The ultrastructure of formed white blood elements (neutrophils) in schizophrenia. A comparative electronmicroscopic study of neutrophils in schizophrenic patients and in healthy control subjects has revealed a certain difference in the ultrastructure of blood cell formation. This difference concerns the coarse-grained chromatin, the osmiophility of nuclear membrane, the enlargement of perinuclear halo, the enlightenment of matrix, the enlargement of glycogen granules, the swelling of mitochondria, the change in the amount of free ribosomes, an increase in the number of azurophilic granules and a decrease in the specific ones.


[The acid-base equilibrium during mouse lymphoblastic leukemia]. In transplantable and in spontaneous lymphoblastic mouse leukemia blood pH, PCO2, TCO2 and BB were examined. Spontaneous Gross-leukemia in AKR mice was found to develop in its final phase respiratory acidosis. Transplantable (TAL) leukemia of AKR mice presents from the onset a tendency toward respiratory acidosis. "L-1210"-leukemia, on the contrary, was shown to alcalise the recipients during the first 8 days after inoculation, later on it can bring about an acidosis. The pH-deviations in "L-1210"-leukemia display a respiratory character in syngeneic DBA/2J recipients, whereas in semiallogeneic recipients the observed changes are metabolic or mixt in nature. This finding strongly argues for the importance of histocompatibility. Intranodally inoculated animals distinctly differ in their parameters in comparison to intravenously and intraperitoneally recipients, hence, an important role of the tissular-milieu which is put first in contact with the leukemic factor must be concluded.


[The cerebral sensitivity to "meningosis-prophylaxis" with 198Au radiogold according to EEG findings]. 35 children (16 girls and 19 boys) at the age of 1 11/12 to 16 11/12 with acute leukaemia were injected intrathecally with 198Au-radiogold colloids (HOECHST-BEHRING) for "prophylaxis of meningosis". The colloid size of the isotope amounted to 5 or 30 nm, the applied activity lay between 1.4 and 3.12 mCi. According to a dosage estimation made with the help of LOEWINGERS formula 1 mCi of radiogold corresponds to approximately 1200 rad. Clinical observations, such as headaches, vomiting or fever up to 39 degrees C, could only be found in 6 children (17.1%) during the first 24 hours. All symptoms subsided quickly and without any sequels. Even retarded complications could not be detected. An electroencephalogram was made from all children before and after applying radiogold (1-8 d afterwards). After the injection of radiogold the majority of children had no change of findings in the electroencephalogram, 11 children even showed a tendency towards an improvement up to normalisation. Only 4 children had a deterioration of findings with unspecific disorders or appearances suspected of peak potential discharges. Simultaneously an accumulation of clinical complaints could be found. Judging from the clinical and electroencephalographic behaviour of our patients no absolute neurotoxity of radiogold could be ensured.


[The occurrence of Heinz bodies in the erythrocytes of persons exposed to low dose radiation during work]. 274 persons were examined by the provocation test of Heinz's bodies according to BEUTLER. 168 of them were working under x-rays exposure. No correlation could be identified between the dosage and the presence of Heinz's bodies in the lower range of the dosage related to the annual x-rays load and the integrated total exposure.


[The question of bone marrow cell proliferation in Di-Guglielmo's syndrome]. In a patient with a di Guglielmo's syndrome DNA was determined cytophometrically in the bone-marrow cells. The results show that the proliferation of paraerythroblasts is increased in the phase of erythremia in comparison to normal erythropoiesis. The proliferation of myeloblasts during the stage of myeloblastomatosis, however, is low similar to the majority of acute leukaemias. The examinations confirm the view that the di Guglielmo's syndrome represents a form of acute leukaemia.


Changes in the blood parameters of an air-breathing fish during different respiratory conditions. Some of the blood parameters recorded in an air-breathing eel, Amphipnous cuchia under normal respiratory condition during non-breeding period (September-April) are haemoglobin (Hb) concentration 19.26%, haematocrit value 56.16%, RBC number 1.71 million/mm3, RBC size 18.86 X 9.70 mum, mean corpuscular haemoglobin (MCH) 113.4 ng, mean corpuscular haemoglobin concentration (MCHC) 34.2%, blood sugar 77 mg% and ascorbic acid 0.435 mg%. The higher concentration of haemoglobin (19.26%) appears to be related to its obligatory air breathing habit and habitat in a water of low oxygen content. Though a definite trend of increase in the haemoglobin and haematocrit concentration with an increase in the body weight of the fish was lacking, variations were clearly marked related to intrinsic activity of the fish connected with different respiratory conditions. Asphyxiation in a submerged but continuous flow of water (liter/h) for 5 1/2 h resulted in an increase in the above-mentioned parameters to an appreciable extent. These increases were 0.23 million/mm3 in the number of erythrocytes, 6.16% in haemoglobin concentration, 10% in haematocrit value, 20% in blood sugar and 35% in ascorbic acid content. The mean corpuscular haemoglobin showed a decline of 6.2%. Exclusive aerial breathing for 5 1/2 h also caused 7.4% increase in haemoglobin concentration, 9.4% in haematocrit value, 0.14 million/mm3 in RBC number, 20% in blood sugar level, 9% in ascorbic acid content but almost no change in mean corpuscular haemoglobin. The average surface area for diffusion of gases appeared to have reduced by 6.8 mum2 per RBC.


[Studies on blood morphology in the European eel (Anguilla anguilla). I. Erythrocytes and their developmental stages]. By means of heart tapping we produced a larger loss of blood in the European eel, Anguilla anguilla. This resulted in enriching the peripheric blood with erythroblasts and proerythrocytes. By panoptic staining erythroblasts, proerythrocytes and mature erythrocytes could be prepared and described. By the aid of supravital staining it was possible to detect mitochondria and Substantia granulofilamentosa in all stages of development. Phase contrast observations proved that the erythrocytes of Anguilla anguilla do not possess any locomotion ability. Cytochemical investigations showed that the erythrocytes weakly or very weakly react to acid phosphatases, lipids and ribonucleic acids and positively to unspecific esterases. The artificially produced loss of blood and the following continuous extraction of blood gave indications of how rapidly the erythrocytes form and how much time their maturing takes. 48 hours after tapping erythroblasts appeared in large amounts. The number of proerythrocytes increased 24 hours later. The transformation of proerythrocytes into mature erythrocytes does not take more than 48 hours. The loss of erythrocytes caused by heart tapping is already compensated six days later by new growth. During this investigation division processes in erythrocytes were observed. These are endomitotic or amitotic division or mitotic ones which are very rarely observed.


[The "free" erythrocytes in newborn infants]. The percentage of non-A and non-AHP erythrocytes in newborns is usually increased and presents individual differences. It does not depend on the A1/A2 subgroups. These individual differences also concern the relationship between the number of non-A erythrocytes and of non-AH P-RBC. After delivery the A and AHP agglutinogens will individually develop in a different manner and independently one from the other.


[The demonstration of HL-A antigens on thrombocytes under various conditions]. Platelets of healthy test persons were gained according to the method of Aster (avital cells) and according to the procedure of BUBE and GMURZYNSKI (vital cells). Because of their tendency towards aggregation vital cells do bind any HL-A antibodies and whereas avital cells will fix antibodies and complement depending on the height of the titre. Thrombocytes stored in the own serum will respond less to the micro-C'-fixation test at + 4 degrees C as well as--196 degrees C, whereas storage in NaN3/NaCl and under the conditions of the own serum will reveal no differences up to--196 degrees C in each case.


The influence of azathioprine on the behaviour of blood platelets in vitro. The influence of azathioprine on human platelets was studied in vitro. Azathioprine in a final concentration of 10(-4) M caused significant inhibition of spontaneous sedimentation and aggregation of platelets, but did not effect platelet adhesiveness or resistance to freezing and thawing. The influence of azathioprine is probably due to thioimidazole, this last being a potent stimulating agent of phosphodiesterase activity, involved in the metabolism of cyclic AMP to 5' AMP. These observations indicate that the widely used platelet function assays could be influenced by drugs. The current therapy should therefore be taken into account when the results of the tests are evaluated for clinical purposes.


[The reactivity of the anticoagulatory system in sympathectomized animals under stress conditions due to acoustic trauma]. In rats of kind of Kruschinskij-Molodkina the number of cells in the stellate ganglion was diminished to 0.5% of the norm after eliminating the sympathetic tonus by means of guanethidine. These animals died of cardiac thrombosis during stress situations. This thrombosis cannot be explained by adrenalin, corticosteroids or thromboplastin spreading in the blood stream. A factor, the nature of which has still to be explained, may be assumed to be responsible for thrombosis to develop during stress situations in animals whose sympathetic tonus has been eliminated by guanethidine.


Failure of tranexamic acid to influence the ellagic acid-induced hypercoagulable state. Tranexamic acid in a dose of 50 mg/kg b.w. was unable to alter the ellagic acid induced hypercoagulable state. No change in the hypercoagulability pattern was observed regardless of the time of administration of the compound (before or after the ellagic infusion). The silicone clotting times after the infusion of ellagic acid were markedly shortened and remained so for about 60 minutes. The results observed in two control groups treated with saline were similar. The euglobulin lysis times were clearly prolonged after the administration of tranexamic acid, whereas no changes were observed after the administration of saline. These results indicate that tranexamic acid has no anti-factor XII activity.


Relations between fibrinogen degradation products and heparinocytes. In the emergency reaction there is a short-time increase of the values of basophilic leukocytes in connection with normal values of fibrinogen degradation products (FDP). After diminuation of basophils FDP are to be detected. In the chronical ill and under different hormonal contraceptives these two processes are overlapped disturbing a unique negative or positive correlation of heparinocytes and FDP. Concerning the situation under standardized conditions or in single cases worthful conclusions are possible about compensation or decompensation of a latent disseminated intravascular coagulation.


Suppression of photo-induced sporulation in Trichoderma viride by inhibitors. The mycelium of Trichoderma viride grown in the dark under submerged conditions and transferred to membrane filters sporulated only after photoinduction. The optimum photoinduction of sporulation was reached when applying daylight for 3 min and near ultraviolet radiation (355 nm) for 10 to 30 sec. After the photoinduction probounced synthesis of DNA, RNA and protein was observed. The photoinduced sporulation was partially or fully inhibited in the presence of phenethyl alcohol, actinomycin D, 5-fluorouracil, cycloheximide and ethidium bromide. The same inhibitors blocked also the photoinduced sporulation of surface growing colonies of Trichoderma viride. Various inhibitiors of synthesis of nucleic acids and protein, inhibitors impairing the function of membranes and certain other compounds were also effective.


Determination of electrolytes in the myocardium as a tool for the post-mortal diagnosis of recent infarction. The K/Na quotient in the myocardium has been compared to the result of the PTAH staining method in an autopsy material of verified myocardial infarctions, suspected but macroscopically unverified infarctions and controls. The control cases regularly had quotion showed ischemic changes in PTAH staining. These and additionally 10 cases had K/Na quotients below 1.2. The electrolyte method thus appears to be a more sensitive method for the diagnosis of recent myocardial infarction.


Experiences with the hematoxylin basic fuchsin picric acid staining method for morphologic diagnosis of myocardial ischemia - an experimental study in forensic pathology. An investigation was performed on 148 medicolegal autopsy cases with the purpose of obtaining experience with the hematoxylin basic fuchsin acid staining method for morphologic diagnosis of early myocardial ischemia. A comparative study was performed on rats with induced myocardial infarcts. The uptake of the basic fuchsin stain in myocardial sections agreed well with eosinophilia, often occurred when myocardial infarction was suspected, but very often yielded false positive and negative results. This lack of reliability probably depended on the high sensitivity of the staining procedure, degree of autolysis, fixation time, thickness of the sections and mainpulative lesions. Although the HBFP-technique does not seem sufficiently reliable in medicolegal autopsy cases it probably produces accurate results under controlled experimental conditions.


Practical approach to the diagnosis of sudden unexpected death of cardiac origin. Experiences concerning the practical demonstration of recent myocardial lesion (infarction) with various conventional and enzyme-histochemical methods are explained. It has been found in our laboratory that besides careful inspection of the heart, additional useful information can be obtained with ordinary H-E staining and beta-OH butyrate dehydrogenase reaction on frozen sections. Myocardial cells are darkly eosinophilic in the areas of infarction. Uneven staining in the dehydrogenase reactions was regarded as a sign of lesion in that section. beta-OH butyrate dehydrogenases revealed the damage more clearly than succinate and malate dehydrogenase. The enzyme reactions were usable as late as 7 days after death if decomposition had not commenced.


[The effect of bencyclane hydrogen fumarate (Fludilate) on the adhesion of tumor cells in vivo and in vitro]. Bencyclane hydrogen fumarate (Fludilat) was tested on the stickiness of tumor cells in vivo and in vitro. It was intended to determine whether Fludilat reduced the cancer cell stickiness in vitro, and if the survival time of cancer cell carrying animals can be increased with Fludilat in vivo, or in combination with a cytostatic. For the in vitro trials, concentrations from 0.001 mg/ml to 1 mg/ml medium were chosen. The survival trial on NMRI-mice with Nemeth-Kellner lymphosarcoma was performed in three groups, each with 4-5 sub-groups: Control group--Fludilat 5 mg, 10 mg, 20 mg/kg bodyweight, Bleomycin--50 mg/kg bodyweight, 100 mg/kg bodyweight, 250 mg/kg bodyweight, Bleomycin 50 mg/kg bodyweight + Fludilat 5 mg/kg bodyweight, Bleomycin 100 mg/kg + Fludilat 10 mg/kg bodyweight, Bleomycin 250 mg/kg + Fludilat 20 mg/kg bodyweight. The sequence of deaths was determined, and the 50% survival time was taken as criterium for the effect of the treatment. The in vitro trials showed a complete removal of the monolayer of the tumor cells from the bottom of the culture flask, in doses of 0.01-1 mg/ml medium. In the in vivo trial an increase in the 50% survival time could be achieved in all groups. The results of combined therapy of Fludilat and Bleomycin were striking. In comparison to the control animals, the treated animals showed that the occurrence of solid abdominal metastases from the Nemeth-Kellner lymphosarcoma could be almost completely prevented, especially at high doses. The Ca++-antagonistic effect, in changing the surface of the cells, is discussed as a mechanism of action.


Effect of bursa Fabricius extracts on antibody production in bursectomized or bursal cell autografted chickens. Restoration and enhancement of immune response against BSA antigen was achieved by 5-day consecutive doses of BF estract from 4-5-week-old chickens, in birds which had been surgically bursectomized or given BF-cell autografts at 17 days of age. A similar 5-day treatment with other tissue extract, i.e. liver, spleen, pancreas or intestine, or with LPS of E. coli, in contrast, failed to provide such restoration or enhancement.


Long-term antibody synthesis in vitro- IV. Independent segregation of antibodies directed to different determinants of an antigen molecule in its native configuration. Independent segregation of antibody populations directed to different portions of E. coli beta-d-galactosidase occurs during the immune response against the enzyme. Anti-enzyme antibodies able to interact and activate a naturally occurring ligand, the mutant-defective enzyme AMEF (Antibody Mediated Enzyme Factor), do not parallel anti-enzyme antibodies which are measured by a coprecipitation assay involving precipitation of the wild-type molecule. Dissociation of the two antibody populations is best achieved in microcultures sustaining long-lasting responses. Similarly, anti-NIP (4-hydroxy-3-iodo-5-nitrophenylacetic acid) antibodies could be elicited without concomitant synthesis of anti-carrier antibodies by short-term challenge in vitro of ovalbumin-NIP-primed lymph nodes with a heterologous conjugate in which the hapten NIP was coupled to a carrier known to be non-immunogenic under the conditions of challenge. The potential applications of these findings are indicated, namely: large-scale production of monospecific antibodies in vitro; and the possibility of studying the regulatory role of antibodies directed towards on portion of the immunogenic molecule on the response to other regions of the same molecule.


Rat parietal yolk sac basement membrane. An investigation of the antigenic determinants using a radioimmunoassay. Previous studies have shown that there is microscopic and biochemical evidence that rat parietal yolk sac synthesizes basement membrane (type IV) collagen; this study shows that a radioimmunoassay may be used for the detection of type IV collagen in such biosynthetic systems. Rat parietal yolk sacs incubated in medium containing (14C) proline either with or without alphaalpha-dipyridyl produced either unhydroxylated or hydroxylated (14C)collagen. The immunological reactivity of these two preparations was investigated using antibodies to bovine type IV collagen in a  radioimmunoassay which demonstrated that the hydroxylated (14C)collagen preparation had a considerably higher level of antigenicity than the unhydroxylated (14C)collagen. Hydroxylated rat type IV (14C)collagen which had been reduced and alkylated was intermediate in antigenicity between hydroxylated and unhydroxylated material. These findings suggest that there are antigenic determinants which depend upon hydroxylation of the collagen molecule, and others dependent upon intact disulphide bonds. In addition, various levels of pepsin extracted unlabelled calf anterior lens capsule collagen caused inhibition of antibody binding to (14C)collagen. Rat type IV (14C)collagen which had been digested with collagenase was inactive in the radioimmunoassay, while pepsin digestion caused no reduction in antigenicity. These findings suggest that the antiserum is directed towards the collagenous part of the molecule and may be a useful tool in the detection of biosynthesized basement membrane collagen.


Regulatory effect of temperature and antigen upon immunity in ectothermic vertebrates. I. Influence of hapten density on the immunological and serological properties of penicilloyl-carrier conjugates. Sodium penicillin was conjugated to sheep erythrocytes and optimal quantities, added to a 5% SRBC suspension, were determined for haemagglutination (12-5 mg/ml) and for haemolysis (50 mg/ml) using carp antibodies and carp complement. The epitope density on the BSA molecule was gradually increased, when increasing amounts of sodium-penicillin were added to a constant quantity of BSA, until a maximum of about thirty penicilloyl groups were bound. Low conjugates, having less than seven haptenic groups per one BSA molecule, were found to stimulate carp for both anti-hapten and anti-carrier antibodies. The higher conjugates having seven and more haptenic groups were found to stimulate carp for anti-panicilloyl antibodies but not for anti-BSA antibodies. A booster dose with native BSA, given to the Pen30 BSA preimmunized carp, gave rise directly to a secondary-like response. In the rabbits, however, both heavy and low conjugates were found to stimulate antibody production for the hapten as well as for the carrier. It was suggested that the modified BSA in the heavy conjugates loses its ability to stimulate B cells, probably due to a decrease in local concentration of antigenic determinants in the BSA molecule, but its ability to stimulate helper cells is not affected for this reason.


Distribution of ad and ay subtypes of hepatitis B surface antigen among hepatitis patients and symptomless carriers in Hungary. A total of 176 HBsAg positive sera from acute and chronic hepatitis patients and symptomless carriers (blood donors) in Hungary were subtyped for ad and ay determinants. The distribution of ad and ay determinants in Hungary was found to be about the same. There were no significant differences among the groups tested.


Specificity of serum from rodents immune to Moloney C-type virus-induced tumours. When cells are infected by C-type viruses such as Moloney sarcoma/leukaemia virus, new antigens appear on the cell membrane. Mice and rats will respond immunologically to the antigen(s). It was uncertain whether the antigens were related to the viral structural proteins or to non-virion, tumour-specific surface antigens (TSSA) or both. In an 125I-antiglobulin binding assay, Moloney virus completely blocked the binding of mouse and rat sera to virus shedding target cells, thus suggesting that mice and rats recognise only viral proteins. Mice responded to type-specific and rats mainly to group-specific determinants on the virus. Individual Moloney viral proteins were prepared using the guanidine HCl method and were used to block the binding of the rat anti-Moloney immune serum to Moloney virus shedding target cells. By this method, it was demonstrated that the rat serum contains specificities for the viral proteins gp70 and p30, but it was not possible to detect any antibodies directed towards non-virion or TSSA-like molecules.


Studies on Epstein-Barr virus-related antigens. I. Indirect single radial immunodiffusion as a useful method for detection and assay of soluble antigen. A useful method for the detection and assay of Epstein-Barr virus (EBV)-related soluble antigen has been developed by the application of the indirect single radial immunodiffusion technique which is frequently used for quantitative measurements of immunoglobulins and other soluble proteins. When the extracts of EBV-determined nuclear antigen (EBNA)-positive non-producer cells (Raji and NC-37) were applied to agar plates containing seropositive human serum, followed by overlay with anti-human IgG serum, ring-shaped precipitates with high specificity were clearly evident. The size of such precipitin rings was proportional to the amount of the antigen. This method is simple and applicable for a quantitative assay of a particular EBV-related soluble antigen and antibody and the sensitivity is equivalent to that seen with the complement fixation test.


In vitro stimulation of mouse lymphoid cells by C1300 neuroblastoma cells or tumor membrane extracts. Spleen lymphoid cells from A/J mice recognize specific antigenic differences on the surface membranes of syngeneic C1300 neuroblastoma cells and incorporate 3H-thymidine into DNA in unidirectional mixed cell cultures in the absence of isologous serum. The response requires an optimal ratio of responder to stimulator cells, and is detectable after 24 h. It is specifically blocked by the presence of a papain-solubilized crude membrane extract from the same neuroblastoma cells, the extent of inhibition being dependent on the concentration of the extract and the time when it is added to the cultures. Spleen cells from mice bearing the neuroblastoma respond earlier and incorporate more 3H-thymidine than cells from unsensitized mice. The enhanced response of the primed spleen cells to the stimulator cells is similar to a secondary immune response and can be induced by soluble crude tumor extracts in the absence of stimulator cells.


Interaction of bleomycin and radiation in combined treatment on mouse L cells. Using an established line of L cells, we studied survival properties and recovery after combined action of bleomycin and radiation. A moderate synergistic effect was observed when bleomycin and radiation were given simultaneously. Cell recovery from radiation damage was not affected by bleomycin. Results seem to be in agreement with previously published data.


A new pyrazolidine derivative - benetazone spofa - in short- and medium-term treatment of rheumatoid arthritis. (Double-blind comparative study with phenylbutazone). The effects of Benetazone Spofa (trimethazone) in a dose of 1000 mg/day with those of phenylbutazone in a dose of 600 mg day in patients with rheumatoid arthritis were tested in a short-term double-blind trial (3 weeks) and in a long-term double-blind trial (12 weeks). The short-term trial failed to disclose a significant superior effect of phenylbutazone, and the continued prolonged therapy showed, on the contrary, the higher effectiveness of the new derivative, but the difference was not statistically significant. The main advantage of Benetazone consisted of its lower toxicity, better tolerance and of the much lower tendency to produce fluid retention compared to phenylbutazone.


Double-blind controlled trial of the new preparation FZ. 560 in the symptomatic treatment of digestive disorders. A controlled double-blind clinical trial was carried out to compare the effects of a new combination FZ. 560 in the symptomatic treatment of digestive disorders including a wide variety of symptoms. The combination contained fentonium bromide, 10 mg, dehydrocholic acid, 25 mg, pancreatin 3FU, 50 mg, and lactulose, 200 mg; it was compared with fentonium bromide 10 mg, and the other components. The three treatment groups included 37 patients, treated for a maximum of 14 days. The severity of the painful and dyspeptic symptoms was recorded daily; the overall daily scores were analyzed using persistence curves. A 50% reduction of the initial overall scores was observed in all the 12 patients in the combination groups by the third treatment day, but not until the 13th day in the fentonium group, while a 50% reduction in symptom intensity was not achieved during the trial in 7.7% of the group treated with the other components. Statistical comparison with the Wilcoxon test showed in fact that the effects of FZ. 560 were significantly superior to those of the two control preparations which did not differ significantly. These results, therefore, demonstrate clearly the advantages of the new combination FZ. 560.


Immunologic identification of M. leprae. Immunofluorescence and complement fixation. A markedly improved immunofluorescent technic employing FITC conjugated IgG antibody prepared from lepromatous serum is described as a means of specific identification of M. leprae. An additional immunologic identification method for M. leprae is presented as a micro-complement fixation technic employing antigen rather than antibody dilution. Studies with these technics suggest that M. leprae specific antigen is probably a surface antigen and has as part of its mosaic a lecithin-phospholipid component. It is not unlikely that it is a protein-glyco-phospholipid with a polysaccharide component. These technics employed with nodular extract (NE) from lepromas, with 15 strains of mycobacteria, with M. leprae from human tissue as well as from previously reported in vitro culture, strongly reinforce the allegation that M. leprae is readily cultivated in vitro in the hyaluronic acid based medium LA-3, previously reported.


The use of non-deparaffinized tissue sections for staining leprosy bacilli. Reduced acid-fast staining of leprosy bacilli occurs during the dewaxing of paraffin sections by xylene and alcohols; the older and more decrepit bacilli being especially affected. By the use of non-deparaffinized sections, the leprosy bacilli which could not be stained with the usual carbol fuchsin are strongly stained. Moreover, non-deparafinized sections can be used for the periodic acid-carbol pararosanilin stain or methenamine silver stain for demonstrating mycobacteria.


Evidence for a role of N-acetylmuramyl-L-alanine amidase in septum separation in Escherichia coli. Septum formation and septum separation have been studied in a chain-forming mutant of Escherichia coli K-12 bearing the envA mutation and its parental strain. In comparison to the wild type, the mutant showed a sixfold reduction in the specific activity of the enzyme, N-acetylmuramyl-L-alanine amidase (EC 3.5.1.28), part of which was associated to the outer membrane. Genetic as well as physiological suppression of chain formation resulted in an increase in amidase activity. The addition of N-acetylmuramyl-L-alanyl-D-glutamyl-meso-diaminopimelic acid to growing wild-type cells and to cells bearing the envA mutation caused an inhibition of cell separation and an increased frequency of visible septa. The kinetics of septum formation and separation was followed in chains by the use of ampicillin and nalidixic acid. The latter drug inhibited initiation of new septa but allowed preformed ones to go to cell separation at a rate corresponding to that of steady-state growing cells. Ampicillin treatment, on the other hand, resulted in a more rapid decrease in the frequency of septa. The disparate effects of ampicillin and nalidixic acid were not explained by a difference in amidase activity but could be due to an inhibitory effect of ampicillin on a septal peptidoglycan fusing activity.


[Histologic data on the lingual glands in various anuran amphibians]. The anuran Amphibians have a well developed tongue which is either protractile or attached to the floor of the buccal cavity according to the species. The lingual glands of the papillary dorsal side consist of simple tubules. The stratified epithelium of the ventral and lateral sides of the tongue is composed of mucous and muco-serous cells inserted between the covering ciliated cells. Filiform pappillae and glands  have a simple epithelium with numerous outer ciliated cells and glandular cells. The latter eleborate a product rich in glycosaminoglycans, more or less acid according to the species, associated with proteins. The lingual gland secretion is composed of proteins; when present, glycosaminoglycans are less abundant and less acid than those produced by the outer glandular cells. The histochemical and ultrastructural characters of the lingual glands of Anurans are compared to those of Apoda and Caudata previously studied by the same techniques.


Inhibition of activated factor XII by antithrombin-heparin cofactor. The activation of Factor XII occurs via fragmentation of this zymogen into a diverse spectrum of enzymatically potent molecular species. To study the interaction of antithrombin-heparin cofactor and heparin with activated Factor XII, we have employed two forms of this enzyme with widely differing physical characteristics and biologic potencies. Antithrombin-heparin cofactor was found to be a progressive, time-dependent inhibitor of both forms. The addition of heparin dramatically accelerated the rates of these interactions. Furthermore, sodium dodecyl sulfate gel electrophoresis of reduced proteins has indicated that antithrombin-heparin cofactor functions by forming an undissociable complex with either species of the enzyme. This complex represents a 1:1 stoichiometric combination of activated Factor XII and inhibitor. In the presence of heparin, both species undergo virtually instantaneous complex formation with antithrombin-heparin cofactor without exhibiting alterations in dissociability or stoichiometry.


Binding of thyroid hormones and their analogues to thyroxine-binding globulin in human serum. The present study was undertaken to study the binding of several thyroid hormones and structurally related compounds to human serum thyroxine-binding alpha-globulin (TBG). The source of TBG was normal human serum diluted 1:100 in 0.035 M barbital buffer, pH 7.4. In the binding assays, 125I-thyroxine, unlabeled thyroxine, and diluted serum were incubated for 20 h at 37 degrees in Plexiglas equilibrium dialysis units. Two orders of binding sites were discerned: a high affinity, low capacity binding site with an affinity constant of approximately 2.5 X 10(9) M-1, and a low affinity, very high capacity binding site with an affinity constant of less than 10(6) M-1. Studies with purified TBG, serum deficient in TBG, and purified human serum albumin indicated that the high affinity site represented binding to TBG and the low affinity site represented binging to albumin. The ability of several groups of thyroid hormone analogues to bind to TBG was then investigated. As a result of these studies, the following structural features of thyroid hormones were found to be important for optimal binding activity: (a) the L-alanine side chain conformation, (b) the presence of a 4'-hydroxyl group, (c) the presence of two substituents in the inner and outer rings (positions 3, 5, 3', and 5'), and (d) the presence of either bromines or iodines in the inner ring and iodines in the outer ring. Of lesser importance was the presence of an oxygen atom in the ether position.


Studies on the structural localization of rabbit H chain allotypic determinants controlled by the a locus. Purification and immunological properties of an immunopeptide bearing a3 allotypic determinants. An immunopeptide bearing a3 allotypic determinant(s) was isolated from the gamma chain of an a3 homozygous rabbit (G222-2) immunized with type III pneumococcal vaccine. Immunocogical properties of peptides were studied using a radioimmunoassay that involved inhibition by these peptides of a reaction between 125I-labeled anti-a3 antibody and Sepharose-bound a3 immunoglobulin G (IgG). The gamma chain was isolated from IgG of restricted heterogeneity and then citraconylated and digested with trypsin. The tryptic digest (TD1) was passed through an anti-a3 immunoabsorbent column either directly or after an intermediate step of Sephadex G-75 chromatography. The bound peptides (T1) were eluted with 0.1 M acetic acid and further digested with trypsin. The digest (TD2) was again run on the anti-a3 immunoabsorbent column to purify the bound immunopeptide T2. In the radioimmunossay this immunopeptide was found to have major a3 determinant(s). Its molecular weight was found to be approximately 6,000, which decreased to about 3,000 after reduction and alkylation. These data, together with NH2- and COOH-terminal analyses and cysteine peptide mapping, demonstrated that T2 is composed of two polypeptide chains linked by a disulfide bond, one from the cysteine 22 region having lysine at the COOH terminus and the other from the cysteine 92 region arginine at the COOH terminus. The lysine peptide was separated from the arginine peptide and its NH2-terminal sequence was found to be Gly-Asx-Glx-Ser-Thr-Cys. Since the cysteine is at position 22, the lysine peptide starts at position 17. It has approximately 22 residues. The framework sequence from 17 to 20 is different from those reported so far. In addition, the heavy chain used in these studies has some other unusual features including a histidine, probably in the first hypervariable region. The presence of histidine in the first hypervariable region of rabbit heavy chain has not been reported previously. The other peptide which is about 30 amino acids in length and ends with arginine 94, probably includes positions 67, 70, 71, 84, and 85 that are believed to have substitutions correlating with a allotypes. In a hypothetical three-deminsional model of the Fv portion of rabbit anti-SIII antibody BS-5, residues 17 to 33 of the lysine peptide and 67 to 79 and 84 to 85 which may be present in the arginine peptide are fully exposed on the surface and are far removed from the antibody combining site.


Flow cytofluorometric analysis of cell cycle distributions using propidium iodide. Properties of the method and mathematical analysis of the data. In order to better characterize the new rapid staining method for flow cytofluorometry proposed by Krishan, we have tested its stability and several other properties, and have carried out a quantitative comparison of the fluorescence histograms obtained using propidium iodide or the acriflavine-Feulgen staining procedure. Using a human hematopoietic cell line in the logarithmic phase of growth, and analyzing the data by means of a mathematical method we have devised, we found that the fluorescence intentsity of cells stained with propidium iodide remains stable for at least 48 h; it is insensitive to dye concentration between 0.025 and 0.10 mg/ml (37-150 muM); it is not affected by incubation with ribonuclease before staining; propidium iodide in 0.1% sodium citrate remains stable for at least 20 days; and quantitative estimates of the fractions of cells in the different phases of the cell cycle are in good agreement with those obtained from acriflavine-Feulgen staining and from autoradiography after pulse labeling with tritiated thymidine. We conclude that this method is useful for the measurement of relative DNA content by flow cytofluorometry, although modifications in the technique are necessary for some cell types which grow in monolayers.


Effect of colchicine on rat mast cells.

Effect of colchicine on rat mast cells. In the mast cell, a well-developed array of microtubules is centered around the centrioles. Complete loss of microtubules is observed when mast cells are treated with 10(-5) M colchicine for 4 h at 37 degrees C. The loss of ultrastructurally evident microtubules is associated with a marked change in the shape of mast cells from spheroids to highly irregular, frequently elongated forms with eccentric nuclei. In colchicine-treated cells the association of nucleus, Golgi apparatus, and centrioles is also lost. Mast cells exposed to 10(-5) M colchicine for 4 h at 37 degrees C retain 80% of their capacity to release histamine when stimulated by polymyxin B. Exocytosis is evident in stimulated cells pretreated with colchicine and lacking identifiable microtubules. When the conditions of exposure of mast cells to colchicine are varied with respect to the concentration of colchicine, the length of exposure, and the temperature of exposure, dissociation between deformation of cell shape and inhibition of histamine secretion is observed. These observations indicate that microtubules are not essential for mast cell histamine release and bring into question the assumption that the inhibitory effect of colchicine on mast cell secretion depends on interference with microtubule integrity.


Apparent anomalies in nuclear feulgen-DNA contents. Role of systematic microdensitometric errors. The Feulgen-DNA contents of human leukocytes, sperm, and oral squames were investigated by scanning and integrating microdensitometry, both with and without correction for residual distribution error and glare. Maximally stained sperm had absorbances which at  lambdamax exceeded the measuring range of the Vickers M86 microdensitometer; this potential source of error could be avoided either by using shorter hydrolysis times or by measuring at an off-peak wavelength. Small but statistically significant apparent differences between leukocyte types were found in uncorrected but not fully corrected measurements, and some apparent differences disappeared when only one of the residual instrumental errors was eliminated. In uncorrected measurements, the apparent Feulgen-DNA content of maximally stained polymorphs measured at lambdamax was significantly lower than that of squames, while in all experimental series uncorrected measurements showed apparent diploid:haploid ratios significantly greater than two. In fully corrected measurements no significant differences were found between leukocytes and squames, and in four independent estimations the lowest diploid:haploid ratio found was 1.99 +/- 0.05, and the highest 2.03 +/- 0.05. Discrepancies found in uncorrected measurements could be correlated with morphology of the nuclei concerned. Glare particularly affected measurements of relatively compact nuclei such as those of sperm, polymorphs and lymphocytes, while residual distribution error was especially marked with nuclei having a high perimeter:area ratio (e.g. sperm and polymorphs). Uncorrected instrumental errors, especially residual distribution error and glare, probably account for at least some of the previously reported apparent differences between the Feulgen-DNA contents of different cell types. On the basis of our experimental evidence, and a consideration of the published work of others, it appears that within the rather narrow limits of random experimental error there seems little or no reason to postulate either genuine differences in the amounts of DNA present in the cells studied, or nonstoichiometry of a correctly performed Feulgen reaction.


Immunofluorescence evidence for the absence of histone H1 in a mitotically dividing, genetically inactive nucleus. Antibodies directed against whole histone and purified lysine-rich histone H1 extracted from isolated macronuclei of the ciliate Tetrahymena were obtained and conjugated to fluorescein isothiocyanate. The fluorescein-antibody conjugates were used to directly label Tetrahymena cells. Both macro- and micronuclei were visibly fluorescent in cells stained with anti-whole histone conjugate. However, the anti-H1 conjugate only labeled macronuclei. This in situ demonstration of the lack of positive immunofluorescent staining of micronuclei with anti-H1 conjugate provide further evidence for the absence of H1 in the genetically inactive, mitotically dividing Tetrahymena micronucleus.


Preclinical hyperthyroidism in multinodular goiter. The thyrotropin (TSH) response to thyrotropin-releasing hormone(TRH) (200 mug iv) was determined in 80 surgical patients with nontoxic multinodular goiter. The TSH reserve was normal in multinodular goiter. The TSH reserve was normal in 55 and elavated in 8 patients. No TSH response to TRH (deltaTSH less than or equal to 1 muU/ml) was detectable in 17 patients (21%). Individual and mean serum T4, FT4I and serum T3 values did not differ from normal in 13 of the TRH unresponsive patients; in 4 patients FT4I or serum T3 was marginally elevated. No statistical differences were noted for I131-uptake, PBI131 and conversion rate between controls and TRH unresponsive patients. All patients who failed to respond to TRH were euthyroid on clinical evaluation. Goiters were large multinodular and long-standing in most instances. In 12 tested subjects TRH responsiveness recovered following partial thyroidectomy. In 3 of 7 TRH unresponsive euthyroid patients tested 9-12 days post surgery a transient lack of TSH to respond to TRH was observed. Recovery of TRH responsiveness was accompanied by a significant (P IS LESS THAN 0, 02) decrease in serum T4and FT4I in the euthyroid range, whereas no change in serum T3 occurred. It is suggested that TRH unresponsiveness represents a state of preclinical hyperthyroidism maintained by autonomously functioning goiter compartments.


A study of possible biohazards in the fluorescent antibody test using adenovirus, coxsackievirus, herpesvirus, and respiratory syncytial virus as antigens. Infectious adenovirus type 5 and coxsackievirus type B5, both nonlipid-containing viruses, were isolated from cells fixed in acetone at 22 degrees C for 15 min, from acetone used for fixation, from the solution used for washing slides during the fluorescent antibody procedure, and after complete processing of antigen preparations with serial twofold dilutions of human antisera and fluorescein-labeled goat anti-human immunoglobulin G. Lipid-containing herpes simplex virus type 1 and respiratory syncytial virus were inactivated by acetone, and infectious virus could not be recovered at any stage in the fluorescent antibody test. Fixation in acetone at 56 degrees C destroyed the infectivity of adenovirus 5 and coxsackievirus B5 within 30 min, but no adverse effect on the antigenic determinants of either virus occurred until after 60 min, thus demonstrating that these antigens can be utilized without the hazard of infectious virus.


Microscopy of stained urine smears to determine the need for quantitative culture. Consecutive specimens (2,564) of urine were cultured quantitatively, and Gram-stained smears were prepared from centrifugates as well as from the uncentrifuged specimens. About half of the specimens harbored organisms, and the quantity seen in smears of the centrifugates correlated reasonably well with the numbers of viable organisms cultured from the specimens. For example, smears of 900 centrifugates had one or more organisms per oil immersion field; 712 of their respective specimens proved to have colony counts of 10(5) or greater, and 120 had 10(4) to less than 10(5). For 188 specimens, however, the smears falsely predicted greater than or equal to 10(5) colony-forming units (many of these fell into the group that had 10(4) to less than 10(5)) but failed to predict clinically significant concentration of 10(5) or greater in only 31. Predictive values are presented also for lesser quantities of organisms seen in such smears. With rare exception, smears of centrifugates were superior to those of whole specimens as predictors of the concentration of viable organisms. Evaluation of smears proved not to be biased by the operator; values did not deviate significantly depending upon whether one or several microbiologists performed the evaluations.


Comparison of machine and manual staining of direct smears for acid-fast bacilli by fluorescence microscopy. Comparisons were made in Lusaka and in London between manual staining and staining in an automatic machine with auramine-phenol of direct smears of sputum and other types of specimen for acid-fast bacilli. No evidence was obtained of carry-over of acid-fast bacilli from positive to negative smears during machine staining. There was improved contrast between bacilli and the background in smears prepared with the machine.


Changes in protease inhibitors after acute myocardial infarction. Plasma levels of fibrinogen, alpha1-antitrypsin, alpha2-macroglobulin, antithrombin III, and C1 inactivator were measured serially for 10 days in 11 patients after acute myocardial infarction. Both fibrinogen and alpha1-antitrypsin rose markedly to reach peak levels 5-7 days after infarction while C1 inactivator levels rose slowly with the highest observed mean level on the 10th postinfarction day. Neither antithrombin III nor alpha2-macroglobulin changed significantly after myocardial infarction. No relationship between C1 inactivator levels and either fibrinogen or alpha1-antitrypsin was found in a study of 30 patients with a variety of disorders while fibrinogen and alpha1-antitrypsin levels were significantly correlated.


The efferent connections of the ventromedial nucleus of the hypothalamus of the rat. The efferent connections of the ventromedial nucleus of the hypothalamus (VMH) of the rat have been examined using the autoradiographic method. Following injections of small amounts (0.4-2.0 muCi) of tritium labeled amino acids, fibers from the VMH can be traced forward through the periventricular region, the medial hypothalamus and the medial forebrain bundle to the preoptic and thalamic periventricular nuclei, to the medial and lateral preoptic areas, to the bed nucleus of the stria terminalis and to the ventral part of the lateral septum. Some labeled axons continue through the bed nucleus of the stria terminalis into the stria itself, and hence to the amygdala, where they join other fibers which follow a ventral amygdalopetal route from the lateral hypothalamic area and ventral supraoptic commissure. These fibers terminate in the dorsal part of the medial amygdaloid nucleus and in the capsule of the central nucleus. A lesser number of rostrally directed fibers from the VMH crosses the midline in the ventral supraoptic commissure and contributes a sparse projection to the contralateral amygdala. Descending fibers from the VMH take three routes: (i) through the medial hypothalamus and medial forebrain bundle; (ii) through the periventricular region; and (iii) bilaterally through the ventral supraoptic commissure. These three pathways are interconnected by labeled fibers so that it is not possible to precisely identify their respective terminations. However, the periventricular fibers seem to project primarily to the posterior hypothalamic area and central gray, as far caudally as the anterior pole of the locus coeruleus, while the medial hypothalamic and medial forebrain bundle fibers apparently terminate mainly in the capsule of the mammillary complex, in the supramammillary nucleus and in the ventral tegmental area. The ventral supraoptic commissure fibers leave the hypothalamus closely applied to the medial edges of the two optic tracts. After giving off their contributions to the amygdala, they continue caudally until they cross the dorsal edge of the cerebral peduncle to enter the zona incerta. Some fibers probably terminate here, but others continue caudally to end in the dentral tegmental fields, and particularly in the peripeduncular nucleus. Within the hypothalamus, the VMH appears to project extensively to the surrounding nuclei. However, we have not been able to find evidence for a projection from the VMH to the median eminence. Isotope injections which differentially label the dorsomedial or the ventrolateral parts of the VMH have shown that most of the long connections (to the septum, amygdala, central tegmental fields and locus coeruleus) originate in the ventrolateral VMH, and there is also some evidence for a topographic organization within the projections of this subdivision of the nucleus.


Origins of axons in the cat's acoustic striae determined by injection of horseradish peroxidase into severed tracts. Origins and terminations of fibers of the dorsal and intermediate acoustic striae were studied by surgically severing these tracts and injecting HRP into the incision. This procedure results in filling the severed axons with HRP. Filled axons were traced to cell groups of origin and to some terminations of the acoustic striae. HRP-labeled terminals were found in the cochlear nuclei as well as in periolivary cell groups. Filling of cells with HRP RANged from being complete, resulting in Golgi-like images, to being barely detectable. Labeled cells were abundant in the dorsal and posteroventral cochlear nucleus adjacent to the injection as well as scattered throughout the periolivary cell groups of both sides, being highest in concentration around the ipsilateral lateral superior olive. On the side contralateral to the injection, labeled cells were found along the medial border of the dorsal cochlear nucleus, in the interstitial nucleus of the stria of Held, and sparsely throughout the ventral cochlear nucleus. The distribution of labeled cells was similar following HRP injections of the dorsal cochlear nucleus, except that these injections revealed additional descending projections from the inferior colliculi and from the ventral nucleus of the trapezoid body of both sides. These additional projections were interpreted as entering the CN by a ventral route. Findings of this study are in accord with physiological recordings made from fibers of the acoustic striae.


The effect of atropine and albuterol aerosols on the human bronchial response to histamine. This study was designed to determine whether histamine-induced bronchoconstriction in human asthmatics is mediated by the parasympathetic nervous system and involves cholinergic pathways. Inhalation challenges were performed on 14 adult asthmatic patients using the standardized procedure for inhalation challenge recently recommended by the Asthma and Allergic Disease Centers panel. The effect of pretreatment with either aerosolized atropine sulfate or aerosolized albuterol, a specific beta-2 adrenergic agonist, was studied. The comulative units of histamine required for induction of a positive bronchial response (20% or greater drop in FEV1 from baseline) was used as the basis of comparison of the effects of these drugs. This value was expressed as the PD20-FEV1 to histamine. Analysis of the data showed that aerosolization of sufficient atropie to effect a cholinergic blockade, as shown by inhibition of the bronchial response to inhaled methacholine, only minimally affected the bronchial response to histamine (p less than 0.05). However, the administration of albuterol markedly shifted the response to histamine (p less than 0.005). Although there was a statistically significant change in the mean PD20-FEV1 to histamine following atropine blockade, this effect was small in comparison to that which could be demonstrated with a beta agonist. It would thus appear that the major influence of histamine is not through cholinergic pathways.


Rat urinary metabolites from O,O-diethyl-O-(3,5,6-trichloro-2-pyridyl) phosphorothioate. Rats metabolized single oral doses of O,O-diethyl-O(3,5,6-trichloro-2-pyridyl-2,6-14C) phosphorothioate to at least six radiolabeled urinary metabolites. The urine contained about 90 percent of the dose. Three of these metabolites were identified as the glucuronide of 3,5,6-trichloro-2-pyridinol (80% the urinary 14C), a glycoside of 3,5,6-trichloro-2-pyridinol (4%), and 3,5,6-trichloro-2-pyridinol (12%).


1,2-Benzanthracene in soil.

1,2-Benzanthracene in soil. Samples from three cultivated soils and one from a roadside, all in Ontario, were found to contain less than 1 to 68 ppb of 1,2-Benzanthracene (BA). Two plots subjected to stubble (residue of wheat crop) burning annually for 15 years did not contain significant amounts of BA, although polyaromatic hydrocarbons, including BA, result from pyrolysis of most organic matter.


Immunocytochemical identification and localization of immunoglobulin A within Paneth cells of the rat small intestine. Light microscopic immunocytochemistry was used to identify Paneth cells by their lysozyme content and to detect immunoglobulin antigens within a subpopulation of these cells. Antisera specific for the heavy chains of rat or human immunoglobulin A and for immunoglobulin light chain antigens produced specific staining of rat Paneth cells. The distribution of immunoglobulin staining varied between adjacent Paneth cells in the same crypt and between Paneth cells in adjacent crypts, as well as between Paneth cell populations of different animals. No staining of rat Paneth cells was detected using antisera specific for the heavy chain of immunoglobulins G or M. The specific staining of Paneth cells for immunoglobulin A and light chain antigens was blocked by absorption of each antiserum with its respective purified antigen. Absorption of these antisera with purified rat lysozyme did not affect staining and thereby eliminated the possibility of immunologic cross-reactivity between lysozyme and immunoglobulin antigens. It is suggested, in light of current concepts of Paneth cell function, that the immunoglobulin staining of Paneth cells may reflect their ability to phagocytize immunoglobulin A-coated microorganisms or immune complexes containing immunoglobulin A.


[Glycoproteins in maternal and fetal blood and amniotic fluid. Variations in normal and pathologic pregnancies]. In this brief note, which is a preliminary report, the authors investigated systematically the variations, during normal and complicated pregnancies, of the concentrations of alpha-1-antitrypsin, orosomucoid, transferrin and alpha feto-protein, simultaneously in maternal serum, fetal serum and amniotic fluid. There exists during normal pregnancies a difference between the concentrations of alpha-1-antitrypsin and of orosomucoid found for primigravidae and for multigravidae. The role of these glycoproteins in preventing the mother from rejecting the fetus is discussed. In addition for Rhesus isoimmunization the clinical value of the variations in the levels of transferrin is discussed.


Virus-enhanced modulation of cell surface antigens: effect on immune lytic susceptibility. Monkey kidney cells, upon progressive subculture, became refractory to complement (C)-dependent immune cytolysis by anti-cell serum. Arbovirus infection restored these cells to a state of lytic susceptibility. Similar results were also abtained with antibody-dependent cellular cytotoxicity (ADCC), which is C independent. Antibodies raised against different subcultures varied considerably in lytic efficiency, indicating changing patterns of host cell expression during continous subculture. Taken together with the fact that arbovirus infection festored the lytic efficiency of all antibody preparations to the same degree suggested some form of host cell antigen re-expression as a mechanism. The results obtained in several exploratory experiments indicated that the antigenic re-expression responsible for the restoration of lysis was probably a local or selective rather than a generalized phenomenon. Thus, the amount of host cell surface antigen, measured by the use of mouse anti-cell serum and 125I anti-mouse globulin, was identical in both uninfected lytic susceptible and refractory cells, and decreased in both functional states following infection. Further, the binding of 125I concanavalin A, used to quantify surface glycoproteins, was similar in both lytic refractory and susceptible cells, and in both cases declined folowing virus infection. This result was incompatible with gross "masking" of cell surface antigens by exuberant production of surface coat material in lytic resistant cells. Finally, brief trypsinization of lytic resistant cells yielded an 8-fold increase in immune lysis, a result further consistent with local rather than generalized surface changes. The data were discussed interms of modulation of cell surface antigens affected both by repeated subculture and arboviral infection, and as a possible in vitro correlate of altered self-reactivity.


[Alpha feto protein. Normal pregnancies and maternal diseases. Apropos of 3,010 radioimmunologic determinations]. Using an immunological technique limits of normal levels of AFP in maternal blood have been worked out during pregnancy from a series of 3010 samples of blood. Comparing mean curves for the levels of AFP in mothers suffering from anaemia and diabetes with normal curves shows that there is a significant rise in these levels in the third trimester of pregnancy. The possible mechanisms and their relationship to the variation levels of AFP are discussed.


Coexistence of helper and suppressor activities in carrier-primed spleen cells. Both helper and suppressor activities for an in vitro IgG anti-hapten (Lac) response to Lac-HRBC were demonstrated in the same population of carrier (HRBC)-primed spleen cells. The relative radiosensitivity of suppressor activity permitted the selective removal of suppression and the demonstration of help. Both activities appear to be T cell-dependent and antigen-specific. Dilution analysis showed that help and suppression are mediated by distinct populations of cells.


Specific, transient suppression of the immune response by HGG tolerant spleen cells. II. Effector cells and target cells. In a previous report, it was shown that spleen cells from mice made tolerant to human gamma-globulin (HGG)5 could specifically inhibit the immune response of normal spleen cells after adoptive transfer to lethally irradiated recipients. However, that report also showed that the suppressive activity was only transiently associated with tolerant spleen cell populations. It was concluded from those experiments that while suppressive activity could be demonstrated in tolerant spleen cells under certain conditions, such activity was not obligatory for the maintainance of the tolerant state. The experiments presented here were performed to determine the nature of the effector cell(s) and the target cell(s) involved in this system of suppression of the immune response. Treatment of cells from tolerant animals with anti-thymocyte serum and complement to remove thymus-derived (T) cells completely abrogated suppresive activity. Removal of adherent cells from tolerant spleen cells by passage over glass wool columns resulted in partial loss of the suppression. The inhibitory activity of the suppressor cells was resistant to 900 R irradiation regardless of whether the tolerant spleen cells were irradiated before or after adoptive transfer. The cellular target(s) for the supprssor cells was examined by using lipopolysaccharide (LPS) as an alternative source of helper activity for the response to HGG. LPS, injected at the time of the initial antigenic challenge of mice that had been reconstituted with tolerant and normal spleen cells, prevented the expression of suppression against bone marrow-derived (B) cells. However, when LPS was presented only at the time of secondary antigenic challenge, it was unable to overcome suppression of the immune response of reconstituted recipients. Thus, LPS could produce a state where the B cells were resistant to suppression, but LPS could not rescue the responsiveness of B cells once the cells in the reconstituted recipient had been suppressed. In addition, the immune response to both the hapten dinitrophenol (DNP) and the carrier (HGG) were suppressed when recipients of tolerant and normal spleen cells were challenged with DNP6HGG. This indicates that T helper cells are also a target for suppression. The results presented in this paper are discussed in relation to a possible mechanism of suppression which proposes that suppressive activity represents the induction of tolerance in immunologically competent cells by HCG which is closely associated with the tolerant spleen cells.


Distinct target determinants on two lymphoblastoid lines derived from the same individual1. Specifically cytotoxic cells were generated in undirectional mixed leukocyte culture against both T and B lymphoblastoid cell lines. The cytotoxicity observed included both specific and less selective components which were differentiated by the interaction analysis based on the two-way analysis of variance. The results indicate that a T cell line and a B cell line derived from the same person possessed distinct target specificities. The antigens detected by direct cytotoxicity were confirmed by inhibition studies with competitor cells.


Reaginic antibody formation in the mouse. VII. Depression of the ongoing IgE antibody formation by suppressor T cells. The ongoing IgE antibody formation against ovalbumin (OA) in high responder mice was depressed by i.v. injections of either native or urea-denatured ovalbumin (UD-OA). Adoptive transfer experiments to determine the helper function of spleen cells from the treated animals showed that helper function for both IgE and IgG antibody responses diminished after treatment. Evidence was obtained that treatment suppressed the expansion of IgE-G memory cells. When the same treatment with OA or UD-OA was given to OA-primed mice before the appearance of IgE antibody in their serum, OA-specific splenic suppressor T cells were demonstrable. Thus, the transfer of splenic T cells from treated mice into normal mice suppressed the primary IgE and IgG antibody responses of the recipeints to DNP-OA. It was also found that the transfer of the splenic T cells from UD-OA-treated mice into OA-primed mice depressed ongoing IgE antibody formation in the recipients. The results suggested strongly that the decrease of helper function and the depression of ongoing IgE antibody formation by repeated injections of UD-OA was caused by generation of antigen (OA)-specific suppressor T cells.


Macrophage-lymphocyte interaction. III. Site of alloantiserum inhibition of T lymphocyte proliferation induced by allogeneic or aldehyde-bearing cells. Inhibition by anti-Ia sera of guinea pig T lymphocyte proliferation induced by allogeneic macrophages (MLR) and NaIO4 or neuraminidase-galactose oxidase-treated macrophages has been investigated in order to identify the target cell upon which the antisera act. Anti-2 and anti-13 alloantisera were found to inhibit both MLR and aldehydeinduced T cell reactivity when directed against the specificity of the stimulatory macrophage. Little or no inhibition was observed when these antisera were directed against the T lymphocyte specificity when cultures were harvested at the time of peak proliferation. In addition, anti-2 serum was found to inhibit macrophage-lymphocyte rosett formation at 20 hr between neuraminidase-galactose oxidase-treated strain 2 macrophages and strain 13 lymphocytes. These findings demonstrate that inhibition of T cell proliferation can be produced by anti-Ia sera directed against the macrophage and raise the possibility that Ir gene products may function in part at the level of the macrophage.


B lymphocyte alloantigen specificities present on cultured lymphoblastoid cell lines. Thirty-three of 34 cultured human lymphoblastoid B cell lines have been shown to express four out of five polymorphic non-HLA specificities expressed by normal B lymphocytes. The specificities were detected by 32 alloantisera produced by absorption with pooled platelets to remove HLA activity and selected from over 400 pregnancy sera. One B group (B2) which was expressed by 23% of a panel of normal B lymphocytes was not found on any of the cultured lines tested. Four T cell lines tested and myeloid line (K562) did not react with any of the B cell alloantisera.


Nucleoside specificity in the carrier IgG-dependent induction of tolerance. Induction of tolerance to nucleoside haptens in BALB/c mice with isologous IgG conjugates bearing four nucleosides simultaneously (A, G, C, T)-IgG was confirmed. A mixture of separate nucleoside-IgG tolerogens (A-IgG, G-IgG, C-IgG, and T-IgG) was as effective or more effective that the (A, G, C,T)-IgG form in suppressing the response to (A, G, C, T)-KLH. The nucleosides acted independently and simultaneously, since tolerogens with varying combinations of nucleosides caused specific suppression of the respones to only those nucleosides present on the tolerogen. Nucleoside-IgG conjugates did not suppress the response to denatured DNA-methylated bovine serum albumin, in which larger oligonucleotide determinants predominate. In varying combinations, guanosine was the dominant nucleoside both for immunization and for induction of tolerance. After three or four immunizations, control immunized animals made mainly IgG anti-nucleoside antibodies and this IgG antibody formation was preferentially suppressed in tolerogen-treated animals. Tolerance could be established before the primary or secondary immunization and it then persisted for at least 75 days through a fourth course of immunization. The same dosage of tolerogen did not reverse a strongly established anti-nucleoside antibody production after a tertiary response.


Nonspecific inhibition of tumor growth in vivo by admixed allogeneic tumor-sensitized lymphoid cells and identical inactivated allogeneic tumor cells. With the in vivo tumor neutralization test (Winn test), growth of a transplanted (KMT-17) from Wistar-King-Aptekman rats was inhibited by allogeneic tumor (AH-66 from Donryu rats)-sensitized syngeneic lymphoid cells admixed with mitomycin C (MMC)-treated AH-66 cells. The observed tumor inhibition may be immunologically nonspecific, since no cross-antigens were detected by membrane immunofluorescence on the surfaces of KMT-17 and AH-66 cells. Close contact among KMT-17, AH-66-sensitized lymphoid cells and MMC-treated AH-66 cells was required for the inhibition of KMT-17 growth. AH-66 cells pretreated with formalin or ultrasonication lost tumor inhibitory activity when they were admixed with AH-66-sensitized lymphoid cells, and only MMC-treatment effectively preserved the tumor inhibitory activity of AH-66 cells. The sensitized spleen cells, draining lymph node, or peripheral blood cells inhibited tumor growth when they were admixed with MMC-treated AH-66 cells, whereas nucleated cells from bone marrow, thymus, or distal lymph node did not. Growths of KMT-17 were inhibited by admixed sensitized spleen cells and MMC-treated AH-66 even when pre-irradiated rats were used as recipients.


Release of chemical mediators from partially purified human lung mast cells. Human lung mast cells dispersed by enzymatic digestion of human lung fragments were concentrated to greater than 50% purity by sedimentation in isopycnic and velocity gradients. The dispersed lung mast cells had a characteristic ultrasturctural appearance including granules with a scroll or reticular structural appearance including granules with a scroll or reticular structure surrounded by perigranular membranes. Histamine and preformed eosinophilotactic activity sedimented with mast cells on isopycnic gradients, and mast cells and these mediators were separated from the bulk of the other lung cells after velocity gradient sedimentation. The histamine content of isolated lung mast cells was calculated to range from 1.0 to 5.5 pg/cell. The quantity of SRS-A generated with anti-IgE or specific antigen was relatively limited but confined to the mast cell-rich fractions and associated with release of histamine and eosinophilotactic activity.


Primary in vitro sensitization of murine lymphocytes against isogeneic and allogeneic cells transformed by simian virus 40. Primary in vitro sensitization of murine lymphocytes to isogeneic and allogeneic cells transformed by simian virus 40 (SV40) is described. The results of specificity studies utilizing cytotoxic effector lymphocytes obtained by in vitro immunization indicate that SV40 transformation results in the expression of tumor-specific antigens which are recognized by cytotoxic effector cells. Moreover, the studies demonstrate that expression of tumor-specific antigens on transformed cells is associated with a reduction in the functional expression of normal histocompatibility antigens.


Characterization of antibodies to the structural polypeptides of HGSAg: evidence for subtype-specific determinants. Antisera prepared in guinea pigs to the structural polypeptides of HBAAg/adw and HBSAg/ayw were examined by a modified passive hemagglutination assay for antibodies to the subtype-specific d and y determinants. All of the isolated polypeptide fractions stimulated antibodies to both group specific and subtype-specific antigens of the native HBSAg particle from which they were derived. These data indicate that the polypeptides have similarities in their immunochemical structure.


The demonstration of cell-associated immunity to viruses. In vitro lymphocyte responsiveness to Varicella-zoster antigen. The demonstration of in vitro lymphocyte responsiveness to common pediatric viruses has previously been fraught with many technical and conceptual problems. Based upon our prior experience in demonstrating cell-associated immunity to mumps, rubella and measles viruses we illustrate our methodology and conceptual framework by documenting in vitro lymphocyte responsiveness to the Varicella-zoster virus, another ubiquitous virus of childhood. We discuss our approach to the problems of reactivity, specificity and reliability in the use of membrane-associated viral antigens.


Measurement of the fractional uptake of macromolecules by the renal vascular bed compared to other vascular beds. Macromolecules resembling soluble immune complexes can be made from heat-aggregated human gamma globulin (AHGG). In 15 rats, we studied vascular trapping of 125I-labeled AHGG (AHGG)-125I) given by constant I.V. infusion over 1 hour while tissue blood flow was marked by intermittant aortic arch injections of 85Sr-labeled microspheres. Red cells labeled with 51Cr (RBC-51Cr) were also infused so that when the tissues were removed at the end of the experiment, the vascular volume of each tissue specimen could be balculated to correct issue 125I for AHGG-125I which was not trapped but simply in transit in the bascular space at the time the tissue was removed. These data permitted us to calculate the fractional uptake of AHGG-125I (FM) for a given tissue in comparison to any other tissue. We chose to compared the FM of each tissue to the FM of renal cortex. This comparison was expressed as a ratio termed the FM ratio for the given tissue. The following tissues had FM ratios significantly greater than 1.00 (i.e., per unit blood flow, these tissues trapped AHGG-125I more avidly than renal cortex): liver, spleen, skin, stomach, fat, testes, and large bowel. The respective ratios were 381 +/- 74, 15.7 +/- 4.0, 11.8 +/- 4.0, 7.47 +/- 1.95, 6.24 +/- 1.0 +/-, 3.03 +/- 0.67, 2.86 +/- 0.72 (all p less than 0.025). The FM ratio for adrenal, heart, thymus, and diaphragm were not significantly different from 1.00. The FM ratio of lung and brain were significantly less than 1.00: 0.014 +/- 0.008 and 0.14 +/- 0.065, respectively (p less than 0.001 for both). In 13 experiments, glomeruli was 23.8 +/- 3.5 per cent as assessed by recovery of the microspheres contained in renal cortex. Compared to whole renal cortex, the isolated glomeruli contained only minor amounts of AHGG-125I. We conclude that tissues vary widely with respect to their ability to trap macromolecules. When uptake is viewed in terms of the amount of complex trapped per unit delivery rate, many organs trap AHGG-125I for more avidly than renal cortex. Furthermore, under the present experimental conditions, glomeruli are not the major intrarenal site of macromolecule uptake.


Wenckebach-type exit block from an ectopic focus as a cause of variable coupling. Intermittent bigeminal and trigeminal ventricular premature beats were recorded in an otherwise healthy 14 year old male. Coupling intervals progressively lengthened until an ectopic beat was dropped. Odd numbers of sinus beats occurred between bigeminal runs. This rhythm is interpreted as being due to Wenckebach-type block in an exit pathway from the ectopic focus, resulting in concealment of the persistently active extrasystolic mechanism.


Prostatic distribution of sex hormone-binding globulin and cortisol-binding globulin in benign hyperplasia. Sex hormone-binding globulin-(SHBG) and cortisol-binding globulin-(CBG) like proteins have been demonstrated in prostatic tissue surgically removed from patients with benign prostatic hyperplasia. These proteins are not easily removed by superfusion of tissue slices. Epithelial tissue was separated from stroma and found not to contain the SHBG- or CBG-like proteins. Substantial amounts of these proteins, however, remained associated with the stroma. It is suggested that they may be constituents of interstitial fluid in this tissue compartment. The possible significance of this in benign prostatic hyperplasia is discussed.


Metal-androgen interrelationships in carcinoma and hyperplasia of the human prostate. Zinc and cadmium concentrations were measured by atomic absorption spectroscopy in normal and pathological human prostates. Our studies confirm the values of zinc in normal tissue [6.84 +/- 1.21 (S.E.M.) mumol/g] and benign prostatic hypertrophy (BPH) (6.9 +/- 1.19 mumol/g) are similar, while in neoplastic tissues zinc concentrations were significantly lower (2.61 +/- 0.45 mumol/g). The Cd2+ levels in BPH (23.11 +/- 3.28 nmol/g) were, on the other hand, considerably higher than those found for normal tissues (5.15 +/- 0.62 nmol/g). In agreement with other published reports, Cd2+ concentrations were found to be markedly increased in carcinomatous tissue (129.79 +/- 22.22 nmol/g). No correlation was however established between the values for the two metals in either type of prostatic tissue. An established specific radioimmunoassay was used for the measurement of testosterone and dihydrotestoesterone (DHT) and a distinct pattern emerged upon comparing these results with those for the zinc and cadmium concentrations. It appears that the concentrations of DHT in benign hypertrophy and of testosterone and DHT in carcinoma were inversely proportional to the levels of Zn2+ in abnormal tissue. In contrast, the DHT levels in the hypertrophied and malignant tissue were proportional to the Cd2+ concentrations.


Androgen levels in the plasma and prostatic tissues of patients with benign hypertrophy and carcinoma of the prostate. Specific radioimmunoassays for testosterone, dihydrotestosterone (DHT) and androstenedione were carried out to measure the concentrations of the three hormones in the plasma and prostatic tissue of ten patients with benign prostatic hypertrophy (BPH) and ten patients with carcinoma of the prostate. The results indicate that there are no significant differences between the peripheral plasma concentrations of testosterone, DHT and androstenedione in BPH [19.7 +/- 2.6, 2.6 +/- 0.9 AND 5.5 +/- 1.7 (S.E.M.) nmol/l respectively] and in carcinoma [16.9 +/- 2.8, 2.4 +/- 0.5, 4.4 +/- 1.1 nmol/l respectively], (in all cases P greater than 0.1). In contrast, the prostate tissue rations DHT: testosterone (3.59 +/- 0.55 for BPH and 0.66 +/- 0.09 for carcinoma) and androstenedione: testosterone (2.83 +/- 0.38 for BPH and 1.07 +/- 0.16 for carcinoma) are significantly less in carcinoma than in benign hypertrophy ( in all cases P less than 0.01). The accumulation of testosterone in the carcinoma, relative to values found in BPH tissue is, therefore, not associated with changes in the concentrations of androgens in the plasma pool but may be related to local factors and metabolic changes within the prostate.


Immunological studies of aging. II. Loss of IgG and high avidity plaque-forming cells and increased suppressor cell activity in aging mice. The magnitude and heterogeneity of the immune response to dinitrophenylated bovine gamma globulin was measured in aged and young mice at a cellular level using an inhibition of plaque-forming cell assay. The primary and secondary responses of 24-mo-old mice were markedly depressed in magnitude and restricted in avidity for the DNP determinant when compared to 2-mo-old animals. Bacterial lipopolysaccharide given at the time of immunization increased the restriction in heterogeneity seen in 12- and 24-mo-old mice. Indirect PFCs were more severely depressed than direct PFCs in 24-mo-old mice. Syngeneic, lethally irradiated, 2-mo-old mice reconstituted with aged spleen cells exhibit the depressed and restricted response to DNP-BGG seen in old mice. When 10(8) young thymus cells were given together with old spleen cells the heterogeneity of the response was increased. When 2-mo- and 24-mo-old spleen cells were transferred together into young recipients the magnitude of the response to the young spleen cells markedly reduced. Thus, there appears to be a loss of thymic-helper cells and an increase in suppressor activity in aged animals.


Alternative pathway of complement: demonstration and characterization of initiating factor and its properdin-independent function. A novel component of the properdin system has been discribed which represents a heretofore unrecognized human serum protein. The protein has been tentatively termed the initiating factor (IF) because it functions in the initial reaction of the properdin pathway. IF is a 170,000 dalton beta-pseudoglobulin which is composed of two presumably identical 85,000 dalton chains linked by disulfide bonds. The protein reacts with antibody to nephritic factor, which is defined by its activity and is found in the serum of patients with certain nephritides. The activity of IF is heat stable. Upon treatment of serum with activators of the alternative pathway, the initial C3 convertase is assembled from IF, Factors D and C, C3, and magnesium without participation of properdin. It is the function of the enzyme to deposit C3b on the surface of the activator particles, thereby affording generation of the solid phase enzymes of the pathway, a process that is a prerequisite for properdin activation. By exposure to low pH, IF assumed the electrophoretic mobility of psi-globulin and acquired the ability to generate without activators a fluid phase C3 convertase in serum. Serum depleted of IF did not allow activation of the properdin pathway. Serum depleted of properdin did permit activation of the pathway and expression of cytolytic activity. These results raise the possibility that IF represents the recognition unit of the pathway.


Ly phenotype of cytotoxic T cells for syngeneic tumor. Our present and previous findings may be summarized as follows: The phenotype of C57BL/6 (B6) cytotoxic cells for allogeneic target cells is Thy-1+, Ly-1- Ly-2/3+, MSLA+, and Ig-. the phenotype of B6 cytotoxic cells for syngeneic tumor cells is Thy-1+, Ly-1+, Ly-2/3+, MSLA+, and Ig-. The phenotype of B6 cytotoxic cells for syngeneic tumor cells is Thy-1+, Ly-1+, Ly-2/3+, MSLA+, AND Ig-. Thus, differences in Ly phenotype appear to be exhibited not only by cytotoxic T cells as opposed to helper T cells, but also within subcategories of cytotoxic T cells.


Independent differentiative pathways of Ly1 and Ly23 subclasses of T cells. Experimental production of mice deprived of selected T-cell subclasses. When B mice are supplied with Ly1 or Ly23 cells they acquire, over the next 6 mo, only the immune functions associated with each of these T-cell subclasses, respectively. The T-cell population of these "B-Ly1" and "B-Ly23" mice mice also remains restricted to the Ly1 and Ly23 subclass phenotypes. Thus the Ly1 and Ly23 populations are derived from two separate lines of differentiation and are not sequential stages of a single differentiative pathway.


Cell-mediated lympholysis to H-2-matched target cells modified with a series of nitrophenyl compounds. The specificity of C57BL/10 cytotoxic effector cells generated by in vitro sensitization with autologous spleen cells modified with a series of related nitrophenyl compounds was investigated. The failure of trinitrophenyl (TNP)-sensitized effector cells to lyse TNP-beta-alanylglycylglycyl(AGG)-modified target cells is presented as evidence contradicting the intimacy or dual receptor model or T-cell recognition in its simplest form. Data are also shown indicating that sensitization with N-(3-nitro-4-hydroxy-5-iodophenylacetyl)-AGG-modified stimulating cells generates noncross-reacting clones of cytotoxic effector cells.


Effects of anti-Ia sera on mitogenic responses. III. Mapping the genes controlling the expression of Ia determinants on concanavalin A-reactive cells to the I-J subregion of the H-2 gene complex. We have shown that the Ia determinants expressed on nylon wool-purified T lymphocytes reactive to concanavalin A (Con A) in serum-free media are coded in a single I subregion of the H-2 gene complex. This region, I-J, is defined by two pairs of intra-H-2 recombinant haplotypes: H-2t3, H-2t4 and H-2i3, H-2i5, carried by B10.HTT, B10.S(9R), B10.A(3R), AND B10.A(5R), respectively. No activity against Con A-reactive T cells has been detected in any antiserum that was produced in strain combinations which shared a common I-J region. This suggests that Ia antigens expressed on Con A-reactive T cells are restricted to the I-J subregion.


Serological detection of variable region (Vh) subgroups of Ig heavy chains. Serological test systems were established for determining the heavy-chain variable region (Vh) subgroups of immunoglobulin heavy chains. Myeloma proteins with known Vh subgroups based on amino acid sequence were utilized as the primary basis of reference for analysis by hemagglutination and hemagglutination inhibition. Good agreement between the chemical and serological typing was obtained and nonoverlapping systems established for the three major Vh subgroups. In a survey of 167 myeloma proteins, all except two were exclusively positive in one of the three systems. The two exceptions may represent a fourth subgroup. There was an overall incidence ratio of 1:2:3 for VhI:VhII:VhIII subgroups. Some differences in the overall ratios were encountered within the immunoglobulin classes. Certain advantages of the serological typing antisera were discussed with special emphasis on their use for studies of Vh antigens on the membranes of lymphocytes.


H-2 restriction of virus-specific cytotoxicity across the H-2 barrier. Separate effector T-cell specificities are associated with self-H-2 and with the tolerated allogeneic H-2 in chimeras. During infection with lymphocytic choriomeningitis or vaccinia virus, F1 irradiation chimeras reconstituted with bone marrow cells from or both parents generate cytotoxic T cells which can lyse targets across the H-2 barrier. However, activity of chimera T cells is H-2 restricted as shown by cold target competition experiments and selective restimulation of a secondary response in vitro; T cells of H-2k specificity which lyse tolerated infected H-2d target cells do not lyse infected H-2k or unrelated target cells and vice versa. Therefore, H-2 restriction of virus-specific cytotoxic T cells probably does not reflect need for like-like self-interactions for lysis to occur. The specificity of virus immune T cells is thus determined by the H-2K and H-2D specificities present in the infected animal and which are probably recognized unidirectionally by T cells. The results are compatible with the idea the T cells are specific for "altered alloantigen", i.e., a complex of cell surface marker and viral antigen. Alternatively, explained with a dual recognition model, T cells may possess two independently, clonally expressed receptors, a self-recognizer which is expressed for one of the syngeneic or tolerated allogeneic K or D "self" markers, and an immunologically specific receptor for viral antigen.


Suppression by autogenous complementary idiotypes: the priority of the first response. Complementary idiotypes or antibodies are considered to have combining site structures which are at least partly directed against each other. Complementary antibodies were induced in A/He mice by immunization with phosphorylcholine (PC)-containing antigens and by immunization with the PC-binding IgA myeloma protein TEPC-15 (T15). Both responses were monitored by enumerating plaque-forming cells (PFC) and assaying serum antibody levels against the corresponding antigens. Mice immunized at least three times with T15 in adjuvants had markedly suppressed responses to subsequent immunization with PC; similarly, mice preimmunized multiple times with PC had suppressed responses to immunizations with T15. In contrast, mice immunized with T15 in the interval between "primary" and "secondary" immunizations with PC had undiminished PFC responses to both antigens but significantly decreased antibody titers to PC. Simultaneous responses were also induced by immunizations with T15 superimposed on weekly immunizations with PC; with this regime, immunization with T15 actually enhanced the PFC response to PC, but serum antibody to PC was significantly lower than for mice immunized with PC only. Levels of serum antibody to PC were probably lower, either because anti-PC antibody was complexed with the complementary antibody directed against T15, or because the antibody directed against T15 prevented synthesis and/or release of anti-PC antibody by cells in vivo. Thus, an established prior autogenous immune response can dramatically suppress a subsequent primary complementary response, but the effects of complementary responses on each other are more complex with different sequences of immunization. Also, the effects of variables such as the amounts and ratios of the classes of antibodies on regulation of complementary responses remain to be defined.


Anti-idiotype sera raised against surface immunoglobulin of human neoplastic lymphocytes. The idiotypic determinants of surface immunoglobulins on B-cell lymphomas and lymphocytic leukemias represent tumor-specific antigens, individually unique for each tumor. As such they have both diagnostic and therapeutic potential, particularly for those neoplasms with no serum monoclonal immunoglobulin arising from synthesis of the protein for export. We describe the raising in animals of anti-idiotype sera directed against two examples of a nonexporting neoplasm, human chronic lymphocytic leukemia. The procedure involves exposing the cells to papain so as to remove the Fab fragments (containing the idiotypic determinants) from the surface immunoglobulin, recovering the Fab on cellulose immunosorbent particles, and immunizing animals with the immunosorbent-Fab complex.


Lysis of RNA tumor viruses by human serum: direct antibody-independent triggering of the classical complement pathway. In earlier studies we found that human serum, but not serum from multiple other species, inactivated and lysed oncornaviruses from a number of diverse sources in the apparent absence of antibody. A detailed analysis of the role of the human complement (C) system in mediating this lytic process indicates that human C1q interacts directly, in the absence of immunoglobulin, with oncornaviruses. Binding of C1 via C1q in this manner leads to activation of C1r, C1s, and thus of the classical C pathway. Integrity of the classical pathway  is an absolute requirement for lysis although activation of the alternative pathway considerably amplifies the amount of lysis obtained, possibly through involvement of the C3b-dependent feedback mechanism. Activation of C is accompanied by deposition of C components on the viral surface and lysis on completion of the C reaction sequence. Thus in this system, the C1q subunit of C1 subserves a specific recognition function normally associated with antibody. This ability of human serum to inactivate oncornaviruses may represent a natural defense mechanism operative in vivo which deters expression of intact oncornaviruses in human malignancies.


The analysis of the monoclonal immune response to influenza virus. II. The antigenicity of the viral hemagglutinin. The antigenicity of the hemagglutinins (HA) of five influenza viruses of the A0 and A1 subtypes has been analyzed by means of monoclonal antibodies of murine origin produced in vitro. Secondary monoclonal anti-HA(PR8) antibodies were able to differentiate 14 antigenic determinants (or groups of determinants) on the HA of five influenza virus strains of the A0 and A1 subtypes. Taking into account that certain pairs of determinants delineated on heterologous HA may reflect the heterogeneity of the humoral immune response to a single homologous determinant, the presence of at least eight determinants (host cell-derived determinants not included) on the homologous HA of PR8 and probably on the HA of influenza viruses in general is postulated. Three types of HA-determinants of A0 and A1 influenza virus strains could be distinguished: strain-specific, partially shared, and determinant(s) common to all five virus strains tested. Roughly 40, 55, and 5%, respectively, of the secondary anti-PR8 antibodies of BALB/c mice were directed against determinants belonging to either of the three types.


Some characteristics of salt-dependent haemagglutinating measles viruses. Several strains of measles virus which did not agglutinate monkey erythrocytes in phosphate-buffered saline did so in buffer containing 0-8 M-ammonium sulphate. Haemadsorption to cells infected with these viruses was also salt-dependent. In a series of tests salt-dependent agglutinin was shown to be a stable structural component of the infectious virion. The relevance of these findings is discussed in the light of previous reports that many measles virus preparations do not agglutinate erythrocytes.


Tryptic cleavage of antibody binding sites from hepatitis B surface antigen particles. The sedimentation of radiolabelled 22 nm hepatitis B surface antigen particles was unaffected by treatment with either trypsin or SDS alone, but combined treatment disrupted the particulate nature of the radiolabelled material. Considerable antibody binding activity by the group-specific determinant (a) was preserved after combined SDS and trypsin treatment but was released from the bulk of the radiolabelled protein; gel filtration indicated an approximate mol. wt. of 5000 to 15000 for the released antibody binding material. This material was precipitated by concanavalin A, suggesting the presence of carbohydrate. Its serological activity was remarkably resistant to boiling and to proteolytic digestion, but was partially sensitive to treatment with 0-01 M-periodate or with mixed carbohydrases and neuraminidase, and was greatly reduced by treatment with reducing agent. These data suggest that the stability of the a determinant is due to the structure of the antibody binding site itself, rather than to involvement in the quaternary structure of the particle, and that intact disulphide bonds and carbohydrate, closely related to the antibody binding site, are necessary for the full expression of serological acitivity.


A comparison of human papovavirus T antigens. A comparison was made of the T antigens induced in transformed cells or infected permissive cells by representatives of three categories of human papovavirus. The transformed hamster cell lines employed contained T antigen induced by either the BK or RF strains of papovavirus associated with human renal allografts; the JC strain of papovavirus from progressive multifocal leukoencephalopathy (PML), or a variant of SV40 virus isolated from PML. The human papovavirus T antigens were also compared with that of a human cell line transformed by SV40 of simian origin. Anti-T antibody prepared in hamsters against each of the hamster cell lines was absorbed with crude T antigen from each cell line, and the unabsorbed and absorbed antisera were tested for residual T antibody against each cell line, or against infected permissive cells by immunoperoxidase (IP) staining and complement-fixation (CF) tests. In unabsorbed antisera, T antibodies from each cell line cross-reacted with all T antigens in IP tests, and CF tests showed that T antisera reacted preferentially with T antigen induced by homologous virus. Absorpminants. T antigens of the two urine-derived strains, BK and RF, were identical or nearly so, but were clearly separable from T antigens of JC virus, PML-derived SV40 or simian-derived SV40. JC T antigen was intermediate, being more closely related to T antigens both of BK virus and SV40 virus than the latter were to each other. The T antigen of PML-derived SV40 could be distinguished from the T antigen of simian-derived SV40 and the T antigen of the SV40 variant from human brain was more closely related to those of the other human-derived papovaviruses than was the T antigen of SV40 from monkey kidney.


Evaluation of the blood-CSF barrier by protein gradients and the humoral immune response within the central nervous system. A linear correlation was found between the serum/cerebrospinal fluid (CSF) concentration ratios of albumin, caeruloplasmin and alpha2-macroglobulin and their hydrodynamic radii in a semilogarithmic plot. This protein gradient is used as a parameter to evaluate the blood-CSF barrier under normal and pathological conditions. Irrespective of vastly different transfer rates, the ratio/size permeation curves of proteins at the blood-CSF barrier and the blood-lymph barrier have comparable characteristics. Therefore the protein gradients found in various disease states are interpreted by means of Renkin's general law of lymph formation. Declined gradients are caused either by an increased permeability of the barrier sites or by a decreased turnover rate of the CSF within the compartment punctured. The concentration ratios of immunoglobulins are related to the gradient that is constructed with the ratios of the barrier-indicative marker proteins. As judged by comparative disc electrophoresis of serum and CSF, those disease states that are dominated by barrier impairment are used to establish the range of concentration ratios, compatible with a passive immunoglobulin transfer in any condition in which the barrier is disordered. A mathematical approach is described, which allows the quantitative evaluation of the minimal immunoglobulin portion that is synthesized within the central nervous system.


Generalized gangliosidosis: acid beta-galactosidase deficiency with early onset, rapid mental deterioration and minimal bone dysplasia. This report concerns a 3-month-old girl with rapidly progressive psychomotor retardation, hepatomegaly, vacuolated lymphocytes, minimal bone dysplasia and normal excretion of acid mucopolysaccharides. A deficiency of acid beta-galactosidase was demonstrated in isolated leucocytes and in a liver biopsy. The diagnosis of generalized gangliosidosis due to deficiency of beta-galactosidase was also based on the absence of the enzyme activity from cultured fibroblasts. The diagnosis was confirmed on autopsy at 16 months by typical histology, electron microscopy and biochemistry of the organs. beta-galactosidase deficiency has been demonstrated in various clinical conditions ranging from generalized gangliosidosis with severe mental retardation to clinical pictures resembling Morquio's disease and normal intelligence. The heterogeneity of the clinical manifestations in beta-galactosidase deficiency could be explained by different residual activities of a structurally mutated enzyme towards its various substrates.