User requirements on CT-based computed dose planning systems in radiation therapy. Presentation of 'check lists'

The expanding use of computers in radiation therapy procedures, especially the rapidly increasing use of digital CT-information, necessitates the coordination of the different systems in order to facilitate their developments. In order to define necessary demands for tomorrow a Nordic cooperation was initiated 1981 by NORDFORSK (Nordic co-operative organization for applied research), and a group of physicians and physicists having their daily work in this field of medicine and physics was invited to produce a report on 'User requirements on CT-based computed dose planning systems on radiation therapy'. The work has been done within the frame of NORDFORSK's activities and has been independent of the existing commissions and associations in the radiology field, but it has taken into consideration recommendations that have been given by or are being produced by other organizations. This report is a short summary of the complete paper which will be published in Acta Radiologica. The aim of this short version is to get an early presentation of the 'requirement lists' (see Appendix) which we think are of immediate importance.     

Flow cytometric multiparameter analysis of proliferating cell nuclear antigen/cyclin and Ki-67 antigen: a new view of the cell cycle

Flow cytometric multiparameter analysis of two proliferation-associated nuclear antigens (proliferating cell nuclear antigen (PCNA)/cyclin and Ki-67) was performed on seven human hematopoietic cell lines. PCNA/cyclin, an S phase-related antigen, was detected using an autoantibody and a fluorescein isothiocyanate-labeled anti-human antibody. The Ki-67 antigen, which in cycling cells is expressed with increasing levels during the S phase with a maximum in the M phase, was detected using a monoclonal antibody and a phycoerythrin-conjugated anti-mouse antibody. In some experiments the PCNA/Ki-67 staining was combined with a DNA stain, 7-amino actinomycin D, and simultaneous detection of the three stains was performed by a single laser flow cytometer. Using this technique four distinct cell populations, representing G1, S, G2, and M, respectively, could be demonstrated in cycling cells on the basis of their PCNA/cyclin and Ki-67 levels. The cell cycle phase specificity could be verified using metaphase (vinblastine, colcemide) and G2 phase (mitoxantrone) blocking agents, as well as by stainings with a mitosis-specific antibody (MPM-2). Also, G0 cells could be discriminated from G1 cells in analysis of a mixture of resting peripheral mononuclear blood cells and a proliferating cell line. This technique can be valuable in detailed cell cycle analysis, since all cell cycle phases can be visualized and calculated using a simple double staining procedure.     

Citric acid production in chemically defined media by Aspergillus niger

Summary The effect of changing the composition of a chemically defined medium on citric acid production by Aspergillus niger was investigated. High and reproducible amounts of citric acid were obtained with deionized commercial sugar solutions, proper phosphate concentrations, low initial pH values and suitable amounts of copper as growth inhibiting agent.Comparison of high and low yielding process parameters showed that under high yielding conditions, (deionized sugar, Cu⁺⁺ addition) besides more citric acid, less mycelium and less mycelial lipids were formed; the consumption of sugar, nitrogen and phosphorus was related to the amount of biomass.Partly presented at the XII. International Congress of Microbiology; München, September 3–8, 1978     

High‐Throughput DNA sequencing of ancient wood

Reconstructing the colonization and demographic dynamics that gave rise to extant forests is essential to forecasts of forest responses to environmental changes. Classical approaches to map how population of trees changed through space and time largely rely on pollen distribution patterns, with only a limited number of studies exploiting DNA molecules preserved in wooden tree archaeological and subfossil remains. Here, we advance such analyses by applying high‐throughput (HTS) DNA sequencing to wood archaeological and subfossil material for the first time, using a comprehensive sample of 167 European white oak waterlogged remains spanning a large temporal (from 550 to 9,800 years) and geographical range across Europe. The successful characterization of the endogenous DNA and exogenous microbial DNA of 140 (~83%) samples helped the identification of environmental conditions favouring long‐term DNA preservation in wood remains, and started to unveil the first trends in the DNA decay process in wood material. Additionally, the maternally inherited chloroplast haplotypes of 21 samples from three periods of forest human‐induced use (Neolithic, Bronze Age and Middle Ages) were found to be consistent with those of modern populations growing in the same geographic areas. Our work paves the way for further studies aiming at using ancient DNA preserved in wood to reconstruct the micro‐evolutionary response of trees to climate change and human forest management.     

Influence of different Sinorhizobium meliloti inocula on abundance of genes involved in nitrogen transformations in the rhizosphere of alfalfa (Medicago sativa L.)

Inoculation of leguminous seeds with selected rhizobial strains is practised in agriculture to ameliorate the plant yield by enhanced root nodulation and nitrogen uptake of the plant. However, effective symbiosis between legumes and rhizobia does not only depend on the capacity of nitrogen fixation but also on the entire nitrogen turnover in the rhizosphere. We investigated the influence of seed inoculation with two indigenous Sinorhizobium meliloti strains exhibiting different efficiency concerning plant growth promotion on nitrogen turnover processes in the rhizosphere during the growth of alfalfa. Quantification of six target genes (bacterial amoA, nirK, nirS, nosZ, nifH and archaeal amoA) within the nitrogen cycle was performed in rhizosphere samples before nodule formation, at bud development and at the late flowering stage. The results clearly demonstrated that effectiveness of rhizobial inocula is related to abundance of nifH genes in the late flowering phase of alfalfa. Moreover, other genes involved in nitrogen turnover had been affected by the inocula, e.g. higher numbers of amoA copies were observed during flowering when the more effective strain had been inoculated. However, the respective gene abundances differed overall to a greater extent between the three plant development stages than between the inoculation variants.     

A chicory cytochrome P450 mono-oxygenase CYP71AV8 for the oxidation of (+)-valencene

Chicory (Cichorium intybus L.), which is known to have a variety of terpene-hydroxylating activities, was screened for a P450 mono-oxygenase to convert (+)-valencene to (+)-nootkatone. A novel P450 cDNA was identified in a chicory root EST library. Co-expression of the enzyme with a valencene synthase in yeast, led to formation of trans-nootkatol, cis-nootkatol and (+)-nootkatone. The novel enzyme was also found to catalyse a three step conversion of germacrene A to germacra-1(10),4,11(13)-trien-12-oic acid, indicating its involvement in chicory sesquiterpene lactone biosynthesis. Likewise, amorpha-4,11-diene was converted to artemisinic acid. Surprisingly, the chicory P450 has a different regio-specificity on (+)-valencene compared to germacrene A and amorpha-4,11-diene.     

Combined metformin-salicylate treatment provides improved anti-tumor activity and enhanced radiotherapy response in prostate cancer; drug synergy at clinically relevant doses

BackgroundThere is need for well-tolerated therapies for prostate cancer (PrCa) secondary prevention and to improve response to radiotherapy (RT). The anti-diabetic agent metformin (MET) and the aspirin metabolite salicylate (SAL) are shown to activate AMP-activated protein kinase (AMPK), suppress de novo lipogenesis (DNL), the mammalian target of rapamycin (mTOR) pathway and reduce PrCa proliferation in-vitro. The purpose of this study was to examine whether combined MET+SAL treatment could provide enhanced PrCa tumor suppression and improve response to RT.MethodsAndrogen-sensitive (22RV1) and resistant (PC3, DU-145) PrCa cells and PC3 xenografts were used to examine whether combined treatment with MET+SAL can provide improved anti-tumor activity compared to each agent alone in non-irradiated and irradiated PrCa cells and tumors. Mechanisms of action were investigated with analysis of signaling events, mitochondria respiration and DNL activity assays.ResultsWe observed that PrCa cells are resistant to clinically relevant doses of MET. Combined MET + SAL treatment provides synergistic anti-proliferative activity at clinically relevant doses and enhances the anti-proliferative effects of RT. This was associated with suppression of oxygen consumption rate (OCR), activation of AMPK, suppression of acetyl-CoA carboxylase (ACC)-DNL and mTOR-p70^s6k/4EBP1 and HIF1α pathways. MET + SAL reduced tumor growth in non-irradiated tumors and enhanced the effects of RT.ConclusionMET+SAL treatment suppresses PrCa cell proliferation and tumor growth and enhances responses to RT at clinically relevant doses. Since MET and SAL are safe, widely-used and inexpensive agents, these data support the investigation of MET+SAL in PrCa clinical trials alone and in combination with RT.     

Spatial separation of litter decomposition and mycorrhizal nitrogen uptake in a boreal forest

Our understanding of how saprotrophic and mycorrhizal fungi interact to re-circulate carbon and nutrients from plant litter and soil organic matter is limited by poor understanding of their spatiotemporal dynamics. In order to investigate how different functional groups of fungi contribute to carbon and nitrogen cycling at different stages of decomposition, we studied changes in fungal community composition along vertical profiles through a Pinus sylvestris forest soil. We combined molecular identification methods with 14C dating of the organic matter, analyses of carbon:nitrogen (C:N) ratios and 15N natural abundance measurements. Saprotrophic fungi were primarily confined to relatively recently (< 4 yr) shed litter components on the surface of the forest floor, where organic carbon was mineralized while nitrogen was retained. Mycorrhizal fungi dominated in the underlying, more decomposed litter and humus, where they apparently mobilized N and made it available to their host plants. Our observations show that the degrading and nutrient-mobilizing components of the fungal community are spatially separated. This has important implications for biogeochemical studies of boreal forest ecosystems.