Gene expression differentiation in the reproductive tissues of Drosophila willistoni subspecies and their hybrids.

Early lineage diversification is central to understand what mutational events drive species divergence. Particularly, gene misregulation in interspecific hybrids can inform about what genes and pathways underlie hybrid dysfunction. In Drosophila hybrids, how regulatory evolution impacts different reproductive tissues remains understudied. Here, we generate a new genome assembly and annotation in Drosophila willistoni and analyse the patterns of transcriptome divergence between two allopatrically evolved D. willistoni subspecies, their male sterile and female fertile hybrid progeny across testis, male accessory gland, and ovary. Patterns of transcriptome divergence and modes of regulatory evolution were tissue-specific. Despite no indication for cell-type differences in hybrid testis, this tissue exhibited the largest magnitude of expression differentiation between subspecies and between parentals and hybrids. No evidence for anomalous dosage compensation in hybrid male tissues was detected nor was a differential role for the neo- and the ancestral arms of the D. willistoni X chromosome. Compared to the autosomes, the X chromosome appeared enriched for transgressively expressed genes in testis despite being the least differentiated in expression between subspecies. Evidence for fine genome clustering of transgressively expressed genes suggests a role of chromatin structure on hybrid gene misregulation. Lastly, transgressively expressed genes in the testis of the sterile male progeny were enriched for GO terms not typically associated with sperm function, instead hinting at anomalous development of the reproductive tissue. Our thorough tissue-level portrait of transcriptome differentiation between recently diverged D. willistoni subspecies and their hybrids provides a more nuanced view of early regulatory changes during speciation.

Ephrin-A5 potentiates netrin-1 axon guidance by enhancing Neogenin availability.

Axonal growth cones are guided by molecular cues in the extracellular environment. The mechanisms of combinatorial integration of guidance signals at the growth cone cell membrane are still being unravelled. Limb-innervating axons of vertebrate spinal lateral motor column (LMC) neurons are attracted to netrin-1 via its receptor, Neogenin, and are repelled from ephrin-A5 through its receptor EphA4. The presence of both cues elicits synergistic guidance of LMC axons, but the mechanism of this effect remains unknown. Using fluorescence immunohistochemistry, we show that ephrin-A5 increases LMC growth cone Neogenin protein levels and netrin-1 binding. This effect is enhanced by overexpressing EphA4 and is inhibited by blocking ephrin-A5-EphA4 binding. These effects have a functional consequence on LMC growth cone responses since bath addition of ephrin-A5 increases the responsiveness of LMC axons to netrin-1. Surprisingly, the overexpression of EphA4 lacking its cytoplasmic tail, also enhances Neogenin levels at the growth cone and potentiates LMC axon preference for growth on netrin-1. Since netrins and ephrins participate in a wide variety of biological processes, the enhancement of netrin-1 signalling by ephrins may have broad implications.

Adropin, nesfatin-1 and angiotensin II receptor expression in the abdominal aorta in ovariectomized rats after nesfatin-1 treatment.

The aim of the research was to assess the effect of nesfatin-1 on the structure, flexibility parameters, and expression of adropin, nesfatin-1, and angiotensin II receptor type 1 (AT1R) in the abdominal aorta in ovariectomized rats. Fragments of aortas were collected after euthanasia of female sham-operated (CONT) and ovariectomized Wistar rats (EXP), which were administered intraperitoneal injection of physiological saline (CONT, n = 7; EXP-O, n = 7) or nesfatin-1 (EXP-N, n = 7) in an amount of 2 µg/kg b.w. once a day for 8 weeks. The samples of aortas were collected for measurement of elasticity as well as histomorphometric, immunohistochemical, FTIR, and Raman spectroscopy analysis. The ovariectomy caused a significant increase in the thickness of the total wall and its particular layers in the aorta, in comparison to the CONT and EXP-N groups. However, the ovariectomy led to a decrease in the amount of elastin, collagen (mature, immature collagen, collagen maturity ratio 1660 - 1690 cm-1), and amides, with a simultaneous increase in lipids, especially in the tunica intima-media of the abdominal aorta compared to the other groups. The use of nesfatin-1 significantly increased the amount of collagen, elastin and amides with a simultaneous decrease in the amount of lipids and the expression of AT1R, adropin and nesfatin-1 in the abdominal aorta of ovariectomized rats. In conclusion, our study showed that the ovariectomy surgery induced changes in the abdominal aorta wall characteristic for aging females. Application of nesfatin-1 may prevent the negative consequences in the vessel wall structure in females in conditions of estrogen deficiency and prevent atherosclerotic changes in the cardiovascular system.

Factors controlling the reactivity of divalent metal ions towards pheophytin a.

In this study, we evaluate the factors which determine the reactivity of divalent metal ions in the spontaneous formation of metallochlorophylls, using experimental and computational approaches. Kinetic studies were carried out using pheophytin a in reactions with various divalent metal ions combined with non- or weakly-coordinative counter ions in a series of organic solvents. To obtain detailed insights into the solvent effect, the metalations with the whole set of cations were investigated in three solvents and with Zn2+ in seven solvents. The reactions were monitored using electronic absorption spectroscopy and the stopped-flow technique. DFT calculations were employed to shed light on the role of solvent in activating the metal ions towards porphyrinoids. This experimental and computational analysis gives detailed information regarding how the solvent and the counter ion assist/hinder the metalation reaction as activators/inhibitors. The metalation course is dictated to a large extent by the reaction medium, via either the activation or deactivation of the incoming metal ion. The solvent may affect the metalation in several ways, mainly via H-bonding with pyrrolenine nitrogens and the activation/deactivation of the incoming cation. It also seems to affect the activation enthalpy by causing slight conformational changes in the macrocyclic ligand. These new mechanistic insights contribute to a better understanding of the "metal-counterion-solvent" interplay in the metalation of porphyrinoids. In addition, they are highly relevant to the mechanisms of metalation reactions catalyzed by chelatases and explain the differences between the insertion of Mg2+ and other divalent cations.

Promoter hypermethylation of SHOX2 and SEPT9 is a potential biomarker for minimally invasive diagnosis in adenocarcinomas of the biliary tract.

BACKGROUND: Biliary tract carcinoma (BTC) is a fatal malignancy which aggressiveness contrasts sharply with its relatively mild and late clinical presentation. Novel molecular markers for early diagnosis and precise treatment are urgently needed. The purpose of this study was to evaluate the diagnostic and prognostic value of promoter hypermethylation of the SHOX2 and SEPT9 gene loci in BTC. METHODS: Relative DNA methylation of SHOX2 and SEPT9 was quantified in tumor specimens and matched normal adjacent tissue (NAT) from 71 BTC patients, as well as in plasma samples from an independent prospective cohort of 20 cholangiocarcinoma patients and 100 control patients. Receiver operating characteristic (ROC) curve analyses were performed to probe the diagnostic ability of both methylation markers. DNA methylation was correlated to clinicopathological data and to overall survival. RESULTS: SHOX2 methylation was significantly higher in tumor tissue than in NAT irrespective of tumor localization (p < 0.001) and correctly identified 71% of BTC specimens with 100% specificity (AUC = 0.918; 95% CI 0.865-0.971). SEPT9 hypermethylation was significantly more frequent in gallbladder carcinomas compared to cholangiocarcinomas (p = 0.01) and was associated with large primary tumors (p = 0.01) as well as age (p = 0.03). Cox proportional hazard analysis confirmed microscopic residual tumor at the surgical margin (R1-resection) as an independent prognostic factor, while SHOX2 and SEPT9 methylation showed no correlation with overall survival. Elevated DNA methylation levels were also found in plasma derived from cholangiocarcinoma patients. SHOX2 and SEPT9 methylation as a marker panel achieved a sensitivity of 45% and a specificity of 99% in differentiating between samples from patients with and without cholangiocarcinoma (AUC = 0.752; 95% CI 0.631-0.873). CONCLUSIONS: SHOX2 and SEPT9 are frequently methylated in biliary tract cancers. Promoter hypermethylation of SHOX2 and SEPT9 may therefore serve as a minimally invasive biomarker supporting diagnosis finding and therapy monitoring in clinical specimens.

[Descending necrotizing mediastinitis : Treatment concept with focus on thoracic surgery]. / Deszendierende nekrotisierende Mediastinitis : Therapiekonzept aus thoraxchirurgischer Sicht.

BACKGROUND: Descending necrotizing mediastinitis (DNM) originates from odontogenic or oropharyngeal infections which spread along preformed cervicothoracic spaces into the mediastinum and requires emergency multidisciplinary treatment. MATERIAL AND METHODS: A total of seven patients were diagnosed with DNM based on typical radiological features in a cervicothoracic computed tomography (CT) scan and subsequently underwent standardized transcervical and open transthoracic radical debridement. RESULTS: The initially detected polymicrobial spectrum of pathogens was dominated by streptococci followed by enterobacteriae. After calculated antibiotic treatment a shift in the spectrum of pathogens was noted and in particular a mycotic superinfection occurred in 43 % of the cases. Anterolateral thoracotomy was performed for radical removal of tissue necrosis and mediastinothoracic drainage extending to the posterior mediastinum was placed. In selected cases, cervico-mediastino-thoracic tubes were transmediastinally placed by the rendevouz technique either in the previsceral or retrovisceral mediastinal space. Despite predominantly advanced mediastinitis (Endo classification type II B) in this patient cohort, the mortality only reached 14 %. CONCLUSION: Rapid diagnosis, anatomical knowledge, understanding of the progression of infections as well as critical care, antimicrobial treatment and multidisciplinary radical surgical therapy are paramount for successful treatment of DNM. We favor anterolateral thoracotomy as the standard open transthoracic approach to the mediastinum. Placement of cervico-mediastino-thoracic irrigation drains can help to limit DNM.

Functional divergence of the miRNA transcriptome at the onset of Drosophila metamorphosis.

MicroRNAs (miRNAs) are endogenous RNA molecules that regulate gene expression posttranscriptionally. To date, the emergence of miRNAs and their patterns of sequence evolution have been analyzed in great detail. However, the extent to which miRNA expression levels have evolved over time, the role different evolutionary forces play in shaping these changes, and whether this variation in miRNA expression can reveal the interplay between miRNAs and mRNAs remain poorly understood. This is especially true for miRNA expressed during key developmental transitions. Here, we assayed miRNA expression levels immediately before (≥18BPF [18 h before puparium formation]) and after (PF) the increase in the hormone ecdysone responsible for triggering metamorphosis. We did so in four strains of Drosophila melanogaster and two closely related species. In contrast to their sequence conservation, approximately 25% of miRNAs analyzed showed significant within-species variation in male expression levels at ≥18BPF and/or PF. Additionally, approximately 33% showed modifications in their pattern of expression bias between developmental timepoints. A separate analysis of the ≥18BPF and PF stages revealed that changes in miRNA abundance accumulate linearly over evolutionary time at PF but not at ≥18BPF. Importantly, ≥18BPF-enriched miRNAs showed the greatest variation in expression levels both within and between species, so are the less likely to evolve under stabilizing selection. Functional attributes, such as expression ubiquity, appeared more tightly associated with lower levels of miRNA expression polymorphism at PF than at ≥18BPF. Furthermore, ≥18BPF- and PF-enriched miRNAs showed opposite patterns of covariation in expression with mRNAs, which denoted the type of regulatory relationship between miRNAs and mRNAs. Collectively, our results show contrasting patterns of functional divergence associated with miRNA expression levels during Drosophila ontogeny.

Functional evidence that a recently evolved Drosophila sperm-specific gene boosts sperm competition.

In many species, both morphological and molecular traits related to sex and reproduction evolve faster in males than in females. Ultimately, rapid male evolution relies on the acquisition of genetic variation associated with differential reproductive success. Many newly evolved genes are associated with novel functions that might enhance male fitness. However, functional evidence of the adaptive role of recently originated genes in males is still lacking. The Sperm dynein intermediate chain multigene family, which encodes a Sperm dynein intermediate chain presumably involved in sperm motility, originated from complex genetic rearrangements in the lineage that leads to Drosophila melanogaster within the last 5.4 million years since its split from Drosophila simulans. We deleted all the members of this multigene family resident on the X chromosome of D. melanogaster by chromosome engineering and found that, although the deletion does not result in a reduction of progeny number, it impairs the competence of the sperm in the presence of sperm from wild-type males. Therefore, the Sperm dynein intermediate chain multigene family contributes to the differential reproductive success among males and illustrates precisely how quickly a new gene function can be incorporated into the genetic network of a species.

KCH, the German preoperative score for isolated coronary artery bypass surgery: is it superior to the logistic EuroSCORE?

BACKGROUND: The accuracy of the logistic EuroSCORE in different patient populations has been questioned. Using the German registry database, the KoronarCHirurgie (KCH) score was introduced as a preoperative risk stratification tool specifically for patients who undergo isolated coronary artery bypass surgery in Germany. However, no direct statistical comparison of this score with the well-established logistic EuroSCORE has been previously performed. The aim of this study was to validate the preoperative German KCH score and to compare it to the logistic EuroSCORE for all coronary artery bypass surgery patients as well as for on-pump and off-pump subgroups. METHODS: We prospectively included all consecutive adult patients admitted to our department between January 1, 2007 and December 31, 2008, who underwent isolated coronary artery bypass surgery. The logistic EuroSCORE and the KCH-3.0 were calculated on admission to hospital. The outcome was defined as 30-day mortality. We performed calibration (Hosmer-Lemeshow test and Anderson-Grunkemeier Observed/Expected "O/E" mortality ratio) and discrimination (receiver operating characteristic "ROC" test) analyses of both scores. The accuracy of the scores was compared using DeLong's test. RESULTS: A total of 1461 patients (23.96 % females, mean age 66.94 ± 9.43 years) were included. The 30-day mortality rate was 2.87 %. The two models were comparable with regard to the prediction of an individual patient's risk of mortality in the whole study population and in the on-pump and off-pump subgroups (according to the ROC test and DeLong's test). Overall, there was no significant difference between observed and expected mortality according to the Hosmer-Lemeshow test ( P > 0.05). However, the KCH-3.0 was far less likely to overpredict mortality than the logistic EuroSCORE, as demonstrated by the observed mortality/expected mortality (O/E) ratios. The O/E ratio was 0.32 for the logistic EuroSCORE and 0.74 for the KCH-3.0. For the on-pump subgroup the O/E ratios were 0.37 and 0.80, respectively, and 0.24 and 0.63, respectively, for the off-pump subgroup. CONCLUSIONS: The KCH-3.0 is more reliable than the logistic EuroSCORE as a preoperative mortality prediction score for patients undergoing isolated coronary artery bypass surgery, providing predicted mortality rates that are closer to the actual mortality rates with a lower overprediction of mortality.

Freezing of the Nb5+ ion dynamics in AgNbO3 studied by linear and nonlinear dielectric response.

Linear and nonlinear dielectric measurements of AgNbO3 ceramics and single crystals were carried out for the M phases (77-673 K). The linear dielectric response is dominated by the contribution of the submillimetre relaxational mode related to the Nb5+ ion dynamics (M2-M3). On the other hand, nonlinear dielectric χ3' susceptibility revealed anomalies at three characteristic temperatures: 90, 325 and 448 K. Two later ones are connected with changes of the Nb5+ ion dynamics. At T(f) = 448 K a partial freezing of the Nb5+ ion displacement to the anti-polar, antiferroelectric array takes place. At 325 K further freezing of Nb and Ag displacements to the polar weak relaxor ferroelectric or dipolar glass transition occurs. This polar state coexists with the ground antiferroelectric one.

Coexistence of antiferromagnetic and spin cluster glass order in the magnetoelectric relaxor multiferroic PbFe 0.5 Nb 0.5 O3.

The coexistence of cluster glass with long-range antiferromagnetic order in the relaxor ferroelectric PbFe 0.5 Nb 0.5 O3 is elucidated. While the transition at T(N) = 153 K on the infinite antiferromagnetic cluster induces 3m symmetry with large EH2 magnetoelectric response, the disconnected subspace of isolated Fe3+ ions and finite clusters accommodates the cluster glass below T(g) = 10.6 K with field-induced m' symmetry and EH-type magnetoelectric response. Critical slowing-down, memory and rejuvenation after aging, occurrence of a de Almeida-Thouless phase line, and stretched exponential relaxation of remanence corroborate the glass nature.

Responses to iron limitation in Hordeum vulgare L. as affected by the atmospheric CO2 concentration.

Elevated atmospheric CO2 treatments stimulated biomass production in Fe-sufficient and Fe-deficient barley plants, both in hydroponics and in soil culture. Root/shoot biomass ratio was increased in severely Fe-deficient plants grown in hydroponics but not under moderate Fe limitation in soil culture. Significantly increased biomass production in high CO2 treatments, even under severe Fe deficiency in hydroponic culture, indicates an improved internal Fe utilization. Iron deficiency-induced secretion of PS in 0.5 to 2.5 cm sub-apical root zones was increased by 74% in response to elevated CO2 treatments of barley plants in hydroponics but no PS were detectable in root exudates collected from soil-grown plants. This may be attributed to suppression of PS release by internal Fe concentrations above the critical level for Fe deficiency, determined at final harvest for soil-grown barley plants, even without additional Fe supply. However, extremely low concentrations of easily plant-available Fe in the investigated soil and low Fe seed reserves suggest a contribution of PS-mediated Fe mobilization from sparingly soluble Fe sources to Fe acquisition of the soil-grown barley plants during the preceding culture period. Higher Fe contents in shoots (+52%) of plants grown in soil culture without Fe supply under elevated atmospheric CO2 concentrations may indicate an increased efficiency for Fe acquisition. No significant influence on diversity and function of rhizosphere-bacterial communities was detectable in the outer rhizosphere soil (0-3 mm distance from the root surface) by DGGE of 16S rRNA gene fragments and analysis of marker enzyme activities for C-, N-, and P-cycles.

IMRT for breast. a planning study.

BACKGROUND AND PURPOSE: To evaluate the performance of ten different treatment-planning systems when intensity modulated (IMRT) plans are designed for breast treatments that include the irradiation of the internal mammary chain. PATIENTS AND METHODS: A dataset of five patients (CT images and volumes of interest) was distributed to design IMRT plans on the ten systems. To minimise biases, the same geometry and clinical planning aims were imposed on the individual plans. Results were analysed in terms of dose distributions and dose volume histograms. RESULTS AND CONCLUSIONS: For target coverage, the volume receiving more than 95% of the prescribed dose ranged from 77% (OTP) to 91% (Eclipse and Pinnacle), the volume receiving more than 107% ranged from 3.3% (Hyperion) to 23.2% (OTP). The mean dose to ipsilateral lung ranged from 13 Gy (Eclipse) to 18 Gy (OTP). The volume of the contralateral breast receiving more than 10 Gy ranged from 3% (Pinnacle) to 26% (Precise). The volume of heart receiving more than 20 Gy ranged from 7% (Eclipse) to 47% (Precise), the maximum significant dose to heart ranged from approximately 27 Gy (XiO) to approximately 49 Gy (Precise). The maximum significant dose to healthy tissue ranged from approximately 51 Gy (Eclipse) to approximately 62 Gy (OTP). It was also possible to show that the treatment geometry proposed here enables to minimise contralateral breast irradiation while keeping minimal ipsilateral lung (or heart) involvement and satisfactory target coverage.

[Pulmonary lymphangioleiomyomatosis as a very rare interstitial lung disease]. / Limfangioleiomiomatoza jako jedna z bardzo rzadkich przyczyn chorób sródmiazszowych pluc.

Lymphangioleiomyomatosis (LAM) is a rare lung disease affecting premenopausal women characterized by an abnormal proliferation of smooth muscle cells that leads to the obstruction of airways, lymph and blood vessels. We present a case of a 46-year-old woman who was admitted to our department with dyspnoea and dry cough. The patient had a history of spontaneous pneumothorax 2 years prior to admission. Physical examination revealed dull percussion note on the lower right side of the chest. The chest X-ray film showed diffuse interstitial parenchymal infiltration and flattened costodiaphragmatic angle on the right side. The high resolution computerized tomography (HRCT) scan showed the numerous air filled cysts, about 25 mm in diameter with thin regular walls and liquid in the right pleural cavity. The effusion in a pleural cavity was chylous. Airway obstruction (FEV1/FVC 57% of predicted), markedly elevated residual volume (140%), and decreased DLCO were observed in functional pulmonary tests, and she underwent diagnostic videothoracoscopy. Pulmonary biopsy specimens confirmed diagnosis of LAM. The patient has been under careful observation, no treatment was instituted. The patient remains clinically stabile. During the last six months of observation she has normal sex hormone levels, therefore there exists a possibility of postmenopausal remission of symptoms.

Basic concepts of CORVUS dose model.

Basic concepts of the dose model utilized in the CORVUS treatment planning system are reviewed. Following the Peacock delivery tool (MIMiC) by NOMOS Corporation, CORVUS "delivers" radiation to a patient by means of narrow x-ray beams (pencil beams), which are subject to lateral electronic disequilibrium. Dose data for such beams are difficult to obtain experimentally. Therefore, the CORVUS dose model uses analytically calculated (rather than experimentally measured) narrow-beam dose data. The model is based on the idea that physical parameters necessary to calculate absorbed dose in narrow x-ray beams can be derived from measured broad-beam dose data. Calculation of central-axis and off-axis absorbed dose in narrow beams as well as a method of generating beam profiles are described.

New analogues of bradykinin containing a conformationally restricted dipeptide fragment in their molecules.

The present paper describes the synthesis and some pharmacological properties of two new bradykinin analogues containing the ethylene-bridged dipeptide Phe-Phe in their molecules. In a further two peptides this modification was combined with acylation of the N-terminus with 1-adamantaneacetic acid. Finally, we synthesized four analogues by removing the Ser6 residue from the four peptides mentioned above. The activity of the new analogues was assayed on isolated rat uterus (RUT) and in rat blood pressure tests (BPT). The results clearly indicate that the proposed modification, alone or in combination with other changes, resulted in either a drop in antiuterotonic activity or even in conversion to an agonism. Although this tendency is not so distinct in blood pressure assays, the antagonistic potency of the new analogues is also diminished. Nevertheless, it was demonstrated that the D-amino acid in position 7 which, until recently, was considered necessary for antagonism, may be replaced, together with the amino acid occupying position 8, by a suitable, sterically restricted L,L-dipeptide unit.

New bradykinin analogs in contraction of rat uterus.

In this study, we evaluated 20 of our previously synthesized peptides on isolated rat uterus by Holton's procedure with minor modifications, and compared their activity with that assessed previously by their ability to inhibit vasodepressor response to exogenous bradykinin (BK) in conscious rats. We used [D-Arg(0), Hyp(3), Thi(5, 8), (D-Phe)(7)]BK, the B(2) antagonist of Vavrek and Stewart as a model when designing our analogs. We observed that, in the case of the rat uterus test, the activity of peptides modified by acylation of the N-terminus with various bulky groups depends substantially on the chemical character of the substituent. We also learned that, contrary to previous examples, acylation of the N-terminus of antagonists, which contain a sterically restricted fragment in the C-terminal part, may not improve their antagonistic potencies. Besides an improved characterization of a series BK analogs, our studies have provided new information on the structure-activity relationship, which in turn may be of value in the design of more potent and selective bradykinin antagonists. The results of our studies appear to support the hypothesis of others about the presence of different subtypes of B(2) receptors in rat uterus and blood vessels.

Silver niobium trioxide, AgNbO3

The present structure determination of silver niobium trioxide at 291 K was performed on a twinned single crystal with a predominant presence [about 93 (1)%] of one twin domain. The sample contained traces of V (about 1 atomic %). This study confirms that the room-temperature phase of AgNbO(3) is isostructural with the room-temperature phase of NaNbO(3), i.e. it is a tilted perovskite. Structural deviation in AgNbO(3) from centrosymmetry was not detected in this study and its structure was refined in Pbcm, though a previous study indicated ferroelectricity below 350 K, in contrast with NaNbO(3).

Coordinate roles for LIM homeobox genes in directing the dorsoventral trajectory of motor axons in the vertebrate limb.

Motor neurons extend axons along specific trajectories, but the molecules that control their pathfinding remain poorly defined. We show that two LIM homeodomain transcription factors, Lim1 and Lmx1b, control the initial trajectory of motor axons in the developing mammalian limb. The expression of Lim1 by a lateral set of lateral motor column (LMC) neurons ensures that their axons select a dorsal trajectory in the limb. In a complementary manner, the expression of Lmx1b by dorsal limb mesenchymal cells controls the dorsal and ventral axonal trajectories of medial and lateral LMC neurons. In the absence of these two proteins, motor axons appear to select dorsal and ventral trajectories at random. Thus, LIM homeodomain proteins act within motor neurons and cells that guide motor axons to establish the fidelity of a binary choice in axonal trajectory.