Quantitation of T and B lymphocytes and cellular immune function in Hodgkin's disease. Peripheral blood T and B lymphocytes were quantitated in 42 patients with untreated Hodgkin's disease and the results compared with the response to phytohemagglutinin (PHA) stimulation and delayed hypersensitivity skin testing. T lymphocytes were identified by an in vitro cytotoxicity assay employing a specific anti-T-cell serum and by spontaneous rosette formation with sheep erythrocytes (E rosettes). The percentage of T cells in the patients was similar to that of normal subjects as judged by the cytotoxicity assay (65 to 90%). In addition, absolute T-lymphocyte counts were normal in 63% of the patients and were generally reduced only in those with lymphopenia. The percentage of T lymphocytes determined by the E-rosette assay was similar to that determined by the cytotoxicity assay in normal controls, but was significantly lower than that determined by the cytotoxicity assay in the patients. Moreover, the decreased response to PHA stimulation in the patients was directly correlated with the decrease in E-rosette formation. These findings suggest that T lymphocytes in the peripheral blood are not generally diminished in untreated Hodgkin's disease. However, a proportion of these cells exhibits altered surface interactions that may account for some aspects of their impaired immunologic function.


Combination chemotherapy of Hodgkin's disease with adriamycin, bleomycin, vinblastine, and imidazole carboxamide versus MOPP. This paper reports the preliminary results of a controlled study randomizing MOPP vs. a new four-drug combination (ABVD) in advanced Hodgkin's disease. ABVD consists of 6 cycles of adriamycin, bleomycin, vinblastine, and imidazole carboxamide. The purpose for designing this new combination was two-fold: to compare the efficacy of ABVD with MOPP, and to demonstrate absence of cross-resistance between the two regimens. Of 60 patients entered into the study, 45 (MOPP25, ABVD20) are presently evaluable for the analysis of remission induction. No patient was previously treated with chemotherapy; 20% had relapsed after primary radiotherapy. Whenever possible, complete remission was defined also through rebiopsy of known organ involvement. Complete remission occurred in 76% of patients treated with MOPP and in 75% of those given ABVD, with no difference between the two regimens as far as stage (IIIB-IIIS and IV), histologic type, and prior irradiation were concerned. Crossover carried out for progressive disease or for relapse after initial remission showed absence of cross-resistance between MOPP and ABVD. Toxic manifestations after ABVD were in general well tolerated and reversible. The percent of optimal dose for each drug was as follows: adriamycin 87%, vinblastine 87%, bleomycin 96%, and imidazole carboxamide 96%. These preliminary results indicate that in terms of complete remission, ABVD could represent a successful alternative to MOPP to be used either in MOPP failures or in sequential combination with MOPP. However, the lack of long-term followup limits at the present time an adequate comparison between the two treatments.


67Ga accumulation in pulmonary lesions associated with bleomycin toxicity. The radionuclide, gallium-67, accumulates in a wide variety of neoplastic and inflammatory lesions. Diffuse bilateral lung localization of 67Ga occurred in two cases of interstitial pneumonitis associated with bleomycin therapy. Clinical symptoms and laboratory evaluation of pulmonary status correlated with scintigraphic studies in these two patients, while discrepancies between scintigraphic and roentgenographic findings were observed.


Enhancement by tetrahydrouridine (NSC-112907) of the oral activity of 5-azacytidine (NSC-102816) in L1210 leukemic mice. 5-Azacytidine is more active when administered parenterally than orally in the treatment of L1210 leukemic mice. Oral coadministration of tetrahydrouridine, a pyrimidine nucleoside deaminase inhibitor with no intrinsic antitumor activity, greatly increases the oral activity of 5-azacytidine. 5-azacytidine (or cytotoxic equivalent) blood levels in BDF mice are much higher after oral administration of the 5-azacytidine-tetrahydrouridine combination than when 5-azacytidine is administered alone by the same route. The therapeutic results (L1210 leukemia) achieved with the oral combination are similar to those observed with parenteral 5-azacytidine alone.


Continuous intravenous bleomycin (NSC-125066) therapy with vinblastine (NSC-49842) in stage III testicular neoplasia. Twenty-three patients with stage III germinal neoplasia of the testis were treated with a variation of our original vinblastine-bleomycin program. This modification consisted of 0.4 mg/kg of vinblastine given in two fractions on Days 1 and 2 followed by continuous intravenous administration of 30 units of bleomycin in 1000 cc of 5% glucose and distilled water over a 24-hour period for 5 successive days beginning on Day 2. Therapy was repeated every 28-35 days as toxicity permitted. There were 17 responses, nine of which were complete (39%). Eight of the complete responses were in patients with massive disease in whom a low complete response rate was expected. Toxic effects consisted of severe leukopenia in 90% thrombopenia in 50%, and unexplained transient hyperbilirubinemia in about 30% of the patients. Bleomycin pneumonitis occurred in one patient and resulted in death. Hypertension was a new and unexpected side reaction experienced by four patients. Further trials are indicated since the complete response rate in patients with advanced massive disease appears to be improved.


Location of glycogen in spermatids and spermatozoa of the shipworm, Bankia australis (Teredinidae, Bivalvia, Mollusca). The periodic acid-thiocarbohydrazide-silver proteinate technique and alpha-amylase digestion were used to locate glycogen in the spermatids and sperms of the bivalve Bankia australis. Glycogen was found in the middle piece and around the acrosome in spermatozoa, apparently randomly scattered throughout the cytoplasm of young spermatids, and in a cytoplasmic bead in old spermatids.


Granulocyte differentiation by Friend leukemia cells. Friend leukemia cells growing in suspension culture are thought to represent a population of primitive erythroid cells which have undergone malignant transformation. We have found that when growing in vivo or in plasma clots in vitro, these suspension culture cells can exhibit morphologic and enzymatic properties which are characteristic of primitive granulocytic cells. The microenvironment in which the tumor cells grow plays a major role in determining the direction of differentiation of these leukemia cells. Hence it appears likely that the Friend cell is in fact a neoplastic pluripotent hematopoietic stem cell.


[Problem in the determination of calcinosis by means of hematoxylin]. The histochemical analysis of a number of dystrophic and metastatic calcifications revealed that the haematoxylinophilia of calcified tissues is not based on the inorganic component, but on their organic component. In most of the cases the material examined contained protein on the basis of its staining properties, as well as neutral and acid mucosubstances and, most probably, also covalently bound phosphate. Exceptionally the organic component appeared to be unequivocally acidic phospholipid, tigtly bound to an inorganic component. The significance of the findings has been discussed.


[Congenital toxoplasmosis with activation of extramedullary hematopoiesis in a premature infant]. An immature neonate surviving for 45 minutes showed a generalized hydrops and hepatosplenomegaly unassociated with blood incompatibility. A marked activation of extramedullary haematopoiesis was observed not only in its liver and spleen, but also in the interstitium of the lungs, pancreas, kidneys, and adrenal glands. In addition, extensive necrosis of the brain and diffuse nonsuppurative myositis and myocarditis were found. All the changes were due to congenital toxoplasmosis, which has been confrimed by the findings of occasional pseudocysts within the placenta, by the result of the parasitologic examination of the brain of the autopsied infant and by serological examination of the mother post partum.


[New histochemical contributions to the study of corticotropic cells in the adenohypophysis of the chick enbryo]. Comparative analysis of results obtained after subsequent treatments of chick embryo adenohypophysis by anti-ACTH 17-39 Is, PAS and Bodian method demonstrates that ACTH cells are PAS positive, Bodian negative and are located in the cephalic lobe. With the histochemical fluorescence method of Falck, the cells marked by anti-ACTH 17-39 Is exhibit an induced fluorescence, after L-DOPA injection. However, after treatment by metopirone, numerous cells of the caudal part of the adenohypophysis take up L-DOPA but they are not corticotrophs.


Permanent pacemaker implantation in infants, children, and adolescents. Long-term follow-up. Twenty-four patients in the pediatric age range who underwent implantation of a cardiac pacemaker for treatment of complete atrioventricular (A-V) block were followed for an average of five years (range 1-12 years). The etiology of the A-V block was surgical in 13 cases, congenital in nine, and acquired in two. Twenty patients had symptoms of cerebrovascular insufficiency and four had congestive heart failure. To date, 18 of the 24 patients studied are alive and well. Death occurred in six patients, five of whom had complex congenital heart defects, and one of whom had Refsum's disease. Death probably was caused by complete heart block despite pacemaker treatment in four patients, and congestive heart failure in two. In 18 of the 24 children with disabling complete A-V block, pacemaker therapy provided relief of symptoms and prolonged life.


Immunological methods for human placental alkaline phosphatase (Regan isoenzyme). Four different immunological methods for the determination of the placental isoenzyme of alkaline phosphatase (Regan isoenzyme) were compared in 64 normal blood donors, 23 healthy laboratory and medical staff workers and 68 pregnant women: a. Inhibition by soluble antibodies to the placental enzyme. b. Precipitation with soluble antibodies. c. Precipitation with immobilized antibodies. d. Measurement of the activity of the placental alkaline phosphatase following binding to an immunoabsorbent that has been obtained by polymerization of anti-placental-alkaline phosphatase gamma-globulin using glutaraldehyde. The immunoabsorbent method yielded the best results. The optimal conditions were evaluated for the measurement of the activity of immunoabsorbent-fixed placental (tumoral) alkaline phosphatase activity. This method was applied to 209 normal blood donors and to 239 patients with different malignant tumors: 25.5 percent of the cancer patients exhibited an elevated Regan -isoenzyme activity in the serum.


Studies on the inhibition of macrophage migration induced by soluble antigen-antibody complexes. Mixtures of serum of Freund's complete adjuvant (FCA) immunized guinea-pigs and tuberculin PPD consistently inhibited the in vitro migration of peritoneal exudate cells (PEC) of normal guinea-pigs. It is shown that this inhibitory effect is due to a soluble complex between an IgG2 antibody and PPD. By separation of PPD on Sephadex G-200 two peaks were obtained, corresponding respectively to molecular weights of at least 800,000 and 25,000, separated by a plateau. Material derived from both peaks and from the plateau was able to form inhibitory complexes with anti-PPD IgG2. When a mixture of small molecular weight PPD and anti-PPD IgG2 was fractionated on Sephadex G-200, the inhibitory activity was recovered in the void region only. The detection of both IgG2 and PPD in the latter was taken as evidence for the presence of a high molecular weight antigen-antibody complex. When the mechanism of complex-induced inhibition of migration was examined it was found that: (1) complexes act directly on macrophages present in the peritoneal exudate; (2) removal of the Fc fragment of IgG2 by pepsin abolishes its ability to form migration inhibitory complexes; (3) passive sensitization of macrophages with anti-PPD IgG2, followed by exposure to PPD does not result in inhibition of migration; (4) in order to obtain migration inhibition, the complexes must be present during the entire migration period. A 2-hr pulse with complexes does not induce permanent inhibition; (5) the migration inhibitory activity of antigen--antibody complexes can be abolished by certain concentrations of puromycin and aminophylline.


Bacterial blocking activity of specific IgG in chronic Pseudomonas aeruginosa infection. A factor has been demonstrated in the serum of some patients with chronic Gram-negative infections which specifically blocks bactericidal activity against the infecting organism. Sera with this factor, that had been obtained from patients suffering from chronic Pseudomonas aeruginosa infections, were fractionated by Sephadex G-200 and DEAE-Sephadex column chromatography. Blocking activity was detected in the Sephadex G-200 '7S' peak and eluted from the DEAE column along with the serum IgG. Immunoelectrophoresis studies of this material along with pure IgG showed that the bactericidal blocking factor in these patients was IgG. The blocking factor was specific in its ability to protect bacteria, and could be absorbed from the serum by the particular bacterial strain isolated from the patient. Possible clinical importance of blocking activity by IgG is suggested by the persistent nature of the Ps. aeruginosa infections in these patients.


Surface antigenic specificities of human thymus-derived lymphocytes. Surface antigenic specificities of human thymus-derived (T) lymphocytes were studied by cytotoxicity tests using a heterologous rabbit anti-human thymus serum. This serum showed higher cytotoxic titres on thymocytes by comparison with peripheral lymphocytes. After proper absorption the antiserum was non-toxic for chronic lymphatic leukaemia cells, but lysed the majority of thymocytes. It also lysed some of peripheral lymphocytes, corresponding to those lymphocytes which bound sheep erythrocytes (E) but not erythrocyte-antibody-complement complexes (EAC). Pretreatment of lymphocytes with the absorbed antiserum and complement completely abrogated rosette formation with E but spared EAC-binding lymphocytes. It also eliminated their reactivity to phytohaemagglutinin and concanavalin A. These findings indicate that the absorbed serum causes selective lysis of T cells. The results obtained from quantitative absorption studies suggest that a certain loss of T-cell antigens is brought about during the differentiation of thymocytes into peripheral T cells.


Binding of 125I-labelled encephalitogenic basic protein to normal lymphocytes. Inhibition with multiple sclerosis serum. A sheep lymphocyte 125I-labelled human encephalitogenic basic protein (125I-BP) binding assay was used to study the correlation of 125I-BP binding with the diagnosis and status of multiple sclerosis (MS) patients. MS serum generally caused significant inhibition of binding, and serum from MS patients in remission caused significantly more inhibition than serum from MS patients in relapse. Serum inhibition increased in association with improvement in neurological status in three out of four MS patients studied for more than 10 months.


Characteristics of the leucocyte-aggregation assay for cell-mediated immunity. The leucocyte aggregation assay detects cell-mediated immunity by the specific adherence of sensitized lymphocytes onto target cell monolayers. Leucocyte aggregates appear to develop by instruction of non-immune cells by sensitized T lymphocytes. B cells may function in a secondary capacity to amplify aggregate formation. The reactions are sensitive to low temperatures and to metabolic inhibitors of protein and RNA synthesis and function. Serum factors alternatively enhanced or blocked the phenomenon, depending upon the immune status of the patient. This assay may prove useful for the dissection of allograft rejection and tumor resistance due to its brevity, reflection of T-cell immunity, and sensitivity to host humoral factors.


E and EAC rosetting lymphocytes in patients with carcinoma of bronchus. II. A sequential study of thirty patients: effect of BCG. T and B lymphocytes have been estimated in thirty patients with carcinoma of bronchus at monthly intervals. The patients have been treated by surgery, radiotherapy or no conventional antineoplastic therapy, with or without monthly intradermal BCG. Significant increase in T-lymphocyte percentages occurred in three out of six patients treated by BCG and no conventional therapy, and similar increases were observed in other BCG-treated patients. Patterns emerged suggesting that the T-lymphocyte percentage may be depressed, directly or indirectly, by tumour and recover after surgery or palliative radiotherapy; that high T-lymphocyte levels may accompany reactive lymphocytosis to tumour and that T-lymphocyte collapse was associated with, and appeared to precede terminal clinical deterioration in many patients.


Quantification of IgG subclasses in sera of normal adults and healthy children between 4 and 12 years of age. The concentration of the four subclasses of IgG was determined in sera of normal adults and healthy children between 4 and 12 years of age, using the radial immunodiffusion technique. A relation between the concentration of IgG subclasses and Gm type was studied in adults. No influence of Gm type on IgG1 concentration could be shown, except that the group of Gm(fb) individuals had a higher level than the others. The mean concentration of IgG2 was higher in sera positive for Gm(n) than in those lacking this genetic marker. High IgG3 concentrations corresponded to the presence of Gm(b). No clearcut evidence was obtained for a relation between IgG4 concentration and Gm factors, although in general Gm(n) positive individuals had higher and Gm (zag) positive individuals lower concentrations of this subclass in their serum. Quantification of IgG subclasses in sera from healthy children of different ages revealed that the amount of IgG2 rises slowly with age, having not yet reached the adult level at the age of 12 years. This also holds for IgG4, although in a lesser degree. No significant differences from the adult level were found for the concentrations of IgG1 and IgG3.


[Jaundice after aprindin (author's transl)]. Viral-hepatitis-like hepatic damage ("aprindin hepatitis") developed in a 50-year-old woman after taking aprindin, the cause of the jaundice being confirmed by the same signs occurring when the drug was taken again after initial withdrawal.


Cheyne-Stokes respiration and sleep: a diurnal polygraphic study. Thirty-four adult patients with Cheyne-Stokes respiration provided 47 routine EEG records and 24 polygraphic afternoon records. These records were analysed and compared with those of 18 Pickwickian patients previously studied. The main points of difference between these two groups of patients were: the Cheyne-Stokes patients were older and their respiratory periodicity (RP) was not strictly sleep-linked; it could be observed in wakefulness as well as sleep (in that case opening and closing the eyes often did not interrupt the ongoing RP); it tended to disappear in REMS. The main points of resemblance were the frequent appearance of Cheyne-Stokes RP at sleep onset and the presence of occasional ineffective respiratory movements during the crescendo phase of RP in sleep.


Sex-dependent differences in the biosynthesis of pyrimidine nucleotides in rat liver after repeated administration of alpha-hexachlorocyclohexane. Utilization of [2-14C]orotic acid for the synthesis of the pyrimidine components of the free nucleotide pool and RNA from isolated cytoplasmic ribosomes was investigated in the liver of female and male rats. In females, labeled orotic acid is incorporated relatively more into uridine than into cytidine nucleotides; the opposite is true for males. The administration of alpha-hexachlorocyclohexane (alpha-HCH) decreases the specific radioactivity of rRNA cytidylic acid more in males than in females. Simultaneously, the level of liver microsomal cytochrome P-450 is increased after the administration of alpha-HCH more in males than in females. After four days of treatment with alpha-HCH, the concentrations of the uridine and cytidine components of the acid-soluble pool are slightly depressed. Following repeated administration of alpha-HCH, the utilization of labeled uridine for the synthesis of cytidine nucleotides of the acid-soluble pool and RNA is depressed, whereas that of labeled cytidine is enhanced. Repeated administration of alpha-HCH decreases the utilization of [2-14C]orotic acid for the synthesis of cytidine nucleotides of DNA; the specific radioactivity of thymidylic acid is increased. The administration of beta-diethylaminoethyl diallylacetate (CFT-1201) to animals that have repeatedly received alpha-HCH decreases the specific radioactivity of DNA thymine.


Estrogenic inhibition of the hepatic synthesis of alpha2u globulin in the rat. The hepatic synthesis of the androgen-dependent urinary protein in the rat, called alpha2u globulin, is strongly inhibited by estrogens. In mature male rats, treatment with estradiol-17beta (0.5 mug/g body weight) completely inhibits alpha2u synthesis within 6-7 days. Following withdrawal of estrogen treatment alpha 2u synthesis is not reinitiated for approximately 20 days. Parabiotic joining of estrogen-suppressed male rats with their normal littermates within this lag period fails to change the preparabiotic pattern of alpha2u synthesis in the respective partners. Besides estradiol-17beta, other estrane derivatives such as estrone, estriol and estradiol-17alpha were also found to inhibit the synthesis of alpha2u globulin. All of the above estrane derivatives which inhibit alpha2u synthesis are also found to inhibit the uptake of 5alpha-dihydrotestosterone by the hepatic cytosol androgen binding protein of the mature male rat. Unlike cycloheximide, a known translational inhibitor, estradiol-17beta does not inhibit alpha 2u synthesis in the perfused rat liver.


Identification of C-cells in normal and goitrous rat thyroid tissues using antiserum to rat thyrocalcitonin and the immunoperoxidase bridge technique. Application of the immunoperoxidase bridge technique to the light microscopic localization of C-cells in rat thyroid tissue is described. Guinea pig antisera to rat thyrocalcitonin (TCT) were produced by the injection of highly purified rat TCT (100-300 MRC U/mg) emulsified in complete Freund's adjuvant. A 1:1000 dilution of the antiserum used in this study gave a strong positive reaction with rat C-cells, and 1 ml of undiluted antiserum provided sufficient material for staining approximately 5000 slides. The substitution of nonimmune guinea pig serum for the anti-rat TCT serum or the prior absorption of anti-rat TCT serum with increasing amounts of highly purified rat TCT both eliminated the staining of thyroid C-cells. Likewise, no staining was observed in tissue sections from rat parathyroid, ovary, pituitary gland, and skeletal muscle. Antiserum to synthetic human TCT also could be used to identify rat thyroid C-cells. The method revealed abundant C-cells in goiters from rats fed a low-iodine diet for more than 1 year. This finding was supported by electron microscopic evaluation of goitrous tissue and by the detection, by radioimmunoassay, of TCT in thyroid tissue and in peripheral blood from goitrous rats.


Purification of form AI and AII DNA-dependent RNA polymerases from rat-liver nucleoli using low-ionic-strength extraction conditions. Recent findings have confirmed the role of form A DNA-dependent polymerase activity as that which is responsible for the transcription of the ribosomal RNA-coding genes. Unfortunately, the form A enzymes have proved to be very labile and difficult to work with, especially under high ionic strength conditions. We have, therefore, investigated a method for the purification of the form AI and AII enzymes from rat liver using mild low-ionic-strength conditions. Since preparations from whole nuclei were found to be grossly contaminated with protein having similar properties, the enzymes are extracted from nucleoli. Forms AI and AII are separated on a phosphocellulose column, purified by further ion-exchange chromatography, and by sedimentation through a glycerol gradient. The purified enzymes each migrate as a single band on native polyacrylamide gels and have the expected characteristics of form A RNA polymerase. Sedimentation rates through glycerol gradients indicate that they both have a similar size to that of Escherichia coli RNA polymerase (Mr about 500,000). The purified enzymes are free of DNase and RNase. A method is also described for the purification of form B from the nucleoplasm remaining after isolation of nucleoli. The presence of form C activity was not detected.


RNA-dependent DNA polymerase from a cell line derived from the bone marrow of a patient with polycythemia vera. A RNA-dependent DNA polymerase was isolated from a human cell line derived from the bone marrow of a patient with polycythemia vera. The purification procedure included chromatography on phosphocellulose and oligo(dT)-cellulose, and glycerol gradient centrifugation. The enzyme could be distinguished from polymerase A by salt elution from phosphocellulose, utilization of poly(rC) - oligo(dG) and its molecular size of about 70000, as determined by centrifugation. Throughout the purification procedure ribonuclease H activity was co-purified. Upon dodecylsulfate-polyacrylamide electrophoresis on microgradient gels two main bands with molecular weights of 68000 and 66000 and three minor bands were detected. The enzyme preferentially used poly(rA) - oligo(dT) as template-primer compared with poly(dA) - oligo(dT). It incorporated dGMP into polymer on poly(rC) - oligo(dG).


Antigenic structure of the cyanogen bromide peptide F-CB3 from fibrinogen alpha-chain. The antigenic properties of the cyanogen bromide peptide F-CB3 from bovine fibrinogen alpha-chain were studied in radioimmune assays with rabbit antibodies to fibrinogen or to peptide F-CB3. Both fibrinogen and peptide F-CB3 were indistinguishable in inhibition and dissociation tests. Modification of the single disulfide bridge in peptide F-CB3 either by reduction or by cleavage with cyanide was not accompanied by loss of serologic activity. Inhibition studies with three individual fragments obtained after cyanide cleavage (molecular weight range 7000 to 23000) indicated the presence of at least three distinct antigenic determinants in peptide F-CB3. After trypsin digestion of peptide F-CB3 still 75% of its maximal inhibiting capacity was retained. Lack of change in antigenic activity of peptide F-CB3 after release from the fibrinogen molecule by cyanogen bromide and upon further fragmentation is presumably due to the presence of several sequential antigenic determinants but the presence of conformational determinants could not be entirely excluded. Since no cross-reaction was observed between bovine and human peptides F-B3 one may expect considerable variation in their amino acid sequence.


Rôle of axonal transport in maintaining central synaptic connections. Synaptic boutons on the somata of rat hypoglossal neurones have been stained with Zinc Iodide-Osmium, and counted using light microscopy. Two weeks after Silastic cuffs containing colchicine or vinblastine were applied to the hypoglossal nerve, the number of boutons on parent nerve cell bodies was significantly lower than controls. These drugs inhibited retrograde axonal transport of horseradish peroxidase, but did not affect neuromuscular conduction or the number of axons in the nerve trunk. Cuffs containing lignocaine did not affect boutons or axonal transport significantly.


Evidence for animal origin of cells and rescued tumour virus from a putative human osteosarcoma cell line. Human osteosarcoma cell lines T 1 and ZT 1 were analyzed for host origin. The results indicated that these lines must have been contaminated with rat x mouse hybrid cells. The isolated virus was identified as an avian sarcoma virus belonging to the C subgroup.


Allograft tolerance as in immunologically active process: analysis in a restricted H-2 incompatibility due to mutation (M504) in mouse strain B10.D2. Cell-free spleen and liver extracts were used for attempted induction of allograft tolerance across a restricted H-2 barrier due to a chemically induced mutation in the B10.D2 mouse strain. When using mice of the latter strain as donors and the new congenic strain carrying the mutant H-2 hyaplotype as recipients, and with long-term administration of the cell-free antigens in adult life up to 100% yields (according to the dose of antigen) of permanently tolerant animals (particularly males) was obtained with spleen extract, but not with liver extract. This contrasts with a higher tolerance-inducing efficiency of liver extracts in various non-H-2 incompatibilities. In the reversed donor-recipient combination, neither spleen nor liver extracts gave positive results. The immune status of the permanently tolerant animals was analyzed and it was shown that with their bone marrow normal responsiveness can be conferred upon lethally irradiated syngeneic hosts whereas the specific allograft tolerance can be adoptively transferred to sublethally irradiated recipients by means of spleen or lymph node cells. It is concluded that in the present system the state of allograft tolerance is maintained by an immunologically active process of peripheral lymphocytes.


Hepatitis B immune serum globulin in prevention of hepatitis B among hospital personnel. Preliminary report from a controlled trial. At Sahlgren Hospital, Göteborg, hepatitis B has been a serious problem for several years among the personnel in the hemodialysis and renal transplantation wards as well as in the chemical laboratories. For this reason a randomized controlled clinical study with conventional gammaglobulin and hepatitis B immune globulin was started in May 1973. The titer of anitbodies against HBsAg by passive hemagglutination was 1:100 and approximately 1:350,000, respectively. Newly employed staff without a history of jaundice and without demonstratable HBsAg (RIA) or HBsAb (RIP) in serum were enrolled in the trial. Assignment of the two preparations was performed by random numbers and the ampoules were labelled only with the serial number of the participant. Three cc of either preparation was administered i.m. within seven days after admission and was repeated three months later. After that no further injections were given. Each participating employee was followed up with liver function tests, IEOP, RIA and RIP tests for at least six months after the last injection. Preliminary results from this double-blind study, which is still going on, are presented.


The use of hepatitis B immunoglobulin in the Netherlands. Hepatitis B immunoglobulin (HBIg) was administered to 241 patients contaminated with HBAg positive material; 116 persons were followed up for 7 months after the HBIg injection. Only 4 (3.4%) cases of hepatitis B with jaundice and demonstrable HBAg occurred and 15 (12.9%) cases of subclinical hepatitis B were observed. HBIg is well tolerated. Passively transferred HBAb were demonstrable for 2-3 months and no chronic carriers of HBAg were seen after administration of HBIg.


Specific lymphocyte sensitization in chronic pancreatitis. Specific lymphocyte sensitization was found in vitro using a lymphocyte transformation test with human pancreatic antigens. It should be possible to develop an immunological diagnostic test for chronic pancreatitis.


Serum proteins in the rat after extensive proximal or distal small bowel resection. Rats underwent surgical removal of either the proximal or distal half of the small bowel. 5-6 weeks later, total protein concentrations and electrophoretic patterns were determined in sera from the two groups of resected animals. The data compared to similar measurements from sham-operated and unoperated control groups, showed no differences between resected or control animals. This indicates re-establishment of overall nutritional adequacy by this time interval after substantial loss of either proximal or distal small intestine.


[Staining of blood smears with rivanol in patients suffering from bronchogenic neoplasms]. Blood smears of patients suffering from bronchiogenic lung cancer were stained with Rivanol and observed by means of the fluorescence microscope. The results were compared with those obtained by FEULGEN's method. It was found that a single intravenous infusion of cyclophosphamide 30 mg/kg did not change the picture of Rivanol-coloured nuclei which is not always true of the results obtained by FEULGEN's method.


Sister chromatid exchange in xeroderma pigmentosum cells that are defective in DNA excision repair or post-replication repair. The formation of sister chromatid exchanges has been postulated to depend upon the action of DNA repair enzymes. Our experiments with various human cell lines show that the yield of sister chromatid exchanges is within normal limits in both excision-repair-defective and post-replication-repair-defective cells from the autosomal recessive disease, xeroderma pigmentosum. These results indicate that hypotheses invoking known DNA repair processes to acconnt for the recombination of sister chromatids are inadequate and that the exact enzymatic processes are as yet unknown.


[Cytochemistry of leukocytes in children. IV. The use of cytochemical tests in the differentiation and prognostic evaluation of acute leukemias]. From 63 children with acute leukaemia the bone-marrow smears were cytochemically examined before the beginning of therapy. The activity of peroxydase was examined according to Sato and Sekya, that of acid phosphatase according to Löffler and Berghoff, that of alpha-naphthyl-acetate-esterase according to Gomori; the evidence of glycogen was examined by means of the PAS-diastase response according to McManus. Among the 63 cases of leukaemia we found 6 cases of paramyeloblastic leukaemia, 2 cases of parapromyelocytic leukaemia, and 3 cases of myelomonocytic leukaemia. 52 cases of leukaemia could not be further differentiated in morphological respect. They represented an immature paraleukoblastic leukaemia. A division according to leading cytochemical criteria was made for them. The therapeutic possibility of influencing the various groups was checked by means of prolonged observations. Children affected with paraleukoblastic leukaemia of the phosphatase type had a significantly low rate of remission similar to the myeloid leukaemia. Paraleukoblastic leukaemia of the PAS type, esterase type and the undifferentiated type revealed no essential differences. The rate of remission, however, was highest in leukaemia of the PAS type amounting to 100%. In one part of patients the prolonged cytochemical observations in 8 children with recidives showed that the cytochemical type under chemotherapy was changed.


Some immunological aspects of the long-term remission in acute lymphoblastic leukemia in children. Nineteen children aged from 3 to 16 years with first long-term hematologic remission were studied. Cell-mediated immunity was assayed by the reaction to dinitrochlorobenzene (DNCB), nitroblue tetrazolium reduction test (NBT) and lysozyme activity. Humoral immunity was determined by immunoglobulin (IgG, IgA, IgM) levels in serum. The DNCB-reaction was positive in 50% of children treated from 12 to 36 months. Percentage and absolute counts of NBT-positive granulocytes and lysozyme/granulocyte ratio systematically increased with time of therapy. In all studied children IgG was normal, IgA and IgM were depressed to 40% and 76% of normal value (with the exception of 2 children after cessation of therapy, in which IgA was normal). It seemed that continuous control of the immunologic status during long-term cytostatic therapy is essential in clinical practice.


A note on the demonstration of nuclear acidic HCl insoluble proteins in peripheral blood smears. The present study deals with the conditions necessary for the specific demonstration of nuclear acidic HCl insoluble proteins is smeared peripheral leucocytes by means of a simple cytochemical procedures. These proteins can be very easily selectively demonstrated by staining with Azure C or Toluidine blue above pH 6, after the fixation of smears with methanol, extraction of histones, nucleic acids (with HCl, TCA) and treatment with formaldehyde. The double fixation with methanol and formaldehyde facilitated specific extraction of nuclear acidic HCl insoluble proteins of smeared leucocytes with pepsin. The nuclear acidic HCl insoluble proteins are present in leucocytes in interchromatin areas and their isoelectric point determined by cytochemical procedure is pH 6.


[The estrogen dependence of the pregnancy associated alpha glycoprotein "SP3" and its occurrence in sera of patients with various malignant and benign diseases]. The pregnancy associated alpha-2-glycoprotein "SP3" is a macroprotein; the serum concentration of which is markedly enhanced by the increase of endogenous and exogenous estrogen. As regards the bronchial and uterine cancer the frequent presence of enhanced SP3 serum concentrations in tumour patients can be explained by an enhanced production of endogenous estrogen. This enhanced production has been described in literature. SP3 does not occur in healthy men and women without application of estrogen. In 26% of the cases it can be proved in cases of benign disease and in 52% in malignant cases having received no therapy of estrogen.


[The india ink phagocytosis by monocytes and granulocytes during acute and chronic consumption coagulopathy]. During generalized intravascular coagulation (GIC) the polimerisation products of fibrinogen are phagocytized by the reticulo-endothelial system of the liver and spleen. Thus an inhibition of the clearance of RES could be detected in a progressed stage of the shock and GIC; this can also be represented by a decrease of India ink storage. In the present work the authors investigate the behavior of monocytes and granulocytes in the peripheral blood in GIC. Approximately 80% of cells will usually phagocytize and concentrate India ink granules, if heparinized blood mixed with India ink is incubated for two hours at 37 degrees C. A decrease of the India ink phagocytosis was detected in 6 patients and in 10 dogs during the shock. This observation corresponds to the behavior of hepatolienal RES during this condition.


Measurement of blood volume in the Indian mud eel Amphipnous cuchia (Ham.) (amphipnoidae; pisces). The blood volume of 10 male and 8 female Amphipnous cuchia (Ham.) were measured through dilution technique method using Evans blue and sodium citrate as anticoagulant. This method was established against heparin. The mean total blood volume and the blood volume per 100 g of body weight for all the weight groups in male were 9.73 +/- 2.45 and 3.10 +/- 0.66, and in female were 10.34 +/- 2.78 and 3.13 +/- 0.66 respectively. The total blood volume of male and female fishes increase as an exponential function of its total body weight.


Haemoglobin levels according to age, haematocrit, transferrin and serum iron level in medical students of comparable socio-economic status. 1. The haemoglobin, haematocrit, transferrin and serum iron of a group of 418 healthy medical students, aging from 18 till 25 years, were determined. The mean Hb for the male students was 9.4 +/- 0.63 mmol/l, for the female students 8.7 +/- 0.56 mmol/l. 2. The correlations between Hb, Ht, SeFe and transferrin have been investigated. A positive correlation exists between Hb and SeFe and Ht and SeFe.


[Effect of ampholines on blood coagulation: 1. Activation of factor VIII (antihemophilic globulin A)]. Polyaminopolycarboxyl acids (Ampholine, LKB producers Sweden) caused the antihaemophilic globulin (factor VIII) to be activated. This increase of activity revealed a linear dependence on Ampholine concentrations. Only those Ampholine molecules of alkaline pH areas positively charged under test conditions turned out to be efficient. The positive charge, therefore, seems to be significant for an activation. The coagulation activities were measured by determining the partial thromboplastin time (PTT) and the factor VIII in an one-phase test.


[Potency determination of horse anti-human-lymphocyte serum (ALS) by means of the lymphocyte resistance test, a modification of Schroder's leukocyte resistance test]. The leukocyte resistance test of Schröder was used for the purpose of modifying the in vitro determination of cytotoxic potency of ALS during the course of immunization. In contrast to the original method, where the test was carried out with whole blood, the incubation was made with pure lymphocyte cell suspensions. The results obtained show that the extensive procedure of examination enables the cytotoxic effectiveness of antilymphocytic antibodies to be evaluated.


[The question of thrombocyte substitution as a prophylactic measure in splenectomy for idiopathic thrombocytopenic purpura (ITP)]. The problem of intraoperative thrombocyte transfusion in splenectomy of patients with ITP is discussed. The indication for splenectomy cannot be equalized with that for intra-operative platelet substitution. Thrombocyte kinetic examinations with a concurrent determination of the thrombocyte turnover enable those patients to be recognized by their bleeding tendency who are particularly endangered by operations. The intraoperative platelet substitution should be limited to those patients with ITP whose turnover is markedly lowered as a manifestation of reduced thrombocytopoiesis. In these cases it is advisable to shift splenectomy to a later date.


[The use of anti-AHP as a test reagent on blood group documentation cards]. In principle it is possible to use anti-AHP upon bedside test cards for the purpose of blood group documentation. In accordance with the individual quality of the anti-AHP substance an optimal concentration has to be determined and retained. With regard to the rapidity of responses and their strength the anti-AHP involves advantages towards the human anti-A test serum. However, the reliability of reading the results will be slightly limited by the appearance of very fine pseudoagglutinates towards O and B blood corpuscles.


[Cytological and biochemical studies in a case of "di Guglielmo's syndrome"]. Cytochemical, biochemical and disc-elektrophoretical studies are done on a patient with Di Guglielmo-syndrom. Cytochemical tests showed the wellknown results as: activity of acid phosphatase and alpha-naphthylacetatesterase at their typical sites, deposition of PAS- and Turnbull-positive material. Biochemical findings are done on LDH, cholinesterase, alkaline and acid phosphatase, G-6-PDH and pyruvatkinase. Abnormal high level of fetal hemoglobin was found only once during the whole observation. There was also an alteration in the isoenzym pattern of the acid phosphatase. The de- or increase at the biochemical examinations, the fact of fetal hemoglobin and the variation at the isoenzymes may be due to the changing in distribution of erythropoetic and granulopoetic cells.


[Cytochemical studies in megakaryocytes in hematologic diseases]. Bone marrow smears of 48 patients consisting of 12 normal cases, 36 patients with different haematological diseases-among them 9 cases of idiopathic thrombopenia, 4 cases of polycythaemia, and 9 cases of Hodgkin's disease - were examined cytochemically. Acid phosphatase, unspecific esterases, naphthol-AS-D-chloroacetate esterase, peroxydase, and leucin-aminopeptidase were represented; in addition the PAS reaction, fastgreen staining at pH 1.1, methyl-green pyronin staining and the lipid representation with Sudan black B were carried out. Besides those responses known from literature the different behaviour of acid megacaryocyte phosphatase in different haematological diseases must be particularly emphasized from all reactions.


[Problems of cytochemistry of leukocytes]. In investigating the atypical granula in myeloic cells cytochemically the positivity of DMAB reaction identifies the specific eosinophilic nature of these granula. The slightly modified methods of representing histones allow those histones which are rich of lysine and arginine to be differentiated. With their help those differences can be grasped which exist between the condensed chromatin of lymphocytes and the mature neutrophils caused by the presence of histones being rich of lysine and arginine. New possibilities of examining the proteins in leukocytes in a cytochemical way as well as in blood- and bone-marrow cells are provided by colouring with palatine fast black, the use of which also points to certain differences in the chromatin of lymphocytes and neutrophils.


[Cytochemical data in chronic monocytic leukemia]. 4 cases of chronic monocytic leukemia were observed during several years. In 2 patients there was a final appearance of blastic crisis. During the course of the disease the activity of the naphthylacetate-esterase in the monocytes increased, but the PAS-reaction was lowered. By the help of two cases for comparison - one patient suffering from a chronic myeloid leukemia and intermediate monocytic phase, the other one from a panmyelopathia and monocytic reaction - the morphological resemblance of these phenomena is demonstrated, which cytochemically cannot be separated.


[Cytochemical studies on blood and bone marrow leukocytes in various age groups]. The present paper refers to statistically ensured normal values of cytochemical findings in leukocytes of the peripheral blood, of the leukocytes concentrate and the bone-marrow which were obtained in determining the activity of peroxydase, alkaline and acid phosphatase as well as the content of polysaccharides and lipids in 814 healthy persons of various age and sex. The data were calculated according to the formulae of Astaldi and Verga [2].


[Cytochemical tests on sediment cells of malignant and benign exudates]. In exsudate cells separated from serous body cavities of 29 tumour patients and 30 patients with inflammatory and congestive effusion in cardiac failure or liver cirrhosis respectively the activities of acid and alkaline phosphatase were determined. In addition to sudanophilia the cell content of glycogen and that of ribonucleinic acid were evaluated. By means of cytochemical findings it could be found that an increase of unspecific esterase, acid phosphatase and ribonucleic acid in atypical cells points to a malignous ethiology of the exudate.


[Ultracytochemical studies on leukemic cells]. The present survey of ultracytochemical investigations in the cells of the Graffi and Rauscher leukaemic systems is intended to throw a light on the place of replication and on the incorporation of cell specific material into the virus. The successful evidence of incorporating certain enzymes associated to the membrane into the virus envelope could have a significance in so far as this process can be used to explain the appearance of cytotropism in certain kinds of virus to a target cell.


[Studies on the lipid composition in serum and lymphocytes in chronic leukemia]. In 33 patients with chronic lymphatic leukaemia (CLL) the lipids were examined quantitatively and qualitatively in the serum and in the lymphocytes. Besides the 5 main categories of lipids, lysolecithin, sphingomyelin, lecithin and cephalin as well as cholesterol ester subfractions were determined according to their degree of saturation of their fatty acids. Patients suffering from CLL were found to have a moderately raise of serum triglycerides and an increase of cholesterol esters with simple unsaturated fatty acids. Simultaneously the fraction of ester with poly-unsaturated fatty acids is lowered. Leukaemic lymphocytes reveal in general a lowered total lipid content. It mainly consists of phosphatides and free cholesterol. Moreover, the leukaemic lymphocytes contrary to normal ones reveal a lowered content of cholesterol esters, diglycerides and triglycerides.


[Cyclophosphamide in the treatment of immune hemocytopenias]. Personal experiences with the immunosuppressive therapy of immunohaemocytopenia by cyclophosphamide are represented. A significant therapeutic success in autoimmunohaemolytic anaemia could be registered in 22 patients treated from one to seven years. From a number of 11 patients 9 were in complete remission. In idiopathic thrombocytopenia, however, only 7 patients from a number of 10 were in a clinical remission, the number of thrombocytes remained unchanged. The control of the complement content and the component C'3 can be used well as an indicator for immunological defense when the activity of the process and the success of the therapy are to be evaluated.


Respiratory function in iron deficiency anaemia before and after treatment. The present studies concern the basic ventilatory indices, the arterial blood gases, the indices of the acid-base balance and the indices of the alveolar-capillary diffusion (DLCO, SS, Dm, theta Vc) in anemia before and after treatment. Substantial changes are recorded after treatment mainly in DLCO, SS and theta Vc. These changes are predominantly due to changes in the concentration of Hb. An evaluation of both methods for the standardization of DLCO at 14,6 g Hb/100 ml blood is also presented.


The influence of magnesium ions on pyruvate kinase-deficient red blood cells. The effect of intravenous and oral administration of magnesium containing drug (Magnesium Diasporal, Protina, München) on the erythrocyte pyruvate kinase (PK) activity in two cases of congenital nonspherocytic hemolytic anemia associated with PK deficiency has been examined. Intravenous injection of the drug has been followed by transient increase in magnesium content. Positive correlation between erythrocyte magnesium ions concentration and PK activity has been found. The authors propose a hypothesis that the observed correlation may be due to the erythrocyte accumulation of some PK activators or to the elimination of some PK inhibitors. The authors did not, however, succeed to maintain the high erythrocyte magnesium concentration inducing the increase in PK activity during the longer period of time.


Species specificity of thromboplastin. A phylogenetic study. Having studied the influence of thromboplastin preparations derived from cold and warm-blooded species on the plasma of 12 vertebrates (carp, frog, turtle, hen, rabbit, rat, mouse, guinea-pig, ground squirrel, dog, sheep and man) we have established that among the species far from each other phylogenetically, the phenomenon of species specificity could be demonstrated in general. It was found that the extrinsic coagulation system measured by Quick times can well be activated in every examined species. Simultaneously, in plasmas of turtle and hen the intrinsic clotting system proved to be deficient.


The preparation of the factors IX, II, and X concentrate by means of DEAE cellulos and its therapeutic effectiveness. Starting from a plasma used for producing cryoprecipitate a concentrate was gained by means of adsorption on DEAE cellulose, the factor IX activity of which was six to thirteen times higher than that of the original plasma. Even the factors X and II are enriched to the same extent approximately. In patients with haemophilia B the concentrate proved to be very efficient and compatible during surgical operations.


[Experiences with polychemotherapy of acute leukemias in adults with special reference to the COAP combination]. The authors give a report on their experiences with polychemotherapy in 77 acute leukaemias in adults. On this occasion the COAP combination was identified  to be well effective and to be an enrichment of therapy because of its low side effects.


[Problems in maintenance therapy in acute myeloid leukemias in adults]. Problems of maintaining therapy for acute myelocytic leukemias in adults are discussed. The analysis of the maintaining therapy in 22 patients affected with an acute myelocytic leukemia and living for more than 6 months revealed that the interval therapy with a high dosage of cytostatic combinations in the sense of the COAP scheme is preferable compared with the daily administration of 6-mercaptopurin, in addition methotrexate twice a week. Reasons for this are discussed.


Semiquantitative cytochemical determination of acid phosphatase, beta-glucuronidase, and beta-glucosaminidase activity in peripheral blood lymphocytes. Activity of acid phosphatase, beta-glucuronidase, and beta-glucosaminidase in peripheral blood lymphocytes was determined cytochemically in 20 normal subjects, 10 male and 10 female, by the use of BARKA and ANDERSON's (1962), HAYASHI et al. (1964) and HAYASHI's (1965) methods, respectively. Results obtained were semiquantitatively according to subdivision of lymphocytes into enzyme-negative and enzyme-positive cells. Enzyme-positive lymphocytes were divided into cells with granular, mixed granular and diffuse enzymatic reaction type. In the first two types of cytochemical reaction a number of enzyme-positive lysosomal granules were counted and expressed in terms of both absolute count and percentage of circulating lymphocytes. Enzyme-positive lymphocytes represented 80.3%, 40.5% and 41.5% of the total lymphocyte count in regard to the presence of acid phosphatase, beta-glucuronidase, and beta-glucosaminidase, respectively.


["Hairy cell" leukemia].

["Hairy cell" leukemia]. By means of individual observations the clinical, cytological and cytochemical marks of the so-called "hairy cell"leukemia are described. Peculiarities in the course of the disease and in prognosis justify this rare variant to be separated from reticular systems diseases. A diagnosis can also be made on the basis of cytomorphological characteristics.


[Results of splenectomy in patients with hypoplastic and aplastic anemia]. The results of spleen exstirpation in 129 patients with hypoplastic and aplastic anaemia are presented. For this purpose the role of splenectomy in connection with blood formation is dealt with on the basis of analysing morphological blood findings. The impact of the duration of the disease, of sex, age, and the dosage of corticosteroids on the postoperative development of the disease is demonstrated. The following observations of the patients which were made within 3 to 14 years after the operation revealed that 63 patients from 129 survived, namely 21 patients up to 5 years, 26 patients up to 10 years, 16 patients up to 14 years. 66 patients died, the highest lethality being observed during the first year after the operation (47 patients).


[Experiences with artificial immunization for the production of anti-HL-A sera]. In 17 cases positive cytotoxic anti-HL-A antibodies were gained from 26 immunizated pairs. On the whole this corresponds to the data to be found in literature [10, 17]. The antibodies against the antigens of the I. locus will appear more rapidly, younger persons will form antibodies more easily. The difference in one single antigen results in favourable findings when the pairs are composed; in spite of that, however, cross-reacting antibodies may frequently be obtained, particularly when boostered to produce sera with a high titre. No side effects were detected in the immunizated group; yet there is a certain reason to suppose that the production of anti-HL-A sera by artificial immunization cannot be compared with the usual production of AB0 sera. The immunopathological significance of the HL-A-system is still elucidated insufficiently; by no means it is confined to histocompatibility. Only those institutes should be concerned with artificial immunization which are able to perform the necessary immunological controls exactly in each case.


[The effect of heating and drugs on HL-A antigens and their reactions with cytotoxins]. In some instances the heating of lymphocytes for 2 to 3 minutes at 56 degrees C enhances HL-A antigens or makes it possible to detect these antigens by a twostage microlymphocytotoxic test on preheated lymphocytes only. Similar phenomenons are observed in some tannin (1: 40 000) or 0.1% phenol-treated lymphocytes and after the addition of these solutions during the first stage of the lymphocytotoxic test. Prolonged heating at 56 degrees C leads to nonspecific polyreactivity of lymphocytes, giving false results with all sera. For similar reasons also higher concentrations of tannin and phenol solutions were found dissatisfactory for the pretreatment of lymphocytes. The pretreatment of lymphocytes with 36-0.5% formaline induces full inhibition of specific cytotoxic reactivity of HL-A antigens and their absorption ability with regards to respective HL-A sera. The addition of formaline at 0.06-0.3% concentration during the first and second stages of the test and at the end of the second stage (or simultaneously with eosine) gives also negative results of the cytotoxic test owing to the inhibitory effect of formaline on rabbit complement and HL-A antigens.


Separation of human antihemophilic factor (AHF, factor VIII) from fibrinogen by means of defibrase. Various attempts have been made in the past to remove fibrinogen from plasma fractions rich in factor VIII. Present studies indicated the Defibrase, a preparation of a thrombin-like enzyme from the venom of Bothrops atrox, might be successfully employed for this purpose. Addition of Defibrase to plasma or low-purity factor VIII concentrates allows for the complete removal of fibrinogen without any apparent reduction in the activity. It was also shown that treatment with Defibrase neither decreases the stability of factor VIII nor apparently affects its antigens properties.


Bovine fibrinogen-induced 14C-serotonin and other compounds release from human blood platelets. Bovine fibrinogen can aggregate human blood platelets by a direct mechanism (first wave) and also indirectly by releasing endogenous ADP (second wave). A primarily non-altered surface structure of the thrombocytes is important for the induction of the first wave. Under certain conditions a second aggregation wave, concomitant with the release and the availability reaction is elicited. Besides the cell's contact, the presence of ADP and calcium ions are necessary for the induction of the release reaction in human platelet rich plasma. The changes induced by bovine fibrinogen thus follows the same pattern as the irreversible phase of aggregation, linked to release as induced by ADP.


[Hemolytic transfusion accident due to the blood group antibody anti-Jka]. A haemolytic transfusion incident caused by anti-Jka is reported about. In a 56 year old patient suffering from chronic iron deficiency anaemia an immunization in the Kidd system has taken place within 9 days. In the course of this immunization the fourth incompatible blood transfusion caused a severe transfusion incident. The necessity of serological compatibility testing by the indirect antiglobulin or enzyme test and the significance of making a strict indication for blood transfusions are referred to.


[The effect of stabilizer composition on thrombocytes and thrombocyte function in stored whole blood. I. The thrombocyte count, thrombocyte aggregation and retraction during storage]. The behaviour of thrombocyte number and thrombocyte function aggregation and retraction in ACD, AcD-A and AcD-AG stabilized blood was examined in 18 apparently healthy test persons for a period of 9 days. On the one hand the addition of adenin or guanosin respectively increased the thrombocyte aggregation, on the other hand, however, a decrease of free, haemostatically efficient thrombocytes could be observed. Under the test conditions chosen retraction does not allow any statement to be made on the degree of the storage damage.


[The effect of stabilizer composition on the thrombocytes and thrombocyte function in stored whole blood. II. Results of the thrombocyte spreading test]. The behaviour of thrombocyte spreading in ACD, AcD-A and AcD-AG stabilized blood was examined in 18 apparently healthy test persons for a storage period of 9 days. Due to an improved energy metabolism of thrombocytes the addition of adenin or guanosin respectively in ACD stabilized blood will cause the thrombocyte spreading to be widely preserved during storage.


In vitro survival and amethopterin effects on the structure of ovarian tissues of the sawfly, Acantholyda nemoralis Thoms. Whole ovaries of the sawfly, Acantholyda nemoralis Thoms., Tenthredinidae, Hymenoptera, were cultivated by the organ culture technique of Fell in the medium of Jones and Cunningham modified by doubling the contents of salts, sugars and lactalbumin hydrolysate and supplemented with an addition of folate. In the experimental conditions the sawfly ovary survived for several days showing mitoses in the follicular epithelium. In some of the nurse cell nuclei of the cultured ovaries nucleolus-like structures developed which showed a positive reaction to the Hamm test for viral polyhedra. Formation of these structures was stimulated by amethopterin. Electron microscopical examination revealed that these structures were not viral polyhedra but enlarged nucleoli whose modified ultrastructure differed considerably from that of the nucleoli of cells of trophic ovaries in normally developing insects. Amethopterin appears to induce enlargement of the granules of the granular component of nucleolonema, probably due to disturbance in RNA biosynthesis. The positive Hamm test obtained in the nucleoli shows that this test cannot be regarded as specific for nuclear polyhedra.


Increase in Feulgen staining intensity by pre-treatment with different reagents. In the experiments sections of rat tissues fixed in 10 per cent buffered neutral formalin for 3 hour and treated for 15 minutes with different chemical reagents such as pyridine, tributylamine, urea, tris sodium nitrite and sodium hydroxide were subjected to hydrolysis in 6 N HCl at 25 degrees C for 20 minutes and stained by the UV Feulgen technique. The results reveal a far more intense staining of the nuclei in tissues treated with any of the chemicals mentioned than in untreated controls. The possible role of these chemicals in enhancing the staining intensity is discussed.


Hot hydrochloric acid hydrolysis and UV Feulgen staining of rat liver sections fixed in CRAF and CRAF-like fixative. Sections of rat liver fixed in CRAF III and Nawaschin's fixative in Dutt's modification were subjected to hydrolysis in 1N HCl at 60 degrees C for different periods of time and to Schiff's staining according to the UV Feulgen technique. The study showed that Feulgen reaction intensity depends upon time of hydrolysis, optimum coloration being possible only after 10-15 min of hydrolysis. Prolongation of hydrolysis beyond this time produced decreased staining intensity which is retained for further 35 min of hydrolysis thus forming a plateau. Further prolongation of hydrolysis results in gradual deterioration of the staining intensity which culminates in utterly pale coloration of the nuclei after one hour's hydrolysis. A possible explanation for this phenomena is suggested.


Alteration of the HL-A antigenic site in situ. The chemical nature of the HL-A antigenic sites on peripheral blood lymphocytes was studied by treatment of these cells with glycolytic enzymes and with sodium metaperiodate, and monitoring the residual antigen expression either by the lymphocyte cytotoxicity test or by quantitative microabsorption of monospecific anti-HL-A sera. Neither the gentle enzymatic nor the chemical treatment of lymphocytes altered expression of the HL-A antigens indicating that carbohydrates are not involved, in a major way, in the HL-A antigenic sites.


Isolation of a VHClambda fragment on tryptic digestion of human IgD myeloma proteins. The tryptic digestion pattern of IgD myeloma proteins is reported. The Fab delta of fragment is shown to be very susceptible to further degradation to yield a novel fragment composed of VH and Clambda domains which are still bound through the interchain disulphide bridge. Reduction results in a change in M.W. from ca. 26,000 to 13,000. The loss of Fd delta of isotypic determinants may be of importance to studies in which membrane bound IgD is detected or measured after release from the cell surface by enzyme digestion.


Molecular construction of Clostridium botulinum type A toxins. Two Clostridium botulinum type A toxic fractions, named large (L) and medium (M) toxins, were eluted from Sephadex G-200. Sucrose density gradient centrifugation resolved L toxin (2.5 X 10(8) to 3.0 X 10(8) mean lethal doses per mg of N) into two fractions, 19S and 16S. The same procedure performed at pH 8resolved it into three fractions; the heavier two were both nontoxic and hemagglutinin positive, and the lightest on (7S) was toxic. M toxin (12S) (4.5 X 10(8) to 5.0 X 10(8) mean lethal doses per mg of N) was homogeneous in electrophoresis and centrifugation at pH 6. The latter procedure performed at pH 8 dmonstrated that it dissociated into uniform 7S components. The nontoxic component of M toxin was free from hemagglutinin. M toxin alone was demonstrated in culture by sucrose density gradient centrifugation at pH 6. Dialysis of the culture supernatant resulted in partial formation of 16S toxin. Centrifugation of the crystalline toxin in 1 MNaCl demonstrated 16S toxin only. The toxic components of L, M, and crystalline toxins were antigenically identical. The nontoxic components of the crystalline and L toxins, consisting of two distinct antigens, were antigenically identical; that of M toxin was identical with one of these two antigens.


Vi antigen from Salmonella typhosa and immunity against typhoid fever. 11. Safety and antigenicity in humans. Safety and antigenicity of a purified preparation of Salmonella typhosa Vi antigen was evaluated in human volunteers. Dosages of Vi antigen at 25, 50, and 100 mug were less toxic than U.S. standard typhoid vaccine (lot 6A) containing 5 X 10(8) bacteria per dosage. Vi antigen in comparison with the standard typhoid vaccine induced higher hemagglutinating antibody but lower bactericidal antibody responses.


Antitoxic immunity in experimental cholera: protection, and serum and local antibody responses in rabbits after enteral and parenteral immunization. The protective effect of enternal and parenteral immunization with cholera toxin antigen against experimental cholera in rabbits was studied by using the small-bowel loop technique. Subcutaneous injection of crude toxin as well as purified toxin or toxoids gave rise to significant protection against toxin challenge. The enhanced resistance to toxin was found to correspond to a many-fold higher magnitude of protection against challenge with live vibrios. In the primary response the protection increased during the first month. Booster immunization gave rise to a further increased immunity which, however, declined rapidly. Multiple oral or repeated intraintestinal antigen administrations also induced protective antitoxic immunity although of less magnitude than that obtained by parenteral immunization. Enteral and, to a lesser extent, parenteral immunization gave rise to increased antitoxic antibody titers and immunoglobulin levels in intestinal washings and mucosa scraping. Immunoglobulin G (IgG) and IgG antitoxins predominated, but after enteral immunization total IgA and specific IgA antibodies occasionally reached levels similar to those for IgG. In serum, significantly increased antibody levels (IgG) were only recorded after parenteral immunization. Both the primary binding and the neutralizing antitoxin titers showed a stayistically significant  correlation with the degree of protection against toxin challenge; however, for the neutralizing antibodies this correlation was not without exceptions. No relation to protection was found for intestinal antibodies. The results of the present study indicate that enternal as well as parenteral immunization with toxin antigen can give rise to effective cholera immunity. After enternal immunization, the protection appears to be medicated by locally synthesized antibodies. After parenteral vaccination both serum-derived and locally produced antibodies seem to be effective.


Antigens of Streptococcus mutans: characterization of a polysaccharide antigen from walls of strain GS-5. A cell wall-associated polysaccharide antigen was isolated from Streptococcus mutans GS-5 and appeared to determine serotype c specificity. Ouchterlony double-diffusion analysis of crude formamide extracts derived from purified cell walls of two serotype c strains (GS-5 and JC-2) showed complete identify when reacted with anti-GS-5 sera. Immunoelectrophoresis of this extract demonstrated the typical mobility for this serotype as described by others. Column chromatography on BioGel P-100 of the crude formamide extracts derived from GS-5 walls resulted in a single antigenic peak being resolved. This material, when loaded onto a diethylaminoethylcellulose column and eluted with a linear gradient of ammonium carbonate (0.0 to 0.2 M), was resolved further into two serologically reactive peaks (I and II). Only two consituents, rhamnose and glucose, were detected in the purified column fractions. Peak 1 had a rhamnoseto-glucose molar ratio of 0.9:1.0, and peak II, the major resolvable fraction, had a molar ratio of 1.7:1.0, The peak II ratio was very similar to that found in the formamide extract residue pellet (1.6:1.0)9 Ouchterlony analysis of the crude formamide extract and the purified fractions revealed only partial identify between peaks I and II but complete identify between peak II and the crude extract. Likewise, immunoelectrophoresis showed no differences in mobility of peak II and the crude extract, whereas peak I moved towards the cathode. Possible structural relationships between the two antigenic fractions are discussed below. Hapten inhibition studies suggested that an alpha-glucosyl group is at the immunodeterminant site of the antigen.


Quantitative assay of diphtherial toxin and of immunologically cross-reacting proteins by reversed passive hemagglutination. A reversed passive hemagglutination (RPHA) assay for diptherial toxin has been developed. Antitoxic antibodies were isolated from commercially available equine diptherial antitoxin by immunoabsorption using highly purified diphtherial toxin covalently linked to Sepharose 4B. Formalinized, tanned sheep erythrocytes sensitized with the purified antitoxic antibodies are specifically agglutinated by diphtherial toxin but are not agglutinated by extracellular antigens of Corynebacterium diptheriae that are unrelated to toxin. The RPHA assay described can detect less than 20 pg of diphtherial toxin and is comparable in sensitivity to intracutaneous tests for toxin. The RPHA assay was shown to be at least 1,000 times more sensitive than quantitative immunological assays for diptherial toxin performed by single radial immunodiffusion or by one-dimensional double diffusion in agar gels. Fragment A prepared from purified diphtherial toxin and nontoxic mutant proteins that cross-react immunologically with toxin can be assayed directly by RPHA, but the sensitivity of the assay for these proteins is less than for native diphtherial toxin. Inhibition of RPHA was also shown to be a sensitive quantitative method for measuring diptherial antitoxin in vitro.


Infection in immunodepressed patients. The approach to diagnosis and treatment. Infection is an important cause of death in patients receiving cytostatic drugs or with any other impairment of host resistance. Such infections are frequently due to opportunist micro-organisms usually belonging to the endogenous flora of the patient. It is often difficult to obtain an exact diagnosis of the cause and localization of the infection. The problems associated with the prevention of infection are manifold. Exogenous infections can be prevented by proper isolation and a sterile diet. Endogenous infections can only be prevented by eradication of the patient's endogeous flora, so-called decontamination. Special attention should be given to treatment of foci of chronic infection and of the carrier state of certain microorganisms. However, the prophylactic use of antibiotics should be avoided. The curative use of antibiotics should be based on the most probable micro-organism. We consider the inventory of the patient's microflora, repeated weekly, of great help in the choice of antibiotics in cases of septicaemia of unknown aetiology. The initial therapy usually consists of a broad-spectrum combination of antibiotics, which should be bactericidal. When the causative bacteria have been isolated and the sensitivity is known, antibiotic therapy should be adjusted to the narrowest spectrum possible.


Antigenic properties of polymers formed by beta-lactam antibiotics. Purified polymers have been isolated from 6-aminopenicillanic acid and from semi-synthetic penicillins and cephalosporins. All the polymers were shown to react with rabbit antibody of penicillins and cephalosporins. All the polymers were shown to react with rabbit antibody of penicilloyl specificity, as demonstrated by passive cutaneous anaphylaxis in guinea pigs, and the reactions were shown to be penicilloyl-specific by hapten inhibition experiments. The cephalosporin-derived polymers in addition reacted with rabbit antibodies raised to the corresponding cephalosporin conjugates of bovine gamma-globulin. Using direct skin tests, passive cutaneous anaphylaxis and an in vitro assay, no evidence was obtained that the polymers induced the formation of specific antibodies in baboons, guinea pigs and rabbits, but in baboons the induction of cell-mediated immunity, demonstrable by delayed skin test reactions, was shown.


Preparation of carbon-14-labelled whole short ragweed pollen antigens. A new method of labelling short ragweed pollen antigens with carbon-14 is described. The known ragweed antigens AgE, Ra3, Ra5 and several other antigens are radiolabelled. These components are biologically active, have a high specific radioactivity and a long half-life.


A clinical trial of intravenous bleomycin in the treatment of brain tumors. The author has carried out the administration of bleomycin, chiefly by the intravenous route, in the postoperative adjuvant treatment of 150 cases of brain tumor since January 1969. The overall response rate of gliomas to bleomycin was more than 50 per cent. No distinct, given tendency toward possible differences in bleomycin effectiveness due to the type and location of tumors was revealed, but it appears that there were fewer effective cases among highly malignant gliomas and metastitic brain tumors. The survival data also showed encouraging results.


Prevalence of carcinoma of the penis with special reference to India. The frequency of carcinoma of the penis throughout the world is tabulated and compared with similar data from 32 hospitals in India. There is a wide variation in the frequency of penile carcinoma in India despite the fact that circumcision is not practiced by Hindus and is by Muslims. There must be some other etiologic factors such as poor sexual hygiene and lack of cleanliness to account for this variability in frequency. There was no association between the frequency of cervical and penile carcinomas.


[The geographic distribution of psoriasis]. On the basis of data published in the literature the geographical distribution of psoriasis vulgaris is described. One of the most interesting results is the increase of the psoriasis frequency in countries such as Japan, Korea, Kazachstan and East Africa. This could be due to the fact, that altered life conditions in connection with an increasing industrialization favour the manifestation of psoriasis genes. How far population differences of the psoriasis frequencies are to be seen exclusively as genetical ones, is hitherto unclear, but rather unlikely. No relations between psoriasis and climatec factors could be seen. Associations between psoriasis and several antigens (HL-A 13 and W 17) of the HL-A-systems seem to be present. These associations could be of considerable importance for the interpretation of the geographical distribution pattern of psoriasis. Further studies in psoriatic families are required, which describe the distribution of the HL-A specifities in the possibly potential psoriatic members of the families, who carry the respective genes.


[Phoinatric and audiological aspects of autism (author's transl)]. The diagnosis of autism or autistic traits always necessitates exclusion of a primary or secondary speech or hearing disorder. The following findings are based on experience gained from 12 children with the diagnosis of early infantile autism. If an acoustic or speaking disorder is caused by brain damage, it will increasingly dominate the total symptomatology as the child matures. The more intensively the organic factor influences the picture, the more frequently a primary acoustic or speech disorder is found. This was observed in 6 cases, which can be diagnosed an "pseudo autistic". From the phoniatric and audiological view the type of autism syndrome as described by Kanner has so far not been confirmed.


Ultrastructural study of Mycoplasma pneumoniae in organ culture. The ultrastructure of Mycoplasma pneumoniae M129 was studied by using specialized staining methods for thin-section transmission electron microscopy. Nucleic acid was shown in the cytoplasmic granules and fibrillar material in the nuclear region. The central filament of the highly structured tip contained basic protein. With one method of fixation, parallel filaments were seen in the central core. M. pneumoniae was enveloped in an extracellular mucoprotein layer that was especially concentrated around its terminal structure.


Flagellar hook protein from Salmonella SJ25. From acid-disintegrated flagellar hooks of Salmonella SJ25 an immunochemically pure preparation of hook protein was obtained by column chromatography. The molecular weight of the protein determined by sodium dodecyl sulfate-gel electrophoresis was 43,000, whereas that of SJ25 flagellin was 56,000. The amino-terminal residue of the hook protein was determined to be seryl. The amino acid composition of the protein was determined, the results being very similar to that for an Escheria coli hook protein reported by Silverman and Simon (1972). Within a wavelength range of 200 to 250 nm, our purified preparation of hook protein gave a circular dichroism spectrum with unusually small amplitudes, suggesting that the alpha-helix content of the protein was very low.


Mitotic chromosome condensation in the sperm nucleus during postfertilization maturation division in Urechis eggs. Changes in the morphology of the sperm nucleus in the egg cytoplasm are mong the immediate events in nucleocytoplasmic interactions during early embryogenesis. Soon after its entrance into the egg cytoplasm, the sperm nucleus of various organisms increases in size with the transformation of condensed chromatin to a diffuse state, resembling the chromatin of an interphase nucleus (2, 13, 15, 16). This is followed by a close association or fusion of male and female pronuclei (2, 13, 15, 16). Cytoplasmic influences on nuclear morphology have also been demonstrated clearly in nuclear transplantation and cell fusion studies (10, 11). Reactivation of the nucleus, such as the transplanted brain nucleus in Xenopus egg cytoplasm or the hen erythrocyte nucleus in interphase cytoplasm of HeLa cells, is accompanied by nuclear enlargement and chromatin dispersion (10, 11). However, premature mitotic-like chromosome condensation takes place in the nuclei of sperm or interphase cells fused with mitotic cells (9, 12). Thus, chromosome dispersion and condensation seem to depend on the state of the cytoplasm in which the nucleus is present. These observations imply that the initial morphological changes in the sperm nucleus after fertilization may very well be dependent on the state of maturation of eggs at the time of sperm entry. Unfertilized eggs of Urechis caupo, a marine echiuroid worm, are stored at the diakinesis stage. These eggs complete maturation division after insemination and this is followed by fusion of male and female pronuclei (5, 8). Therefore, Urechis caupo is a suitable organism in which to study the response of the sperm nucleus to the changing state of the egg cytoplasm during and after postfertilization maturation division.


In vitro formation of gap junction vesicles. A method is described that uses trypsin digestion combined with collagenase-hyaluronidase which produces a population of gap junction vesicles. The hexagonal lattice of subunits ("connexons") comprising the gapjunctions appears unaltered by various structural criteria and by buoyant density measurements. The gap junction vesciles are closed by either a single or a double profile of nonjunctional "membrane," which presents a smooth, particle-free fracture face. Horseradish peroxidase and cytochrome c studies have revealed that about 20% of the gap junction vesicles are impermeable to proteins 12,000 daltons or larger. The increased purity of the trypsinized junction preparation suggests that one of the disulfide reduction products of the gap-junction principal protein may be a nonjunctional contaminating peptide. The gap junction appears to be composed of a single 18,000-dalton protein, connexin, which may be reduced to a single 9,000-dalton peak. The number of peptides in this reduced peak are still unknown.


Transformation of cells by rous sarcoma virus: cytoplasmic vacuolization. Chick embryo cells transformed by the Bryan "high titer" strain of Rous sarcoma virus (RSV-BH) are heavily vacuolated. A variety of microscopic techniques have been used demonstrating that the vacuoles are cytoplasmic, bounded by membrane, and are composed largely of water. Proteins, lipids, glycoproteins, glycolipids, glycosaminoglycans, glycogen, and nucleic acids were undetectable in the vacuoles. Physiological requirements for development of the vacuoles, and reversal of vacuolization, were examined in cells infected with a virus mutant, RSV-BH-Ta, which induces reversible temperature-dependent transformation. Na+ was the only component of the cell culture medium found essential for both the development and reversal of vacuoles. Glucose depletion or dinitrophenol treatment inhibited vacuolization, suggesting a possible energy requirement in the vacuolization process. Ouabain, an inhibitor of Na+-K+ ATPase, enhanced vacuolization, but a variety of other substances affecting cell surface components were in active. Two sugars, glucosamine and mannosamine, prevented the disappearance of vacuoles. The observations suggest that cellular vacuolization may be a normal physiological response to an increase in water and Na+, and, in the specific case of transformation by RSV-BH, may be relevant to the physiological basis for malignancy.


Lipid staining for the electron microscope: a new method. Tissues fixed in osmium tetroxide or in combined osmium and glutaraldehyde (Hinde), embedded in Spurr's medium, cut at 0-5-I mum and mounted in Farrants' gum medium containing ethyl gallate, show good staining of lipid-contaning structures (droplets of triglyceride, membranes, mitochondria, etc.) in the light microscope. Such preparations show moderate contrast in the electron microscope without further staining. But a specific increase in contrast in lipid-rich structures is obtained by partition of the tissues, before embedding, in 70% ethanol saturated with the monoterpene hydrocarbon myrcene, with or without the addition of 0-I % ethyl gallate, followed by osmium tetroxide. This method will visualize both saturated and unsaturated lipids, including waxes.


An ultrastructural study of acid phosphatase localization in Phaseolus vulgaris xylem by the use of an azo-dye method. The localization of acid phosphatase during xylem development has been examined in the bean, Phaseolus vulgaris. The azo dye, the final reaction product, is initially prominent in the dictyosomes, vesicles apparently participating in secondary wall formation, and in the middle lamella of the young vessel element. Final reaction particles are also present in mitochondria, chloroplasts, and certain vacuoles and are sparsely scattered in the cytoplasm. At a later stage of vessel differentiation, the azo dye is concentrated in the disintegrating cytoplasm and along the fibrils of the partially hydrolysed primary wall and middle lamella. In the mature vessel element, the azo dye is still present along the disintegrated primary wall at the side of the vessel and covers the secondary wall. In the parenchyma cell adjacent to the vessel element, acid phosphatase localization is found in the dictyosomes, endoplasmic reticulum, mitochondria, small vacuoles, and the middle lamella. The controls from all stages of vessel element development were free of azo dye particles. The concentration of acid phosphatase along the secondary walls of the mature vessels and in the middle lamella between other cells indicates that this enzyme has other functions besides autolysis of the cytoplasm and primary cell wall. Acid phosphatase may participate in the formation of the secondary wall and may also have a role in the secretion and transport of sugars.


Appearances of microtubules after various fixative procedures, and comparison with the appearances of tobacco mosaic virus. Microtubules in crane-fly spermatids appeared altered when the glutaraldehyde-fixed cells were not postfixed with osmium tetroxide. The cytoplasmic microtubules were altered more than the doublet microtubules. Addition of osmium tetroxide after dehydration did not produce appearances identical with those of microtubules postfixed directly after glutaraldehyde, and thus at least some alterations occurred during dehydration, possibly due to extraction of microtubule-associated lipid. The omission of osmium tetroxide postfixation did not cause drastic alterations in the appearances of either tobacco mosaic virus (TMV), or polymerized tobacco mosaic virus protein (without RNA), suggesting that microtubule stability is different from TMV stability (with respect to the embedment procedure). The electron-dense stain associated with embedded-sectioned TMV is predominantly outside the TMV protein, as demonstrated by the known distribution of TMV protein compared with the dimensions of sectioned TMV and negatively stained TMV. The same might hold true for microtubules, as evidenced by the dimensions of negatively stained, isolated brain microtubules compared with those of embedded and sectioned brain microtubules.


Cell surface saccharides of Trypanosoma lewisi. I. Polycation-induced cell agglutination and fine-structure cytochemistry. Trypanosoma lewisi bloodstream and culture forms were agglutinated differentially with low concentrations of the cationic compounds: ruthenium red, ruthenium violet, Alcian blue chloride, 1-hexadecylpyridinium chloride, lanthanum chloride, and cationized ferritin. The bloodstream form trypanosomes gave the highest agglutination levels with each of the compounds tested. Ruthenium red was the most effective inducer of cell agglutination among the several cations used. Trypsin-treated bloodstream forms were agglutinated less in the presence of ruthenium red than untreated controls. Ruthenium red-induced cell agglutination also was lowered with chondroitin sulphate and dextran sulphate, but not with alpha-D-glucose, alpha-D-mannose or with several methyl glycosides. Treatment of the bloodstream trypanosomes with alpha-amylase, dextranase, or neuraminidase had little effect on agglutination levels obtained with ruthenium red. Fine-structure cytochemical staining with ruthenium red, ruthenium violet, and Alcian blue-lanthanum nitrate was used to ascertain the presence and distribution of presumptive carbohydrates in the trypanosome cell surface. The extracellular surface coat of the bloodstream forms stained densely with each of the polycationic dyes. Trypsin treatment removed the surface coat from bloodstream trypanosomes; however, the surface membranes of the organisms were stained densely with the several dyes. Similar surface-membrane staining was obtained with the cationic compounds and the culture forms, which lack a cell surface coat. Cationized ferrin was used at the fine-structure level to visualize the negative surface charge present in the cell surface coat and external membrane of the several trypanosome stages. Results obrained from the agglutination and cytochemistry experiments indicate that complex polysaccharides are present in the surface membranes and cell surface coat of T. lewisi bloodstream forms. Similar conclusions also pertain to the surface membranes of the T. lewisi culture from trypanosomes. The carbohydrates probably represent glycopeptide and glycoprotein structural components of the surface membrane of this organism.


Thyroidal triiodothyronine and thyroxine in Graves' disease: correlation with presurgical treatment, thyroid status, and iodine content. To evaluate the potential contribution of thyroidal secretion to the relative excess of triiodothyronine (T3) production in hyperthyroidism and to investigate the effects of treatment, iodine (127I), T3 and thyroxine (T4) were measured in digests of thyroid tissue obtained at surgery from 13 patients with Graves' disease. In 11 normal human thyroid glands, 127I content was 630 +/- 60 (all values mean +/- SE in mug/ wet weight) T4, 254 +/- 39 and T3 21 +/-3. The T4I was 26 +/- 3% of the total iodine and the molar ratio T4/T3 was 11 +/- 1. The 13 patients with Graves' disease were divided into three groups. Eleven were clinically euthyroid (Groups I and II) and had received either iodide or iodide plus a thiourea derivative before surgery. Two subjects (Group III) received only propranolol. In Group I (n = 8), mean thyroidal 127I content was 320 +/- 50, T4 was 115 +/- 9 and T3 22 +/- 4. The molar ratio T4/T3 was 5.9 +/- 1 and T4I was 26 +/- 2% of the total. Group II patients (3) had the lowest preoperative serum T4 (less than 2.5 mug/dl) and T3 (less than ng/dl) concentrations with TSH elevated in only one (7 muU/ml). Thyroidal 127I was 100 +/- 26, T49 +/- and T3 1.3 +/- 0.3. The % T4I was 5 +/-2. The two chemically hyperthyroid subjects had a mean tissue 127I of 450; T4, 295 and T3, 56, the T4/T3 ratio was 4.5 and % T4I was 42. There was no correlation between tissue 127I and T4/T3 within either the normal or Graves' disease group. Since adequate clinical and chemical control of hyperthyroidism with antithyroid drugs and iodine was attained in the 8 Group I subjects without a decrease in the % T4I or T3I below that of normal thyroids, it suggests that inhibition of iodotyrosine coupling is not required for this effect. The % T4I was below normal only in patients with marked suppression of serum T4 and T3 concentraions. The lack of correlation between tissue 127I and T4/T3 ratio in the treated patients suggests that the lower T4/T3 ratio in Graves' thyroids is independent of intrathyroidal iodine concentrations. This hypothesis is strengthened by the similarly low T4/T3 ratio in untreated subjects with near normal tissue 127I content. Assuming that the thyroid hormones are secreted in the ratios present in these digests, one can estimate that direct secretion by the thyroid could contribute most, of not all, of the excess T3 production in Graves' disease.


Degradation of blood group antigens in human colon ecosystems. I. In vitro production of ABH blood group-degrading enzymes by enteric bacteria. Human feces contain enzymes produced by enteric bacteria that degrade the A, B, and H blood group antigens of gut mucin glycoproteins. We have studied their production in fecal cultures to determine if such cultures can be a source for enzyme purification and to explore how blood group antigen-degrading enzymes are adapted in individual human colon ecosystems. They were present in fecal cultures from each of 27 healthy subjects, including ABH nonsecretors. Heat-sensitive obligate anaerobes are their major source. From 39 to 85% of the total enzyme activity produced by growing cultures was extracellular. Commercial hog gastric mucin and salivary glycoproteins, including Lea saliva which lacks A, B, and H antigens, enhance production of A-, B-, and H-degrading activity in anaerobic fecal cultures irrespective of the glycoprotein's blood group specificity. There is evidence that the host's ABO blood type and secretor status affects the specificity of blood group-degrading enzymes produced by his fecal bacteria in vitro. Thus, fecal inocula from B secretors incubated with hog gastric mucin (A and H specificity) or with Lea saliva produced greater levels of B-degrading than A- or H-degrading activity, and inocula from A secretors in similar media produced greater levels of A-degrading than B- or H-degrading activity. Blood group-degrading enzymes produced in fecal cultures are glycosidases and not proteases. The B-degrading enzyme cleaves the B antigenic determinant alpha-D-galactose from the oligosaccharide side chains of mucin glycoproteins with B specificity. Anaerobic fecal cultures containing blood group substances are a feasible source for purifying blood group antigen-degrading enzymes. Prior adaptation to blood group antigens in the gut mucins of type A and type B secretors affects the specificity of the enzymes produced in vitro.


Degradation of blood group antigens in human colon ecosystems. II. A gene interaction in man that affects the fecal population density of certain enteric bacteria. The autosomal dominant ABH secretor gene together with the ABO blood type gene control the presence and specificity of A, B, and H blood group antigens in human gut mucin glycoproteins. Certain obligate anaerobes in feces produce extracellular antigen-specific glycoside structures. We estimated the populations of these bacteria in feces of 22 healthy subjects by determining the greatest dilution of feces that yielded A, B, or H blood group-degrading enzyme activity after 24 h incubation in anaerobic cultures. Comparatively small populations of fecal bacteria produce blood group-degrading enzymes; their estimated populations were 10(8) per g or less in 21 subjects. Fecal populations of B-degrading bacteria were stable over time, and their population density averaged 50,000-fold greater in blood group B secretros than in other subjects. We present evidence that the greater fecal populations of B-degrading bacteria in B secretors is due in part to a competitive nutritional advantage gained by their ability to enzymatically cleave the B antigenic determinant alpha-D-galactose from gut mucins of B secretors. Fecal populations of bacteria producing A and H antigen-degrading enzyme activities were comparable in all subjects to the fecal population of B-degrading bacteria in B secretors. The large populations of fecal anaerobes may be an additional source of A antigen substrate for A-degrading bacteria; thus, antigens cross-reacting with A antigen were detected on cell walls of anaerobic bacteria from 3 of 10 cultures inoculated with 10(-10) g feces. Bacteria producing B-degrading activity likely represent a separate population from those producing A- or H-degrading activity since their fecal populations differed numerically in 14 subjects. These findings suggest that adaptation of blood group-degrading enzymes to mucin structures in human colon ecosystems is chiefly by mutation-selection of comparatively small populations of constitutive enzyme-producing strains rather than by substrate induced enzyme synthesis in many strains.


Intracellular mucous inclusions. A feature of malignant cells in effusions in the serous cavities, particularly due to carcinoma of the breast. Three cases of carcinoma of the breast are described, in which similar tumour cells were found in pleural or peritoneal fluid. The cells were characterized by the frequent presence of 'bull's-eye' vacuoles, in which a central spot was deeply stained by Giemsa, eosin or periodic acid-Schiff (PAS). In one of these it was shown by electron microscopy that the vacuoles are lined by microvilli, and that their mucinous content is condensed in the central area, presumably as a result of failure to discharge the secretion.


Effect of Bacteroides fragilis on the human erythrocyte membrane: pathogenesis of Tk polyagglutination. Incubation of normal human erythrocytes with the supernatant of centrifuged culture of some strains of Bacteroides fragilis results in the exposure of Tk polyagglutination determinants on the erythrocyte membrane. These determinants are present on non papain-labile structures and are probably exposed by an enzyme mechanism. Other strains of B. fragilis were found to produce neuraminidase, haemolysins, and protease. It is suggested that early recognition of Tk determinants may be an aid to the diagnosis of B. fragilis infection.


A note on the use of immunofluorescent methods for the detection of Pseudomonas aeruginosa in bronchitic sputum. By immunofluorescent microscopy of sputum from 67 cases of bronchitis Pseudomonas aeruginosa was detectable in 14 as compared with nine by cultural methods.