Molecular Mechanisms of Vascular Disease in Patients with Rare Variants in MYH11

Thoracic aortic aneurysms and dissections (TAAD) are the primary disease affecting the thoracic ascending aorta, with an incidence rate of 10.4/100,000. Although about 20% of patients carry a mutation in a single gene that causes their disease, the remaining 80% of patients may also have genetic factors that increase their risk for developing TAAD. Many of the genes that predispose to TAAD encode proteins involved in smooth muscle cell (SMC) contraction and the disease-causing mutations are predicted to disrupt contractile function. SMCs are the predominant cell type in the ascending aortic wall. Mutations in MYH11, encoding the smooth muscle specific myosin heavy chain, are a rare cause of inherited TAAD. However, rare but recurrent non-synonymous variants in MYH11 are present in the general population but do not cause inherited TAAD. The goal of this study was to assess the potential role of these rare variants in vascular diseases. Two distinct variants were selected: the most commonly seen rare variant, MYH11 R247C, and a duplication of the chromosomal region spanning the MYH11 locus at 16p13.1. Genetic analyses indicated that both of these variants were significantly enriched in patients with TAAD compared with controls. A knock-in mouse model of the Myh11 R247C rare variant was generated, and these mice survive and reproduce normally. They have no structural abnormalities of the aorta or signs of aortic disease, but do have decreased aortic contractility. Myh11R247C/R247C mice also have increased proliferative response to vascular injury in vivo and increased proliferation of SMCs in vitro. Myh11R247C/R247C SMCs have decreased contractile gene and protein expression and are dedifferentiated. In fibroblasts, myosin force generation is required for maturation of focal adhesions, and enhancers of RhoA activity replace enhancers of Rac1 activity as maturation occurs. Consistent with these previous findings, focal adhesions are smaller in Myh11R247C/R247C SMCs, and there is decreased RhoA activation. A RhoA activator (CN03) rescues the dedifferentiated phenotype of Myh11R247C/R247C SMCs. Myh11R247C/R247C mice were bred with an existing murine model of aneurysm formation, the Acta2-/- mouse. Over time, mice carrying the R247C allele in conjunction with heterozygous or homozygous loss of Acta2 had significantly increased aortic diameter, and a more rapid accumulation of pathologic markers. These results suggest that the Myh11 R247C rare variant acts as a modifier gene increasing the risk for and severity of TAAD in mice. In patients with 16p13.1 duplications, aortic MYH11 expression is increased, but there is no corresponding increase in smooth muscle myosin heavy chain protein. Using SMCs that overexpress Myh11, we identified alterations in SMC phenotype leading to excessive protein turnover. All contractile proteins, not just myosin, are affected, and the proteins are turned over by autophagic degradation. Surprisingly, these cells are also more contractile compared with wild-type SMCs. The results described in this dissertation firmly establish that rare variants in MYH11 significantly affect the phenotype of SMCs. Further, the data suggests that these rare variants do increase the risk of TAAD via pathways involving altered SMC phenotype and contraction. Therefore, this study validates that these rare genetic variants alter vascular SMCs and provides model systems to explore the contribution of rare variants to disease.

Potential of RapidEye image data for sediment monitoring of the German tidal flats

The Wadden Sea is located in the southeastern part of the North Sea forming an extended intertidal area along the Dutch, German and Danish coast. It is a highly dynamic and largely natural ecosystem influenced by climatic changes and anthropogenic use of the North Sea. Changes in the environment of the Wadden Sea, natural or anthropogenic origin, cannot be monitored by the standard measurement methods alone, because large-area surveys of the intertidal flats are often difficult due to tides, tidal channels and unstable underground. For this reason, remote sensing offers effective monitoring tools. In this study a multi-sensor concept for classification of intertidal areas in the Wadden Sea has been developed. Basis for this method is a combined analysis of RapidEye (RE) and TerraSAR-X (TSX) satellite data coupled with ancillary vector data about the distribution of vegetation, mussel beds and sediments. The classification of the vegetation and mussel beds is based on a decision tree and a set of hierarchically structured algorithms which use object and texture features. The sediments are classified by an algorithm which uses thresholds and a majority filter. Further improvements focus on radiometric enhancement and atmospheric correction. First results show that we are able to identify vegetation and mussel beds with the use of multi-sensor remote sensing. The classification of the sediments in the tidal flats is a challenge compared to vegetation and mussel beds. The results demonstrate that the sediments cannot be classified with high accuracy by their spectral properties alone due to their similarity which is predominately caused by their water content.

Bird count in Mastbos forest (Breda, Netherlands) in the years 1950 to 1981

Notes from Henrik de Nie: The project started as a phenological study in cooperation with the (Dutch) meteorological institute (KNMI) to register the time of arrival of Fitis and Tjiftaf. During 1951 to 1969 he went every day to the wood (except 1966, in this year his wife died). Thereafter he went no more daily, but because he knew the wood very well and he was free to choice the day on which he did a survey, therefore he choose days with relatively good weather. He did not observe very common bird species, maybe because they are dependent on nest boxes and he did not want to be dependent on the management of the nest box-people (in fact I forgot precisely his arguments, and now I cannot ask him this): Common Starling; Eurasian Tree Sparrow (not common); Great Tit; Eurasian Blue Tit Pieter mentioned 14 species that scored many zero values or only one observation: Stock Dove; Common Cuckoo; Lesser Spotted Woodpecker; Eurasian Golden Oriole; Eurasian Nuthatch; Short-toed Treecreeper; Common Nightingale; Marsh Warbler; Lesser Whitethroat; Goldcrest; Common Firecrest (after 1970 he had difficulties in hearing these two species); Spotted Flycatcher; Eurasian Bullfinch; Black Woodpecker He also mentioned species that he found much fewer as: European Greenfinch; European Pied Flycatcher; Long-eared Owl; Red Crossbill; Sedge Warbler; Icterine Warbler; Eurasian Woodcock; Eurasian Siskin; European Green Woodpecker; Great Spotted Woodpecker; Eurasian Hobby; Western Barn Owl; Woodlark; Common Wood Pigeon; Little Owl; European Crested Tit; Hawfinch. But for these species I think that observations are strongly dependent on the number of visits to the wood. Also here, many zeros and few 1 x during the whole series of visits.

Changes in Scleral Architecture in Chronic Vogt-Koyanagi-Harada Disease.

PURPOSE: To describe scleral changes in chronic VKH. METHODS: Medical records of patients with chronic VKH were retrospectively reviewed. Change of scleral architecture was defined as progressive posterior bowing on OCT, axial length elongation, and/or increased myopia more than -1.0 D, not explicable by other etiologies. RESULTS: In total, 28 eyes (16 patients) with mean age of disease onset 32.5 ± 14.0 years were included in the study. Disease duration was 15.1 ± 10.2 years. Eight eyes (28.6%) showed progressive scleral architectural changes. Five eyes (18%) developed scleral changes on OCT, not seen on prior imaging (2-12 years earlier). One eye had posterior bowing on OCT with increased axial length, both eyes of a bilateral pseudophake developed increased myopia with increased axial length. Well-circumscribed chorioretinal atrophy within the arcade was associated with progressive scleral change. CONCLUSIONS: Progressive scleral change may develop as a late complication of VKH. The association with well-circumscribed chorioretinal atrophy suggests that chronic choroidal inflammation may be responsible.

Politics of the 1945 Constitution:Democratization and Its Impact on Political Institutions in Indonesia

Introduction : Before 1998, no one could think about the amendment of the 1945 Constitution. The 1945 Constitution was a product of nationalist who had hard fought for independence from the Dutch colonization. This historical background made it the symbol of independence of the Indonesian nation. Thus, it has been considered as forbidden to touch contents of the 1945 Constitution whereas political leaders have legitimized their authoritarian rulership by utilizing a symbolic character of the Constitution. With the largest political turmoil since its independence, that is, a breakdown of authoritarian regime and democratic transformation in 1998-1999, however, a myth of the "sacred and inviolable" constitution has disappeared. A new theme has then aroused: how can the 1945 Constitution be adapted for a new democratic regime in Indonesia? 　　　The Indonesian modern state has applied the 1945 Constitution as the basic law since its independence in 1945, except for around 10 years in the 1950s. In the period of independence struggle, contrary to the constitutional provision that a kind of presidential system is employed, a cabinet responsible for the Central National Committee was installed. Politics under this institution was in practice a parliamentary system of government. After the Dutch transferred sovereignty to Indonesia in 1949, West European constitutionalism and party politics under a parliamentary system was fully adopted with the introduction of two new constitutions: the 1949 Constitution of Federal Republic of Indonesia and the 1950 Provisional Constitution of Republic of Indonesia. Since a return from the 1950 Constitution to the 1945 Constitution was decided with the Presidential Decree in 1959, the 1945 Constitution had supported two authoritarian regimes of Soekarno's "Guided Democracy" and Soeharto's "New Order" as a legal base. When the 32-year Soeharto's government fell down and democratization started in 1998, the 1945 Constitution was not replaced with a new one, as seen in many other democratizing countries, but successively reformed to adapt itself to a new democratic regime. In the result of four constitutional amendments in 1999-2002, political institutions in Indonesia are experiencing a transformation from an authoritative structure, in which the executive branch monopolized power along with incompetent legislative and judicial branches, to a modern democratic structure, in which the legislative branch can maintain predominance over the executive. However, as observed that President Abdurrahman Wahid, the first president ever elected democratically in Indonesian history, was impeached after one and a half years in office, democratic politics under a new political institution has never been stable. 　　　Under the 1945 Constitution, how did authoritarian regimes maintain stability? Why can a democratic regime not achieve its stability? What did the two constitutional amendments in the process of democratization change? In the first place, how did the political institutions stipulated by the 1945 Constitution come out? Through answering the above questions, this chapter intends to survey the historical continuity and change of political institutions in Indonesia along with the 1945 Constitutions and to analyze impact of regime transformation on political institutions. First, we examine political institutions stipulated by the original 1945 Constitution as well as historical and philosophical origins of the constitution. Second, we search constitutional foundations in the 1945 Constitution that made it possible for Soekarno and Soeharto to establish and maintain authoritarian regimes. Third, we examine contents of constitutional amendments in the process of democratization since 1998. Fourth, we analyze new political dynamics caused by constitutional changes, looking at the impeachment process of President Abdurrahman Wahid. Finally, we consider tasks faced by Indonesia that seeks to establish a stable democracy.

Monitoring Neurological disease in Phonation

It is well known that many neurological diseases leave a fingerprint in voice and speech production. The dramatic impact of these pathologies in life quality is a growing concert. Many techniques have been designed for the detection, diagnose and monitoring the neurological disease. Most of them are costly or difficult to extend to primary services. The present paper shows that some neurological diseases can be traced a the level of voice production. The detection procedure would be based on a simple voice test. The availability of advanced tools and methodologies to monitor the organic pathology of voice would facilitate the implantation of these tests. The paper hypothesizes some of the underlying mechanisms affecting the production of voice and presents a general description of the methodological foundations for the voice analysis system which can estimate correlates to the neurological disease. A case of study is presented from spasmodic dysphonia to illustrate the possibilities of the methodology to monitor other neurological problems as well.

Neurological Disease Detection and Monitoring from Voice Production

The dramatic impact of neurological degenerative pathologies in life quality is a growing concern. It is well known that many neurological diseases leave a fingerprint in voice and speech production. Many techniques have been designed for the detection, diagnose and monitoring the neurological disease. Most of them are costly or difficult to extend to primary attention medical services. Through the present paper it will be shown how some neurological diseases can be traced at the level of phonation. The detection procedure would be based on a simple voice test. The availability of advanced tools and methodologies to monitor the organic pathology of voice would facilitate the implantation of these tests. The paper hypothesizes that some of the underlying mechanisms affecting the production of voice produce measurable correlates in vocal fold biomechanics. A general description of the methodological foundations for the voice analysis system which can estimate correlates to the neurological disease is shown. Some study cases will be presented to illustrate the possibilities of the methodology to monitor neurological diseases by voice

Data mining for the diagnosis of type 2 diabetes

Diabetes is the most common disease nowadays in all populations and in all age groups. diabetes contributing to heart disease, increases the risks of developing kidney disease, blindness, nerve damage, and blood vessel damage. Diabetes disease diagnosis via proper interpretation of the diabetes data is an important classification problem. Different techniques of artificial intelligence has been applied to diabetes problem. The purpose of this study is apply the artificial metaplasticity on multilayer perceptron (AMMLP) as a data mining (DM) technique for the diabetes disease diagnosis. The Pima Indians diabetes was used to test the proposed model AMMLP. The results obtained by AMMLP were compared with decision tree (DT), Bayesian classifier (BC) and other algorithms, recently proposed by other researchers, that were applied to the same database. The robustness of the algorithms are examined using classification accuracy, analysis of sensitivity and specificity, confusion matrix. The results obtained by AMMLP are superior to obtained by DT and BC.

An ICT-based closed-loop model to improve care for heart failure patients

Durante las últimas décadas se ha producido un fenómeno global de envejecimiento en la población. Esta tendencia se puede observar prácticamente en todos los países del mundo y se debe principalmente a los avances en la medicina, y a los descensos en las tasas de fertilidad y mortalidad. El envejecimiento de la población tiene un gran impacto en la salud de los ciudadanos, y a menudo es la causa de aparición de enfermedades crónicas. Este tipo de enfermedades supone una amenaza y una carga importantes para la sociedad, especialmente en aspectos como la mortalidad o los gastos en los sistemas sanitarios. Entre las enfermedades cardiovasculares, la insuficiencia cardíaca es probablemente la condición con mayor prevalencia y afecta a 23-26 millones de personas en todo el mundo. Normalmente, la insuficiencia cardíaca presenta un mal pronóstico y una tasa de supervivencia bajas, en algunos casos peores que algún tipo de cáncer. Además, suele ser la causa de hospitalizaciones frecuentes y es una de las enfermedades más costosas para los sistemas sanitarios. La tendencia al envejecimiento de la población y la creciente incidencia de las enfermedades crónicas están llevando a una situación en la que los sistemas de salud no son capaces de hacer frente a la demanda de la sociedad. Los servicios de salud existentes tendrán que adaptarse para ser efectivos y sostenibles en el futuro. Es necesario identificar nuevos paradigmas de cuidado de pacientes, así como mecanismos para la provisión de servicios que ayuden a transformar estos sistemas sanitarios. En este contexto, esta tesis se plantea la búsqueda de soluciones, basadas en las Tecnologías de la Información y la Comunicación (TIC), que contribuyan a realizar la transformación en los sistemas sanitarios. En concreto, la tesis se centra en abordar los problemas de una de las enfermedades con mayor impacto en estos sistemas: la insuficiencia cardíaca. Las siguientes hipótesis constituyen la base para la realización de este trabajo de investigación: 1. Es posible definir un modelo basado en el paradigma de lazo cerrado y herramientas TIC que formalice el diseño de mejores servicios para pacientes con insuficiencia cardíaca. 2. El modelo de lazo cerrado definido se puede utilizar para definir un servicio real que ayude a gestionar la insuficiencia cardíaca crónica. 3. La introducción, la adopción y el uso de un servicio basado en el modelo definido se traducirá en mejoras en el estado de salud de los pacientes que sufren insuficiencia cardíaca. a. La utilización de un sistema basado en el modelo de lazo cerrado definido mejorará la experiencia del usuario de los pacientes. La definición del modelo planteado se ha basado en el estándar ISO / EN 13940- Sistema de conceptos para dar soporte a la continuidad de la asistencia. Comprende un conjunto de conceptos, procesos, flujos de trabajo, y servicios como componentes principales, y representa una formalización de los servicios para los pacientes con insuficiencia cardíaca. Para evaluar el modelo definido se ha definido un servicio real basado en el mismo, además de la implementación de un sistema de apoyo a dicho servicio. El diseño e implementación de dicho sistema se realizó siguiendo la metodología de Diseño Orientado a Objetivos. El objetivo de la evaluación consistía en investigar el efecto que tiene un servicio basado en el modelo de lazo cerrado sobre el estado de salud de los pacientes con insuficiencia cardíaca. La evaluación se realizó en el marco de un estudio clínico observacional. El análisis de los resultados ha comprendido métodos de análisis cuantitativos y cualitativos. El análisis cuantitativo se ha centrado en determinar el estado de salud de los pacientes en base a datos objetivos (obtenidos en pruebas de laboratorio o exámenes médicos). Para realizar este análisis se definieron dos índices específicos: el índice de estabilidad y el índice de la evolución del estado de salud. El análisis cualitativo ha evaluado la autopercepción del estado de salud de los pacientes en términos de calidad de vida, auto-cuidado, el conocimiento, la ansiedad y la depresión, así como niveles de conocimiento. Se ha basado en los datos recogidos mediante varios cuestionarios o instrumentos estándar (i.e. EQ-5D, la Escala de Ansiedad y Depresión (HADS), el Cuestionario de Cardiomiopatía de Kansas City (KCCQ), la Escala Holandesa de Conocimiento de Insuficiencia Cardíaca (DHFKS), y la Escala Europea de Autocuidado en Insuficiencia Cardíaca (EHFScBS), así como cuestionarios dedicados no estandarizados de experiencia de usuario. Los resultados obtenidos en ambos análisis, cuantitativo y cualitativo, se compararon con el fin de evaluar la correlación entre el estado de salud objetivo y subjetivo de los pacientes. Los resultados de la validación demostraron que el modelo propuesto tiene efectos positivos en el cuidado de los pacientes con insuficiencia cardíaca y contribuye a mejorar su estado de salud. Asimismo, ratificaron al modelo como instrumento válido para la definición de servicios mejorados para la gestión de esta enfermedad. ABSTRACT During the last decades we have witnessed a global aging phenomenon in the population. This can be observed in practically every country in the world, and it is mainly caused by the advances in medicine, and the decrease of mortality and fertility rates. Population aging has an important impact on citizens’ health and it is often the cause for chronic diseases, which constitute global burden and threat to the society in terms of mortality and healthcare expenditure. Among chronic diseases, Chronic Heart Failure (CHF) or Heart Failure (HF) is probably the one with highest prevalence, affecting between 23 and 26 million people worldwide. Heart failure is a chronic, long-term and serious condition with very poor prognosis and worse survival rates than some type of cancers. Additionally, it is often the cause of frequent hospitalizations and one of the most expensive conditions for the healthcare systems. The aging trends in the population and the increasing incidence of chronic diseases are leading to a situation where healthcare systems are not able to cope with the society demand. Current healthcare services will have to be adapted and redefined in order to be effective and sustainable in the future. There is a need to find new paradigms for patients’ care, and to identify new mechanisms for services’ provision that help to transform the healthcare systems. In this context, this thesis aims to explore new solutions, based on ICT, that contribute to achieve the needed transformation within the healthcare systems. In particular, it focuses on addressing the problems of one of the diseases with higher impact within these systems: Heart Failure. The following hypotheses represent the basis to the elaboration of this research: 1. It is possible to define a model based on a closed-loop paradigm and ICT tools that formalises the design of enhanced healthcare services for chronic heart failure patients. 2. The described closed-loop model can be exemplified in a real service that supports the management of chronic heart failure disease. 3. The introduction, adoption and use of a service based on the outlined model will result in improvements in the health status of patients suffering heart failure. 4. The user experience of patients when utilizing a system based on the defined closed-loop model will be enhanced. The definition of the closed-loop model for health care support of heart failure patients have been based on the standard ISO/EN 13940 System of concepts to support continuity of care. It includes a set of concept, processes and workflows, and services as main components, and it represent a formalization of services for heart failure patients. In order to be validated, the proposed closed-loop model has been instantiated into a real service and a supporting IT system. The design and implementation of the system followed the user centred design methodology Goal Oriented Design. The validation, that included an observational clinical study, aimed to investigate the effect that a service based on the closed-loop model had on heart failure patients’ health status. The analysis of results comprised quantitative and qualitative analysis methods. The quantitative analysis was focused on determining the health status of patients based on objective data (obtained in lab tests or physical examinations). Two specific indexes where defined and considered in this analysis: the stability index and the health status evolution index. The qualitative analysis assessed the self-perception of patients’ health status in terms of quality of life, self-care, knowledge, anxiety and depression, as well as knowledge levels. It was based on the data gathered through several standard instruments (i.e. EQ-5D, the Hospital Anxiety and Depression Scale, the Kansas City Cardiomyopathy Questionnaire, the Dutch Heart Failure Knowledge Scale, and the European Heart Failure Self-care Behaviour Scale) as well as dedicated non-standardized user experience questionnaires. The results obtained in both analyses, quantitative and qualitative, were compared in order to assess the correlation between the objective and subjective health status of patients. The results of the validation showed that the proposed model contributed to improve the health status of the patients and had a positive effect on the patients’ care. It also proved that the model is a valid instrument for designing enhanced healthcare services for heart failure patients.

Inhibition of the electrostatic interaction between β-amyloid peptide and membranes prevents β-amyloid-induced toxicity

The accumulation of β-amyloid peptides (Aβ) into senile plaques is one of the hallmarks of Alzheimer disease. Aggregated Aβ is toxic to cells in culture and this has been considered to be the cause of neurodegeneration that occurs in the Alzheimer disease brain. The discovery of compounds that prevent Aβ toxicity may lead to a better understanding of the processes involved and ultimately to possible therapeutic drugs. Low nanomolar concentrations of Aβ1-42 and the toxic fragment Aβ25-35 have been demonstrated to render cells more sensitive to subsequent insults as manifested by an increased sensitivity to formazan crystals following MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) reduction. Formation of the toxic β-sheet conformation by Aβ peptides is increased by negatively charged membranes. Here we demonstrate that phloretin and exifone, dipolar compounds that decrease the effective negative charge of membranes, prevent association of Aβ1-40 and Aβ25-35 to negatively charged lipid vesicles and Aβ induced cell toxicity. These results suggest that Aβ toxicity is mediated through a nonspecific physicochemical interaction with cell membranes.

The solution structure of the N-terminal domain of α2-macroglobulin receptor-associated protein

The three-dimensional structure of the N-terminal domain (residues 18–112) of α2-macroglobulin receptor-associated protein (RAP) has been determined by NMR spectroscopy. The structure consists of three helices composed of residues 23–34, 39–65, and 73–88. The three helices are arranged in an up-down-up antiparallel topology. The C-terminal 20 residues were shown not to be in a well defined conformation. A structural model for the binding of RAP to the family of low-density lipoprotein receptors is proposed. It defines a role in binding for both the unordered C terminus and the structural scaffold of the core structure. Pathogenic epitopes for the rat disease Heymann nephritis, an experimental model of human membranous glomerulonephritis, have been identified in RAP and in the large endocytic receptor gp330/megalin. Here we provide the three-dimensional structure of the pathogenic epitope in RAP. The amino acid residues known to form the epitope are in a helix–loop–helix conformation, and from the structure it is possible to rationalize the published results obtained from studies of fragments of the N-terminal domain.

Glycosylation differences between the normal and pathogenic prion protein isoforms

Prion protein consists of an ensemble of glycosylated variants or glycoforms. The enzymes that direct oligosaccharide processing, and hence control the glycan profile for any given glycoprotein, are often exquisitely sensitive to other events taking place within the cell in which the glycoprotein is expressed. Alterations in the populations of sugars attached to proteins can reflect changes caused, for example, by developmental processes or by disease. Here we report that normal (PrPC) and pathogenic (PrPSc) prion proteins (PrP) from Syrian hamsters contain the same set of at least 52 bi-, tri-, and tetraantennary N-linked oligosaccharides, although the relative proportions of individual glycans differ. This conservation of structure suggests that the conversion of PrPC into PrPSc is not confined to a subset of PrPs that contain specific sugars. Compared with PrPC, PrPSc contains decreased levels of glycans with bisecting GlcNAc residues and increased levels of tri- and tetraantennary sugars. This change is consistent with a decrease in the activity of N-acetylglucosaminyltransferase III (GnTIII) toward PrPC in cells where PrPSc is formed and argues that, in at least some cells forming PrPSc, the glycosylation machinery has been perturbed. The reduction in GnTIII activity is intriguing both with respect to the pathogenesis of the prion disease and the replication pathway for prions.

The Vibrio cholerae genome contains two unique circular chromosomes

Vibrio cholerae, the etiologic agent of the diarrheal disease cholera, is a Gram-negative bacterium that belongs to the γ subdivision of the family Proteobacteriaceae. The physical map of the genome has been reported, and the genome has been described as a single 3.2-Mb chromosome [Majumder, R., et al. (1996) J. Bacteriol. 178, 1105–1112]. By using pulsed-field gel electrophoresis of genomic DNA immobilized in agarose plugs and digested with the restriction enzymes I-CeuI, SfiI, and NotI, we have also constructed the physical map of V. cholerae. Our analysis estimates the size of the genome at 4.0 Mb, 25% larger than the physical map reported by others. Our most notable finding is, however, that the V. cholerae chromosome appears to be not the single chromosome reported but two unique and separate circular megareplicons.

Vascular tumors in livers with targeted inactivation of the von Hippel–Lindau tumor suppressor

von Hippel–Lindau (VHL) disease is a pleomorphic familial tumor syndrome that is characterized by the development of highly vascularized tumors. Homozygous disruption of the VHL gene in mice results in embryonic lethality. To investigate VHL function in the adult we have generated a conditional VHL null allele (2-lox allele) and null allele (1-lox allele) by Cre-mediated recombination in embryonic stem cells. We show here that mice heterozygous for the 1-lox allele develop cavernous hemangiomas of the liver, a rare manifestation of the human disease. Histologically these tumors were associated with hepatocellular steatosis and focal proliferations of small vessels. To study the cellular origin of these lesions we inactivated VHL tissue-specifically in hepatocytes. Deletion of VHL in the liver resulted in severe steatosis, many blood-filled vascular cavities, and foci of increased vascularization within the hepatic parenchyma. These histopathological changes were similar to those seen in livers from mice heterozygous for the 1-lox allele. Hypoxia-inducible mRNAs encoding vascular endothelial growth factor, glucose transporter 1, and erythropoietin were up-regulated. We thus provide evidence that targeted inactivation of mouse VHL can model clinical features of the human disease and underline the importance of the VHL gene product in the regulation of hypoxia-responsive genes in vivo.

Germ line deletion of the CD1 locus exacerbates diabetes in the NOD mouse

Quantitative and qualitative defects in CD1-restricted natural killer T cells have been reported in several autoimmune-prone strains of mice, including the nonobese diabetic (NOD) mouse. These defects are believed to be associated with the emergence of spontaneous autoimmunity. Here we demonstrate that both CD1d-null NOD and CD1d-null NOD/BDC2.5 T cell receptor transgenic mice have an accelerated onset and increased incidence of diabetes when compared with CD1d+/− and CD1d+/+ littermates. The acceleration of disease did not seem to result from changes in the T helper (Th)1/Th2 balance because lymphocytes purified from lymphoid organs and pancreatic islets of wild-type and CD1d-null mice secreted equivalent amounts of IFN-γ and IL-4 after stimulation. In contrast, the pancreata of CD1d-null mice harbored significantly higher numbers of activated memory T cells expressing the chemokine receptor CCR4. Notably, the presence of these T cells was associated with immunohistochemical evidence of increased destructive insulitis. Thus, CD1d-restricted T cells are critically important for regulation of the spontaneous disease process in NOD mice.