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La pandemia COVID ha hecho de las consultas telemáticas una herramienta básica en la práctica diaria. El objetivo principal del estudio es valorar los resultados de la aplicación de consultas telemáticas para limitar la movilidad de los pacientes.

Hydrocracking catalysts with large porosity need to be developed to treat heavy oil feedstocks rich in large molecular weight components such as asphaltenes and withstand deactivation due to coke formation. In this work, catalysts were prepared by impregnation of varying NiMo loadings on a mesoporous Al2O3support. The effect of metal loading on the hydrocracking of a vacuum residue at three temperatures (400, 425, and 450 °C) was studied in a batch microbomb reactor. Catalysts were reutilized in a second reaction with fresh feed to assess their activity following the initial period where results are dominated by a large carbon deposition. The textural properties and the coke content on the spent catalysts were evaluated after both reactions. It was found that the reaction temperature had an important effect on the conversion of the fraction with a boiling point above 450 °C, whereas metal loading had minimal impact. On the other hand, metal loading had a significant effect on hydrodeasphaltenization (HDA); higher asphaltene conversions were obtained with higher metal loading. Reaction temperature had an influence on HDA, particularly for lower metal loadings, as catalysts with higher loadings showed significant activity at the lower temperatures studied. It was observed that coke deposits were mainly formed during the initial hour of reaction with little additional coke being formed in the reutilization of the catalysts. More deposits were obtained at lower reaction temperature, as coke precursors, polyaromatic hydrocarbons, polymerize into coke. No evidence of pore mouth plugging was observed, indicating that catalysts could accommodate coke while retaining most of their textural properties. Catalysts with higher metal loadings took longer to reach a stable amount of deposits, but they stabilized at an overall smaller coke deposition and retained significant HDA activity.

Fasting is commonly used in aquaculture to empty the gut before slaughter, but little is known about how feeding frequency before fasting affects the stress response of trout. To find out more, 240 rainbow trout (Oncorhynchus mykissj were separated into three groups with different feeding schedules during the final month of fattening, from 26 September to 28 October 2013 (daily, every two days or every four days) and two durations of pre-slaughter fasting (two days of fasting; 24.3 degree days, to nine days of fasting; 102 degree days). After slaughter, a number of stress-related parameters were measured, such as liver glycogen, skin/gill colour and haematological parameters (Cortisol, glucose, lactate, triglycerides, lactate dehydrogenase and creatine phosphokinase). Trout given food every two days on the farm had lower levels of Cortisol and higher levels of triglycerides and liver glycogen than the other treatments after two days of fasting; indicating that habituating trout to feed once every two days in the final month of fattening lowered their stress response to two days of fasting before slaughter. [ABSTRACT FROM AUTHOR]

Allogeneic hemopoietic stem cell transplantation (HSCT) is the only method currently available to cure transfusion-dependent thalassemia major that has been widely used worldwide. To verify transplantation distribution, demography, activity, policies and outcomes inside the European Group for Blood and Marrow Transplantation (EBMT), we performed a retrospective non-interventional study, extracting data from the EBMT hemoglobinopathy prospective registry database. We included 1493 consecutive patients with thalassemia major transplanted between 1 January 2000 and 31 December 2010. In total, 1359 (91%) transplants were performed on patients <18 years old, 1061 were from a human leukocyte Ag-identical sibling donor. After a median observation time of 2 years, the 2-year overall survival (OS) and event-free survival (EFS; that is, thalassemia-free survival) were 88±1% and 81±1%, respectively. Transplantation from a human leukocyte Ag-identical sibling offered the best results, with OS and EFS of 91±1% and 83±1%, respectively. No significant differences in survival were reported between countries. The threshold age for optimal transplant outcomes was around 14 years, with an OS of 90–96% and an EFS of 83–93% when transplants were performed before this age. Allogeneic HSCT for thalassemia is a curative approach that is employed internationally and produces excellent results.

Prosthetic devices in dentistry are commonly made of acrylic resin. This material has been continuously improved and nanotechnology is now an important part of the process. The main objective of dentures is their restorative function, while preventing the adherence of microorganisms present in the oral mucosa. Recently, nanopigments, fibers and silver nanoparticles were incorporated into poly(methyl methacrylate) formulations, thus obtaining dental materials with a reduced porosity, increased flexural strength and showing a lower Candida albicans adherence. The modified acrylic resin was subjected to in vitro cytotoxicity and genotoxicity tests, resulting in a biocompatible option for denture bases. The history, challenges, innovations and recent patents based on nanoparticles in prosthetic materials for dentures are reviewed in this paper.

Surface engineering provides a powerful tool to create model systems to investigate the complex interactions between cells and the extracellular matrix (ECM). Cell adhesive peptide ligands (e.g. Arginine-Glycine-Aspatate (RGD) sequence) and carbohydrates are abundant in the ECM, and both are involved in mediating cell adhesion. However, how these interact is not well understood in vivo. Model systems to investigate their collective roles are rare. We have developed a robust surface patterning method allowing spatial control of carbohydrate (i.e. mannose) and RGD on glass while overcoming non-specific biomolecular interactions. The method effectively combines a traditional photolithographic process with two robust chemical pathways, namely click chemistry and silanization, in order to tailor the surface chemistry. This method is especially promising for the quantitative immobilization of different molecules and holds potential for the investigation of interactions between carbohydrates and common ECM proteins on cell adhesion.

The electrooxidation of methanol was studied at elevated temperature and pressure by cyclic voltammetry and constant potential experiments at real fuel cell electrocatalysts. Ruthenium core and platinum shell nanoparticles were synthesized by a sequential polyol route, and characterized electrochemically by CO stripping at room temperature to quickly confirm the structure of the synthesized core–shell structure as compared to pure commercial Pt/C and Pt–Ru/C alloy catalysts. A significant promotional effect of Pt decorated Ru cores in the methanol oxidation was found at elevated temperatures and rather high‐electrode potentials. A negative potential shift of the methanol oxidation peak is observed for the Ru@Pt/C core–shell catalyst at moderate temperatures, while a significant shift to positive potentials of the methanol oxidation peak occurs for Pt/C catalysts. The onset potential for methanol oxidation is lowered some 200 mV from room temperature and up to 120 °C for all electrocatalysts, indicating that it is the thermal activity of water adsorption that dictates the onset potential. Direct methanol fuel cell experiments showed only small performance differences between Ru@Pt/C and Pt/C anode electrocatalysts, suggesting the necessity of render possible the formation of surface oxygen species at lower electrode potentials.

To evaluate the utility of a post-discharge pharmaceutical care programme.

Analizar el efecto de la proteína BMP-7 en el tratamiento de los defectos óseos con transporte óseo.

Rhipicephalus (Boophilus) microplusis an important cause of economic losses in Thailand through direct effects of feeding on cattle and pathogen transmission. Current tick control methods rely on expensive chemical acaricides that result in environmental contamination, residues in food animal products and acaricide-resistant ticks. Anti-tick vaccines based on concealed antigens have shown promising results in the control of cattle tick. Thus, recombinant Bm95 (rBm95) from Thai R. microplus(KU-VAC1) was cloned and expressed to test as an anti-tick vaccine in Thailand. The objective of this study was to compare antibody responses induced by KU-VAC1 to that obtained after vaccination with Gavac that is based on the Bm86 homologue. Four groups of six cattle each were immunized with KU-VAC1, Gavac, adjuvant or phosphate-buffered saline, and boosted three times at 21-day intervals. Enzyme-linked-immunosorbent serologic assay were used to measure the humoral antibody responses specific to Thai rBm95. Cattle immunized with either KU-VAC1 or Gavac showed significantly greater antibody production than the controls. Antibody titres were detected after the first immunization and peaked after the seventh week. These results indicated that KU-VAC1 and Gavac are similarly immunogenic, and that further studies are warranted to compare performance parameters of ticks fed on immunized cattle.

Molecules secreted from the tick salivary gland modulate the vertebrate host immune response, thus representing potential targets for novel tick control measures. Tick salivary gland serine protease inhibitor (Serpin) is one such molecule that may facilitate tick feeding, blood meal digestion and pathogen transmission. The objective of this study was to determine the immunogenicity and protection of recombinant Rhipicephalus (Boophilus) microplussalivary gland Serpin (rSerpin) in rabbits. Rabbits were injected with rSerpin, adjuvant or phosphate-buffered saline (PBS) alone, and challenge infested with 500 R. micropluslarvae that were allowed to continuously feed and moult through the adult stage. All immunized rabbits generated antibodies to rSerpin in the second week after immunization. Ticks fed on immunized rabbits resulted in 83 reduction in adult engorgement and 34 reduction in egg mass weight compared with the PBS control. These results indicated that this tick Serpin is immunogenic to rabbits, and suggested that this vaccine candidate antigen can confer protective immunity against cattle ticks in this experimental model.

The objective of this study is to analyse the available evidence regarding the effectiveness of the strategy of induction maintenance with boosted protease inhibitors with ritonavir in adult HIV patients as compared to conventional treatment.

Pt and Au are not miscible within a whole range of concentrations. To obtain PtAu alloys, severe thermal treatments are required that to provide aggregation phenomena. However, it is possible to synthesize bimetallic PtAu nanoparticles provided the proper synthesis route is employed. When they are prepared from water-in-oil microemulsions or with the impregnation technique, it is possible to obtain nanosized bimetallic PtAu particles. In contrast, other colloidal routes have been seen to be adequate for the synthesis of other bimetallic Pt-based particles, affording segregated samples with Pt- or Au-enriched zones. When alloyed, bimetallic PtAu nanoparticles display unique physicochemical properties that are different from those of monometallic and nonalloyed solids. Thus, the performance of alloyed PtAu samples as electrocatalysts for the oxygen reduction reaction is superior to that of the PtAu-segregated samples. In fact, the ability of carbon-supported bimetallic PtAu samples in the oxygen reduction reactions equals or even surpasses that of archetypal Pt/C electrocatalysts.

Human granulocytic anaplasmosis (HGA), an emerging disease of public health concern in many areas of the world, is caused by Anaplasma phagocytophilum.Small animal models of A phagocytophilumin laboratory mice have been developed and used to study the pathogenesis of HGA. In this study, we characterized the pathologic changes in acute infection of C3H/HeJ mice experimentally infected with the NY18 isolate of A phagocytophilum.Although no clinical signs were noted, acute infection was associated with gross splenomegaly, microscopic inflammatory lesions in the lung and liver, hyperplastic lesions on the spleen, and clinical pathology abnormalities including neutropenia and monocytosis. This study emphasizes the use of well-defined animal models as a valuable tool for the study of A phagocytophiluminfections.

ABSTRACTIn the context of a serosurvey conducted on the Anaplasma marginaleprevalence in Swiss cattle, we suspected that a serological cross-reactivity between A. marginaleand A. phagocytophilummight exist. In the present study we demonstrate that cattle, sheep and horses experimentally infected with A. phagocytophilumnot only develop antibodies to A. phagocytophilum(detected by immunofluorescent-antibody assay) but also to A. marginale(detected by a competitive enzyme-linked immunosorbent assay). Conversely, calves experimentally infected with A. marginalealso developed antibodies to A. phagocytophilumusing the same serological tests. The identity of 63% determined in silico within a 209-amino-acid sequence of major surface protein 5 of an isolate of A. marginaleand one of A. phagocytophilumsupported the observed immunological cross-reactivity. These observations have important consequences for the serotesting of both, A. marginaleand A. phagocytophiluminfection of several animal species. In view of these new findings, tests that have been considered specific for either infection must be interpreted carefully.

The labeling of targeting molecules with nanoparticles has revolutionized the visualization of cellular or tissue components by electron microscopy. A particularly desirable target is the nucleus, because the genetic information is there. To date, utilizing nanoparticles for nuclear targeting has not proved very successful due to the impermeable nature of the plasma and nuclear membranes; thus nanoparticle design and synthesis is a critical factor. We report in this article the synthesis of water-soluble gold nanoparticles functionalized with a Tat protein-derived peptide sequence by a straightforward and economical methodology. The particles were subsequently tested in vitro with a human fibroblast cell line by optical and transmission electron microscopy to determine the biocompatibility of these nanoparticles and whether the functionalization with the translocation peptide allowed particles to transfer across the cell membrane and locate in the nucleus.

External mass-transfer coefficients (kf) during supercritical fluid extraction (SCFE) of high-solubility solutes, under solvent upflow conditions and low superficial velocities, can be small because of the negative influence of natural convection phenomena. A shrinking-core model for mass transfer was used to estimate best-fit values of kf for data on SCFE of lipids from prepressed rapeseeds. Values of kf at a high Reynolds number (Re = 14.1) were similar when using solvent upflow or downflow, but kf at lower Re (1.57) was 3.6 times smaller when using solvent upflow than that predicted from a literature correlation for downflow conditions. These kf's are consistent with values estimated by fitting literature data, or gathered from various sources under similar, nonadequate conditions (solvent upflow under low Re) for the extraction of both fatty and essential oils. Care is advisable when employing best-fit values of kf from laboratory data for process design purposes, especially when sizing of the solvent pumps for the experiments is questionable.

The tick-borne intracellular pathogen Anaplasma marginale (Rickettsiales: Anaplasmataceae) develops persistent infections in cattle and tick hosts. While erythrocytes appear to be the only site of infection in cattle, A. marginale undergoes a complex developmental cycle in ticks and transmission occurs via salivary glands during feeding. Many geographic isolates occur that vary in genotype, antigenic composition, morphology and infectivity for ticks. In this chapter we review recent research on the host–vector–pathogen interactions of A. marginale. Major surface proteins (MSPs) play a crucial role in the interaction of A. marginale with host cells. The MSP1a protein, which is an adhesin for bovine erythrocytes and tick cells, is differentially regulated and affects infection and transmission of A. marginale by Dermacentor spp. ticks. MSP2 undergoes antigenic variation and selection in cattle and ticks, and contributes to the maintenance of persistent infections. Phylogenetic studies of A. marginale geographic isolates using msp4 and msp1α provide information about the biogeography and evolution of A. marginale: msp1α genotypes evolve under positive selection pressure. Isolates of A. marginale are maintained by independent transmission events and a mechanism of infection exclusion in cattle and ticks allows for only the infection of one isolate per animal. Prospects for development of control strategies by use of pathogen and tick-derived antigens are discussed. The A. marginale/vector/host studies described herein could serve as a model for research on other tick-borne rickettsiae.

A Chlamydophila abortus-induced abortion model was carried out on the basis of the experimental infection of ewes at day 75 of gestation. The infection induced abortions and the birth of weak lambs during the last 3 weeks of pregnancy. To study the kinetics of the infection in the placenta and in other organs, infected ewes were killed at 105, 120, and 130 days of gestation and also several days after abortion or parturition. Infected ewes developed a systemic infection that caused a mild and transient pneumonia and focal hepatitis. Pathologic changes were observed in placentas at 120 day of gestation, although the lesions varied between animals and even between placentomes of the same placenta. The first placental area infected was the maternal stroma and epithelium next to the intercaruncular areas, where neutrophilic response seemed to control the infection. A substantial degree of multiplication of C. abortuswas then observed in the trophoblast cells of the placentome, periplacentomal choriallantoic membranes, and hilius, with an inflammatory exudate composed mainly of neutrophils, some macrophages, and very scarce lymphocytes. After abortion, the lesions affected the intercotyledonary areas of the aborted placentas, whereas in the uterus significant lymphocyte infiltration was observed, together with a rapid decrease of the C. abortusantigen in the degenerated caruncular tissues.

In addition to the known mutisicoumaranones A (1) and B (2), the methanolic extract of the aerial parts of the shrub Mutisia friesiana afforded two new 5-methylcoumaranones, mutisicoumaranones C (3) and D (4). Their structures were elucidated by spectroscopic methods. ¹³C NMR data for mutisicoumaranones A and B are reported for the first time. All compounds showed antifungal activity against the phytopathogenic fungus Cladosporium cucumerinum and bactericidal activity against Staphylococcus aureus. The presence of 5-methylcoumaranones A−D is biosynthetically related to 5-methylcoumarins and phenolic derivatives previously isolated from M. friesiana.

Aldose reductase (ALR2) has been implicated in the etiology of diabetic complications, including blindness. Because of the limited number of currently available drugs for the prevention of these long-term complications, the discovery of new ALR2 inhibitors appears highly desirable. In this study, a polybrominated diphenyl ether (1) naturally occurring in a marine sponge was found to inhibit recombinant human ALR2 with an IC50 of 6.4 μM. A series of polyhalogenated analogues that were synthesized and tested in vitro to explore the structure−activity relationships displayed various degrees of inhibitory activity. The most active compounds were also capable of preventing sorbitol accumulation inside human retinal cells. In this cell-based assay, the most potent synthesized analogue (16) showed a 17-fold increase in inhibitory activity compared to that of sorbinil (IC50 = 0.24 vs 4 μM). A molecular representation of human ALR2 in complex with the natural product was built using homology modeling, automated docking, and energy refinement methods. AMBER parameters for the halogen atoms were derived and calibrated using condensed phase molecular dynamics simulations of fluorobenzene, chlorobenzene, and bromobenzene. Inhibitor binding is proposed to cause a conformational change similar to that recently reported for zenarestat. A free energy perturbation thermodynamic cycle allowed us to assess the importance of a crucial bromine atom that distinguishes the active lead compound from a much less active close natural analogue. Remarkably, the spatial location of this bromine atom is equivalent to that occupied by the only bromine atom present in zenarestat.

The objective of this study was to examine the time during the postfertilization period that gene expression patterns in in vitro-cultured bovine embryos diverge from those of their in vivo-cultured counterparts. Presumptive bovine zygotes were produced by in vitro maturation and fertilization of immature oocytes collected from the ovaries of slaughtered animals. Approximately 20 h post insemination (hpi), zygotes were denuded and randomly divided into two groups for culture either in vitro, in synthetic oviduct fluid medium, or in vivo, in the ewe oviduct. Embryos were recovered from both systems at approximately 30 hpi (2-cell), 2 (4-cell), 3 (8-cell), 4 (16-cell), 5 (early morula), 6 (compact morula), or 7 (blastocyst) days post insemination. On recovery, they were examined for stage of development and snap frozen in liquid nitrogen for the analysis of transcript abundance using real-time polymerase chain reaction. The transcripts studied were glucose transporter 5, sarcosine oxidase, mitochondrial Mn-superoxide dismutase, connexin 43, interferon tau, insulin-like growth factor II, apoptosis regulator box-α and insulin-like growth factor-I receptor, most of which are known from our previous work to differ in terms of transcript abundance in blastocysts derived from culture in vitro or in vivo. The results demonstrate that the relative abundance of the transcripts studied varies throughout the preimplantation period and is strongly influenced by the culture environment. In addition, the data demonstrate that changes in transcript abundance in blastocyst stage embryos are in many cases a consequence of perturbed transcription earlier in development. Depending on the transcript, these differences may be evident by as little as 10 h of initiation of culture. Such information has implications not only for basic biology but also for human assisted reproduction in which there is a move toward culturing embryos to the blastocyst stage, necessitating prolonged culture in vitro under potentially deleterious conditions.

The reproductive tract of Chu‐Lin, a giant panda that had died at Madrid Zoo, was removed. Four hours postmortem the epididymides was cut with a blade, and the spermatozoa were collected and immersed in a freezing medium. The sample was kept under refrigeration at 5°C. After 4 hr the sperm suspension was aspirated into 0.25‐mL straws, which were held for 7 min in nitrogen vapors before they were plunged into liquid nitrogen. To characterize sperm population and distribution, a morphometric study of the triple‐stained smears of the thawed semen sample was undertaken with the aid of computerized image analysis. Spermatozoa with intact acrosomes were selected and assessed using an immersion lens (1,000×) and bright‐field illumination. The software made it possible to take six basic linear measurements of each spermatozoon: maximum head width (HW), base of the head (BH), maximum head length (HL), midpiece length (ML), midpiece width (MW), and terminal piece length (TL). The system was connected to a data base from which derived parameters could be calculated: head form factor (HL/HW), total flagellum length (TFL), total sperm length (TSL), and the proportion of each of the sections in relation to the whole. The results of the morphometric study did not demonstrate the existence of sperm subpopulations, as the measurements of the 100 spermatozoa analyzed were relatively homogeneous with regard to their distribution for each of the parameters considered. The sperm head of the giant panda displayed a relatively rounded morphology, and the HL/HW was 1.24±0.03. Mean head length was 5.05±0.08 µ, and mean head width was 4.07±0.08 µ. The mean total length of the giant panda spermatozoon was 49.26±0.67 µ. Zoo Biol 22:355–364, 2003. © 2003 Wiley‐Liss, Inc.

We have previously shown that, while the intrinsic quality of the oocyte is the main factor affecting blastocyst yield during bovine embryo development in vitro, the main factor affecting the quality of the blastocyst is the postfertilization culture conditions. Therefore, any improvement in the quality of blastocysts produced in vitro is likely to derive from the modification of the postfertilization culture conditions. The objective of this study was to examine the effect of the presence or absence of serum and the concentration of BSA during the period of embryo culture in vitro on 1) cleavage rate, 2) the kinetics of embryo development, 3) blastocyst yield, and 4) blastocyst quality, as assessed by cryotolerance and gene expression patterns. The quantification of all gene transcripts was carried out by real-time quantitative reverse transcription-polymerase chain reaction. Bovine blastocysts from four sources were used: 1) in vitro culture in synthetic oviduct fluid (SOF) supplemented with 3 mg/ml BSA and 10% fetal calf serum (FCS), 2) in vitro culture in SOF + 3 mg/ml BSA in the absence of serum, 3) in vitro culture in SOF + 16 mg/ml BSA in the absence of serum, and 4) in vivo blastocysts. There was no difference in overall blastocyst yield at Day 9 between the groups. However, significantly more blastocysts were present by Day 6 in the presence of 10% serum (20.0%) compared with 3 mg/ml BSA (4.6%, P< 0.001) or 16 mg/ml BSA (11.6%, P< 0.01). By Day 7, however, this difference had disappeared. Following vitrification, there was no difference in survival between blastocysts produced in the presence of 16 mg/ml BSA or those produced in the presence of 10% FCS; the survival of both groups was significantly lower than the in vivo controls at all time points and in terms of hatching rate. In contrast, survival of blastocysts produced in SOF + 3 mg/ml BSA in the absence of serum was intermediate, with no difference remaining at 72 h when compared with in vivo embryos. Differences in relative mRNA abundance among the two groups of blastocysts analyzed were found for genes related to apoptosis (Bax), oxidative stress (MnSOD, CuZnSOD, and SOX), communication through gap junctions (Cx31 and Cx43), maternal recognition of pregnancy (IFN-τ), and differentiation and implantation (LIF and LR-β). The presence of serum during the culture period resulted in a significant increase in the level of expression of MnSOD, SOX, Bax, LIF, and LR-β. The level of expression of Cx31 and Cu/ZnSOD also tended to be increased, although the difference was not significant. In contrast, the level of expression of Cx43 and IFN-τ was decreased in the presence of serum. In conclusion, using a combination of measures of developmental competence (cleavage and blastocyst rates) and qualitative measures such as cryotolerance and relative mRNA abundance to give a more complete picture of the consequences of modifying medium composition on the embryo, we have shown that conditions of postfertilization culture, in particular, the presence of serum in the medium, can affect the speed of embryo development and the quality of the resulting blastocysts. The reduced cryotolerance of blastocysts generated in the presence of serum is accompanied by deviations in the relative abundance of developmentally important gene transcripts. Omission of serum during the postfertilization culture period can significantly improve the cryotolerance of the blastocysts to a level intermediate between serum-generated blastocysts and those derived in vivo. The challenge now is to try and bridge this gap.

Photoreduction of 3-phenylquinoxalin-2-ones, XNQ, by triethylamine, TEA, gives a metastable semireduced quinoxalin-2-one via an electron−proton−electron transfer, with unit quantum yields at high amine concentrations. During the photoreduction, an ion-radical pair, XNQ^-•/TEA^+•, a neutral-radical pair, XNQH^•/TEA-H^•, and an ion-pair, XNQH^-/TEA-H⁺, are formed. We present time-resolved spectroscopic data and quantum mechanical semiempirical AM1, PM3, and ZINDO/S results for the transient species formed during the flash photolysis of quinoxalin-2-ones in the presence of amines. These calculations show that the neutral-radical pair and the ion-pair are similar in energy, and that the calculated spectra of all the transient species should have similar absorption bands near 400 nm in agreement with experimental results. The ZINDO/S calculated spectra of the XNQH^-/iminium ion pair fit the experimental spectra and explain the lack of visible or near-IR absorption of the metastable compound. Energy changes between the species involved are of interest with regard to the possible use of quinoxalin-2-one/amine systems for light to chemical energy conversion or as temporal data storage devices.

Using reverse transcriptase-amplified fragment length polymorphism (RT-AFLP) analysis of differential mRNA expression and semiquantitative reverse transcriptase-polymerase chain reaction, we compared mRNA expression in bovine blastocysts from 4 sources, known to differ in quality in terms of their ability to withstand cryopreservation: 1) in vitro culture in synthetic oviduct fluid of in vitro-matured (IVM)/in vitro fertilized (IVF) zygotes; 2) in vitro culture in TCM-199 supplemented with granulosa cells (coculture) of IVM/IVF zygotes; 3) in vivo culture in the ewe oviduct of IVM/IVF zygotes; or 4) superovulation, artificial insemination, and nonsurgical embryo recovery. Total mRNA was isolated from pools of blastocysts and reverse transcription was performed. Triplicate reactions from each sample were displayed, and only consistent banding variations were recorded. Using AFLP-differential display assay, we found that cDNA banding patterns are highly conserved between the 4 groups of blastocysts studied; however, there was a difference of 7% in bands either missing or expressed across the groups. Fifty bands were reamplified, and a sequence comparison search revealed similarity of 14 isolated fragments to ribosomal and mitochondrial genes, 16 matched to described cDNA, and 20 corresponded to unknown sequences that may represent novel genes. The study of 7 differentially expressed mRNAs known to be involved in developmental process in the embryo suggests roles for apoptosis, oxidative stress, gap junctions, and differentiation in the determination of embryo quality. The aberrant transcription patterns detected in in vitro-produced bovine embryos compared with those produced in vivo may explain their reduced quality in terms of viability after cryopreservation.

¹H and ¹³C NMR spectra of copolymers of methyl methacrylate (M) and butyl acrylate (B) were analyzed in terms of sequence distribution and stereoregularity of monomer units. The copolymers were prepared by atom transfer radical polymerization (ATRP) in bulk and 25% (v/v) benzonitrile solution. The different experimental conditions resulted in different levels of polymerization control. Under our experimental conditions, the experimental data agreed well with data calculated from the Mayo−Lewis terminal model and Bernoullian statistics with monomer reactivity ratios of rM = 1.44 ± 0.11 and rB = 0.56 ± 0.06 and an isotacticity parameter of σM = 0.23 and coisotacticity parameter of σ = 0.30. Therefore, the copolymer compositions and microstructures are independent of the controllability of the ATRP process.

The choline-binding domain (ChoBD) of the carboxy-terminal region of the Streptococcus pneumoniae amidase LYTA (C-LYTA) presents a strong affinity for tertiary amines. We report a method for single-step purification of proteins expressed in the methylotrophic yeast Pichia pastoris based on the fusion of C-LYTA to the protein of interest. We show that C-LYTA can be efficiently expressed and secreted in this host. Tagged proteins fused to this binding domain can be purified on inexpensive DEAE matrices. It therefore provides a useful system for the purification of recombinant proteins with high specificity suitable for industrial purposes. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 74: 164–171, 2001.

Five new withanolides, 5α,6α:22,26:24,25-triepoxy-16α,26-dihydroxy-18(13→17)-abeo-ergosta-2,13-dien-1-one (salpichrolide N, 1), 5α,6α:22,26:24,25-triepoxi-15α,26-dihydroxyergosta-2,16-dien-1-one (salpichrolide L, 2), 5α,6α:22,26-diepoxi-24,25,26-trihydroxy-17(13→18)-abeo-ergosta-2,13,15,17-tetraen-1-one (salpichrolide M, 3a), 5α,6α:22,25:22,26-triepoxy-24-hydroxy-17(13→18)-abeo-ergosta-2,13,15,17-tetraen-1-one (salpichrolide J, 4), and 5α,6α:22,26-diepoxy-22,24,25-trihydroxy-17(13→18)-abeo-ergosta-2,13,15,17-tetraen-1-one (salpichrolide K, 5), were isolated from the leaves of Salpichroa origanifolia and characterized by a combination of spectroscopic (1D and 2D NMR, MS) and chemical methods.

The effect of glucose in the medium used during in vitro culture on the sex ratio of bovine blastocysts derived from in-vitro-matured andin-vitro-fertilized oocytes was evaluated. Oocytes were matured and inseminated with mixed sperm from three bulls and were cultured in vitro in modified synthetic oviducal fluid medium with 10% fetal calf serum, with or without glucose supplementation. The overall rate of cleaved embryos that developed to expanded blastocyst in the medium without glucose (27.0%) was significantly greater (P&lt;0.05) than the percentage observed when embryos were cultured in medium with glucose (17.5%). Analysis of variance was performed to analyse the effect of glucose on the proportion of male embryos reaching the blastocyst stage (or arrested at the morula stage) during Days 7 to 10. Regardless of the presence or absence of glucose in the medium, significantly (P&lt;0.05) more male than female embryos were harvested as expanded blastocysts on Day 7 and on Day 8 of culture. On Days 9 plus 10 of culture, a sex ratio imbalance only occurred in the absence of glucose in the culture medium (P&lt;0.05). Glucose did not produce any significant effect on the sex ratio of the overall number of expanded blastocysts harvested by Day 10 of in vitro culture. However a significantly greater proportion of females (P&lt;0.01) were found among those embryos that developed only to the morulae stage after 10 days in vitro. These results show that glucose supplementation of culture media produces a preferential loss of female embryos during culture to the blastocyst stage.