Long-term ex vivo expansion of human fetal liver primitive haematopoietic progenitor cells in stroma-free cultures

Successful expansion of haematopoietic cells in ex vivo cultures will have important applications in transplantation, gene therapy, immunotherapy and potentially also in the production of non-haematopoietic cell types. Haematopoietic stem cells (HSC), with their capacity to both self-renew and differentiate into all blood lineages, represent the ideal target for expansion protocols. However, human HSC are rare, poorly characterized phenotypically and genotypically, and difficult to test functionally. Defining optimal culture parameters for ex vivo expansion has been a major challenge. We devised a simple and reproducible stroma-free liquid culture system enabling long-term expansion of putative haematopoietic progenitors contained within frozen human fetal liver (FL) crude cell suspensions. Starting from a small number of total nucleated cells, a massive haematopoietic cell expansion, reaching > 1013-fold the input cell number after approximately 300 d of culture, was consistently achieved. Cells with a primitive phenotype were present throughout the culture and also underwent a continuous expansion. Moreover, the capacity for multilineage lymphomyeloid differentiation, as well as the recloning capacity of primitive myeloid progenitors, was maintained in culture. With its better proliferative potential as compared with adult sources, FL represents a promising alternative source of HSC and the culture system described here should be useful for clinical applications.

Long-term safety and effectiveness of lopinavir/ritonavir in antiretroviral-experienced HIV-1-infected children.

AIM: To evaluate the long-term safety and effectiveness of lopinavir/ritonavir (LPV/r) in a population-based cohort of HIV-1-infected children. METHODS: All children enrolled in the Swiss Mother and Child HIV Cohort Study, treated with LPV/r-based combination antiretroviral treatment (cART) between November 2000 and October 2008, were included. RESULTS: 88 children (25 (28%) protease inhibitor (PI)-naive, 16 (18%) ART-naive) were analysed (251 patient-years on LPV/r). After 48 weeks on LPV/r, 70 children had a median (interquartile range (IQR)) decrease in HIV-1 viral load of 4.25 log (5.45-3.17; PI-naive, n=17) and 2.53 (3.68-1.38; PI-experienced, n=53). Median (IQR) increase in CD4 count was 429 (203-593; PI-naive) and 177 (21-331; PI-experienced) cells/microl. These effects remained stable throughout 192 weeks for 25 children. Treatment was stopped for viral rebound in seven and suspected toxicity in 12 children. CONCLUSION: Long-term treatment with LPV/r-based cART is safe and effective in HIV-1-infected children.

Impairment of mossy fiber long-term potentiation and associative learning in pituitary adenylate cyclase activating polypeptide type I receptor-deficient Mice

The pituitary adenylate cyclase activating polypeptide (PACAP) type I receptor (PAC1) is a G-protein-coupled receptor binding the strongly conserved neuropeptide PACAP with 1000-fold higher affinity than the related peptide vasoactive intestinal peptide. PAC1-mediated signaling has been implicated in neuronal differentiation and synaptic plasticity. To gain further insight into the biological significance of PAC1-mediated signaling in vivo, we generated two different mutant mouse strains, harboring either a complete or a forebrain-specific inactivation of PAC1. Mutants from both strains show a deficit in contextual fear conditioning, a hippocampus-dependent associative learning paradigm. In sharp contrast, amygdala-dependent cued fear conditioning remains intact. Interestingly, no deficits in other hippocampus-dependent tasks modeling declarative learning such as the Morris water maze or the social transmission of food preference are observed. At the cellular level, the deficit in hippocampus-dependent associative learning is accompanied by an impairment of mossy fiber long-term potentiation (LTP). Because the hippocampal expression of PAC1 is restricted to mossy fiber terminals, we conclude that presynaptic PAC1-mediated signaling at the mossy fiber synapse is involved in both LTP and hippocampus-dependent associative learning.

Influence of the Premium Subsidy on Farmers’ Crop Insurance Coverage Decisions, April 2005

The Agricultural Risk Protection Act greatly increased the expected marginal net benefit of farmers buying high-coverage crop insurance policies by coupling premium subsidies to coverage level. This policy change, combined with cross-sectional variations in expected marginal net benefits of high-coverage policies, is used to estimate the role that premium subsidies play in farmers’ crop insurance decisions. We use county data for corn, soybeans, and wheat to estimate regression equations that are then used to obtain insight into two policy scenarios. We first estimate that eventual adoption of actuarially fair incremental premiums, combined with current coupled subsidies, would increase farmers’ purchase of high-coverage policies by almost 400 percent from 1998 levels across the three crops and two plans of insurance included in the analysis. We then estimate that a return to decoupled subsidies would decrease farmers’ high-coverage purchase decisions by an average of 36 percent.

Effects of long-term averaging of quantitative blood pressure traits on the detection of genetic associations.

Blood pressure (BP) is a heritable, quantitative trait with intraindividual variability and susceptibility to measurement error. Genetic studies of BP generally use single-visit measurements and thus cannot remove variability occurring over months or years. We leveraged the idea that averaging BP measured across time would improve phenotypic accuracy and thereby increase statistical power to detect genetic associations. We studied systolic BP (SBP), diastolic BP (DBP), mean arterial pressure (MAP), and pulse pressure (PP) averaged over multiple years in 46,629 individuals of European ancestry. We identified 39 trait-variant associations across 19 independent loci (p < 5 × 10(-8)); five associations (in four loci) uniquely identified by our LTA analyses included those of SBP and MAP at 2p23 (rs1275988, near KCNK3), DBP at 2q11.2 (rs7599598, in FER1L5), and PP at 6p21 (rs10948071, near CRIP3) and 7p13 (rs2949837, near IGFBP3). Replication analyses conducted in cohorts with single-visit BP data showed positive replication of associations and a nominal association (p < 0.05). We estimated a 20% gain in statistical power with long-term average (LTA) as compared to single-visit BP association studies. Using LTA analysis, we identified genetic loci influencing BP. LTA might be one way of increasing the power of genetic associations for continuous traits in extant samples for other phenotypes that are measured serially over time.

Long-term outcome of preterm infants treated with nasal continuous positive airway pressure

RESUME La ventilation en pression positive continue (Continuous Positive Airway Pressure, CPAP), utilisée pour la première fois chez. les prématurés en 1971, est une technique actuellement très largement utilisée dans les unités néonatales. L'utilisation de la CPAP présente de nombreux avantages à court terme: diminution de la fraction maximale inspirée d'oxygène, de la durée de l'oxygénothérapie, du taux d'intubation et donc du recours à une ventilation mécanique, réduction de l'utilisation des amines, des curares et de la morphine, possible prévention de l'apparition d'une bronchodysplasie pulmonaire, et possibles réductions du nombre d'infections postnatales et des entérocolites nécrosantes. Mais peu d'études existent concernant les effets à long terme de la CPAP sur le neurodéveloppement et la croissance, qui constituent l'objectif de la présente étude. L'utilisation systématique de la CPAP comme alternative à la ventilation mécanique a été introduite à Lausanne en 1998. La population cible de cette étude est constituée des prématurés nés à moins de 32 semaines de gestation ou pesant moins de 1500 g à la naissance; ils ont été systématiquement suivis jusqu'en âge préscolaire dans le cadre de l'étude de cohorte «Unité de Développement, CHUV». L'originalité de ce travail réside dans le fait d'évaluer le neurodéveloppement et la croissance à long terme d'enfants prématurés traités préférentiellement avec la CPAP, en comparaison avec un groupe historique contrôle traité par d'autres modes ventilatoires, en tenant compte de nombreux autres paramètres néonataux. Du point de vue du neurodéveloppement, l'usage, de la CPAP montre une tendance à diminuer l'incidence d'hémorragie intraventriculaire et le risque d'avoir un mauvais neurodéveloppement à 6 mois. Ces résultats positifs sur le neurodéveloppement s'estompent à l'âge de 18 mois, où persiste néanmoins une valeur plus élevée du quotient de développement, et disparaissent complètement en âge préscolaire. Du point de vue de la croissance, l'utilisation de la CPAP ne montre aucun effet sur le poids, mais par contre un effet positif sur la taille à 6 et 18 mois et sur le périmètre crânien à 6 mois, 18 mois et en âge préscolaire. Malgré le caractère non randomisé de cette étude, les résultats positifs obtenus ici permettent sans conteste de s'assurer d'une utilisation de la CPAP sans effet négatif sur le neurodéveloppement et la croissance, et fournissent donc un argument supplémentaire pour l'utilisation systématique de la CPAP chez les prématurés.

Incidence and risk factors for Contegra graft infection following right ventricular outflow tract reconstruction: long-term results.

OBJECTIVES: The aim of this study was to evaluate the risk factors associated with Contegra graft (Medtronic Minneapolis, MN, USA) infection after reconstruction of the right ventricular outflow tract. METHODS: One hundred and six Contegra grafts were implanted between April 1999 and April 2010 for the Ross procedure (n = 46), isolated pulmonary valve replacement (n = 32), tetralogy of Fallot (n = 24), double-outlet right ventricle (n = 7), troncus arteriosus (n = 4), switch operation (n = 1) and redo of pulmonary valve replacement (n = 2). The median age of the patients was 13 years (range 0-54 years). A follow-up was completed in all cases with a median duration of 7.6 years (range 1.7-12.7 years). RESULTS: There were 3 cases of in-hospital mortality. The survival rate during 7 years was 95.7%. Despite the lifelong endocarditis prophylaxis, Contegra graft infection was diagnosed in 12 (11.3%) patients at a median time of 4.4 years (ranging from 0.4 to 8.7 years). Univariate analysis of preoperative, perioperative and postoperative variables was performed and the following risk factors for time to infection were identified: female gender with a hazard ratio (HR) of 0.19 (P = 0.042), systemic-to-pulmonary shunt (HR 6.46, P < 0.01), hypothermia (HR 0.79, P = 0.014), postoperative renal insufficiency (HR 11.97, P = 0.015) and implantation of permanent pacemaker during hospitalization (HR 5.29, P = 0.075). In 2 cases, conservative therapy was successful and, in 10 patients, replacement of the infected valve was performed. The Contegra graft was replaced by a homograft in 2 cases and by a new Contegra graft in 8 cases. Cox's proportional hazard model indicated that time to graft infection was significantly associated with tetralogy of Fallot (HR 0.06, P = 0.01), systemic-to-pulmonary shunt (HR 64.71, P < 0.01) and hypothermia (HR 0.77, P < 0.01). CONCLUSION: Contegra graft infection affected 11.3% of cases in our cohort, and thus may be considered as a frequent entity that can be predicted by both intraoperative and early postoperative factors. After the diagnosis of infection associated with the Contegra graft was confirmed, surgical treatment was the therapy of choice.

Long-term Plan for Concrete Pavement Research and Technology: The CP Road Map, November 2004, Volume 1 of 2

An Iowa State University–led team facilitated development of the CP Road Map. They developed a database of existing research. They gathered input, face to face, from the highway community. They identified gaps in research that became the basis for problem statements, which they organized into a cohesive, strategic research plan.

Long-term Plan for Concrete Pavement Research and Technology: The CP Road Map, November 2004, Volume 2 of 2

An Iowa State University–led team facilitated development of the CP Road Map. They developed a database of existing research. They gathered input, face to face, from the highway community. They identified gaps in research that became the basis for problem statements, which they organized into a cohesive, strategic research plan.

Long-term Plan for Concrete Pavement Research and Technology: The CP Road Map, November 2004

An Iowa State University–led team facilitated development of the CP Road Map. They developed a database of existing research. They gathered input, face to face, from the highway community. They identified gaps in research that became the basis for problem statements, which they organized into a cohesive, strategic research plan.

Intrauterine growth restriction and absent or reverse end-diastolic blood flow in umbilical artery (Doppler class II or III): A retrospective study of short- and long-term fetal morbidity and mortality.

OBJECTIVE: Absent or reverse end-diastolic flow (Doppler II/III) in umbilical artery is correlated with poor perinatal outcome, particularly in intrauterine growth restricted (IUGR) fetuses. The optimal timing of delivery is still controversial. We studied the short- and long-term morbidity and mortality among these children associated with our defined management. STUDY DESIGN: Sixty-nine IUGR fetuses with umbilical Doppler II/III were divided into three groups; Group 1, severe early IUGR, no therapeutic intervention (n = 7); Group 2, fetuses with pathological biophysical profile, immediate delivery (n = 35); Group 3, fetuses for which expectant management had been decided (n = 27). RESULTS: In Group 1, stillbirth was observed after a mean delay of 6.3 days. Group 2 delivered at an average of 31.6 weeks and two died in the neonatal period (6%). In Group 3 after a mean delay of 8 days, average gestational age at delivery was 31.7 weeks; two intra uterine and four perinatal deaths were observed (22%). Long-term follow-up revealed no sequelae in 25/31 (81%) and 15/18 (83%), and major handicap occurred in 1 (3%) and 2 patients (11%), respectively, for Groups 2 and 3. CONCLUSION: Fetal mortality was observed in 22% of this high risk group. After a mean period of follow-up of 5 years, 82% of infants showed no sequelae. According to our management, IUGR associated with umbilical Doppler II or III does not show any benefit from an expectant management in term of long-term morbidity.

Short term impact of Tribulus terrestris intake on doping control analysis of endogenous steroids.

Tribulus terrestris is a nutritional supplement highly debated regarding its physiological and actual effects on the organism. The main claimed effect is an increase of testosterone anabolic and androgenic action through the activation of endogenous testosterone production. Even if this biological pathway is not entirely proven, T. terrestris is regularly used by athletes. Recently, the analysis of two female urine samples by GC/C/IRMS (gas chromatography/combustion/isotope-ratio-mass-spectrometry) conclusively revealed the administration of exogenous testosterone or its precursors, even if the testosterone glucuronide/epitestosterone glucuronide (T/E) ratio and steroid marker concentrations were below the cut-off values defined by World Anti-Doping Agency (WADA). To argue against this adverse analytical finding, the athletes recognized having used T. terrestris in their diet. In order to test this hypothesis, two female volunteers ingested 500 mg of T. terrestris, three times a day and for two consecutive days. All spot urines were collected during 48 h after the first intake. The (13)C/(12)C ratio of ketosteroids was determined by GC/C/IRMS, the T/E ratio and DHEA concentrations were measured by GC/MS and LH concentrations by radioimmunoassay. None of these parameters revealed a significant variation or increased above the WADA cut-off limits. Hence, the short-term treatment with T. terrestris showed no impact on the endogenous testosterone metabolism of the two subjects.

Alliance patterns over the course of short-term dynamic psychotherapy: The shape of productive relationships.

The shape of alliance processes over the course of psychotherapy has already been studied in several process-outcome studies on very brief psychotherapy. The present study applies the shape-of-change methodology to short-term dynamic psychotherapies and complements this method with hierarchical linear modeling. A total of 50 psychotherapies of up to 40 sessions were included. Alliance was measured at the end of each session. The results indicate that a linear progression model is most adequate. Three main patterns were found: stable, linear, and quadratic growth. The linear growth pattern, along with the slope parameter, was related to treatment outcome. This study sheds additional light on alliance process research, underscores the importance of linear alliance progression for outcome, and also fosters a better understanding of its limitations.