Biomedical subjects

Walter Seelentag

Publications and source records attributed to Walter Seelentag.

5 recordsLinked to original sources

PubMed2006

An agarose matrix facilitates sectioning of tissue microarray blocks.

Tissue microarray (TMA) is a powerful, high-throughput technique for in situ investigation of biomarkers in many tissue samples in a paraffin block by immunohistochemistry or fluorescence in situ hybridization (FISH), and has rapidly become the standard in marker studies. One of the difficult steps in the procedure is the sectioning of array blocks and mounting of sections using special slides and/or adhesive-coated tape, which demands specific experience and is time-consuming. We report an arraying method that allows melting of the receiving paraffin block and subsequent sectioning like any ordinary paraffin-embedded tissue block. The major difference from the standard microarray technique is the use of an agarose matrix in the recipient block. The agarose matrix allows melting of the paraffin without disturbing the array, resulting in perfect integration of the tissue cores. The agarose-paraffin TMA blocks limit tissue core loss during cutting, mounting, or immunohistochemical or FISH staining and better maintains the array.

ErbB Receptors↗

PubMed2006

A cost-effective algorithm for hereditary nonpolyposis colorectal cancer detection.

Colorectal cancer with microsatellite instability (MSI) may occur sporadically or be inherited in cases of hereditary nonpolyposis colorectal cancer (HNPCC) syndrome. However, there is no consensus as to which patients must be tested and how to test MSI. In this study, MSI was tested by immunohistochemical analysis and by polymerase chain reaction in 148 cases of colorectal cancer, and methylation of the hMLH1 promoter was examined. MSI status was correlated with tumor phenotype. We found that localization, tumor infiltrating lymphocytes, and mucinous differentiation were predictive of high-frequency MSI (MSI-H) colorectal cancer and might be used to select cases for MSI analysis. Immunohistochemical analysis detected most MSI-H colorectal cancer and might constitute the first step in MSI detection. Absence of hMLH1 promoter methylation in MSI-H colorectal cancer could be predictive of hereditary colorectal cancer, and, hence, methylation analysis might constitute the second step in the identification of patients with HNPCC.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Immunohistochemical expression of endothelial markers CD31, CD34, von Willebrand factor, and Fli-1 in normal human tissues.

Few systematic studies have been published comparing the expression and distribution of endothelial cell (EC) markers in different vascular beds in normal human tissues. We investigated by immunohistochemistry the expression of CD31, CD34, von Willebrand factor (vWF), and Fli-1 in EC of the major organs and large vessels. Tissue samples obtained from autopsies and biopsy specimens were routinely processed and stained immunohistochemically for CD31, CD34, and vWF. Biopsy material was also stained immunohistochemically for Fli-1, D2-40, and Lyve-1. The expression pattern of the markers was heterogeneous in some of the organs studied. In the kidney, fenestrated endothelium of the glomeruli strongly expressed CD31 and CD34 but was only focally positive or completely negative for vWF. Alveolar wall capillaries of the lung strongly stained for CD31 and CD34 but were usually negative for vWF. The staining intensity for vWF increased gradually with the vessel caliber in the lung. Sinusoids of the spleen and liver were diffusely positive for CD31. They were negative for CD34 in the spleen and only expressed CD34 in the periportal area in the liver. Fli-1 was expressed in all types of EC but also in lymphocytes. D2-40 stained lymphatic endothelium only. Lyve-1 immunostaining was too variable to be applied to routinely processed tissues. The expression of EC markers CD31, CD34, and vWF in the vascular tree is heterogeneous with a specific pattern for individual vessel types and different anatomic compartments of the same organ. D2-40 labels lymphatic EC only.

Antigens, CD34↗

PubMed2004

Immunohistochemical localization of hTERT protein in human tissues.

Telomerase is a ribonucleoprotein complex mainly composed of a reverse transcriptase catalytic subunit (telomerase reverse transcriptase gene, hTERT) that copies a template region of its RNA subunit to the end of the telomere. For detecting telomerase activity in a tissue specimen the TRAP assay is a relatively sensitive and specific method, but it can be used only on fresh tissue extracts and offers no information at the single cell level. Immunohistochemistry (IHC) allows to detect hTERT protein expression at an individual cell level in human tissues. We have tested commercially available anti-hTERT antibodies in formalin-fixed and paraffin-embedded human tissues by IHC. Only one monoclonal antibody (NCL-hTERT; Novacastra) was sufficiently specific and this was applied to human tissues in which telomerase activity had been shown by TRAP assay and hTERT mRNA expression by RT-PCR. hTERT protein localized diffusely in the nucleoplasm and more intensely in the nucleoli of cancer cells and proliferating normal cells. Mitotic cells showed diffuse staining of the entire cell. Granular cytoplasmic staining was occasionally found in some tumor cells. In telomerase-positive tumors not all the tumor cells showed hTERT immunoreactivity. A significantly heterogeneous hTERT protein expression was observed in human tumor tissues. The hTERT immunostaining in fixed tissues was concordant with telomerase activity and hTERT mRNA expression in corresponding non-fixed samples. Quantitative RT-PCR of microdissected sections showed that hTERT mRNA expression was higher in cells with nuclear expression than in those with cytoplasmic expression. Double staining with the M30 antibody showed that a subpopulation of hTERT-negative cells is apoptotic. We conclude that: (1) hTERT protein can be detected by IHC in fixed human tissues, but the choice of the antibody, tissue processing, and reaction conditions are critical, (2) hTERT protein localizes in the nucleoplasm, more strongly in the nucleolus, and occasionally in the cytoplasm, (3) telomerase-positive tumors show significant heterogeneity of hTERT protein expression, and (4) a subpopulation of hTERT protein negative tumor cells is identified as apoptotic cells.

Adenocarcinoma↗

PubMed2002

Importance of tumor regression assessment in predicting the outcome in patients with locally advanced rectal carcinoma who are treated with preoperative radiotherapy.

BACKGROUND: Locally advanced rectal carcinoma has a poor prognosis. However, since the introduction of preoperative radiotherapy, the outcome of patients with rectal carcinoma has been reported to have improved. Nevertheless, to the authors' knowledge few data are available regarding the histopathologic response to radiotherapy as assessed on surgical specimens as a potential predictive factor for outcome. METHODS: To estimate the effect of radiotherapy on rectal carcinoma, the authors retrospectively reviewed the surgical specimens of 102 patients with T3-4, N0 or > or = N1 rectal carcinoma and 1 patient with T2 but N1 rectal carcinoma. All patients were treated preoperatively with a hyperfractionated accelerated radiotherapy schedule in a prospective protocol (Trial 93-01). Using a standardized approach, tumor regression was graded using a system that varies from Grade 1 (tumor regression Grade [TRG] 1) when complete tumor regression is observed to Grade 5 (TRG5) when no tumor regression is observed. RESULTS: Radiotherapy resulted in tumor downstaging in 43% of the patients. There were 2 pT1 tumors (2%), 21 pT2 tumors (20%), 66 pT3 tumors (64%), and 14 pT4 tumors (14%) after treatment. Regional lymph nodes were involved in 55 patients (53%). None of the patients demonstrated a complete tumor regression after radiotherapy, but in 79% of the specimens a partial tumor regression was observed (TRG1: 0%; TRG2: 20%; TRG3: 39%; TRG4: 20%; and TRG5: 21%). The median actuarial overall survival (OS) and disease-free survival (DFS) were 52 months. Actuarial local recurrence rates at 2 years and 5 years were 6.4% and 7.6%, respectively. Univariate analysis showed the actuarial DFS to be significantly lower in patients with lymph node metastases (P = 0.0004) and advanced pT stages (pT3-4) (P = 0.03). A favorable outcome for OS, DFS, and local control was observed in patients with TRG2-4 (i.e., responders) compared with patients with TRG5 (i.e., nonresponders), but also in patients with low residual tumor cell density (TRG2, 3, and 4). On multivariate analysis, TRG remained an independent prognostic indicator for local tumor control. CONCLUSIONS: Tumor regression as well as residual tumor cell density were found to be predictive factors of survival in rectal carcinoma patients after preoperative radiotherapy. Even after preoperative radiotherapy, the pathologic stage of the surgical specimen remained a prognostic factor. The use of a standardized approach for pathologic evaluation must be implemented to allow comparison between the results of various treatment approaches.

Adult↗