Enhanced in vivo angiogenesis within a model tissue engineered construct using biodegradable microspheres containing encapsulated VEGF

Introduction. Tissue engineering techniques offer a potential means to develop a tissue engineered construct (TEC) for the treatment of tissue and organ deficiencies. However, a lack of adequate vascularization is a limiting factor in the development of most viable engineered tissues. Vascular endothelial growth factor (VEGF) could aid in the development of a viable vascular network within TECs. The long-term goals of this research are to develop clinically relevant, appropriately vascularized TECs for use in humans. This project tested the hypothesis that the delivery of VEGF via controlled release from biodegradable microspheres would increase the vascular density and rate of angiogenesis within a model TEC. ^ Materials and methods. Biodegradable VEGF-encapsulated microspheres were manufactured using a novel method entitled the Solid Encapsulation/Single Emulsion/Solvent Extraction technique. Using a PLGA/PEG polymer blend, microspheres were manufactured and characterized in vitro. A model TEC using fibrin was designed for in vivo tissue engineering experimentation. At the appropriate timepoint, the TECs were explanted, and stained and quantified for CD31 using a novel semi-automated thresholding technique. ^ Results. In vitro results show the microspheres could be manufactured, stored, degrade, and release biologically active VEGF. The in vivo investigations revealed that skeletal muscle was the optimal implantation site as compared to dermis. In addition, the TECs containing fibrin with VEGF demonstrated significantly more angiogenesis than the controls. The TECs containing VEGF microspheres displayed a significant increase in vascular density by day 10. Furthermore, TECs containing VEGF microspheres had a significantly increased relative rate of angiogenesis from implantation day 5 to day 10. ^ Conclusions. A novel technique for producing microspheres loaded with biologically active proteins was developed. A defined concentration of microspheres can deliver a quantifiable level of VEGF with known release kinetics. A novel model TEC for in vivo tissue engineering investigations was developed. VEGF and VEGF microspheres stimulate angiogenesis within the model TEC. This investigation determined that biodegradable rhVEGF 165-encapsulated microspheres increased the vascular density and relative rate of angiogenesis within a model TEC. Future applications could include the incorporation of microvascular fragments into the model TEC and the incorporation of specific tissues, such as fat or bone. ^

The combined effect of pravastatin and aspirin on clinical cardiovascular events

The 3-hydroxy-3methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, or statins, can achieve significant reductions in plasma low-density lipoprotein (LDL)-cholesterol levels. Experimental and clinical evidence now shows that some statins interfere with formation of atherosclerotic lesions independent of their hypolipidemic properties. Vulnerable plaque rupture can result in thrombus formation and artery occlusion; this plaque deterioration is responsible for most acute coronary syndromes, including myocardial infarction (MI), unstable angina, and coronary death, as well as coronary heart diseaseequivalent non-hemorrhagic stroke. Inhibition of HMG-CoA reductase has potential pleiotropic effects other than lipid-lowering, as statins block mevalonic acid production, a precursor to cholesterol and numerous other metabolites. Statins' beneficial effects on clinical events may also thus involve nonlipid-related mechanisms that modify endothelial function, inflammatory responses, plaque stability, and thrombus formation. Aspirin, routinely prescribed to post-MI patients as adjunct therapy, may potentiate statins beneficial effects, as aspirin does not compete metabolically with statins but acts similarly on atherosclerotic lesions. Common functions of both medications include inhibition of platelet activity and aggregation, reduction in atherosclerotic plaque macrophage cell count, and prevention of atherosclerotic vessel endothelial dysfunction. The Cholesterol and Recurrent Events (CARE) trial provides an ideal population in which to examine the combined effects of pravastatin and aspirin. Lipid levels, intermediate outcomes, are examined by pravastatin and aspirin status, and differences between the two pravastatin groups are found. A modified Cox proportional-hazards model with aspirin as a time-dependent covariate was used to determine the effect of aspirin and pravastatin on the clinical cardiovascular composite endpoint of coronary heart disease death, recurrent MI or stroke. Among those assigned to pravastatin, use of aspirin reduced the composite primary endpoint by 35%; this result was similar by gender, race, and diabetic status. Older patients demonstrated a nonsignificant 21% reduction in the primary outcome, whereas the younger had a significant reduction of 43% in the composite primary outcome. Secondary outcomes examined include coronary artery bypass graft (38% reduction), nonsurgical bypass, peripheral vascular disease, and unstable angina. Pravastatin and aspirin in a post-MI population was found to be a beneficial combination that seems to work through lipid and nonlipid, anti-inflammatory mechanisms. ^

William Osler: Original Papers 1898-1906

Part 1: 1898-1899 On Chronic Symmetrical Enlargement of the Salivary and Lachrymal Glands, 1898 Leprosy in the United States, with the Report of a Case, 1898 An Acute Myxaedematous Condition, with Tachycardia, Glycosuria, Melaena, Mania and Death, 1898 On some of the Intestinal Features of Typhoid Fever, 1898 Cerebro-Spinal Fever, 1898 The Arthritis of Cerebro-Spinal Fever, 1898 In Memoriam, William Pepper, 1899 After Twenty-Five Years, 1899 The Diagnosis of Typhoid Fever, 1899 Interstitial Processes in the Central Nervous System, 1899 Part 2: 1900 The Home Treatment of Consumption, 1900 On Splenic Anaemia, 1900 The Chronic Intermittent Fever of Endocarditis, 1900 A Case of Multiple Gangrene in Malarial Fever, 1900 Latent Cancer of the Stomach, 1900 On the Study of Tuberculosis, 1900 Fatal Angina Pectoris without Lesions of the Coronary Arteries of a Young Man, 1900 On the Advantages of a Trace of Albumin and a Few Casts in the Urine of Certain Men above Fifty Years of Age, 1900 Part 3: 1901-1902 Congenital Absence of the Abdominal Muscles with Distended Hypertrophied Urinary Bladder, 1901 Intermittent Claudication, 1902 On the Diagnosis of Bilateral Cystic Kidney, 1902 On Amebic Abscess of the Liver, 1902 Note on the Occurrence of Ascites in Solid Abdominal Tumors, 1902 Amebic Dysentery, 1902 Notes on Aneurism, 1902 William Beaumont; a Pioneer American Physiologist, 1902 Part 4: 1903 On the Educational Value of the Medical Society, 1903 On obliteration of the Superior Vena Cava,1903 Chronic Cyanosis, with Polycythemia and Enlarged Spleen: A New Clinical Entity, 1903 The Home and its Relation to the Tuberculosis Problem, 1903 Unity, Peace, and Concord, 1903 Typhoid Fever and Tuberculosis, 1903 Part 5: 1904-1906 Ochronosis, 1904 The “Phthisiologia” of Richard Morton, M.D., 1904 On the Surgical Importance of the Visceral Crises In the Erythema Group of Skin Diseases, 1904 Aneurysm of the Abdominal Aorta, 1905 Back Notes

William Osler: Original Papers 1907-1919

Part 1: 1907-1908 The Royal Medical Society of Edinburg, 1907 On the Library of a Medical School, 1907 On Telangiectasis Circumscripta Universalis, 1907 A Clinical Lecture on Abdominal Tumours Associated with Disease of the Testicle, 1907 A Clinical Lecture on Erythraemia, 1908 Vienna after Thirty-Four Years, 1908 Endocardites Infectieuses Chroniques, 1908 Part 2: 1909 Chronic Infectious Endocarditis, 1909 What the Public Can Do in the Fight Against Tuberculosis, 1909 Annual Oration on the Occasion of the Opening of the New Building of the Medical and Chirurgical Faculty of the State of Maryland, May 13, 1909 The Medical Library in Post-Graduate Work, 1909 The Treatment of Disease, 1909 Part 3: 1910-1911 The Pupil Symptoms in Thoracic Aneurysm, 1910 The Lumleian Lectures on Angina Pectoris, 1910 Certain Vasomotor, Sensory, and Muscular Phenomena Associated with Cervical Rib, 1910 An Address on the Hospital Unit in University Work, 1911 Sulle Telangiectasie Emorragiche Ereditarie, 1911 Transient Attacks of Aphasia and Paralyses in States of High Blood Pressure and Arterio-Sclerosis, 1911 The Pathological Institute of a General Hospital, 1911 Part 4: 1912-1914 An Address on High Blood Pressure: its Associations, Advantages, and Disadvantages, 1912 Specialism in the General Hospital, 1913 Syphilis of the Liver with the Picture of Banti’s Disease, 1913 An Introductory Address on Examinations, Examiners, and Examinees, 1913 The Medical Clinic: a retrospect and a Forecast, 1914 Part 5: 1915-1919 Remarks on the Diagnosis of Polycystic Kidney, 1915 The War and Typhoid Fever, 1914/15 The Cerebro-Spinal Fever in Camps and Barracks, 1915 Remarks on Arterio-Venous Aneurysm, 1915 Nerve & “Nerves”, 1915 Intensive Work in Science at the Public Schools in Relation to the Curriculum, 1916 Creators, Transmuters, and Transmitters, 1916 Annual Oration on the Campaign Against Syphilis, 1917 The First Printed Documents relating to Modern Surgical Anaesthesia, 1918 Observations on the Severe Anaemias of Pregnancy and the Post-Partum State, 1919 Typhoid Spine, 1919

Health outcomes, quality improvement and cost-effectiveness of the Community Diabetes Education (CoDE) program: Results of a randomized controlled trial

The objectives of this dissertation were to evaluate health outcomes, quality improvement measures, and the long-term cost-effectiveness and impact on diabetes-related microvascular and macrovascular complications of a community health worker-led culturally tailored diabetes education and management intervention provided to uninsured Mexican Americans in an urban faith-based clinic. A prospective, randomized controlled repeated measures design was employed to compare the intervention effects between: (1) an intervention group (n=90) that participated in the Community Diabetes Education (CoDE) program along with usual medical care; and (2) a wait-listed comparison group (n=90) that received only usual medical care. Changes in hemoglobin A1c (HbA1c) and secondary outcomes (lipid status, blood pressure and body mass index) were assessed using linear mixed-models and an intention-to-treat approach. The CoDE group experienced greater reduction in HbA1c (-1.6%, p<.001) than the control group (-.9%, p<.001) over the 12 month study period. After adjusting for group-by-time interaction, antidiabetic medication use at baseline, changes made to the antidiabetic regime over the study period, duration of diabetes and baseline HbA1c, a statistically significant intervention effect on HbA1c (-.7%, p=.02) was observed for CoDE participants. Process and outcome quality measures were evaluated using multiple mixed-effects logistic regression models. Assessment of quality indicators revealed that the CoDE intervention group was significantly more likely to have received a dilated retinal examination than the control group, and 53% achieved a HbA1c below 7% compared with 38% of control group subjects. Long-term cost-effectiveness and impact on diabetes-related health outcomes were estimated through simulation modeling using the rigorously validated Archimedes Model. Over a 20 year time horizon, CoDE participants were forecasted to have less proliferative diabetic retinopathy, fewer foot ulcers, and reduced numbers of foot amputations than control group subjects who received usual medical care. An incremental cost-effectiveness ratio of $355 per quality-adjusted life-year gained was estimated for CoDE intervention participants over the same time period. The results from the three areas of program evaluation: impact on short-term health outcomes, quantification of improvement in quality of diabetes care, and projection of long-term cost-effectiveness and impact on diabetes-related health outcomes provide evidence that a community health worker can be a valuable resource to reduce diabetes disparities for uninsured Mexican Americans. This evidence supports formal integration of community health workers as members of the diabetes care team.^

Dynamic contrast agent -enhanced magnetic resonance imaging assessment of tumor microvasculature

Dynamic contrast agent-enhanced magnetic resonance imaging (DCE MRI) data, when analyzed with the appropriate pharmacokinetic models, have been shown to provide quantitative estimates of microvascular parameters important in characterizing the angiogenic activity of malignant tissue. These parameters consist of the whole blood volume per unit volume of tissue, v b, transport constant from the plasma to the extravascular, extracellular space (EES), k1 and the transport constant from the EES to the plasma, k2. Parameters vb and k1 are expected to correlate with microvascular density (MVD) and vascular permeability, respectively, which have been suggested to serve as surrogate markers for angiogenesis. In addition to being a marker for angiogenesis, vascular permeability is also useful in estimating tumor penetration potential of chemotherapeutic agents. ^ Histological measurements of the intratumoral microvascular environment are limited by their invasiveness and susceptibility to sampling errors. Also, MVD and vascular permeability, while useful for characterizing tumors at a single time point, have shown less utility in longitudinal studies, particularly when used to monitor the efficacy of antiangiogenic and traditional chemotherapeutic agents. These limitations led to a search for a non-invasive means of characterizing the microvascular environment of an entire tumor. ^ The overall goal of this project was to determine the utility of DCE MRI for monitoring the effect of antiangiogenic agents. Further applications of a validated DCE MRI technique include in vivo measurements of tumor microvascular characteristics to aid in determining prognosis at presentation and in estimating drug penetration. DCE MRI data were generated using single- and dual-tracer pharmacokinetic models with different molecular-weight contrast agents. The resulting pharmacokinetic parameters were compared to immunohistochemical measurements. The model and contrast agent combination yielding the best correlation between the pharmacokinetic parameters and histological measures was further evaluated in a longitudinal study to evaluate the efficacy of DCE MRI in monitoring the intratumoral microvascular environment following antiangiogenic treatment. ^

Historias de vida y desencadenantes emocionales del síndrome coronario agudo : Inicio de un proyecto de investigación en el hospital El Cruce

Las enfermedades cardiovasculares son consideradas la principal causa de muerte de hombres y mujeres en todo el mundo, así como también en nuestro país. Existe en la bibliografía un número identificado de factores de riesgo que incluye el sedentarismo, el consumo de tabaco, una inadecuada dieta alimenticia, el distrés personal y laboral, la tendencia a la ira-hostilidad, la ansiedad y la depresión, los cambios sociales vinculados a los niveles socioeconómicos, y las catástrofes naturales y artificiales. Si bien estos factores de riesgo explican una condición favorable a su aparición, dando cuenta de una enfermedad arterial crónica, la presentación clínica del evento cardiovascular es habitualmente abrupta. Más de la mitad de los pacientes con infarto no son precedidos por patología previa o siquiera síntomas. La literatura científica ha estudiado la activación de los síndromes coronarios agudos, prestando un especial interés a la idea de que los comportamientos y las emociones pueden desencadenar ('gatillar") eventos cardíacos en personas susceptibles. La situación psicológica desencadenante puede ser un acontecimiento aparentemente banal si se la considera y valora de manera objetiva. Sin embargo, su importancia radica en su repercusión psicológica y en la significación que adquiere para el propio paciente en el contexto de sus vivencias actuales y pasadas. La investigación cualitativa aportada por los casos explorados a través de la corriente narrativa en medicina y psicología, y por los casos investigados mediante métodos psicoanalíticos constituyen las fuentes posibles de información acerca de los conflictos recientes vividos por los pacientes con síndrome coronario agudo. Objetivos y metodología: En el Servicio de Cardiología del Hospital de Alta Complejidad en Red 'El Cruce', de Florencio Varela, hemos comenzado a desarrollar un proyecto de investigación clínica con el propósito de evaluar en una serie de pacientes ingresados con diagnóstico confirmado de infarto agudo de miocardio o angina inestable, la prevalencia de situaciones emocionales negativas intensas estereotípicas en las horas o días previos al evento coronario. Para tal objetivo son realizadas entrevistas semidirigidas a cargo de un profesional psicólogo, a pacientes menores de 65 años de edad, en las cuales se toma en especial consideración el relato de la historia de vida reciente del paciente junto a posibles situaciones conflictivas previas que puedan acompañar el cuadro coronario agudo, la trama familiar y las personas significativas, el contexto e historia laboral, y la vida sexual del paciente. El estado de conciencia inadecuado para el interrogatorio y la existencia de eventos coronarios previos constituyen los criterios de exclusión para esta investigación. A su vez, se evaluará la correlación entre los hallazgos obtenidos en las entrevistas y una serie de test y cuestionarios reconocidos en la exploración de los factores emocionales en este tipo de pacientes: el Inventario de Hostilidad de Buss-Durkee y el score de situación de ira, la Escala de Depresión y Ansiedad Hospitalaria (HADS), el cuestionario Beck de depresión y el Cuestionario de Eventos Vitales. La reiteración en un porcentaje relevante de los casos de la identificación de la situación conflictiva estereotípica permitiría un avance significativo en esta línea de investigación fisiopatológica y psicológica, la ampliación del conocimiento relativo al síndrome coronario agudo, y facilitaría, a su vez, la elaboración de estrategias psicoterapéuticas orientadas al tratamiento y a la modificación pronóstica de la enfermedad

Historias de vida y desencadenantes emocionales del síndrome coronario agudo : Inicio de un proyecto de investigación en el hospital El Cruce

Las enfermedades cardiovasculares son consideradas la principal causa de muerte de hombres y mujeres en todo el mundo, así como también en nuestro país. Existe en la bibliografía un número identificado de factores de riesgo que incluye el sedentarismo, el consumo de tabaco, una inadecuada dieta alimenticia, el distrés personal y laboral, la tendencia a la ira-hostilidad, la ansiedad y la depresión, los cambios sociales vinculados a los niveles socioeconómicos, y las catástrofes naturales y artificiales. Si bien estos factores de riesgo explican una condición favorable a su aparición, dando cuenta de una enfermedad arterial crónica, la presentación clínica del evento cardiovascular es habitualmente abrupta. Más de la mitad de los pacientes con infarto no son precedidos por patología previa o siquiera síntomas. La literatura científica ha estudiado la activación de los síndromes coronarios agudos, prestando un especial interés a la idea de que los comportamientos y las emociones pueden desencadenar ('gatillar") eventos cardíacos en personas susceptibles. La situación psicológica desencadenante puede ser un acontecimiento aparentemente banal si se la considera y valora de manera objetiva. Sin embargo, su importancia radica en su repercusión psicológica y en la significación que adquiere para el propio paciente en el contexto de sus vivencias actuales y pasadas. La investigación cualitativa aportada por los casos explorados a través de la corriente narrativa en medicina y psicología, y por los casos investigados mediante métodos psicoanalíticos constituyen las fuentes posibles de información acerca de los conflictos recientes vividos por los pacientes con síndrome coronario agudo. Objetivos y metodología: En el Servicio de Cardiología del Hospital de Alta Complejidad en Red 'El Cruce', de Florencio Varela, hemos comenzado a desarrollar un proyecto de investigación clínica con el propósito de evaluar en una serie de pacientes ingresados con diagnóstico confirmado de infarto agudo de miocardio o angina inestable, la prevalencia de situaciones emocionales negativas intensas estereotípicas en las horas o días previos al evento coronario. Para tal objetivo son realizadas entrevistas semidirigidas a cargo de un profesional psicólogo, a pacientes menores de 65 años de edad, en las cuales se toma en especial consideración el relato de la historia de vida reciente del paciente junto a posibles situaciones conflictivas previas que puedan acompañar el cuadro coronario agudo, la trama familiar y las personas significativas, el contexto e historia laboral, y la vida sexual del paciente. El estado de conciencia inadecuado para el interrogatorio y la existencia de eventos coronarios previos constituyen los criterios de exclusión para esta investigación. A su vez, se evaluará la correlación entre los hallazgos obtenidos en las entrevistas y una serie de test y cuestionarios reconocidos en la exploración de los factores emocionales en este tipo de pacientes: el Inventario de Hostilidad de Buss-Durkee y el score de situación de ira, la Escala de Depresión y Ansiedad Hospitalaria (HADS), el cuestionario Beck de depresión y el Cuestionario de Eventos Vitales. La reiteración en un porcentaje relevante de los casos de la identificación de la situación conflictiva estereotípica permitiría un avance significativo en esta línea de investigación fisiopatológica y psicológica, la ampliación del conocimiento relativo al síndrome coronario agudo, y facilitaría, a su vez, la elaboración de estrategias psicoterapéuticas orientadas al tratamiento y a la modificación pronóstica de la enfermedad

Historias de vida y desencadenantes emocionales del síndrome coronario agudo : Inicio de un proyecto de investigación en el hospital El Cruce

Las enfermedades cardiovasculares son consideradas la principal causa de muerte de hombres y mujeres en todo el mundo, así como también en nuestro país. Existe en la bibliografía un número identificado de factores de riesgo que incluye el sedentarismo, el consumo de tabaco, una inadecuada dieta alimenticia, el distrés personal y laboral, la tendencia a la ira-hostilidad, la ansiedad y la depresión, los cambios sociales vinculados a los niveles socioeconómicos, y las catástrofes naturales y artificiales. Si bien estos factores de riesgo explican una condición favorable a su aparición, dando cuenta de una enfermedad arterial crónica, la presentación clínica del evento cardiovascular es habitualmente abrupta. Más de la mitad de los pacientes con infarto no son precedidos por patología previa o siquiera síntomas. La literatura científica ha estudiado la activación de los síndromes coronarios agudos, prestando un especial interés a la idea de que los comportamientos y las emociones pueden desencadenar ('gatillar") eventos cardíacos en personas susceptibles. La situación psicológica desencadenante puede ser un acontecimiento aparentemente banal si se la considera y valora de manera objetiva. Sin embargo, su importancia radica en su repercusión psicológica y en la significación que adquiere para el propio paciente en el contexto de sus vivencias actuales y pasadas. La investigación cualitativa aportada por los casos explorados a través de la corriente narrativa en medicina y psicología, y por los casos investigados mediante métodos psicoanalíticos constituyen las fuentes posibles de información acerca de los conflictos recientes vividos por los pacientes con síndrome coronario agudo. Objetivos y metodología: En el Servicio de Cardiología del Hospital de Alta Complejidad en Red 'El Cruce', de Florencio Varela, hemos comenzado a desarrollar un proyecto de investigación clínica con el propósito de evaluar en una serie de pacientes ingresados con diagnóstico confirmado de infarto agudo de miocardio o angina inestable, la prevalencia de situaciones emocionales negativas intensas estereotípicas en las horas o días previos al evento coronario. Para tal objetivo son realizadas entrevistas semidirigidas a cargo de un profesional psicólogo, a pacientes menores de 65 años de edad, en las cuales se toma en especial consideración el relato de la historia de vida reciente del paciente junto a posibles situaciones conflictivas previas que puedan acompañar el cuadro coronario agudo, la trama familiar y las personas significativas, el contexto e historia laboral, y la vida sexual del paciente. El estado de conciencia inadecuado para el interrogatorio y la existencia de eventos coronarios previos constituyen los criterios de exclusión para esta investigación. A su vez, se evaluará la correlación entre los hallazgos obtenidos en las entrevistas y una serie de test y cuestionarios reconocidos en la exploración de los factores emocionales en este tipo de pacientes: el Inventario de Hostilidad de Buss-Durkee y el score de situación de ira, la Escala de Depresión y Ansiedad Hospitalaria (HADS), el cuestionario Beck de depresión y el Cuestionario de Eventos Vitales. La reiteración en un porcentaje relevante de los casos de la identificación de la situación conflictiva estereotípica permitiría un avance significativo en esta línea de investigación fisiopatológica y psicológica, la ampliación del conocimiento relativo al síndrome coronario agudo, y facilitaría, a su vez, la elaboración de estrategias psicoterapéuticas orientadas al tratamiento y a la modificación pronóstica de la enfermedad

Fulminant hepatic and multiple organ failure following acute viral tonsillitis: a case report.

BACKGROUND Pyogenic tonsillitis may often be observed in the general Western population. In severe cases, it may require antibiotic treatment or even hospitalization and often a prompt clinical response will be noted. Here we present an unusual case of progressive multiple organ failure including fulminant liver failure following acute tonsillitis initially mistaken for "classic" pyogenic (that is bacterial) tonsillitis. CASE PRESENTATION A 68-year-old previously healthy white man was referred with suspicion of pyogenic angina. After tonsillectomy, he developed acute liver failure and consecutive multiple organ failure including acute hemodynamic, pulmonary and dialysis-dependent renal failure. Immunohistopathological analysis of his tonsils and liver as well as serum polymerase chain reaction analyses revealed herpes simplex virus-2 to be the causative pathogen. Treatment included high-dose acyclovir and multiorgan supportive intensive care therapy. His final outcome was favorable. CONCLUSIONS Fulminant herpes simplex virus-2-induced multiple organ failure is rarely observed in the Western hemisphere and should be considered a potential diagnosis in patients with tonsillitis and multiple organ failure including acute liver failure. From a clinical perspective, it seems important to note that fulminant herpes simplex virus-2 infection may masquerade as "routine" bacterial severe sepsis/septic shock. This persevering condition should be diagnosed early and treated goal-oriented in order to gain control of this life-threatening condition.

Xanthine oxidase activity associated with arterial blood pressure in spontaneously hypertensive rats

Recent evidence in vivo indicates that spontaneously hypertensive rats (SHR) exhibit an increase in oxyradical production in and around microvascular endothelium. This study is aimed to examine whether xanthine oxidase plays a role in overproduction of oxidants and thereby may contribute to hypertensive states as a consequence of the increasing microvascular tone. The xanthine oxidase activity in SHR was inhibited by dietary supplement of tungsten (0.7 g/kg) that depletes molybdenum as a cofactor for the enzyme activity as well as by administration of (−)BOF4272 [(−)-8-(3-methoxy-4-phenylsulfinylphenyl)pyrazolo(1,5-α)-1,3,5-triazine-4-monohydrate], a synthetic inhibitor of the enzyme. The characteristic elevation of mean arterial pressure in SHR was normalized by the tungsten diet, whereas Wistar Koto (WKY) rats displayed no significant alteration in the pressure. Multifunctional intravital videomicroscopy in mesentery microvessels with hydroethidine, an oxidant-sensitive fluoroprobe, showed that SHR endothelium exhibited overproduction of oxyradicals that coincided with the elevated arteriolar tone as compared with WKY rats. The tungsten diet significantly repressed these changes toward the levels observed in WKY rats. The activity of oxyradical-producing form of xanthine oxidase in the mesenteric tissue of SHR was ≈3-fold greater than that of WKY rats, and pretreatment with the tungsten diet eliminated detectable levels of the enzyme activity. The inhibitory effects of the tungsten diet on the increasing blood pressure and arteriolar tone in SHR were also reproducible by administration of (−)BOF4272. These results suggest that xanthine oxidase accounts for a putative source of oxyradical generation that is associated with an increasing arteriolar tone in this form of hypertension.

Role of substance P and the neurokinin 1 receptor in acute pancreatitis and pancreatitis-associated lung injury

Substance P, acting via the neurokinin 1 receptor (NK1R), plays an important role in mediating a variety of inflammatory processes. However, its role in acute pancreatitis has not been previously described. We have found that, in normal mice, substance P levels in the pancreas and pancreatic acinar cell expression of NK1R are both increased during secretagogue-induced experimental pancreatitis. To evaluate the role of substance P, pancreatitis was induced in mice that genetically lack NK1R by administration of 12 hourly injections of a supramaximally stimulating dose of the secretagogue caerulein. During pancreatitis, the magnitude of hyperamylasemia, hyperlipasemia, neutrophil sequestration in the pancreas, and pancreatic acinar cell necrosis were significantly reduced in NK1R−/− mice when compared with wild-type NK1R+/+ animals. Similarly, pancreatitis-associated lung injury, as characterized by intrapulmonary sequestration of neutrophils and increased pulmonary microvascular permeability, was reduced in NK1R−/− animals. These effects of NK1R deletion indicate that substance P, acting via NK1R, plays an important proinflammatory role in regulating the severity of acute pancreatitis and pancreatitis-associated lung injury.

Severe reduction in leukocyte adhesion and monocyte extravasation in mice deficient in CC chemokine receptor 2

CC chemokine receptor 2 (CCR2) is a prominent receptor for the monocyte chemoattractant protein (MCP) group of CC chemokines. Mice generated by gene targeting to lack CCR2 exhibit normal leukocyte rolling but have a pronounced defect in MCP-1-induced leukocyte firm adhesion to microvascular endothelium and reduced leukocyte extravasation. Constitutive macrophage trafficking into the peritoneal cavity was not significantly different between CCR2-deficient and wild-type mice. However, after intraperitoneal thioglycollate injection, the number of peritoneal macrophages in CCR2-deficient mice did not rise above basal levels, whereas in wild-type mice the number of macrophages at 36 h was ≈3.5 times the basal level. The CCR2-deficient mice showed enhanced early accumulation and delayed clearance of neutrophils and eosinophils. However, by 5 days neutrophils and eosinophils in both CCR2-deficient and wild-type mice had returned to near basal levels, indicating that resolution of this inflammatory response can occur in the absence of macrophage influx and CCR2-mediated activation of the resident peritoneal macrophages. After intravenous injection with yeast β-glucan, wild-type mice formed numerous large, well-defined granulomas throughout the liver parenchyma, whereas CCR2-deficient mice had much fewer and smaller granulomas. These results demonstrate that CCR2 is a major regulator of induced macrophage trafficking in vivo.

Synthesis, Storage, and Release of Vascular Endothelial Growth Factor/Vascular Permeability Factor (VEGF/VPF) by Human Mast Cells: Implications for the Biological Significance of VEGF206

Mast cells have been implicated in various diseases that are accompanied by neovascularization. The exact mechanisms by which mast cells might mediate an angiogenic response, however, are unclear and therefore, we have investigated the possible expression of vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) in the human mast cell line HMC-1 and in human skin mast cells. Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that mast cells constitutively express VEGF121, VEGF165, and VEGF189. After a prolonged stimulation of cells for 24 h with phorbol 12-myristate 13-acetate (PMA) and the ionophore A23187, an additional transcript representing VEGF206 was detectable, as could be verified by sequence analysis. These results were confirmed at the protein level by Western blot analysis. When the amounts of VEGF released under unstimulated and stimulated conditions were compared, a significant increase was detectable after stimulation of cells. Human microvascular endothelial cells (HMVEC) responded to the supernatant of unstimulated HMC-1 cells with a dose-dependent mitogenic effect, neutralizable up to 90% in the presence of a VEGF-specific monoclonal antibody. Flow cytometry and postembedding immunoelectron microscopy were used to detect VEGF in its cell-associated form. VEGF was exclusively detectable in the secretory granules of isolated human skin mast cells. These results show that both normal and leukemic human mast cells constitutively express bioactive VEGF. Furthermore, this study contributes to the understanding of the physiological role of the strongly heparin-binding VEGF isoforms, since these were found for the first time to be expressed in an activation-dependent manner in HMC-1 cells.

Agrin in Alzheimer’s disease: Altered solubility and abnormal distribution within microvasculature and brain parenchyma

Agrin is a heparan sulfate proteoglycan that is widely expressed in neurons and microvascular basal lamina in the rodent and avian central nervous system. Agrin induces the differentiation of nerve-muscle synapses, but its function in either normal or diseased brains is not known. Alzheimer’s disease (AD) is characterized by loss of synapses, changes in microvascular architecture, and formation of neurofibrillary tangles and senile plaques. Here we have asked whether AD causes changes in the distribution and biochemical properties of agrin. Immunostaining of normal, aged human central nervous system revealed that agrin is expressed in neurons in multiple brain areas. Robust agrin immunoreactivity was observed uniformly in the microvascular basal lamina. In AD brains, agrin is highly concentrated in both diffuse and neuritic plaques as well as neurofibrillary tangles; neuronal expression of agrin also was observed. Furthermore, patients with AD had microvascular alterations characterized by thinning and fragmentation of the basal lamina. Detergent extraction and Western blotting showed that virtually all the agrin in normal brain is soluble in 1% SDS. In contrast, a large fraction of the agrin in AD brains is insoluble under these conditions, suggesting that it is tightly associated with β-amyloid. Together, these data indicate that the agrin abnormalities observed in AD are closely linked to β-amyloid deposition. These observations suggest that altered agrin expression in the microvasculature and the brain parenchyma contribute to the pathogenesis of AD.