Dose-response analysis of microvasculature changes in the murine fetal brain and the maternal extremities due to prenatal ethanol exposure.

SIGNIFICANCE: Prenatal exposure to ethanol causes several morphological and neurobehavioral deficits. While there are some studies on the effects of ethanol exposure on blood flow, research focusing on acute changes in the microvasculature is limited. AIM: The first aim of this study was to assess the dose-dependent changes in murine fetal brain microvasculature of developing fetuses in response to maternal alcohol consumption. The second aim was to quantify changes in vasculature occurring concurrently in the mother's hindlimb and the fetus's brain after maternal exposure to alcohol. APPROACH: Correlation mapping optical coherence angiography was used to evaluate the effects of prenatal exposure to different doses of ethanol (3, 1.5, and 0.75 g / kg) on murine fetal brain vasculature in utero. Additionally, simultaneous imaging of maternal peripheral vessels and the fetal brain vasculature was performed to assess changes of the vasculature occurring concurrently in response to ethanol consumption. RESULTS: The fetal brain vessel diameters (VDs) decreased by ∼47 % , 30%, and 14% in response to ethanol doses of 3, 1.5, and 0.75 g / kg, respectively. However, the mother's hindlimb VD increased by 63% in response to ethanol at a dose of 3 g / kg. CONCLUSIONS: Results showed a dose-dependent reduction in vascular blood flow in fetal brain vessels when the mother was exposed to ethanol, whereas vessels in the maternal hindlimb exhibited concurrent vasodilation.

Assessing the acute effects of prenatal synthetic cannabinoid exposure on murine fetal brain vasculature using optical coherence tomography.

Marijuana is one of the most commonly abused substances during pregnancy. Synthetic cannabinoids (SCBs) are a group of heterogeneous compounds that are 40- to 600-fold more potent than Δ9 -tetrahydrocannabinol, the major psychoactive component of marijuana. With SCBs being legally available for purchase and the prevalence of unplanned pregnancies, the possibility of prenatal exposure to SCBs is high. However, the effects of prenatal SCB exposure on embryonic brain development are not well understood. In this study, we use complex correlation mapping optical coherence angiography to evaluate changes in murine fetal brain vasculature in utero, minutes after maternal exposure to an SCB, CP-55940. Results showed a significant decrease (P < 0.05) in fetal brain vessel diameter, length fraction and area density when compared to the sham group. This preliminary study shows that acute prenatal exposure to an SCB resulted in significant fetal brain vasoconstriction during the peak period for brain development.

The effect of light exposure on the cleavage rate and implantation capacity of preimplantation murine embryos.

During assisted reproduction the embryos are subjected to light. We investigated the relationship between light exposure and the developmental- and implantation capacity of mouse embryos. In vitro cultured embryos were exposed to white or red filtered light, then transferred to the uteri of pseudo-pregnant females. The mice were sacrificed on day 8.5 and implantation sites were counted. The number of nucleic acid containing (PI+) extracellular vesicles (EVs) in culture media of light-exposed and control embryos, as well as, the effect of the EVs on IL-10 production of CD8+ spleen cells was determined by flow cytometry. DNA fragmentation in control and light exposed embryos was detected in a TUNEL assay. The effect of light on the expression of apoptosis-related molecules was assessed in an apoptosis array. Light exposure significantly reduced the implantation capacity of the embryos. The harmful effect was related to the wavelength, rather than to the brightness of the light. Culture media of light exposed groups contained significantly higher number of PI + EVs than those of the control embryos, and failed to induce IL-10 production of spleen cells. The number of nuclei with fragmented DNA, was significantly higher in embryos treated with white light, than in the other two groups. In conclusion exposure to white light impairs the implantation potential of in vitro cultured mouse embryos. These effects are partly corrected by using a red filter. Since there is no information on the light sensitivity of human embryos, embryo manipulation during IVF and ICSI should be performed with caution.

Minimum volume Spatula MVD vitrification method improves embryo survival compared to traditional slow freezing, both for in vivo and in vitro produced mice embryos.

This study was conducted to compare the effect of minimum volume Spatula MVD vitrification (VIT) versus traditional slow freezing (SLF) of mouse embryos. A total of 2,617 8-cell in vivo derived and 2-cell in vitro produced B6D2 mouse embryos were subjected to freezing/thawing or vitrification/warming, while fresh embryos were used as control group. Embryo recovery, survival and development rate, pregnancy rate and offspring production were analyzed. In Experiment 1, 8-cell in vivo derived embryos were subjected to in vitro culture, resulting in greater survival and development rates at 3.5 days post coitum stage in VIT than in SLF group (P < 0.05). Although both methods reached an acceptable hatching rate (41.0% and 49.7% for VIT and SLF, respectively; P=NS), it was significantly lower respect to the control group (67.8%, P < 0.01). In Experiment 2, 2-cell in vitro produced mouse embryos showed a similar recovery rate from the device after freezing/thawing or vitrification/warming (∼84%), however survival rate was significantly higher for vitrified/warmed (94.7%) than frozen/thawed embryos (85.1%; P < 0.01). Vitrified/warmed and control fresh embryos were transferred to surrogate mothers, revealing no differences both in pregnancy and offspring production rates. Our data demonstrate that minimum volume Spatula MVD method is a simple home-made useful technique for vitrification of 2-cell and 8-cell mouse embryos produced either in vitro or in vivo.

Avaliação do extrato etanólico de casca de Punica granatum (romã) na diminuição da replicação viral do BoHV-1 Colorado em embriões murinos experimentalmente infectados / Evaluation of ethanol extract of Punica granatum (Pomegranate) peel decrease in viral replication of BoHV-1 in Colorado murine embryos experimentally infected

O objetivo do trabalho foi avaliar a diminuição da replicação viral (BoHV-1 Colorado) em embriões murinos após tratamento do extrato etanólico da casca de Punica granatum (EEPg). Camundongos fêmeas Swiss com idade entre 6 e 8 semanas foram superovuladas com 0,2 mL a 5 UI de hormônios (eCG e hCG), e acasaladas com machos da mesma idade. Após 18 horas, as fêmeas sofreram eutanásia em câmara de CO2 e, através de abertura no peritônio, os zigotos foram coletados e lavados com solução de pronase 0,5%.Os zigotos foram divididos em quatro grupos: G1 (controle), G2 (expostos aos vírus BoHV-1 Colorado a 108 TCID50/mL), G3 (expostos ao EEPg) e G4 (expostos aos vírus e ao EEPg). Os grupos foram mantidos a 37,5ºC em meio TCM199 (100µL) com 10% de soro fetal bovino em estufa a 5% de CO2 e 95% de umidade. Após 24 h, analisamos a taxa de clivagem (teste exato de Fisher; p<0,05), a morfologia (por microscopia óptica), a nested-PCR e a titulação dos embriões em cocultura com células MDBK após mais 72 h do tratamento (teste de Mann-Whitney; p<0,05) e microscopia eletrônica de transmissão (ME). Os embriões murinos tratados com EEPg apresentaram resultados satisfatórios: sem alterações morfológicas, taxa de clivagem semelhante ao controle e, apesar da detecção da presença do vírus pela nested-PCR e ME, houve diminuição do título viral após tratamentos com esse extrato, o que sugere interferência desse tratamento no ciclo viral do BoHV-1 Colorado sem alterar o desenvolvimento dos embriões.(AU)

The aim of this study was to evaluate the reduction of viral replication (Colorado BoHV-1) in murine embryos after the treatment of ethanol extract of Punica granatum peel (PgEE). Swiss female mice aged 6 to 8 weeks were superovulated with 0.2 mL of the 5 UI hormones (eCG and hCG) and mated with males of the same age. After 18 hours, the females were euthanized in a CO2 chamber, and through the opening in the peritoneum, zygotes were collected and washed with 0.5% pronase solution. The zygotes were divided into four groups: G1 (control), G2 (exposed to the virus Colorado BoHV -1 to 108 TCID50/mL), G3 (exposed to PgEE) and G4 (exposed to the virus and to PgEE). The groups were maintained at 37.5ºC in TCM199 (100 mL) with 10% fetal bovine serum in an incubator at 5% CO2 and 95% humidity. After 24 h, we analyzed the cleavage rate (Fisher's exact test; p<0.05), the morphology (by light microscopy), the nested-PCR and the titration of embryos in co-culture with MDBK cells after over 72 h of treatment (Mann-Whitney test; p<0.05) and transmission electron microscopy (TEM). The murine embryos treated with PgEE showed satisfactory results: no morphological changes, cleavage rate similar to controls, despite the detection of the presence of virus by nested PCR and TEM, there was a decrease of the viral titer after the treatment with this extract, which suggests interference of this treatment in the viral cycle BoHV-1 Colorado without altering the embryo development.(AU)