Report on the driver’s license issuance program administered by the Department of Transportation and County Treasurers’ offices for the year ended June 30, 2009

Report on the driver’s license issuance program administered by the Department of Transportation and County Treasurers’ offices for the year ended June 30, 2009

A cell therapy approach for erythropoietin delivery using encapsulated human primary fibroblasts

Summary Cell therapy has emerged as a strategy for the treatment of various human diseases. Cells can be transplanted considering their morphological and functional properties to restore a tissue damage, as represented by blood transfusion, bone marrow or pancreatic islet cells transplantation. With the advent of the gene therapy, cells also were used as biological supports for the production of therapeutic molecules that can act either locally or at distance. This strategy represents the basis of ex vivo gene therapy characterized by the removal of cells from an organism, their genetic modification and their implantation into the same or another individual in a physiologically suitable location. The tissue or biological function damage dictates the type of cells chosen for implantation and the required function of the implanted cells. The general aim of this work was to develop an ex vivo gene therapy approach for the secretion of erythropoietin (Epo) in patients suffering from Epo-responsive anemia, thus extending to humans, studies previously performed with mouse cells transplanted in mice and rats. Considering the potential clinical application, allogeneic primary human cells were chosen for practical and safety reasons. In contrast to autologous cells, the use of allogeneic cells allows to characterize a cell lineage that can be further transplanted in many individuals. Furthermore allogeneic cells avoid the potential risk of zoonosis encountered with xenogeneic cells. Accordingly, the immune reaction against this allogeneic source was prevented by cell macro- encapsulation that prevents cell-to-cell contact with the host immune system and allows to easy retrieve the implanted device. The first step consisted in testing the survival of various human primary cells that were encapsulated and implanted for one month in the subcutaneous tissue of immunocompetent and naturally or therapeutically immunodepressed mice, assuming that xenogeneic applications constitute a stringent and representative screening before human transplantation. A fibroblast lineage from the foreskin of a young donor, DARC 3.1 cells, showed the highest mean survival score. We have then performed studies to optimize the manufacturing procedures of the encapsulation device for successful engraftment. The development of calcifications on the polyvinyl alcohol (PVA) matrix serving as a scaffold for enclosed cells into the hollow fiber devices was reported after one month in vivo. Various parameters, including matrix rinsing solutions, batches of PVA and cell lineages were assessed for their respective role in the development of the phenomenon. We observed that the calcifications could be totally prevented by using ultra-pure sterile water instead of phosphate buffer saline solution in the rinsing procedure of the PVA matrix. Moreover, a higher lactate dehydrogenase activity of the cells was found to decrease calcium depositions due to more acidic microenvironment, inhibiting the calcium precipitation. After the selection of the appropriate cell lineage and the optimization of encapsulation conditions, a retroviral-based approach was applied to DARC 3.1 fibroblasts for the transduction of the human Epo cDNA. Various modifications of the retroviral vector and the infection conditions were performed to obtain clinically relevant levels of human Epo. The insertion of a post-transcriptional regulatory element from the woodchuck hepatitis virus as well as of a Kozak consensus sequence led to a 7.5-fold increase in transgene expression. Human Epo production was further optimized by increasing the multiplicity of infection and by selecting high producer cells allowing to reach 200 IU hEpo/10E6 cells /day. These modified cells were encapsulated and implanted in vivo in the same conditions as previously described. All the mouse strains showed a sustained increase in their hematocrit and a high proportion of viable cells were observed after retrieval of the capsules. Finally, in the perspective of human application, a syngeneic model using encapsulated murine myoblasts transplanted in mice was realized to investigate the roles of both the host immune response and the cells metabolic requirements. Various loading densities and anti-inflammatory as well as immunosuppressive drugs were studied. The results showed that an immune process is responsible of cell death in capsules loaded at high cell density. A supporting matrix of PVA was shown to limit the cell density and to avoid early metabolic cell death, preventing therefore the immune reaction. This study has led to the development of encapsulated cells of human origin producing clinically relevant amounts of human EPO. This work resulted also to the optimization of cell encapsulation technical parameters allowing to begin a clinical application in end-stage renal failure patients. Résumé La thérapie cellulaire s'est imposée comme une stratégie de traitement potentiel pour diverses maladies. Si l'on considère leur morphologie et leur fonction, les cellules peuvent être transplantées dans le but de remplacer une perte tissulaire comme c'est le cas pour les transfusions sanguines ou les greffes de moelle osseuse ou de cellules pancréatiques. Avec le développement de la thérapie génique, les cellules sont également devenues des supports biologiques pour la production de molécules thérapeutiques. Cette stratégie représente le fondement de la thérapie génique ex vivo, caractérisée par le prélèvement de cellules d'un organisme, leur modification génétique et leur implantation dans le même individu ou dans un autre organisme. Le choix du type de cellule et la fonction qu'elle doit remplir pour un traitement spécifique dépend du tissu ou de la fonction biologique atteintes. Le but général de ce travail est de développer .une approche par thérapie génique ex vivo de sécrétion d'érythropoïétine (Epo) chez des patients souffrant d'anémie, prolongeant ainsi des travaux réalisés avec des cellules murines implantées chez des souris et des rats. Dans cette perpective, notre choix s'est porté sur des cellules humaines primaires allogéniques. En effet, contrairement aux cellules autologues, une caractérisation unique de cellules allogéniques peut déboucher sur de nombreuses applications. Par ailleurs, l'emploi de cellules allogéniques permet d'éviter les riques de zoonose que l'on peut rencontrer avec des cellules xénogéniques. Afin de protéger les cellules allogéniques soumises à une réaction immunitaire, leur confinement dans des macro-capsules cylindriques avant leur implantation permet d'éviter leur contact avec les cellules immunitaires de l'hôte, et de les retrouver sans difficulté en cas d'intolérance ou d'effet secondaire. Dans un premier temps, nous avons évalué la survie de différentes lignées cellulaires humaines primaires, une fois encapsulées et implantées dans le tissu sous-cutané de souris, soit immunocompétentes, soit immunodéprimées naturellement ou par l'intermédiaire d'un immunosuppresseur. Ce modèle in vivo correspond à des conditions xénogéniques et représente par conséquent un environnement de loin plus hostile pour les cellules qu'une transplantation allogénique. Une lignée fibroblastique issue du prépuce d'un jeune enfant, nommée DARC 3 .1, a montré une remarquable résistance avec un score de survie moyen le plus élevé parmi les lignées testées. Par la suite, nous nous sommes intéressés aux paramètres intervenant dans la réalisation du système d'implantation afin d'optimaliser les conditions pour une meilleure adaptation des cellules à ce nouvel environnement. En effet, en raison de l'apparition, après un mois in vivo, de calcifications au niveau de la matrice de polyvinyl alcohol (PVA) servant de support aux cellules encapsulées, différents paramètres ont été étudiés, tels que les procédures de fabrication, les lots de PVA ou encore les lignées cellulaires encapsulées, afin de mettre en évidence leur rôle respectif dans la survenue de ce processus. Nous avons montré que l'apparition des calcifications peut être totalement prévenue par l'utilisation d'eau pure au lieu de tampon phosphaté lors du rinçage des matrices de PVA. De plus, nous avons observe qu'un taux de lactate déshydrogénase cellulaire élevé était corrélé avec une diminution des dépôts de calcium au sein de la matrice en raison d'un micro-environnement plus acide inhibant la précipitation du calcium. Après sélection de la lignée cellulaire appropriée et de l'optimisation des conditions d'encapsulation, une modification génétique des fibroblastes DARC 3.1 a été réalisée par une approche rétrovirale, permettant l'insertion de l'ADN du gène de l'Epo dans le génome cellulaire. Diverses modifications, tant au niveau génétique qu'au niveau des conditions d'infection, ont été entreprises afin d'obtenir des taux de sécrétion d'Epo cliniquement appropriés. L'insertion dans la séquence d'ADN d'un élément de régulation post¬transcriptionnelle dérivé du virus de l'hépatite du rongeur (« woodchuck ») ainsi que d'une séquence consensus appelée « Kozak » ont abouti à une augmentation de sécrétion d'Epo 7.5 fois plus importante. De même, l'optimisation de la multiplicité d'infection et la sélection plus drastique des cellules hautement productrices ont permis finalement d'obtenir une sécrétion correspondant à 200 IU d'Epo/10E6 cells/jour. Ces cellules génétiquement modifiées ont été encapsulées et implantées in vivo dans les mêmes conditions que celles décrites plus haut. Toutes les souris transplantées ont montré une augmentation significative de leur hématocrite et une proportion importante de cellules présentait une survie conservée au moment de l'explantation des capsules. Finalement, dans la perspective d'une application humaine, un modèle syngénique a été proposé, basé sur l'implantation de myoblastes murins encapsulés dans des souris, afin d'investiguer les rôles respectifs de la réponse immunitaire du receveur et des besoins métaboliques cellulaires sur leur survie à long terme. Les cellules ont été encapsulées à différentes densités et les animaux transplantés se sont vus administrer des injections de molécules anti-inflammatoires ou immunosuppressives. Les résultats ont démontré qu'une réaction immunologique péri-capsulaire était à la base du rejet cellulaire dans le cas de capsules à haute densité cellulaire. Une matrice de PVA peut limiter cette densité et éviter une mort cellulaire précoce due à une insuffisance métabolique et par conséquent prévenir la réaction immunitaire. Ce travail a permis le développement de cellules encapsulées d'origine humaine sécrétant des taux d'Epo humaine adaptés à des traitements cliniques. De pair avec l'optimalisation des paramètres d'encapsulation, ces résultats ont abouti à l'initiation d'une application clinique destinée à des patients en insuffisance rénale terminale.

Strategic behaviours in healthcare : evidence from primary care and pharmaceutical markets

We analyse the strategic behaviours of agents in a market through the appropriate¬ness of their skills to the market. If agents' skills are well adapted to market and they can reach their target, they will not need to adopt strategic behaviours. The agents will behave as selfish individuals. However, if their skills are not well adapted and they cannot attain their target alone, they will adopt strategic behaviours to reach their objectives. These behaviours will have a different impact on the utilities of other agents, depending on the skills and the objectives of the agent. If these agents need other agents to reach their objectives, they will behave as altruistic individuals who internalise the utilities of other agents in reaching their objectives and will adopt cooperative behaviours. However, if these agents fear that other agents could prevent them from reaching their target because they can foresee that the skills of other agents are better adapted than their own skills, the agents will then behave as predator individuals and will adopt destructive behaviours to attain their objective. It is in the interests of these agents to manipulate information to increase disorder and dissimulate their lack of skills. They will reproduce the strategies of animals that modify their appearance to escape predators or simulate being bait to attract their prey. These agents will seek to induce chaos into the behaviours of other agents to amplify the impact of their strategies. The appropriateness of skills to the market allows an understanding of the emer-gence of networks and associated strategies. The members of a networks are inputs who are excluded when their costs are higher than their benefits. A network simul-taneously allows cooperation and selfish, predatory behaviours among its members. A network may adopt informational strategies when seeking to become the leader in a market or when it cannot survive. The creation of networks and the manipulation of information are two overlapping evolutionary strategies, with the first strategy favouring the second. In our model, an agent does not behave like a firm that aims only to maximise the profits of the firm but rather as a member of a network who adopts strategic behaviours as a function of the interests of this network. If his skills are well adapted to the market and he can innovate, he will not invest in erroneous input; in contrast, if his skills are not adapted, the agent will invest in the erroneous input of information into the market in order to survive. Therefore, when any informational asymmetries between the agents and their principals characterise the market, the price cannot be the main element that allows equilibrium to be reached in the market; instead, the appropriateness of skills to the market enables equilibrium. We will now apply these hypotheses to explain the strategic behaviours of physicians and pharmaceutical companies.

Backseat drivers: the hidden influence of microbial viruses on disease.

Because viral replication depends on the vigour of its host, many viruses have evolved incentives of fitness to pay their keep. When the viral host is a human pathogen, these fitness factors can surface as virulence: creating a Russian doll of pathogenesis where pathogens within pathogens complicate the disease process. Microbial viruses can even be independently immunogenic, as we recently reported for leishmania-virus. Thus, the incidence of this 'hyperpathogenism' is becoming an important clinical consideration and by appreciating the microbial-virus as a backseat driver of human disease, we could exploit its presence as a diagnostic biomarker and molecular target for therapeutic intervention. Here we discuss the prevalence of clinically relevant hyperpathogenism as well as the environmental sanctuaries that breed it.

Geoecological drivers of cerrado heterogeneity and 13C natural abundance in oxisols after land-use change

The 13C natural abundance technique was applied to study C dynamics after land-use change from native savanna to Brachiaria, Pinus, and Eucalyptus in differently textured Cerrado Oxisols. But due to differences in the d13C signatures of subsoils under native savanna and under introduced species, C substitution could only be calculated based on results of cultivated soils nearby. It was estimated that after 20 years, Pinus C had replaced only 5 % of the native C in the 0-1.2 m layer, in which substitution was restricted to the top 0.4 m. Conversely, after 12 years, Brachiaria had replaced 21 % of Cerrado C to a depth of 1.2 m, where substitution decreased only slightly throughout the entire profile. The high d13C values in the subsoils of the cultivated sites led to the hypothesis that the natural vegetation there had been grassland rather than Cerrado sensu stricto, in spite of the comparable soil and site characteristics and the proximity of the studied sites. The hypothesis was tested using aerial photographs of 1964, which showed that the cultivated sites were located on a desiccated runoff head. The vegetation shift to a grass-dominated savanna formation might therefore have occurred in response to waterlogging and reduced soil aeration. A simple model was developed thereof, which ascribes the different Cerrado formations mainly to the plant-available water content and soil aeration. Soil fertility is considered of minor significance only, since at the studied native savanna sites tree density was independent of soil texture or nutrient status.

Comment dépister les syndromes gériatriques au cabinet [How to screen geriatric conditions in primary care?].

The ongoing aging of the population will lead to an increasing prevalence of chronic diseases and functional limitation. Preventative measures need to be promoted to prevent health care utilization and health costs explosion. Among these measures, those aimed at promoting early recognition of chronic conditions associated with functional decline, will have to be reinforced. This paper proposes simple, feasible, and efficient procedures to screen, in primary care practices, common geriatric conditions, as cognitive impairment, gait impairment, hearing and vision impairment or functional limitation.

Report on a special investigation of certain driver’s license collections at the Sioux County Treasurer’s Office for the period July 1, 2009 through May 31, 2010

Report on a special investigation of certain driver’s license collections at the Sioux County Treasurer’s Office for the period July 1, 2009 through May 31, 2010

The impact of transmission clusters on primary drug resistance in newly diagnosed HIV-1 infection.

OBJECTIVES: To monitor HIV-1 transmitted drug resistance (TDR) in a well defined urban area with large access to antiretroviral therapy and to assess the potential source of infection of newly diagnosed HIV individuals. METHODS: All individuals resident in Geneva, Switzerland, with a newly diagnosed HIV infection between 2000 and 2008 were screened for HIV resistance. An infection was considered as recent when the positive test followed a negative screening test within less than 1 year. Phylogenetic analyses were performed by using the maximum likelihood method on pol sequences including 1058 individuals with chronic infection living in Geneva. RESULTS: Of 637 individuals with newly diagnosed HIV infection, 20% had a recent infection. Mutations associated with resistance to at least one drug class were detected in 8.5% [nucleoside reverse transcriptase inhibitors (NRTIs), 6.3%; non-nucleoside reverse transcriptase inhibitors (NNRTIs), 3.5%; protease inhibitors, 1.9%]. TDR (P-trend = 0.015) and, in particular, NNRTI resistance (P = 0.002) increased from 2000 to 2008. Phylogenetic analyses revealed that 34.9% of newly diagnosed individuals, and 52.7% of those with recent infection were linked to transmission clusters. Clusters were more frequent in individuals with TDR than in those with sensitive strains (59.3 vs. 32.6%, respectively; P < 0.0001). Moreover, 84% of newly diagnosed individuals with TDR were part of clusters composed of only newly diagnosed individuals. CONCLUSION: Reconstruction of the HIV transmission networks using phylogenetic analysis shows that newly diagnosed HIV infections are a significant source of onward transmission, particularly of resistant strains, thus suggesting an important self-fueling mechanism for TDR.

Prévention primaire et dépistage chez l'adulte: mise à jour 2006 [Primary prevention and screening in adults: update 2006]

Several diseases can be prevented either by primary prevention, such as immunisation or behavioural counselling, or secondary prevention such as screening. The new clinical recommendations include screening of abdominal aortic aneurysm among male smokers and ex-smokers aged between 65 and 75 years and the extension of breast cancer screening by mammography for women aged between 40 and 49 years, as well as screening for diabetes among patients with hypertension or dyslipidemia.

Target blood pressure attainment with antihypertensive therapy in Swiss primary care.

Introduction: The control of high blood pressure (BP) remains insufficient in developed as well as in developing countries. We conducted a cross-sectional survey to investigate the management of hypertension and the achievement of target BPs in a large population of hypertensive patients treated by Swiss primary care physicians. Methods. Data from 4594 hypertensive patients were collected and assessed for demographic data, mode of treatment and BP achievements for the overall population and for high-risk patients such as diabetics and patients with impaired renal function (CKD patients). Furthermore, we analysed the achieved BP in patients receiving single pill combinations or dual free combinations for the three most commonly prescribed substances. Results. In this large patient population, 84% of patients were receiving an antihypertensive treatment of which 54% showed BP control (< 140/90 mmHg or < 130/80 mmHg for diabetics and patients with CKD). Considering the higher BP target in the elderly, 60.6% of treated patients were on target. In contrast, 28.8% of treated diabetics and 29.7% of patients with impaired renal function met BP goals. Diuretics and blockers of the renin-angiotensin system were the most commonly prescribed substances. High-risk patients and patients at advanced age (≥ 80 years) received dual free combination more frequently than younger patients. The use of diuretics was particularly high because of the prescription of single pill formulations. Differences in the pattern of drug prescription were found according to the linguistic areas. Conclusion. The control of hypertension in the Swiss hypertensive population is relatively high but still insufficient particularly among high cardiovascular risk patients such as diabetics and patients with impaired renal function. A further improvement of BP control could perhaps be achieved with a greater use of single pill combinations particularly in patients with complicated hypertension.

Development and validation of a clinical prediction rule for chest wall syndrome in primary care

Le "Chest wall syndrome" (CWS) est défini comme étant une source bénigne de douleurs thoraciques, localisées sur la paroi thoracique antérieure et provoquées par une affection musculosquelettique. Le CWS représente la cause la plus fréquente de douleurs thoraciques en médecine de premier recours. Le but de cette étude est de développer et valider un score de prédiction clinique pour le CWS. Une revue de la littérature a d'abord été effectuée, d'une part pour savoir si un tel score existait déjà, et d'autre part pour retrouver les variables décrites comme étant prédictives d'un CWS. Le travail d'analyse statistique a été effectué avec les données issues d'une cohorte clinique multicentrique de patients qui avaient consulté en médecine de premier recours en Suisse romande avec une douleur thoracique (59 cabinets, 672 patients). Un diagnostic définitif avait été posé à 12 mois de suivi. Les variables pertinentes ont été sélectionnées par analyses bivariées, et le score de prédiction clinique a été développé par régression logistique multivariée. Une validation externe de ce score a été faite en utilisant les données d'une cohorte allemande (n= 1212). Les analyses bivariées ont permis d'identifier 6 variables caractérisant le CWS : douleur thoracique (ni rétrosternale ni oppressive), douleur en lancées, douleur bien localisée, absence d'antécédent de maladie coronarienne, absence d'inquiétude du médecin et douleur reproductible à la palpation. Cette dernière variable compte pour 2 points dans le score, les autres comptent pour 1 point chacune; le score total s'étend donc de 0 à 7 points. Dans la cohorte de dérivation, l'aire sous la courbe sensibilité/spécificité (courbe ROC) est de 0.80 (95% de l'intervalle de confiance : 0.76-0.83). Avec un seuil diagnostic de > 6 points, le score présente 89% de spécificité et 45% de sensibilité. Parmi tous les patients qui présentaient un CWS (n = 284), 71% (n = 201) avaient une douleur reproductible à la palpation et 45% (n= 127) sont correctement diagnostiqués par le score. Pour une partie (n = 43) de ces patients souffrant de CWS et correctement classifiés, 65 investigations complémentaires (30 électrocardiogrammes, 16 radiographies du thorax, 10 analyses de laboratoire, 8 consultations spécialisées, et une tomodensitométrie thoracique) avaient été réalisées pour parvenir au diagnostic. Parmi les faux positifs (n = 41), on compte trois angors stables (1.8% de tous les positifs). Les résultats de la validation externe sont les suivants : une aire sous la courbe ROC de 0.76 (95% de l'intervalle de confiance : 0.73-0.79) avec une sensibilité de 22% et une spécificité de 93%. Ce score de prédiction clinique pour le CWS constitue un complément utile à son diagnostic, habituellement obtenu par exclusion. En effet, pour les 127 patients présentant un CWS et correctement classifiés par notre score, 65 investigations complémentaires auraient pu être évitées. Par ailleurs, la présence d'une douleur thoracique reproductible à la palpation, bien qu'étant sa plus importante caractéristique, n'est pas pathognomonique du CWS.