Characteristics of the T-dependent alpha(1 leads to 6) glucosyl (dextran) antibody response induced in mice with isomaltohexaose coupled to chicken gamma-globulin.

Isomaltohexaose flavazole coupled to chicken gamma-globulin (IM6-CGG) induced T cell-dependent anti-alpha(1 leads to 6) dextran-specific IgM and IgG responses in CBA, BALB/c and A strain mice. The IgG responses were of restricted heterogeneity as judged by isoelectric focusing, and belonged mostly to the IgG1 subclass with a minor IgG3 component in the case of BALB/c and CBA mice. All four subclasses of IgG were produced in A strain mice. In contrast, native dextran B 512 induces exclusively T cell-independent IgM responses of the same specificity for all 3 strains. All BALB/c mice immunized with different doses of IM6-CGG either in Freund's adjuvant or with Al(OH)3 plus Bordetella pertussis showed the same spectrotypes by isoelectric focusing. Sera obtained at different times after immunization of BALB/c mice failed to show spectrotype variations. Late, but not early, bleedings from CBA mice showed a tendency towards more uniform isoelectric focusing patterns.


Substance P infusion into substantia nigra of the rat: behavioural analysis and involvement of striatal dopamine.

The purpose of the present study was to characterize the nature of the behavioural response to substance P (SP) infusion into the substantia nigra and to evaluate this response in 6-hydroxydopamine (6-OHDA) caudate lesioned rats. The effects of SP infusions (3 microgram in 1 microliter bilaterally) were assessed in an open field. Two groups of rats were used: one with 6-OHDA lesions in the caudate nucleus, and one with sham lesions. In sham rats, the first infusion produced a strong increase in sterotyped rearing and sniffing, with no concurrent enhancement of locomotion. With three subsequent infusions (made at interval of two days) the rearing response disappeared and a tendency to groom emerged. All SP-induced behavioural stimulation was blocked in the caudate-lesioned rats; an effect of the lesion itself was reduced rearing. These results suggest that the response to SP infusion is mediated through the nigro-striatal dopamine system. The behavioural profiles which emerge after SP infusion into the substantia nigra and ventral tegmental area are compared. In general, the behavioural studies of SP effects support the concept that the A9 and A10 dopamine systems can be behaviourally differentiated.


Aprotinin induces surface changes in malignant cells in culture: a possible mode of antitumour action.

Treatment of TRES cells with aprotinin (Trasylol) produced marked changes in their surface properties. There was a concentration-dependent increase in the surface charge density of the cells. The agglutinability of the cells was inhibited by the antiproteinase. The cells became less adhesive to concanavalin A-linked plates. The adhesion of treated TRES cells appeared to be mediated by lectin receptors with lower specificity for concanavalin A. These observations and an analysis of the kinetics of adhesion have suggested the possibility that a new class of concanavalin A receptors appears on the surface of aprotinin-treated TRES cells, which might increase immunogenicity of the cells.


Differential location of different types of intermediate-sized filaments in various tissues of the chicken embryo.

The location of constitutive proteins of different types of intermediate-sized (about 10 mm) filaments (cytokeratin, vimentin, desmin, brain filament protein) was examined in various tissues of 11--20 day chick embryos, using specific antibodies against the isolated proteins and immunofluorescence microscopy on frozen sections and on isolated serous membrane. The tissues studied which contained epithelia were small intestine, gizzard, esophagus, crop, liver, kidney, thymus, mesenteries, and epidermis. The results show that the different intermediate filament proteins, as seen in the same organ, are characteristic of specific lines of differentiation: Cytokeratin filaments are restricted to--and specific for--epithelial cells; vimentin filaments are seen--at this stage of embryogenesis--only in mesenchymal cells, including connective tissue, endothelial and blood cells, and chondrocytes; filaments containing protein(s) related to the subunit protein prepared from gizzard 10 nm filaments (i.e., desmin) are significant only in muscle cells; and intermediate filament protein of brain, most probably neurofilament protein, is present only in nerve cells. We conclude that for most tissues the expression of filaments of cytokeratin, vimentin, desmin, and neurofilament protein is mutually exclusive, and that these protein structurees provide useful markers for histochemical and cytochemical differentiation of cells of epithelial, mesenchymal, myogenic, and neurogenic differentiation.


Identification and characterization of epithelial cells in mammalian tissues by immunofluorescence microscopy using antibodies to prekeratin.

The occurrence of intermediate-sized filaments containing prekeratin-like proteins ('cytokeratins') has been examined in various organs of rat and cow by electron microscopy and by immunofluorescence microscopy on frozen sections using antibodies to defined constitutive proteins of various types of intermediate-sized filaments (prekeratin, vimentin, desmin). Positive cytokeratin reaction and tonofilament-like structures have been observed in the following epithelia: epidermis; ductal, secretory, and myoepithelial cells of sweat glands; mammary gland duct; myoepithelial cells of lactating mammary gland; milk secreting cells of cow; ductal, secretory, and myoepithelial cells of various salivary glands; tongue mucosa; bile duct; excretory duct of pancreas; intestinal mucosa; urothelium; trachea; bronchi; thymus reticulum, including Hassall corpuscles; mesothelium; uterus; and ciliated cells of oviduct. None of the epithelial cells mentioned has shown significant reaction with antibodies to vimentin, the major component of the type of intermediate-sized filaments predominant in mesenchymal cells. The widespread, if not general occurrence of cytokeratin filaments in epithelial cells is emphasized, and it is proposed to use this specific structure as a criterion for true epithelial character or origin.


Carcinoembryonic proteins in gastric carcinoma metastatic to the liver.

Two cases of liver metastases from gastric carcinoma are described in which the simultaneous occurrences of alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA) and carcinoplacental alkaline phosphatase (CPALP) were demonstrated in the sera and tumor tissues. AFP was detected not only in the tumor tissues but also in the liver tissue adjacent to the tumor, while the other 2 carcinoembryonic proteins were not detected in the non cancerous liver tissues. The characteristics of CPALP in Case 1 were almost similar to the Nagao isoenzyme, based on enzyme tests involving L-leucine, L-phenylalanine and EDTA inhibitions, heat-stability and Michaelis constant, except for electrophoretical slower moving, while that in Case 2 were identical to variant type CPALP (Warnock).


Topographical distribution and association of tumor--associated antigens and their role in antigenicity in rat tumor cells.

The antigenicity of tumor-associated antigen (TAA) was compared between 3-methylcholanthrene-induced rat fibrosarcoma KMT-17 and Friend murine leukemia virus-infected KMT-17 (FV-KMT-17) cells. The antigenicity was classified into immunosensitivity and immunogenicity, and studied from the viewpoint of humoral immunity. The immunosensitivity to anti TAA antibodies increased by artificial infection of KMT-17 cells with Friend virus. The results suggested that an increase of lateral mobility of TAA on the cell surface contributes to the increase of immunosensitivity. Concerning the immunogenicity of TAA, an augmentation of anti-TAA antibody formation was demonstrated when FV-KMT-17 cells were used as the immunogen. In this case, a physical association between TAA and virus-associated antigen (VAA) was suggested to be very important to increase the immunogenicity of TAA.


Effect of modification of reactive lysine on antitryptic and antichymotryptic activity of proteinase inhibitor from cow colostrum.

40% of the primary structure of the cow colostrum proteinase inhibitor (CTI) is homologous with the structure of the trypsin kallikrein inhibitor (TKI) from bovine organs; the positions of the reactive lysine residues are also the same in both inhibitors. Both CTI and TKI were modified by carbamoylation and the fully labeled derivatives were isolated by ion-exchange chromatography. The effect of the modification on the antitryptic and antichymotryptic activity of both inhibitors was investigated. The antichymotryptic activity of both inhibitors is not decreased after the modification. The antitryptic activity of modified TKI is retained, yet the dissociation constant of the complex of the modified inhibitor with trypsin is considerably increased; nevertheless, modified TKI is a good trypsin inhibitor. The antitryptic activity of modified CTI is hardly detectable. We explain this difference in the behaviour of both inhibitors by a replacement of basic residues Arg-17 and Arg-39 in TKI by neutral amino acids Ala-20 and Gln-42 in CTI.


Protease inhibitors in porcine serum and their immunological relationships to human protease inhibitors.

A close molecular relationship exists between the protease inhibitors of porcine serum and those of human serum as shown by studying their immunological cross-reactivities with gel diffusion and immunoelectrophoretic methods. On studying seven different antisera to human protease inhibitors, five were found to cross-react with porcine serum, and on this bisis it was possible to identify alpha 2 -macroglobulin f, alpha 2 -macroglobulin s, alpha 1 -protease inhibitor, inter-alpha-trypsin inhibitor, antithrombin and alpha 2 -antiplasmin in porcine serum. Antisera to four of these porcine serum inhibitors (alpha 2 -macroglobulin f, alpha 2 -macroglobulin s, alpha 1 -protease inhibitor and inter-alpha-trypsin inhibitor) were produced and were shown to react immunologically with their human serum protease inhibitor counterparts.


Identification and characterization of trypsin, chymotrypsin and elastase inhibitors in porcine serum.

Characterization of the trypsin-, chymotrypsin- and elastase-inhibiting properties of porcine serum was carried out by gel filtration on Ultrogel, AcA 44, and agarose gel electrophoresis with subsequent processing for protease-inhibiting activity. Moreover, by allowing the fractions obtained from gel filtration to react with antibodies to porcine serum protease inhibitors, the specific inhibiting properties of these inhibitor molecules were identified. At least six protease inhibitors were identified and partially characterized in porcine serum. Two alpha 2 -macroglobulins (alpha 2 Mf and alpha 2 Ms), homologues to human alpha 2 -macroglobulin, with slightly different electrophoretic mobilities, were both found to exhibit trypsin, chymotrypsin and elastase inhibiting activity. Alpha 1 -Protease inhibitor (Mr 51 000), a homologue to human alpha 1 -protease inhibitor (alpha 1 -antitrypsin), also showed trypsin-, chymotrypsin- and elastase-inhibiting properties. Inter-alpha-trypsin inhibitor (Mr 162 000 and 129000), a porcine serum counterpart to human inter-alpha-trypsin inhibitor, showed trypsin- and chymo-trypsin-inhibiting properties. In addition, a specific trypsin inhibitor, alpha 2 -antigrypsin (Mr 58 000), and a specific elastase inhibitor, beta-elastase inhibitor, were characterized in porcine serum, and these seem to have no counterparts in human serum.


[Purification and characterization of the free secretory piece of human colostrum (author's transl)].

The free secretory piece is isolated from human colostrum by gel filtration and ion-exchange chromatography in high yield (200 mg/l colostrum). DEAE-Cellulose chromatography separates the free secretory piece in two fractions which are electrophoretically distinct, but otherwise have the same characteristics, like molecular weight, antigenic determinants, N-terminal sequence, peptide map and amino acid composition. It was therefore concluded that the protein part of the secretory piece is homogenous.


Histochemistry in the diagnosis of early prostatic carcinoma.

Twenty-seven prostatic adenocarcinomas, most of them well differentiated, were examined for acid mucoproteins by the Kreyberg and alcian blue-periodic acid-Schiff methods. Seventeen (63 per cent) showed varying amounts of positive material in the lumens of the malignant glands. The procedures can be performed on routinely fixed and paraffin blocked tissues, and are useful in confirming the presence of malignant disease in equivocal cases in which crush artifact or small amounts of very well differentiated tumor acini are present.


The application of immunoperoxidase methodology for the visualization of prolactin binding sites in human prostate tissue.

In previous studies the immunoperoxidase method was used to detect intracellular prolactin binding sites in epithelial cells of normal and neoplastic rat prostate. As an extension of this work, the same approach was used to test for and to localize prolactin binding sites in autopsy and biopsy specimens of formalin fixed, paraffin embedded human prostate. Rehydrated tissue sections were exposed to varying concentrations of human placental lactogen or human prolactin and then to human placental lactogen or human prolactin antisera. The loci of hormone binding were visualized by an immunoperoxidase staining sequence. Hormone pretreatment produced immunospecific, dose related staining inside epithelial cells of normal, hyperplastic, and neoplastic human prostate indicative of intracellular prolactin binding. The patterns of intracellular hormone binding in human prostate were similar to those seen previously in rat prostate tissue. The possible involvement of prolactin in prostatic cancer and the potential diagnostic value of the immunoperoxidase approach to hormone binding are discussed.


Serum and cellular biologic tumor markers in patients with urologic cancer.

During the past several years the development of radioimmunoassay and immunocytochemical techniques to detect small amounts of marker in the sera and cancer cells of cancer patients has made a significant impact on the diagnosis and management of certain cancers. Among these markers alpha-fetoprotein, human chorionic gonadotropin, and pregnancy specific beta-1 glycoprotein have been useful in the staging, detection of recurrence, prognosis, and management of testicular cancer. In this article the recent developments and future perspectives concerning these and other newer markers are discussed.


MA 160 and EB33 cell lines: HeLa cell contaminants, hybrids or prostatic epithelial cells?

Studies of acid phosphates produced by cell lines MA 160 and EB 33 demonstrated immunochemically their prostatic origin. MA 160 and EB 33, rather than being HeLa contaminants, may be hybrids of prostatic epithelial and HeLa cells or true prostatic cell lines with chromosomal changes common to all long-term cultivated cell lines.


Surface-associated host proteins on virulent Treponema pallidum.

A surface coat of host serum proteins was detected on virulent Treponema pallidum by sodium dodecyl sulfate-gel electrophoresis. The loosely associated serum proteins could be removed by repeated washings in a protein-free medium. Washed T. pallidum retained the ability to readsorb numerous host proteins from rabbit serum as well as iodinated rabbit or human albumin. In addition, various avidly associated host serum proteins including albumin, alpha(2)-macroglobulin, transferrin, ceruloplasmin, immunoglobulin G, immunoglobulin M, and C3 were identified on the outer envelope of washed treponemes by an immunoadsorbent technique with protein A-bearing staphylococcus. Hyaluronidase treatment did not remove the avidly associated host proteins from the surface of washed treponemes, whereas trypsin treatment resulted in decreased levels of agglutination. Electrophoretic patterns of trypsin-treated treponemes showed that treponemal proteins as well as adsorbed host proteins were released concurrently by protease digestion. Reacquisition studies involving alpha(2)-macroglobulin and transferrin suggested the presence of noncompetitive binding sites for serum proteins on the treponemal outer envelope. Finally, differences among the T. pallidum preparations from individual rabbits with respect to incorporation of [(35)S]methionine, extent of agglutination with antisera, and length of time required for removal of avidly associated host proteins by trypsin treatment indicated biological variability among the treponemal populations.


Antigenic heterogeneity of the non-serogroup antigen structure of Neisseria gonorrhoeae lipopolysaccharides.

Studies of the antigenic structure of the polysaccharide component of gonococcal lipopolysaccaride (LPS) indicated that the non-serogroup antigen structure is antigenically heterogeneous. Immunodiffusion studies of Gc4 strain 8551 indicated that in addition to the Gc serogroup determinent, this polysaccharide contains two other sets of determinants, one which is shared with the other five Gc serogroups and a second which is shared by the Gc1 and Gc3 serogroups. Enzyme-linked immunosorbent assay (ELISA) studies confirmed the immunodiffusion, documented qualitative differences, and suggested that quantitative differences may exist in the common determinant from serogroup to serogroup. As measured by ELISA inhibition, native LPS showed the same antigenic arrangement as the purified LPS polysaccharides. Studies with LPS-derived polysaccharides from nonprototype strains in immunodiffusion gave results identical to the prototype strains. This antigen variation, although distinct from serogroup specificity, was related to the serogroup determinant.


Generation of specfic T-cell suppressor function induced by Streptococcus mutans in monkeys and mice.

Specific suppressor cells in mice and monkeys can be induced with high doses of Streptococcus mutans antigen. When these cells are further cultured with a low dose of S. mutans antigen in vitro, they secrete specific suppressor factors which decrease the cooperative responses to this antigen. In view of the sensitivity of these cells to anti-Thy 1 and complement, and as they are not retained on nylon-wool, these suppressor cells are T cells. The suppressor factor exerts its effect on T helper cells and not on B cells. Suppressor cells and factors may regulate antibody responses to S. mutans and might be of significance in determining the dose and frequency of immunization and the type of adjuvant to be used.


Alpha-fetoprotein and gamma-glutamyltranspeptidase in mice. Effect of Raf gene.

The regulation of the high physiological level of serum alpha-fetoprotein (AFP) in BALB/c/J mice was studied. Serum AFP concentration in 7- to 12-week-old BALB/c/J mice varied from 460 ng/ml up to 2,790 ng/ml and in other strains tested from 10 ng/ml to 400 ng/ml. In contrast to the difference in serum AFP level, activity of the fetal enzyme gamma-glutamyltranspeptidase (GGT) in the liver of BALB/c/J mice was similar to that found in other mice. In all newborn mice, the initially high GGT activity decreased almost 100-fold during the first 10 days. The serum AFP concentration stayed relatively stable during this period. AFP decreased to the adult level between the 10th and 30th days. Partial hepatectomy and administration of carbon tetrachloride increased the serum AFP level in all mice. The elevation of AFP was 10 times higher in BALB/c/J mice than in mice with low basal AFP level. A slightly elevated GGT activity in the liver was observed in only 10 out of 40 mice after carbon tetrachloride treatment, but none of the mice after partial hepatectomy showed an elevation. These results suggest that the gene ("Raf" gene) controlling the reduced decrease of serum AFP level in young BALB/c/J mice enhances the "turning on" of production of AFP in the regenerating liver. This gene does not, however, control the expression of the other carcinofetal liver marker, GGT.


Epstein-Barr virus-specific antibody-dependent cellular cytotoxicity in patients with Burkitt's lymphoma.

Coded sera from 54 patients with African Burkitt's lymphoma (BL) were titrated for antibodies against an Epstein-Barr virus (EBV)-induced membrane antigen in the antibody-dependent cellular cytotoxicity (ADCC) assay. The titers were then correlated with the progression of lymphoma growth following chemotherapy. In 74% of the patients with high ADCC titers (greater than 3,840), lymphomas showed partial or complete regression following therapy. In the medium-titered group (240-3,840), 36% of the lymphomas showed some response to therapy, while only 29% of the lymphomas in the low group (less than 240) responded to treatment. These preliminary results indicated that, as previously reported for patients with nasopharyngeal carcinoma, ADCC titers may be a prognotic value in patients with this EBV-associated disease. In an attempt to determine the identity of the ADCC antigen, some of these sera were examined for antibody to the four major MA components so far identified in the membrane of EBV-infected Raji cells. Sera with high ADCC titers in general contained antibody to the four major MA components, while low-titered sera usually contained antibody to three or less of these proteins. There were exceptions to this pattern, however, indicating that the ADCC antigen might differ from the four EBV-induced membrane components so far identified.


Hepatitis B virus antigens in human primary hepatocellular carcinoma tissues.

The presence of hepatitis B virus (HBV) antigens was examined in specimens of liver tissue obtained at necropsy from black Senegalese patients suffering from primary hepatocellular carcinoma (PHC). The results were correlated with markers of hepatitis B infection in serum. Hepatitis B surface antigen (HBsAg) and core antigen (HBcAg) were sought for in 15 liver extracts. HBsAg was found in the liver in 10 of 12 cases with HBsAg-positive serum. HBcAg was detected in three livers. The HBsAg was detected in seven of eight livers by immunofluorescence and orcein staining. HBsAg-positive cells were mainly located in the peri-tumoral cirrhotic tissue, although positive hepatocytes were also found in tumour nodules in liver from one of the patients. HBcAg was found in five of seven cases by  immunofluorescence in hepatocytes of the cirrhotic areas. HBcAg fluorescence was primarily nuclear but, in some lobules, a patchy cytoplasmic fluorescence was observed. This suggests a cytoplasm-nucleus pathway in the synthesis of the HBV core antigen. Electron microscopy was performed on two HBsAg- and HBcAg-positive cases. Fibrillar and crystalline cytoplasmic inclusions were observed in tumour cells. In the same cells, 20-25 nm virus-like particles were present in swollen cisternae of the endoplasmic reticulum.


In vivo induction of endogenous retroviruses in BALB/c mouse hepatocytes by successive treatments with carbon tetrachloride and bromodeoxyuridine.

The effects of bromodeoxyuridine (BrdUrd) incorporation into hepatocytes of carbon tetrachloride (CCl4)-induced regenerating liver of BALB/c mice on the induction of endogenous retroviruses was examined. From the nucliec acid hybridization studies, the maximum levels of hybridization were obtained for both N- and X-tropic BALB/c endogenous retrovirus specific [3H]-cDNAs with liver RNA from animals receiving BrdUrd at 40 and 44 h post-CCl4 treatment, and killed on the fourth day following BrdUrd injection. Media from NIH-3T3 (Swiss mouse) and mink cell cultures, infected with liver homogenates from animals treated as above, gave significant levels of reverse transcriptase activity. The observations made in the present study show that BrdUrd incorporation into cell DNA can cause induction of both N- and X-tropic endogenous retroviruses in BALB/c mouse hepatocytes in vivo, and such induction is probably a transient event.


Isolation of a SV40-like Papovavirus from a human glioblastoma.

A human glioblastoma multiforme (M27) tested in early cell cultures by indirect immunofluorescence staining showed SV40-related tumor (T)-antigen, 95% of the cells being positive. SV40-related viral capsid (V)-antigen was absent in all cells tested. Experiments to rescue this virus were performed by fusing M27 cells with CV-I monkey cells, which were permissive for SV40, using polyethylene glycol (PEG) as fusion factor. We succeeded in isolating virus particles SV40-GBM which electron microscopy showed to correspond in size and morphology to papovaviruses. Serological tests (hemagglutination, neutralization, fluorescent antibody) revealed that the virus is indistinguishable from SV40. Despite this apparent antigenic identity SV40-GBM differs slightly from SV40 wild type. This virus can propagate and produce CPE in both CV-I cells and primary fetal human kidney cells. Furthermore digestion of SV40-GBM DNA with the HindII/III restriction endonucleases revealed minor differences compared with the SV40 DNA. Therefore the virus SV40-GBM obtained from glioblastoma cells seems to be closely related to the SV40-PML viruses described earlier.


The localization of Hodgkin's disease in lymph nodes. A study with immunohistological, enzyme histochemical and rosetting techniques on frozen sections.

Lymphoid tissue of 51 patients with Hodgkin's disease was studied with immunohistological, enzyme histochemical and rosetting techniques for the detection of B and T cells in frozen sections. In lymph nodes of patients with lymphocyte predominance type of Hodgkin's disease, the majority of the lymphocytes in the involved areas were normal B lymphocytes of polyclonal origin. This was also true for nodular sclerosis cases with a predominance of lymphocytes. Surrounding Sternberg-Reed cells small clusters of T lymphocytes could be demonstrated. In mixed cellularity and also in nodular sclerosis with a mixed cellular pattern only small residual areas of B lymphocytes were present, whereas relatively large numbers of T lymphocytes were found in the involved areas. In lymphocyte depletion B lymphocytes were scarce and T lymphocytes were present in small number. It is concluded that different patterns of lymphocyte population can be discerned in the subtypes of Hodgkin's disease. A predominance of B lymphocytes is found in cases with lymphocyte predominance and thus is a prognostic favourable sign. A predominance of T lymphocytes as found in cases with a mixed cellular pattern with or without nodular sclerosis is therefore not a favourable sign in general but may indicate progressive disease. The possible reasons for the presence of large numbers of B or T lymphocytes in lymphoid tissue affected by Hodgkin's disease are discussed.


Rauscher leukemia virus-induced tumor antigens: complete separation from gp70, p30 and H-2.

The possibility that some or all of the viral proteins, gp70, p30, and the histocompatibility antigen, H-2, function as the tumor-specific transplantation antigen (TSTA) of the R-MuLV-induced leukemia, RBL-5, and also in the secondary in vitro induction of cytotoxic T lymphocytes (CTL), was investigated. The antigen was obtained by isolating the plasma membranes of RBL-5 cells and solubilizing with sodium deoxycholate (DOC) followed by gel filtration chromatography. A fraction containing excellent tumor-rejection activity but low amounts of gp70, p30 and H-2 was chromatographed on goat anti-gp 70 goat anti-p 30 and sheep anti-H-2b immunoaffinity columns. The data obtained indicate that gp 70, p 30 or H-2 do not function as TSTA of RBL-5 leukemia, individually or as a complex. Similarly, the antigen responsible for the specific secondary induction of CTL in vitro is distinct from these three proteins.


Abortive expression of the Epstein-Barr virus (EBV) cycle in a variety of EBV DNA-containing cell lines, as reflected by nucleic acid hybridization in situ.

A variety of Epstein-Barr virus (EBV) DNA-containing cell lines have been tested for the expression of the EBV-associated antigens EBNA (nuclear antigen), EA (early antigen), and VCA (viral capsid antigen), and for the presence of cells containing disproportionate amounts of EBV DNA. The antigen tests utilized immunofluorescence and 125I-labelled antibodies combined with autoradiography. EBV-DNA was detected by in situ hybridization with 3H-labelled EBV RNA complementary to P3HR-1 EBV DNA (P-EBVcRNA). The P-EBVcRNA has been shown to represent the majority of the P3HR-1 EBV DNA sequences. It was concluded that EBV DNA-containing cell lines can be divided into those that express only EBNA, those that express EBNA and EA and those that express EBNA, EA and VCA and also contain cells that undergo disproportionate EBV DNA synthesis. Consequently, in some cell lines there is an abortive expression of the EBV cycle in that some cells spontaneously express EA but fail to continue further to viral DNA synthesis. A similar pattern can be found after experimental induction of the EBV cycle, suggesting that related mechanisms govern the spontaneous expression of the EBV cycle and the extent of its inducibility.


Characteristics and genetic control of NK-cell-mediated cytotoxicity activated by naturally acquired infection in the mouse.

Most and perhaps all natural-cell-mediated cytotoxicity (NCMC) may be activated by a response to exogenous infections. Specific-pathogen-free (SPF) mice have little or no natural killer (NK) cell activity, but when they are relocated in conventional conditions, they develop strong NCMC within 2 to 3 days. Unlike other SPF animals, hypothymic nude mice display good NCMC which is further augmented upon their entry into a pathogenic environment. The ontogeny, genetic control and other features of pathogen-activated NCMC resemble those previously described for NK cells. An H2-D region NCMC regulatory gene is active in either a homozygous or heterozygous state, but to be operative seems to require the presence of an additional complementing locus, which maps outside the H-2 complex. It is proposed that H-2 influences NCMC levels by affecting the expression or immunogenicity of NK cell-activating determinants. Recently activated NK cells were non-adherent and lacked Ia and Thy-1 determinants. NCMC was not affected by treatment with monoclonal high-titer Thy-1.2 antisera and complement. However, an Ly-6.2 antiserum did contain anti-NK cytotoxic activity, which was shown by absorption analysis to be distinct from anti-Ly-6.2 activity. The NK antigen has a strain distribution pattern distinct from Ly-5 and other Ly markers, but may be related or identical to NK-1.


Enhancement of feline leukemia virus-induced leukemogenesis in cats exposed to methylnitrosourea.

The effect of methylnitrosourea (MNU), a potent resorptive carcinogen, was evaluated for its influence on the susceptibility of adult cats to infection and induction of oncornavirus disease by feline leukemia virus (FeLV). Young adult cats at an age previously demonstrated to be highly resistant to FeLV, were injected intravenously with moderately toxic doses (15-20 mg/kg) of MNU alone or with infections FeLV (Rickard strain). Following exposure to virus and chemical, cats were monitored for antibody to the feline oncornavirus-associated cell membrane antigen (FOCMA), viremia by direct infectivity and the presence of gsa in peripheral blood leukocytes, and for toxic effects of MNU by hemogram analyses on peripheral blood. Of 8 cats injected with MNU + FeLV, 6 developed persistent viremia, 5 of which became debilitated from thymic lymphoma. Only 1 of 6 non-MNU-treated and infected cats of the same age became transiently viremic. FOCMA antibody development was markedly depressed in MNU + FeLV inoculated cats compared with cats inoculated with FeLV alone. Results show that MNU was apparently responsible for the obliteration of age-related susceptibility in cats to FeLV infection and induction of FeLV-related disease, and suggest that in nature exposure to toxic chemical carcinogens may act as factors which determine susceptibility to feline oncornaviruses in the cat.


The effect of mazindol on metabolic and regulatory changes in obese women during weight reduction.

The effect of mazindol (1 mg/day) on some metabolic and regulatory values was investigated in ten adult obese women and compared with the same number of controls during weight reduction in hospital (a five-times repeated cycle of 5 days complete starvation and 3 days on a 500 kcal(2.1MJ/day diet.) Mazindol caused a rise of hydroxyacyl CoA dehydrogenase in striated muscle by 70 per cent and a marked decline in malic dehydrogenase. Mazindol also produced higher levels of non-esterified fatty acids--significantly higher during the fifth starvation period; a small decrease in blood glucose, IRI and glucose/IRI ratio being unaffected, a significantly two-fold greater decrease of serum cholesterol; a significant increase of the noradrenaline elimination compared with a decrease in controls and in increase in triiodothyronine binding globulin towards the upper range of normal.


Nutritive value of N-acetyl-L-tryptophan in man.

The nutritive value of N-acetyl-L-tryptophan was investigated by loading tests with deuterated L-tryptophan and deuterated N-acetyl-L-tryptophan in three healthy adults, and by a 7 day-long feeding experiment with deuterated N-acetyl-L-tryptophan in one healthy adult. After loading with deuterated N-acetyl-L-tryptophan, serum levels of deuterated tryptophan and kynurenine were only about 10%, and the amounts of deuterated 5-hydroxyindole-3-acetic acid excreted in 24 h approximately 30% of those found after deuterated tryptophan loading. During the feeding experiment with deuterated N-acetyl-L-tryptophan, the serum level of deuterated tryptophan remained below 20% of total serum tryptophan, and serum protein fell below the normal range. It is concluded that the nutritive value of N-acetyl-L-tryptophan is inferior to that of L-tryptophan. Contradictory findings of earlier studies are discussed.


Orcein, collastin and pseudo-elastica: a re-investigation of Unna's concepts.

Orcein has been recommended for identification of elastin. Since other traditional elastica stains proved to be unspecific, it was deemed of interest to determine the selectivity of orcein and to review pertinent literature. Orcein was employed as a textile dye in ancient Egypt and was used for dyeing of wool and silk until the early 20th century. It was introduced into histological technic in 1878 as a stain for cytoplasm. Unna recommended it for demonstration of elastic tissue in 1890 and retracted claims for its specifity in 1894 because orcein colored also certain collagen fibers. Unna suggested the term collastin for collagen fibers which share the affinity of elastin for acid orcein. Other orcein solutions were used as selective stains for collagen. In histochemical studies, the staining properties of resorcin-fuchsin and orcein were very similar; elastin and various collagen fibers were strongly colored. Unna's collastin is apparently identical with the pseudo-elastica described in sections stained with resorcin-fuchsin. Both dyes react with meshworks of fine fibers, embryonic, experimentally or pathologically altered collagens. It is suggested to use the term collastin, instead of pseudo-elastica, for collagenous fibers which bind the traditional elastica stains.


Ruthenium red as a stain for electron microscopy. Some new aspects of its application and mode of action.

Commercial ruthenium red has been tested for its purity by spectrophotometry. Impurities detected by this method could be abolished by nitric acid-precipitation of ruthenium brown. This substance has no effect on cell surface staining and converts almost completely to ruthenium red under the conditions used in electron microscopy. It was found, by photometric analysis, that in the ruthenium red-osmium tetroxide-cacodylate combination, generally used for cell surface staining, chemical reactions between ruthenium red and osmium tetroxide occur. As aerial oxidation of hexammineruthenium2+ leads to a product with some surface staining capability, it is suggested that an oxidized product of ruthenium red is responsible for binding to cellular components, and that a reduced product of osmium tetroxide gives an additional contrast enhancement. In ruthenium red-osmium dioxide combinations ruthenium red seems to bind to cell surfaces without any molecular alteration, and contrast is gained by the model proposed by Blanquet (1976b). The latter method could open a way for investigating the binding of ruthenium red to certain natural compounds involved in calcium transport, as postulated by a number of authors. Both ruthenium-osmium combinations differ in their cell surface staining ability. The ruthenium red-osmium dioxide combination tends to form distinct subunits, whereas the osmium tetroxide variety stains homogeneously. In combination with osmium dioxide, the surface staining is affected by EDTA, and, in contrast to osmium tetroxide, a successive application of ruthenium red and osmium dioxide as possible.


The ionic components of normal human oesophageal epithelium.

The distribution of cations and anions in normal human oesophageal epithelium has been investigated with the pyroantimonate and silver-osmium tetroxide techniques. There is a discontinuous distribution of both ions in the intercellular space. The ions are associated with various organelles, as has already been described in the literature. Specifically, in the oesophageal epithelium, there are a few deposits of pyroantimonate and occasional silver in the membrane coating granules, but here is no apparent relationship of either ion with the tonofilaments or glycogen particles. The superficial cells are leaky and contain fewer ions than the deeper functional layer cells.


The histochemical demonstration of brush border endopeptidase.

A histochemical method for the demonstration of a brush border endopeptidase is described based on results of biochemical and histochemical experiments. The substrate of choice is Glut-Ala-Ala-Ala-MNA which displays a very good localization ability and suitable kinetic properties. Km estimated in rat kidney homogenate amounts to 2.35 X 10(-4) M. pH optimum of this endopeptidase associated with the brush border membrane is in the alkaline range. The activity is dependent on the buffer used. In phosphate and cacodylate buffers of pH 7.2 about 30% lower activity in rat kidney and about 25% lower activity in rat small intestine than in Tris-HCl buffer of the same pH was found. The most suitable diazonium salt for the detection "in situ" is Fast Blue B. It inhibits the endopeptidase activity of rat kidney by about 85% at pH 7.2 AND BY ABOUT 55% AT PH 6.0. The best results are obtained in cryostat sections adherent to semipermeable membranes treated with chloroform-acetone before the incubation. A microdensitometric evaluation of the reaction product is possible and results are in good agreement with those of the biochemical determination. When Suc-Ala-Ala-Ala-INA is used as substrate hexazonium-p-rosaniline is the most suitable coupling agent although it inhibits more than Fast Blue B. The reaction using acylated trialanyl naphthylamides as substrates runs in two steps. Endopeptidase sets free Ala-NA which is attacked by aminopeptidase M. Aminopeptidase M is not reaction rate or localization limiting factor because its activity in the brush border is very high and the enzyme is anchored to the cell membrane very closely to endopeptidase. In homogenates of rat kidney and jejunal mucosa the endopeptidase activity was inhibted by EDTA (2X10(-3) M) by 75% in the kidney and by 68% in the jejunum, by DFP (10(-3) M) by 41% in the kidney and by 35% in the intestine, by Mn2+ (5X10(-3) M) by 25% in the kidney and by 30% in the intestine. No inhibition was exerted by E 600. In sections the results were similar. 1,10-phenanthroline (10(-2) M) caused a substantial inhibition. Endopeptidase activity was detected in the brush border of cells of proximal convuluted tubules of the kidney and in the brush border of differentiated enterocytes of the small intestine. In the same species enterocytes display a lower activity than kidney tubular cells. There are species differences in the distribution pattern of endopeptidase in the kidney. In the rabbit and man the positive reaction occurs in the whole cortex. It is distributed unevenly, however. In the rat the tubules of the inner cortex display a very high activity. In the outer cortex straight portions react strongly. In the rabbit kidney cells of the parietal layer of Bowman's capsule display a weak reaction as well. No sex differences were found in the distribution pattern of endopeptidase in the rat kidney. In the intestine of all species examined a proximo-distal gradient was found...


The binding of streptomycin to ribonucleotides.

Incubation of streptomycin (SM) with [32P]5'-ribonucleotides at pH 7.0 produces fractions that migrate towards the cathode in high voltage electrophoresis (HVE) separations at pH 3.5. SM appears to interact with pG, pA and pC but not with pU. The appearance of these [32P]-labeled fractions is dependent on incubation time and SM concentration. Incubation of nucleotides with dihydrostreptomycin (DSM) or SM reduced with sodium cyanoborohydride (NaBH3CN) at pH 5.0, does not produce detectable changes in [32P] nucleotide mobility on HVE; however, incubation with SM reduced with NaBH3CN at pH 7.0 does produce [32P]-labeled fractions migrating with a net positive charge. Elution of [32P]-labeled material migrating towards the cathode from SM-5'-nucleotide incubations and re-electrophoresis results in nucleotides migrating with pG, pA and pC markers. These data indicate a reversible interaction between the SM-streptose aldehyde and amino-group containing nucleotides. This type of interaction may form an additional binding site for SM to RNA, relative to DSM.


Response to bleomycin of Escherichia coli mutants deficient in DNA repair.

The effect of bleomycin on the colony forming ability of Escherichia coli K12 strains in exponential growth at 37 degrees C was not affected by introducing recA13, lexA1, polA1 and uvrA6 mutations. For cells starved for amino acids, wild type strains became ten-fold more resistant to bleomycin, but again introducing lexA1, polA1 and uvrA6 strains did not change the effect on colony forming ability; however, starved recA13 cells were now four-fold more sensitive. Strains with recA13, lexA1 and polA1 mutations were always more sensitive than wild type to gamma rays under the same conditions as used for the bleomycin treatment. It is suggested that bleomycin-induced lesions may be concentrated in that part of the bacterial genomes at the cell wall, near the replication forks.


Studies of metallobleomycins by electronic spectroscopy, electron spin resonance spectroscopy, and potentiometric titration.

The 1:1 bleomycin-A2-Cu(II) complex shows an absorption maximum at 595 nm (epsilon 120), circular dichroism extrema at 555 nm (delta epsilon + 1.21) and 665 nm (-0.61), and electron spin resonance (ESR) signal with g = 2.211, g = 2.055, and A = 178 x 10(-4) cm-1. The formation constant (log K = 12.630) and deprotonation constant (PKc = 3.585) of the 1:1 bleomycin-Cu(II) complex were determined by computer analysis of potentiometric data. The results of potentiometric titration also indicate that the stability of bleomycin-metal complexes is in the order Fe(II) less than Co(II) less than Ni(II) less than Cu(II) greater than Zn(II) and that these divalent metal complexes have a similar coordination environment. The bleomycin-Cu(II) complex has substantially a square-pyramidal configuration in which the secondary amine nitrogen, pyrimidine(N-1) ring nitrogen, deprotonated peptide nitrogen of histidine residue, and histidine imidazole(N-1) nitrogen coordinate to Cu(II) as planar ligand donors, and the alpha-amino nitrogen as axial donor. The specific Cu(II)-binding site of bleomycin has been compared with that of human serum albumin.


Low resolution crystal structures of Taka-amylase A and its complexes with inhibitors.

The molecular structure of Taka-amylase A, an alpha-amylase from Aspergillus oryzae, has been studied at 6 A resolution by X-ray diffraction analysis. The electron density map showed a non-crystallographic three-fold screw arrangement of the molecules in the crystal. The molecule is an ellipsoid with approximate dimensions of 80 x 45 x 35 A and contains a hollow which may correspond to the active center. The inhibitor molecules bind to Taka-amylase A at four different sites, one of which is located in the hollow of the enzyme. The probable position of a thiol group is discussed in connection with heavy atom binding.


Structure of a major oligosaccharide of Taka-amylase A.

The structures of a major oligosaccharide of Taka-amylase A, shown below, is proposed based on the results of chemical (methylation and acetolysis) and enzymatic (digestions with exo and endo-glycosidases) analyses. This structure is an amendment of that proposed by Yamaguchi et al. (1971) (J. Biochem. 70, 587-594), in which one more mannose residue is attached (Formula: see text) through an alpha 1,2 linkage to the mannose residue which is alpha 1,3-linked to the intermost mannose residue.


Fine structural observations of calcium storage in human dental pulp cells in primary culture.

Primary culture of explants of human dental pulp tissue allows the study of the cytophysiology and differentiation of the cultured cells over a two-week period. The distribution of calcium was found in two different experimental conditions : with and without a calcium loading, by mean of a lead technique checked by microprobe analysis. The existence of two cell populations was revealed. Intra-mitochondrial ring-like granules characterize type 1 cells when overloaded, while a strong calcium storage is detected in the rough endoplasmic reticulum, Golgi apparatus and mitochondrial (without any inner organization of the deposits) of the type 2 cells. Our results also show the presence of calcium on gap-junctions (revealed) by lanthanum method), and on the extracellular matrix (collagen fibres and complex carbohydrates). The ability of some mitochondria to store calcium (ring-like granules) suggests that the type 1 cells are fully differentiated in odontoblast-like cells and perhaps engaged in mineralization processes. The calcium binding sites, localized on the extracellular matrix may therefore be considered as the earliest foci of calcification.


Practical considerations in using counterimmunoelectrophoresis to identify the principal causative agents of bacterial meningitis.

Many clinical laboratories are currently using counterimmunoelectrophoresis (CIE) as an aid in the rapid diagnosis of bacterial meningitis. Because cross-reactions among causative agents have been reported, the present study was undertaken to explore the problems that might occur when reference and commercial antisera are used in CIE. Broth cultures of 35 bacterial strains were tested with 76 reference and commercial antisera by CIE. Some of the antisera tested failed to react with their homologous strains. Furthermore, several cross-reactions between genera, as well as within species, were noted. These findings suggest that precautions must be taken to insure that all materials used in CIE tests are of high quality. If properly performed and interpreted, CIE may be a valuable adjunct in the identification of organisms causing bacterial meningitis, but it is, nevertheless, a presumptive test and should not be used to replace the Gram stain and culture techniques.


Neutralizing antibody responses of humans and mice to vaccination with Venezuelan encephalitis (TC-83) virus.

Laboratory workers were vaccinated with Venezuelan encephalitis virus, strain TC-83. After 2 to 3 years, they were bled, and their sera were tested for neutralizing antibody to all known Venezuelan encephalitis subtypes and varieties. The results indicated higher titers to epizootic than to enzootic Venezuelan encephalitis viruses and suggested that individuals vaccinated with TC-83 do not produce significant neutralizing antibody to heterologous subtypes. Mice vaccinated, bled, and tested in a similar manner produced much the same antibody profiles as did the humans and resisted challenge with all Venezuelan encephalitis viruses tested. In addition, the serum dilution plaque reduction neutralization test used was shown to be highly specific and adequate for diagnosis of infections with Venezuelan encephalitis viruses.


Comparison of auramine-rhodamine B and acridine orange for staining of acid-fast bacteria.

In cooperation of 6 laboratories in Czechoslovakia and in the GDR, the efficiency of auramine-rhodamine B (AR) and acridine orange (AO) (short-time method) for staining of acid-fast bacilli was compared. Whereas a former comparison of AR and AO (original method) pointed out the superiority of AR, the investigation of both methods used as short-time procedures showed significantly more acid-fast rods after using AO. The number of "false positive" results was somewhat higher on AR staining. However the results depend not only on the method used but also on the procedure of staining and the optical equipment, and they are essentially influenced by the experience and proficiency of the microscopist. Taking into account the results of both studies both auramine-rhodamine B and acridine orange can be proposed for the staining of slides for microscopical detection of acid-fast rods. In case of AO, the short-time method is superior to the original long-time procedure.


Genetic variation in the M antigen of group A streptococci: reassortment of type-specific markers and possible antigenic drift.

The phylogenetic relationships of 73 strains of group A Streptococcus isolated over a six-year period from a population with a high prevalence of streptococcal infections were analyzed with use of four serotype-specific markers: the serum opacity factor (OF), the T antigen, the determinant of the M antigen that precipitates with antibody, and the M antiphagocytic determinant. OF inhibition tests divided the strains into five subtypes: a, b, c, d, and e. Strains within subtypes a, b, and c shared a previously unidentified M precipitin. The identification of this new common M precipitin was based on absorption studies, double agar immunodiffusion, and immunoelectrophoresis analysis. Antisera directed against representative strains from subtypes a, b, and c contained cross-reactive bactericidal antibody, and absorption studies indicated that determinants of the M precipitins were identical, whereas determinants of resistance to phagocytosis were closely related but not identical. The association of a common M precipitin with three different OF antigens, two different T antigens, and three different, yet similar, antiphagocytic determinants is discussed in light of the possibility that the genes that code for these antigens have reassorted or recombined. Moreover, the possibility that differences in the antiphagocytic determinants of these strains have resulted from antigenic drift in a common gene is considered.


Immunoprecipitation of Triton X-100-solubilized Mycoplasma mycoides proteins.

Mycoplasma mycoides subsp. mycoides (PG1 and strain Y) proteins were solubilized in Triton X-100, and the antigenic proteins were precipitated from this complex mixture by addition of antiserum and then separated by two-dimensional gel electrophoresis. Of the 300 proteins solubilized, about 10 were precipitated. Proteins of PG1, a slow-growing, small colony (SC) strain, were precipitated by antiserum to PG1 and by antiserum to strain Y, a fast-growing, large colony (LC) strain. Similarly, strain Y proteins were precipitated by antiserum to PG1 and by antiserum to strain Y. The few proteins precipitated in this way gave similar patterns after two-dimensional gel electrophoresis indicating that many of the dominant protein antigens of PG1 and strain Y are shared. Antiserum to Mycoplasma mycoides subsp. capri (PG3) also precipitated some proteins of strain Y. Antiserum to Mycoplasma gallisepticum gave no reaction with any M. mycoides antigens. It was concluded that, in addition to the polysaccharide antigens, there are proteins in M. mycoides that are antigenic and that some of these are found in both the SC and LC strains of subsp. mycoides and also in subsp. capri.


Mechanism of the inhibitory action of linoleic acid on the growth of Staphylococcus aureus.

Linoleic acid, but not stearic acid, inhibited the growth of Staphylococcus aureus NCTC 8325. Growth inhibition was associated with an increase in the permeability of the bacterial membrane. The presence of a plasmid conferring resistance to penicillin (PC plasmid, e.g. pI258blaI-) increased the growth inhibitory and membrane permeability effects of linoleic acid. Under growth inhibitory conditions, linoleic acid was incorporated into the lipid of both PC plasmid-containing and PC plasmid-negative bacteria and there was little difference between these cultures in the uptake or fate of linoleic acid. Experiments using a glycerol auxotroph of S. aureus suggested that free linoleic acid, rather than lipid containing this acid, inhibits growth. Linoleic acid probably inhibits growth by increasing the permeability of the bacterial membrane as a result of its surfactant action, and the presence of the PC plasmid increases these effects.


Susceptibility to butirosin and neomycin B in Bacillus circulans, the butirosin-producing organism.

Butirosin, an aminoglycoside antibiotic, is produced by Bacillus circulans B-3312. Experiments using recombined ribosomal and supernatant fractions from this strain and from B. megaterium KM have shown that the ribosome of both are sensitive to butirosin. The aminoglycoside 3'-phosphotransferase present in B. circulans modifies butirosin and neomycin in vitro but confers resistance only to the former in vivo. The phosphotransferase does not modifya detectable amount of extracellular butirosin while mediating resistance to the antibiotic. In vitro, however, the enzyme appears to protect against inhibition by butirosin by inactivating the bulk of the antibiotic in the system. An extrachromosomal element of unknown function has been detected in B. circulans.


Analysis of hepatitis B surface antigen components solubilized with Triton X-100.

Three glycoproteins of intact hepatitis B surface antigen (HBsAg) with mol. wt. of 32 000, 30 000 and 28 000 respectively were identified by reaction with 125I-concanavalin A (Con A) after separation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The antigen was effectively disrupted with Triton X-100 to produce a structure with a sedimentation coefficient of 3.9S. Affinity chromatography of disrupted HBsAg using concanavalin A-Sepharose 4B (Con A-Sepharose) resulted in two fractions. The first contained material which did not bind to the lectin and consisted of a single polypeptide of mol. wt. 64 000. Further studies revealed this component to be serologically identical to serum albumin and to lack any affinity for antibody to HBsAg. A comparison of the tryptic peptide map of this polypeptide with that of purified serum albumin demonstrated identical amino-acid sequences. The second fraction contained material which bound to Con A and contained two polypeptides with mol. wt. of 28000 and 23000 respectively. HBsAg reactivity was associated with this fraction. This procedure allows the prepartion of HBsAg sub-units in milligram quantities for further immunological studies.


Characterization of adenovirus protein IX.

Protein IX from adenovirus type 2 was purified by two methods, one from groups of nine hexons obtained by disrupting purified virus by heating in the presence of deoxycholate, and the other by a previously published method. The purified protein was used to obtain a monospecific antiserum. Protein IX was found to possess both sub-group- and type-specific antigenic determinants which were apparently accessible within the groups of nine hexons. Approximately 15 molecules of IX were found per group of nine hexons and from considerations of symmetry it seemed possible that IX was located at the 'corner to edge' contacts between hexons in the icosahedron. The protein in infected cells was found to possess approximately neutral charge as determined by immunoelectrophoresis. This was consistent with the amino acid composition, which showed it to be rich in serine, alanine and leucine with approximately half of its glutamic and aspartic acid residues amidified, and the isoelectric point of 6.0, as determined by two dimensional gel analysis. No free N-terminal amino acid was detectable. It is suggested that a unique tryptophan residue is located at around position 70 from the blocked N-terminus, on the basis of chemical cleavage by BNPS-skatole. Based on one tryptophan residue a total of 107 amino acids and a mol. wt. of 11200 was deduced. Analysis of 35S-methionine-labelled infected cell extracts in a two-dimensional gel system showed that the synthesis of polypeptide IX could be detected early in infection, i.e. in the presence of an inhibitor of DNA synthesis.


Antigenic and structural relationships between avian paramyxoviruses isolated from ducks in Hong Kong and Mississippi, U.S.A.

Representative isolates of the paramyxoviruses duck/Hong Kong/75 and duck/Mississippi/75 were shown to be serologically closely related by haemagglutination and neuraminidase inhibition tests. The structural polypeptides of these viruses were also shown to be similar. For each of the isolates tested, polyacrylamide gel electrophoresis in the presence of SDS revealed a similar polypeptide pattern consisting, under reducing conditions, of seven polypeptides with apparent mol. wt. ranging from 46000 to 190000. Each virus had two glycosylated polypeptides with apparent mol. wt. of 56000 and 71000 to 72000 under reducing conditions and 62000 to 63000 and 135000 to 142000 under non-reducing conditions.


Physicochemical heterogeneity of hepatitis B e antigen detected in asymptomatic carriers and carriers in a hemodialysis unit.

Physicochemical studies of hepatitis B e antigen (HBeAg) revealed a clear cut difference between e1 and e2 antigen. The e1 antigen was found to have a MW of Ca 150,000 and a pI of 6.4-7.2, whereas both the MW and pI of the e2 antigen were heterogeneous depending upon the source of serum. Sera obtained from asymptomatic carriers were characterized by low titers of HBs antigen, HBc antigen and DNA polymerase and contained e2 antigen of larger molecular weight (200,000-300,000) with a narrow distribution range and a pI of 4.8 to 5.2 (type 1). On the other hand, the sera from patients in a hemodialysis unit who were HBs antigen carriers and had high titers of HBs antigen, HBc antigen and DNA polymerase contained e2 antigen of heterogeneous distribution in MW (from 300,000 to 70,000) and pI (type 2 and 3). The e2 antigen obtained from the higher MW type 3 serum had lower isoelectric points (pI 4.5 to 5.2) as was the case with e2 antigen obtained from asymptomatic carriers whereas relatively wide range of isoelectric points (pI 5.1 to 8.2) was found with the lower molecular weight e2 antigen.


Changes in brain dopamine, norepinephrine and serotonin associated with convulsions induced by pinealectomy in the gerbil.

In three separate experiments, adult male gerbils 10 weeks of age were pinealectomized (PX), observed for convulsions, and sacrificed at various intervals after PX. Norepinephrine (NE), dopamine (DA) and serotonin (5-HT) in selected brain parts were determined by fluorometric procedures. No changes in DA were observed. Serotonin levels were shown to be elevated in sham-operated and PX animals, possibly due to stress phenomena associated with anesthesia or surgical procedures. Norepinephrine was significantly depressed over control and sham-operated animals by 1 hour after surgery with a post-operative duration of at least 12 hours. Results in this study did not indicate whether the decreased NE was the cause or an effect of the PX-induced convulsive activity.


Charcot-Marie-Tooth disease associated with retinal pigment dystrophy and protanopia. Neurological, ophthalmological and genetic study of a family.

Neurological, ophthalmological and genetic investigations were performed on a family, a member of which presented with a rare association of tapeto-retinal degeneration, protanopia and Charcot-Marie-Tooth disease (CMT), and asked for genetic counseling. The neurological enquiry was completed by measurement of motor nerve conduction velocity in several completed by measurement of motor nerve conduction velocity in several members of the family. The propositus was submitted to a muscle biopsy. The ophthalmological examination included ophthalmoscopy, fluorescein angiography, electroretinogram and electrooculogram. The propositus, a woman aged 40, had typical CMT disease and her father also had a mild form of it. She had protanopia as had her father, her son and her nephew. In addition she had large macular pigmented changes, described as retinal dystrophy, "flavus flavimaculatus." Her mother had only senile pigmented modification of the fundus and her three daughters had mild macular pigmented changes, like "salt and pepper." Two genes are probably involved: one for protanopia with X linked recessive inheritance, the other responsible of CMT and tapeto-retinal degeneration, with an autosomal dominant inheritance, giving a 50% risk of recurrence.


Treatment of myasthenia gravis. Report on 139 patients.

In the treatment of myasthenia gravis (MG) considerable progress has recently been achieved. Our experience is based on the observation of 139 patients with an average follow-up of 3 years and 4 months. A treatment plan and results are presented. Indications for thymectomy: all cases of MG in adult life, apart from ocular myasthenia without radiological thymoma and without electrophysiological and pharmacological signs of generalization; before puberty only cases with radiological thymoma and severely incapacitating or life-threatening signs. Median sternotomy is preferable for thymoma, the transcervical approach with a sternal split for non-neoplastic thymus. Mediastinal radiotherapy is indicated after removal of an invasive or adhesive thymoma. Indications for corticosteroids: 1) before thymectomy: respiratory weakness; 2) soon after thymectomy: life-threatening signs; 3) later after thymectomy: incapacitating or life-threatening signs; 4) as an alternative to thymectomy: when surgery cannot be performed or it is not indicated. Oral Prednisone was nearly always preferred: alternate-day high single dose (75 to 115 mg) has given good results in most cases even if in some cases a small dose was required in the "off day"; inversely a lower alternate-day or daily dose was often sufficient. Long-term results: following this schedule for adult patients good results were scored in 67% of thymomas, in 94% of hyperplasias, and in 62% of unthymectomized patients: in prepuberal life the few cases of severe MG have all shown a favorable evolution.


Cerebral atrophy and long-term response to levodopa in Parkinson's disease.

In 92 parkinsonian patients (42 men, 50 women) aged from 37-79 years (mean 62.8) the impact of cerebral atrophy as assessed by computed tomography on the course of the clinical symptomatology under levodopa during a period of 1 to 5 years was investigated. It could be shown that patients suffering from medium to severe degrees of atrophy-independent of its special location--have a less favorable response than those with normal CT findings. The data do not allow a definite decision whether cortical artrophy or ventricular enlargement are of major importance. At any rate, the width of the third ventricle seems to have no influence on the course of the clinical symptoms. After 3 years of levodopa treatment a dissociation between cerebral atrophy and therapeutic effectiveness can be observed.


The prognosis of intraventricular hemorrhage.

A series of 40 patients with blood in the cerebral ventricular system demonstrated by computed tomography (CT) is presented. Overall prognosis was found to be better than expected from the literature. Nearly half of the patients are still alive; 12 of the 40 patients in this study had no significant residual disability. The results suggest that the rupture into the ventricular system by itself is of minor importance. However the clinical condition of the patient on admission, site and origin of the ruptured intracerebral hematoma and possibly the age of the patient seem to play an important role in predicting the outcome.


Diagnostic value of brief microzone electrophoresis of unconcentrated CSF and agar gel electrophoresis of concentrated and unconcentrated CSF.

This paper reports a comparison between brief microzone electrophoresis and agar gel electrophoresis of CSF proteins; 41 different CSFs were examined simultaneously by all three methods. Fractionation of gamma-globulins was found by brief microzone electrophoresis in 53.7% of unconcentrated CSF. This fractionation was found in 63.4% of unconcentrated CSF after agar gel electrophoresis, in 48.8% of concentrated CSF. Serum-like pherogram and the rapid alpha-globulin type were found equally often.


Vertigo and drop attacks caused by acute transient monocular disequilibrium (Halpern's syndrome).

Among the causes of acute vertigo the syndrome of sensorimotor induction in unilateral disequilibrium (Halpern's syndrome) should be considered. This syndrome was found in a patient and described in detail. The major features of this syndrome are the displacement of vertical and horizontal axes induced by looking with the "affected" eye only, which are further aggravated by applying red filters to the eyes only, which are further aggravated by applying red filters to the eyes and corrected by blue filters. The symptomatology is described and discussed in detail. Theories causing this syndrome are discussed.


Differentiation of normal pressure hydrocephalus and cerebral atrophy by computed tomography and spinal infusion test.

The diagnostic value of computed tomography (CT) and spinal infusion test (SIT) was investigated in 27 patients with normal pressure hydrocephalus (NPH) and 35 patients with cerebral atrophy. The most consistent CT finding of NPH was dilatation of the temporal horns, that of cerebral atrophy widening of the convexity sulci. However, 43% of patients with cerebral atrophy demonstrated no cortical atrophy. The SIT showed an excellent relation with isotope cisternography and continuous intracranial pressure recording. NPH and cerebral atrophy were correctly differentiated in 71% by CT and SIT. A normal SIT and a CT scan without the typical features of NPH exclude impairment of cerebrospinal fluid absorption. An abnormal SIT and a CT scan showing ventricular enlargement without dilatation of convexity sulci, require isotope cisternography and possibly intracranial pressure recording to determine the degree of the absorption deficit.


The symptomatology with the most severe clinical course of spontaneous subarachnoid hemorrhage.

The symptomatology of 18 patients with the severest clinical course after subarachnoid hemorrhage (SAH) is described and analyzed. Seventeen patients died, five with an acute bulbar syndrome with cardiac arrest, and 12 with irreversible breakdown of brain function. One patient had an apallic syndrome with minimal signs of remission, who died 4 months after the first rebleeding. The cases are divided into five clinical groups on the basis of the acute brainstem symptomatology which set in instantly or after temporary adaptation, or following a second hemorrhage. Pathological examination revealed that 16 patients had a brainstem pressure cone, 14 with marked edema and signs of herniation; one patient had only cisternal tamponade while another had predominantly brain edema with herniation. There was striking parenchymatous damage of the brain of the patient with the apallic syndrome. There was a marked analogy between the material reported and the symptomatology described by Plum and Posner (1972).


Peripheral neuropathy after administration of tetanus toxoid.

A reversible paresis of several muscles of the right arm was observed in a 20-year-old female, 3 weeks after the intragluteal injection of tetanus toxoid and 6--7 weeks after multiple injuries. The EMG findings indicated a transitory conduction block of nerve fibers to the deltoid and pectoralis muscles and a more severe axonal-demyelinating lesion of fibers to the biceps muscle. The cause of the palsy was probably a reaction to the administration of tetanus toxoid; other possible causes are discussed.


Electrophysiological findings in workers exposed to N-heptane fumes.

Electrophysiological examination of the peroneal nerve was performed in a group of individuals exposed for professional reasons to vapours of n-heptane. The results suggest a probably specific role of this hydrocarbon compound in provoking "minimal" peripheral nerve damage.


Genetic segregation of multiple sclerosis and histocompatibility (HLA) haplotypes.

The possibility that a gene determining susceptibility to multiple sclerosis (MSS) may be closely linked to the major histocompatibility locus (HLA) is suggested by observation of a loose association between multiple sclerosis (MS) and certain HLA determinants. In the present study, the possible association was analyzed by studying the segregation of MS and the HLA haplotypes in families with more than one case of MS. Analysis of 48 published families revealed that the haplotype shared by those with MS within the family was also shared by those without clinical signs of MS at close to the 50% frequency expected by chance. Thus, we were unable to demonstrate that MS is associated with one HLA defined parental haplotype. We discussed reasons for this apparent failure to demonstrate existence of an MSS gene using available multiplex MS families.


Familial cerebellar ataxia with hypogonadism.

A brother and sister with congenital cerebellar ataxia, anosmia, oligophrenia, hypogonadism and anomalies of amino acid distribution are reported. Ties between the different symptoms are difficult to establish. It seems to be a new syndrome rather than a new disease. Once more these associations emphasize the need for metabolic and biochemical research in heredodegenerative diseases. The evolution of the disease in these cases might make it possible to classify and locate it more accurately.


Pattern reversal evoked cortical responses in normals. A study of different methods of stimulation and potential reproducibility.

Using a commercially available television set for stimulation of 78 healthy subjects, the upper limit of normal for the latency of the major positive wave (p 100) of checkerboard pattern reversal evoked potentials is practically the same as that obtained by employing slide projector combined with a rotating mirror. Potentials evoked by a small pattern, for purely foveal stimulation, were often difficult to evaluate exactly. Fixation of a large pattern at the upper border of the stimulus field brought no advantages as compared to the usual fixation in the center. Continuous prolonged recordings, with successive averaging of 64 pattern reversals, confirmed statements that only the major positive wave and its latency are constantly reproducible without being influenced by fatigue or inattention. Repeated examinations in the same subjects at intervals of 2 weeks showed a considerable spread of the latencies from one session to the other but the absolute values always ranged within the normal. More or less large latency differences (up to 12ms with large pattern stimulation) were found in every subject at least once. A longer latency found at a control examination, or a newly appearing latency difference, therefore do not prove a fresh optic nerve lesion as long as the absolute values are still within the limits of normal. From repeated examinations it could also be seen that foveal stimulation with a small pattern is not suitable for routine examinations because of high variation in the results.


Benign encephalitis: electro-oculographic analysis of opsoclonus.

Two cases of benign encephalitis are described, which showed opsoclonus associated with myoclonic jerks of the body and face, and with cerebellar dystaxia in one case. EOG and EEG analysis during the course of these cases showed: 1. opsoclonic eye movements were triggered by saccadic and non-saccadic eye movements, 2. decrease of luminancy and loss of fixation were releasing, but not triggering opsoclonic bursts during the initial stages, 3. eye closure had the strongest trigger effect, which lasted very long and allowed an objective description of the (benign) course of the illness, 4. besides conjugate, mostly horizontal, spontaneous eye movements oblique and vertical ones were also obtained. Monocular and disconjugate eye movements were seen only during the initial stages, 5. statistical analysis of the frequency of single eye movements during periods of opsoclonic bursts showed that, in the course of the disease the variability of frequencies decreased and the degree of fast frequencies increased, particularly with open eyes, 6. the frequent coincidence of opsoclonus and lesions of the cerebellum or of the cerebellar pathways is striking, but the initially distinct EEG changes, the very different trigger modes and certain pathoanatomically described cases without any cerebellar lesion indicate the more general character of the disturbance. 7. the entity of opsoclonus, body tremulousness, and benign encephalitis has to be differentiated from other syndromes including the sign opsoclonus by recording the EEG and EOG during the course of the disease; this might lead to very useful diagnostic and prognostic information.


Calcium and cyclic nucleotide interaction in secretion of amylase from rat pancreas in vitro.

1. Interaction of Ca2+ and cyclic nucleotides in stimulus-secretion coupling in rat pancreas in vitro was studied utilizing the divalent cation inophore A23187. phosphodiesterase inhibitors, cyclic nucleotides and cholera toxin. 2. Amylase secretion was increased by the ionophore in the presence of extracellular Ca2+ in a dose-dependent fashion. Activation of CCK-PZ receptors simultaneously with induction of amylase secretion by A23187 did not alter amylase secretion whereas theophylline or caffeine had effects additive to A23187. Dibutyryl cyclic AMP potentiated the effect of ionophore whereas dibutyryl cyclic GMP had no effect on basal or ionophore-induced amylase secretion. Cholera toxin by itself did not effect amylase secretion whereas it potentiated the effect of ionophore. 3. A23187 increased bidirectional fluxes of 45Ca and increased efflux of 45Ca in a fashion similar to CCK-PZ. Theophylline did not alter basal efflux of 45Ca. Dibutyryl cyclic AMP increased the basal efflux of 45Ca whereas, cholera toxin, dibutyryl cyclic GMP and sodium butyrate had no effect. 4. Theophylline increased basal cyclic AMP levels with a peak effect observed at 5 min. Combination of theophylline and ionophore did not lead to an increase in levels of cyclic AMP greater than that observed with theophylline alone. Cholera toxin increased cyclic AMP levels at 30 and 60 min of incubation. 5. Ionophore and CCK-PZ increased tissue cyclic GMP levels significantly greater than that obtained with theophylline alone. This effect was dependent on extracellular Ca2+. The effect of ionophore on tissue levels of cyclic GMP could be dissociated from its effect on 45Ca efflux and amylase secretion. 6. It is concluded from these studies that Ca2+ plays a predominant role in regulating amylase secretion with interactions occurring between Ca2+ and cyclic AMP and Ca2+ and cyclic GMP. It appears that by themselves cyclic AMP and cyclic GMP do not play a significant role in regulating enzyme secretion.


Activation and inhibition of calcium-dependent histamine secretion by ATP ions applied to rat mast cells.

1. The concentration dependence on ATP of mast cell histamine secretion in the presence of various concentrations of Mg2+ and Ca2+ confirms that the agonist form of ATP is the free form of ATP (ATP(free) not bound to divalent cations, i.e. ATP4-. It induces 50% activation at about 1.2 microM, maximal secretion at about 2.7 microM and 50% self-inhibition at about 4.4 microM. 2. The divalent cations Mg2+ and Ca2+ were used to buffer ATP(tree) in the range 1-8 microM in the presence of much higher concentrations of ATP(total). In addition to its effect as a buffer for ATP, Ca2+ is required for secretion. 3. With ATP(free) at 1 microM, the time-course of histamine secretion is characterized by a delay of about 10 min before secretion commences. With increasing concentration of ATP(free) the delay becomes shorter (less than 5 min with ATP(free) at 2 microM). 4. Secretion commences promptly on addition of Ca2+ to cells which have been pretreated with low concentrations of ATP(free) (less than 2 microM). This observation suggests that the delay normally observed represents the time taken for Ca2+ sensitivity to develop (i.e. probably the time taken for Ca2+ channels to open). 5. Late addition of Ca2+ to cells pretreated with higher concentrations of ATP(free) (greater than 2 microM) results in a reduced amount of histamine secretion compared with that which normally occurs. This reduction (which increases with time of exposure to ATP) and the self-inhibition due to higher concentrations of ATP(free) may be two facets of a common inhibitory mechanism. 6. These results are discussed in the light of other experiments which show that mast cells treated with ATP(free) at self-inhibitory concentrations become permeable to phosphorylated metabolites and nucleotides.


Amylase secretion in the rabbit parotid gland when stimulating the sympathetic nerves during parasympathetic activity.

1. In anaesthetized rabbits amylase secretion from the parotid gland was investigated. Secretion was evoked by sympathetic nerve stimulation, either alone or superimposed on a parasympathetic background secretion, imitating the resting secretion present in the waking animal.2. Sympathetic nerve stimulation at frequencies below 1 Hz was alone subthreshold for fluid secretion, but could greatly increase the amounts of amylase present in fluid secretion produced by parasympathetic nerve stimulation. The amylase output due to sympathetic nerve stimulation alone at 10 Hz did not exceed that seen in response to a stimulation at 1 Hz superimposed on parasympathetic activity.3. The amylase output in response to superimposed sympathetic stimulation was not influenced by the rate of fluid secretion, which was altered by stimulating the parasympathetic nerves at different frequencies.4. Sympathetically-evoked amylase secretion was abolished after beta(1)-block. The amylase secretion remaining on parasympathetic activation was sparse.5. It is concluded that secretion of amylase in response to sympathetic nerve stimulation requires the presence of a parasympathetic fluid secretion to be washed along the glandular ducts. Parasympathetic activity may also augment the sympathetic effect on amylase secretion.


Delayed neurotoxicity of subchronic oral administration of leptophos to hens: recovery during four months after exposure.

Daily oral administration of small doses of technical grade O-methyl O-4-bromo-2,5-dichlorophenyl phenylphosphonothioate (leptophos, 0.5-20.0 mg/kg) caused delayed neurotoxicity in hens. Severity of clinical condition and progression or improvement of signs of delayed neurotoxicity depended on the dose and duration of administration. Hens given 20.0 mg/kg suffered ataxia, paralysis, and death. Intermediate doses (5 and 10 mg/kg) caused ataxia, with most treated hens showing no change in clinical condition during the 4-mo observation period. Hens given small doses (2.5 and 1.0 mg/kg) demonstrated regression of neurological deficits after administration of leptophos was stopped. Hens given the smallest tested dose (,.5 mg/kg) developed mild ataxia and showed total recovery during the observation period. Days of administration and total administered dose before onset of ataxia depended on the daily dose. Degeneration of axons and myelin i, the spinal cord was the most consistent histopathologic change and was identical to that observed in tri-o-cresyl phosphate (TOCP) control hens. Only one hen, which died early in the treatment period, showed peripheral nerve degeneration. Controls consisted of 3 groups of hens given a daily oral dose of 10.0 mg/kg TOCP, 1.0 mg/kg O,O-diethyl O-4-nitrophenyl phosphorothioate (parathion), or an empty gelatin capsule. TOCP-treated hens developed delayed neurotoxicity, whereas those given parathion showed initial leg weakness but subsequently recovery without developing delayed neurotoxicity. Controls given gelatin capsules remained normal.


[Determination of LH and FSH levels before and after prostatic adenomectomy (author's transl)].

Plasma testosterone, LH, and FSH levels were measured in 29 patients with adenoma of the prostate, before and after adenomectomy. Blood testosterone levels showed significant variations which were probably related to operation stress, as after a moderate reduction--they returned to pre-operative levels. No significant modifications in plasma LH or FSH levels were observed.


[Radiomanometric study of the anatomic sites of the pressure profile (author's transl)].

The analysis of the urethral pressure profile was realised on 30 normal patients. The technique of radiomanometry permitted to visualize the anatomic sites of the urethral and the corresponding pressure.


Dietary control of the renal absorption and excretion of alpha 2u-globulin.

The kidneys are an important factor in controlling the excretion of the sex-dependent protein, alpha 2u globulin, in the urine. On a protein-free diet (0% casein), when the plasma level was reduced by 60%, the daily excretion of alpha 2u in the urine was reduced from a normal of 11 mg to 0.2 mg; 97% of the alpha 2u filtered by the kidneys was reabsorbed. When a normal-protein diet (20% casein) was restored, the serum alpha 2u levels were returned to normal within 48 hours. The urinary excretion, on the other hand, remained below normal until the eighth day after repletion, whereas the renal concentrations of alpha 2u reached higher than normal values by the fourth day. Thus, during protein deficiency the kidney reduced excessive losses of protein in the urine by an almost total reabsorption of alpha 2u. On a high-protein diet (50% casein), the serum levels of alpha 2u were normal, yet its excretion was elevated 80% above the normal (20 mg/day compared with 11 mg/day). To effect this increased loss, the reabsorption of the protein was reduced to 26% of the normal.


Double staining technique for rat foetus skeletons in teratological studies.

Cartilage and bone were differentiated using alcian blue and alizarin red S respectively. Anomalies of both cartilaginous and bony parts of the skeleton could be examined.


[Determination of the lymphocytic and oncofetal antigen subpopulations in patients with adenocarcinomas of the stomach and of the colon and rectum (author's transl)].

The authors present a study of 50 patients with adenocarcinomas of the colon and rectum, patients with gastric adenocarcinomas, and 30 healthy individuals as a control group. In all subjects the following parameters were determined: total number of lymphocytes in the peripheral blood, T lymphocytes, T-active lymphocytes, and B lymphocytes. A study of the test for lymphoblastic transformation (TTL) with phytohemagglutinin (PHA) stimulation and the determination of alpha-fetoprotein (AFP) and carcino-embryonic antigen (CEA) were also carried out. In patients with gastric adenocarcinoma the results revealed a lymphopenia, especially at the expense of T and T-active lymphocytes, as well as a depression (in 73 per cent) of the lymphocytic response to the PHA stimulation. Patients with carcinoma of the colon showed significant results in the T-active lymphocyte population. In both neoplastic situations the determination for alpha-fetoprotein was negative, while the CEA presented a clear correlation with the evolutive stage of the tumor, being more demonstrative in the tumors located in the colon and rectum.


[Systemic lupus erythematosus induced by anticonvulsivant drug (author's transl)].

A 20-year-old patient with previous history since 9 years of age of epileptic crises, was followed since then by anticonvulsivant therapy. Eight months before her admittance to the hospital ethosuximide was added to the anticonvulsivant treatment with good results. The development of fever, pain in the joints, and pleuropericarditis decided her admittance, being the clinical evaluation analytically and immunologically compatible with a systemic lupus erythematosus. The discontinuance of the anticonvulsivant medication determined at the end of the 3rd week the clinical and analytical recovery of the patient. The immunological study was normal after 3 months. The addition of other anticonvulsivant drugs, without having modified the normal condition of the patient, establishes a cause-effect relationship after the administration of ethosuximide. The infrequency of this observation is commented on, and the present knowledge of the possible mechanisms implicated in the drug-induction of a systemic lupus erythematosus is discussed.


[The control of culicidae in Djibouti town. An experience of 7 years (author's transl)].

The control of culicidae in Djibouti is based first on an antilarval action with suppression of breeding areas and use of Dursban and Abate 5 times a year. Baygon, the use of which is more expensive, is employed for imagicidal control in well delimitated foci.


Projection system with brightness amplifier for biology and medicine.

Some results of investigation of biological systems using laser projection microscope is given. The scheme permitted to work both in reflected and transmitting light. The image of a living cell on a large screen was observed.


Staining of DNA-phosphate groups with a mixture of azure A and acridine yellow.

This paper deals with staining of DNA-phosphate groups with a mixture of an equal parts of aqueous solution of azure A and acridine yellow in a 1:1 proportion and also embodies a study of the absorption properties of the stained nuclei. It also embodies results of sequential staining of nuclei stained first with azure A followed by staining with acridine yellow and vice versa, after extraction of RNA with cold phosphoric acid. The results indicate that the absorption peaks of nuclei differ from those of nuclei stained for DNA-aldehyde molecules with azure A-SO2 or acridine yellow-SO2. The in vitro absorption characteristics of an aqueous solution of azure A and those of an aqueous solution of acridine yellow are also presented herein. The conclusion obtained from this study is that all the phosphate groups of DNA do not take part in the staining process when staining is carried out with azure A or acridine yellow alone after after RNA has been extracted. This is because the nuclei stained with these dyes sequentially show the presence of acridine yellow-DNA and azure A-DNA complex.


Staining of acid mucopolysaccharides appearing on and in various cell types by DAPI.

It could be shown that DAPI, a newer fluorochrome (4',6-diamidino-2-phenylindole), is able to stain various types of acid mucopolysaccharides, e.g. heparin and agar mucopolysaccharides. Due to their content of heparin, mast cells and basophil cells could be stained rapidly. Other cells e.g. mononuclear phagocytes and malignant epithelial cells, could be loaded with these acid mucopolysaccharides and subsequently stained by DAPI.


[Changes in the physiologic properties of a chemostat culture of Propionibacterium shermanii during growth limitation by a deficiency of sodium lactate].

A chemostat curve has been constructed for changes in the Propionibacterium shermanii population density with an increase in the flow rate of the medium from 0.04 to 0.55 h-1 upon limitation of the growth with sodium lactate deficiency. The curve differs from the classic chemostat curve described with the Monod equation, thus indicating changes in metabolism at different growth rates. Fermentation of lactate to acetic and propionic acids by the culture is most complete at low flow rates of the medium (D = 0.04 h-1). The rates of oxidation processes increase at average flow rates (D = 0.1--0.33 h-1). Lactate is both fermented to yield volatile acids, and oxidized. The growth rate D = mu = 0.55 h-1 approaches the maximal one. The rate of fermentation by the culture at this growth rate decreases while the rate of oxidation increases. The ratio between the rates of these two processes shifts towards oxidation. The cells display now a very high synthetic activity: constructive metabolism of the cells is realized practically only at the account of the nitrogen from ammonium rather than the nitrogen of amino acids. The chemical composition of the cells along the chemostat curve is relatively constant. The content of protein and DNA does not change; the content of RNA slightly rises with an increase in the growth rate, and the protein synthesizing activity of RNA is directly proportional to D = mu.


Effects of a chromatin low molecular weight peptidic fraction on differentiation markers and virus production in Friend leukemia cells.

Low molecular weight chromatin peptides exert a dose-dependent inhibition of Dimethylsulfoxide (DMSO)-induced erythroid differentiation of murine Friend Leukemia Cells (FLC). This effect correlates with the degree of purification of the peptide fractions. Crot analysis of globin mRNA amounts in DMSO-treated FLC given the peptides showed a 4-5-fold decrease of messenger RNA in the cytoplasma with no nuclear storage of globin transcripts. Spectrin accumulation in "induced" FLC is inhibited as well. The effects of the peptides on erythroid markers are reversible upon removal of the compounds. They also appear to be specific for induced gene expression as (1) no effects are observed on cell growth and RNA synthesis in normal non-differentiating cell lines; and (2) no changes have been detected with regard to the expression of integrated viral genes coding for continuous shedding of viral particles.


[Diagnosis value of lipid content in prostatic cancer].

Authors have demonstrated presence of lipide droplets in the tumour cells in 16 cases out of 22 of the carcinoma of prostata. In 8 cases the tumour was of the xanthomatous variant of Kinoshita. Metastases were examined in 4 cases. Cells of the metastatic tumours contained lipide alike the primaer ones. Authors believe, that lipide content of the metastatic carcinoma may help in recognition of the original tumour, i.e. prostatic carcinoma.


[Modified trichrome staining].

Author in the process of the trichrom staining method instead of Orange-G, acid fuchsine and aniline-blue applied other dyes. For the staining of thin sections ponceau-S was found more suitable than acid-fuchsine. For the staining of thick sections the cerommends acid-fuchsine, light-green and fast-green FCF.


A simple method for staining semi-thick tissue sections to demonstrate fungal elements.

A simple method of staining epon embedded semi-thick sections with Schiff-reagent is described. This technique allows fungal elements in tissue to be readily observed. The combination of Schiff-reagent with a Methylene blue counterstain for demonstrating fungal elements and tissue structures is also described.


Shape change of blood platelets brought about by myelin basic protein and other basic polypeptides.

Basic proteins and polypeptides (BPP) such as myelin basic protein (MBP), polyornithine (M.W. 40,000), polylysine and protamine, which are known to cause neuronal depolarization in the central nervous system, induced a shape change reaction in blood platelets of various species, including man. This reaction was not accompanied by platelet aggregation or marked alterations of 5-hydroxytryptamine release. Cyclic nucleotide levels were also unchanged. The shape change induced by polyornithine was inhibited by heparin but not by antagonists of 5HT, catecholamines or gamma-aminobutryic acid, substances which are known to have no effect on the MBP-induced neuronal depolarization. Other basic substances, e.g. low molecular weight polyornithine (M.W. 4,000), cytochrome c, spermine and spermidine, did not induce either platelet shape change or (as shown before) neuronal depolarization. It is concluded, that 1) the shape change reaction of platelets seems to be a sensitive and simple means of detecting those BPP which induce functional changes in mammalian cells and 2) the use of platelets as models for neurons can be extended to include the action of BPP on the plasma membranes.


Longitudinal contraction of isolated guinea-pig ileum induced by rapid cooling.

1. Rapid change of bath temperature from 37 degrees C to 27 degrees C and vice versa caused longitudinal contraction of the isolated guinea-pig ileum. 2. Tetrodotoxin, tropicamide, noradrenaline, isoprenaline, morphine, and the met-enkephalin analogue FK 33-824 depressed the responses or accelerated the fade of the contraction induced by rapid cooling when added after the response had reached its maximum. 3. Hexamethonium had no influence on the responses. 4. Physostigmine potentiated all responses and reversed the fade of contraction induced by rapid cooling when added after this contraction had reached its maximum. 5. The effects of rapid cooling or warming were not altered in preparations made tachyphylactic to substance P; the response to rapid warming, but not cooling, was partially inhibited under tachyphylaxis to 5-hydroxytryptamine. 6. Antazoline, phentolamine, naloxone, and indomethacin did not block the responses. 7. Capsaicin firt potentiated and subsequently depressed the responses to both rapid cooling and warming. 8. The results indicate that rapid change of bath temperature induces longitudinal contraction by excitation of postganglionic cholinergic fibres.


Preferential replication of murine xenotropic type-C virus in human lymphosarcoma-derived cell lines.

Murine xenotropic virus, designated 698/X, was recovered by implantation of human lymphosarcoma-derived cells U-698M into nude mice of Giovanella's colony. The budded and extracellular particles revealed typical type-C morphology, the latter possessing reverse transcriptase (RT) activity and exhibiting a buoyant density 1.17 g/ml in sucrose gradient. In competitive radioimmunoassay using iodinated p30 of Rauscher MuLV, the 698X viral concentrate and cell extracts of both implanted lymphosarcoma cells (U-698M-N-1 and U-715M-N-1) were as effective as Gross MuLV, thus indicating the murine origin of the virus. The propagation of the 698/X virus in five human, four mouse (permissive for N- and B-tropic MuLV), two rat and one bovine cell lines was followed by RT, XC syncytia assays and EM investigations. The replication of the 698/X virus seems to be restricted mainly to both human lymphosarcoma-derived cell lines U-698M and U-715M. The new recovery of the virus from the nude mouse by implantation of U-175M cells has asserted its high tropism to human lymphosarcoma cells and its murine origin. The comparative response of mouse, rabbit and rat cells exposed to both NZB and 698/X xenotropic murine viruses exhibited host range differences between these viruses. The rabbit SIRC and rat embryonic cells REC were fully permissive for the murine xenotropic NZB virus, while low viral production was detected by RT assay only in 698/X virus infected SIRC cells.


Demonstration of alpha 2-pregnancy associated-glycoprotein (alpha 2-PGA) in serum of men with different localization of cancer.

By counterimmunoelectrophoresis in agarose gel the presence of alpha 2-PGA in the sera of 28 untreated patients with different localization of cancer was demonstrated. In 17 (60.7%) a precipitation line between the well with patient serum and that with monospecific anti-alpha 2-PGA serum, diluted 1 : 7 was obtained. At 1 : 7 dilution, with the sera from 28 healthy persons precipitation lines were not obtained. The determination of alpha 2-PAG can be made quickly and contributes to cancer diagnosis in men.


Combined regional chemotherapy and radiation therapy in the treatment of epidermoid carcinoma in the oro-facial region.

Followed treatment of oro-facial epidermoid carcinoma by combined chemo- and radiation therapy with eventual surgical intervention. Cytostatic drugs were applied intraarterially. After a monocytostatic treatment trial with Methotrexat (MTX), a combined cytostatic program was worked out utilizing two cytostatic drugs, viz. MTX and Bleomycin (BLM) in a cyclic alternation. Usefulness of chemotherapy and its inclusion into the treatment of epidermoid carcinoma in the oro-facial region was found justified principally in the case of combined therapy. However, the selected intraarterial mode of cytostatic administration is attended by certain drawbacks, mainly of a technical nature and is not suitable for routine application. For this reason, the authors have abandoned the i. a. mode of cytostatic administration in epidermoid oro-facial carcinoma and have adopted systemic application and this in combination with radiation, eventually surgical therapy with the utilization of chemotherapy.


Effect of transection on choline acetyltransferase, thyrotropin releasing hormone and substance P in the cat cervical cord.

The effects of a transection on the choline acetyltransferase activity, the thyrotropin releasing hormone and substance P contents in the cat cervical spinal cord have been investigated. Seven days after the hemitransection at the C1 level, the grey matter of the C6-7 levels of the spinal cord were dissected for the biochemical measurements. The choline acetyltransferase activity and the thyrotropin releasing hormone content remained unchanged in any regions in the grey matter following the high cervical transection. On the other hand, the substance P content was decreased by approx. 70% in the ventral horn. These results suggest that the fibers originating the supraspinal structures and terminating in the grey matter of the spinal cord, contain the substance P-releasing fibers, whereas there seems to be little cholinergic or thyrotropin releasing hormone-containing fibers.