Biomedical subjects

Sabine Herterich

Publications and source records attributed to Sabine Herterich.

8 recordsLinked to original sources

PubMed2006

Exclusion/confirmation of ataxia-telangiectasia via cell-cycle testing.

Ataxia telangiectasia (AT) is an autosomal recessive multisystem disorder with increased radiosensitivity and cancer susceptibility. The responsible gene (ATM) consists of 66 exons and a coding region of 9171 bp which precludes direct sequencing as a screening assay for confirmation or exclusion of the clinical suspicion of AT. Peripheral blood mononuclear cells of 330 patients referred for the exclusion of AT were exposed to ionizing radiation (IR) and incubated for 72 h in the presence of phytohemagglutinin. Using bivariate BrdU-Hoechst/ethidium bromide flowcytometry, the following cell cycle parameters were ascertained: (1) proportion of non-proliferating (G0,G1) cells as a measure of mitogen response, (2) proportion of first-cycle G2-phase cells relative to the growth fraction (G2/GF) as a measure of radiosensitivity. Of the cases tested, 94.2% could be unequivocally assigned either to the AT-negative or the AT-positive group of patients. Of the AT-positive cases, 11 were confirmed by ATM mutation analysis. Nineteen cases presented with non-conclusive results, mostly due to poor mitogen response; however, a combination of cell-cycle data with serum AFP concentrations led to the exclusion of AT in all but two of the uncertain cases. Substitution of ionizing radiation by the radiomimetic bleomycin was additionally tested in a small series of patients. We conclude that cell-cycle testing complemented by serum AFP measurements fulfills the criteria as a rapid and economical screening procedure for the differential diagnosis of juvenile ataxias.

Adolescent↗

PubMed2006

Prenatal exclusion/confirmation of Fanconi anemia via flow cytometry: a pilot study.

OBJECTIVE: To explore the potential of flow cytometry in the prenatal exclusion or confirmation of Fanconi anemia (FA). METHODS: Indications for prenatal diagnosis were (1) FA-negative family history, but suspicious ultrasound findings such as radial ray aplasia, (2) FA-positive family history, but without knowledge of the affected gene and/or mutation. Amniotic fluid (AF) cell cultures and umbilical cord (UC) blood cultures were assayed for typical cell cycle changes (G2-phase accumulations) without and with mitomycin C (MMC) treatments using single- and dual-parameter (BrdU-Hoechst) flow cytometry. RESULTS: Single-parameter flow cytometry correctly identified 2 positive and 9 negative cases on the basis of MMC sensitivity of cultivated AF cells. Likewise, 8 negative and 2 positive cases were correctly predicted using bivariate flow cytometry of 72-hour UC blood cultures. In contrast, bivariate flow cytometry applied to AF cells grown in the presence of bromodeoxyuridine (BrdU) yielded false-positive and false-negative results. CONCLUSIONS: Single-parameter flow cytometry of AF cell cultures and bivariate flow cytometry of UC cell cultures have the potential to correctly predict the affected status in cases at risk for FA, whereas bivariate flow cytometry proved unreliable when applied to BrdU-substituted AF cell cultures. Cases with a low a priori risk (e.g. sonographic finding of radial ray abnormalities and negative family history) would benefit most from flow cytometry as a rapid and economical prenatal screening procedure.

Amniocentesis↗

PubMed2006

Analysis of SAGE data in human platelets: features of the transcriptome in an anucleate cell.

A comprehensive SAGE (serial analysis of gene expression) library of purified human platelets was established. Twenty-five thousand (25,000) tags were sequenced, and after removal of mitochondrial tags, 12,609 (51%) non-mitochondrial-derived tags remained, corresponding to 2,300 different transcripts with expression levels of up to 30,000 tags per million. This new, highly purified SAGE library of platelets is enriched in specific transcripts. The complexity in terms of tag distribution is similar to cells that are still able to replenish their mRNA pool by transcription. We show that our SAGE data are consistent with recently published microarray data but show further details of the platelet transcriptome, including (i) longer UTR regions and more stable folding in the enriched mRNAs, (ii) biologically interesting new candidate mRNAs that show regulatory elements, including elements for RNA stabilization or for translational control, and (iii) significant enrichment of specific, highly transcribed mRNAs compared to a battery of SAGE libraries from other tissues. Among several regulatory mRNA elements known to be involved in mRNA localization and translational control, CPE elements are in particular enriched in the platelet transcriptome. mRNAs previously reported to be translationally regulated were found to be present in the library and were validated by real-time PCR. Furthermore, specific molecular functions such as signal transduction activity were found to be significantly enriched in the platelet transcriptome. These findings emphasize the richness and diversity of the platelet transcriptome.

3' Untranslated Regions↗

PubMed2005

A NOS-III haplotype that includes functional polymorphisms is associated with bipolar disorder.

The pleiotropic messenger molecule nitric oxide (NO) has been implicated in a variety of higher CNS functions, including learning, memory, and emotionality. In the human brain, NO is predominantly formed by neuronal NO synthase (NOS-I), while the so-called 'endothelial' isoform NOS-III also contributes to NO generation. We recently reported that NOS-III knockout mice display decreased adult neurogenesis and reduced responsiveness in a learned helplessness paradigm. To examine whether NOS-III plays a role in affective disorders as well, we tested a NOS-III gene haplotype, consisting of three functional polymorphisms, for an association with bipolar disorder and major depression. A significant global haplotype association with bipolar disorder (n = 284 controls; n = 91 patients; p(global) = 0.021; p(t-a-g) < 0.001), but not unipolar depression (n = 45) was detected. Our results thus suggest that the NOS-III genotype may convey a modest genetic risk to develop bipolar disorder. This finding should be further clarified by the use of within-family designs and in samples of other ethnicity.

Adult↗

PubMed2005

Understanding platelets. Lessons from proteomics, genomics and promises from network analysis.

New large-scale analysis techniques such as bioinformatics, mass spectrometry and SAGE data analysis will allow a new framework for understanding platelets. This review analyses some important options and tasks for these tools and examines an outline of the new, refined picture of the platelet outlined by these new techniques. Looking at the platelet-specific building blocks of genome, (active) transcriptome and proteome (notably secretome and phospho-proteome), we summarize current bioinformatical and biochemical approaches, tasks as well as their limitations. Understanding the surprisingly complex platelet regarding compartmentalization, key cascades, and pathways including clinical implications will remain an exciting and hopefully fruitful challenge for the future.

Animals↗

PubMed2004

Lack of sensitivity of primary Fanconi's anemia fibroblasts to UV and ionizing radiation.

Clinical observations and theoretical considerations suggest some degree of radiosensitivity in Fanconi's anemia (FA), but experimental evidence remains controversial. We tested the sensitivity of primary skin fibroblast cultures from all known FA complementation groups to ionizing radiation and ultraviolet light using conventional cell growth and colony formation assays. In contrast to previous studies, and because FA fibroblasts grow and clone poorly at ambient oxygen, we performed our sensitivity tests under hypoxic cell culture conditions. Fibroblast strains from healthy donors served as negative controls and those from patients with ataxia telangiectasia (AT) and Cockayne syndrome (CS) as positive controls. We observed interstrain variation but no systematic difference in the response of FA and non-FA control fibroblasts to ionizing radiation. After exposure to UV radiation, only complementation group A, G and D2 strains displayed values for colony formation EC50 that were intermediate between those for the negative and positive controls. Because of considerable interstrain variation, minor alterations of the response of individual FA strains to ionizing and UV radiation should be interpreted with caution and should not be taken as evidence for genotype-specific sensitivities of primary FA fibroblasts. All together, our data indicate neither systematic nor major sensitivities of primary FA fibroblast cultures of any complementation group grown under hypoxic cell culture conditions to ionizing or UV radiation.

Cell Division↗

PubMed2003

Yeast two-hybrid screens imply involvement of Fanconi anemia proteins in transcription regulation, cell signaling, oxidative metabolism, and cellular transport.

Mutations in one of at least eight different genes cause bone marrow failure, chromosome instability, and predisposition to cancer associated with the rare genetic syndrome Fanconi anemia (FA). The cloning of seven genes has provided the tools to study the molecular pathway disrupted in Fanconi anemia patients. The structure of the genes and their gene products provided few clues to their functional role. We report here the use of 3 FA proteins, FANCA, FANCC, and FANCG, as "baits" in the hunt for interactors to obtain clues for FA protein functions. Using five different human cDNA libraries we screened 36.5x10(6) clones with the technique of the yeast two-hybrid system. We identified 69 proteins which have not previously been linked to the FA pathway as direct interactors of FANCA, FANCC, or FANCG. Most of these proteins are associated with four functional classes including transcription regulation (21 proteins), signaling (13 proteins), oxidative metabolism (10 proteins), and intracellular transport (11 proteins). Interaction with 6 proteins, DAXX, Ran, IkappaBgamma, USP14, and the previously reported SNX5 and FAZF, was additionally confirmed by coimmunoprecipitation and/or colocalization studies. Taken together, our data strongly support the hypothesis that FA proteins are functionally involved in several complex cellular pathways including transcription regulation, cell signaling, oxidative metabolism, and cellular transport.

Cell Cycle Proteins↗

PubMed2002

A mouse model for Sorsby fundus dystrophy.

PURPOSE: Sorsby fundus dystrophy (SFD) is a rare, late-onset macular dystrophy caused by mutations in the tissue inhibitor of metalloproteinases-3 (TIMP3) gene. The known mutations introduce potentially unpaired cysteine residues in the C terminus of the protein and result in the formation of higher-molecular-weight protein complexes of as yet unknown composition and functional consequences in the pathologic course of SFD. To facilitate in vivo investigation of mutant TIMP3, the authors generated a knock-in mouse carrying a disease-related Ser156Cys mutation in the orthologous murine Timp3 gene. METHODS: Site-directed mutagenesis and homologous recombination in embryonic stem (ES) cells was used to generate mutant ES cells carrying the Timp3(S156C) allele. Chimeric animals were obtained, of which two displayed germline transmission of the mutated allele. Molecular genetic, biochemical, electron microscopic, and electrodiagnostic techniques were used for characterization. RESULTS: At 8 months of age, knock-in mice showed abnormalities in the inner aspect of Bruch's membrane and in the organization of the adjacent basal microvilli of the retinal pigment epithelium (RPE). Changes resembling those in the mutant animals were also present to some extent in normal littermates, but only at an advanced age of 30 months. Long-term electrodiagnostic recordings indicated normal retinal function throughout life. The biochemical characteristics of the mutant protein appear similar in humans and knock-in mice, suggesting common molecular pathways in the two species. The localization of the mutant protein in the eye is normal, although there is evidence of increased Timp3 levels in Bruch's membrane of mutant animals. CONCLUSIONS: The knock-in mice display early features of age-related changes in Bruch's membrane and the RPE that may represent the primary clinical manifestations of SFD. In addition, our immunolabeling studies and biochemical data support a model proposing that site-specific excess rather than absence or deficiency of functional Timp3 may be the primary consequence of the known Timp3 mutations.

Animals↗