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RESEARCH QUESTION: What impact do variations in embryo transfer catheter loading and movement procedures have on temperature and pH fluctuations during embryo transfer? DESIGN: Mock embryo transfers were conducted to test the impact of air flow/movement, use of catheter coverings, and the type of workstation used for catheter loading on catheter temperature. A thermocouple probe inserted into the tip of the outer catheter or taped to the exterior of the inner catheter recorded temperature within the catheter every 5 s from time of mock embryo loading (TL) to 60 s (TLâ¯+â¯60 s) or from the start of transit (TT). Fluctuations in culture medium pH in embryo transfer dishes were monitored. RESULTS: The rate of cooling during transit was faster (all P < 0.05) when catheters were uncovered compared with all covering methods tested. This resulted in a lower catheter temperature at TLâ¯+â¯20 s (28.43 ± 0.30 °C) compared with catheters covered by plastic tubing (31.4 ± 0.30 °C), paper (31.0 ± 0.26 °C) or paperâ¯+â¯thumb (31.1 ± 0.78 °C; all P ≤ 0.05). Temperature was maintained more effectively when catheters were loaded in a crib compared with a heated stage, until initiation of transit, when the rate of temperature decrease was similar. Culture medium pH increased more rapidly when embryo transfer dishes remained on a heated stage during the procedure compared with in an open crib. CONCLUSIONS: Temperature loss during the embryo transfer procedure can be mitigated by reducing the transit time and using catheter coverings. Use of a crib for catheter loading only improved temperature stability while the catheter remained in the crib, not during transit, and reduced pH fluctuations during the procedure.
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Transferência Embrionária , Temperatura , Transferência Embrionária/métodos , Concentração de Íons de Hidrogênio , Humanos , Catéteres , Feminino , Meios de Cultura , Técnicas de Cultura Embrionária/métodosRESUMO
INTRODUCTION: Per-oral pyloromyotomy (POP or G-POEM) provides significant short-term improvements in symptoms and objective emptying for patients with medically refractory gastroparesis, but it is unclear if patients with gastroparesis and co-existing dysmotility (small bowel or colonic delay) also benefit. In this study, we used wireless motility capsule (WMC) data to measure outcomes in patients with isolated gastroparesis (GP) and gastroparesis with co-existing dysmotility (GP + Dys) who underwent POP. METHODS: We retrospectively analyzed patients who had POP and completed WMC data during their evaluation of intestinal dysmotility. WMC data were reviewed to identify patients who demonstrated isolated GP or GP + Dys. Each patient's pre-op and post-op Gastroparesis Cardinal Symptom Index (GCSI) and 4-h solid-phase scintigraphy gastric emptying studies (GES) scores were compared to evaluate improvement. RESULTS: Of the entire cohort (n = 73), 89% were female with a mean age of 47.0 ± 15.0 years old. Gastroparesis etiologies were divided among idiopathic (54.8%), diabetic (26%), postsurgical (8.2%), autoimmune (5.5%), and multifactorial (5.5%). Forty-one patients (56%) had GP and 32 patients (44%) had GP + Dys. GCSI improved after POP whether the patient had isolated GP (- 12.31, p < 0.001) or GP + Dys (- 9.58, p < 0.001); however, there was no significant difference in total GCSI improvement between the two groups. A subset of patients had postoperative GES available (n = 47). In the isolated GP and GP + Dys cohorts, 15/28 (54%) and 12/19 (63%) patients had normal post-op 4-h GES, respectively, but no statistical difference between the two groups. CONCLUSION: Patients with medically refractory gastroparesis with and without concomitant gastrointestinal dysmotility show short-term subjective and objective improvement after POP. Concomitant small bowel or colonic dysmotility should not deter physicians from offering POP in carefully selected patients with gastroparesis.
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Gastroparesia , Piloromiotomia , Adulto , Contraindicações , Feminino , Esvaziamento Gástrico , Gastroparesia/complicações , Gastroparesia/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Piloromiotomia/efeitos adversos , Estudos Retrospectivos , Resultado do TratamentoRESUMO
RESEARCH QUESTION: Does single embryo culture under atmospheric or reduced oxygen alter preimplantation metabolism and post-implantation development compared with culture in groups? DESIGN: Mouse embryos were cultured under 5% or 20% oxygen, individually or in groups of 10. Spent media were analysed after 48, 72 and 96 h of culture. Blastocysts were assessed by outgrowth assay or transferred to pseudo-pregnant recipients, and fetal and placental weight, length and morphology were assessed. RESULTS: Compared with group culture, individually cultured blastocysts had lower net consumption of glucose and aspartate and higher glutamate production. Atmospheric oxygen reduced uptake of glucose and aspartate and increased production of glutamate and ornithine compared with 5% oxygen. Combining 20% oxygen and single culture resulted in further metabolic changes: decreased leucine, methionine and threonine consumption. Under 5% oxygen, individual culture decreased placental labyrinth area but had no other effects on fetal and placental development or outgrowth size compared with group culture. Under 20% oxygen, however, individual culture reduced outgrowth size and fetal and placental weight compared with group-cultured embryos. CONCLUSIONS: Preimplantation metabolism of glucose and amino acids is altered by both oxygen and individual culture, and fetal weight is reduced by individual culture under atmospheric oxygen but not 5% oxygen. This study raises concerns regarding the increasing prevalence of single embryo culture in human IVF and adds to the existing evidence regarding the detrimental effects of atmospheric oxygen during embryo culture. Furthermore, these data demonstrate the cumulative nature of stress during embryo culture and highlight the importance of optimizing each element of the culture system.
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Blastocisto/efeitos dos fármacos , Blastocisto/metabolismo , Técnicas de Cultura Embrionária/métodos , Desenvolvimento Embrionário/efeitos dos fármacos , Oxigênio/farmacologia , Animais , Células Cultivadas , Meios de Cultura/farmacologia , Embrião de Mamíferos , Metabolismo Energético/efeitos dos fármacos , Feminino , Glucose/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , GravidezRESUMO
Single embryo culture is suboptimal compared with group culture, but necessary for embryo monitoring, and culture systems should be improved for single embryos. Pronucleate mouse embryos were used to assess the effect of culture conditions on single embryo development. Single culture either before or after compaction reduced cell numbers (112.2 ± 3.1; 110.2 ± 3.5) compared with group culture throughout (127.0 ± 3.4; P < 0.05). Reduction of media volume from 20 µl to 2 µl increased blastocyst cell numbers in single embryos cultured in 5% oxygen (84.4 ± 3.2 versus 97.8 ± 2.8; P < 0.05), but not in 20% oxygen (55.2 ± 2.9 versus 57.1 ± 2.8). Culture in microwell plates for the EmbryoScope and Primo Vision time-lapse systems changed cleavage timings and increased inner cell mass cell number (24.1 ± 1.0; 23.4 ± 1.2) compared with a 2 µl microdrop (18.4 ± 1.0; P < 0.05). Addition of embryo-conditioned media to single embryos increased hatching rate and blastocyst cell number (91.5 ± 4.7 versus 113.1 ± 4.4; P < 0.01). Single culture before or after compaction is therefore detrimental; oxygen, media volume and microwells influence single embryo development; and embryo-conditioned media may substitute for group culture.
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Blastocisto , Técnicas de Cultura Embrionária/métodos , Oxigênio/metabolismo , Animais , Meios de Cultivo Condicionados , Feminino , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBARESUMO
Embryos are routinely cultured individually, although this can reduce blastocyst development. Culture in atmospheric (20%) oxygen is also common, despite multiple detrimental effects on embryos. Although frequently occurring together, the consequences of this combination are unknown. Mouse embryos were cultured individually or grouped, under physiological (5%) or atmospheric (20%) oxygen. Embryos were assessed by time-lapse and blastocyst cell allocation. Compared with the control group (5% oxygen group culture), 5-cell cleavage (t5) was delayed in 5% oxygen individual culture and 20% oxygen group culture (59.91 ± 0.23, 60.70 ± 0.29, 63.06 ± 0.32 h post-HCG respectively, P < 0.05). Embryos in 20% oxygen individual culture were delayed earlier (3-cell cleavage), and at t5 cleaved later than embryos in other treatments (66.01 ± 0.40 h, P < 0.001), this delay persisting to blastocyst hatching. Compared with controls, hatching rate and cells per blastocyst were reduced in 5% oxygen single culture and 20% oxygen group culture (134.1 ± 3.4, 104.5 ± 3.2, 73.4 ± 2.2 cells, P < 0.001), and were further reduced in 20% oxygen individual culture (57.0 ± 2.8 cells, P < 0.001), as was percentage inner cell mass. These data indicate combining individual culture and 20% oxygen is detrimental to embryo development.
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Blastocisto/fisiologia , Técnicas de Cultura Embrionária , Desenvolvimento Embrionário , Animais , Blastocisto/química , Blastocisto/citologia , Microambiente Celular , Fase de Clivagem do Zigoto/citologia , Fase de Clivagem do Zigoto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Oxigênio/química , Imagem com Lapso de TempoRESUMO
OBJECTIVE: Doctors of Osteopathic Medicine (DOs) are sparsely represented within plastic and reconstructive surgery (PRS) and recent changes including the elimination of step 1 scoring have further disadvantaged DO applicants. The demographics, degrees, and scholarly output of DO PRS trainees were compared to that of Doctors of Medicine (MDs) to identify areas of focus which could be used to increase competitiveness of DO applications. DESIGN: A cross-sectional study was created, including ACGME-accredited PRS program trainees during the 2020 to 2021 academic year. DO and MD trainee demographics and scholarly accomplishments were compared using t-test and chi-squared analysis. SETTING: Web-based publicly available information was collected for subjects. PARTICIPANTS: A total of 1092 PRS MD and DO trainees were identified. DOs made up only 2.7% (nâ¯=â¯30) and MDs made up 97.3% (nâ¯=â¯1062). RESULTS: More DOs trained in independent programs (63.3%) than integrated (36.7%) compared to MDs (88.2% v. 11.8%, p < 0.001) and more DOs trained at lower ranked PRS programs (60.0% of DOs and 18.1% of MDs trained at Q4 programs, p < 0.001). DOs had fewer publications (median, IQR: 1, [0-2]) compared to MDs (3, [1-8]), fewer citations (0, [0-6]) vs. (10, [1-56]) and lower H-index (1, [0-1]) vs. (1, [1-3]). CONCLUSION: DO candidates should consider research years in the field of PRS and optimize clinical experience opportunities to increase the competitiveness of their PRS applications. Special attention should be paid to providing networking and research opportunities to DOs who lack home institutions.
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Internato e Residência , Medicina Osteopática , Cirurgia Plástica , Humanos , Estados Unidos , Medicina Osteopática/educação , Estudos Transversais , DemografiaRESUMO
To assess the value of deep learning in selecting the optimal embryo for in vitro fertilization, a multicenter, randomized, double-blind, noninferiority parallel-group trial was conducted across 14 in vitro fertilization clinics in Australia and Europe. Women under 42 years of age with at least two early-stage blastocysts on day 5 were randomized to either the control arm, using standard morphological assessment, or the study arm, employing a deep learning algorithm, intelligent Data Analysis Score (iDAScore), for embryo selection. The primary endpoint was a clinical pregnancy rate with a noninferiority margin of 5%. The trial included 1,066 patients (533 in the iDAScore group and 533 in the morphology group). The iDAScore group exhibited a clinical pregnancy rate of 46.5% (248 of 533 patients), compared to 48.2% (257 of 533 patients) in the morphology arm (risk difference -1.7%; 95% confidence interval -7.7, 4.3; P = 0.62). This study was not able to demonstrate noninferiority of deep learning for clinical pregnancy rate when compared to standard morphology and a predefined prioritization scheme. Australian New Zealand Clinical Trials Registry (ANZCTR) registration: 379161 .
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PURPOSE: The use of acellular dermal matrix changed the breast reconstruction algorithm facilitating implant coverage and direct to implant technique. This study aims to evaluate the effect of the ADM surgical complications, breast aesthetics, and patient satisfaction. METHODS: In a tertiary hospital, patients that underwent implant-based breast reconstructions during a three-year period had their charts retrospectively reviewed, received post-operative BreastQ, and had their post-operative photos evaluated by a three-member panel using a multi-parameter breast specific scale (scored 1-5). The complication information was analysed per reconstructed breast while the analysis of aesthetic and patient-reported outcomes was done per patient. RESULTS: A total of 501 patients (990 breasts) were evaluated. In the complication analysis group, 20.3% of the breasts had ADM. Overall complications and major complications were more frequent in the ADM group. During the first 30 postoperative days the most frequent complications were: skin necrosis/delayed wound healing and haematoma, after 30 days was infection, and complications after 1 year being less than 1%. On the outcome analysis group, ADM was used in 21.5% patients, 44% had post-operative photos, and 29% answered the BreastQ. Factors associated with higher appearance score were bilateral reconstruction, prophylactic surgery, and nipple presence. ADM demonstrated no difference in satisfaction with breasts. CONCLUSION: In implant-based breast reconstructions ADM has been shown to increase rate of complications. The use of acellular dermal matrix did not influence the overall appearance or the patient-reported outcome. A good aesthetic outcome is positively influenced by bilateral reconstructions with preservation of the nipple.
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Derme Acelular , Implante Mamário , Implantes de Mama , Neoplasias da Mama , Mamoplastia , Humanos , Feminino , Estudos Retrospectivos , Mamoplastia/efeitos adversos , Mamoplastia/métodos , Implante Mamário/efeitos adversos , Implante Mamário/métodos , Estética , Mamilos , Neoplasias da Mama/cirurgia , Complicações Pós-Operatórias/epidemiologiaRESUMO
Female anti-Müllerian hormone (AMH) overexpressing (Thy1.2-AMHTg/0) mice experience fetal resorption (miscarriage) by mid-gestation. This study examined whether the ovary, uterine implantation sites and hypothalamus are potential sites of AMH action, as AMH type-2 receptor (AMHR2) expression is reported in each tissue. Pregnancy in Thy1.2-AMHTg/0 mice was compared to wild-type (WT) mice via histological examination of implantation sites, hormone assays, embryo culture and embryo transfer. Uterine AMH and AMHR2 expression was examined by RT-qPCR and immunohistochemistry. The first signs of fetal resorption in the Thy1.2-AMHTg/0 dams occurred at embryonic day 9.5 (E9.5) with 100% of fetuses resorbing by E13.5. Cultured embryos from Thy1.2-AMHTg/0 dams had largely normal developmental rates but a small proportion experienced a minor developmental delay relative to embryos from WT dams. However, embryos transferred from WT donor females always failed to survive to term when transferred into Thy1.2-AMHTg/0 dams. Amh and Amhr2 mRNA was detected in the gravid uterus but at very low levels relative to expression in the ovaries. Progesterone and estradiol levels were not significantly different between WT and Thy1.2-AMHTg/0 dams during pregnancy but luteinizing hormone (LH) levels were significantly elevated in Thy1.2-AMHTg/0 dams at E9.5 and E13.5 relative to WT dams. Collectively, these experiments suggest that AMH overexpression does not cause fetal resorption through an effect on oocytes or preimplantation embryo development. The Thy1.2-AMHTg/0 fetal resorption phenotype is nearly identical to that of transgenic LH overexpression models, suggesting that neuroendocrine mechanisms may be involved in the cause of the miscarriage.
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Aborto Espontâneo , Hormônio Antimülleriano , Aborto Espontâneo/metabolismo , Animais , Hormônio Antimülleriano/genética , Hormônio Antimülleriano/metabolismo , Transferência Embrionária , Feminino , Reabsorção do Feto/metabolismo , Humanos , Camundongos , Oócitos/metabolismo , GravidezRESUMO
INTRODUCTION: Hantaviruses are a group of globally distributed rodent-associated viruses, some of which are responsible for human morbidity and mortality. Sin Nombre orthohantavirus, a particularly virulent species of hantavirus associated with Peromyscus spp. mice, is actively monitored by the Department of Public Health in California (CDPH). Recently, CDPH documented high (40%) seroprevalence in a potentially novel reservoir species, the cactus mouse (Peromyscus eremicus) in Death Valley National Park. METHODS: This study was performed in the extremely isolated Mojave Desert Amargosa River valley region of southeastern Inyo County, California, 105 km from Death Valley, approximately over the same time interval as the CDPH work in Death Valley (between 2011 and 2016). Similar rodent species were captured as in Death Valley and were tested for select hantaviruses using serology and RT-PCR to assess risk to human health and the conservation of the endemic endangered Amargosa vole. RESULTS: Among 192 rodents tested, including 56 Peromyscus spp., only one seropositive harvest mouse (Reithrodontomys megalotis) was detected. DISCUSSION: These data highlight the heterogeneity in the prevalence of hantavirus infection even among nearby desert communities and suggest that further studies of hantavirus persistence in desert environments are needed to more accurately inform the risks to public health and wildlife conservation.
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Anticorpos Antivirais/sangue , Roedores/sangue , Roedores/virologia , Vírus Sin Nombre/imunologia , Animais , California , Reservatórios de Doenças/virologia , Testes Sorológicos/veterináriaRESUMO
Anti-Müllerian hormone (AMH) is an ovarian regulator that affects folliculogenesis. AMH inhibits the developmental activation of the dormant primordial follicles and the oocyte within. In more mature follicles, AMH reduces granulosa cell sensitivity to follicle-stimulating hormone (FSH). We examined the effects of AMH overexpression on the stages of ovarian folliculogenesis, and the development of embryos, with a transgenic mouse that overexpresses human AMH in central nervous system neurons under the control of the mouse Thy1.2 promoter (Thy1.2-AMHTg mice). These mice are severely sub-fertile, despite relatively normal ovulation rates. The embryos of Thy1.2-AMHTg females exhibited delayed preimplantation development and extensive mid-gestation fetal resorption. Young Thy1.2-AMHTg mouse ovaries exhibited only a slight reduction in the rate of primordial follicle activation but large declines in the number of developing follicles surviving past the primary stage. It was expected that Thy1.2-AMHTg mice would retain more primordial follicles as they aged, but at 5 months, their number was significantly reduced relative to wild-type females. These data indicate that moderate elevations in AMH levels can severely restrict reproductive output and the number of developing follicles in the ovary. This evidence suggests that early antral follicles are a target for AMH signaling, which may regulate early follicle survival.
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Hormônio Antimülleriano/genética , Folículo Ovariano/fisiologia , Animais , Hormônio Antimülleriano/fisiologia , Sobrevivência Celular/genética , Células Cultivadas , Técnicas de Cultura Embrionária , Perda do Embrião/genética , Perda do Embrião/patologia , Embrião de Mamíferos , Feminino , Humanos , Tamanho da Ninhada de Vivíparos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Transgênicos , Ovulação/genética , Ovulação/fisiologia , GravidezRESUMO
OBJECTIVE: In vitro culture of preimplantation embryos is improved by grouping embryos together in a drop of media. Individually cultured embryos are deprived of paracrine factors; with this in mind, we investigated whether the addition of a single embryo-secreted factor, interleukin-6 (IL-6), could improve the development of individually cultured embryos. METHODS: Mouse embryos were cultured individually in 2 µL of G1/G2 media in 5% oxygen and supplemented with a range of doses of recombinant mouse or human IL-6. RESULTS: Mouse IL-6 increased hatching at doses of 0.01 and 10 ng/mL compared to the control (93% and 93% vs. 78%, p<0.05) and increased the total number of cells at a dose of 0.1 ng/mL compared to the control (101.95±3.36 vs. 91.31±3.33, p<0.05). In contrast, the highest dose of 100 ng/mL reduced the total number of cells (79.86±3.29, p<0.05). Supplementation with human IL-6 had a different effect, with no change in hatching or total cell numbers, but an increase in the percentage of inner cell mass per embryo at doses of 0.1, 1, and 100 ng/mL compared to the control (22.9%±1.1%, 23.3%±1.1%, and 23.1%±1.1% vs. 19.5%±1.0%, p<0.05). CONCLUSION: These data show that IL-6 improved mouse embryo development when cultured individually in complex media; however, an excess of IL-6 may be detrimental. Additionally, these data indicate that there is some cross-species benefit of human IL-6 for mouse embryos, but possibly through a different mechanism than for mouse IL-6.
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One of the hypotheses that may help explain the loss of honey bee colonies worldwide is the increasing potential for exposure of honey bees to complex mixtures of pesticides. To better understand this phenomenon, two multi-residue methods based on different extraction and cleanup procedures have been developed, and compared for the determination of 11 relevant pesticides in honey bees, pollen, and wax by gas chromatography-quadrupole mass spectrometry. Sample preparatory methods included solvent extraction followed by gel permeation chromatography (GPC) cleanup and cleanup using a dispersive solid-phase extraction with zirconium-based sorbents (Z-Sep). Matrix effects, method detection limits, recoveries, and reproducibility were evaluated and compared. Method detection limits (MDL) of the pesticides for the GPC method in honey bees, pollen, and wax ranged from 0.65 to 5.92 ng/g dw, 0.56 to 6.61 ng/g dw, and 0.40 to 8.30 ng/g dw, respectively, while MDLs for the Z-Sep method were from 0.33 to 4.47 ng/g dw, 0.42 to 5.37 ng/g dw, and 0.51 to 5.34 ng/g dw, respectively. The mean recoveries in all matrices and at three spiking concentrations ranged from 64.4% to 149.5% and 71.9% to 126.2% for the GPC and Z-Sep methods, with relative standard deviation between 1.5-25.3% and 1.3-15.9%, respectively. The results showed that the Z-Sep method was more suitable for the determination of the target pesticides, especially chlorothalonil, in bee hive samples. The Z-Sep method was then validated using a series of field-collected bee hive samples taken from honey bee colonies in Virginia.
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Abelhas/química , Exposição Ambiental/análise , Poluentes Ambientais/análise , Cromatografia Gasosa-Espectrometria de Massas/métodos , Praguicidas/análise , Agricultura , Animais , Cromatografia em Gel/métodos , Limite de Detecção , Reprodutibilidade dos TestesRESUMO
Large regions of recurrent genomic loss are common in cancers; however, with a few well-characterized exceptions, how they contribute to tumor pathogenesis remains largely obscure. Here we identified primate-restricted imprinting of a gene cluster on chromosome 20 in the region commonly deleted in chronic myeloid malignancies. We showed that a single heterozygous 20q deletion consistently resulted in the complete loss of expression of the imprinted genes L3MBTL1 and SGK2, indicative of a pathogenetic role for loss of the active paternally inherited locus. Concomitant loss of both L3MBTL1 and SGK2 dysregulated erythropoiesis and megakaryopoiesis, 2 lineages commonly affected in chronic myeloid malignancies, with distinct consequences in each lineage. We demonstrated that L3MBTL1 and SGK2 collaborated in the transcriptional regulation of MYC by influencing different aspects of chromatin structure. L3MBTL1 is known to regulate nucleosomal compaction, and we here showed that SGK2 inactivated BRG1, a key ATP-dependent helicase within the SWI/SNF complex that regulates nucleosomal positioning. These results demonstrate a link between an imprinted gene cluster and malignancy, reveal a new pathogenetic mechanism associated with acquired regions of genomic loss, and underline the complex molecular and cellular consequences of "simple" cancer-associated chromosome deletions.
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Deleção Cromossômica , Cromossomos Humanos Par 20 , Regulação da Expressão Gênica , Impressão Genômica , Alelos , Animais , Linhagem da Célula , Proteínas Cromossômicas não Histona/genética , Feminino , Inativação Gênica , Heterozigoto , Humanos , Proteínas Imediatamente Precoces/genética , Macaca , Macropodidae , Masculino , Modelos Genéticos , Família Multigênica , Transtornos Mieloproliferativos/genética , Neoplasias/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Repressoras , Transcrição Gênica , Proteínas Supressoras de TumorRESUMO
A trade-off exists between beneficial n-3 long-chain polyunsaturated acids and toxic persistent halogenated hydrocarbons (PHHs), both of which primarily originate from fish oil commonly used in fish feeds. Alternative lipid sources are being investigated for use in fish feeds to reduce harmful contaminant accumulation, hence, research is needed to evaluate PHHs in fish feeds with various lipid compositions. An analytical method was developed for PHHs including nine organochlorine insecticides (OCPs), 26 polychlorinated biphenyls (PCBs) and seven polybrominated diphenyl ethers (PBDEs) in fish feeds with differing proportions of fish oils and alternative lipid sources by GC-ECD after accelerated solvent extraction, gel permeation chromatography (GPC), and sulfuric acid cleanup. The GPC removed the majority of the neutral lipids and sulfuric acid treatment effectively destroyed the polar lipids. Thus, the combination of the two methods removed approximately 99.7% of the lipids in the extracts. The method detection limits were less than 5 ng/g dry weight (dw) for most PHHs, while recoveries were 75-118%, 67-105%, 69-92%, 63-100% and 94-144% with relative standard deviations of 0.2-39%, 0.3-20%, 0.5-12%, 1.5-18% and 1.5-15% for PHHs in five types of fish feeds made from different lipid sources. Although the source of lipid showed no impact on cleanup efficiency and the developed method worked well for all feeds, fish feeds with 100% fish oil contained background PHHs and more interference than feeds containing alternative lipids.
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Óleos de Peixe/análise , Peixes/metabolismo , Hidrocarbonetos Halogenados/análise , Lipídeos/análise , Animais , Cromatografia Gasosa , Cromatografia em Gel , Éteres Difenil Halogenados/análise , Humanos , Hidrocarbonetos Clorados/análise , Inseticidas/análise , Bifenilos Policlorados/análiseRESUMO
The oxygen concentration used during embryo culture can influence embryo development and quality. Reducing the oxygen concentration in the atmosphere to 2% during post-compaction culture of mouse embryos perturbs embryonic gene expression. This study examined the effect of culturing mouse embryos under different oxygen concentrations on subsequent fetal and placental development. Embryos were cultured from the zygote to morula stage under 7% oxygen, followed by 20, 7 or 2% oxygen to the blastocyst stage. Cultured and in vivo developed blastocysts were transferred into pseudopregnant recipients. Fetal and placental outcomes were analysed at day 18 of pregnancy. Implantation rate was not influenced by embryo culture conditions, but resorption rates were increased in embryos cultured under 2% oxygen, compared with 7% oxygen. Day 18 fetal weights were reduced following culture under 2%, compared with 7 or 20% oxygen, or in vivo development. Placental weight was not influenced by culture conditions. No differences in the proportion of junctional or labyrinthine exchange regions within the placenta or the morphometry of the labyrinthine region were detected. Surface density (surface area/gram labyrinth) of trophoblast available for exchange was reduced in placentas developed from embryos cultured under 2% oxygen, compared with 7% oxygen. Placental gene expression of Slc2a1, Slc2a3, Igf2, Igf2r and H19 was not influenced by oxygen conditions during embryo culture. Thus, exposure to 2% oxygen during post-compaction pre-implantation embryo development has adverse consequences for fetal development in the mouse. Oxygen is a significant component of the embryonic environment and reductions in oxygen availability can influence both embryonic gene expression and subsequent fetal development.
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Desenvolvimento Embrionário/fisiologia , Desenvolvimento Fetal/fisiologia , Peso Fetal/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Oxigênio/metabolismo , Placentação , Envelhecimento/fisiologia , Animais , Camundongos , Técnicas de Cultura de ÓrgãosRESUMO
Gonadotropins are routinely administered to produce multiple oocytes for clinical in vitro fertilization (IVF) treatment, laboratory research, and livestock industries. Studies in mice have shown gonadotropin stimulation using equine chorionic gonadotropin (eCG) affects the endometrium, implantation, and fetal development. Evidence from clinical studies also indicates that stimulation with recombinant human follicle-stimulating hormone (rhFSH) may be detrimental to the endometrium and implantation rates. We investigated the effect of rhFSH in mice on maternal plasma hormone concentrations and uterine gene and protein expression and the effect of a stimulated maternal environment on pregnancy. Adult females were stimulated with rhFSH or eCG, followed by human chorionic gonadotropin (hCG). On day 4 of pseudopregnancy, mice either had embryos transferred to the uterus or were killed, and blood and uterine samples were collected. Pregnancy outcomes were examined on day 15. Gonadotropin stimulation increased plasma progesterone concentrations on day 4 compared with controls, whereas estradiol concentrations were unaffected. Stimulation also reduced uterine leukemia inhibitory factor (Lif) mRNA, but the expression of estrogen and progesterone receptors (Esr1 and Pgr), homeobox gene Hoxa10, and Vegf mRNA were unchanged. Furthermore, distribution of uterine PGR protein expression was altered by stimulation, but LIF protein was unchanged. Stimulated embryo transfer recipients had lower pregnancy rates than controls, and fetuses from the rhFSH group had reduced weight, length, and maturity. These results demonstrate that gonadotropin stimulation with rhFSH or eCG alters the preimplantation maternal environment, which results in reduced pregnancy rates and fetal development in the mouse.