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Biomedical subjects

J Roth

Publications and source records attributed to J Roth.

At least 271 records · Page 15Linked to original sources
PubMed1994

[Pronounced flush symptoms in carcinoid syndrome without liver metastases].

The case of a 78 years old patient with impressive flush symptoms is reported. 5-Hydroxyindoleacetic acid and serotonin were elevated. The responsible pathomorphological structure is a 8 x 4.5 x 6 cm ileocaecal neoplasm. Pre-, intra- and postoperatively liver metastases could not be detected during a 18 month period. After surgery of the carcinoid flush disappeared, so that the tumour seize resulting in enormous secretion of metabolites with endocrine activity should be claimed responsible for causing this symptom.

Aged↗
PubMed1994

[Tumors--disorders of cell adhesion].

Cell adhesion molecules are of pivotal importance during organogenesis and fulfil important functions in maintaining normal organ architecture in the adult. Furthermore, they are involved in diverse and specialized functions such as migration and activation of leukocytes in immunosurveillance and inflammation, and homophilic as well as heterophilic interactions of cells with others and the extracellular matrix. In tumors, various cell adhesion molecules play a role in the complex process of tumor dissemination including metastatic spread. In addition, immunohistochemical detection of certain cell adhesion molecules in tumors has been shown to be of value in predicting the clinical course of the tumor disease. This short review focuses on the polysialic acid of the neural cell adhesion molecule NCAM, integrins and CD44.

Animals↗
PubMed1994

[Comparison of histology, immunohistochemistry, RT-PCR, in situ hybridization, and in situ RT-PCR for demonstration of hepatitis C virus in paraffin-embedded liver biopsies].

To compare immunohistochemical and molecular methods for the detection of hepatitis C virus (HCV) infection in archival liver biopsies we analyzed formalin-fixed and paraffin-embedded liver specimens of 10 patients with serologically confirmed HCV infection. Methods employed included indirect FITC-immunofluorescence, reverse-transcriptase polymerase chain reaction (RT-PCR) using extracted RNA and Southern blotting with chemiluminescence-based detection, non-radioactive in situ hybridization (ISH) with digoxigenin-labeled oligo- and cRNA probes, direct in situ RT-PCR with incorporation of labeled nucleotides into PCR-products, and indirect in situ RT-PCR using subsequent ISH for the visualization of intracellular PCR-products. Our results indicate that: (1) using the histological criteria described by Lefkowitch et al. (Gastroenterology 1993; 104-595) together with clinical data, most chronic HCV infections can be diagnosed by conventional histology, if liver biopsies are representative; (2) the commercially available mAB TORDJI-22 appears to cross-react with non-HCV epitopes; (3) molecular methods performed on routinely fixed and processed liver biopsies frequently yield false negative results due to sampling problems, low viral copy number and RNA degradation in infected cells; (4) analysis of HCV-RNA by RT-PCR of extracted total RNA is more sensitive than indirect in situ RT-PCR or ISH; and (5) direct in situ RT-PCR is not reliable despite the use of modifications such as DNase pretreatment and hot-start procedures. Further studies are required to define both optimal methods for sample processing and improvements of protocols, in order to increase detection sensitivity and specificity of HCV infection by immunohistochemical and molecular methods.

Biopsy↗
PubMed1994

Expression of murine VCAM-1 in vitro and in different models of inflammation in vivo: correlation with immigration of monocytes.

VCAM-1 (vascular cell adhesion molecule-1) is a cytokine-inducible adhesion molecule which is known to mediate adhesion of mononuclear cells to endothelial cells in vitro via binding to the integrin VLA-4 (very late antigen-4). To further elucidate the role and regulation of VCAM-1 in vivo, we compared in vitro and in vivo expression of VCAM-1 in response to cytokines and investigated immunohistochemically the expression of VCAM-1 in three murine models of experimental inflammation. These models differed with regard to the pathogenetic mechanism and the subsequent infiltrate: allergic contact dermatitis (ACD) to DNFB as a T cell-controlled, DTH type of inflammation, cutaneous infection with Leishmania major as a chronic granulomatous inflammation and the cauterized cornea as a model for acute inflammation. VCAM-1 was found to be markedly enhanced on vascular endothelia in all types of inflammation and after subcutaneous administration of LPS and TNF-alpha. Administration of IL-4, however, failed to induce VCAM-1 both in vivo and in vitro. The increased VCAM-1 expression in the inflammatory models correlated with the appearance of infiltrating monocytes/macrophages. A concomitant influx of CD4-positive/CD8-positive lymphocytes was only observed in ACD and Leishmaniasis.

Animals↗
PubMed1994

Detection of RET proto-oncogene point mutations in paraffin-embedded pheochromocytoma specimens by nonradioactive single-strand conformation polymorphism analysis and direct sequencing.

The suitability of formalin-fixed and paraffin-embedded tumor material was evaluated for molecular analysis of the RET proto-oncogene. We analyzed exons 10, 11, and 16 for point mutations in seven sporadic and six multiple endocrine neoplasia (MEN) 2A-associated pheochromocytomas by a nonradioactive single-strand conformation polymorphism assay followed by nonradioactive direct sequencing of PCR-amplified DNA using an automated DNA sequencer. All MEN 2A-associated pheochromocytomas contained a heterozygous missense germline mutation within cystine codons of the cysteine-rich extracellular domain encoded by exons 10 and 11. Mutations were located in codon 619 (TGC-->TCC; Cys-->Ser) in one, in codon 635 (TGC-->CGC; Cys--Arg) in three, and in codon 635 (TGC-->TAC; Cys-->Tyr) in two pheochromocytomas. No tumor-specific (somatic) mutations were detected in exons 10, 11, and 16 of the sporadic pheochromocytomas. These data support recent findings that germline point mutations that are clustered in distinct cysteine codons of the RET proto-oncogene are involved in the neoplastic phenotype of the MEN 2A syndrome. Our results demonstrate that both nonradioactive single-strand conformation polymorphism and direct sequencing are suitable methods to detect single base substitutions in DNA extracted from archival material.

Adolescent↗
PubMed1994

[A rare etiology for HBs-Ag negative acute hepatitis B--coinfection by hepatitis B and delta].

An unusual case of a 25-year-old male Italian is reported. The patient endured an acute hepatitis without detectable HBs-antigen by coinfection with hepatitis-B and Delta. Coincidently, a cured hepatitis-C was present. Firstly hepatitis-B-virus DNA could be demonstrated in a small quantity by serodiagnosis (6 pg/ml, hybridization technique). Subsequently, the identification of B-virus DNA was only possible in liver tissue (PCR-technique), but no longer by serodiagnosis. The probable enduring inhibition of hepatitis-B-virus replication by Delta virus resulted in a self limitation of the disease within 2 months (HDV-RNA negative, HBs-Ag and HBe-Ag negative; Anti-HBs negative, Anti-HBe and Anti-HBc positive). In spite of negativation of replication markers for hepatitis-B a subsequent reactivation of the infection was possible by viral material which persisted in liver tissue.

Acute Disease↗
PubMed1994

In vitro and in vivo growth of clonal sublines of human small cell lung carcinoma is modulated by polysialic acid of the neural cell adhesion molecule.

BACKGROUND: Polysialic acid (poly Sia) of the neural cell adhesion molecule (N-CAM) is an oncodevelopmental antigen and is found in small cell lung carcinomas (SCLC) as well as cell lines derived from these tumors. EXPERIMENTAL DESIGN: Cell heterogeneity in poly Sia expression was observed in primary SCLC and cell cultures of SCLC by immunostaining using a directly gold-labeled monoclonal antibody against poly Sia (MAb 735) and antibodies against N-CAM. Clonal sublines of the N-CAM-positive SCLC cell line, NCI-H69 were established to study the basis of this heterogeneity. The resulting sublines were examined for the proportion of cells expressing poly Sia, the stability of poly Sia expression, and the possible involvement of DNA methylation. Two of the sublines that expressed poly Sia on 0 and 95% of the cells were used in three independent in vitro assays to investigate the importance of poly Sia in cell-cell aggregation, disaggregation and cell to substrate adherence. Finally, clonogenic growth of these sublines was studied in soft agar and in the nude mouse. RESULTS: The proportion of cells immunoreactive for poly Sia was stable in serial subculture in these clones and was not affected by reducing DNA methylation. In aggregation and disaggregation assays, poly Sia was shown to modulate both calcium-dependent and independent cell-cell adhesion. No measurable differences in the attachment rates to various substrates (collagen type IV, laminin, heparan sulfate, and poly-L-lysine) were detected between the sublines. Cells from the poly Sia-positive clonal subline formed significantly more colonies in semisolid media and more intracutaneous metastasis in the nude mouse. CONCLUSIONS: Poly Sia does not occur on all N-CAM immunoreactive cells of SCLC. Poly Sia on SCLC cells is a clonable trait and high poly Sia expression correlates with reduced cell-cell adherence, a greater clonogenic ability in semisolid media, and a significantly higher incidence of intracutaneous metastases in nude mice.

Animals↗
PubMed1993

Cataract formation after bone marrow transplantation.

OBJECTIVE: To evaluate the incidence, time course, and factors associated with cataract formation in bone marrow transplant recipients. DESIGN: Prospective cohort study. SETTING: University Hospitals, Basel, Switzerland. PATIENTS: 197 patients treated with allogeneic or autologous bone marrow grafts at least 180 days before the start of the study. INTERVENTION: Three regimens for bone marrow transplant were used: 74 patients received single-dose, total-body irradiation (TBI), 90 patients received fractionated TBI, and 33 received chemotherapy alone. RESULTS: Three and one half years after single-dose TBI, 51 of the 74 patients (69%) were alive and cataracts had developed in all of these 51 patients. Cataracts developed in 18 of the 90 (20%) patients treated with fractionated TBI, with an 83% (95% CI, 63% to 100%) risk for lens opacification at 6 years. Cataracts developed in only 1 of the 33 (3%) patients treated with chemotherapy alone. Incidence of cataracts is higher and lens opacification occurs earlier after single-dose TBI than after fractionated TBI (P < 0.01). With Cox regression analysis, the use of irradiation (relative risk, 21.0), the mode of irradiation (relative risk, 7.4), and the use of steroid treatment (relative risk, 2.9) for more than 3 months after bone marrow transplantation increased the risk for cataract formation. In contrast, age, sex, and chronic graft-versus-host disease did not influence the rate of cataract development. The probability of requiring cataract surgery after 6 years was 85% (CI, 75% to 95%) for the patients treated with single-dose TBI and 20% (CI, 0% to 49%) for those prepared with fractionated irradiation. CONCLUSIONS: Patients treated with TBI, regardless of fractionation, are likely to have cataracts within 10 years, and some will need surgical repair. Long-term steroid treatment accelerates cataract formation. Preventive measures, such as lens shielding during TBI, should be considered.

Adolescent↗
PubMed1993

MRP8 and MRP14, S-100-like proteins associated with myeloid differentiation, are translocated to plasma membrane and intermediate filaments in a calcium-dependent manner.

MRP8 and MRP14 are two Ca(2+)-binding proteins of the S-100 family expressed by myelomonocytic cells. Both proteins assemble to noncovalently associated complexes in a Ca(2+)-dependent manner. Members of the S-100 family are known to play a role in cytoskeletal-membrane interactions; therefore, we investigated the subcellular distribution of MRP8/MRP14 and their complexes in human monocytes. Using differential centrifugation and subsequent Western blot or enzyme-linked immunosorbent assay analysis, we found that MRP8/MRP14 were almost completely translocated from the cytoplasma to membrane and cytoskeletal structures in a Ca(2+)-dependent manner. Using a cross-linking technique, complexed forms of MRP8/MRP14 were found to be associated with the plasma membrane. Analysis of MRP-transfected L132 cells showed that the MRP8 as well as the MRP14 component of the MRP8/MRP14 complex may independently bind to membrane and cytoskeletal structures. Furthermore, immunogold electron microscopy showed a colocalization of MRP8/MRP14 and the intermediate filament type III protein vimentin in A23187-treated monocytes. Our data indicate that, in analogy to other S-100-like proteins, MRP8 and MRP14 play a role in Ca(2+)-dependent cytoskeletal-membrane interactions. Restriction of MRP8/MRP14 expression to distinct stages of myelomonocytic differentiation suggests that these proteins are involved in highly specific pathways of intracellular signaling in phagocytes.

Antibodies, Monoclonal↗
PubMed1993

Expression of calcium-binding proteins MRP8 and MRP14 is associated with distinct monocytic differentiation pathways in HL-60 cells.

MRP8 and MRP14 are two calcium-binding proteins of the S-100 family which are expressed during distinct stages of monocytic maturation. To further investigate their regulation the human leukemic cell line HL-60 which can be induced to differentiate to monocytes/macrophages by 12-O-tetradecanoylphorbol 13-acetate (TPA), 1,25-(OH)2 Vitamin D3 (VD3), tumor necrosis factor-alpha (TNF-alpha) and interferon-g (IFN-g) were analyzed for expression of MRP8/MRP14. Employing Northern blotting, a sandwich enzyme-linked immunosorbent assay and immunocytochemical analysis we determined MRP8/MRP14 mRNA and protein levels, which were found to be elevated after VD3 and reduced after TPA treatment. TNF-alpha and IFN-g did not affect MRP8/MRP14 levels. Western blot analysis revealed that formation of MRP8/MRP14 to biologically active complexes which has previously been shown to be a calcium-mediated process is not dependent on the differentiation stages of HL-60 cells. Restriction of MRP8/MRP14 expression to only distinct pathways of monocytic differentiation in HL-60 cells may thus reflect different functional phenotypes of monocytes/macrophages in vivo.

Calcium-Binding Proteins↗
PubMed1993

A novel chicken homeobox-containing gene expressed in neurulating embryos.

We have isolated a cDNA clone from chicken embryo that contains a homeobox sequence (CHox-cad2). Analysis at the nucleotide and amino acid levels revealed closest similarity to the Xenopus Xcad1 and to other homeoboxes related to Drosophila caudal. RNA blot analysis showed hybridization of CHox-cad2 to two transcripts of 2.6 and 1.5 kb, present at day 1 of embryogenesis (E1). Using the highly sensitive polymerase chain reaction (PCR) to amplify cDNAs from embryonic RNAs from E0 (unincubated blastoderm) to E4, we confirmed the restricted expression of this homeobox sequence to the period of neurulation (E1).

Amino Acid Sequence↗
PubMed1993

Expression and complex assembly of calcium-binding proteins MRP8 and MRP14 during differentiation of murine myelomonocytic cells.

In the present study we analyzed expression and biochemical properties of the two recently cloned calcium-binding proteins MRP8 and MRP14, both members of the S-100 family, in murine myelomonocytic cells. Expression of MRPs was found to be restricted to granulocytes and distinct stages of macrophage differentiation when compared to the expression of other markers (Mac-1, F4/80) in transformed macrophage cell lines (M1, RMB.TG, WEHI.TG, J774A, P388D) and resident and exudate peritoneal and tissue macrophages. Similarly, mRNA and protein levels of MRP8/MRP14 in murine bone marrow cells decreased during culture in L cell-conditioned medium. Applying a cross-linking technique, the formation of noncovalently associated heteromeric MRP8/MRP14 complexes of 25, 35, and 48 kd was demonstrated. Our data suggest that MRP8/MRP14 expression and complexation are characteristic for granulocytes and distinct stages of macrophage differentiation in mice. Analysis of the MRP8/MRP14+ phenotype of macrophages may provide further insights into the mechanisms underlying macrophage differentiation.

Animals↗
PubMed1993

Cellular sialoglycoconjugates: a histochemical perspective.

Sialic acids are found at the non-reducing terminus of oligosaccharide side-chains of most glycoproteins and glycolipids. Their expression is often developmentally regulated during organogenesis and in differentiating cells. Further, differential expression of various sialyltransferases results in organ- and cell-type specific expression of sialylated glycosylation sequences. The availability of highly specific lectins and of monoclonal antibodies has permitted histochemical investigations of the various cellular and subcellular aspects of the process of sialylation. This review provides a survey on the developmentally regulated and organ- as well as cell-type specific expression of various sialoglycoconjugates and their tumour-associated occurrence with special reference to intestine and kidney.

Animals↗
PubMed1993

Use of Phaseolus vulgaris leukoagglutinating lectin in histochemical and blotting techniques: a comparison of digoxigenin- and biotin-labelled lectins.

An increase in the number of beta 1,6 branches of the trimannosyl core of asparagine-linked oligosaccharides has been shown to be directly correlated with the metastatic potential of cultured tumour cells. The Phaseolus vulgaris leukoagglutinating lectin (PHA-L) binds to beta 1,6 branches of tri- and tetra-antennary oligosaccharides. We have applied digoxigenin- and biotin-conjugated PHA-L to establish a non-radioactive detection system for beta 1,6 branches, which can be used in lectin blotting as well as light and electron microscopic cytochemistry. For this purpose the HCT116 human colon carcinoma cell line and colon carcinoma tissue were investigated. Digoxigenin-conjugated PHA-L in conjunction with alkaline phosphatase-conjugated anti-digoxigenin antibodies was superior to biotin-conjugated PHA-L in lectin blotting with respect to sensitivity and specificity. Similarly, the digoxigenin conjugated PHA-L in conjunction with gold-labelled anti-digoxigenin antibodies resulted in more intense specific staining and lower background compared to biotin-conjugated PHA-L visualized with a streptavidin immunogold complex. The specificity of lectin binding in blotting and cytochemical studies was demonstrated by the absence of staining when the lectin was omitted or preabsorbed with glycoprotein, and following pretreatment of the cellular homogenates or tissue sections by N-glycosidase F. Our results demonstrate that digoxigenin-conjugated PHA-L provides high sensitivity and specificity for histochemical and blotting techniques and is amenable for quantification. The technique should have applications in tumour research.

Biotin↗
PubMed1993

Adaptation of a non-radioactive in situ hybridization method to electron microscopy: detection of tenascin mRNAs in mouse cerebellum with digoxigenin-labelled probes and gold-labelled antibodies.

In this study we describe a method for the detection of mRNAs at the ultrastructural level using a non-radioactive in situ hybridization method based on digoxigenin-labelled cRNA probes and gold-labelled digoxigenin-specific antibodies. We applied this protocol to an analysis of the expression of the extracellular matrix protein tenascin in the developing cerebellar cortex of the mouse. To gain an impression of the sensitivity attainable with digoxigenin-labelled probes, we first established at the light microscopic level that the hybridization signal obtained with the non-radioactive probe is as sensitive as that obtained with a 35S-labelled probe. The non-radioactive hybridization protocol was then combined with electron microscopic post-embedding and immunogold detection techniques. Tenascin-specific, digoxigenin-labelled cRNA probes were hybridized to ultrathin sections of Lowicryl K4M-embedded tissue and the probe/target mRNA hybrids were detected using gold-labelled antibodies to digoxigenin. In agreement with the observations from in situ hybridization at the light microscopic level, specific labelling was observed in Golgi epithelial cells in the region of the Purkinje cell layer and cells in the internal granular layer, which could be identified as astrocytes by ultrastructural criteria. Labelling was detectable in association with free ribosomes and ribosomes of the rough endoplasmic reticulum. In addition, focal hybridization signals were occasionally found in the nucleus. No signal was observed in Golgi epithelial cells or astrocytes using sense or in any other cerebellar cell type using either sense or anti-sense probes. The described in situ hybridization technique uses ultrastructural criteria to associate the presence of a given mRNA species with a particular cell type. Additionally, it provides information about the target mRNA's subcellular distribution, thus offering the possibility to study intracellular transport of particular mRNAs.

Acrylic Resins↗
PubMed1993

Monoclonal antibodies that recognize the trisaccharide epitope Gal alpha 1-3Gal beta 1-4GlcNAc present on Ehrlich tumor cell membrane glycoproteins.

Monoclonal antibodies were prepared against the trisaccharide Gal alpha 1-3Gal beta 1-4GlcNAc, a sequence which occurs on the surface of Ehrlich ascites tumor cells as well as in thyroglobulin, laminin and a variety of other proteins. This was accomplished by immunizing BALB/c mice with the fraction of Ehrlich cell membrane glycoproteins obtained by affinity chromatography on a Griffonia simplicifolia I (GS I) column which selectively binds alpha-D-galactosyl-terminated structures. Detection of Gal alpha 1-3Gal beta 1-4GlcNAc-specific antibodies was accomplished by employing glycoproteins containing the trisaccharide sequence; fusion with spleen cells from an immunized mouse was accomplished in the presence of polyethylene glycol (PEG1500). An enzyme-linked immunosorbent assay (ELISA) system was used to identify two clones (2.10G and 6.8E), which recognized the desired trisaccharide conjugate. These clones also recognized a thyroglobulin fraction isolated by GS I affinity chromatography and murine laminin, both of which possess the Gal alpha 1-3Gal beta 1-4GlcNAc sequence. Inhibition of antibody-trisaccharide reactivity, examined employing an ELISA assay, revealed that two trisaccharides, Gal alpha 1-3Gal beta 1-4GlcNAc/Glc, were the best inhibitory haptens; Gal beta 1-4GlcNAc (LacNAc), Gal alpha 1-3Gal and Gal beta 1-4Glc (lactose) were poor inhibitors. Indirect immunofluorescence staining of unfixed Ehrlich cells using the monoclonal antibody at 4 degrees C revealed fluorescence over the entire cell surface. Indirect immunogold labeling of semithin and ultrathin sections of aldehyde fixed and Lowicryl K4M-embedded Ehrlich cells resulted in specific labeling of the cell surface and internal structure.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗