Microtubule-associated protein 2 (MAP2)-immunoreactive neurons in the retina of Bufo marinus: colocalisation with tyrosine hydroxylase and serotonin in amacrine cells

Summary Neuron populations in the retina of the toad, Bufo marinus, were labelled with a monoclonal antibody raised against microtubule-associated protein 2 (MAP2). A subpopulation of cones, probably corresponding to the blue-sensitive small single cones, large diameter amacrine cells in the most proximal row of the inner nuclear layer and some large ganglion cells in the ganglion cell layer were labelled. Double labelling experiments were carried out to establish the colocalisation of MAP2 with known putative transmitter substances of the anuran amacrine cells. MAP2 was colocalised in a subpopulation of serotonin-immunoreactive and in all tyrosine hydroxylase-immunoreactive amacrine cells. The results indicate, that the MAP2 content in the neurons of the anuran retina can be correlated with other well-defined neurochemical and/or physiological properties.On leave from Department of Zoology, Attlia József University, Szeged, Hungary     

Modulation of differentiation markers in human choriocarcinoma cells by extracellular matrix: on the role of a three-dimensional matrix structure

Abstract. During spontaneous or chemically induced differentiation human choriocarcinoma cells express typical characteristics of the normal differentiating trophoblast: 1) increased production of peptide and steroid hormones (chorionic gonadotropin, placental lactogen, estrogens, progesterone); 2) increased activity of cellular alkaline phosphatase; 3) morphological transition from cytotrophoblast to syncytiotrophoblast-like cells; and 4) arrested cell proliferation. Since the extracellular matrix is known to control gene expression we have examined the effects of different substrates composed of matrix macromolecules on the differentiation of BeWo choriocarcinoma cells. Matrices tested were; fibronectin, laminin, collagens type I and type IV, the basement membrane-like complex matrix Matrigel, and a complex matrix extracted from human term placenta. Irrespective of the type of molecule(s), it was consistently found that, whenever the matrix molecules were presented as threedimensional structures (as opposed to protein coatings on tissue culture plastic) the response of affected differentiation markers monitored was highly pronounced. Morphology was changed from monolayers to rounded colonies, cell proliferation was reduced, and the secretion of chorionic gonadotropin was increased up to tenfold. Heterogeneous effects were observed on progesterone secretion and on the activity of cellular alkaline phosphatase. Cell adhesion to matrix molecules, however, did not depend on the structure of the matrix. This study demonstrates that gene expression in these tumor cells can be modified by extracellular matrix and highlights that not only the presence of effector molecules in the matrix but also the three-dimensional structure of the matrix is important for the induction of differentiation.     

Fast one-step procedure for the detection of nucleic acids in situ by primer-induced sequence-specific labeling with fluorescein-12-dUTP.

We provide fast, simple, one-step procedures for sequence-specific detection of nucleic acids in situ. Tandem repeat sequences in DNA are stained within 30 min, and mRNA is stained within 2 h. The procedures are based on the incorporation of the newly available fluorescein-labeled dUTP into DNA synthesized in situ by primed in situ labeling, with denatured fragments of cloned DNA or oligonucleotides as primers. The extreme speed and simplicity of the reaction make it attractive for automatization in routine laboratory procedures and opens up new diagnostic possibilities.     

Analysis of proliferative activity in colorectal mucosa by immunohistochemical detection of proliferating cell nuclear antigen (PCNA). Methodological aspects and application to routine diagnostic material.

Immunohistochemical detection of proliferating cell nuclear antigen (PCNA) has been suggested as a new approach for determining proliferative activity in paraffin-embedded tissue. In a prospective study PCNA immunostaining was performed in 284 colorectal biopsies using monoclonal antibodies 19F4 (Ogata et al. 1987) and PC10 (Waseem and Lane 1990) and compared with the Ki67 method. From each site three biopsies were taken and a variety of fixation regimens for frozen and paraffin-embedded samples tested. For frozen biopsies methanol fixation at -20 degrees C proved best. In paraffin sections PCNA could be detected after methacarn fixation as well as after controled fixation at 4 degrees C in 4% paraformaldehyde for 1 h and in most biopsies routinely fixed with 10% formalin. However, the latter fixation regimens revealed additional PCNA-positive cells in the normal superficial colonic mucosal epithelium. Although the percentage of cells positive for PCNA was generally lower than for Ki67, the rates correlated in a highly significant fashion, both in frozen methanol-fixed biopsies, and in paraformaldehyde-fixed paraffin-embedded samples. PCNA immunohistochemistry revealed a similar proliferative activity in different parts of the large bowel. A higher proliferative activity was found in inflamed mucosa, adenomas, carcinomas and even in normal mucosa from patients with colorectal neoplasms. In routinely fixed biopsies, the monoclonal antibody PC10 was superior to 19F4 because of considerably less background staining. However, in the routine material only a rough estimate of the proliferative activity was possible by PCNA immunohistochemistry using these antibodies, because unpredictable numbers of non-S-phase cells were also stained.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serial studies of autologous antibody reactivity to squamous cell carcinoma of the head and neck

Summary In previous studies we evaluated the incidence and specificity of autologous antibody reactivity against squamous cell carcinoma of the head and neck (SCCHN). We were able to demonstrate that autologous antibody reactivity is present in native sera but was usually of too low a titer to allow further analysis. Dissociation of immune complexes by acidification and ultrafiltration of serum augmented autologous antibody reactivity in nine out of nine autologous systems tested. Native antibody and antibody derived from immune complexes produced by the host and reactive with autologous tumor cells may be directed against physiologically relevant antigens. Therefore, correlations of antibody titers with clinical course may provide insight into the nature of the host response to cancer. In the present analysis, serological studies of six patients with SCCHN were performed with serum samples obtained over many months. Results of serial serological assays were correlated to tumor progression and clinical course. Fluctuations in autologous antibody reactivity were noted over time. In four cases, rises in autologous antibody titers preceded the clinical diagnosis of recurrence by several months. Drops in autologous antibody reactivity were noted in two cases following surgery or radiation therapy. In two cases of long-term survivors, no correlation between antibody reactivity and clinical course was noted. Specificity analysis of the six autologous systems demonstrated reactivity against autologous and allogeneic SCCHN as well as melanoma cell lines. These sera did not react with glioma, neuroblastoma, renal cell, breast, bladder and colon carcinoma cell lines nor with fetal calf serum, pooled lymphocytes, red blood cells and platelets. Autologous serial serological studies may provide a means by which to evaluate the host/tumor relationship in patients with SCCHN.     

Differences in anti-oncogene p53 expression in human monocytes and lymphocytes in vitro]

The p53 gene has been associated with malignant transformation as well as "anti-oncogene" activity. In the present report expression of p53 in resting and activated human blood monocytes and lymphocytes is analyzed. It is found that human monocytes freshly isolated by continuous percoll gradient centrifugation contained detectable level of p53 mRNA. Stimulation of monocytes by potent activation inducer Staphylococcus Aureus Cowan I for 3-5 hr caused disappearance of r53 mRNA. In contrast, induction of high level of TNF-alpha mRNA was detected. Addition of cycloheximide had no effect on p53 mRNA content in stimulated monocytes, and caused disappearance of mRNA in resting cells. In lymphocytes cultures p53 mRNA was absent in freshly isolated cells and in resting lymphocytes cultured for 20 hr. Activation of lymphocytes by lectin caused accumulation of p53 mRNA. We suggest that r53 gene regulation and functions might be different in human monocytes and lymphocytes.     

Use of commercial highly sensitive tests for detecting HbsAg in healthy persons]

The study has revealed the possibility of the contamination of serum with HBsAg under laboratory conditions during its treatment and preparation for analysis, which may be the cause of false positive results of HBsAg detection by the enzyme immunoassay (EIA). Excluding the factors of contamination, the authors demonstrate the efficacy of commercial systems, such as the passive hemagglutination test system "Gorky" and the EIA systems "Gorky" and "DIAplus", for the detection of HBsAg in the blood of donors in Moscow and Tashkent.     

The phages of halophilic vibrios and their use]

The range of the lytic activity of 46 phages of parahemolytic vibrios isolated from lysogenic strains, sea water samples, crabs and mussels has been studied. The phages are represented by virions belonging to morphological groups II, IV, V according to the phage classification currently used in Russia and to different serological groups. No relationship between the sensitivity of vibrio strains to the phages under study and the specificity of serotypes O and K has been established. The preparation of diagnostic phage [see text] suitable for the identification of 82% of strains of parahemolytic vibrios has been proposed.