Nuclear metabolism of benzo(a)pyrene and of (±)-trans-7,8-dihydroxy-7,8,-dihydrobenzo(a)pyrene: Comparative chromatographic analysis of alkylated DNA

Rat liver nuclei were incubated with [¹⁴C]benzo(a)pyrene (BP) or [³H](±)-trans-7,8-dihydrodiol of BP (³H-BP-7,8-diol) in the presence of a NADPH-generating system. The nuclei were able to form from BP the 9,10-, 4,5- and 7,8-dihydrodiols, the 3,6- and 1,6-quinones as well as the 3- and 9-phenols. The total nuclear metabolism was stimulated 11-fold by prior administration to the rats of 3-methylcholanthrene (3MC). BP-7,8-dihydrodiol formation, under these circumstances, was enhanced 29-fold. The rat liver nuclei were also able to form from [³H]BP-7,8-diol, (±)-7β,8α-dihydroxy-9β,10β-epoxy-7,8,9,10-tetrahydro BP (diol epoxide 1), (±)-7β,8α-dihydroxy-9α,10α-epoxy-7,8,9,10-tetrahydro BP (diol epoxide 2), as well as three unknown metabolites. Diol epoxides 1 and 2 represented 23 and 65% of the total metabolites produced during the control nuclear incubation. Pretreatment of the rats with 3MC resulted in 4-fold increase in nuclear metabolic activity. Under the latter circumstances, the diol epoxides 1 and 2 represented 43 and 38%, respectively, of the total nuclear metabolites. Incubation of liver nuclei with labeled BP or BP-7,8-diol in the presence of NADPH resulted in alkylation of DNA. The alkylated deoxyribonucleosides were separated by Sephadex LH-20 chromatography. Two peaks of radioactivity were noted after incubation with the parent polycyclic hydrocarbon while only one peak was seen after incubation with the diol derivative. These results emphasize the importance of nuclei in the metabolism of BP and in the subsequent alkylation of DNA, reactions which may be related to mutagenesis or carcinogenesis.     

Mammalian target of rapamycin (mTOR): a central regulator of male fertility?

Mammalian target of rapamycin (mTOR) is a central regulator of cellular metabolic phenotype and is involved in virtually all aspects of cellular function. It integrates not only nutrient and energy-sensing pathways but also actin cytoskeleton organization, in response to environmental cues including growth factors and cellular energy levels. These events are pivotal for spermatogenesis and determine the reproductive potential of males. Yet, the molecular mechanisms by which mTOR signaling acts in male reproductive system remain a matter of debate. Here, we review the current knowledge on physiological and molecular events mediated by mTOR in testis and testicular cells. In recent years, mTOR inhibition has been explored as a prime strategy to develop novel therapeutic approaches to treat cancer, cardiovascular disease, autoimmunity, and metabolic disorders. However, the physiological consequences of mTOR dysregulation and inhibition to male reproductive potential are still not fully understood. Compelling evidence suggests that mTOR is an arising regulator of male fertility and better understanding of this atypical protein kinase coordinated action in testis will provide insightful information concerning its biological significance in other tissues/organs. We also discuss why a new generation of mTOR inhibitors aiming to be used in clinical practice may also need to include an integrative view on the effects in male reproductive system.     

Generation of goats by nuclear transfer: a retrospective analysis of a commercial operation (1998–2010)

At LFB USA, Inc., the ultimate use for transgenic cloned goats is for the production of recombinant human protein therapeutics in their milk. This retrospective analysis of the Somatic Cell Nuclear Transfer (SCNT) program, spanning from 1998 to 2010, examined parameters potentially affecting the outcomes and efficiencies in this commercial operation. Over 37,000?+?ova were utilized in the SCNT protocol producing a total of 203 cloned goats. Fifty one (51) clones were produced from non-transfected (transgenic and non-transgenic animal donor) cell lines and 152 clones were produced from transfected cell lines. Comparisons and summaries of (a) transfected versus non-transfected cell lines, (b) relationship of SCNT parameters to offspring produced, (c) skin versus fetal cells, (d) fresh versus cryopreserved cells, (e) parameters from all cell lines used versus those producing SCNT offspring, (f) variation among cell sources, (g) methods of SCNT parturition management and effects on live offspring, and lastly (h) SCNT variation by program are reported. Findings indicate that (a) non-transfected cell lines were more efficient versus transfected cell lines in generating viable cloned offspring on a per reconstructed embryo transferred basis, (b) transfected fetal fibroblasts had improved efficiency versus transfected skin fibroblasts, (c) the percentage of non-transfected cell lines that produced offspring was statistically higher than transfected cell lines, (d) and induction of parturition improved the percentage of viable offspring. In summary, this retrospective analysis on the SCNT process has identified certain parameters for improved efficiency in producing viable cloned goats in a commercial setting.

     

Comment on Altaba (2015): a case of species misidentification?

Previously in this journal, Altaba (2015) presented an original hypothesis on the dispersal of small wetland snails in Central Europe after the Last Glacial Maximum, based on the discovery of rare species of the genus Vallonia at the foot of the Devín Hill (Slovakia). On the basis of the evidence available to us, it is our opinion that the existence of Vallonia declivis, Vallonia suevica, Vallonia enniensis, and Vallonia tenuilabris at Devín Gate has not been established, and that the hypothesis advanced in Altaba (2015) is therefore unsubstantiated by hard evidence.     

Klaus Apel (1942–2017): a pioneer of photosynthesis research

We present here a Tribute to Klaus Apel (1942–2017), a photosynthesis pioneer—an authority on plant molecular genetics—in five parts. The first section is a prologue. The second section deals with a chronological discussion of Apel’s research life, prepared by the editor Govindjee; it is based on a website article at the Boyce Thompson Institute (BTI) by Patricia Waldron (https://btiscience.org/explore-bti/news/post/bti-says-goodbye-klaus-apel/), as approved for use here by Keith C. Hannon and David Stern of BTI. The third section, which focuses on Apel’s pioneering work on singlet oxygen-mediated EXECUTER-dependent signaling in plants, is written by two of us (J-DR and CK). The fourth section includes three selected reminiscences, one from BTI and two from ETH (Eidgenössische Technische Hochschule). This tribute ends with section five, which is a very brief presentation of Klaus Apel’s personal life, by Wiebke Apel.     

QSAR study on pK(a) vis-à-vis physiological activity of sulfonamides: a dominating role of surface tension (inverse steric parameter)

The paper describes the dominating role of surface tension (ST) on the modeling, monitoring, and estimating pK(a) for a large series of 43 substituted sulfonamides. Because of the direct correlation of ST with parachor (Pc) vis-a-vis molecular volume (MV), ST is considered as a steric parameter. Single as well as multi-parametric regressions have indicated that ST has a dominating role in QSAR of the set of sulfonamides used and that excellent results are obtained in multi-parametric regression analysis. The results are discussed critically on the basis of statistical parameters.     

Evolution of dark colour in toucans (Ramphastidae): a case of molecular adaptation?

In the last decades, researchers have been able to determine the molecular basis of some phenotypes, to test for evidence of natural selection upon them, and to demonstrate that the same genes or genetic pathways can be associated with convergent traits. Colour traits are often subject to natural selection because even small changes in these traits can have a large effect on fitness via camouflage, sexual selection or other mechanisms. The melanocortin‐1 receptor locus (MC1R) is frequently associated with intraspecific coat colour variation in vertebrates, but it has been far harder to demonstrate that this locus is involved in adaptive interspecific colour differences. Here, we investigate the contribution of the MC1R gene to the colour diversity found in toucans (Ramphastidae). We found divergent selection on MC1R in the clade represented by the genus Ramphastos and that this coincided with the evolution of darker plumage in members of this genus. Using phylogenetically corrected correlations, we show significant and specific relationships between the rate of nonsynonymous change in MC1R (dN) and plumage darkness across Ramphastidae, and also between the rate of functionally significant amino acid changes in MC1R and plumage darkness. Furthermore, three of the seven amino acid changes in MC1R that occurred in the ancestral Ramphastos branch are associated with melanism in other birds. Taken together, our results suggest that the dark colour of Ramphastos toucans was related to nonsynonymous substitutions in MC1R that may have been subject to positive selection or to a relaxation of selective pressure. These results also demonstrate a quantitative relationship between gene and phenotype evolution, representing an example of how MC1R molecular evolution may affect macroevolution of plumage phenotypes.     

Spawning aggregations of squid (Sepioteuthis australis) populations: a continuum of ‘microcohorts’

The aim of this study was to determine how size, age, somatic and reproductive condition, abundance and egg production of southern calamary spawning aggregations changed during the spawning season in each of 2 years. During the spawning period in at least one of the years there was a decline as much as 20% in average size, 50% in somatic condition, 28–34% in size-at-age, 26–29% in reproductive status, as well as abundance and reproductive output of the stock declining during the spawning season. However, this change was not a function of the population becoming reproductively exhausted, as the aggregation was composed of different individuals with different biological characteristics. In each month the average age of individuals was ca. 6 mo, indicating that squid that had hatched at different times had entered the spawning aggregations, suggesting that the aggregation was made-up of a succession of microcohorts. Currently, management of many squid populations assumes that there is a single cohort in the aggregation. Therefore, estimating stock biomass at the start of the spawning season cannot be used as the population is constantly changing as micro-cohorts move into the aggregation. An instantaneous estimate of the spawning biomass, independent of fishing activity may be obtained by quantifying the density of deposited eggs. The strategy of individuals with a diversity of life history characteristics coming together in a single spawning aggregation may ensure the phenotypic and genetic diversity required to guarantee successful recruitment of this short-lived species. Therefore, temporally structured protection from harvest throughout the spawning season will ensure maintenance of this population diversity.     

Generation of transgenic goats by pronuclear microinjection: a retrospective analysis of a commercial operation (1995–2012)

Production of transgenic founder goats involves introducing and stably integrating an engineered piece of DNA into the genome of the animal. At LFB USA, the ultimate use of these transgenic goats is for the production of recombinant human protein therapeutics in the milk of these dairy animals. The transgene or construct typically links a milk protein specific promoter sequence, the coding sequence for the gene of interest, and the necessary downstream regulatory sequences thereby directing expression of the recombinant protein in the milk during the lactation period. Over the time period indicated (1995–2012), pronuclear microinjection was used in a number of programs to insert transgenes into 18,120, 1- or 2- cell stage fertilized embryos. These embryos were transferred into 4180 synchronized recipient females with 1934 (47%) recipients becoming pregnant, 2594 offspring generated, and a 109 (4.2%) of those offspring determined to be transgenic. Even with new and improving genome editing tools now available, pronuclear microinjection is still the predominant and proven technology used in this commercial setting supporting regulatory filings and market authorizations when producing founder transgenic animals with large transgenes (> 10 kb) such as those necessary for directing monoclonal antibody production in milk.     

Dissemination of the entomopathogenic fungus verticillium lecanii (Zimmermann) Viégas (Hyphomycetales: Moniliaceae) in a population of Frankliniella occidentalis (Pergande, 1895) (Thysanoptera: Thripidae)

Tests were carried out to investigate the dissemination of the entomopathogenic fungus Verticillium lecanii (Zimmermann) Viégas in a population of Frankliniella occidentalis. The tested factors, which influence the efficacy of the fungus against the pest insect, have been the population density of the thrips at the application moment as well as the temperature. The population density influenced the dissemination of the fungal spores in the population. The higher the density has been, the higher the insetting control effect has been as well. The temperature influenced the speed of the fungal effect, too. The higher the temperature has been, the earlier the control effect started. However, an increase of the natural mortality was found as well.     

Temporal relationships of a pulse of prolactin (PRL) to a pulse of a metabolite of PGF2α in mares

Hourly blood samples were collected from 10 mares during 24 h of each of the preluteolytic, luteolytic, and postluteolytic periods. The autocorrelation function of the R program was used to detect pulse rhythmicity, and the intra-assay CV was used to locate and characterize pulses of prolactin (PRL) and a metabolite of prostaglandin F2α (PGFM). Rhythmicity of PRL and PGFM concentrations was detected in 67% and 89% of mares, respectively. Combined for the three periods (no difference among periods), the PRL pulses were 5.2 ± 0.4 h (mean ± SEM) at the base, 7.5 ± 1.5 h between nadirs of adjacent pulses, and 12.3 ± 1.5 h from peak to peak. The peaks of PRL pulses were greater (P < 0.05) during the luteolytic period (46 ± 14 ng/mL) and postluteolytic period (52 ± 15 ng/mL) than during the preluteolytic period (17 ± 3 ng/mL). Concentrations of PRL during hours of a PGFM pulse were different (P < 0.003) within the luteolytic period and postluteolytic period and were greatest at the PGFM peak; PRL concentrations during a PGFM pulse were not different during the preluteolytic period. The frequency of the peak of PRL and PGFM pulses occurring at the same hour (synchrony) was greater for the luteolytic period (65%, P < 0.01) and postluteolytic period (50%, P < 0.001) than for the preluteolytic period (17%). This is the first report in mares on characterization and rhythmicity of PRL pulses, synchrony between PRL and PGFM pulses, and greater PRL activity during the luteolytic and postluteolytic periods than during the preluteolytic period.     

Genetic erosion in a stenotopic heathland ground beetle (Coleoptera: Carabidae): a matter of habitat size?

Two centuries ago large areas of north-west Europe were covered by coherent heathlands which hosted numerous specialized species. Changes in land use made heathlands fragmented and rare, consequently, they are in the focus of nature conservation efforts today. But how large should remaining heathland patches be in order to secure the survival of populations of specialized species? We investigated the genetic diversity at five allozyme loci of Poecilus lepidus, a flightless and stenotopic heathland ground beetle. 29 populations from differently sized heathland patches in north-west Germany were analyzed. Results show a weak but significant genetic differentiation and no evidence for isolation by distance or other patterns of spatial autocorrelation. Linear regression analysis revealed significant relationships between patch size, allelic richness, number of alleles and expected heterozygosity. These findings are explained by severe habitat fragmentation together with strong fluctuations in population size which have been reported for this species in the past. To conserve the vast majority of the species’ genetic diversity for a period of 100 years we suggest to maintain heathland patches of at least 50 ha in size.     

Isolation of a Neurotoxin (α-colubritoxin) from a Nonvenomous Colubrid: Evidence for Early Origin of Venom in Snakes

The evolution of venom in advanced snakes has been a focus of long-standing interest. Here we provide the first complete amino acid sequence of a colubrid toxin, which we have called -colubritoxin, isolated from the Asian ratsnake Coelognathus radiatus (formerly known as Elaphe radiata), an archetypal nonvenomous snake as sold in pet stores. This potent postsynaptic neurotoxin displays readily reversible, competitive antagonism at the nicotinic receptor. The toxin is homologous with, and phylogenetically rooted within, the three-finger toxins, previously thought unique to elapids, suggesting that this toxin family was recruited into the chemical arsenal of advanced snakes early in their evolutionary history. LC-MS analysis of venoms from most other advanced snake lineages revealed the widespread presence of components of the same molecular weight class, suggesting the ubiquity of three-finger toxins across advanced snakes, with the exclusion of Viperidae. These results support the role of venom as a key evolutionary innovation in the early diversification of advanced snakes and provide evidence that forces a fundamental rethink of the very concept of nonvenomous snake.     

Leaf variegation in Caladium steudneriifolium (Araceae): a case of mimicry?

The leaves of Caladium steudneriifolium (Araceae) of the understorey of a submontane rainforest in the Podocarpus National Park (South East Ecuador, 1,060 m a.s.l.) are plain green or patterned with whitish variegation. Of the 3,413 individual leaves randomly chosen and examined in April 2003, two-thirds were plain green, whereas one third were variegated (i.e., whitish due to absence of chloroplasts). Leaves of both morphs are frequently attacked by mining moth caterpillars. Our BLAST analysis based on Cytochrome-c-Oxidase-subunit-1 sequences suggests that the moth is possibly a member of the Pyraloidea or another microlepidopteran group. It was observed that the variegated leaf zones strongly resemble recent damages caused by mining larvae and therefore may mimic an attack by moth larvae. Infestation was significantly 4–12 times higher for green leaves than for variegated leaves. To test the hypothesis that variegation can be interpreted as mimicry to deter ovipositing moths, we first ruled out the possibility that variegation is a function of canopy density (i.e., that the moths might be attracted or deterred by factors unrelated to the plant). Then plain green leaves were artificially variegated and the number of mining larvae counted after 3 months. The results on infestation rate (7.88% of green leaves, 1.61% of the variegated leaves, 0.41% of white manipulated leaves and 9.12% of uncoloured manipulated leaves) suggest that ovipositing moths are deterred by the miner-infestation mimicry. Thus, variegation might be beneficial for the plants despite the implicated loss of photosynthetically active surface.     

Vietnamese species of Taumacera viridis–group with a description of new species (Coleoptera: Chrysomelidae: Galerucinae)

The species of Taumacera viridis species–group from Vietnam are revised. Four species are recognized. Taumacera indica (Jacoby, 1889), T. occipitalis (Laboissière, 1933) and T. variceps (Laboissière, 1933), are redescribed. Taumacera phuquoca sp. nov. from Phu Quoc National Park (southern Vietnam) is described. An identification key is provided for all Vietnamese species.www.zoobank.org/urn:lsid:zoobank.org:pub:396862F9-8A47-4101-A27C-CD0625E68FA1.     

Fracture risk and the use of a diuretic (indapamide sr) ± perindopril: a substudy of the Hypertension in the Very Elderly Trial (HYVET)

Background

Chronic pancreatitis is an inflammatory disease which is characterized by an irreversible conversion of pancreatic parenchyma to fibrous tissue. Beside obstructive jaundice and pseudocyst formation, about half of the patients need surgical intervention due to untreatable chronic pain during the course of the disease. In most of the patients with chronic pancreatitis, the head of the pancreas is the trigger of the chronic inflammatory process. Therefore, resection of pancreatic head tissue must be the central part of any surgical intervention. However, it is unclear to which extent the surgical procedure must be radical in order to obtain a favourable outcome for the patients.

Design

A single centre randomized controlled, superiority trial to compare two techniques of duodenum preserving pancreatic head resection. Sample size: 65 patients will be included and randomized intraoperatively. Eligibility criteria: All patients with chronic pancreatitis and indication for surgical resection and signed informed consent. Cumulative primary endpoint (hierarchical model): duration of surgical procedure, quality of life after one year, duration of intensive care unit stay, duration of hospital stay. Reference treatment: Resection of the pancreatic head with dissection of the pancreas from the portal vein and transsection of the gland (Beger procedure). Intervention: Partial Resection of the pancreatic head without transsection of the organ and visualization of the portal vein (Berne procedure). Duration: September 2003-October 2007.

Organisation/responsibility

The trial is conducted in compliance with the protocol and in accordance with the moral, ethical, regulatory and scientific principles governing clinical research as set out in the Declaration of Helsinki (1989) and the Good Clinical Practice guideline (GCP). The Center for Clinical Studies of the Department of Surgery Heidelberg is responsible for planning, conducting and final analysis of the trial.