Christine Steinhoff - Academia.edu (original) (raw)

Papers by Christine Steinhoff

Tumor Biology, Jan 8, 2015

The version in the Kent Academic Repository may differ from the final published version. Users ar... more The version in the Kent Academic Repository may differ from the final published version. Users are advised to check http://kar.kent.ac.uk for the status of the paper. Users should always cite the published version of record.

PubMed, May 15, 2001

Testicular germ cell tumors (GCT) characteristically display two chromosome 12 abnormalities: the... more Testicular germ cell tumors (GCT) characteristically display two chromosome 12 abnormalities: the isochromosome i(12p) and concomitant deletions of the long arm. Some genes important in the control of the G(1)-S cell cycle checkpoint G(1)-S, i.e., cyclin-dependent kinases 2 and 4, cyclin D2 are located on this chromosomal region. Therefore, testicular GCTs were analyzed as to the expression of CDK2, CDK4, CDK6, and the expression of their catalytic partners cyclins D1, D2 and E by semiquantitative reverse transcription-PCR. Cyclin D2, located on 12p, was overexpressed in 69% (31 of 45) of the tumors by a mean factor of 8, including all histological subtypes. In addition, the cyclin D2 partner CDK4 was increased in 41% (21 of 51) of all tumors by a factor of 6, most strongly in embryonal carcinomas. Sixty-four percent of the seminomas and 23% of the non-seminomas had decreased expression of CDK6 by a mean factor of 5 (P = 0.009). Statistical analysis using configural frequency analysis and regression analysis revealed that cyclin D2 and CDK4 expression were strongly correlated (r(2) = 0.682; P = 0.000052), whereas expression of CDK6 did not correlate with either of them (r(2) = 0.382; P = 0.00085). CDK2 and its catalytic partner cyclin E were down-regulated in 40% (19 of 47) and 42% (19 of 45) of the tumors, respectively, by a factor of 7 each. Western blots and immunohistochemical experiments confirmed cyclin D2 and CDK4 overrepresentation and reduced expression of cyclin E and CDK2 tumors in the few tumors under protein study. Despite its localization on 12q13, a hot spot for loss of heterozygosity in testicular GCTs (>40%), Southern blotting revealed no gross DNA alteration of the CDK2 gene. Because up-regulation of the cyclin D2/CDK4 complex and down-regulation of cyclin E/CDK2 complex were found in seminomas as well as in non-seminomas and in all tumor stages, these findings seem to be early events during tumorigenesis of testicular GCTS: Together with previous findings that retinoblastoma mRNA and protein expression is strongly decreased in these tumors, these data suggest an unusual deregulated G(1)-S checkpoint as a decisive event for germ cell tumors.

Molecular Genetics and Genomics, Oct 7, 2003

Although LINE-1 (L1) sequences constitute the most important family of retrotransposons in the hu... more Although LINE-1 (L1) sequences constitute the most important family of retrotransposons in the human genome, their transcriptional regulation is poorly understood. Specifically, their unusual internal promoter is incompletely characterized. Current promoter prediction programs fail to identify the promoter in the 5¢UTR of the active LINE-1 element L1.2B. Experimental investigation of this promoter using reporter gene assays in various human and murine cell types confirmed that the promoter consists of two segments, and demonstrated that the distal portion is essential for cell-type-independent activity. No differences in promoter activity were found between normal and transformed cells. The complete promoter was shown to possess %20% of the activity of the strong early promoter of cytomegalovirus, and to be capable of directing the expression of levels of p53 sufficient to kill normal and transformed human cells. Thus, active LINE-1 elements contain highly active promoters capable of driving cell-type-independent expression, which are of potential use in mammalian expression constructs. In vitro methylation of the promoter at Hpa II sites decreased its activity independently of cell type, but this repression was alleviated in MBD2)/) cells. Surprisingly, mutation of specific Hpa II sites was also found to reduce promoter activity. Thus, efficient repression of the L1.2B promoter by DNA methylation may involve MBD2 binding, but at least one Hpa II site also appears to be involved specifically in transcriptional activation. Since neither promoter activity nor the efficiency of repression by methylation differed between normal and tumor cells, the re-activation of LINE-1 sequences observed in tumor cells is probably caused by hypomethylation of the promoter.

International Journal of Cancer, Feb 19, 2003

In many common cancers such as transitional cell carcinoma (TCC), specific genes are hypermethyla... more In many common cancers such as transitional cell carcinoma (TCC), specific genes are hypermethylated, whereas overall DNA methylation is diminished. Genome-wide DNA hypomethylation mostly affects repetitive sequences such as LINE-1 retrotransposons. Methylation of these sequences depends on adequate expression of DNA methyltransferase I (DNMT1) during DNA replication. Therefore, DNMT1 expression relative to proliferation was investigated in TCC cell lines and tissue as well as in renal carcinoma (RCC) cell lines, which also display hypomethylation, as indicated by decreased LINE-1 methylation. Cultured normal uroepithelial cells or normal bladder tissue served as controls. In all tumor cell lines, DNMT1 mRNA as well as protein was decreased relative to the DNA replication factor PCNA, and DNA hypomethylation was present. However, the extents of hypomethylation and DNMT1 downregulation did not correlate. Reporter gene assays showed that the differences in DNMT1 expression between normal and tumor cells were not established at the level of DNMT1 promoter regulation. Diminished DNMT1:PCNA mRNA ratios were also found in 28/45 TCC tissues but did not correlate with the extent of DNA hypomethylation. In addition, expression of the presumed de novo methyltransferases DNMT3A and DNMT3B mRNAs was investigated. DNMT3B overexpression was observed in about half of all high-stage TCC (DNMT3B vs. tumor stage, chi(2): p = 0.03), whereas overexpression of DNMT3A was rarer and less pronounced. Expression of DNMT3A and DNMT3B in most RCC lines was higher than in TCC lines. Our data indicate that DNMT1 expression does not increase adequately with cell proliferation in bladder cancer. This relative downregulation probably contributes to hypomethylation of repetitive DNA but does not determine its extent alone.

The Journal of Neuroscience, Jun 30, 2004

The molecular changes underlying neural progenitor differentiation are essentially unknown. We ap... more The molecular changes underlying neural progenitor differentiation are essentially unknown. We applied cDNA microarrays with 13,627 clones to measure dynamic gene expression changes during the in vitro differentiation of neural progenitor cells that were isolated from the subventricular zone of postnatal day 7 mice and grown in vitro as neurospheres. In two experimental series in which we withdrew epidermal growth factor and added the neurotrophins Neurotrophin-4 or BDNF, four time points were investigated: undifferentiated cells grown as neurospheres, and cells 24, 48, and 96 hr after differentiation. Expression changes of selected genes were confirmed by semiquantitative RT-PCR. Ten different groups of gene expression dynamics obtained by cluster analysis are described. To correlate selected gene expression changes to the localization of respective proteins, we performed immunostainings of cultured neurospheres and of brain sections from adult mice. Our results provide new insights into the genetic program of neural progenitor differentiation and give strong hints to as yet unknown cellular communications within the adult subventricular zone stem cell niche.

Genes & Development, Feb 1, 2010

Klinische Padiatrie, Apr 1, 2010

Conclusion: Muscular dystrophy is associated with significant cardiomyopathy, which may shorten l... more Conclusion: Muscular dystrophy is associated with significant cardiomyopathy, which may shorten life expectancy. This unique population can benefit from careful cardiac surveillance with echocardiography to detect cardiomyopathy and allow early pharmacological treatment.

Genes & Development, Mar 15, 2010

Urology, Oct 1, 2000

To evaluate the expression of p27(KIP1) and p21(CIP1) and the prognostic values of both markers i... more To evaluate the expression of p27(KIP1) and p21(CIP1) and the prognostic values of both markers in urothelial carcinoma. The expression of the cyclin-dependent kinase inhibitor p27(KIP1) characterizes early-stage and well-differentiated carcinomas of the colon, breast, and prostate and is associated with an improved prognosis. In urothelial carcinoma, its expression has not been as well investigated. Another cyclin-dependent kinase inhibitor, p21(CIP1), is expressed in early-stage bladder tumors, but published data on its prognostic value are contradictory. Expression of p27(KIP1) and p21(CIP1) was analyzed by immunohistochemistry in 114 urothelial carcinoma specimens from 77 patients. The Ki67 index was determined as an indicator of cell proliferation. The expression of the markers was correlated with tumor recurrence and progression during an average follow-up period of 3.9 years. Expression of p27(KIP1) was significantly more frequent in superficial than in muscle-invasive tumors (chi-square test, P = 0.012; Fisher's exact test, P = 0.014). Although similar overall, the expression pattern of p21(CIP1) did not match on a tumor-by-tumor basis. No correlation was seen with the Ki67 index. Patients with tumors displaying strong positive staining for p27(KIP1) or p21(CIP1) had fewer recurrences and progression events, but the difference was not statistically significant. Instead, a Ki67 index of less than 10% was significantly (P = 0.0335) related to a lack of recurrence. Neither p27(KIP1) nor p21(CIP1) appear to be good predictors of tumor progression in urothelial carcinoma, even though their expression is strongly decreased in muscle-invasive tumors.

Stem Cells and Development, Aug 15, 2014

True tendon regeneration in human patients remains a vision of musculoskeletal therapies. In comp... more True tendon regeneration in human patients remains a vision of musculoskeletal therapies. In comparison to other mesenchymal lineages the biology of tenogenic differentiation is barely understood. Specifically, easy and efficient protocols are lacking that might enable tendon cell and tissue differentiation based on adult (stem) cell sources. In the murine mesenchymal progenitor cell line C3H10T½, overexpression of the growth factor bone morphogenetic protein 2 (BMP2) and a constitutively active transcription factor, Smad8 L + MH2, mediates tendon cell differentiation in vitro and the formation of tendon-like tissue in vivo. We hypothesized that during this differentiation secreted factors involved in extracellular matrix formation exert a major impact on tendon development. Gene expression analyses revealed four genes encoding secreted factors that are notably upregulated: periostin, C-type lectin domain family 3 (member b), RNase A4, and follistatin-like 1. These factors have not previously been implicated in tendon biology. Among these, periostin showed a specific expression in tenocytes of adult mouse Achilles tendon and in chondrocytes within the nonmineralized fibrocartilage zone of the enthesis with the calcaneus. Overexpression of periostin alone or in combination with constitutively active BMP receptor type in human mesenchymal stem cells and subsequent implantation into ectopic sites in mice demonstrated a reproducible moderate tenogenic capacity that has not been described before. Therefore, periostin may belong to the factors contributing to the development of tenogenic tissue.

The Prostate, 2000

BACKGROUND. Alterations of DNA methylation are very frequent in prostatic carcinoma. A possible c... more BACKGROUND. Alterations of DNA methylation are very frequent in prostatic carcinoma. A possible cause underlying altered DNA methylation could be an insufficient level of Sadenosylmethionine as a consequence of nutritional imbalances or of weaker alleles of genes for its synthesis, i.e., encoding methylene-tetrahydrofolate reductase (MTHFR), methionine synthase (MS), and ␤-cystathione synthetase (CBS). Therefore, homozygosity or heterozygosity for such weaker alleles may underlie susceptibility to prostatic carcinoma. METHODS. The distribution of the two most frequent MTHFR, MS, and CBS alleles was determined in 132 prostatic carcinoma patients and 150 population controls by restriction fragment length polymorphism-(RFLP) PCR. RESULTS. In the controls, a Hardy-Weinberg equilibrium was observed for each allele pair. No significant differences were observed with respect to age or gender. No significant differences for single genes or combinations were found between prostatic carcinoma patients and controls, although the MTHFR Val allele was slightly overrepresented among the tumor patients. Neither did the allele distribution significantly differ among the prostatic carcinoma patients stratified according to age, clinical stage, or presence of metastases. However, the MTHFR Val allele tended to be associated with higher tumor grade. CONCLUSIONS. In general, the data do not support the hypothesis that weaker alleles in methyl group metabolism genes constitute a major factor in the high prevalence of DNA methylation alterations found in prostatic carcinoma. However, a potential association with the MTHFR genotype deserves further study.

Briefings in Bioinformatics, Mar 7, 2006

Array-based gene expression studies frequently serve to identify genes that are expressed differe... more Array-based gene expression studies frequently serve to identify genes that are expressed differently under two or more conditions. The actual analysis of the data, however, may be hampered by a number of technical and statistical problems. Possible remedies on the level of computational analysis lie in appropriate preprocessing steps, proper normalization of the data and application of statistical testing procedures in the derivation of differentially expressed genes. This review summarizes methods that are available for these purposes and provides a brief overview of the available software tools.

Stem Cells, Jun 1, 2005

We recently established that two midgestation-derived stromal clones-UG26-1B6, urogenital ridge-d... more We recently established that two midgestation-derived stromal clones-UG26-1B6, urogenital ridge-derived, and EL08-1D2, embryonic liver-derived-support the maintenance of murine adult bone marrow and human cord blood hematopoietic repopulating stem cells (HSCs). In this study, we investigate whether direct HSC-stroma contact is required for this stem cell maintenance. Adult bone marrow ckit + Ly-6C − side population (K6-SP) cells and stromal cells were cocultured under contact or noncontact conditions. These experiments showed that HSCs were maintained for at least 4 weeks in culture and that direct contact between HSCs and stromal cells was not required. To find out which factors might be involved in HSC maintenance, we compared the gene expression profile of EL08-1D2 and UG26-1B6 with four HSC-nonsupportive clones. We found that EL08-1D2 and UG26-1B6 both expressed 21 genes at a higher level, including the putative secreted factors fibroblast growth factor-7, insulin-like growth factor-binding proteins 3 and 4, pleiotrophin, pentaxin-related, and thrombospondin 2, whereas 11 genes, including GPX-3 and HSP27, were expressed at a lower level. In summary, we show for the first time long-term maintenance of adult bone marrow HSCs in stroma noncontact cultures and identify some secreted molecules that may be involved in this support.

European Urology Supplements, 2002

Urology, Feb 1, 2004

Objectives. The Fas-Fas ligand system is an important regulator of apoptosis and is involved in t... more Objectives. The Fas-Fas ligand system is an important regulator of apoptosis and is involved in tumor development. Invasive cancers downregulate Fas expression to evade antitumor immune responses. Fas is a transcriptional target of p53, which is often mutated in bladder cancers. Therefore, Fas expression and its relation to p53 mutation was investigated. Methods. Expression of Fas protein and p53 status was studied by immunohistochemistry in 83 bladder cancer specimens. In addition, mRNA levels for soluble (decoy) and membrane-bound forms of Fas were compared between 10 bladder cancer cell lines and primary uroepithelial cells by quantitative TaqMan polymerase chain reaction. Mutational analysis of the death domain of the Fas gene was performed in all cell lines. Results. Organ-confined tumors maintained specific Fas staining at the cell membrane and often also in the cytoplasm. In higher stage carcinomas, Fas expression became restricted to a smaller fraction of cells or was lacking entirely. The correlation of Fas staining with tumor stage was highly significant but no correlation to tumor grade or survival was found. Furthermore, no statistically significant relationship was observed with either the presence or lack of mutated p53 accumulation. Membrane-bound Fas mRNA was decreased in most, and soluble Fas was increased in all transitional cell carcinoma lines compared with primary uroepithelial cells. No mutations in the death domain were detected. Conclusions. Fas downregulation occurring in advanced bladder cancer is unrelated to p53 mutations. The results of immunohistochemistry and mRNA studies of soluble and membrane-bound Fas in transitional cell carcinoma lines support the hypothesis of immune evasion in advanced bladder cancer. UROLOGY 63: 392-397, 2004.

Journal of Cellular and Molecular Medicine, May 14, 2010

The RNA-binding protein Musashi1 (MSI1) is a marker of progenitor cells in the nervous system fun... more The RNA-binding protein Musashi1 (MSI1) is a marker of progenitor cells in the nervous system functioning as a translational repressor. We detected MSI1 mRNA in several bladder carcinoma cell lines, but not in cultured normal uroepithelial cells, whereas the paralogous MSI2 gene was broadly expressed. Knockdown of MSI1 expression by siRNA induced apoptosis and a severe decline in cell numbers in 5637 bladder carcinoma cells. Microarray analysis of gene expression changes after MSI1 knockdown significantly up-regulated 735 genes, but down-regulated only 31. Up-regulated mRNAs contained a highly significantly greater number and density of Musashi binding sites. Therefore, a much larger set of mRNAs may be regulated by Musashi1, which may affect not only their translation, but also their turnover. The study confirmed p21 CIP1 and Numb proteins as targets of Musashi1, suggesting additionally p27 KIP1 in cell-cycle regulation and Jagged-1 in Notch signalling. A significant number of up-regulated genes encoded components of stress granules (SGs), an organelle involved in translational regulation and mRNA turnover, and impacting on apoptosis. Accordingly, heat shock induced SG formation was augmented by Musashi1 down-regulation. Our data show that ectopic MSI1 expression may contribute to tumorigenesis in selected bladder cancers through multiple mechanisms and reveal a previously unrecognized function of Musashi1 in the regulation of SG formation.

Developmental Dynamics, May 1, 2004

To obtain a deeper insight into the genes and gene networks involved in the development of placen... more To obtain a deeper insight into the genes and gene networks involved in the development of placentopathies, we have assessed global gene expression in three different models of placental hyperplasia caused by interspecies hybridization (IHPD), cloning by nuclear transfer, and mutation of the Esx1 gene, respectively. Comparison of gene expression profiles of approximately 13,000 expressed sequence tags (ESTs) identified specific subsets of genes with changed expression levels in IHPD, cloned, and Esx1 mutant placentas. Of interest, only one gene of known function and one EST of unknown function were found common to all three placentopathies; however, a significant number of ESTs were common to IHPD and cloned placentas. In contrast, only one gene was shared between IHPD and Esx1 mutant, and cloned and Esx1 mutant placentas, respectively. These genes common to different abnormal placental growth genotypes are likely to be important in the occurrence of placentopathy. Developmental Dynamics 230: 149-164, 2004.

Cell and Tissue Research, Jan 31, 2003

Bone marrow stromal cells (BMSC) have gained increased attention because of their multipotency an... more Bone marrow stromal cells (BMSC) have gained increased attention because of their multipotency and adult stem cell character. They have been shown to differentiate into other cell types of the mesenchymal lineage and also into non-mesenchymal cells. The exact identity of the original cells, which are isolated from bone marrow by their selective adherence to plastic, remains unknown to date. We have established and characterized mouse BMSC cultures and analyzed three independent samples by cDNA microarrays. The expression profile was compared with two previous expression studies of human BMSC and revealed a high degree of concordance between different techniques and species. To gain clues about the positional context and biology of the isolated cells within the bone marrow stroma, we searched our data for genes that encode proteins of the extracellular matrix, cell adhesion proteins, cytoskeletal proteins and cytokines/cytokine receptors. This analysis revealed a close association of BMSC with vascular cells and indicated that BMSC resemble pericytes.