Eicosapentaenoic acid supplemented to in vitro maturation medium results in lesser lipid content and intracellular reactive oxygen species in blastocysts of cattle.

Sub-optimal cattle embryo development to the blastocyst stage still is a problem when conducting in vitro production (IVP) procedures. Supplementation of in vitro maturation (IVM) medium with omega 3-polyunsaturated eicosapentaenoic acid (EPA) is an approach that might have positive effects on lipid metabolism of cattle oocytes, potentially improving subsequent embryo development. The aim of this study was to evaluate effects of EPA addition to serum-free IVM medium on pronuclear formation after in vitro fertilization, cleavage, and blastocyst rates. Effects of EPA on lipid accumulation and intracellular reactive oxygen species (ROS) generation with IVP of cattle embryos was also investigated. In all experiments, cumulus-oocyte complexes were matured in IVM medium supplemented with 0 nM, 1 nM, or 1 µM EPA for 24 h. Pronuclear formation, cleavage, and blastocyst rates were similar for embryos when there was supplementation of EPA at all concentrations to those of the control group (P > 0.05). The inclusion of 1 nM EPA in medium resulted in a greater lipid content and less intracellular ROS in day 8-embryos compared with those of the Control group (P < 0.05). There were no differences, however, when there was inclusion of 1 µM EPA compared to embryos of the Control group at the day 8 developmental stage (P > 0.05). In conclusion, supplementation with IVM medium with the 1 nM EPA concentration resulted in a lesser blastocyst lipid and intracellular ROS concentration, without modifying embryo development, therefore, EPA could be a desirable supplement to improve embryo quality in cattle.

Funic potassium chloride injection during intact dilation and evacuation.

OBJECTIVE: To describe effectiveness of funic potassium chloride (KCl) injection for fetal demise during intact dilation and evacuation (D&E). STUDY DESIGN: We abstracted medical records of patients who underwent standard or intact D&E to identify those who had intact D&E from 20 weeks 0 days through 23 weeks 6 days from February 2016 to August 2017 at one academic center. We extracted data on time to asystole following KCl injection, as well as incidents including hemorrhage, infection, uterine perforation, and unplanned admissions for up to 6 months following each procedure. RESULTS: Of 57 procedures, 32 (56%) were intact. Median time from KCl injection to fetal asystole was 48 seconds (interquartile range [IQR] 34-100). Median time to asystole at weeks 20 and 21 (46 seconds [IQR 34-60 seconds]) did not differ significantly from weeks 22 and 23 (85 seconds [IQR 34-200 seconds]), p = 0.15. Asystole occurred in less than four minutes in all cases but one. No adverse events occurred with either the funic KCl injections or the intact D&E procedures. CONCLUSION: Funic KCl injection for intact D&E usually causes fetal asystole in less than 4 minutes. IMPLICATIONS: Funic KCl injection under direct visualization is a feasible method to induce fetal asystole at the time of intact dilation and evacuation.

Reproductive hormones influence zinc homeostasis in the bovine cumulus-oocyte complex: Impact on intracellular zinc concentration and transporters gene expression.

Zinc (Zn) is a vital trace element for the body and its bioavailability influences numerous reproductive events. However, the mechanisms that regulate Zn homeostasis in the cumulus-oocyte complex (COC) are yet to be elucidated. The aim of this study was to investigate the role of estradiol 17-beta (E2), FSH and LH in Zn homeostasis regulation in bovine COC matured in vitro and Zn transporters gene expression. For this purpose, intracellular Zn levels in oocytes and cumulus cells (CC) were assessed using a Zn-specific fluorescent indicator. In addition, gene expression and sequencing of six Zn transporters (Slc39a6, Slc39a8, Slc39a14, Slc30a3, Slc30a7 and Slc30a9) were assessed. Our results demonstrated that the simultaneous presence of E2, FSH, and LH during oocyte maturation altered intracellular zinc levels and transporters expression in both oocytes and CC. Transporter's gene expression was different in oocytes and CC, possibly due to cell-specific changes in Zn levels during maturation. The interaction effects of Zn with hormonal treatments influenced the results. This study emphasizes that Slc39a6 is highly sensitive to hormone induction. Overall, the hormonal modulation of Zn homeostasis in the COC was evidenced. Also, a preponderant role of FSH as a modulator of Zn intracellular levels and transporter gene expression is suggested.

Transversus abdominis plane block reduces pain and narcotic consumption after robot-assisted distal pancreatectomy.

BACKGROUND: Minimizing pain and disability are key postoperative objectives of robot-assisted distal pancreatectomy (RADP). This study tested effects of bupivacaine transversus abdominis plane (TAP) block on opioid consumption and pain after RADP. METHODS: Retrospective case-control study (June 2012 -Oct 2017) evaluating bilateral intraoperative bupivacaine TAP block as an interrupted time series. Linear regression evaluated opioid consumption in terms of intravenous (IV) morphine milligram equivalents (MME) and controlled for preoperative morbidity. Secondary outcomes included numerical rating scale (NRS) pain scores. RESULTS: 81 RADP patients met eligibility, 48 before and 33 after implementation of TAP. Baseline characteristics were equivalent with a trend toward higher age, Charlson comorbidity, and ASA score among the TAP cohort. TAP patients consumed on average 4.52 fewer IV MME than controls during the first six postoperative hours (p = 0.032) and reported lower mean NRS scores at six (p = 0.009) and 12 h (p = 0.006) but not at 24 h (p = 0.129). Postoperative morbidity and lengths of stay (LOS) were equivalent (5 vs. 6 days, p = 0.428). CONCLUSION: Bupivacaine TAP block was associated with significant reductions in opioid consumption and pain after RADP but did not shorten hospital LOS consistent with bupivacaine's limited half-life.

PHYTOCHROME-INTERACTING FACTOR 3 mediates light-dependent induction of tocopherol biosynthesis during tomato fruit ripening.

Tocopherols are important antioxidants exclusively produced in plastids that protect the photosynthetic apparatus from oxidative stress. These compounds with vitamin E activity are also essential dietary nutrients for humans. Although the tocopherol biosynthetic pathway has been elucidated, the mechanisms that regulate tocopherol production and accumulation remain elusive. Here, we investigated the regulatory mechanism underlying tocopherol biosynthesis during ripening in tomato fruits, which are an important source of vitamin E. Our results show that ripening under light conditions increases tocopherol fruit content in a phytochrome-dependent manner by the transcriptional regulation of biosynthetic genes. Moreover, we show that light-controlled expression of the GERANYLGERANYL DIPHOSPHATE REDUCTASE (SlGGDR) gene, responsible for the synthesis of the central tocopherol precursor phytyl diphosphate, is mediated by PHYTOCHROME-INTERACTING FACTOR 3 (SlPIF3). In the absence of light, SlPIF3 physically interacts with the promoter of SlGGDR, down-regulating its expression. By contrast, light activation of phytochromes prevents the interaction between SlPIF3 and the SlGGDR promoter, leading to transcriptional derepression and higher availability of the PDP precursor for tocopherol biosynthesis. The unraveled mechanism provides a new strategy to manipulate fruit metabolism towards improving tomato nutritional quality.

The presence of acylated ghrelin during in vitro maturation of bovine oocytes induces cumulus cell DNA damage and apoptosis, and impairs early embryo development.

The aim of this study was to investigate the effects of acylated ghrelin supplementation during in vitro maturation (IVM) of bovine oocytes. IVM medium was supplemented with 20, 40 or 60 pM acylated ghrelin concentrations. Cumulus expansion area and oocyte nuclear maturation were studied as maturation parameters. Cumulus-oocyte complexes (COC) were assessed with the comet, apoptosis and viability assays. The in vitro effects of acylated ghrelin on embryo developmental capacity and embryo quality were also evaluated. Results demonstrated that acylated ghrelin did not affect oocyte nuclear maturation and cumulus expansion area. However, it induced cumulus cell (CC) death, apoptosis and DNA damage. The damage increased as a function of the concentration employed. Additionally, the percentages of blastocyst yield, hatching and embryo quality decreased with all acylated ghrelin concentrations tested. Our study highlights the importance of acylated ghrelin in bovine reproduction, suggesting that this metabolic hormone could function as a signal that prevents the progress to reproductive processes.

Hydrolases of the ILR1-like family of Arabidopsis thaliana modulate auxin response by regulating auxin homeostasis in the endoplasmic reticulum.

Amide-linked conjugates of indole-3-acetic acid (IAA) have been identified in most plant species. They function in storage, inactivation or inhibition of the growth regulator auxin. We investigated how the major known endogenous amide-linked IAA conjugates with auxin-like activity act in auxin signaling and what role ILR1-like proteins play in this process in Arabidopsis. We used a genetically encoded auxin sensor to show that IAA-Leu, IAA-Ala and IAA-Phe act through the TIR1-dependent signaling pathway. Furthermore, by using the sensor as a free IAA reporter, we followed conjugate hydrolysis mediated by ILR1, ILL2 and IAR3 in plant cells and correlated the activity of the hydrolases with a modulation of auxin response. The conjugate preferences that we observed are in agreement with available in vitro data for ILR1. Moreover, we identified IAA-Leu as an additional substrate for IAR3 and showed that ILL2 has a more moderate kinetic performance than observed in vitro. Finally, we proved that IAR3, ILL2 and ILR1 reside in the endoplasmic reticulum, indicating that in this compartment the hydrolases regulate the rates of amido-IAA hydrolysis which results in activation of auxin signaling.