Biomedical subjects

F Hofstaedter

Publications and source records attributed to F Hofstaedter.

At least 37 records · Page 2Linked to original sources

PubMed1995

Microsatellite instability: new aspects in the carcinogenesis of colorectal carcinoma.

Very recently a new molecular mechanism in the tumorigenesis of colorectal carcinoma has been described which is closely linked to hereditary non-polyposis colonic cancer (HNPCC). Ubiquitous changes in the length of simple repetitive DNA sequences between constitutional and tumour DNA occur in about 90% of cases of HNPCC and in about 15% of cases of non-familial, sporadic colorectal carcinoma. Such microsatellite instabilities have been shown to be the phenotypical marker of mutations in the human homologues of prokaryotic mismatch repair genes (MutS, MutL, MutH). These data provide crucial new tools in the detection of patients at high risk of developing colon cancer and other HNPCC-related carcinomas. In addition, these developments provide new insights into a new, presumably primary event in oncogenesis, i.e. the occurrence of mutations in genomic stability genes leading to an increased cellular mutation rate ("mutator phenotype") and thus to cancer.

Carcinogens↗

PubMed1995

Mechanisms of shockwave action in the human kidney.

The effects on the human kidney parenchyma of high-energy shockwaves (HESW) with different energy densities were examined. Kidneys of patients treated by radical nephrectomy for renal cell carcinoma were perfused with cold HTK solution immediately after nephrectomy and kept in hypothermia (8 degrees C) for a maximum of 4 hours. The tumor-free parenchyma was treated with 2000 shocks at energy outputs of 15 kV (16 MPa, 0.15 mJ/mm2), 17 kV (32 MPa, 0.25 mJ/mm2), 19 kV (50 MPa, 0.4 mJ/mm2), and 21 kV (65 MPa, 0.6 mJ/mm2) in an experimental electromagnetic shockwave system (Siemens Co., Erlanger, Germany). Resulting tissue effects were analyzed by histologic and immunohistochemical examinations and confocal laser scanning microscopy. Different sensitivities of cell components, blood vessels, and tubules were found. Laser scanning microscopy revealed nuclear alterations in the vicinity of the focus up to a distance of approximately 10 mm. Severe histologic changes were found in a smaller zone, while immunohistochemistry studies revealed negative collagen IV staining in an area of approximately 4 x 4 mm (all distances measured within the plane perpendicular to the acoustic axis). From these results, it can be concluded that HESW directly damage the tubules and the vascular system, which might explain the clinical changes after extracorporeal shockwave lithotripsy in human patients. The extent of these effects seems to be dependent on the applied energy.

Carcinoma, Renal Cell↗

PubMed1995

Treatment of human renal cell carcinoma with high-energy shock waves--a new in vivo/in vitro model.

The effects of high-energy shock waves (HESW) on the human renal cell carcinoma were examined. The kidneys were available from 32 patients treated by radical nephrectomy due to renal cell carcinoma. Immediately after nephrectomy the kidneys were perfused with cold HTK solution and stored for a maximum of 4 h in hypothermia at 8 degrees C. The tumors were treated with 4,000 shocks (65 mPa = 0.6 mJ/mm2) in an electromagnetic lithotriptor (Siemens Co., Erlangen, Germany). Microscopic and immunohistological examinations of the tumors were performed after treatment, and cell proliferation rates of treated and untreated specimens were analyzed by cell cultures in 10 cases. HESW induce severe microscopic damage in the tumor tissue as complete rupture of the vessel walls and destruction of the tubular-formed tumor masses in the focal area. Immunohistochemistry shows intact immune reactive endothelial cells by factor 8-associated antibodies until the border to histological damage. Around this region a zone of negative antibody reaction against collagen type 4 is found. In cell cultures the proliferation rates of treated specimens were significantly lower compared to untreated. The human renal cell carcinoma seems to be susceptible for treatment with shock waves. HESW induce direct damage of tumor cells and vascular damage in the tumor which may be the primary cause of tumor necrosis.

Aged↗

PubMed1995

In vitro and ex vivo expression of nucleolar proteins B23 and p120 in benign and malignant epithelial lesions of the prostate.

The expression of two specific nucleolar antigens, p120 and B23, has been investigated in the prostatic carcinoma cell line LNCaP as well as in 40 frozen and 40 formalin-fixed tissue samples of benign and malignant prostatic lesions (15 benign hyperplasias, 5 grade 1, 15 grade 2, and 5 grade 3 carcinomas). In vitro, immunoreactivity of p120 was confined to nucleoli of proliferating cells, with virtually no negative staining during S and G2/M phases. Unlike p120, B23 was expressed in the nucleoli of all LNCaP cells independently of growth and cell cycle phases. Hence, B23 was detectable in all stromal as well as in normal and malignant epithelial prostatic cells, both in fresh and in formalin-fixed tissue sections after microwave treatment. In contrast, the immunoreactivity of p120 was almost completely restricted to the nucleoli of prostate carcinoma cells: frozen sections of benign prostatic hyperplasia (n = 15) were either totally negative for p120 (n = 13) or had a low percentage of positively stained cells (labeling index = 3.3% in 3 cases). In the carcinoma group 76% (19/25) of the specimens were p120 positive, and there was a significant rise of labeling index from 18.1% in grade 1 to 82.2% in grade 3 carcinomas (P < 0.001). In contrast to B23, p120 could not be reliably demonstrated in formalin-fixed and paraffin-embedded tissue. We therefore conclude that anti-B23 is a general marker of nucleoli, whereas expression of p120 appears to correlate with "hyperactivity" of the nucleolus and provides a new tool for flow cytometrical and immunohistochemical assessment of nucleolar activity in tumor pathology.

Adenocarcinoma↗

PubMed1994

Flow cytometric detection and quantitation of the epidermal growth factor receptor in comparison to Scatchard analysis in human bladder carcinoma cell lines.

The epidermal growth factor receptor (EGFR) is considered a tumor-related marker with potential diagnostic and prognostic value. In order to assess the sensitivity of flow cytometry to detect EGFR and to quantify receptors objectively, two human bladder carcinoma cell lines with different urothelial differentiation, RT4 and J82, were grown in vitro, and their membrane EGFR content was measured by flow cytometry. Exponential monolayers showed decrease of EGFR content after 20 min pulses with 10 ng/ml EGF in medium, as detected with the antibody EGFR1 in a double staining technique with propidium iodide for DNA evaluation. Further decrease of green fluorescence intensity was seen in cells constantly exposed to EGF. Absolute receptor numbers were determined by Scatchard analysis with radioactive EGF and resulted in relatively low receptor numbers for both cell lines (approximately 3-4 x 10(4) EGFR/cell), as well as one affinity class. These findings could be matched by absolute receptor quantification by flow cytometry, adding beads with defined antigenic sites (Quantum Simply Cellular, Microbead Corporation) to the cell suspension for staining. Our data suggest that flow cytometric EGFR detection and quantitation may be supplied to in vivo tumor samples and that measurements by multiparameter analysis may define subpopulations valuable for tumor diagnosis and judgment on tumor progression.

Biomarkers, Tumor↗

PubMed1994

The genomic structure of the human AP-2 transcription factor.

The transcription factor AP-2 is encoded by a gene located on chromosome 6 near the HLA locus. Here we describe the genomic organization of the AP-2 gene including an initial characterization of the promoter. We have mapped two mRNA initiation sites, the entire exon-intron structure and located two polyadenylation sites. The mature AP-2 mRNA is spliced from 7 exons distributed over a region of 18 kb genomic DNA. A recently cloned inhibitory AP-2 protein is generated by alternative usage of a C-terminal exon. The proline-rich transactivation motif is encoded by a single exon within the N-terminal region in contrast to the complex DNA binding and dimerization motif which involves amino acid residues located on four different exons. The sites of mRNA initiation are located 220 and 271 bases upstream from the ATG translation start site. Although the promoter contains no canonical sequence motifs for basal transcription factors, such as TATA-, CCAAT- or SP-1 boxes, it mediates cell-type-specific expression of a CAT reporter gene in PA-1 human teratocarcinoma cells and is inactive in murine F9 teratocarcinoma cells. We demonstrate that the promoter of the AP-2 gene is subject to positive autoregulation by its own gene product. A consensus AP-2 binding site is located at position -622 with respect to the ATG. This site binds specifically to bacterially expressed AP-2 as well as to multiple proteins, including AP-2, present in PA-1 and HeLa cell nuclear extracts. A partial AP-2 promoter fragment including the AP-2 consensus binding site is approximately 5-fold transactivated by cotransfection of an AP-2 expression plasmid.

Animals↗

PubMed1994

Proliferation of tumor spheroids after shock-wave treatment.

Multicellular tumor spheroids (MCTS) grown from the bladder cancer cell line RT112 and from the prostate cancer cell line PCA were exposed to 200 or 800 electromagnetically generated focused ultrasound shock waves. RT112 cells showed a distinct but transient decrease in proliferation whereas the effect of PCA cells was less pronounced. Flow-cytometric measurements of DNA content and Ki67 expression revealed no significant changes in the cell cycle distribution. The capacity of RT112 cells exposed to 800 shock waves to re-grow as MCTS was markedly decreased, indicating an alteration of intercellular adhesion.

Cell Division↗

PubMed1994

[Molecular cloning of a new AP-2 transcription factor, AP-2beta, and its function in cell differentiation].

Transcription factor AP-2 has been previously shown to play an important function in embryonal development and cell differentiation. We have investigated the possibility that AP-2 function in embryonic development is exerted by a multigene family of AP-2 related transcription factors. Here we describe the molecular cloning of such an AP-2 related gene, AP-2 beta, and prove that it encodes for a functional transcription factor. In situ hybridizations of murine embryo sections revealed a temporally restricted and tissue-specific expression pattern that indicates a function of AP-2 beta in the development of the midbrain in the differentiation of sensory neurons for taste, olfaction and palpation.

Animals↗

PubMed1993

Cloning and characterization of a fourth human somatostatin receptor.

We have isolated a gene coding for a fourth human somatostatin (somatotropin release-inhibiting factor) receptor. This additional somatostatin receptor (hSSTR4) is specifically expressed in human fetal and adult brain and lung tissue. The deduced amino acid sequence of the receptor displays both sequence and structural homology to three cloned somatostatin receptors as well as to other members of the family of GTP-binding-protein-coupled seven-helix transmembrane-spanning receptors. Pharmacological characterization of the expressed receptor reveals specific, high-affinity binding of somatostatin 14 and somatostatin 28. Surprisingly, several well-characterized synthetic somatostatin analogs fail to exhibit high-affinity binding to hSSTR4, indicating the existence of pharmacologically different receptor subtypes. Our data suggest that the diverse biological effects exerted by somatostatin are mediated by a family of receptors with discrete patterns of expression and different pharmacological properties.

Amino Acid Sequence↗

PubMed1993

Effects of high-energy shock waves on the viable human kidney.

Between September 1990 and July 1991, we treated 17 patients with renal-cell carcinoma by radical nephrectomy and two patients with urothelial carcinoma of the kidney pelvis by ureteronephrectomy. Immediately after nephrectomy, perfusion of the kidneys with cold HTK solution was performed and the organs were kept in hypothermia of 8 degrees C. The tumor-free parenchyma of the kidneys was treated 4 h later with shock waves of different energy levels in an experimental shock-wave system (Siemens Company, Erlangen). Light microscopy and examinations by scanning laser microscopy were performed after treatment. High-energy shock waves (HESW) produce significant changes in the tubulary and blood-vessel system of the viable human kidney, depending on the energy applied. Although our model is limited by hypothermia of the explanted kidneys, the effects of shock waves on the organs can be studied. Our model is suitable for testing the effects of different lithotriptors on the human kidney.

Aged↗

PubMed1993

Determination of the energy-dependent extent of vascular damage caused by high-energy shock waves in an umbilical cord model.

To determine the spatial extent of shock-wave-induced vascular damage human umbilical cords were exposed to electromagnetically generated, focused ultrasound waves of different energy densities. During treatment macroscopically visible hematoma and superficial holes appeared. Following exposure specimens were fixed and examined histologically. In addition to vessel wall necrosis and rupture, complete detachment of endothelial cells in defined regions was observed. A correlation of the extent of the damage with the energy density distribution revealed that a local energy density of 0.3 mJ/mm2 is the lower threshold for the occurrence of severe vascular damage.

Biophysical Phenomena↗

PubMed1993

Proliferating cell nuclear antigen in normal urothelium and urothelial lesions of the urinary bladder: a quantitative assessment using a true color image analysis system.

To evaluate proliferating cell nuclear antigen (PCNA) staining for assessing proliferative activity in routine pathology specimens of urinary bladder, the bladder carcinoma cell line J82 and a total of 122 specimens of normal bladder and urothelial lesions were stained with the antibody clone PC10 against proliferating cell nuclear antigen. In in vitro plateau cultures the proportion of PCNA-positive cells exceeded that of Ki-67-positive cells, and only very few cells were negative. In formalin-fixed tissues, the PCNA staining pattern, which should be confined to replicon units in the nucleus, was optimized by 1 h postfixation in an organic solvent (methacarn). Sections showed positive nuclear staining confined to basal and some suprabasal cells in normal urothelium and grade 1 dysplasias, but more generalized nuclear staining in all other neoplastic lesions. In addition, stromal cells adjacent to invasive tumors showed nuclear positivity in some instances. Using quantitative true color image analysis of sections counterstained with hemalum, the degree of brown staining of the PCNA reaction product is contrasted with the blue staining of the nuclear area. With this method low contrast specific staining not appreciated optically can be reliably detected. Image analysis data confirmed observations made on noncounterstained sections and showed significant differences between grade 1 and 2 dysplasias as well as between grade 1 dysplasia and all grades of papillary tumor. Furthermore, a significant difference in PCNA staining indices was found between grade 1 and 3 bladder carcinomas. The results indicate that PCNA staining using the PC10 antibody is not confined to the proliferative fraction of neoplastic urothelium.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Neoplasm↗

PubMed1993

[In vivo and ex vivo expression of nucleolar proliferation-associated antigens (p120, B23) in the prostate].

Expression of two nucleolar antigens, p120 and B23, was studied in a prostatic carcinoma cell line (LNCaP) and in frozen and paraffin embedded tissue sections of 40 benign and malignant prostatic lesions. The percentage of p120 negative G0/G1 phase cells rose significantly during transition from exponential to plateau growth phase in vitro (from 9% to 32%). In contrast, B23 was equally expressed throughout different cell cycle and growth phases. Thus, nucleoli of almost all stromal and epithelial cells were stained by B23 in tissue sections. P120, however, selectively stained nucleoli of proliferating prostatic epithelium. Whereas 88% (13/15) of benign hyperplasia were p120 negative this was the case in only 24% (6/25) of carcinomas. Using microwave procedure both MoAbs reacted in paraffin sections, but the percentage of p120 negative cases doubled. A routine application to formalin fixed and paraffin embedded tissue cannot be recommended thus far.

Cell Cycle↗

PubMed1993

[Differential regulation of androgen receptor promoter in hormone-sensitive and -insensitive prostate carcinoma cells].

The sensitivity of prostate epithelial cells to androgens is mediated by the androgen receptor (AR). Analyses of cell lines in vitro and prostate carcinomas in vivo provide evidence that loss of AR expression parallels the loss of sensitivity to androgens. The aim of our work is to analyze the genetic control elements of the AR gene and answer the question of why these elements fail to function in hormone-insensitive prostate cancer cells. Therefore, we have cloned and fully sequenced a 5700 base pair sequence 5'-upstream of the AR gene. This DNA fragment contains promoter and enhancer elements that are active in androgen-sensitive but inactive in androgen-insensitive prostate cancer cell lines.

Androgens↗

PubMed1993

[Multiparameter analysis using flow cytometry as additional tool for bladder cancer diagnosis].

In order to find additional ways to classify bladder cancer, multiparameter analysis with antibodies against urothelial associated-glycoproteins (UAGAb: Uro1, -5, -9, -10; Signet) and cytokeratins (CKAb: KL1, Immunotech) were used in parallel with DNA staining. Single cell suspensions of 21 bladder cancer specimens (4pTaG1, 9pTaG2, 1pT1G2, 2 > pT2G2, 5 > pT2G3) were stained. Preliminary data showed that the proportion of UAGAb positive cells have to be related to the pan-urothelial marker Uro5, since percentage of urothelial cells was variable (30-97%). Phenotypic differences found in different stages of tumor will be described. Selection of tumor cells by UAG did result in higher precision to determine tumor S-phase fraction, and helped select tetraploid tumors. The methodology is best applicable to pTa and pT1-tumors and prospective analysis of these tumors has started.

Antigens, Neoplasm↗

PubMed1993

[Molecular biological mechanisms in prostatic neoplasms].

An overview is given on three topics of prostate carcinoma: 1. New data on genetic factors in the pathogenesis, 2. molecular activation mechanisms in metastasis and 3. endocrine dedifferentiation during the course of the disease causing androgen independent state. Recent epidemiological analyses have clearly shown, that a specific part of prostate cancer is inherited. By DNA-interphase analyses specific allelic losses have been demonstrated. Metastatic progression is both associated with oncogene ras activation and chromosomal deletions. Endocrinological dedifferentiation cannot be simply explained by an overgrowth by androgen-receptor negative tumor cells, but by a more complex deregulation with strong influences by stromal-epithelial interactions. Downregulation of the promoter region of the androgen receptor may play a crucial role in the development of "endocrine deafness".

Chromosome Deletion↗

PubMed1993

[Distribution pattern of urothelial associated glycoproteins in human bladder cancer cell lines in in vitro culture].

The distribution of four antibodies against urothelial-associated glycoproteins (UAGAb: Uro1, -5, -9, -10, SIGNET) was studied by dual parameter flow-cytometry (UAGAb and DNA staining) in normal non-urothelial cancer cell lines (2), normal urothelial cell lines (2) and bladder cell lines (5). When cell lines were compared as plateau monolayers, high proportions of Uro1 and Uro10 were found in all cell lines, in contrast to Uro5 and Uro9, both showing specificity for urothelial cells. Though being described as a pan-urothelial marker, Uro5 is not found in all cell lines. In two cell lines (RT4 and J82) three different growth states (exponential and plateau monolayer and multicellular spheroids were compared, and dependence on spatial configuration could be shown for Uro9. Cell cycle relation of expression was found for Uro1 and Uro10.

Cell Line↗

PubMed1993

[AP-2: a nuclear effector of malignant transformation by ras oncogene].

We have applied a series of cell clones established from the human teratocarcinoma cell line PA-1 to study the effect of malignant transformation by ras-oncogenes on the regulation of cell growth and differentiation. A particular aim of this study was to identify nuclear gene-regulatory factors that are affected by ras-transformation. We show that a key nuclear target of ras is the transcription factor AP-2. AP-2 function is inhibited through the ras-controlled signal transduction cascade by at least two different mechanisms, i.e. inhibition of an AP-2 coregulatory factor and by expression of an alternatively spliced inhibitory AP-2 protein.

Alternative Splicing↗