Immunohistochemical localization of hTERT protein in human tissues

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Serval ID
serval:BIB_9AD22B30EF18
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Immunohistochemical localization of hTERT protein in human tissues
Journal
Histochemistry and Cell Biology
Author(s)
Yan  P., Benhattar  J., Seelentag  W., Stehle  J. C., Bosman  F. T.
ISSN
0948-6143 (Print)
Publication state
Published
Issued date
2004
Volume
121
Number
5
Pages
391-397
Notes
PT - Comparative Study PT - Journal Article PT - Research Support, Non-U.S. Gov't
Abstract
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
Keywords
Adenocarcinoma/enzymology/genetics/Antibodies,Monoclonal/immunology/Apoptosis/Breast Neoplasms/Pathology/Cell Nucleolus/metabolism/Cell Nucleus/Colonic Neoplasms/Cytoplasm/DNA-Binding Proteins/Female/Gene Expression/Humans/Immunohistochemistry/methods/Keratins/analysis/Lung Neoplasms/Lymphocytes/Male/Neoplasms/RNA,Neoplasm/isolation & purification/Reverse Transcriptase Polymerase Chain Reaction/Sarcoma/Spermatogonia/Telomerase/Urinary Bladder Neoplasms
Pubmed
Web of science
Open Access
Yes
Create date
29/01/2008 19:36
Last modification date
01/10/2019 7:18
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