.4 to 2.6 for those without comorbid diabetes, and from 3.3 to 5.5 for those with comorbid diabetes. Our study thus confirms prior research demonstrating elevated CKD risk in hypertensive patients, and suggests that this risk varies substantially in relation to other metabolic conditions, especially diabetes.

Cytotoxicity of different selective serotonin reuptake inhibitors (SSRIs) against cancer cells.

Cell membrane ion transporters expression and activity are altered in cancer cells and these phenotypic alterations offer potential targets for cancer therapies. Among the therapeutic agents affecting cell membrane transporters, serotonin reuptake inhibitors (SSRIs) have been shown to have anticancer potential. In this work, we have compared two SSRIs, one very specific for serotonin reuptake transporters (paroxetine) and another which also inhibit norepinephrine and dopamine transporters (venlafaxine), for their ability to counteract growth of various murine and human cancer cell lines. We found that paroxetine has cytotoxic activity against tumor cells, both of murine or human origin in the micromolar concentration range, whereas venlafaxine has not. A neurotransmitter receptor mediated mechanism of action appears thus unlikely for SSRIs cytotoxicity on cancer cells. With ranges of SSRIs cytotoxicity on cancer cells defined, limits in their possible applicability in cancer therapy is discussed.

Pro-apoptotic effect of a nitric oxide-donating NSAID, NCX 4040, on bladder carcinoma cells.

Nitric oxide-releasing non steroidal anti-inflammatory drugs (NO-NSAIDs) are a promising class of compounds that cause cell cycle perturbations and induce apoptosis in cell lines from different tumors. We investigated the activity of a recently developed NO-NSAID (NCX 4040) in bladder cancer cell lines (HT1376 and MCR). Cells were treated with different drug concentrations for different exposure times. Cytostatic and cytocidal activity was tested by SRB assay and apoptosis was evaluated by TUNEL analysis, ANNEXIN V assay and fluorescence microscopy. To further investigate the cell death-inducing mechanisms of NCX 4040, we analyzed gp-170, caspase expression and mitochondrial membrane potential (Delta Psi) depolarization. NCX 4040 showed a striking cytocidal activity in both cell lines, reaching LC(50) at a 10-microM and 50-microM concentrations in HT1376 and in MCR cells, respectively, after an exposure of only 6 h followed by an 18-h washout. Apoptosis was triggered in up to 90% of cells and was associated with active caspase-3 expression and Delta Psi depolarization in both cell lines after a 6-h exposure. In conclusion, NCX 4040, which probably causes apoptosis via a mitochondrial-dependent mechanism, could prove to be a useful agent for improving bladder cancer treatment.

Modulation of drug cytotoxicity by Iressa (ZD1839) in pancreatic cancer cell lines.

High expression of the epidermal growth factor receptor (EGFR) family confers a growth advantage on malignant cells in various tumor types. Most pancreatic cancers express EGFR, which seems to play an important role in the acquisition of aggressive clinical behaviour and in tumor invasion. Iressa (ZD1839), a quinazoline tyrosine kinase inhibitor selective for the EGF receptor, has shown good anti-tumor activity in both preclinical and clinical studies. Using two pancreatic cancer cell lines that express different EGFR and ErbB-2 levels, we analyzed the activity of Iressa and evaluated its modulation effect on four conventional cytotoxic drugs: gemcitabine, oxaliplatin, docetaxel and SN38. Iressa was tested at scalar doses up to the plasma peak level concentration and showed a similar weak cytostatic effect in both cell lines. Conversely, an additive or weak synergistic effect was observed when the drug was administered simultaneously with or following cytotoxic drugs. Our data show that Iressa has only a weak activity at doses within the plasmatic peak concentration and that its effect is independent of EGFR and p42/p44 expression and phosphorylation levels. This is in agreement with recent literature data that attribute an essential role to a specific EGFR mutation in mediating response to Iressa. This mutation was absent in both pancreatic cell lines tested.

To stay or not to stay: the contribution of tactile and thermal cues to coming to rest in newborn rabbits.

Newborn rabbits, like other altricial mammals, demonstrate thermotaxis and when placed on a thermal gradient locate and come to rest at physiologically appropriate temperatures. Little is known, however, about the sensory-motor components contributing to the in energetic terms important decision of the young to cease locomotion and come to rest. We investigated the behavior of newborn rabbits on two thermal gradients; linear in which pups could use tactile cues from the arena wall, and concentric in which pups were unable to use such cues. On both gradients pups located the warm, thermal-neutral area within the 200-s test time, thereby demonstrating their ability to orient appropriately using thermal cues alone. Unexpectedly, however, pups on the concentric gradient failed, or took significantly longer, to come to rest than pups on the linear gradient. Since the speed of locomotion of pups on the linear gradient was significantly slowed when they were in contact with the arena wall, and in most cases they came to rest in contact with it, we suggest that not only thermal but also tactile cues may be important in bringing young mammals to rest in a thermally appropriate environment.

Initial sequencing and analysis of the human genome.

The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.

Proton stopping powers averaged over beam energy spectra.

Stopping powers averaged over the proton energy spectra at various depths in water and a tissue-like material were calculated for proton beams with initial energies between 50 and 250 MeV. The analysis was made for proton beams with a spread-out Bragg peak (SOBP) to assess in particular the variations of stopping power with depth in the SOBP plateau region. Nuclear interactions were accounted for in determination of the proton energy spectra. In modulated beams, stopping power varies considerably with depth along the plateau region of the spread-out peak. Moreover, stopping powers at the same water depth may differ from each other by up to 20% if referring to modulated or non-modulated beams. Calculations of water-air mass stopping power ratios were also performed, with and without the inclusion of nuclear interactions, for modulated and non-modulated beams. The stopping power ratios do not depend significantly on proton energy, and this makes the effect of accounting for nuclear interactions in proton fluence calculation negligible. For the same reason the stopping power ratios for modulated beams do not appreciably differ from those referred to non-modulated beams with the same Emax at the same depth in water. The present results also show that the dependence of stopping power on energy spread and spatial divergence for proton beams is not negligible in some conditions. Some conclusions are drawn on the possibility of using the residual range as a descriptor of the proton beam quality for different experimental beam conditions.

Building health plan databases to risk adjust outcomes and payments.

OBJECTIVES: To highlight the types and sources of data on medical risk and outcomes routinely collected by managed care organizations over time; to summarize the quality and consistency of these data; and to describe some of the difficulties that arise in collecting, pooling, and using these data. DESIGN: Synthesis of the experiences of two risk-adjustment modeling projects in assembling large volumes of demographic, diagnostic, and expense data from several health maintenance organizations (HMOs) over multiple years. SETTING: Six large HMOs from the Northwest, North Central, and Northeast regions of the USA. INTERVENTIONS: Health plans were approached to participate in a risk-adjustment study, presented with an extensive variable-by-variable data request, and, if willing to participate, asked to specify a desired process for extracting, copying, and transferring selected variables to the study site for purposes of research. Depending on local circumstances, three different approaches were used: (i) health plan staff obtained the data and organized them into the requested study format; (ii) study staff were provided access to health plan data systems to perform the extractions directly; and (iii) health plans hired contract programmers to perform the extractions under the direction of the study team. Key measures of risk and cost were extracted and merged into analysis files. MAIN OUTCOME MEASURES: Complete and consistent eligibility maps, demographic information, inpatient and outpatient diagnoses, and total health plan expense for each enrollee. RESULTS: We have been successful in collecting and integrating complete utilization, morbidity, demographic, and cost data on total memberships of five large HMOs as well as a subset from a sixth HMO, all for multiple years. CONCLUSION: While HMOs vary greatly in the quality and comprehensiveness of their data systems, these attributes have been improving across the board over time. Automated health plan data systems represent potentially valuable sources of data on health risks and outcomes and can be used to benchmark disease management programs and risk adjust capitation payments and medical outcomes.