Immunopathology of the lens. I. Humoral and cellular immune responses to heterologous lens antigens and their roles in ocular inflammation. In experimental rabbits heterologous soluble lens proteins consisting of alpha-, beta-, and gamma-crystallins were found to be antigenic; they stimulated a marked antibody response compared to a rather weak T-cell response. The serum antibodies to alpha-crystallins appeared first, to be followed by antibodies to beta- and gamma-crystallins in that order. The rabbits did not respond to heterologous gamma-crystallins unless these were injected with Freund's adjuvant containing mycobacteria. Incomplete Freund's adjuvant (i.e., without mycobacteria) was found to be an inferior immunoaccelerator so far as lens antigens are concerned. The response to lenticular antigens in both magnitude and duration varied in different rabbits, which suggested to us the important role played by a central control mechanism involving the immune response (Ir) genes. Some of the antibodies in potent lens antisera cross-reacted with mitochondria, endoplasmic reticulum (i.e., microsomes), contractile organelles, and cell nuclei. This explains for the first time at least in part the reasons for the widely observed phenomenon of the reactivity of lens antisera with ocular and extraocular structures. Antibodies to soluble lens proteins as detected by immunofluorescence and immunoperoxidase techniques were shown to be of the IgG class. Systemic heterologous immunisation followed by discission of the lens does not lead to the typical changes of phakoallergic endophthalmitis in the rabbit.


The role of axoplasmic transport in the pathogenesis of retinal cotton-wool spots. Small retinal arterioles in the pig retina were occluded by argon laser photocoagulation and the morphology and topography of the resulting lesions studied by ophthalmoscopy, histology, and electron microscopy. Two days after laser coagulation ischaemic necrosis of the inner retina was observed in the territory of occluded arterioles, and swollen axon terminals packed with cytoplasmic organelles were found in the retinal nerve-fibre layer on the peripheral border of the infarcts and their border nearest to the disc. These swellings gave rise to localised zones of opacification (cotton-wool spots) at the borders of the retinal infarcts, and similar zones developed in relation to the laser burns. Amino-acid uptake and transport in retinal ganglion cells was studied in these animals by autoradiography following an intravitreal injection of tritiated leucine. When 3H-leucine was injected at the same time as laser coagulation and arteriolar occlusion, label became concentrated in the swollen axon-terminals on the peripheral aspect of 2-day-old infarcts but was absent from the terminals on the disc side of infarcts. The accumulation of 3H-leucine and organelles was attributed to interruption of orthograde axoplasmic transport. When 3H-leucine was injected into the vitreous 2 days prior to laser occlusion, label became distributed throughout the axoplasm and then accumulated in swollen axon terminals on both sides of infarcts after a further 2 days. An obstruction to both orthograde and retrograde axoplasmic flow was thus demonstrated. We concluded that cotton-wood spots should be redefined as accumulations of cytoplasmic debris in the retinal nerve-fibre layer caused by obstruction of orthograde or retrograde axoplasmic transport in ganglion cell axons.


Activities of lectins and their immobilized derivatives in detergent solutions. Implications on the use of lectin affinity chromatography for the purification of membrane glycoproteins. The effects of several commonly used detergents on the saccharide-binding activities of lectins were investigated using lectin-mediated agglutination of formalin-fixed erythrocytes and affinity chromatography of glycoproteins on columns of lectins immobilized on polyacrylic hydrazide-Sepharose. In the hemagglutination assays, Ricinus communis I (RCA1) and II (RCAII), concanavalin A (Con A), and the agglutinins from peanut (PNA), soybean (SBA), wheat germ (WGA), and Limulus polyphemus (LPA) were tested with several concentrations of switterionic, cationic, anionic, and nonionic detergents. It was found that increasing detergent concentrations eventually affected hemagglutination titers in both test and control samples, and the highest detergent concentrations not affecting lectin hemagglutinating activities were determined. The effects of detergents on specific binding of [3H]fetuin and asialo[3H]fetuin to and elution from columns of immobilized lectins were less severe when compared with lectins in solution, suggesting that the lectins are stabilized by covalent attachment to agarose beads. Nonionic detergents did not affect the binding efficiency of the immobilized lectins tested at concentrations used for membrane solubilization while cationic and zwitterionic detergents caused significant inhibition of Con A- and SBA-Sepharose activities. In sodium deoxycholate (greater than 1%) only RCAI-Sepharose retained its activity, whereas the activities of the other lectins were reduced dramatically. Low concentrations of sodium dodecyl sulfate (0.05%) inhibited only the activity of immobilized SBA, but at higher concentration (0.1%) and prolonged periods of incubation (16 h, 23 degrees C) most of the lectins were inactivated. These data are compared with previous reports on the use of detergents in lectin affinity chromatography, and the conditions for the optimal use of detergents are detailed.


Immunological determination of the order of folding of portions of the molecule during air oxidation of reduced ribonuclease. An immunological method is used to follow the folding of different portions of the reduced bovine pancreatic ribonuclease molecule during air oxidation. Antibodies that react specifically with segments 1-13, 31-79, and 80-124 of native ribonuclease, as they are folded, were purified by affinity chromatography, using antiserum to native ribonuclease and columns to which the ribonuclease fragments were attached. The kinetics of reaction between these prufied antibodies and refolded portions that are produced when reduced rebonuclease is oxidized by air demonstrate the presence of intermediate states of folding, and are consistent with folding of the anti-genic determinants in the order 80-124, 1-13, and 31-79. The relative stabilities of each of these segments to thermal denaturation in the native protein provide additional evidence that the native conformation of region 80-124 is a very stable one in the intact molecule. On the basis of these two types of evidence, it appears that segment 80-124 contains a nucleation site for the folding of the protein molecule.


Kinetics of the photoreduction of cytochrome b-559 by photosystem II in chloroplasts. The kinetics of the photoreduction of cytochrome b-559 and plastoquinone were measured using well-coupled spinach chloroplasts. High potential (i.e, hydroquinone reducible) cytochrome b-559 was oxidized with low intensity far-red light in the presence of N-methyl phenazonium methosulfate or after preillumination with high intensity light. Using long flashes of red light, the half-reduction time of cytochrome b-559 was found to be 100 +/- 10 ms, compared to 6-10 ms for the photoreduction of the plastoquinone pool. Light saturation of the photoreduction of cytochrome b-559 occurred at a light intensity less than one-third of the intensity necessary for the saturation of ferricyanide reduction under identical illumination conditions. The photoreduction of cytochrome b-559 was accelerated in the presence of dibromothymoquinone with a t 1/2 = 25-35 ms. The addition of uncouplers, which caused stimulatory effect on ferricyanide reduction under the same experimental conditions resulted in a decrease in the rate of cytochrome b-559 reduction. The relatively slow photoreduction rate of cytochrome b-559 compared to the plastoquinone pool implies that electrons can be transferred efficiently from Photosystem II to plastoquinone without the involvement of cytochrome b-559 as an intermediate. These results indicate that it is unlikely that high potential cytochrome b-559 functions as an obligatory redox component in the main electron transport chain joining the two photosystems.


Structure of pyruvate dehydrogenase complex. Comparison between freeze-etching and negative staining. Pyruvate dehydrogenase complex (pyruvate : lipoate oxidoreductase (decarboxylating and acceptor-acetylating), EC 1.2.4.1), from pig heart, was studied by spray freeze etching and negative staining. From freeze etching experiments an average particle weight of 7-10(6) was estimated. Negative staining after glutaraldehyde fixation and freeze etching of unfixed and prefixed enzyme solutions yielded no significant difference in particle dimensions: the majority of the isometric complex molecules measured approximately 400 A in diameter. Tantalum tungsten shadowed freeze etch replicas indicated that the surface of the complex is built up of globular units. The relative positions of these units are in good agreement with the model still under discussion.


The secondary structure of myelin basic protein extracted by deoxycholate. Because of the implication of myelin basic protein in some neurological diseases its in vivo structure is of particular interest. The protein is usually isolated using organic solvents and acid solutions and has previously been shown to contain little alpha-helical or beta-structure; but it is not known how the extraction methods influence the structure. Following recent observations that deoxycholate generally causes minimal structural perturbation when used to dissolve membrane proteins, this detergent has been used to extract the basic protein from bovine myelin. The protein contained in deoxycholate washes of myelin has been purified by gel chromatography and its secondary structure examined by circular dichroism spectroscopy. This protein and conventionally prepared bovine and human basic protein to which 1% deoxycholate has been added appear to have the same structure: they contain 8-14% more helical structure than the chloroform/methanol-extracted protein in pH 4.8 acetate buffer or in pH 9.15 Tris buffer. This conformational change is unaffected by addition of 0.25 M NaCl. The helical content will approach the upper limit if, as is expected, these ordered segments are short. It is suggested that basic protein may adopt this more ordered structure in myelin and possess activity not apparent in its water-soluble unordered conformation. Retention of its encephalitogenic activity following severe treatment may result from an ability to rapidly refold to the original conformation rather than from this activity being inherent in the unordered form.


[Immunoelectrophoretic analysis of mink sera lipoproteins fractionated by preparative ultracentrifugation]. Using the method of preparative ultracentrifugation, four lipoprotein fractions were obtained differing in hydrate density. By means of immunoelectrophoretic analysis of the fractions, it is found that mink sera (except chilomicrons) contains at least five lipoproteins; two of them, lipoproteins 2 and 3, bind the major bulk of lipids. Lipoprotein 1 is of very low density. Lipoprotein 2 mostly belongs to the fraction with density less than 1.100 g/ml, although antigenically identical molecules with higher density were identified. Lipoprotein 3 is much more heterogeneous antigenically, electrophoretically and in its density (from 1.006 to 1.210 g/ml and higher). This protein seems to contain several structurally similar, though not identical, apoproteins. Lipoproteins 4 and 5 are very dense (more than 1.210 g/ml) and are weakly stained with Sudan black. These data, together with the results obtained in our previous studies, indicate that lipoproteins 2 and 4 are marked with genetically alloantygenic determinants.


[Membranes of bacteria and mechanism of action of the antibiotic gramicidin S]. The cyclopeptide antibiotic gramicidin S taken at a concentration of 100--200 mkg/mg membrane protein rapidly increases the permeability of M. lysodeikticus protoplast membranes for substrates of respiratory chain and exogenous cytochromes c. Prolonged incubation of gramicidin S with protoplasts results in their lysis which is more fast at low temperatures. In contrast to natural gramicidin, a derivative of gramicidin S with acetylated amino groups does not inhibit either the micrococcus membrane dehydrogenase or the whole of respiratory chain and does not affect the osmotic barrier of protoplasts. Aliphatic diamines (at concentrations up to 0.1 M) and Ca2+ ions (10(-2) M) do not affect the functioning of the respiratory chain in isolated micrococcus membranes. Another derivative of the antibiotic with an increased distance of loaded amino groups from the cyclopeptide framework (diglycyl gramicidin S) affects the membrane in a way similar to that of natural gramicidin. Washing of gramicidin-treated membranes with NaCl enhances the inhibitory effect of the antibiotic on membrane enzymes. The data obtained suggest that in addition to ionic interactions some hydrophobic interactions also occur during gramicidin S binding to the bacterial membrane, probably at the expense of a hydrophobic peptide ring. It is assumed that gramicidin S, similar to Ca2+ and some other membranotropic agents provides for phase separation of negatively charged phospholipids from other groups of phospholipids, manifesting itself in an appearance of "frozen" sites on the membrane which destroys its barrier properties. This is due to the formation of ionic bonds of negatively charged phospholipids. Simultaneously, unlike Ca2+, gramicidin S, when interacting with membrane proteins, prevents their redistribution in more liquid parts of the membrane, which results in a situation when the respiratory enzymes become surrounded by alkyl chains with restricted motion.


Rhodopsin and other proteins in artificial lipid membranes. Some basic aspects of incorporation of hydrophobic peptides and proteins in artificial lipid membranes are discussed. As examples valinomycin as a carrier model and gramicidin A as a channel former in lipid vesicles and in planar lipid membranes are presented. In the second part of the lecture some examples of incorporation of membrane proteins into lipid vesicles and planar lipid membranes are reported. The interaction with artificial lipid membranes of the Ca++ ATPase from the sarcoplasmic reticulum, of Rhodopsin, and of Bacteriorhodopsin is presented.


[Effect of glucocorticoids on alpha-fetoprotein synthesis and liver structure in mice acutely poisoned with carbon tetrachloride]. Experiments on male C57BL mice demonstrated that the CCl4 inhalation for 15 minutes in a concentration of 0.05 ml to 4 l of air caused deep dystrophic lesions of the liver with the signs of necrobiosis. The signs of regeneration and cellular infiltration appeared almost in 24 hours accompanied by the secretion of alpha-fetoprotein into the blood. The foci of necrobiosis disappeared completely in 4 days, being replaced by the lymphoid cells with large hepatocytes in the circumference. In the animals given hydrocortisone (20 mg/kg) or dexamethasone (2 mg/kg) for therapeutic or prophylactic purpose the regenerative processes were decreased, and the number of animals producing alpha-fetoproteins fell from 92 to 60--65%.


[Adaptation to the action of several teratogens as a consequence of preliminary administration of pesticides to females]. The effect of DDT and lindane pesticides on the intensity of the teratogenic action of sodium acetylxalicylate (SA) and of the cabomate benlate group of pesticides was studied on Wistar rats given the mentioned pesticides from the onset of pregnancy. The teratogens were administered on the 10th and the 12th days of gestation, respectively. Preliminary administration of these pesticides was found to weaken the teratogenic and the embryotoxic action of benlate given in a dose of 250 mg/kg, and of SA administered in a dose of 400 mg/kg. When SA was given in a dose of 600 mg/kg preliminary administration of the pesticides decreased the postimplantation mortality of the embryos, but the number of fetuses with developmental anomalies was the same as in the isolated action of the preparation given in this dose.


[Rhythmicity in the structure-functional changes in hepatocytes following exposure to pesticides]. The processes of dursban biotransformation during the liver perfusion in rats exposed to CCl4 and milbex for 4, 8 and 15 days (microsomal enzyme inhibitor and inducer, respectively) were studied. The ultrastructure of hepatocytes was examined in the animals treated with the mentioned poisons. Three stages of the structural and functional changes were revealed. Reconstruction of the dursban biotransformation ways occurred from the 8th day, i.e. the dialkylation processes were intensified, resulting in production of less toxic metabolites.


[Morphologic and histochemical characteristics of frog skeletal muscle following application of colchicine to a motor nerve]. Experiments with application of colchicine to the muscle motor nerve carried out; this was done for the purpose of disturbance of rapid axoplasmic transport. A reduction of the areas of transverse sections of the muscle fibers, an increase in the number of fibers with a low succinic dehydrogenase (SDH) activity a greater homogeneity of the muscle fibers by the degree of optic density in staining for detection of the SDH activity was noted. Analogous changes were revealed under conditions of section of the motor nerve. However, denervation was accompanied by the block of conductivity and by degenerative changes in the nerve endings. As to the preparations treated with colchicine, transmission of excitation in the nerve and through the synapse was retained and was recorded by the end plate miniature potentials, end plate potentials and the action potentials of the muscle fibers. A conclusion was drawn that rapid axoplasmic transport brought substances maintaining differentiated state of the muscle fibers.


[In vitro and inhibition of anaphylactic reactions 1-methyl-3-isobutylxanthine]. 1-methyl-3-isobutylxanthine (MIBX), a potent antiphosphodiesterase drug, inhibited the anaphylactic reactions both in vitro and in vivo: anaphylactic histamine release from the isolated mast cells of rats, anaphylactic contracture of the isolated trachea of guinea pigs, passive cutaneous anaphylaxis (PCA) in mice and anaphylactic bronchial constriction (ABC) in guinea pigs. MIBX inhibited the anaphylactic reaction of the trachea, PCA and ABC more than the corresponding reactions induced by histamine.


[In vitro synthesis of alpha-fetoprotein by individual hepatocytes and microcolonies of human hepatocytes]. Alpha-fetoprotein (AFP) produced by individual hepatocytes and hepatocyte microcolonies was detected with microelectrophoresis-precipitation in polyacrylamide gel. Hepatic cells of 6--13-week-old human embryos were cultivated in vitro for 2 to 5 days. 23 of 28 individual cells, and 89 of 91 microcolonies, built up of 2--35 cells, were demonstrated to produce AFP within the range of 70--800 pg per cell.


[Several structural characteristics of vital dyes]. Lymphocytes and macrophages of normal rats, cells of lymphosarcoma and ovarian carcinoma of rats (CC strain), cells of sarcoma 37 and lymphosarcoma Nk/Ly of mice were investigated for their capacity for vital staining by 86 days in concentrations within the range of 10(-5)--3-10(-2) M. It was found that alkylation of amino groups of the basic dyes increased their capacity for penetrating the living cells and depositing in their cytoplasm in cases of thiazine, oxazine, acridine, xanthene, diazine, and triphenylmethane compounds. The acid radicals decreased the capacity of the dye to vital staining of the cells considerably. The degree of basicity of the dye molecule and its amino groups played no decisive role in the process of vital staining.


[Modification of the method for staining DNA with basic dyes]. The author studied the mechanisms and the applicability in histochemistry of the sodium bisulfate treatment with subsequent toluidine and methylene blue staining after Felgen's hydrolysis. Bisulfite treatment proved to increase the reaction intensity 11/2-fold; the stain is bound stoichiometrically. Toludidine blue results in a metachromatic and anisotropic staining of the cell nuclei. The method is recommended as a sensitive test for DNA in cytochemical investigations and for the study of dichroism of the DNA-containing structures.


[Advantages in immunofluorescence of dichlortriazinylaminofluorescein conjugates isolated by single-step gel filtration]. The author describes a simple method obtaining dichlortriazinylaminofluoresceine (DCTAF) conjugates to human immunoglobulins and separation of their optimally labeled fractions. The single-stage method of the conjugate release from the nonbound flurorchrome and of a simultaneous separation of the optimally labeled fractions was carried out on Sephadex G-25 at pH 5.6--6.6 and at low ionic power of the eluate. Proteins overloaded with the dye are sorbed on gel according to their isoelectrical points.


Abnormalities in granule formation in acute myelogenous leukemia. Granule formation was investigated in differentiating neutrophils of a patient with acute myelogenous leukemia (AML) by means of the combined techniques of electron microscopy and peroxidase cytochemistry. Two important pathologic features were observed: first, an abnormal concentration and packaging of peroxidase into Auer rods in leukemic promyelocytes, and second, the presence of Auer rods surrounded by single-unit membranes in some mature polymorphonuclear leukocytes (PMN). An additional unexpected finding was the discovery of two distinct populations of PMN circulating concurrently; a minor (less than 5%) normal one that contained both peroxidase-positive azurophilic and peroxidase-negative specific granules and a major abnormal one characterized by the absence of specific granules. None of these abnormalities was observed during the two remissions of this patient's disease. During relapse a "hiatus leukemicus" occurred, which also revealed two populations of cells, a majority population of leukemic blasts, and a minority population of a few normal PMN. These findings documented several developmental abnormalities in the differentiating cells of myelogenous leukemia and also suggested that concurrent normal and abnormal populations of PMN may be a helpful diagnostic feature of a leukemic process.


Group-specific human granulocyte antigens on a chronic myelogenous leukemia cell line with a Philadelphia chromosome marker. Group-specific human granulocyte antigens are serologically detectable with granulocytotoxic-positive human alloantisera on a cell line, K562, of chronic myelogenous leukemia origin which bears a Philadelphia chromosomal marker. The same cell line lacks serologically detectable HLA, B2 microglobulin, and B-lymphocyte antigens. Granulocyte antigens are important cell markers for cell lines of suspected myeloid lineage.


Combination chemotherapy of advanced non-Hodgkin lymphoma with bleomycin, adriamycin, cyclophosphamide, vincristine, and prednisone (BACOP). Seventy-three patients with advanced non-Hodgkin lymphoma were treated with bleomycin, Adriamycin, cyclophosphamide, vincristine (Oncovin) and prednisone (BACOP), administered intensively during a 7-wk induction course followed by intermittent cycles every 3 wk for a total of 28 wk. The objective response in 44 evaluable nonleukemic patients with diffuse histology was 86%, with 66% achieving a complete remission (CR), varying from 80% for diffuse poorly differentiated lymphocytic (DPDL) to 56% for diffuse histiocytic (DH) lymphoma. In patients with nodular histology 89% (8/9) achieved a CR with a projected 75% of patiients in CR at 14 mo. Median follow-up from time of CR for nodular histology was 17 mo. The projected median duration of CR in diffuse histology was 14 mo. with median survival 14 mo. Patients with a partial response survived a median of 7 mo, compared to 3 mo for nonresponders. Of 29 patients with diffuse histology, 17 (59%) have remained disease free for 5-34 mo with a median follow-up of 12 mo. Survival beyond 20 mo has been projected for 42% of patients with diffuse histology (58% with DPDL and 32% with DH). The central nervous system (CNS) was involved in a total of 11/44 (25%) patients with diffuse histology, including 5 with primary CNS relapse. BACOP resulted in a higher CR rate and longer survival than a previous three-drug program (COP), especially in patients with diffuse histology.


Eosinophil colony formation in semisolid cultures of human bone marrow cells. A method for in situ staining and scoring of eosinophil colonies and clusters in the whole culture dish has been developed for agar cultures of human marrow cells. Cultures stimulated by human placental conditioned medium were found to develop the same proportion of eosinophil, neutrophil, and monocyte colonies as cultures stimulated by conventional peripheral white cell underlayers. Eosinophil colonies and clusters always contained pure populations of eosinophils, and no mixed colonies of eosinophils with other cells were observed. Eosinophil colony formation was delayed in onset relative to neutrophil or monocyte colony formation, but by day 14 eosinophil colonies comprised 20% of all colonies. Eosinophil colonies were absent in cultures of marrow cells from patients with acute myeloid leukemia in relapse, but eosinophil colonies and clusters were found with higher than normal frequency in cultures from acute leukemic patients in remission. A high frequency of eosinophil colonies and clusters was also observed in cultures from three patients with idiopathic thrombocytopenic purpura. The new staining procedure should permit a wide range of studies on eosinophil precursor populations in the marrows of patients with various diseases.


[Problems of conductor demonstration in haemophilia A: error possibilities in the determination of factor-VIII-associated antigen]. Difficulties in the detection of haemophilia A carriers by the determination of the ratio factor VII  coagulation activity to factor VII antigen concentration are shown in part one of our investigations. The second part evaluates methodical parameters of the quantitative immunoelectrophoretic technique. According to our results reliable data can only be obtained under the following conditions: 1. At least three determinations of each sample are required and the mean value should be used as the final result. 2. Each gel plate should be calibrated separately 3. The calibration line should include values less than 100%. Factor VIII concentrates are not suitable for determination of the calibration curve, as higer dilution results in lower calculated concentration of the undiluted concentrate. Storage of pooled normal plasma at--25 degrees C is possible without significant loss of antigen concentration over 5 months. However, plasma samples of a single person show an unpredictable variation in the antigen concentration during storage over the same time.


Lead levels in small mammals and selected invertebrates associated with highways of different traffic densities. Lead levels in small mammals were studied in 6 areas -- 3 associated with highways of different traffic densities, and 3 control sites in forested areas. Lead levels were also studied in insects from 2 traffic areas and in earthworms from different distances from 2 traffic areas. Lead levels were low in insects ranging from 3.84 microng/g to 3.35 microng/g. Lead levels in earthworms were highest in those recovered closest to highways and from the area with the higher traffic volume. Lead levels in mammals generally was highest in mammals from areas of higher traffic densities. Higher levels of lead were found in shrews than in Peromyscus and Microtus.


Prostatic obstruction in Sudan.

Prostatic obstruction in Sudan. Prostatic obstruction is not uncommon among Africans. Senile hypertrophy of the prostate is a common condition in Northern Sudan, while it is rare in the south. This difference in incidence is probably due to environmental rather than genetic factors. Carcinoma of the prostate exists in both regions.


Homocytotropic antibodies (IgE) to Neisseria gonorrhoeae in the rat and a cross-reactivity of heterologous gonococcal strains. Outbred Wistar rats were immunised with a single intraperitoneal injection of a mixture of 30 mg of A1(OH)3 and 100microng of gonococcal zeolite antigen (ZA). Ten days after immunisation, ZA prepared from T1 and T4 colonies of Neisseria gonorrhoeae strain GC6 (GC6-T1 ZA and GC6-T4 ZA) were able to elicit a reaginic (IgE) response which declined to low levels by 35 days. There was a significant amount of histamine release from the mast cells of actively-sensitised rats on challenge with a specific gonococcal antigen. The antisera heated at a temperature of 56 degrees C for four hours failed to elicit a passive cutaneous anaphylaxis (PCA) reaction, indicating a lack of IgGa antibody. In addition antisera to GC6-T1 ZA gave a positive PCA reaction with GC6-T4 ZA and ZA prepared from T1 colonies of six heterologous gonococcal strains suggesting that these strains of N. gonorrhoeae share common antigenic determinants.


Axoplasmic flow in myelinated and unmyelinated nerves. [14C]Leucine was injected into the floor of the IVth ventricle of rabbits in order to label the axoplasmic flow of proteins in the efferent fibres of the vagus nerve. At intervals from 6 h to 180 days after injection, the rabbits were sacrificed and the cervical and abdominal vagus and recurrent laryngeal nerves removed. The radioactivity in the protein, lipid and low molecular weight fractions of the nerve segments was estimated by liquid scintillation counting. Two rates of axoplasmic flow were thus demonstrated in the abdominal vagus nerve; a fast rate of about 450 mm/day and a slow rate of 30 mm/day. This agrees very well with previous work. Peaks of activity in the recurrent laryngeal nerve corresponded to rates of 9 and 270 mm/day, but their level was low, being not significantly above sciatic nerve background levels. The number of efferent fibres in the two nerves and the corresponding axonal areas were calculated in order to estimate the amount of labelled material in the flow. The abdominal vagus was shown to contain 4-5 times as much label per unit area of axoplasm as the recurrent laryngeal. A difference was therefore demonstrated in the same anatomical nerve between myelinated fibres in the recurrent laryngeal and unmyelinated fibres in the abdominal vagus nerve, both in the rates of flow and in the amounts of material contained in the flow.


Release of protein from axons during rapid axonal transport: an in vitro preparation. An in vitro system from the frog was used to study fast axonal transport and determine if transported protein is released from the axons. This preparation included the eighth and ninth dorsal root ganglia with their roots, sciatic nerve and gastrocnemius muscle. The preparation was placed in three-compartment chamber with each compartment separated by a silicone grease barrier. The dorsal root ganglia were incubated in [14C]leucine for 5 h in compartment A. The labeled protein was transported down the axon from compartment A to compartment B. The sciatic nerve in compartment B was superfused with frog Ringer. This solution was collected in hourly samples and dialyzed to remove unincoprorated leucine before counting. Incubating the ganglia in 100 microng/ml cycloheximide in frog Ringer blocked the release of labeled protein from the axon. Superfusing compartment B with solution containing 100 microng/ml cycloheximide inhibited axonal and Schwann cell protein synthesis, but did not block the release of labeled protein. It was concluded that the labeled protein released into the superfusing solution was synthesized in the ganglia and transported to the axon before release. SDS acrylamide gels were used to separate the labeled proteins. Sectioning the gels in 2 mm slices and determining the radioactivity showed that 80-85% of the counts were contained in two fast moving bands.


A positive difference in nature of envelopes of thymus- and uterus-derived leukemia viruses of AKR mice. Thymus-derived leukemia virus of AKR/J mice was inactivated by anti-theta antiserum. But it was not inactiviated by the antiserum which had been absorbed with intact thymus cells of AKR/J or RF/J mice, and by anti-uterus-derived leukemia virus antiserum. In contrast, uterus-derived leukemia virus of the strain was not inactivated by anti-theta antiserum, but was neutralized by anti-uterus-derived leukemia virus antiserum. The results suggest the possibility that some constitutents of the envelope of thymus-derived leukemia virus are derived from the plasma membrane of thymus cells of AKR mice at the time of budding and that such constituents are not associated with the enveloped of uterus-derived leukemia virus.


Combination chemotherapy of MOPP-resistant Hodgkin's disease with adriamycin, bleomycin, dacarbazine and vinblastine (ABDV). Twenty-four patients with advanced Hodgkin's disease, resistant to the MOPP regimen, were treated with a combination of Adriamycin, bleomycin, dacarbazine and vinblastine (ABDV). Fifteen (63%) achieved objective remission, 14 partial remissions and one complete remission. The median duration of remission in this group of patients was 6.5 months; four of the 15 patients are still in remission (8+, 8+, 9+, 10+ months). Objective remission occurred rapidly within 1.5 months. Regression was evident in disease within nodes, lung, liver and bone. Toxic manifestations caused by ABDV were well tolerated and reversible. In one patient death was directly attributed to drug toxicity. This combination has produced a better rate and duration of remission than that reported with single agent chemotherapy in MOPP-resistant patients with Hodgkin's disease. In our hands, ABDV did not equal the recent results reported with Bleomycin-CCNU-Velban in  a seemingly similar group of patients.


Bleomycin serum pharmacokinetics as determined by a radioimmunoassay and a microbiologic assay in a patient with compromised renal function. Serum and plasma bleomycin concentrations were determined in a patient with renal dysfunction at two creatinine clearances. The results obtained with a new radioimmunoassay and the microbiologic assay were compared. It was shown: 1) that the clearance of bleomycin from the blood is markedly retarded in severe renal dysfunction, 2) that clearance of bleomycin varies with creatinine clearance, 3) that bleomycin is probably not dialyzable, 4) that determinations on serum and plasma were equivalent, and 5) that the radioimmunoassay and microbiologic assays gave equivalent results (P less than 0.001).


A phase II study of adriamycin (NSC 123127) in patients with hepatocellular carcinoma from Zambia and the United States. Nineteen Zambian and 22 American patients with hepatocellular carcinoma were treated with Adriamycin every three weeks in intravenous doses ranging from 20-75 mg/m2 (depending upon their initial serum bilirubin levels). Four of 16 (25%) "good risk" Zambian and American patients who received 75 mg/m2 had objective responses, while in five additional patients there was evidence of either transient tumor regression or disease stabilization. In contrast three of 25 "poor risk" patients who received 20-60 mg/m2 had objective responses. Even in this latter group, however, transient, objective signs of tumor regression were noted in four patients. The results of the present study confirm previous reports suggesting anti-tumor activity for high doses of Adriamycin in hepatocellular carcinoma. Since those responses seen were generally incomplete and transient, further clinical trials of this agent used in combination or sequentially with other agents are indicated.


Biochemical evidence for viral-like characteristics in cerebrospinal fluids of brain tumor patients. Previous work done in this laboratory has shown that human Central Nervous System tumors possess attributes in common with those found in RNA tumor viruses. Because of the intimate physical relationship between the circulating cerebro-spinal fluid and the Central Nervous System, particles derived from tumors in this system may very well be shedded into the surrounding fluid system thus displaying similar biochemical attributes. Twenty samples of cerebro-spinal fluids were examined. Fifteen were from patients with various types of tumors in various location of the Central Nervous System. Five were from patients with unrelated diseases. Eleven or 74% of those from tumor patients were found to be positive when simultaneous detection assays were done on their core structures. None of the control was positive. The [3H]DNA synthesized from core structures hybridized readily to polysomal RNAs from a solid tumor. Particles from them were found to have a density of 1.867 g/ml sucrose gradient.


Properties of cell lines established from transitional cell cancers of the human urinary tract. Characterization studies have been carried out on eight cell lines (253J, 192B, 639V, 647V, 486P, 575A,743E, and 751G) established from transitional cell cancers of the human urinary tract. Although subtle morphological differences exist among individual lines, each has an epithelial morphology and exhibits multilayering. The doubling times for the cells range from 20 to 56 hr, and at least a 1-to-3 split can be achieved when they are subcultured every 4th day. Karyotypic analysis revealed a hyperdiploid stemline for each cell line, and presence of a Y chromosome was confirmed by Q banding in five of the lines. The tumorigenic nature of the cell lines was demonstrated by their production of tumors in hamsters and confirmed by colony formation in agar. The transitional cell cancer lines were shown to be free of Mycoplasma, and their glucose-6-phosphate dehydrogenase mobility patterns and their Karyotypes prove that they are not HeLa cells.


Isolation and identification of human lung tumor-associated antigens. Three human lung tumor-associated antigens (TAA's) have been identified in soluble and membrane-solubilized extracts of human squamous cell lung carcinoma with the use of antisera raised in rabbits. The antigens were identified and partially characterized by means of an agarose adsorption technique. These antigens, termed lung TAA's 1,2, and 3, are all soluble in 50% ammonium sulfate, are antigenically distinct, and do not cross-react with carcinoembryonic antigen or alpha-fetoprotein. Lung TAA's 1 and 2 are oncofetal antigens demonstrable in soluble extracts from 24-week-old but not from 26-week-old fetal lungs. Rabbit antibodies to these lung TAA's were not adsorbed by types A, B, and O human red blood cells, serum proteins as well as soluble or insoluble lung preparations. Of several commercial antisera to human proteins, none cross-reacted with lung TTA 1, but anti-human liver ferritin cross-reacted with lung TAA 2, and anti-human lactoferrin cross-reacted with lung TAA 3. Lung TAA 1 was partially adsorbed and cross-reacted with certain normal serum or plasma preparations used and appears to be a normal serum protein in Cohn Fraction IV-4. Lung TAA 2 and 3 appear only in lung tumor-soluble extracts, whereas the lung TAA 1 was demonstrable in soluble extracts of breast, colon, cervical and head and neck carcinoma. All may be tumor markers of value in immunodiagnosis.


Effect of bleomycin on the synthesis and function of RNA. Bleomycin inhibits cellular RNA synthesis and the inhibition is nonspecific. The ratio of polyadenylate- [poly(A)] containing RNA to non-poly(A)-containing RNA in the drug-treated human lymphocytic cells, line Wil2, was the same as that in untreated cells. Poly(A) RNA isolated from untreated cells was used as a template for reverse transcriptase to synthesize complementary DNA, which was then used as a probe to assay the sequence diversity of poly(A)RNA's from treated and untreated cells. It was found that essentially all of the poly(A) RNA's in the untreated cells were also present in the treated cells. The effect of bleomycin on the biological activity of messenger RNA (mRNA) was tested with globin mRNA in a wheat germ embryo translation system. Although bleomycin inhibited protein synthesis at high concentrations, the inhibition was not due to a modification of mRNA. This was evidenced by the fact that no decrease in the ability of mRNA to function in the test system was found when globin mRNA was pretreated with high concentrations of bleomycin followed by removal of the drug.


Inhibition of the lethality of bleomycin A5 in L-cells by hirudonine. The lethality of several individual bleomycin derivatives, i.e., the spermidine derivative (A5), The dimethyl sulfonium aminopropyl derivative (A2), and the agmatine derivative (B2), was compared on mouse fibroblasts. The spermidine derivative, bleomycin A5, was the most toxic, producing more than a 2-log drop in clonability in 50 hr at 20 microng/ml. 6-Azauridine, a relatively nonlethal inhibitor of RNA synthesis and cell multiplication, produced a 60% decrease of adenine incorporation into nucleic acids without inhibiting the lethal action of A5. This result differed from the effects of inhibition of RNA synthesis on the lethality of A5 in Escherichia coli. Hirudonine (1,8-diamidino-spermidine) markedly and specifically inhibited the lethal effects of A5 in L-cells but not in E. coli. However, hirudonine did not affect the toxicity of A2 and B2, separately or together, as it did in the mixture used clinically. Nor did arcaine (diamindinoputrescine) reduce the lethality of the agmatine (monoamidinoputrescine) derivative, B2.


In vitro stimulation and inhibition of tumor cell growth mediated by different lymphoid cell populations. Spleen, lymph node and thymus cells from normal C57BL/6 mice or mice given injections 6 to 20 days previously with syngeneic methylocholanthrene-induced fibrosarcoma (B6MCA) cells were tested for their ability to mediate growth stimulation and/or inhibition of B6MCA cells in vitro. At an effector cell to tumor cell ratio of 10:1,, nucleated spleen cells from mice bearing the tumor for 6 days enhanced tumor cell growth. Neither lymph node nor thymus cells from tumor-bearing mice were stimulatory. Tumor cell growth was inhibited by either spleen or lymph node cells only when a ratio of 1000:1 was exceeded. Thymocytes, however, were inhibitory at a ratio of 100:1,. Lymphoid cells from normal mice were not stimulatory at low ratios or inhibitory at any ratio tested. Both stimulatory and inhibitory activities disappeared by Day 20. Filtration of sensitized spleen cells through a glass wool column did not remove stimulatory or inhibitory activities. Subsequent passage through a nylon wool column resulted in a loss of stimulation, but not inhibition, of tumor cell growth. Stimulatory activity was completely abrogated by treatment with either anti-theta or rabbit anti-mouse gamma-globulin serum. A 1:1 mixture of spleen cells treated with either antisera did not restore stimulation. Spleen cells from T-cell- or B-cell-deficient mice bearing the tumor for 6 days were also not stimulatory. The results suggest that immunostimulation of tumor growth in vitro is mediated by a lymphoid cell that is distinct from the cytotoxic effector cell.


Inhibition of the reverse transcriptase of bovine leukemia virus by antibody in sera from leukemic cattle and immunological characterization of the enzyme. Sera from some leukemic cattle contain an antibody that inhibits the reverse transcriptase activity of the bovine leukemia virus. The antibody is not directed against the synthetic template or the major internal and envelope viral antigens. The antibody failed to inhibit the DNA polymerases of the murine leukemia virus, simian sarcoma-associated virus, avian myeloblastosis virus, or Escherichia coli. Conversely, the bovine leukemia virus enzyme was not inhibited by antibody against the reverse transcriptases of other C-type viruses. These findings agree with previous results showing that the major internal bovine leukemia virus protein lacks the known interspecies- and intraspecies-specific antigenic determinants indentified in the homologous proteins of other oncornaviruses.


In vitro cytotoxic and biochemical effects of 5-aza-2'-deoxycytidine. The in vitro effect of 5-aza-2'-deoxycytidine (5-aza-CdR) on cytotoxicity and macromolecular synthesis in A(T1)C1-3 hamster fibrosarcoma cells was investigated. The in vitro concentrations that produce 50% cell kill for 5-aza-CdR were about 1.0 and 0.01 microng/ml for a 2- and 24-hr exposure, respectively. 5-aza-CdR inhibited the growth of the fibrosarcoma cells by 40% at a concentration of 0.05 microng/ml. Deoxycytidine, but not cytidine, was a potent antagonist of the cytotoxicity produced by 5-aza-CdR. At cytotoxic concentrations 5-aza-CdR did not appear to inhibit DNA, RNA, or protein synthesis during a 1-hr incubation as measured by the incorporation of radioactive thymidine, uridine,, or leucine into acid-insoluble material. At a concentration of 10 microng/ml, 5-aza-CdR stimulated the incorporation of radioactive thymidine into DNA by more than 50%. These results indicate that 5-aza-CdR is a very potent cytotoxic agent to tumor cells in vitro at concentrations that do not inhibit macromolecular synthesis.


Increased primary cell-mediated immunity in culture subsequent to adriamycin or daunorubicin treatment of spleen donor mice. Spleen cell populations from mice treated with Adriamycin or daunorubicin were found to develop a greater complement-independent cellular cytotoxic immune response during culture with allogeneic tumor cells than spleen cells from untreated or cyclophosphamide-treated animals. A temporal and drug dose dependence of this effect was demonstrated. The changes in spleen cell population occurring in the donor mice consequent to drug treatment were evident in the nylon woll-adherent fraction of the spleen cells. The results are consistent with the possibility that the concentration of specific progenitor or accessory cells in the spleen is increased consequent to drug treatment.


Reactivity of human brain antiserum with neuroblastoma cells and nonreactivity with thymocytes and lymphoblasts. Brain-associated antigens have been detected on human and mouse thymocytes. Also, murine neuroblasts and brain cells have common antigens. In this study we compared the reactivity of rabbit anti-human brain (HB) serum with neoplastic neuroblasts and normal and neoplastic lymphoid cells. The binding of HB antiserum to viable cells was assessed by immunofluorescence and an indirect radiolabeled antibody assay. HB antiserum reacted with greater than 80% of neuroblasts derived from two human cell lines and five children with neuroblastoma, but with less than 1% of human thymocytes, bone marrow lymphoid cells, and lymphocytic leukemia cells. HB antiserum also reacted with 5 to 10% of peripheral blood lymphocytes. Absorption with neuroblasts did not alt-r this reactivity. Rabbit antisera raised against normal human thymocytes and leukemic T-cells specifically bound to thymocytes but did not bind to neuroblasts. The reactivity of anti-HB serum against SK-N-SH neuroblasts was removed by absorption with HB, but not with human kidney or liver, or mouse and guinea pig brain. We conclude that human neuroblastoma cells possess cell-surface antigens that are present on HB. These antigens appear to be species specific and are not present on normal or malignant thymic cells. Conversely, thymus-associated antigens are not expressed on neuroblasts.


Macrophage processing of antigen for induction of tumor immunity. Previous studies have indicated, that, after in vitro incubation of antigen with macrophages, the "processed" antigen preferentially induces cell-mediated immunity. To investigate this phenomenon with tumor antigens, mycobacteria-stimulated macrophages were incubated with irradiated syngeneic EMT6 tumor cells for varying lengths of time and injected into normal mice. On subsequent challenge with EMT6, there was a significant increase in protection in mice immunized with macrophage-processed tumor antigen over control animals. Mineral oil-stimulated macrophages were also capable of processing irradiated EMT6, but macrophages induced by thioglycollate or proteose peptone were not. Freeze-thawed mycobacteria-stimulated macrophages were nearly as effective as viable macrophages in processing tumor antigen, but heat-treated macrophages lost this capacity. The immunity generated was specific and could be passively transferred by immune cells but not by immune serum. The results indicate that incubation of tumor antigen with appropriately activated macrophages leads to the enhanced induction of immunity to the tumor. Macrophage enzymes may degrade tumor antigens to fragments with few antigenic determinants that preferentially induce cell-mediated immunity.


The clinical and physiological implications of hepatoma B12-binding proteins. A serum B12-binding protein with increased sialic acid content (termed hepatoma B12-binding protein) that causes elevations of serum B12 and unsaturated B12-binding capacity has been found in some patients with hepatocellular carcinoma (hepatoma). We now report another patient with hepatoma with initial near-normal, unsaturated B12-binding capacity that increased 400-fold as the disease progressed and then fell 50% with response to chemotherapy. A perfusate of the tumor in the liver had 5 times more B12-binding protein than did the serum and was immunologically the same as the serum hepatoma B12-binding protein isolated from previous cases. A cell line derived from hepatoma produced significant amounts of B12-binding protein similar to hepatoma B12-binding protein, whereas cell lines from normal liver and other neoplasia did not. The hepatoma sera, perfusate, and media from the hepatoma cell line contained elevated sialyltransferase activity. These data suggest that some hepatomas produce increased hypersialylated B12-binding protein that is cleared slowly from the plasma and accumulates there as hepatoma B12-binding protein.


Effects of bleomycin on the epidermal content of growth-regulatory substances (chalones). Hairless male mice were given 2 mg Bleomycin i.p. on two successive days. At different time intervals from 1 to 10 days after the last Bleomycin injection, groups of animals were killed and water extracts of hemogenized skin were made. These extracts, supposed to contain the epidermal G1 and G2 chalones, were injected into female hairless mice, and their growth inhibitory potency determined by two methods. 5 mg of lyophilized crude skin extract were injected i.p. together with Colcemid, and the animals killed 4 hr later. The number of Colcemid-arrested mitoses was determined, and was considered to be a measure of the G2 inhibitor present in the skin extracts. 10 mg of the same extracts were injected i.p., and these animals also got 3H-TdR i.p. 12 hr later, and were killed after a subsequent 30 min. The epidermal LI was determined, and was considered to be a measure of the epidermal G1 factor in the skin extracts. The results obtained were compared to the effect of Bleomycin alone and to the effects of skin extracts from non-Bleomycin-treated animals. The results show that Bleomycin provoked slight alterations in the growth-inhibitory potency of the G1 chalone, whereas significant effects were seen in the G2 chalone, There was an increased amount of growth-inhibiting factors on days 2 and 3, and on days 8-10. The results are discussed and it is concluded that the most probable hypothesis is that Bleomycin, in addition to its known inhibition by accumulation of cells with high growth inhibitory potency. An initial, additional direct effect of Bleomycin on the chalone system cammot be excluded.


[Solubility of the neurosecretory products of Crepidula fornicata Phil., a prosobranch mullusk]. Study of solubility of neurosecretory products in Crepidula fornicata shows that the presence of ethanol in the fixative is incompatible with good fixation. This character permits the choice of a more effective extraction technique. An experimental study in organ culture shows that the action of differentiation of brain appears in the extract obtained by this method.


[The neurosecretory cells of the cerebral ganglia and the nerve chain in the isopod crustacean Asellus aquaticus L. Cyclic variations in the beta type cells as a function of reproduction]. Asellus aquaticus L. cerebral ganglions and nervous cord contain seven types of neurosecretory cells; two of these are similar to Malacostraca beta and alpha cells. Female beta cells are the seat of cyclic acidophil variations which are connected to ovocyte growing; this acidophil secretion is superimposed on the basophil constant secretion in male and female beta pericaryons.


[Stimulation by isoprinosine of the antiviral effect of interferon]. With a single dose of interferon, isoprinosine greatly enhances the antiviral protection of Mice against the lethal effect of encephalomyocarditis virus (100 LD 50).


[Isolation, from anti-dinitrophenyl sera taken at the beginning of immunization, of a protein combining with the hapten and which is not related to immunoglobulins]. A protein, not antigenically related to the immunoglobulins, has been isolated by affinity chromatography from the serum of rabbits immunized with dinitrophenylated human gamma-globulin. Its concentration in the serum reaches maximum during the first three days which follow the injection. On "Sephadex G-200", it is eluted after the ribonuclease. Its electrophoretic mobility is slower than that of the IgG.


Improved staining characteristics of serum lipids after halogenation and esterification of thin-layer chromatograms. We describe a procedure for treatment of thin-layer chromatographic serum lipid patterns so that they may be stained by dyes for evaluation by densitometry. After development of the chromatogram [Clin. Chem. 18, 384 (1972)] the plates are dried and sprayed with butyryl chloride. This esterifies the free cholesterol. After drying, the plate is treated with iodine monobromide, to add iodline to the double bonds. The triacylglycerols (triglycerides), cholesterol esters, free cholesterol, and phospholipids are now all in the form of esters with no unsaturated double bonds. The free fatty acids are also now saturated. The chromatogram is now stained with basic fuchsine in acetate buffer (0.1 mol/liter, pH 5.0). Excess stain is removed with a buffered solution of guanidine hydrochloride. Erythrosine B may also be used. With basic fuchsine the background will be a uniform pink. With erythrosine B the background is white, but the stain tends to be washed out of the free fatty acids. The chromatograms are evaluated by densitometry, with use of a 540-nm filter for basic fuchsine and a 520-nm filter for erythrosine B. The stained chromatograms and densitometric scans accurately represent the relative concentration of the various lipid fractions as compared to that of an internal standard, and correlate with the nature of the disease being explored.


Prenatal diagnosis of Meckel syndrome: alpha-feto protein and beta-trace protein in amniotic fluid. Meckel syndrome was diagnosed prenatally by alpha-feto protein and beta-trace protein determinations in amniotic fluid. No central nervous system anomalies were detected in the affected fetus, who presented with large polycystic kidneys and polydactyly. An excessive synthesis of these fetal proteins by the dysplastic kidneys is suggested, allowing for the possibility of prenatal diagnosis of polycystic kidneys in families at risk for this disease. The present family emphasized the importance of amniocentesis in pregnancies at risk for Meckel syndrome, regardless of the presence of a defect in neural tube closure.


Cytological assessment of climacteric patients. The cytohormonal method using the upper lateral vaginal wall is an inexpensive, rapid, painless, effective and simple technique for measuring hormone effect in the human. With proper safeguards, one can evaluate the proliferative effect of oestrogens. The astute experienced clinician can use this test as one of several parameters to study the menopause and some of its related metabolic and neoplastic conditions. The cytohormonal proliferated condition (KI) cannot, and should not, be used as the only criterion for diagnosis and therapy. The presence of cells showing low levels of proliferation would be a valid measure of oestrogen lack and clinical need. This evaluation of the low end of the proliferation scale (low intermediate and parabasal cells) would more likely be useful and certainly more consistent with cytohormonal reality.


Protein clearances in chronic renal failure: a contribution to the pathophysiology of tubular dysfunction. Glomerular selectivity is still conserved in chronic renal failure (CRF), although it is poor when compared with non-CRF. In addition, clearances per unit nephron of 15 plasma proteins (Cprot./Ccr X 100) are markedly elevated in CRF. This reflects an effect of osmotic diuresis on tubular reabsorption of proteins, which may partially account for the tubular dysfunction shown by the augmented excretion of the low molecular proteins, such as lysozyme and insulin. Besides this, there exists certainly tubular damage, since tubuloglomerular protein ratio (Clysozyme/Ctransferrin) rises with the progress of glomerular injury.


A systematic approach to the bedside differentiation of cardiac murmurs and abnormal sounds. Fortified by knowledge gained from cardiac catheterization studies, a more precise physiologic approach to cardiac auscultation has evolved in recent years. This system involves observing changes in the characteristics of murmurs and abnormal sounds during respiration, the Valsalva maneuver, exercise, changes in posture, after pharmacologic agents and during arrhythmias, These techniques will aid in the accurate identification of the origin of the murmur or abnormal sound but do not necessarily indicate the presence or severity of cardiac disease. The auscultatory results must be interpreted in the light of the complete clinical evaluation, which may include the ECG, chest x-ray and echocardiogram. Thus, the careful physiologic approach to the physical examination represents a powerful noninvasive tool that, when combined with other noninvasive studies, can accurately diagnose cardiac disease in many patients without the necessity for cardiac catheterization.


The effect of cations on C-band formation in human chromosomes. A number of salt solutions were tested for their ability to replace 2 X SSC in the production of C-bands with the so-called CT technique. Those containing monovalent cations, except for the ones with Ag+ ions, were capable of inducing C-bands in Ba(OH)2-pretreated human chromosomes. Cesium cations were the most effective in this respect. In contrast to the effect of the monovalent cations studied, di- or polyvalent cations (and Ag+ cations) were incapable of producing C-bands, but appeared to stabilize the Ba(OH)2-induced reverse-banding pattern, This stabilizing effect was obtainable with relatively low cation concentrations and persisted even when C-band-producing cations were present at the same time. The experiments clearly show that the cations present during salt incubations, generally used in the CT- and several other banding methods, can exert a pronounced influence on chromosome behavior.


Studies on sex chromosomes of four hamster species: Cricetus cricetus, Cricetulus griseus, Mesocricetus auratus, and Phodopus sungorus. In this paper, we present an analysis of the sex chromosomes of four hamster species after application of different staining techniques. The mitotic X chromosomes show a striking similarity in G-banding pattern but rather great differences in their C-banding patterns. A presumably homologous euchromatic segment that exhibits two distinct G-bands appears in the X chromosome of each species. The Y chromosome of Cricetus cricetus is in contrast to those of the other species, because it reveals a relatively well-differentiated G- and C-banding pattern. In meiotic metaphase I, interstitial chiasmata can be found in the sex bivalents of Cricetus cricetus and Cricetulus griseus, whereas the gonosomes of Mesocricetus auratus and Phodopus sungorus sungorus are terminally associated. The regions that are involved in pairing or association are always heterochromatic.


Optical Studies of the interaction of 4'-6'-diamidino-2-phenylindole with DNA and metaphase chromosomes. The optical absorption and fluorescence characteristics of 4'-6-diamidino-2-phenylindole (DAPI) with DNA and chromosomes were studied. There is a decrease in extinction coefficient and chift in the absorption spectra to a higher wavelength when the dye binds to DNA. The fluorescence of DAPI is enhanced by both A-T and G-C base-pairs. The enhancement by A-T rich is significantly greater than by G-C rich DNA. The chromosomes and the constrictions of human chromosomes 1 and 16; these regions are known to contain A-T rich DNA and show dull fluorescence when treated with quinacrine. This dye may be useful for identifying A-T rich region in chromosomes. The fluorescence of DAPI bound to polynucleotides or chromosomes is partially quenched by the introduction of BrdU. This suppression of dye fluorescence allows optical detection of sister chromatid exchanges and chromosome region containing DNA with an unequal distribution of thymidine between polynucleotide chains after BrdU incorporation.


Selective staining of Y chromosomal loops in Drosophila hydei, D. neohydei, and D. eohydei. Modified Giemsa procedures have been developed which elicit differential and highly selective staining of individual Y chromosomal lampbruch loops in spermatocyte nuclei of Drosophila hydei, D. NEOHYDEI, AND D. eohydei. In all three species the Y loop pair known as the "clubs" stains a brilliant dark red with Giemsa at pH 10. With the same treatment other loop pairs either remain unstained, e.g. the "threads", or show a differentiation between light blue and pink staining material, e.g. "pseudonucleolus" and "cones" in D. hydei and D. eohydei. With eosin at pH 2.8 the "threads" in D. hydei can be stained intensely, as well as one matrical component of th "pseudonucleolus". Pretreatment with RNase or TCA removes all stainability from the Y loops with Giemsa at Ph 10. TCA treatment enhances eosin staining at pH 2.8. These and other variations of Giemsa may be utilized to establish homologies between Y loops in different species. The molecular basis of the staining reactions remains to be elucidated.


[Treatment of inoperable oesophageal malignancies with a thermal probe (author's transl)]. In inoperable stenosing tumours of the oesophagus the problem arises of maintaining oral food intake. As malignant cells are more cold-sensitive than normal cells oesophageal tumours may be "cold-coagulated" with special probes. Preliminary results with this treatment in 28 patients are reported.


Maturation of evoked potentials and visual preference in 6-45-day-old infants: effects of check size, visual acuity, and refractive error. Visual evoked potentials (VEPs) and the percentage time fixated (PTF) were investigated in response to checkerboard light flashes in 10 human infaed as a function of the size of check in the evoking stimulus (diffuse light, 11, 22, 45, 90 and 180 min of arc), the refractive lens strength the checkerboards were viewed through (-6 to +6 diopters), and the age of the infants (6-26 or 27-45 days). Check size significantly influenced VEP amplitude in infants as young as 6 days. The 11' checks evoked greater responses that diffuse light suggesting a visual acuity of better than 20/220. Only the 27--45-day-olds behaviorally discriminated the checks, PTF  indicating an acuity of 20/120. Evoked potential refraction with spherical lention between VEP amplitude and check size measured from different VEP components at different ages indicated the function contained two modes or components. The first mode was inverted "U-shaped" and was obtained in response to check sizes less than 45'. It was primarily due to changes in amplitude of the early VEP components (less than 210 msec after the evoking stimulus) and was poorly correlated with the behavioral PTF measure. It was proposed that this mode reflected subcortical activity. The second mode was a linear increase in amplitude as check size was increased from 45' to 180'. It was primarily due to changes in the amplitude of late VEP components (240--400 msec after the evoking stimulus) and was highly correlated with the percentage time the infants fixated the various check sizes. It was proposed that this mode reflected cortical activity. Age selectively influenced the late VEP components and the PTF behavioral measure, these measures being influenced by check size only in the 27--45-day-old infants. This change in responsivity of late VEP components and the transition from passive to more active and discriminating visual preference, suggest the onset of increased cortical function between 28 and 45 days of age.


Spike-wave rhythms in cat cortex induced by parenteral penicillin. I. Electroencephalographic features. Surface and depth recordings were made in 21 cats with generalized, parenteral penicillin induced epileptiform activity often assuming spike-wave forms, to obtain information on the structural substrate of "spontaneous" spike-wave rhythms. Recordings were made from neocortex, medial and lateral thalamus, hippocampus and brainstem reticular formation. Epileptiform activity first appeared in cortex and subsequently projected to depth structures. Occasionally, focal discharges could be seen in subcortical structures, but these generally did not spread to cortex. No consistent "pacemaker" was identified in cortex or depth. Bilateral applications of penicillin to cortex produced synchronous spike-wave bursts. In contrast, ventriculocisternal perfusion of penicillin solutions yielded synchronous cortical potentials which differed clearly in morphology and frequency from parenterally induced spike-wave bursts. During generalized activity, transcortical, intrahemispheric lesions interfered with ipsilateral synchrony. After transcortical section, projection of localized cortical bursts to thalamus did not necessarily result in "recruitment" of diffuse cortical regions into epileptiform activity. These results emphasize the importance of cortical circuitry and corticocortical connections in the genesis and synchronization of spike-wave rhythms.


Spike-wave rhythms in cat cortex induced by parenteral penicillin. II. Cellular features. Epileptiform potentials, consisting of spontaneous, generalized bursts frequently assuming a 3/sec spike-wave form and tonic clonic electrographic seizures were produced in 32 lightly anesthetized cats by parenteral injections of penicillin. The activity of 83 identified pyramidal tract cells and 207 cortical non-pyramidal tract cells was correlated with the surface EEG. The majority of both cell types generated depolarizations and action potentials with the EEG spike. Hyperpolarizations, during which cells were inhibited, followed the depolarizations. The depolarizations responded to injected current as if they were generated by excitatory synapses; and hyperpolarizations to injected current and chloride ions as if generated by proximal inhibitory synapses. Attempts to identify a class of neurons firing during the surface-negative wave (presumed inhibitory interneurons) were unsuccessful. Forty-two units were recorded during tonic-clonic seizures. Intracellular records disclosed tonic oscillations of membrane potential, phased bursting with "depolarization shifts", abortive action potentials and post-ictal hyperpolarizations. Cell somata often depolarized to the point of inactivation, but axons continued to fire at high rates. These results emphasize the role of EPSP-IPSP sequences in the generation of spike-wave rhythms.


Pathophysiology of generalized penicillin epilepsy in the cat: the role of cortical and subcortical structures. I. Systemic application of penicillin. The mechanism of precipitation of generalized epileptiform discharges in feline generalized penicillin epilepsy, a model of human generalized corticoreticular ('centrencephalic') epilepsy, was studied in acute and chronic experiments in cats with implanted skull and intracerebral electrodes. Single shock and low frequency repetitive stimulation of subcortical sites from which prior to penicillin administration spindle activity and recruiting responses could be elicited, readily triggered epileptiform discharges in the same animals after penicillin. These structures comprised the intralaminar and midline thalamic nuclei, the neostriatum, and some posterior thalamic association nuclei (Pulvinar and nucleus lateralis posterior). Subcortical and cortical structures which prior to penicillin elicited neither spindle activity nor recruiting responses were significantly less effective in  triggering generalized epileptic bursts after penicillin injection. The probability with which such bursts were elicited from these structures was still, however, in many instances above chance level. It is concluded that the generalized epileptiform discharges in feline generalized penicillin epilepsy can be triggered from a large number of brain sites, but most reliably so from subcortical nuclei involved in spindle generation and recruiting responses. The experimental evidence presented still does not allow one to determine whether epileptic alteration of neuronal function in this form of epilepsy primarily resides in cortical or subcortical nerve cells or in both.


Sources of frequency following responses (FFR) in man. In order to study the sources and pathways which are responsible for the frequency following response (FFR), records were made in control subjects and in patients with special types of lesion and response. It has already been shown that the FFR in normal subjects to tone bursts with single onset phases is made up of a short latency cochlear microphonic potential (CM) and a longer latency neural component (neural FFR). No neural FFR could be recorded in patients with upper brain-stem lesions (absence of click-evoked responses from the inferior colliculus along with clinical signs of such a lesion). Their FFR was exclusively a cochlear microphonic potential, thus demonstrating that the neural FFR with a latency of 6 msec is generated in the region of the inferior colliculus. Also in subjects with large post-auricular muscle (PAM) responses, the PAM can contribute to the FFR, with a latency of 10 msec. In patients with high-tone hearing loss due to acoustic trauma, no CM could be recorded while a neural FFR with a latency of 6 msec was present. This indicates that the CM recorded by this technique may be generated in the basal turn. It also demonstrates that the pathway of the neural FFR begins in the apical turn of the cochlea.


Electroencephalographic changes in the dominant hemisphere during memorizing and reasoning. The EEG was recorded with bipolar technique in ten normal subjects in the frontal, temporal and occipital regions of the dominant hemisphere in four situations: (1) during an auditory memory test, (2) during 'auditory rest' (listening to white noise), (3) during a visual reasoning test and (4) during 'visual rest' (watching a black dot on a white screen). Computer analysis of the EEG was made by (a) an on-line automatic EEG analyser yielding measures of mean power, mean frequency and frequency spread (complexity) and (b) an off-line period-amplitude (P-A) analysis, which gave per cent activity time and mean voltage in 21 frequency bands. As compared to auditory rest the auditory memory test gave an amplitude increase frontally in the alpha, theta, and delta bands. During the visual reasoning test there was in addition an amplitude increase in the alpha band in the temporal region. During the auditory test a tendency to a decrease of the alpha activity was seen in all three regions but this decrease occurred only occipitally during the visual test. The two types of mental activity thus induced two patterns of regional EEG changes. These showed principal similarities to regional cerebral blood flow patterns which have been recorded during visual and auditory tests of about the same types as those used in the present study.


The action of lithium carbonate on the sleep-waking cycle in the cat. The therapeutic effect of lithium carbonate on manic-depressive psychosis is now universally recognized. In view of this positive action on the cyclic endogenous manifestations, it was interesting to investigate the possible effects of lithium on the sleep-waking cycle in the cat. Polygraphic recording from three adults cats was carried out during 24 h periods before, and on different days after, beginning treatment with lithium carbonate. With low doses (30 and 50 mg/kg/day), important morphological changes were observed 5 days later while a new balance of the sleep-waking stages occurred. The EEG of each stage was characterized by slowing and amplitude increase of the different frequencies. The rhythms which appeared in long runs during quiet wakefulness and paradoxical sleep (PS) in the somaethetic cortex (mu rhythm) and the visual areas (alpha rhythm) were increased, slowed and almost continuously present. From the quantitative point of view, the percentage of time of deep slow wave sleep (DSWS) was increased from 38% to 55%. Conversely, the waking and PS times were both reduced, the latter from 18% to 10%. In contrast with human data, the mean duration of PS episodes was unchanged. Furthermore, lithium induced a slight dissociation between EEG activity and waking behaviour. With toxic dose (90 mg/kg/day) all the above changes were again observed, but more conspicuously. During wakefulness and SWS, bursts of generalized paroxysmal events appeared, in frequent association with a myoclonic jerk. SP became atypical and its percentage of time was drastically reduced.


Regional cerebral blood flow in children. A rheoencephalographic study of the modifications induced by reading. Cerebral impedance changes (F changes) indicating cerebral blood flow were studied in 17 children aged 6-11 years and in 10 normal adults. At rest, large values were found in children, who never exhibited the 'hyperfrontality' pattern of the adults. Reading induced an F increase in both the dominant and non-dominant hemispheres, in contrast with a larger increase in the dominant hemisphere in adults. A spatial test was designed in adults which triggered a larger increase in the non-dominant hemisphere.


Reversible profound depression of cerebral electrical activity in hyperthermia. Transient major reduction of EEG activity in an hyperpyrexic patient (rectal temperature 42.5 degrees C) and transient isoelectric ECoG during accidental hyperthermia (rectal temperature 41.8 degrees C) in a Rhesus monkey are reported. Since recovery of electrocortical activity occurred in both instance this implies that in hyperthermia, as well as in hypothermia, an isoelectric EEG may not indicate irreversible brain damage.


Effects of selective attention on the late components of evoked potentials in man. Vertex and specific evoked potentials to visual and cutaneous stimuli were recorded in 6 subjects. The delivery of roughly equal numbers of flashes and shocks was paced by the subjects themselves. They also had to choose which stimulus they would be given next; 9 times out of 10 the stimulus matched the subject's expectation and stimuli from the unexpected modality were given randomly 1 time out of 10 on the average. No task was required after the stimulus occurrence. Cortical evoked potentials (CEPs) to unexpected stimuli were averaged and compared with CEPs the major difference consisted in large N2 and P3 waves in response to unexpected stimuli; no difference was observed in the amplitude of the N1 wave. In specific CEPs unexpected stimuli also elicited N2 and P3 waves; in addition N1 showed a slight tendency to be higher in CEPs to unexpected stimuli,


VEP and AEP variability: interlaboratory vs. intralaboratory and intersession vs. intrasession variability. The VEP and AEP of a normal adult were recorded at single sessions in five laboratories that used different instrumentation. The results were compared with VEP and AEP data recorded at eight weekly testing sessions at one of the labs. The interlaboratory standard deviation was found to be over twice as large as the intralaboratory standard deviation measured over an eight week period. Intrasession EP variability was significantly less than intersession variability.


Sequential discharges of phasic waves in the pontine oculomotor system during paradoxical sleep in chronic mesencephalic and decorticate cats. The temporal organization of sequences of phasic activity (PGO) in the pontine oculomotor system during paradoxical sleep was compared in normal cats, in "mesencephalic" preparations, and in completely decorticated animals. After both types of operation, PGO discharges lost their random character, becoming grouped in bursts separated by long intervals. These observations suggest that the cerebral cortex is involved in the regulation of patterns of PGO sequences.


General-purpose timer from transistor logic to auxiliary equipment. This paper describes a low-cost circuit for the control and timing of auxiliary experimental equipment by a variety of signals from 5 V, integrated-circuit logic. The circuit is designed for variable timing and switching up to 240 AC V at 3 A, with two independent timing parameters, delay and duration, in response to positive-going or to negative-going logic signal transitions.


Identification of beta-cells in dissociated rat pancreatic cell suspensions. beta-Cells may be demonstrated in pancreas sections as well as in monolayer cultures. Until now, however, it has been impossible to differentiate quickly among individual cell types in freshly dissociated cell suspensions prepared for pancreatic monolayer cultures. Rapid identification of endocrine cells within the total cell population can be achieved by means of vital staining with neutral red. Moreover, the direct observation of unfixed and unstained cell suspensions under dark-field illumination allows an immediate identification of granulated beta-cells.


Comparison of diagnostic accuracy of different peripheric parameters of thyroid function. Diagnostic value of a scope of peripheric parameters of thyroid function was assessed in an unselected group of untreated patients with suspected thyroid disorder, further in untraeted selected patients (without cardiac involvement) and in treated patients. Comparison of relative values  of individual tests was performed, based on relation to plasma thyroid hormone level represented by PBI. It was found that: 1. The diagnostic value of heart rate, plasma cholesterol level, B. M. R. and Hegglin's sign (T-2s interval) is of a very limited degree. 2. The diagnostic accuracy of AJT, Q-Kd interval and PEP was found to be of considerable interest even in unselected patients. Values of IRVD and D indices found for these tests are comparable and allow the immediate estimation of thyroid function in bedside diagnosis. 3. The diagnostic value of PEP could be enhanced by exclusion of patients with suspected or proved cardiac disorder or myocardial failure; this may be useful for physiologic studies. 4. While heart rate is profoundly and inconsistently influenced by beta-blockade, AJT is influenced to a minimal degree only and Q-Kd and PEP are uniformly shifted to higher values, allowing thus diagnostic evaluation during this form of treatment also. 5. As Q-Kd is considerably age-and height-dependent, AJT and PEP are believed to be the most suitable test for immediated clinical diagnosis.


An investigation of mistranslation in vivo induced by streptomycin by an examination of the susceptibility of abnormal proteins to degradation. Proteolysis rates in vivo were measured in Escherichia coli cultures during treatment with dihydrostreptomycin and under various other conditions. Dihydrostreptomycin treatment caused an increase in the proteolysis rate, compared to untreated controls. The proteolytic system in vivo responsible for the elevated proteolysis in the early stages of dihydrostreptomycin treatment, or that during canavanine and puromycin treatment, were not inhibited by addition of phenylmethanesulphonyl fluoride. This agent did inhibit proteolysis rates in cultures whose growth was inhibited by starvation, or had been completely stopped by dihydrostreptomycin. It seems, therefore, that the extremely high proteolysis rates in cultures at this stage of dihydrostreptomycin treatment were due to the action of two protease systems: the one concerned with the breakdown of abnormal proteins, and the other concerned with normal protein turnover and active during a non-specific decline of growth. The proteolytic rate at complete growth inhibition brought about by dihydrostreptomycin was intermediate between those induced by canavanine and puromycin at the same stage of treatment. This indicated a similar hierarchy in the extent and nature of abnormality in the proteins synthesized under these conditions. The relationship between the abnormality of proteins induced by dihydrostreptomycin and the importance of this in the antibiotic mechanism is discussed.


Aberrant CSF protein fractions found by electrofocusing in multiple sclerosis. A study of 26 cases with clinically verified or probable multiple sclerosis and 2 cases with optic neuritis. The CSF gamma-globulin patterns found by electrofocusing in multiple sclerosis can be classified into five (a-e) mean patterns, where the individual bands lie very close together. However, in some patients with verified or probable multiple sclerosis conspicuous bands are found. The electrofocusing patterns from 28 such cases were compared with those reported by Kjellin and Vesterberg and with 20 cases of verified multiple sclerosis collected by chance. Differences were found with regard to the frequency of the different mean patterns, the occurrence of a double fraction in region '5', a single fraction in region '7' and a highly alkaline fraction. The electrofocusing findings have been compared with the clinical data of the 28 patients. There seems to be some correlation between the patterns of conspicuous bands and the duration and course of the disorder as well as probable sites of the CNS lesions. No indication was found, that either a high CSF gamma-globulin value and/or in a high degree of barrier damage was a prerequisite for the occurrence of conspicuous bands in the CSF gamma-globulin region. The aberrant electrofocusing patterns might be genetically determined or caused by some unusual agents (possible viral) or autoimmune reactions, modified by other than gentically divergent factors.


Multiple sclerosis: a clinico-biological study of 100 cases in Spain. A study was carried out on 100 cases with the clinical diagnosis of multiple sclerosis. Several clinical and analytical factors were considered and correlated. Special attention was paid to data obtained from cerebrospinal fluid (CSF) electrophoresis. The degree of severity of the disease does not affect the selective evaluation frequency of gamma-globulin or its average values. A greater frequency of definite gamma-globulin increase is observed in patients with a stationary development than in those a progressive development although, in both instances, the average values of gamma-globulin do not show any significant difference. There is no correlation between the CSF data and the presumed clinical site of the lesion. Those patients with a single lesion have a lower incidence of pathological data in CSF than those with multiple lesions. A higher proportion of pathological results is seen in CSF of patients with a chronic evolution from the onset than in chronic-intermittent cases. The cases studied during a relapse show a marked decrease in the frequency of definite gamma-globulin increase and paretic colloidal curves. This group of patients gives lower average values of gamma-globulin, total protein and cells, than those patients examined out of relapse. These results are in agreement with recent immunopathological observations.


An immunofluorescent study of alpha-fetoprotein. II. Its significance in some nonhepatoma patients. alpha-Fetoprotein was investigated in nonhepatoma patients, whose definitive diagnoses had been confirmed at surgery. Its changes in serum level and localization in tissues were reported and the possible etiology and pathogenesis of these diseases were discussed with an emphasis on morphological observation.


Influence of physiological saline, dextran 70, hydroxyethyl starch, degraded gelatin, and fat emulsion solutions on screen filtration pressure. The effect of 0.9% NaCl solution, dextran 70, hydroxyethyl starch, degraded gelatin and fat emulsion on Hb, Hct, platelet count and screen filtration pressure has been studied in the rabbit. Dextran 70, hydroxyethyl starch and degraded gelatin caused hemodilution and decreased platelet count. Screen filtration pressure increased after infusion of degraded gelatin and tended to decrease after dextran 70 and hydroxyethyl starch. 0.9% NaCl and fat emulsion caused less pronounced changes. Intraportal infusion of 7 ml/kg body weight of the same solutions gave results which at present are difficult to interpret.


Colchicine inhibits stimulated release of gastric histamine but not activation of histidine decarboxylase. In the rat, gastric mucosal histamine is mobilized and histidine decarboxylase activated by treatment with insulin or pentagastrin. Colchicine pretreatment prevented the histamine release without preventing the enzyme activation. The results suggest a) that histamine release and histidine decarboxylase activation are independent events, and b) that microtubules are involved in the release of histamine.


Gamma globulins in the mice vaginal fluids: cyclic and experimental variations. The study of the gamma globulins in the vaginal fluids reveals that the lowest values were found in mice at estrus and in castrated mice 8 days after the estrogen treatment. We suggest that the variation of gamma globulins in the vaginal fluids is influenced both under physiological and experimental conditions by estrogens.


A simple system for inspection of microelectrophoretic patterns. An inexpensive device for inspection of electrophoretic patterns on microgels is described. A common slide frame is modified in order to hold inside microgels immersed in liquid, so that the gels may then be projected on paper screens. The electrophoretic patterns may be drawn on the basis of the gel images. Staff-meeting presentation of microelectrophoretic patterns seems to be another application of the system.


[Characteristics of the cardiotropic effect of gutimine]. Experiments proved gutimine capable of intensify the collateral coronary circulation, intraventricular pressure and the rate of its accretion (dp/dt), the excitability and permeability of the heart, as well as to raise the cardiac contractility in dogs during an acute period of myocardial ischemia. In rats with pituitrin-isadrine induced cardiopathy the drug drastically depressed the permeability of the histohematogenous barriers of the myocardium and lengthened somewhat the life span of the animals. Parallel with is rising doses gutimine depressed the excitability threshold of the frog's heart "in situ" in a test with a liminal current producing extrasystole.


[Effect of lanthanum sulfate on cardiac activity and blood supply]. In concentration of 1:50 000, 1:25 000 and 1:1000 lanthanum sulphate is shown to reduce and in that of 1:100 -- to increase the amount of perfusion fluid from the vessels of an isolated and arrested rabbit heart. In doses of 20--40 mg/kg introduced intravenously it tended to increase the coronary circulation rate, this being attended by an insignificant fall of the arterial pressure. In these doses the compound did not change significantly the heart rate, but prevented the development of pituitrin-induced cardiac disorders in rabbits.


HL-A antigens in pustular psoriasis.

HL-A antigens in pustular psoriasis. HL-A typing was performed on 97 patients with pustular psoriasis. HLA-B27 was found increased for the combined three subgroups: localized psoriasis of palms and soles, acrodermatitis continua and generalized pustular psoriasis, who were associated with a high incidence of arthritis. These subgroups have this in common with Reiter's disease indicating a link between the entities. In persistent palmo-plantar pustulosis an increased incidence of HLA-Bw35 was found. HLA-B13, HLA-B17 and HLA-Bw37 which are found markedly increased in psoriasis vulgaris were in acrodermatitis continua, generalized pustular psoriasis and persistent palmo-plantar pustulosis either absent or not increased as compared with the control population. 7 of 30 patients with localized psoriasis of palms and soles had one of these antigens. Our findings confirm that psoriasis vulgaris and pustular psoriasis as such, seem to be different aetiological entities. Some patients with localized psoriasis of palms and soles may be true psoriatics which besides their psoriasis have a tendency to develop a pustular reaction in their palms and soles similar to persistent palmo-plantar pustulosis.


The use of alpha fetoprotein in prenatal diagnosis. Maternal or blood levels of alpha fetal protein are elevated in the presence of certain fetal anomalies, particularly those of the neural tube in which protein from the cerebrospinal fluid may leak into the amniotic fluid. Since this is a nonspecific elevation, other diseases are potentially screened for by this determination. Alpha fetal protein levels should be measured on any amniotic fluid samples obtained during the middle trimester of pregnancy.


Hepatitis B surface antigenemia in patients with hepatocellular carcinoma in relation to clinical course and alpha-fetoprotein. Hepatitis B surface antigen was determined in sera of 122 cases of hepatocellular carcinoma seen in Japan, using both the counterimmunoelectrophoresis and radioimmunoassay (RIA) techniques. It was positive in 49.2% of the patients with RIA, but the level of antigen in serum was relatively low since positivity rate by counterimmunoelectrophoresis was only 10.7%, The degree of antigenemia as assessed from the count relative to the cut-off value in RIA, was increased during the clinical course in 75% of the patients. The antigen tended to rise in concentration when the tumor grew at a rapid rate, when damage to liver parenchyma was extensive, or in patients receiving chemotherapy. There was also a tendency for less frequent positive antigen tests in patients with higher alpha-fetoprotein levels. Illustrative cases are presented with discussion on the possible explanation for the change in the degree of antigenemia.


Whipple's disease with minimal intestinal involvement. An uncharacteristic case of Whipple's disease is reported, in which, although overt intestinal involvement was absent, and there was only a patchy histological lesion, the diagnosis was confirmed by electron-microscopic examination of peroral intestinal biopsies.


Feulgen staining of rat liver fixed in different fixatives. In the present study rat liver pieces fixed in 1) 10 per cent buffered neutral formalin, 2) 4 per cent glutaraldehyde, 3) Heidenhain's-Susa fixative and 4) Flemming's fluid, and following hydrolysis in 1-0 N HC1 at 60degreesC for varying time periods have been stained with the UV Feulgen procedure. The results of this study reveal that following hydrolysis for different time periods the tissue material fixed in formalin show the same staining pattern as those fixed in glutaraldehyde. The material fixed in Heidenhain's-Susa displays an intense Feulgen staining after two different times of hydrolysis, and that fixed in Flemming's fluid shows particular staining intensity for a prolonged time period thus indicating better preservation of DNA than in the materials fixed in the other three fixtatives.


Immunoperoxidase: a sensitive immunohistochemical technique as a "special stain" in the diagnostic pathology laboratory. The present communication is a brief review of the origins, theory, and applications of a relatively new immunohistochemical technique that can be performed on routinely fixed, paraffin embedded tissues in which visualization by bright field light microscopy is feasible and which thus can be readily adapted to routine diagnostic work. The chief concern of this presentation is the practical diagnostic application of the immunoperoxidase technique, a method whose reputation as a sensitive investigative tool is well established.


Immunopathology of skeletal muscle. The value of direct immunofluorescence in the diagnosis of connective tissue disease. Whitaker and Engel in 1972 first described immunoglobulin deposition in muscle biopsy specimens in connective tissue disorders. In order to confirm and extend their observations and in the hope of identifying features that may differentiate skeletal muscle involvement in connective tissue diseases from other neuromuscular diseases, a series of 80 muscle biopsy specimens were examined using direct immunofluorescence. Three distinctive direct immunofluorescence patterns consisting of vascular, sarcolemma-basement membrane, and fiber staining were identified in skeletal muscle biopsy specimens from 35 patients, 31 of whom had a connective tissue disease. A vascular staining pattern showing a granular deposition of immunoglobulin or complement was seen in 15 patients, all of whom had a connective tissue disease. Routine stains generally did not reveal vessel abnormalities. A sarcolemma-basement membrane staining pattern was demonstrated around the sarcoplasmic membrane in 29 cases. Twenty-six of these patients had a connective tissue disease. There was no correlation with inflammation, fiber necrosis, or degree of connective tissue replacement. Fibers staining for immunoglobulin or complement, seen in 14 cases, generally occurred in morphologically normal fibers. Thirteen of these patients had a connective tissue disease. Since the pathologic change in muscle in the collagen vascular diseases often consists of the nonspecific findings of focal fiber necrosis frequently without inflammatory infiltrates, direct immunofluorescence may be useful in the diagnosis and classification of muscle diseases in the collagen vascular disorders. Furthermore, the findings of immunoglobulin deposition either within vessels or within individual muscle fibers suggest that immunological mechanisms may be responsible for muscular abnormalities in the connective tissue diseases.


N-band polymorphism of human acrocentric chromosomes and its relevance to satellite association. With the aid of Q- and N-banding techniques we investigated the relationship between the length of satellite stalks, the appearance of N-bands and the frequency of satellite association of individual acrocentric chromosomes in the cells of seven individuals, including one male with a satellited and small Y-chromosomes. The appearance of N-bands, seemed to be a constant and characteristic property of individual acrocentric chromosomes, independent of the status of concentration of the chromosomes at metaphase. The homolog with longer satellite stalks had larger N-bands and participated in satellite association at a higher frequency than the one with shorter stalks. It appeared that N-bands were present along the whole length of the satellite stalk, the size of which could possibly reflect the amount of rDNA present in the nucleolar organizers in human chromosomes.


Banding and spiralization of human metaphase chromosomes. A new technique is described which produces spiralization of human metaphase chromosomes. The important feature is heat followed by trypsin treatment. By varying conditions, it is possible to produce bands, spirals and intermediate stages. This provides a new approach to the understanding of banding and chromosome structure.


DIPI and DAPI: fluorescence banding with only negliglible fading. DIPI and DAPI produce distinct fluorescent bands in human chromosomes similar to quinacrine banding patterns. Additionally, the AT rich secondary constrictions in the chromosomes Nos. 1, 9 and 16 are brightly fluorescent. On the other hand the brilliantly fluorescent regions after staining with quinacrine mustard in the chromosomes Nos. 3 and 4, satellites and some other regions in the acrocentric chromosomes are less striking. The distal part of the Y, however, is clearly discernible. Thus DIPI and DAPI seem to be strictly AT specific fluorochromes like Hoechst 33258. In interphase nuclei the Y chromosome can be identified. However, quinacrines are superior for Y-body analysis in buccal, hair cell and sperm smears. BrdU labeled chromatids show reduced fluorescence intensity. The difference, however, is less apparent than after staining with Hoechst 33 258. DAPI and especially DIPI are highly resistant to UV-irradiation; there is almost no fading within 30 min when using DIPI. Moreover, fluorescence intensity is stronger than in quinicrines. When photographing, exposure times may be reduced to about one quarter compared to quinacrine mustard.


Fluorescent C bands of human chromosomes with 33 258 Hoechst stain. Air-dried preparations of human metaphase chromosomes normally exhibit a Q band fluorescence pattern with 33 258 Hoechst stain while the C band regions of 1,9,16, and most acrocentric short arms appear dull. If these stained and mounted slides are stored at room temperature in the dark for several days, a spontaneous change from C-negative to C-positive band some times occurs. We postulate that the pH of the buffered saline mounting medium during storage of the slides causes the C band shift since the results can be duplicated experimentally by lowering the pH of the mounting buffer from 5.5 to 4.0.


Mithramycin and DIPI: a pair of fluorochromes specific for GC-and AT-rich DNA respectively. The AT specificity of the fluorochromes DIPI and DAPI and the GC specificity of mithramycin are evidenced by observations in human, mouse, and bovine chromosomes. DIPI and DAPI produce a pattern similar to Hoechst 33258 in all three species, whereas mithramycin results in a reverse pattern. The AT-rich centromeric heterochromatin in mouse is brilliantly stained by DIPI or DAPI and remains nearly invisible after mithramycin staining. In the GC-rich centromeric heterochromatin of cattle the opposite behavior is observed.