Cell culture studies on neurofibromatosis (von Recklinghausen)

Summary The growth of strains of fibroblasts derived from patients with neurofibromatosis (NF) was compared with that of strains from appropriate controls in culture medium containing 1% or 15% fetal calf serum. The means of the ratios of final to initial cell numbers do not differ significantly between NF strains and control strains. Weakly significant differences are, however, obtained after conversion of the results to mean numbers of cell population doublings, the NF strains showing the higher numbers. The ratios of final to initial amounts of protein also differ significantly under both sets of growth conditions. High growth parameters occur significantly more frequently among our smaple of 11 NF strains than among our sample of 13 control strains. The possibility of the expression of the NF genotype(s) on the level of the cultured fibroblast-like cells and the possible causes of the large ranges of inter-and intra-individual variations of the results are discussed.     

Identification and characterization of CYP79D6v4, a cytochrome P450 enzyme producing aldoximes in black poplar (Populus nigra)

After herbivore feeding, poplar trees produce complex volatile blends containing terpenes, green leaf volatiles, aromatics, and nitrogen-containing compounds such as aldoximes and nitriles. It has been shown recently that volatile aldoximes released from gypsy moth (Lymantria dispar) caterpillar-damaged black poplar (Populus nigra) trees attract parasitoids that are caterpillar enemies. In western balsam poplar (P. trichocarpa), volatile aldoximes are produced by 2 P450 monooxygenases, CYP79D6v3 and CYP79D7v2. A gene fragment with high similarity to CYP79D6/7 was recently shown to be upregulated in herbivore-damaged leaves of P. nigra. In the present study we report the cloning and characterization of this gene, designated as CYP79D6v4. Recombinant CYP79D6v4 was able to convert different amino acids into the corresponding aldoximes, which were also found in the volatile blend of P. nigra. Thus, CYP79D6v4 is most likely involved in herbivore-induced aldoxime formation in black poplar.     

Fish oil slows S phase progression and may cause upstream shift of DHFR replication origin ori-beta in CHO cells

Fish oils (FOs) have been noted to reduce growth and proliferation of certain tumor cells, effects usually attributed to the content of polyunsaturated fatty acids of the n-3 family, which are thought to modulate cellular signaling pathways. We investigated the influence of FO on cell cycle kinetics of cultured Chinese hamster ovary cells. Exponentially growing cells were labeled with 5-bromo-2'-deoxyuridine (BrdU) and analyzed by flow cytometry after 5-day treatment with exogenous fat. Bivariate BrdU-DNA analysis indicated slower progression through S phase and thus longer S phase duration time in FO- but not corn oil-treated or control cells. We hypothesize that FO treatment might interfere with spatial/temporal organization of replication origins. Therefore, we mapped the well-characterized replication origin ori-beta downstream of the dihydrofolate reductase gene with the nascent strand length assay. Three DNA marker segments with known positions relative to this origin were amplified by PCR. By quantitatively assessing DNA length of the fragments in all fractions containing these markers, the location of ori-beta was established. In control or corn oil-treated cells, the location of ori-beta was consistent with previous studies. However, in FO-treated cells, DNA replication appears to start from a new site located farther upstream from ori-beta, suggesting a different replication initiation pattern. This study suggests novel mechanism(s) by which fats affect cell proliferation and DNA replication in mammalian cells.     

An european pupil project linked to the scientific aims of the experiment AQUARIUS-XENOPUS on the taxi Soyuz flight Andromede to ISS

The German-French biological experiment AQUARIUS-XENOPUS which flew on the Soyuz flight Andromede to the International Space Station ISS (launched October 21, 2001 in Baikonour/Kazakhstan) was extended by an outreach project. Pupils of class 10 to 12 from Ulm/D and Nancy-Tomblaine/F studied swimming behavior of Xenopus tadpoles on ground. They were instructed to perform all experimental steps following the protocol of similar video recordings on ISS. After the flight, they evaluated the kinetics of swimming of both ground controls and space animals. The pupil project included theoretical components to introduce them to the field of gravitational biology. One feature of the project was the exchange of ideas between pupils by meetings which took place in Ulm (June 2001), Nancy (February 2002) and Paris (May 2002). We consider our approach as a successful way to include young people in space experiments on a cheap cost level and to bring ideas of gravitational biology into the curricula of European schools.     

Advanced glycation endproducts influence the mRNA expression of RAGE, RANKL and various osteoblastic genes in human osteoblasts

Glycation reactions resulting in the generation and accumulation of advanced glycation endproducts (AGEs) are potential mechanisms by which bone protein may be altered in vivo. AGEs accumulate in the bone increasingly with age come into close contact with osteoblasts or osteoclasts. The direct effect of AGEs on bone cells has not been thoroughly investigated. This study aimed to examine whether glycated bovine serum albumin (AGE - BSA) as an AGE modulate the mRNA expression of various genes in primary human osteoblast cultures. The following parameters were included: RAGE (receptor for AGEs), alkaline phosphatase, osteocalcin, osterix and RANKL (receptor activator of nuclear factor-kappaB ligand). Primary human osteoblast cultures were obtained from bone specimens of six patients with osteoarthrosis. Human osteoblasts were treated in AGE - BSA or control-BSA (non-glycated BSA) containing medium (5 mg/ml each) over a time course of seven days. After RT-PCR the mRNA expression was measured by real-time PCR. Related to control - BSA exposure, the mRNA expression of RAGE, RANKL and osterix increased during AGE - BSA treament. For alkaline phosphatase and osteocalcin a tendency of down-regulation was found. In summary, the study presents evidence that advanced glycation end products accumulated in bone alter osteoblasts by activation the AGE - RAGE pathway (RAGE mRNA up-regulation), inducing enhanced osteoclastogenesis (RANKL mRNA up-regulation) and impaired matrix mineralization (down-regulation of alkaline phosphatase and osteocalcin mRNA). Thus, AGEs may play a functional role in the development of bone diseases (e.g. osteoporosis).     

Plant community diversity influences allocation to direct chemical defence in Plantago lanceolata

Background

Forecasting the consequences of accelerating rates of changes in biodiversity for ecosystem functioning requires a mechanistic understanding of the relationships between the structure of biological communities and variation in plant functional characteristics. So far, experimental data of how plant species diversity influences the investment of individual plants in direct chemical defences against herbivores and pathogens is lacking.

Methodology/Principal Findings

We used Plantago lanceolata as a model species in experimental grasslands differing in species richness and composition (Jena Experiment) to investigate foliar concentrations of the iridoid glycosides (IG), catalpol and its biosynthetic precursor aucubin. Total IG and aucubin concentrations decreased, while catalpol concentrations increased with increasing plant diversity in terms of species or functional group richness. Negative plant diversity effects on total IG and aucubin concentrations correlated with increasing specific leaf area of P. lanceolata, suggesting that greater allocation to light acquisition reduced the investment into these carbon-based defence components. In contrast, increasing leaf nitrogen concentrations best explained increasing concentrations of the biosynthetically more advanced IG, catalpol. Observed levels of leaf damage explained a significant proportion of variation in total IG and aucubin concentrations, but did not account for variance in catalpol concentrations.

Conclusions/Significance

Our results clearly show that plants growing in communities of varying species richness and composition differ in their defensive chemistry, which may modulate plant susceptibility to enemy attack and consequently their interactions with higher trophic level organisms.