Crosstalk of the mitotic spindle assembly checkpoint with p53 to prevent polyploidy

Treatment of cells with microtubule inhibitors results in activation of the mitotic spindle assembly checkpoint, leading to mitotic arrest before anaphase. Upon prolonged treatment, however, cells can adapt and exit mitosis aberrantly, resulting in the occurrence of tetraploid cells in G1. Those cells subsequently arrest in postmitotic G1 due to the activation of a p53-dependent G1 checkpoint. Failure of the G1 checkpoint leads to endoreduplication and further polyploidization. Using HCT116 and isogenic p53-deficient or spindle checkpoint compromised derivatives, we show here that not only p53 but also a functional spindle assembly checkpoint is required for postmitotic G1 checkpoint function. During transient mitotic arrest, p53 stabilization and activation is triggered by a pathway independent of ATM/ATR, Chk1 and Chk2. We further show that a prolonged spindle checkpoint-mediated mitotic arrest is required for proper postmitotic G1 checkpoint function. In addition, we demonstrate that polyploid cells are inhibited to re-enter mitosis by an additional checkpoint acting in G2. Thus, during a normal cell cycle, polyploidization and subsequent aneuploidization is prevented by the function of the mitotic spindle checkpoint, a p53-dependent G1 checkpoint and an additional G2 checkpoint.     

Immunocytochemical characterization of rat brainstem neurons with vagal afferent input from the stomach challenged by acid or ammonia

Exposure of the gastric mucosa to backdiffusing acid is signalled to the brainstem via vagal afferents. This study examined whether exposure of the Sprague-Dawley rat stomach to hydrochloric acid (HCl) or ammonium hydroxide (NH4OH), a noxious chemical produced by Helicobacter pylori, activates different vagal afferent pathways as reflected by different circuitries in the medullary brainstem. Two hours after intragastric treatment with HCl or NH4OH the activation of neurons in the nucleus tractus solitarii at the rostrocaudal extension of the area postrema (NTSAP) was visualized by c-Fos immunohistochemistry and their chemical coding characterized by double-labelling immunohistochemistry. Exposure of the rat gastric mucosa to HCl (0.15-0.5 M) or NH4OH (0.1-0.3 M) led to a concentration-dependent expression of c-Fos in the NTSAP. The number and distribution of NTSAP neurons activated by 0.35 M HCl and 0.3 M NH4OH were similar; the highest number of activated neurons occurring in the medial part of the NTSAP. Some 60% of the NTSAP neurons activated by intragastric HCl and NH4OH stained for the high affinity glutamate transporter EAAC1, while some 30% contained calbindin or neuropeptide Y. Glutamate receptors of the N-methyl-D-aspartate type were found on approximately 50% of the c-Fos-positive cells in the NTSAP, whereas tachykinin NK1, NK2 and NK3 receptors were present on 5-10% of the activated neurons. The similar number and distribution of c-Fos-expressing neurons within the NTSAP and their identical chemical coding indicate that exposure of the rat stomach to backdiffusing concentrations of HCl and NH4OH activates the same vagal afferent-NTSAP pathway.     

Reconstituted high-density lipoprotein exhibits neuroprotection in two rat models of stroke

BACKGROUND: Reconstituted high-density lipoprotein (rHDL) is prepared from apolipoprotein A-I, isolated from human plasma, and soybean-derived phosphatidylcholine and exhibits biochemical and functional characteristics similar to endogenous nascent high-density lipoprotein (HDL). This study tested the hypothesis that pretreatment with rHDL may reduce neuronal damage in 2 experimental rat models of stroke. METHODS: In the first model, an excitotoxic lesion was induced by unilateral injection of N-methyl-D-aspartate (NMDA) in the right striatum (excitotoxic lesion model). In the second model, temporary occlusion of the middle cerebral artery (MCA) was attained by inserting a nylon thread through the carotid artery and blood flow was restored 30 min later (MCAo model). In both models, either rHDL (120 mg/kg) or saline (control) were infused over 4 h, starting 2 h before the injection of NMDA or the induction of ischemia, respectively. 24 h after the interventions, the rats were sacrificed and the brains removed for histochemical preparation. The necrotic area was delimited using an image analysis system. In addition, the levels of reactive oxygen species (ROS) in human endothelial (ECV 304) and neuroblastoma (SK-N-BE) cell lines were measured fluorometrically as 2',7'-dichlorofluorescein fluorescence in the presence and absence of rHDL and under basal and stress-induced conditions. RESULTS: In the excitotoxic lesion and MCAo models, pretreatment with rHDL significantly reduced the brain necrotic area by 61 and 76%, respectively (p < 0.01). Overnight incubation of ECV 304 and SK-N-BE cells with 0.5 mg/ml rHDL decreased basal and stress-induced ROS levels by 73 and 72% (ECV 304) and by 76 and 43% (SK-N-BE), respectively (p < 0.01). CONCLUSION: These results suggest that rHDL reduces neuronal damage after onset of ischemic stroke, possibly by involving an anti-oxidative mechanism. Thus, rHDL may be a powerful neuroprotective tool for the treatment of cerebrovascular diseases.     

Effects of intraduodenal application of peppermint oil (WS(R) 1340) and caraway oil (WS(R) 1520) on gastroduodenal motility in healthy volunteers

Enteric-coated preparations containing a fixed peppermint oil/caraway oil combination (e.g. Enteroplant(R)) are widely used in patients with functional dyspepsia. The effect of a separate intraduodenal application of the active ingredients of Enteroplant(R) (90 mg peppermint oil (WS(R) 1340), 50 mg caraway oil (WS(R) 1520) per capsule) and of a hydrophobic phase galenic auxiliary material contained in the Enteroplant(R) formulation (dose as contained per capsule) on gastroduodenal motility were studied with stationary manometry in healthy volunteers. The carrier demanded by the experimental setup also served as a control. The results showed that: (1) during phase III of the migrating motor complex (MMC) the frequency and duration of contractions showed a significant decrease in the duodenum for WS(R) 1340; (2) WS(R) 1520 significantly reduced the contraction amplitudes in the duodenum during phase I and II of the MMC; (3) trends for decreased values were seen for WS(R) 1340 in the gastric corpus and duodenum and for WS(R) 1520 in the gastric antrum; (4) in the gastric corpus the duration of contractions was significantly reduced after application of WS(R) 1340 during phases I and II of the MMC; (5) WS(R) 1520 significantly reduced the contraction amplitudes and the duration of contractions in the gastric corpus during phase III of the MMC; (6) for the hydrophobic phase a moderate but significant decrease of duration of contractions in the duodenum and of frequency of contractions in the gastric corpus was seen. No adverse events were observed during the study. In conclusion, it could be shown that both WS(R) 1340 and WS(R) 1520 contribute to the efficacy of Enteroplant(R). They act locally in the stomach and duodenum to produce smooth-muscle relaxation. The effects of the active ingredients WS(R) 1340 and WS(R) 1520 substantially exceed the effects observed with the galenic auxiliary material and the carrier, respectively.     

Structure activity relationship studies of cinnamic acid derivatives as inhibitors of human neutrophil elastase revealed by ligand docking calculations

Structure-activity relationship of cinnamic acid derivatives as inhibitors of the human neutrophil elastase is reported. Comparison of the inhibitory concentrations (IC50 values) with the results of the ligand docking calculations revealed that the structure element of the aromatic ortho-dihydroxy groups combined with a lipophilic residue seems to be a prerequisite for an optimal binding within the active site.     

Chemokine-induced cell death in CCR5-expressing neuroblastoma cells

CCR5 is expressed in neurons but its function in this cellular context is hitherto poorly understood. We have generated CCR5-expressing SH-SY5Y neuroblastoma cells. CCR5 ligands induced cell death in these cells, but not in control neuroblastoma cells or in CCR5-expressing fibroblasts. CCR5-dependent killing of neuroblastoma cells occurred through apoptosis, since it was accompanied by caspase-3 activation and could be prevented by a caspase-3 inhibitor. Finally, cell killing by activated microglia was more rapid and extensive in CCR5-expressing neuroblastoma cells than in control cells. In summary, CCR5 may act as a death receptor in cells of neuronal lineage and therefore be involved in inflammatory neurodegeneration.     

Zur Wirkungsweise neuer insektizider Stoffe

Ohne ZusammenfassungHerrn Professor Dr. Wolfgang Heubner zum 70. Geburtstag gewidmet.     

The impact of an additional ecotoxicity test on ecological quality standards

The present study aims to support decisions on whether or not to perform an extra toxicity test in order to improve environmental quality standards (EQSs). The impact of an additional ecotoxicity test was analyzed by predicting new ecotoxicity values with three different estimation methods and adding them to existing species sensitivity distributions (SSDs) on which the EQSs are based. The results show that EQSs are likely to increase due to increasing sample size, but the change also depends on the number of toxicity values available, the estimation method used and the representativeness of the species tested. The management consequences are illustrated in a case study on contaminated freshwater sediment in the Netherlands. It is shown that a slight increase of the EQS can result in a large reduction of sediment remediation costs without impairing regulatory protection levels. The paper identifies indicators that can be used to evaluate the potential impact of an extra ecotoxicity test.     

Pterine als Wirkstoffe und Pigmente

Ohne Zusammenfassung Mit 5 Abbildungen