Antiepileptic drugs affect neuronal androgen signaling via a cytochrome P450-dependent pathway

Recent data imply an important role for brain cytochrome P450 (P450) in endocrine signaling. In epileptic patients, treatment with P450 inducers led to reproductive disorders; in mouse hippocampus, phenytoin treatment caused concomitant up-regulation of CYP3A11 and androgen receptor (AR). In the present study, we established specific in vitro models to examine whether CYP3A isoforms cause enhanced AR expression and activation. Murine Hepa1c1c7 cells and neuronal-type rat PC-12 cells were used to investigate P450 regulation and its effects on AR after phenytoin and phenobarbital administration. In both cell lines, treatment with antiepileptic drugs (AEDs) led to concomitant up-regulation of CYP3A (CYP3A11 in Hepa1c1c7 and CYP3A2 in PC-12) and AR mRNA and protein. Inhibition of CYP3A expression and activity by the CYP3A inhibitor ketoconazole or by CYP3A11-specific short interfering RNA molecules reduced AR expression to basal levels. The initial up-regulation of AR signal transduction, measured by an androgen-responsive element chloramphenicol-acetyltransferase reporter gene assay, was completely reversed after specific inhibition of CYP3A11. Withdrawal of the CYP3A11 substrate testosterone prevented AR activation, whereas AR mRNA expression remained up-regulated. In addition, recombinant CYP3A11 was expressed heterologously in PC-12 cells, thereby eliminating any direct drug influence on the AR. Again, the initial up-regulation of AR mRNA and activity was reduced to basal levels after silencing of CYP3A11. In conclusion, we show here that CYP3A2 and CYP3A11 are crucial mediators of AR expression and signaling after AED application. These findings point to an important and novel function of P450 in regulation of steroid hormones and their receptors in endocrine tissues such as liver and brain.     

Die Unfallchirurgie und die Deutsche Gesellschaft für Unfallchirurgie

Die Unfallchirurgie -     

TMPRSS2-ERG Fusions Are Strongly Linked to Young Patient Age in Low-grade Prostate Cancer

Based on next-generation sequencing of early-onset prostate cancer (PCa), we earlier demonstrated that PCa in young patients is prone to rearrangements involving androgen-regulated genes—such as transmembrane protease, serine 2 (TMPRSS2)–v-ets avian erythroblastosis virus E26 oncogene homolog (ERG) fusion—and provided data suggesting that this situation might be caused by increased androgen signaling in younger men. In the same study, an accumulation of chromosomal deletions was found in cancers of elderly patients. To determine how age-dependent molecular features relate to cancer phenotype, an existing data set of 11 152 PCas was expanded by additional fluorescence in situ hybridization analyses of phosphatase and tensin homolog (PTEN), 6q15 and 5q21. The results demonstrate that the decrease in TMPRSS2–ERG fusions with increasing patient age is limited to low-grade cancers (Gleason ≤3 + 4) and that the significant increase in the deletion frequency with age was strictly limited to ERG-negative cancers for 6q15 and 5q21 but to ERG-positive cancers for PTEN. These data suggest that the accumulation of non–androgen-linked genomic alterations with advanced patient age may require an appropriate microenvironment, such as a positive or negative ERG status. The strong link of ERG activation to young patient age and low-grade cancers may help to explain a slight predominance of low-grade cancers in young patients.     

Expression of modulators of extracellular matrix structure after anterior cruciate ligament injury

The ability of the anterior cruciate ligament (ACL) to heal after injury declines within the first 2 weeks after ACL rupture. To begin to explore the mechanism behind this finding, we quantified the expression of genes for collagen I and III, decorin, tenascin‐C, and alpha smooth muscle actin, as well as matrix metalloproteinase (MMP)‐1 and ‐13 gene expression within multiple tissues of the knee joint after ACL injury in a large animal model over a 2‐week postinjury period. Gene expression of collagen I and III, decorin, and MMP‐1 was highest in the synovium, whereas the highest MMP‐13 gene expression levels were found in the ACL. The gene expression for collagen and decorin increased over the 2 weeks to levels approaching that in the ligament and synovium; however, no significant increase in either of the MMPs was found in the provisional scaffold. This suggests that although the ACL and synovium up‐regulate both anabolic and catabolic factors, the provisional scaffold is primarily anabolic in function. The relative lack of provisional scaffold formation within the joint environment may thus be one of the key reasons for ACL degradation after injury.     

Massive blood transfusion after the first cut in liver transplantation predicts renal outcome and survival

Introduction

Transfusion requirements of blood products may provide useful prognostic factors for the prediction of short-term patient mortality and renal outcome after liver transplantation.

Patients and methods

Two hundred ninety-one consecutive liver transplants in adults were analysed retrospectively. Combined and living-related liver transplants were excluded. The amount of transfused packed red blood cells (PRBC) and units of platelets (UP) within the first 48 h were investigated as prognostic factors to predict short-term patient mortality and renal outcome. Receiver operating characteristic (ROC) curve analysis with area under the curve (AUC), Hosmer-Lemeshow tests and Brier scores were used to calculate overall model correctness, model calibration and accuracy of prognostic factors. Cut-off values were determined with the best Youden index.

Results

The potential clinical usefulness of PRBC as a prognostic factor to predict 30-day mortality (cut-off 17.5 units) and post-transplant haemodialysis (cut-off 12.5 units) could be demonstrated with AUCs >0.7 (0.712 and 0.794, respectively). Hosmer-Lemeshow test results and Brier scores indicated good overall model correctness, model calibration and accuracy. The UP proved as an equally clinically useful prognostic factor to predict end-stage renal disease (cut-off 3.5 units; AUC?=?0.763). The association of cut-off levels of PRBC with patient survival (p?<?0.001, log-rank test) and dialysis-free survival (p?<?0.001, log-rank test) was significant (cut-off levels 17.5 and 12.5 units, respectively) as well as the association of UP with dialysis-free survival (p?<?0.001, log-rank test) (cut-off level 3.5 units).

Conclusions

The impressive discriminative power of these simple prognostic factors for the prediction of outcome after liver transplantation emphasizes the relevance of strategies to avoid excessive transfusion requirements.     

The HIV 5′ Gag Region Displays a Specific Nucleotide Bias Regulating Viral Splicing and Infectivity

Alternative splicing and the expression of intron-containing mRNAs is one hallmark of HIV gene expression. To facilitate the otherwise hampered nuclear export of non-fully processed mRNAs, HIV encodes the Rev protein, which recognizes its intronic response element and fuels the HIV RNAs into the CRM-1-dependent nuclear protein export pathway. Both alternative splicing and Rev-dependency are regulated by the primary HIV RNA sequence. Here, we show that these processes are extremely sensitive to sequence alterations in the 5’coding region of the HIV genomic RNA. Increasing the GC content by insertion of either GFP or silent mutations activates a cryptic splice donor site in gag, entirely deregulates the viral splicing pattern, and lowers infectivity. Interestingly, an adaptation of the inserted GFP sequence toward an HIV-like nucleotide bias reversed these phenotypes completely. Of note, the adaptation yielded completely different primary sequences although encoding the same amino acids. Thus, the phenotypes solely depend on the nucleotide composition of the two GFP versions. This is a strong indication of an HIV-specific mRNP code in the 5′ gag region wherein the primary RNA sequence bias creates motifs for RNA-binding proteins and controls the fate of the HIV-RNA in terms of viral gene expression and infectivity.