Gonadotropin-releasing hormone messenger ribonucleic acid levels are unaltered with changes in the gonadal hormone milieu of the adult male rat

Testicular function is regulated by the negative feedback effect of sex hormones acting at the brain and pituitary to inhibit the secretion of LH and FSH. An important component of this feedback axis is presumed to involve regulation of secretion and possibly synthesis of GnRH by the brain. We tested the hypothesis that the castration-induced increase in gonadotropin secretion is subserved, at least in part, by increased synthesis of GnRH. Using in situ hybridization and an oligonucleotide probe to pro-GnRH messenger RNA (GnRH mRNA), we compared the level of cellular GnRH mRNA and the relative number of GnRH mRNA-containing neurons between intact and 21-day castrate adult male rats. To derive estimates of the number of GnRH cells and the cellular GnRH mRNA content, coronal sections from each animal were anatomically matched between intact and castrate groups. All identifiable cells within these sections were counted and analyzed with the aid of a computerized image analysis system, by an observer unaware of the animal's experimental group and were assigned an anatomical location for reference. In an initial experiment, we observed no difference in cellular GnRH mRNA signal level between intact (n = 4) and castrate (n = 5) animals (129 +/- 8 vs. 139 +/- 5 grains per cell); however, we did find a statistical difference between the intact and castrated groups in the relative number of GnRH mRNA-containing cells (intact: 212 +/- 15 vs. castrate: 320 +/- 18). To confirm this observation, we repeated the experiment by again comparing the number of GnRH mRNA-positive cells between intact (n = 4) and castrate (n = 4) rats. In this second experiment, we found no difference in the number of identifiable GnRH mRNA-containing cells between intact and castrate animals (272 +/- 14 vs. 274 +/- 36, respectively); this was the case for the total cell count as well as when the data were analyzed by anatomical region. To clarify the conflicting results on cell counts of Exps 1 and 2, we repeated the experiment a third time, again comparing both the number of GnRH mRNA-containing cells and the cellular content of GnRH mRNA. In this experiment, we observed that neither cell number nor content of GnRH mRNA differed between the intact and castrate groups. Again, this was the case for total cell count, as well as when the data were analyzed by anatomical region.(ABSTRACT TRUNCATED AT 400 WORDS)     

High-Content Screening Microscopy Identifies Novel Proteins With a Putative Role in Secretory Membrane Traffic

Here we describe the establishment of microscope-based functional screening assays in intact cells that allow us to systematically identify new proteins involved in secretory membrane traffic, and proteins that can influence the integrity of the Golgi complex. We were able to identify 20 new proteins that affected either secretory transport, Golgi morphology, or both, when overexpressed in cells. Control experiments with human orthologs to yeast proteins with a role in membrane traffic, or already well characterized mammalian regulators of the secretory pathway, confirmed the specificity and significance of our results. Proteins localized to the Golgi complex or endoplasmic reticulum (ER) showed preferential interference in our assays. Bioinformatic analysis of the new proteins interfering with membrane traffic and/or Golgi integrity revealed broad functional variety, but demonstrated a bias towards proteins with predicted coiled-coil domains and repeat structures. Extending our approach to a much larger set of novel proteins in the future will be an important step toward a more comprehensive understanding of the molecular basis of the secretory pathway. It will also serve as an example for similar microscope-based screens addressing different biological questions.     

Phase I trial of the combination of 6-methylmercaptopurine riboside and 5-fluorouracil

6-Methylmercaptopurine riboside (MMPR) and 5-fluorouracil (5-FU) were administered sequentially to 12 patients in a phase I clinical trial. Toxicities included mild nausea and vomiting, as well as reversible leukopenia and thrombocytopenia. Maximal accumulation of 6-methylmercaptopurine ribonucleoside 5′-monophosphate (MMPR-P), the active metabolite of MMPR, in patients’ erythrocytes occurred between 2 and 6 h after the administration of MMPR and the degree of accumulation was dose-related. At 96 h after MMPR administration, MMPR-P was still detectable in patients’ erythrocytes. Although no clinical responses were documented, a modified dosage schedule of these drugs should be pursued based on the pharmacokinetic data obtained.     

Application of poly-lactide-co-glycolide-microspheres in the transarterial chernoembolization in an animal model of hepatocellular carcinoma

AIM: To introduce an animal model of hepatocellularcarcinoma (HCC) in ACI-rats, and to evaluate the therapeuticeffects of Poly-lactide-co-glycolide(Plcg)-microspheres in thetransarterial chemoembolization (TACE) in this model, aswell the value of this model in the experiments ofinterventional therapy.METHODS: Subcapsular implantation of a solid MorrisHepatoma 3 924A (1 mm3) in the livers was carried out in11 male ACI-rats. The tumor volume (V1) was measured bymagnetic resonance imaging (MRⅠ) (13 days afterimplantation). After laparotomy and retrograde placementof catheter into the gastroduodenal artery (14 days afterimplantation), the following protocols of interventionaltreatment were performed: (A) mitomycin C+Poly-lactide-co-glycolide(Plcg)-microspheres (n＝4); (B) 0.9 % NaCl(control group, n＝7). 13 days after these therapies the changeof the tumor volume (V2) was determined by MRI again.RESULTS: The success rate of tumor implantation reachedto 100 %. The mean tumor volume before TACE (V1) were0.082 cm3 in group A and 0.096 cm3 in group B respectively.The mean tumor volume after TACE (V2) were 0.230 cm3 ingroup A and 1.347 cm3 in group B respectively. The meanV2/V1 were 2.860 in group A and 27.120 in group Brespectively. Compared to the control group (group B),groups A showed a significant reduction of tumor growth(P＝0.004) in the period of observation.CONCLUSION: The growth of liver tumor could be obviouslyprevented by utilizing Plcg-mitomycin-microspheres in TACEin animal model. This rat model of HCC is suitable for theexperimental studies of interventional therapy.     

IFNβ1 secreted by breast cancer cells undergoing chemotherapy reprograms stromal fibroblasts to support tumour growth after treatment

Chemotherapy (CTX) remains the standard of care for most aggressive tumours, including breast cancer (BC). In BC chemotherapeutic regimens, the maximum tolerated dose of cytotoxic drugs is administered at regular intervals, and cancer cells can re‐grow or adapt during the resting periods between cycles. The impact of the tumour microenvironment on the fate of cancer cells after CTX remains poorly understood. Here, we show that paracrine signalling from CTX‐treated cancer cells to stromal fibroblasts can drive cancer cell recovery after cytotoxic drug withdrawal. Interferon β1 (IFNβ1) secreted by cancer cells following treatment with high doses of CTX instigates the acquisition of an anti‐viral state in stromal fibroblasts. This state is associated with an expression pattern here referred to as interferon signature (IFNS), which encompasses several interferon‐stimulated genes (ISGs), including numerous pro‐inflammatory cytokine genes. This crosstalk is an important driver of the expansion of BC cells after CTX, and IFNβ1 blockade in tumour cells abrogated their fibroblast‐dependent recovery potential. Analysis of human breast carcinomas supported a link between CTX‐induced IFNS in tumour stroma and poor response to CTX treatment. First, IFNβ1 expression in human breast carcinomas was found to inversely correlate with recurrence free survival (RFS). Second, using laser capture microdissection data sets, we show a higher expression of IFNS in the stromal tumour compartment compared to the epithelial one and this signature was found to be more prominent in more aggressive subtypes of BC (basal‐like), pointing to a pro‐tumorigenic role of this signature. Moreover, IFNS was associated with higher recurrence rates and a worse outcome in BC patients. Our study unravels a novel form of paracrine communication between cancer cells and fibroblasts that ultimately results in CTX resistance. Targeting this axis has the potential to improve CTX outcomes in patients with BC.     

Uptake of nickel from 316L stainless steel into contacting osteoblastic cells and metal ion interference with BMP-2-induced alkaline phosphatase

Bone cells contacting nickel (Ni)-containing implant materials may be affected by Ni species via disturbed signaling pathways involved in bone cell development. Here we analyze effects of the Ni-containing steel 316L and major metal constituents thereof on bone morphogenetic protein-2 (BMP-2)-induced alkaline phosphatase (ALP) of MC3T3-E1 cells. While cells grew normally on 316L, cellular Ni content increased 10-fold vs. control within 4 days. With respect to the major components of 316L, Ni2+ (3-50 microM) was most inhibitory to BMP-2-induced ALP, whereas even 50 microM Fe3+, Cr3+, Mo5+, or Mn2+ had no such effect. In line with this, BMP-2-induced ALP was significantly reduced in cells on 316L. This effect was not prevented by the metal ion chelator diethylenetriaminepentaacetic acid (DTPA). Instead, DTPA abolished the stimulatory effect of BMP-2 on ALP, pointing to chelatable metal ions involved. Zn2+, as one possible candidate, antagonized the Ni2+ inhibition of BMP-2-induced ALP in both MC3T3-E1 and human bone marrow stromal cells. Results show that cells contacting 316L steel are exposed to increased concentrations of Ni which suffice to impair BMP-2-induced ALP activity. Zn2+, as a competitor of this inhibition, may help to restore normal osteoblastic function and bone development under these conditions.     

"Nursing care visit: possibilities and limits". An empirical study in Bremen, Hamburg, Mecklenburg-Vorpommern, Lower Saxony and Schleswig-Holstein

This study examines the distribution, understanding and objectives of nursing visits as well as the implementation and the experience with nursing visits in 87 hospitals in North Germany (Bremen, Lower Saxony, Hamburg, Schleswig-Holstein and Mecklenburg-Western Pomerania). The survey was conducted using a questionnaire. It was found that nursing visits are implemented in 31% of the departments. By means of the nursing visit the patients are involved in the nursing process. The objectives of nursing visits are patient orientation. and the improvement of the nursing quality. The nursing visit is criticised because of both the level of organisation and the time spent on nursing visits as well as the lack of training, maintaining confidentiality and questions of data protection. The study contributes to a better understanding of nursing visits and a clearer definition of the term. It is the first empirical study of this type that has been conducted in Germany.