Über den primären Energiespeicher der Muskelkontraktion

Ohne Zusammenfassung     

Urinary collecting system invasion reflects adverse long-term outcome and is associated with simultaneous metastatic spread at the time of surgery and with multilocular dissemination during postsurgical follow-up in renal cell cancer

Objectives  

To assess the prognostic implication of urinary collecting system invasion (CSI) in renal cell cancer (RCC).     

Prognostic impact of molecular genetic alterations in hepatoblastoma

BACKGROUND: During recent years, we identified characteristic genetic alterations in sporadic hepatoblastoma (HB). These include loss of heterozygosity (LOH) at the chromosomal region 11p15.5, LOH on chromosome arms 1p and 1q, activating mutations in exon 3 of the beta-catenin gene, as well as overexpression of the c-met oncogene, which encodes the hepatocyte growth factor receptor. We now wanted to evaluate the prognostic relevance of these abnormalities concerning the outcome in a large group of patients. PROCEDURE: All but 7 of 56 patients with HB were treated either with primary resection for small tumors, or neo-adjuvant chemotherapy with ifosfamide, cisplatin and doxorubicin (IPA) before delayed surgery in case of extended disease and additional adjuvant IPA therapy in all cases. Seven tumors were primarily resected and adjuvantly treated with different cytotoxic drugs. LOH 11p15.5, LOH 1p, and LOH 1q were detected in tumor tissue in comparison to normal liver and/or peripheral blood leukocytes by PCR based microsatellite analysis. Beta-catenin mutations were analysed with SSCP, deletion screening and direct sequencing. RT-PCR was used for identification of c-met mRNA overexpression. The results were correlated with the tumors' stage, histological type and epithelial differentiation, as well as with the patients' outcome. RESULTS: Overall disease-free survival after median follow-up of 5 years was 43/56 patients (77%). LOH 11p15.5 was found in 15/56, LOH1p in 11/53, LOH1q in 7/53 and beta-catenin mutations in 25/55 HB. 13/23 HB had a c-met overexpression. LOH 11p15.5 and LOH 1p were significantly more often found in embryonal HB, while there was no correlation of other genetic alterations with histological type or differentiation. Furthermore, statistical analysis revealed no correlation of any of these disorders with initial tumor stage, nor with the patients' outcome. CONCLUSION: None of the investigated molecular genetic alterations are suited to serve as a prognostic indicator in HB. Further investigations, especially genetic screening of tumor tissue may reveal prognostic markers for this neoplasm.     

Identification of 36 novel Jagged1 (JAG1) mutations in patients with Alagille syndrome

Alagille syndrome (AGS) is an autosomal dominant disorder characterized by five major symptoms: cholestasis, vertebral deformity, heart malformations, ocular defects and peculiar facial appearance. The previously described Jagged1 (JAG1) gene on chromosome 20p12 has been identified as being responsible for AGS. JAG1 encodes a transmembrane protein acting as ligand for the evolutionarily conserved Notch signaling pathway. Here we report 36 novel mutations in the JAG1 gene. We identified 12 novel deletions, 4 insertions, 8 missense, 7 nonsense and 5 splice site mutations. All mutations map to the sequence encoding the extracellular part of the Jagged1 protein. The mutations spread over the entire gene with slightly increased rates in exons 2 to 6 and exon 23 and 24. Eight novel missense mutations map to the Delta-Serrate-Lag2 (DSL) domain and adjacent sequences which are important for ligand-receptor interaction. Inheritance was determined in 27 families. Sixteen mutations (55%) were de novo and eleven mutations (45%) were transmitted. Altogether 226 different JAG1 mutations have been described in association with AGS, including our novel 36 mutations. AGS variants are spread over the entire gene with only a few mutations in exon 26. A relatively high number of mutations are clustered in exons 2 to 6. This sequence region shows high interspecies conservation and encodes the Notch receptor-binding region (DSL domain).     

Bone marrow-derived cells contribute to cell turnover in aging murine hearts

The paradigm that cardiac myocytes are non-proliferating, terminally differentiated cells was recently challenged by studies reporting the ability of bone marrow-derived cells (BMCs) to differentiate into cardiomyocytes after myocardial damage. However, little knowledge exists about the role of BMCs in the heart during physiological aging. Twelve-week-old mice (n=36) were sublethally irradiated and bone marrow from littermates transgenic for enhanced green fluorescent protein (eGFP) was transplanted. After 4 weeks, 18 mice were sacrificed at the age of 4 months and served as controls (group A); the remaining mice were sacrificed at the age of 18 months (group B). Group A did not exhibit a significant number of eGFP+ cells, whereas 9.4±2.8 eGFP+ cells/mm2 was documented in group B. In total, only five eGFP+ cardiomyocytes were detected in 20 examined hearts, excluding a functional role of BM differentiation in cardiomyocytes. Similarly, a relevant differentiation of BMCs in endothelial or smooth muscle cells was excluded. In contrast, numerous BM-derived fibroblasts and myofibroblasts were observed in group B, but none were detected in group A. The present study demonstrates that BMCs transdifferentiate into fibroblasts and myofibroblasts in the aging murine myocardium, suggesting their contribution to the preservation of the structural integrity of the myocardium, while they do not account for regenerative processes of the heart.     

Comprehensive sequence analysis of the NR5A1 gene encoding steroidogenic factor 1 in a large group of infertile males

The steroidogenic factor 1 (SF1) protein, encoded by the NR5A1 gene, plays a central role in gonadal development and steroidogenesis. Mutations in NR5A1 were first described in patients with primary adrenal insufficiency and 46,XY disorders of sexual development and later also in men with hypospadias, bilateral anorchia and micropenis and women with primary ovarian insufficiency. Recently, heterozygous missense mutations were found in 4% of infertile men with unexplained reduced sperm counts living in France, but all mutation carriers were of non-Caucasian ancestry. Therefore, we performed a comprehensive NR5A1 sequence analysis in 488 well-characterised predominantly Caucasian patients with azoo- or severe oligozoospermia. Two-hundred-thirty-seven men with normal semen parameters were sequenced as controls. In addition to several synonymous variants of unclear pathogenicity, three heterozygous missense mutations predicted to be damaging to SF1 protein function were identified. The andrological phenotype in infertile but otherwise healthy mutation carriers seems variable. In conclusion, mutations altering SF1 protein function and causing spermatogenic failure are also found in men of German origin, but the prevalence seems markedly lower than in other populations.     

NTRK testing: First results of the QuiP‐EQA scheme and a comprehensive map of NTRK fusion variants and their diagnostic coverage by targeted RNA‐based NGS assays

Gene fusions involving the three neurotrophic tyrosine receptor kinase genes NTRK1, NTRK2, or NTRK3 were identified as oncogenic drivers in many cancer types. Two small molecule inhibitors have been tested in clinical trials recently and require the detection of a NTRK fusion gene prior to therapeutic application. Fluorescence in situ hybridization (FISH) and targeted next‐generation sequencing (tNGS) assays are commonly used for diagnostic profiling of gene fusions. In the presented study we applied an external quality assessment (EQA) scheme in order to investigate the suitability of FISH and RNA‐/DNA‐based tNGS for detection of NTRK fusions in a multinational and multicentric ring trial. In total 27 participants registered for this study. Nine institutions took part in the FISH‐based and 18 in the NGS‐based round robin test, the latter additionally subdivided into low‐input and high‐input NGS methods (regarding nucleic acid input). Regardless of the testing method applied, all participants received tumor sections of 10 formalin‐fixed and paraffin‐embedded (FFPE) tissue blocks for in situ hybridization or RNA/DNA extraction, and the results were submitted via an online questionnaire. For FISH testing, eight of nine (88.8%) participants, and for NGS‐based testing 15 of 18 (83.3%) participants accomplished the round robin test successfully. The overall high success rate demonstrates that FISH‐ and tNGS‐based NTRK testing can be well established in a routine diagnostic setting. Complementing this dataset, we provide an updated in silico analysis on the coverage of more than 150 NTRK fusion variants by several commercially available RNA‐based tNGS panels.     

Characterization of a chemokine receptor CCR5-negative T cell line and its use in determining human immunodeficiency virus type 1 phenotype

A human CD4-positive T cell line from a donor homozygous negative for the chemokine receptor CCR5 was established, characterized, and used for determining the coreceptor usage of human immunodeficiency virus type 1 (HIV-1) isolates. Clones of this IL-2 dependent human T-cell lymphotropic virus type 1 (HTLV-I) immortalized cell line, named IsnoR5 clones 1 and 2, are susceptible to infection by HIV-1 isolates that use CXCR4 as a coreceptor but resistant to infection by CCR5 tropic HIV-1 viruses. HIV-1 isolates whose replication is inhibited in IsnoR5 cells in the presence of the bicyclam AMD 3100, a CXCR4 specific inhibitor, utilize a coreceptor distinct from CCR5 and CXCR4. Using a panel of primary HIV-1 isolates we have shown that a single T cell line is sufficient to discriminate between use of CCR5, CXCR4 or an alternative coreceptor. As IsnoR5 clone 1 cells revealed the existence of even minor populations of CXCR4-using virus variants, they could be useful for the early identification of changes in coreceptor usage in HIV infected individuals facilitating the timely introduction of appropriate clinical treatments.