scholarly journals Porcine embryo development and fragmentation and their relation to apoptotic markers: a cinematographic and confocal laser scanning microscopic study

Reproduction ◽  
2005 ◽  
Vol 129 (4) ◽  
pp. 443-452 ◽  
Author(s):  
Bart Mateusen ◽  
Ann Van Soom ◽  
Dominiek G D Maes ◽  
Isabelle Donnay ◽  
Luc Duchateau ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Embryo Morphology ◽  
Fragmentation Pattern ◽  
Cell Stage ◽  
Developmental Potential ◽  
Developmental Arrest ◽  
Porcine Embryo

Porcine embryo selection prior to transfer is mainly influenced by morphological criteria. However, the relationship between embryonic morphology, developmental potential and cell death by apoptosis in porcine embryos is still unclear. The aim of this study was to establish embryo quality parameters for in vivo fertilised porcine embryos based on timing of development in vitro, embryo morphology and the presence of apoptosis. The kinetics of development and morphological parameters were investigated in a time-lapse cinematographic experiment. Possible links between embryo morphology and apoptosis were examined via a confocal laser scanning experiment, analysing nuclear changes, annexin V and terminal dUTP nick-end labelling. The timing of early cleavages was firmly linked to embryo developmental competence in vitro. Attainment of at least the 5-cell stage before 77 h post insemination and attainment of the morula stage before 102 h post insemination significantly increased the odds for reaching the early blastocyst stage. Overall, a negative effect of fragmentation percentage and fragmentation pattern on subsequent embryonic development was observed, but the developmental potential of embryos experiencing slight fragmentation (0–5%) was not different from embryos without fragmentation. Correlations detected between developmental arrest and fragmentation, and fragmentation and apoptosis were 0.60 and 0.87 (P < 0.05) respectively. Only a minority of the embryos arrested between the 1- and 4-cell stage displayed biochemical characteristics of apoptosis. Consequently, a significant correlation (0.57) between developmental arrest and apoptosis could only be established for embryos arrested after embryonic genome activation.

Consequences of topoisomerase II inhibition in early embryogenesis of Drosophila revealed by in vivo confocal laser scanning microscopy

1993 ◽  
Vol 104 (4) ◽  
pp. 1175-1185 ◽  
Author(s):  
P. Buchenau ◽  
H. Saumweber ◽  
D.J. Arndt-Jovin
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Topoisomerase Ii ◽  
Metaphase Plate ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser

The regulation of DNA topology by topoisomerase II from Drosophila melanogaster has been studied extensively by biochemical methods but little is known about its roles in vivo. We have performed experiments on the inhibition of topoisomerase II in living Drosophila blastoderm embryos. We show that the enzymatic activity can be specifically disrupted by microinjection of antitopoisomerase II antibodies as well as the epipodophyllotoxin VM26, a known inhibitor of topoisomerase II in vitro. By labeling the chromatin of live embryos with tetramethylrhodamine-coupled histones, the effects of inhibition on nuclear morphology and behaviour was followed in vivo using confocal laser scanning microscopy. Both the antibodies and the drug prevented or hindered the segregation of chromatin daughter sets at the anaphase stage of mitosis. In addition, high concentrations of inhibitor interfered with the condensation of chromatin and its proper arrangement into the metaphase plate. The observed effects yielded non-functional nuclei, which were drawn into the inner yolk mass of the embryo. Concurrently, undamaged nuclei surrounding the affected region underwent compensatory division, leading to the restoration of the nuclear population, and thereby demonstrating the regulative capacity of Drosophila blastoderm embryos.

scholarly journals Percentage of Gutta-Percha-, Sealer-, and Void-Filled Areas in Oval-Shaped Root Canals Obturated with Different Filling Techniques: A Confocal Laser Scanning Microscopy Study

2020 ◽  
Vol 14 (01) ◽  
pp. 008-012
Author(s):  
Vinicio Hidemitsu Goto Hirai ◽  
Ricardo Machado ◽  
Maria Carolina Lucato Budziak ◽  
Lucila Piasecki ◽  
Alexandre Kowalczuck ◽  
...  
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Continuous Wave ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser ◽  
Root Canals ◽  
Gutta Percha

Abstract Objective This study compared different obturation techniques, analyzing percentage of areas filled with gutta-percha, sealer, and voids (PGFA, PSFA, and PVFA, respectively) in oval-shaped root canals. Materials and Methods A total of 60 extracted human mandibular central incisors were decoronated, instrumented, and irrigated using the same protocol. After drying, the root canal was filled with AH Plus labeled with 0.1% rhodamine B dye using a Lentulo spiral. The filling procedure was performed by dividing the teeth into four groups according to the respective technique: G1, cold lateral condensation; G2, continuous wave of condensation; G3, modified cold lateral condensation using an F3 master cone; and G4, modified continuous wave of condensation using an ISO (International Organization for Standardization) sized 30 gutta-percha cone. Then, slices measuring 1.5 mm in thickness were obtained 3 and 6 mm from the apex and evaluated by confocal laser scanning microscopy to determine PGFA, PSFA, and PVFA. Statistical Analysis The data were analyzed statistically with analysis of variance and Games-Howell’s tests (p = 0.05). Results The groups showed no significant differences in the apical third (3 mm from the apex). In the middle third (6 mm from the apex), G3 and G1 showed higher PGFA and PVFA, respectively. G3 showed lower PSFA than G2 and G4. Both cold techniques (G1 and G3) promoted lower PSFA than both warm techniques (G2 and G4). Conclusions Notwithstanding the limitations of this in vitro study, PGFA, PSFA, and PVFA ranged significantly only in the middle third, as observed by the different filling techniques. Higher PGFA and PVFA values were obtained for G3 and G1, respectively. Both cold techniques promoted lower PSFA than both warm techniques.

scholarly journals Increased mitochondrial distribution in early-cleaving embryos indicate successful pre-implantation development

2018 ◽  
Vol 14 (4) ◽  
pp. 512-514
Author(s):  
Nor Shahida Abdul Rahman ◽  
Mimi Sophia Sarbandi ◽  
Wan Hafizah Wan Jusof ◽  
Zolkapli Eshak ◽  
Salina Othman ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Close Association ◽  
Mitochondrial Fission ◽  
Polar Body ◽  
Blastocyst Stage ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Cell Stage ◽  
Second Polar Body

The timing of the first zygotic cleavage is an accurate predictor of embryo quality. Embryos that cleaved early have higher developmental viability compared to their late counterparts. During embryonic development, cleavage is affected by cellular metabolic processes performed by mitochondria and its synergistic interaction with endoplasmic reticulum (ER). However, in depth study on differences of mitochondria and ER ultrastructures in early- cleaving (EC) versus late- cleaving (LC) embryos is limited. This study compares mitochondria and ER ultrastructures of EC versus LC embryos using Confocal Laser Scanning Microscopy (CLSM) and Transmission Electron Microscopy (TEM). Embryos were obtained from female ICR superovulated mice, 28-30 hours post hCG. Two-cell embryos were categorized as early-cleaving (EC), while zygotes with the second polar body and two pronuclei present were categorized as late-cleaving (LC). The LC embryos were cultured in vitro until the 2- cell stage. In EC embryos, mitochondria were mostly found at the perinuclear region and closely associated with dense ER. Meanwhile, mitochondria of LC embryos were distributed uniformly within the cytoplasm. Mitochondrial fluorescence intensity was significantly higher in EC versus LC [(18.7 ± 0.4) versus (14.6 ± 0.4)] x 105 pixel, (p<0.01). Development to the blastocyst stage was also significantly higher in EC compared to LC embryos (96.7% versus 60.9%) (p<0.01). Higher viability of EC embryos is attributed to the close association of their mitochondria to ER. This contributed to better mitochondrial fission, resulting in enhanced energy generating processes and preimplantation development. 

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