Serum- free culture of dendritic cells from patients with chronic myeloid leukemia in vitro and estimation of their cytotoxicity

Objective To establish a serum- free culture system of dendritic cells (DCs) from chronic myeloid leukemia (CML) cells so that DCs vaccine may be applied to the adoptive immunotherapy of CML in the near future.Methods Fetal calf serum, serum- free medium and autologous serum were used for culture of DCs. The usage of cytokines was classified into two groups: group A (stem cell factor, granulocyte/macrophage colony- stimulating- factor, tumor necrosis factor- α and interleukin- 4) and group B (granulocyte/macrophage colony- stimulating- factor, tumor necrosis factor- α and interleukin- 4). The phenotypes of DCs were analyzed by using indirect immunofluorescence and flow cytometry. Mixed leukocyte responses were performed by methyl thiazolyl tetrazolium (MTT) assay. Chromosome analysis of DCs can be achieved by displaying G banding. T cells from CML patients were stimulated with autologous DCs and T- cell cytotoxicity was measured by (MTT) assay. Results CD34(+) cells or mononuclear cells were obtained from peripheral blood or bone marrow samples of eight patients of chronic- phase CML. Group A of serum- free medium was better than group B in expansion of total cell numbers and the rate of DCs. These results of serum- free medium were not significantly different from those of fetal calf serum medium, but the results of autologous serum medium were inferior to two groups above. The expression of major histocompatibility complex class Ⅱ antigen on the surface of DCs was notable (＞50%), but the expression of CD83 and the costimulatory molecules CD86 was not noticeable (10%-50%). Although CD1a+/CD14- DCs were potent stimulators of allogeneic lymphocytes, expansion of T cells from normal volunteers were not significant (average 27. 2 fold at DCs∶T cells ratio of 1∶10). At day 12, CD1a+ cells from three patients were studied by displaying G banding and Ph+ cells in these populations were 100%, 98% and 60%, respectively. At an effector∶target ratio of 40∶1, 32% to 45% cytotoxicity was noted with DC- stimulated T cells against autologous leukemia cells. Conclusions A stable serum- free culture system of CML- DCs was established. The expression of CD83 and CD86 on the surface of CML- DCs and DCs’ potent stimulation of allogeneic lymphocytes were not notable. DCs in CML patients can be derived from the malignant clone and these malignant DCs could induce anti- leukemic reactivity in autologous T lymphocytes without the necessity for additional exogenous antigens.     

Quantitative evaluation of opsonin-independent phagocytosis by alveolar macrophages in monolayer using polystyrene microspheres

Macrophages can bind and engulf a variety of particles in the absence of specific opsonins. Polystyrene-type microspheres are often employed to quantitate opsonin-independent phagocytic activities of macrophages in vitro. Reliable measurement of this cell function, however, requires the ability of the investigator to distinguish between particles that are merely attached to the cell surface and those that are actually internalized. We have developed a simple, rapid, and reproducible method for quantitating phagocytosis using polystyrene microspheres and adherent alveolar macrophages. Basically, particles associated with macrophages after a given incubation time are microscopically quantitated on a cell-by-cell basis before and after toluene dissolution of external particles. Particle/macrophage values obtained after toluene treatment exclusively index phagocytosis.     

Phänomenologie zykloider Achsensyndrome und ihre Abgrenzung gegen eine schizophrene Kerngruppe

Zusammenfassung Der Begriff „zykloide Psychosen” bezeichnet eine zun?chst nach dem Erscheinungsbild abzugrenzende Art innerhalb der Gattung „schizophrener Formenkreis”. Ausgehend von der klinischen Beobachtung wird in der vorliegenden ph?nomenologisch-konzeptuellen Arbeit die Auffassung vertreten, da? die Leonhardschen Subtypen bzw. deren jeweilige Pole als Achsensyndrome zu begreifen sind, die w?hrend ein und derselben Episode gemeinsam auftreten und ineinander übergehen k?nnen. Die „inneren” Zusammenh?nge dieser Achsensyndrome werden herausgearbeitet. Neben dem kongruenten Verweisungscharakter ihrer Aufbauelemente erweist sich dabei das Fehlen struktureller Verformungen 1. von Affektausdruck und Affekt, 2. des Denkens und 3. der Bewegungsimpulse und -abl?ufe als entscheidende Differentia specifica gegenüber einer schizophrenen Kerngruppe. Unseren pr?zisierten Ph?nomenbegriff machen wir zur Grundlage einer Kritik bisheriger Operationalisierungsvorschl?ge, für die wir eine Alternative anbieten.      

Ethologic implications of the skin and its disturbances

From the beginning of its simplest forms, animal life appears to be organized according to specialization by inner functions and surface functions. The passage from prokaryotic to eukaryotic cells, from bacteria to unicellular organisms, consists essentially of a functional specialization of surfaces, accompanied by an extraordinary development of plasma membrane from which originate membranous surfaces that have become specialized into endoplasmic reticulum and nuclear envelope, transferred inside the cell.Whereas the inner functions of cells are prevalently synthetic or transformational, the surface functions are concerned with chemical, physical, or informational exchanges with the environment. With the progressive increase in complexity that accompanies the passage from unicellular to multicellular conditions and the formation of systems of organs, the functions of chemical exchange are performed by surface elements which progressively move inside the body and become specialized (lungs, intestines, kidneys), whereas the outer surface conserves the function of relations with both the physical and the living environment. The function of the outer surface is mainly a protective one. Among the functional protection from the physical environment are the control of osmotic equilibrium, protection from attrition, and the maintenance of body temperature. To perform these many functions in many different environments, the integumentary system presents a wide range of variations both in the skin and in its appendages (cuticles, scales, hair, feathers, beaks, nails and claws, and many others).Equally important functions of the integuments are those connected with relations between organisms and the living environment. The skin and its appendages are the principal signal transmitters for olfactory, visual, and tactile communications. Regarding tactile communication, moreover, the skin also functions as a receptor organ.     

Effect of selenium on glutathione metabolism. Induction of gamma-glutamylcysteine synthetase and glutathione reductase in the rat liver

The effect of sodium selenite (Na₂SeO₃, Se) on cellular glutathione metabolism was examined, particularly with respect to its ability to alter the activities of γ-glutamylcysteine synthetase and glutathione disulfide (GSSG) reductase. The treatment of rats with Se (5, 10 and 20 μmoles/kg) caused time- and dose-dependent increases in the activities of the synthetase and the reductase in the liver. The activity of γ-glutamylcysteine synthetase, the rate-limiting enzyme of the glutathione (GSH) biosynthesis, was particularly susceptible to Se treatment. The Se-mediated increases in the activities of the above enzymes were inhibited by puromycin and the increases could not be elicited in vitro. Selenium treatment caused time-dependent perturbations in the levels and ratio of GSSG and GSH in the liver. When compared to the control animals, rats treated for 3 hr with 10 and 20 μmoles Se/kg showed increased cellular levels of GSSG; in contrast, 24 hr after Se treatment the concentration of GSH was increased significantly. The activity of γ-glutamyl transpeptidase, which catalyzes the initial reaction in GSH breakdown, was unaltered by Se treatment. Repeated administration of low doses of Se (7.0 μmoles/kg, three times) also increased the activities of the reductase and the synthetase as well as the cellular levels of hepatic GSH and GSSG. It is suggested that the Se-mediated increases in the activities of γ-glutamylcysteine synthetase and GSSG-reductase represent cellular responses to Se-mediated perturbations in the levels and ratio of GSH and GSSG.     

Digitalis arrhythmias: Role of oscillatory afterpotentials

Digitalis-induced OAP provide a mechanism of automaticity that may be responsible for many arrhythmias induced by cardiac glycosides. In response to digitalis, OAP occur in tissues of the specialized conducting systems of both ventricles and atria and, under the influence of tension, occasionally in ventricular myocardium. Digitalis, in toxic doses, suppresses “normal” pacemaker activity possibly in part by enhancing overdrive suppression. In contrast to “normal” pacemaker activity, OAP exhibit, both in magnitude and rate of depolarization, postpacing acceleration. This plus the coupled nature of OAP are important characteristics in the generation of complex arrhythmias by OAP. Conduction disturbances may also be related to OAP. At early stages of intoxication OAP may speed conduction of superimposed beats relative to earlier or later beats. More advanced stages of intoxication are associated with conduction block.

The occurrence of digitalis-induced OAP is promoted by high concentrations of calcium, low concentrations of potassium, and moderate stretch. OAP can be suppressed by high concentrations of potassium, reduction of extracellular calcium, and exposure to antiarrhythmic agents including diphenylhydantoin, verapamil, and aprindine. The effectiveness of the latter two agents may be related to ability to block transmembrane calcium currents. Digitalis-induced OAP in atrial tissue can be abolished by acetylcholine.

A transmembrane current possibly but not necessarily carried by calcium appears to underly the occurrence of OAP. This current demonstrates kinetic properties different from those of the slow inward current associated with the plateau of the cardiac action potential. Calcium is intimately involved in the mechanism causing OAP and may be responsible for aftercontractions observed in conjunction with OAP. Aftercontractions greatly affect contractility and may be responsible at least in part for some of the inotropic actions of digitalis. Thus the occurrence of OAP may be linked to the inotropic actions of digitalis.

Digitalis-induced OAP provide a mechanism of automaticity with characteristics paralleling automatic behavior observed in intact animals intoxicated with digitalis. The relative importance of OAP in the genesis of clinically important arrhythmias awaits further investigation.     

Cellular and molecular mechanisms in wound healing: selected concepts

The interactions among cells, structural and other proteins, and large and small molecules in the dermis is an extremely complex, and largely unknown subject in normal skin, so that these interactions in wounded skin are incapable of complete comprehension at present. The problem is compounded by the further reaction and interaction of formed structures within the dermis, such as blood and lymphatic vessels and nerves of all types, and adnexal structures, as well as interactions with the epidermis, which are separately reviewed in the following chapter. In spite of all of this, there are several areas in which research has provided rather extensive insight into the wound healing process. These areas will now be reviewed. A separate review has been provided in this volume of the results obtained using in vitro systems for studies of the wound healing process (Chapter 6).Wound healing has been traditionally divided into three phases: (1) the inflammatory phase, also known as the exudative, lag, or substrate phase; (2) the fibroblastic phase, also known as the connective tissue or proliferative phase; and (3) the remodeling phase, also known as the resorptive or differentiating phase. Although these phases are somewhat arbitrary, we will use them to organize our approach to the problem, dividing our discussion into the inflammatory phase—to which we shall give emphasis—and later developments. Various aspects of the phases of wound healing overlap extensively.