Flow cytometry with a monoclonal antibody to the low density lipoprotein receptor compared with gene mutation detection in diagnosis of heterozygous familial hypercholesterolemia

We used a fluorescence flow cytometry assay with a monoclonal low density lipoprotein (LDL) receptor-specific antibody to detect LDL receptor expression on blood T lymphocytes and monocytes. We prepared peripheral blood mononuclear cells from patients with genetically verified LDL receptor-defective (Trp66-Gly mutation, n = 17) or receptor-negative (Trp23-stop mutation, n = 17) heterozygous familial hypercholesterolemia (FH) and from healthy individuals (n = 24). The cells were stimulated to express the maximum amount of LDL receptor by preincubation in lipoprotein-deficient medium. A dual-labeling technique allowed flow cytometric analysis of LDL receptor expression on cells identified by fluorescently conjugated surface marker antibodies. Knowing the LDL receptor gene mutation of the FH patients allowed us to compare the diagnostic capability of this functional assay with the DNA diagnosis and to validate the assay with molecular genetics instead of clinical indices of heterozygous FH. T lymphocytes expressed more LDL receptors and gave better diagnostic results than monocytes, and cells from patients with either the Trp66-Gly or the Trp23-stop mutation had variable but significantly reduced LDL receptor expression. The data indicate that this fluorescence flow cytometry assay is unsuitable for diagnosis of individual cases of heterozygous FH but that it may be useful for functionally characterizing mutations in the LDL receptor gene.     

Radiation exposure in abdominal radiography with digital luminescence radiography and conventional screen-film system: an experimental animal study]

Progress in the care of the critically ill patient with life-threatening infection has been hampered by inconsistent, often confusing terminology. The clinical syndrome of sepsis-familiar to all yet definable by none-describes a highly heterogeneous group of disorders with different causes and differing prognoses. The imminent availability of mediator-directed therapy has created a sense of urgency to develop better methods for delineating discrete clinical syndromes and to modulate the host response, which may bring both benefit and harm, depending on the clinical circumstances. The term systemic inflammatory response syndrome (SIRS) was introduced several years ago to describe the familiar clinical syndrome of sepsis, independent of its cause. SIRS can result from trauma, pancreatitis, drug reactions, autoimmune disease, and a host of other disorders; when it arises in response to infection, sepsis is said to be present. SIRS describes a dynamic process that has adaptive survival value for the host. The maladaptive consequence of this process in the critically ill patient is the development of progressive but potentially reversible remote organ dysfunction-the multiple organ dysfunction syndrome. The development of cogent conceptual frameworks for classification of the septic response in critically ill patients is more than a question of linguistic pedantry. Optimal therapy presupposes identification of an homogeneous patient population with a characteristic disease process and a predictable response to an intervention. Although progress has been made in identifying such groups of critically ill patients, the disappointing results of clinical trials of agents that so clearly demonstrate efficacy in animal models indicates that considerable work remains.     

Laminin-like proteins are differentially regulated during cerebellar development and stimulate granule cell neurite outgrowth in vitro

OBJECTIVE: To review data supporting the hypothesis that syndrome X plays a major role in the pathogenesis of coronary artery disease (CAD), and the effects of lifestyle factors and pharmacologic interventions on insulin, other metabolic parameters, and outcomes. DATA SOURCES: MEDLINE (January 1966-August 1997) and Current Contents database searches identified applicable English-language experimental trials, epidemiologic studies, reviews, and editorials. STUDY SELECTION AND DATA EXTRACTION: Studies that were included addressed the role of insulin resistance and hyperinsulinemia in the pathogenesis of CAD or the effects of lifestyle factors and pharmacologic interventions on metabolic parameters and outcomes. DATA SYNTHESIS: The main characteristics of syndrome X are hyperinsulinemia and insulin resistance. These result in secondary syndrome X features, including hyperglycemia, increased very-low-density lipoprotein concentrations, decreased high-density lipoprotein cholesterol, and hypertension. Insulin resistance is worsened by obesity, and insulin has been shown to contribute to the development of hypertension. Other studies demonstrate that smoking adversely affects glucose and insulin concentrations. Animal studies have linked hyperinsulinemia and atherogenesis. These animal data have been confirmed by several large prospective and population studies that have identified associations between hyperinsulinemia and CAD. CONCLUSIONS: Strong evidence links insulin resistance and hyperinsulinemia to CAD. Lifestyle modifications play an important role in decreasing cardiovascular risk, and clinicians should strongly encourage such changes. Clinicians must also carefully consider the effects of antihypertensive, antihyperglycemic, and antidyslipidemic agents on patients' metabolic profiles when choosing appropriate therapeutic regimens. However, outcome data on many potentially beneficial agents, including calcium antagonists, alpha 1-adrenergic antagonists, angiotensin-converting enzyme inhibitors, metformin, acarbose, and troglitazone, are not yet available.     

Lidocaine-induced CNS toxicity--a case report

One hundred and twenty-six cases of cerebellopontine angle tumors with various histologies are presented. Results of 75 operated vestibular neurinomas, 22 meningiomas, and 16 tumors with other histologies are discussed. The method of irradiation and non-radical surgery may be an alternative for treatment.     

Efficient expression of the gene for spinach phosphoribulokinase in Pichia pastoris and utilization of the recombinant enzyme to explore the role of regulatory cysteinyl residues by site-directed mutagenesis

Phosphoribulokinase (PRK), unique to photosynthetic organisms, is regulated in higher plants by thioredoxin-mediated thiol-disulfide exchange in a light-dependent manner. Prior attempts to overexpress the higher plant PRK gene in Escherichia coli for structure-function studies have been hampered by sensitivity of the recombinant protein to proteolysis as well as toxic effects of the protein on the host. To overcome these impediments, we have spliced the spinach PRK coding sequence immediately downstream from the AOX1 (alcohol oxidase) promoter of Pichia pastoris, displacing the chromosomal AOX1 gene. The PRK gene is now expressed, in response to methanol, at 4-6% of total soluble protein, without significant in vivo degradation of the recombinant enzyme. This recombinant spinach PRK is purified to homogeneity by successive anion-exchange and dye-affinity chromatography and is shown to be electrophoretically and kinetically indistinguishable from the authentic spinach counterpart. Site-specific replacement of all of PRK's cysteinyl residues (both individually and in combination) demonstrates a modest catalytically facilitative role for Cys-55 (one of the regulatory residues) and the lack of any catalytic role for Cys-16 (the other regulatory residue), Cys-244, or Cys-250. Mutants with seryl substitutions at position 55 display non-hyperbolic kinetics relative to the concentration of ribulose 5-phosphate. Sulfate restores hyperbolic kinetics and enhances kinase activity, presumably reflecting conformational differences between the position 55 mutants and wild-type enzyme. Catalytic competence of the C16S-C55S double mutant proves that mere loss of free sulfhydryl groups by oxidative regulation cannot account entirely for the accompanying total inactivation.     

Delivery of salbutamol to nonventilated preterm infants by metered-dose inhaler, jet nebulizer, and ultrasonic nebulizer

An isocratic reversed-phase high-performance liquid chromatographic method with ultraviolet detection at 230 nm has been developed for the determination of paclitaxel in human plasma. Plasma samples were prepared by a selective one-step liquid-liquid extraction involving a mixture of acetonitrile-n-butyl chloride (1:4, v/v). Paclitaxel and the internal standard docetaxel were separated using a column packed with ODS-80A material, and a mobile phase consisting of water-methanol-tetrahydrofuran-ammonium hydroxide (37.5:60:2.5:0.1, v/v). The calibration graph for paclitaxel was linear in the range 10-500 ng/ml, with a lower limit of quantitation of 10 ng/ml, using 1 ml plasma samples. The extraction recoveries of spiked paclitaxel and docetaxel to drug-free human plasma were 89.6+/-8.52 and 93.7+/-5.0%, respectively. Validation data showed that the assay for paclitaxel is sensitive, selective, accurate and reproducible. The assay has been used in a single pharmacokinetic experiment in a patient to investigate the applicability of the method in vivo.     

Factors influencing survival after gamma knife radiosurgery for patients with single and multiple brain metastases

BACKGROUND: Significant changes are restructurng the U.S. health care delivery system. National health reform is now extending itself into the public sector. Increased health and medical costs by federal and state governments are forcing a reevaluation of major entitlement programs, especially Medicaid. METHODS/RESULTS: Because Medicaid is the single largest item in many state budgets, states are now enrolling Medicaid patients into managed and coordinated care arrangements as a means to control costs and increase access to care. HMOs are not only competing for private patients but also actively seeking the Medicaid population. Nationally, almost one-fourth of all Medicaid patients are now enrolled in managed care plans. Various models and approaches have been developed by individual states. CONCLUSIONS: Because managed care enrollment in the Medicaid program has increased substantially in recent years, selected services including vision care are no longer rendered by any practitioner willing to accept Medicaid fees. Freedom of choice is now restricted to pre-selected and panel practitioners participating with the managed care program. The rules, regulations, billing procedures, fees, and program requisites will differ under managed care programs. Private optometric practitioners must consider entering economic and organizational relationships and linkages that make them attractive to managed care organizations.     

Thymic overexpression of CD40 ligand disrupts normal thymic epithelial organization

We characterized the distribution of CD40 and CD40 ligand (CD40-L) in the adult and developing murine thymus. Before birth, CD40 was almost exclusively localized to scattered foci of medullary cells. By birth there was a dramatic upregulation of CD40 expression by cortical epithelial cells, which was accompanied by a consolidation of medullary epithelial foci. CD40-L+ thymocytes displayed a medullary location. Analysis of mice deficient in CD40-L expression indicated that CD40-L/CD40 interactions were not required for development of the medullary compartment. Overexpression of CD40-L targeted to thymocytes altered thymic architecture, as reflected by a dramatic loss of cortical epithelial cells, expansion of the medullary compartment, and extensive infiltration of the capsule with a mixture of CD3+ cells, B-cells, and macrophages/dendritic cells. Reconstitution of lethally irradiated normal mice with lck CD40-L bone marrow cells also resulted in loss of cortical epithelium and expansion of the medullary compartment. Disruption of the normal pattern of thymic architecture and epithelial differentiation as a consequence of increased intrathymic levels of CD40-L expression points to a role for CD40-L/CD40 interactions in the normal pattern of epithelial compartmentalization/differentiation within the thymic environment.