Internal iliac and uterine arteries Doppler ultrasound in the assessment of normotensive and chronic hypertensive pregnant women

The objective of this work was to compare Doppler flows pulsatility index (PI) and resistance indexes (RI) of uterine and internal iliac arteries during pregnancy in low risk women and in those with stage-1 essential hypertension. From January 2010 and December 2012, a longitudinal and prospective study was carried out in 103 singleton uneventful pregnancies (72 low-risk pregnancies and 31 with stage 1 essential hypertension)at the 1(st), 2(nd) and 3(rd) trimesters. Multiple linear regression models, fitted using generalized least squares and whose errors were allowed to be correlated and/or have unequal variances, were employed; a model for the relative differences of both arteries impedance was utilized. In both groups, uterine artery PI and RI exhibited a gestational age related decreasing trend whereas internal iliac artery PI and RI increased. The model testing the hemodynamic adaptation in women with and without hypertension showed similar trend. Irrespective of blood pressure conditions, the internal iliac artery resistance pattern contrasts with the capacitance pattern of its immediate pelvic division, suggesting a pregnancy-related regulatory mechanism in the pelvic circulation

Uterine artery Doppler in the management of early pregnancy loss: a prospective, longitudinal study

The pharmacological management of early pregnancy loss reduced substantially the need for dilation and curettage. However, prognostic markers of successful outcome were not established. Thus the major purpose of this study was to determine the sensitivity and specificity of the uterine artery pulsatility (PI) and resistance (RI) indices to detect early pregnancy loss patients requiring dilation and curettage after unsuccessful management

Dispersively detected Pauli Spin-Blockade in a Silicon Nanowire Field-Effect Transistor

We report the dispersive readout of the spin state of a double quantum dot formed at the corner states of a silicon nanowire field-effect transistor. Two face-to-face top-gate electrodes allow us to independently tune the charge occupation of the quantum dot system down to the few-electron limit. We measure the charge stability of the double quantum dot in DC transport as well as dispersively via in-situ gate-based radio frequency reflectometry, where one top-gate electrode is connected to a resonator. The latter removes the need for external charge sensors in quantum computing architectures and provides a compact way to readout the dispersive shift caused by changes in the quantum capacitance during interdot charge transitions. Here, we observe Pauli spin-blockade in the high-frequency response of the circuit at finite magnetic fields between singlet and triplet states. The blockade is lifted at higher magnetic fields when intra-dot triplet states become the ground state configuration. A lineshape analysis of the dispersive phase shift reveals furthermore an intradot valley-orbit splitting $\Delta_{vo}$ of 145 $\mu$eV. Our results open up the possibility to operate compact CMOS technology as a singlet-triplet qubit and make split-gate silicon nanowire architectures an ideal candidate for the study of spin dynamics

Mini-Percoll processing of domestic ruminant frozen-thawed semen dispenses the use of heparin in capacitating medium.

Sperm capacitation is a prerequisite for mammal successful fertilization. Although usually a capacitating substance such as heparin is used during sheep in vitro fertilization, evidences suggest that the cryopreservation process and Percoll technique could induce spontaneous capacitation. This study aimed to compare ovine, caprine and bovine frozen-thawed sperm after mini-Percoll processing on sperm parameters, receiving or not heparin supplementation. In conclusion, frozen-thawed ovine, caprine and bovine spermatozoa processed with mini-Percoll behave similarly regarding to capacitation status and does not require heparin supplementation during in vitro incubation to achieve capacitation. [Processamento pela técnica de mini-Percoll em sêmen congelado/descongelado de ruminantes domésticos dispensa o uso da heparina em meio capacitante].Edição dos anais do XXII Congresso Brasileiro de Reprodução Animal (CBRA), Santos, SP, Brasil, maio 2017

Mini-percoll technique induces Similar capacitation features in domestic ruminant frozen-thawed spermatozoa regardless of the presence of heparin.

Background: Sperm capacitation is a process consists of a series of functional, biochemical, and biophysical modifications that render the ejaculated sperm competent for oocyte fertilization. Secreted by the female reproductive tract epithelium, heparin promotes capacitation by binding to and removing seminal plasma proteins, which are adsorbed to the sperm PM and would inhibit capacitation. There is substantial evidence that cryopreservation promotes capacitation-like changes in bull, ram and buck sperm. Our general hypotheses were: (a) cryopreserved ram sperm suffer capacitation more quickly than buck and bull sperm under the same conditions; (b) the capacitation status of ruminant cryopreserved sperm is similar whether or not heparin is present after the mini-Percoll technique; and (c) ruminant frozen-thawed sperm selected by mini-Percoll and incubated within media without heparin supplementation is not impaired in terms of capacitation status and sperm agglutination. This study aimed to compare sperm parameters of ovine, caprine, and bovine frozen-thawed sperm after mini-Percoll processing followed by incubation with or without heparin supplementation. Materials, Methods & Results: Commercial semen of all species were used. Sperm samples were selected by mini-Percoll and supplemented (or not) with heparin within an incubation medium for 18 h. Sperm kinematics (CASA system analyzes), capacitation status (CTC staining) and sperm agglutination were evaluated after thawing, mini-Percoll, 1.5 h, 3 h, 6 h and 18 h. In comparison with post-thawing analysis, ovine species demonstrated a reduction (P 0.05). In caprine and bovine species, a lower (P < 0.05) rate of sperm agglutination was observed in the presence of heparin at 18 h of incubation. In the absence of heparin, ovine samples showed a higher (P < 0.05) agglutination rate compared to the bovine species after long incubation period. Discussion: The present study compared sperm parameters (sperm kinematics, agglutination rate and capacitation status) of ruminant frozen-thawed sperm after mini-Percoll selection followed by in vitro incubation with or without heparin supplementation. In this study, it was observed the same rate of capacitated cells after the sperm selection (min-Percoll) between ruminant species. This indicate that the capacitation process occurs similarly between ruminant species, refuting the first hypothesis of this study. The presence of heparin did not influence the capacitation status of ruminant frozen-thawed sperm after mini-Percoll selection, it demonstrates that the second hypothesis was supported by this study making more economic and practical the use of ruminant frozen-thawed semen. The absence of heparin in the incubation medium did not harmed the capacitation status and sperm agglutination of ruminant frozen-thawed sperm. This supported the third hypothesis of the current study and indicate that the use of mini-Percoll technique regardless the presence of heparin could be a useful alternative for the preparation of ruminant frozen-thawed sperm. In conclusion, the capacitation status of ruminant frozen-thawed sperm is similar whether or not heparin is present after the mini-Percoll technique

Mancha-de-xanthomonas: nova doença do cajueiro.

bitstream/CNPAT-2010/11914/1/bd-24.pd

L-carnitine supplementation during vitrification did not improve survival and quality rates, but altered CrAT and PRDX1 expression in in vivo-produced ovine embryos.

Embryo cryodamage is observed mainly at metabolic and molecular aspects and it impairs post warming quality and survival rates. This study aimed to evaluate the effect of L-carnitine (LC) supplementation during either vitrification or post warming solutions on the 6-7th day of in vivo-produced ovine embryos. LC (3.72 mM) was added to vitrification (Experiment 1; C1: control; LC1: supplemented embryos) or warming solutions (Experiment 2; C2; LC2). In vitro culture (IVC) of warmed embryos was performed for 72 h at 38,5 °C, 5% CO2 and 5% O2 to evaluate survival rates in both Experiments. In Experiment 1, reactive oxygen species (ROS) levels were measured by CellROX Green staining, total cell number (TCN) by Hoechst 33342, number of apoptotic cells by caspase-3 immunofluorescence staining protocol, apoptotic index evaluation in both groups. Gene expression analysis of carnitine palmitoyltransferase 1 and 2 (CPT1 and CPT2), carnitine O-acyltransferase (CrAT) and peroxiredoxin 1 (PRDX1), were performed by RT-qPCR (ACTB as endogenous control) in Experiments 1 and 2 and results were compared to fresh embryos (FE). Averages of survival rates were compared by the Chi-Square test. Means of TCN, apoptotic cells, apoptotic index and fluorescence intensity were compared by Student's t-test, at 5% significance level. Survival rates were similar between groups (p> 0.05) in Experiments 1 (68.7%, C1 vs 81.8%, LC1) and 2 (48.5%, C2 vs 64.7%, LC2). In Experiment 1, ROS levels at 24 h of IVC (85.83 ± 68.37 x 1010, C1 vs 89.04 ± 84.48 x 1010, LC1), total cell number at 24 h (89 ± 22, C1 vs 82.2 ± 28, LC1) and 72 h (86 ± 19.9, C1 vs 68.5 ± 25.26, LC1), apoptotic cells (3.75 ± 1.48, C1 vs 4.50 ± 4.72, LC1) and apoptotic index (4.37 ± 1.45, C1 vs 5.23 ± 4.72, LC1) at 72 h of IVC did not differ (p> 0.05) between C1 and LC1. Gene expression analysis showed no differences in CPT1 and CPT2 mRNA relative abundance in embryos of both experiments compared to FE, however, CrAT was downregulated (p< 0.05) in C1 and PRDX1 was downregulated (p< 0.05) in both C1 and LC1, compared to FE. Moreover, CrAT and PRDX1 were upregulated (p< 0.05) in C2 and CrAT was downregulated (p< 0.05) in LC2, in relation to FE. In conclusion, although the short-term LC supplementation at 3.72 mM during cryopreservation did not improve post-warming survival and morphological parameters of the evaluated embryos, it was able to modulate expression of genes related to energy homeostasis (CrAT) and oxidative stress (PRDX1), proving to be beneficial, in both forms of supplementation, to in vivo-produced ovine embryos.".Proceedings of the 31st Annual Meeting of the Brazilian Embryo Technology Society (SBTE); Cabo de Santo Agostinho, PE, Brazil, August 17th to 19th, 2017. Abstracts

Efeito do protocolo de sincronização celular na produção de clones bovinos.

Os métodos de sincronização da célula doadora de núcleo para uso em clonagem baseiam-se no bloqueio reverslvel do ciclo celular em pontos especlficos. Idealmente, espera-se que esse bloqueio não provoque prejulzo à viabilidade do embrião reconstituldo após a transferência de núcleo. Este trabalho teve por objetivo avaliar a eficiência do processo de clonagem (taxa de eletrofusão, produção de blastocistos e estabelecimento de prenhez) a partir de dois protocolos de sincronização celular: privação de soro fetal bovino (SFB) ou cultivo celular até confluência. Oócitos bovinos foram recuperados por aspiração folicular postmortem e maturados in vitro (MIV) por 18h em TCM-199 suplementado com 10% de SFB, 1 ug/mL FSH, 50ug/ mL hCG, 1 ug/mL estradiol, 0,2mM piruvato de sódio e 83,4~g/mL amicacina em atmosfera úmida de 5% de CO2 em ar a 38,5°C. Oócitos apresentando extrusão do primeiro corpúsculo polar foram selecionados, corados com 10ug/mL de Hoechst 33342 e enucleados em fluido sintético de oviduto (SOF) tamponado com HEPES, suplementado com 10% de SFB e 7,5ug/mL de citocalasina B. A reconstituição oocitâria foi realizada com dois grupos de fibroblastos bovinos: Grupo A - cultivados sob privação de SFB por 3 a 5 dias (Dulbecco's Modified Eagle Medium -DMEM suplementado com 0,5% de SFB); ou Grupo B -cultivados até confluência (DMEM suplementado com 10% de SFB). Conjuntos citoplasto+fibroblasto foram eletrofundidos em solução de manitol 0,28M, com dois pulsos de corrente direta de 2,OKv/cm e duração de 20s cada. A ativação foi realizada por exposição à ionomicina (5 minutos, 5~M) seguida de incubação em SOF suplementado com 20mM cloreto de estrôncio e .\ O~g/mL de citocalasina B por 6 horas. Os provâveis zigotos foram co-cultivados com células da granulosa em SOF suplementado com 2,5% de SFB e 0,5% de BSA em atmosfera úmida de 5% de CO2 em ar a 38,5°C por 7 dias. Alguns blastocistos de cada grupo foram transferidos para fêmeas receptoras previamente sincronizadas. As taxas de eletrofusão e produção de blastocistos dos dois grupos foram submetidas à ANOVA, com nlvel de significância de 5%. Houve diferença signiticativaentre os grupos A e B quanto às taxas de eletrofusão: 3,66o/a:!:137,% (549 fundido sem 1631 reconstituidos) contra 40,07% % 11,95% (696 fundidos em 1737 reconstituidos), respectivamente; e quanto ao desenvolvimento até blastocisto: 20,58% % 13,84% (113 blastocistos em 549) contra 34,77o/a:!:1 0,47% (242 blastocistos em 696), respectivamente. Tais diferenças proporcionaram produção média de 5,65 e 12,74 blastocistos por sessão de micromanipulação nos grupos A e B, respectivamente. No grupo A não foram diagnosticadas prenhezes aos 30 dias em 25 receptoras inovuladas; no grupo B foram diagnosticadas 4 gestações a partir de II inovulações (36,4%). Embora a privação de SFB permita o desenvolvimento a termo de clones bovinos (Campbell et al., Nature, 380:64, 1996), existem relatos de maior fragmentação de DNA após utilização da privação (Gibbons et aI., Biol Reprod, 66:895, 2002). Ainda, as menores taxas de eletrofusão, produção de blastocistos e estabelecimento de prenhez observadas no grupo A em comparação ao B, indicam que o uso de fibrob'astos submetidos à privação de SFB não é justiticâvel para o procedimento de clonagem em bovinos

Expressão de DNA metiltransferases em blastocistos bovinos produzidos in vivo e in vitro.

Nos mamiferos, a metilação do DNA é essencial na regulação da expressão gênica e na diferenciação celular. Uma proporção elevada de embriões produzidos por transferência nuclear (TN) é incapaz de estabelecer e sustentar a gestação a termo. Há grande consenso que alterações epigenéticas, primariamente causadas por alterações nos padrões de metilação do DNA, resultam em expressão gênica anormal em embriões e fetos clonados, tomando os indices de sucesso da clonagem frustrantes. Este trabalho objetivou analisar os padrões de expressão das DNA metiltransferases (DNMT) I, 3A e 3B em blastocistos bovinos produzidos in vivo (grupo IV) ou in vilro por FIV (grupo FlV) e transferência nuclear a partir de fibroblastos submetidos à privação de soro fetal bovino (SFB) durante o cultivo (grupo TN-S), ou cultivados até a confluência (grupo TN-C). Uma vez que a linhagem celular doadora de núcleo era proveniente de uma fêmea, os blastocistos dos grupos IV e FIV foram sexados por PCR e somente aqueles do sexo feminino foram utilizados nas etapas posteriores. Após remoção da zona pelucida em PBS pH 2,5, os blastocistos foram individualmente submetidos a um ciclo de amplificação linear do RNA mensageiro utilizando o "Superscript RNA amplification system" (L-l 016, Invitrogen). A transcrição reversa de volume correspondente a I/IOdo RNA de um embrião foi realizada com 6,75JlM de hexâmeros randômicos e transcriptase reversa (Improm-lI Reverse Transcriptase, Promega) seguindo instruções do fabricante. Os ensaios de quantificação relativa dos transcritos dos genes DNMTI, DNMT3A e DNMT3B foram realizados por PCR em tempo real com sistema de detecção TaqMan* (Applied Biosystems); utilizando o gliceraldeido-3-fosfato-desidrogenase (GAPDH) como controle endógeno. Foram utilizados oito embriões por grupo, amplificados em quadruplicata. A eficiência média das amplificações por PCR foi estimada para cada gene utilizando-se uma regressão linear do logaritmo da tluorescência a cada ciclo (Ramakers et aI., Neurosci. Lett., 339:62, 2003); e as razões de expressão calculadas de acordo com metodologia descrita por Livak e Schmittgen (Methods, 25:402, 2001). Para verificar as diferenças significativas foi utilizado o "Pair wise fixed reallocation randomization tes(' (Pfaffi et ai., Nucleic Acids Res., 30:e36. 2002). Todas as razões de expressão foram normalizadas pelo GAPDH e calibradas em função do grupo IV. Não foram detectados transcritos da DNMTI nos blastocistos do grupo TN-S, em contra partida. não se verificaram diferenças estatisticas (P>0,05) entre as razões de expressão dos grupos FIV (0,95) e TN-C (0,77). comparados ao grupo IV. Os grupos de transferência nuclear (TN-C e TN-S) apresentaram razões de expressão numericamente superiores para a DNMT3A (1,89 e 1,99; respectivamente) e para a DNMT3B (1,31 e 2,08; respectivamente) quando comparados aos grupos fertilizados (IV e FIV: 1,00 e 1,50 para DNMT3A; e 1,00 e 1,29 para DNMT3B; respectivamente). no entanto, sem diferenças significativas (P>0,05). A ausência de expressão da DNMTI no grupo TN com fibroblastos submetidos à privação de SFB nos leva a sugerir que embriões clonados a partir deste protocolo sejam menos viáveis do que aqueles oril,lndos de fibroblastos cultivados até confluência. A falta de DNMTl compromete a metilação do DNA a cada ciclo celular e toma os embriões inaptos a manterem as marcações epigcnéticas após cada mitose e sustentar o desenvolvimento fetal

Livestock-Forest integrated system attenuates deleterious heat stress effects in bovine oocytes.

Global warming poses signiî±cant challenges to the î–ertility oî– tropical dairy cattle. One promising approach to mitigate heat stress eî–î–ects on reproductive function and reduce the carbon î–ootprint is the use oî– integrated livestock-î–orest (ILF) systems. The aim oî– this study was to investigate the effects of two different systems, namely Full Sun (FS) and ILF, on maternal hyperthermia and oocyte quality oî– Holstein and Girolando heiî–ers during the tropical summer season. The temperature-humidity index (THI) data revealed intense heat stress during the experiment. Both the system (P<0.01) and the breed (P<0.01) î–actors had a signiî±cant impact on vaginal temperature, being hyperthermia more pronounced in the FS system and in the Holstein breed. Over the î±ve time points collected at a 33-day interval, we observed distinct patterns for ILF (P=0.65) and FS (P<0.001) systems, suggesting an adaptive response in animals kept in FS systems. Furthermore, oocyte quality assessment revealed an eî–î–ect oî– the system î–or oocyte diameter (P<0.001) and levels oî– IGFBP2 (P<0.001), and caspase 3 levels showed a decrease in ILF compared to FS î–or both Holstein (P<0.001) and Girolando (P<0.001) breeds. Collectively, these parameters indicate that oocyte quality during the summer months was superior in animals maintained in the ILF system. In conclusion, the ILF system demonstrated promising results in attenuating maternal hyperthermia and mitigating its eî–î–ects on oocyte quality. Additionally, our observations suggest that animals in the FS system may exhibit an adaptive response to heat stress