scholarly journals A Review of Automated Microinjection of Zebrafish Embryos

Micromachines ◽  
2018 ◽  
Vol 10 (1) ◽  
pp. 7 ◽  
Author(s):  
Yuliang Zhao ◽  
Hui Sun ◽  
Xiaopeng Sha ◽  
Lijia Gu ◽  
Zhikun Zhan ◽  
...  
Keyword(s):  
Visual Servoing ◽  
Zebrafish Embryo ◽  
Polar Body ◽  
Cell Transfection ◽  
Zebrafish Embryos ◽  
High Cell ◽  
The Third ◽  
Cell Position ◽  
First Polar Body ◽  
Small Cell Size

Cell microinjection is a technique of precise delivery of substances into cells and is widely used for studying cell transfection, signaling pathways, and organelle functions. Microinjection of the embryos of zebrafish, the third most important animal model, has become a very useful technique in bioscience. However, factors such as the small cell size, high cell deformation tendency, and transparent zebrafish embryo membrane make the microinjection process difficult. Furthermore, this process has strict, specific requirements, such as chorion softening, avoiding contacting the first polar body, and high-precision detection. Therefore, highly accurate control and detection platforms are critical for achieving the automated microinjection of zebrafish embryos. This article reviews the latest technologies and methods used in the automated microinjection of zebrafish embryos and provides a detailed description of the current developments and applications of robotic microinjection systems. The review covers key areas related to automated embryo injection, including cell searching and location, cell position and posture adjustment, microscopic visual servoing control, sensors, actuators, puncturing mechanisms, and microinjection.

scholarly journals Bushen Tiaojing (II and III) Decoctions Activate MAPK14 and MAPK3/1 to Promote the Expression of Cumulus Expansion-Related Factors in Mice

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Xiao Liang ◽  
Xue Tong ◽  
Hui-lan Du ◽  
Ming He ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Western Blot ◽  
Polar Body ◽  
Cumulus Cell ◽  
Serum Levels ◽  
Control Group ◽  
Distilled Water ◽  
Related Factors ◽  
Cumulus Expansion ◽  
Sequential Administration ◽  
First Polar Body

Background. Bushen Tiaojing Decoctions (BSTJ-II-D and BSTJ-III-D) are used to assist pregnancy in clinical practice. In this study, we explored the ability of sequential administration of BSTJ-II-D and BSTJ-III-D to promote cumulus cell (CC) expansion and its underlying mechanisms in controlled ovarian hyperstimulation (COH) mice. Methods. Kunming mice were randomly divided into three groups. The normal group was injected intraperitoneally with saline, and distilled water was administered orally by gavage. As the COH model, mice were injected with GnRHa, eCG, and hCG. Subsequently, the BSTJD group received BSTJ-II-D and BSTJ-III-D orally by gavage, while the control group received distilled water. We evaluated CC expansion and oocyte first polar body (PB1) extrusion under a stereomicroscope. Serum levels of follicle-stimulating hormone (FSH) were detected by radioimmunoassay. The expression of the CC expansion-related factors PTX3 and PTGS2 was detected by immunofluorescence, western blot, and quantitative real-time-polymerase chain reaction analyses (qRT-PCR). Expression of p-MAPK14, p-MAPK3/1, MAPK14, and MAPK3/1 was detected by western blot analysis. Results. Sequential administration of BSTJ-II-D and BSTJ-III-D promoted cumulus expansion and oocyte PB1 extrusion and upregulated PTX3 and PTGS2 expression at the mRNA and protein levels. Furthermore, the levels of p-MAPK14/MAPK14, p-MAPK3/1/MAPK3/1 proteins, and serum FSH in the BSTJD group were higher than those in the normal and control groups. Conclusions. Sequential administration of BSTJ-II-D and BSTJ-III-D promotes cumulus expansion and oocyte maturation in COH mice by increasing FSH expression and activating the MAPK14 and MAPK3/1 signalling pathways, thereby increasing expression of PTX3 and PTGS2.

Triphenyltin chloride induces spindle microtubule depolymerisation and inhibits meiotic maturation in mouse oocytes

2014 ◽  
Vol 26 (8) ◽  
pp. 1084 ◽  
Author(s):  
Yu-Ting Shen ◽  
Yue-Qiang Song ◽  
Xiao-Qin He ◽  
Fei Zhang ◽  
Xin Huang ◽  
...  
Keyword(s):  
Polar Body ◽  
Meiotic Maturation ◽  
Dependent Manner ◽  
Germinal Vesicle Breakdown ◽  
Mouse Oocytes ◽  
First Polar Body ◽  
Triphenyltin Chloride ◽  
Dose Dependent

Meiosis produces haploid gametes for sexual reproduction. Triphenyltin chloride (TPTCL) is a highly bioaccumulated and toxic environmental oestrogen; however, its effect on oocyte meiosis remains unknown. We examined the effect of TPTCL on mouse oocyte meiotic maturation in vitro and in vivo. In vitro, TPTCL inhibited germinal vesicle breakdown (GVBD) and first polar body extrusion (PBE) in a dose-dependent manner. The spindle microtubules completely disassembled and the chromosomes condensed after oocytes were exposed to 5 or 10 μg mL–1 TPTCL. γ-Tubulin protein was abnormally localised near chromosomes rather than on the spindle poles. In vivo, mice received TPTCL by oral gavage for 10 days. The general condition of the mice deteriorated and the ovary coefficient was reduced (P < 0.05). The number of secondary and mature ovarian follicles was significantly reduced by 10 mg kg–1 TPTCL (P < 0.05). GVBD decreased in a non-significant, dose-dependent manner (P > 0.05). PBE was inhibited with 10 mg kg–1 TPTCL (P < 0.05). The spindles of in vitro and in vivo metaphase II oocytes were disassembled with 10 mg kg–1 TPTCL. These results suggest that TPTCL seriously affects meiotic maturation by disturbing cell-cycle progression, disturbing the microtubule cytoskeleton and inhibiting follicle development in mouse oocytes.

H1foo is essential for in vitro meiotic maturation of bovine oocytes

Zygote ◽  
2014 ◽  
Vol 23 (3) ◽  
pp. 416-425 ◽  
Author(s):  
Yan Yun ◽  
Peng An ◽  
Jing Ning ◽  
Gui-Ming Zhao ◽  
Wen-Lin Yang ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Polar Body ◽  
Linker Histone ◽  
Meiotic Maturation ◽  
Control Group ◽  
Bovine Oocyte ◽  
First Polar Body ◽  
Effective Sirnas ◽  
Bovine Oocytes

SummaryOocyte-specific linker histone, H1foo, is localized on the oocyte chromosomes during the process of meiotic maturation, and is essential for mouse oocyte maturation. Bovine H1foo has been identified, and its expression profile throughout oocyte maturation and early embryo development has been established. However, it has not been confirmed if H1foo is indispensable during bovine oocyte maturation. Effective siRNAs against H1foo were screened in HeLa cells, and then siRNA was microinjected into bovine oocytes to down-regulate H1foo expression. H1foo overexpression was achieved via mRNA injection. Reverse transcription polymerase chain reaction (RT-PCR) results indicated that H1foo was up-regulated by 200% and down-regulated by 70%. Based on the first polar body extrusion (PB1E) rate, H1foo overexpression apparently promoted meiotic progression. The knockdown of H1foo significantly impaired bovine oocyte maturation compared with H1foo overexpression and control groups (H1foo overexpression = 88.7%, H1foo siRNA = 41.2%, control = 71.2%; P < 0.05). This decrease can be rescued by co-injection of a modified H1foo mRNA that has escaped from the siRNA target. However, the H1e (somatic linker histone) overexpression had no effect on PB1E rate when compared with the control group. Therefore we concluded that H1foo is essential for bovine oocyte maturation and its overexpression stimulates the process.

Maturation promoting factor in ascidian oocytes is regulated by different intracellular signals at meiosis I and II

Development ◽  
1996 ◽  
Vol 122 (7) ◽  
pp. 1995-2003 ◽  
Author(s):  
G.L. Russo ◽  
K. Kyozuka ◽  
L. Antonazzo ◽  
E. Tosti ◽  
B. Dale
Keyword(s):  
Kinase Activity ◽  
Phase 1 ◽  
Polar Body ◽  
Calcium Transients ◽  
Phase 2 ◽  
Phase 3 ◽  
First Polar Body ◽  
Second Polar Body ◽  
Polar Body Extrusion ◽  
Maturation Promoting Factor

Using the fluorescent dye Calcium Green-dextran, we measured intracellular Ca2+ in oocytes of the ascidian Ciona intestinalis at fertilization and during progression through meiosis. The relative fluorescence intensity increased shortly after insemination in a single transient, the activation peak, and this was followed by several smaller oscillations that lasted for approximately 5 minutes (phase 1). The first polar body was extruded after the completion of the phase 1 transients, about 9 minutes after insemination, and then the intracellular calcium level remained at baseline for a period of 5 minutes (phase 2). At 14 minutes postinsemination a second series of oscillations was initiated that lasted 11 minutes (phase 3) and terminated at the time of second polar body extrusion. Phases 1 and 3 were inhibited by preloading oocytes with 5 mM heparin. Simultaneous measurements of membrane currents, in the whole-cell clamp configuration, showed that the 1–2 nA inward fertilization current correlated temporally with the activation peak, while a series of smaller oscillations of 0.1-0.3 nA amplitude were generated at the time of the phase 3 oscillations. Biochemical characterization of Maturation Promoting Factor (MPF) in ascidian oocytes led to the identification of a Cdc2-like kinase activity. Using p13suc1-sepharose as a reagent to precipitate the MPF complex, a 67 kDa (67 × 10(3) Mr) protein was identified as cyclin B. Histone H1 kinase activity was high at metaphase I and decreased within 5 minutes of insemination reaching a minimum level during phase 2, corresponding to telophase I. During phase 3, H1 kinase activity increased and then decayed again during telophase II. Oocytes preloaded with BAPTA and subsequently inseminated did not generate any calcium transients, nonetheless H1 kinase activity decreased 5 minutes after insemination, as in the controls, and remained low for at least 30 minutes. Injection of BAPTA during phase 2 suppressed the phase 3 calcium transients, and inhibited both the increase in H1 kinase activity normally encountered at metaphase II and second polar body extrusion.

scholarly journals The Relationship between Dissolution Behavior and the Toxicity of Silver Nanoparticles on Zebrafish Embryos in Different Ionic Environments

Nanomaterials ◽  
2018 ◽  
Vol 8 (9) ◽  
pp. 652 ◽  
Author(s):  
Wang Lee ◽  
Eungwang Kim ◽  
Hyun-Ju Cho ◽  
Taejoon Kang ◽  
Bongsoo Kim ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Polyethylene Glycol ◽  
Zebrafish Embryo ◽  
Antibacterial Properties ◽  
Dissolution Behavior ◽  
Aquatic Environments ◽  
Zebrafish Embryos ◽  
Negative Effects ◽  
The Relationship ◽  
Water Filters

A silver nanoparticle is one of the representative engineered nanomaterials with excellent optical, electrical, antibacterial properties. Silver nanoparticles are being increasingly used for medical products, water filters, and cosmetics, etc. However, silver nanoparticles are known to cause adverse effects on the ecosystem and human health. To utilize silver nanoparticles with minimized negative effects, it is important to understand the behavior of silver nanoparticles released to the environment. In this study, we compared toxicity behaviors of citrate-stabilized silver nanoparticles with polyethylene glycol coated silver nanoparticles in two different ionic environments, which are aquatic environments for developing zebrafish embryo. Depending on the composition of the ionic environment, citrate-stabilized silver nanoparticles and polyethylene glycol coated silver nanoparticles exhibited different behaviors in dissolution, aggregation, or precipitation, which governed the toxicity of silver nanoparticles on zebrafish embryos.

Effects of oocyte age, cumulus cells and injection methods on in vitro development of intracytoplasmic sperm injection rabbit embryos

Zygote ◽  
2004 ◽  
Vol 12 (1) ◽  
pp. 75-80 ◽  
Author(s):  
Yue-Liang Zheng ◽  
Man-Xi Jiang ◽  
Yan-Ling Zhang ◽  
Qing-Yuan Sun ◽  
Da-Yuan Chen
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Cumulus Cells ◽  
Fertilization Rate ◽  
First Polar Body ◽  
Injection Methods ◽  
Blastocyst Rate ◽  
Repeated Aspiration ◽  
Rabbit Embryos

This study assessed the effects of oocyte age, cumulus cells and injection methods on in vitro development of intracytoplasmic sperm injection (ICSI) rabbit embryos. Oocytes were recovered from female rabbits superovulated with PMSG and hCG, and epididymal sperm were collected from a fertile male rabbit. The oocyte was positioned with the first polar body at 12 o'clock position, and a microinjection needle containing a sperm was inserted into the oocyte at 3 o'clock. Oolemma breakage was achieved by aspirating ooplasm, and the aspirated ooplasm and sperm were re-injected into the oocyte. The injected oocytes were cultured in M199 medium containing 10% fetal calf serum at 38 °C with 5% CO2 in air. The results showed that oocytes injected at 1 h post-collection produced a higher (p<0.05) fertilization rate than those injected at 4 or 7 h post-collection. Blastocyst rate in the 1 h group was higher (p<0.05) than in the 7 h group. Denuded oocytes (group A) and oocytes with cumulus cells (group B) were injected, respectively. Rates of fertilization and development of ICSI embryos were not significantly different (p<0.05) between the two groups. Four ICSI methods were applied in this experiment. In methods 1 and 2, the needle tip was pushed across half the diameter of the oocyte, and oolemma breakage was achieved by either a single aspiration (method 1) or repeated aspiration and expulsion (method 2) of ooplasm. In methods 3 and 4, the needle tip was pushed to the oocyte periphery opposite the puncture site, and oolemma breakage was achieved by either a single aspiration (method 3) or repeated aspiration and expulsion (method 4) of ooplasm. Fertilization rate in method 2 was significantly higher (p<0.05) than in methods 1 and 3. Blastocyst rates were not significantly different (p<0.05) among methods 1, 3 and 4, but method 2 produced a higher (p<0.05) blastocyst rate than method 3.

scholarly journals Next Generation Sequencing Detects Premeiotic Errors in Human Oocytes

2022 ◽  
Vol 23 (2) ◽  
pp. 665
Author(s):  
Harita Ghevaria ◽  
Sioban SenGupta ◽  
Roy Naja ◽  
Rabi Odia ◽  
Holly Exeter ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Polar Body ◽  
Independent Study ◽  
Comparative Genomic ◽  
Next Generation ◽  
Consistent Finding ◽  
Autosomal Aneuploidy ◽  
First Polar Body ◽  
Germinal Mosaicism ◽  
Generation Sequencing

Autosomal aneuploidy is the leading cause of embryonic and foetal death in humans. This arises mainly from errors in meiosis I or II of oogenesis. A largely ignored source of error stems from germinal mosaicism, which leads to premeiotic aneuploidy. Molecular cytogenetic studies employing metaphase fluorescence in situ hybridization and comparative genomic hybridisation suggest that premeiotic aneuploidy may affect 10–20% of oocytes overall. Such studies have been criticised on technical grounds. We report here an independent study carried out on unmanipulated oocytes that have been analysed using next generation sequencing (NGS). This study confirms that the incidence of premeiotic aneuploidy in an unselected series of oocytes exceeds 10%. A total of 140 oocytes donated by 42 women gave conclusive results; of these, 124 (88.5%) were euploid. Sixteen out of 140 (11.4%) provided evidence of premeiotic aneuploidy. Of the 140, 112 oocytes were immature (germinal vesicle or metaphase I), of which 10 were aneuploid (8.93%); the remaining 28 were intact metaphase II - first polar body complexes, and six of these were aneuploid (21.4%). Of the 16 aneuploid cells, half contained simple errors (one or two abnormal chromosomes) and half contained complex errors. We conclude that germinal mosaicism leading to premeiotic aneuploidy is a consistent finding affecting at least 10% of unselected oocytes from women undergoing egg collection for a variety of reasons. The importance of premeiotic aneuploidy lies in the fact that, for individual oocytes, it greatly increases the risk of an aneuploid mature oocyte irrespective of maternal age. As such, this may account for some cases of aneuploid conceptions in very young women.

Patterning of angiogenesis in the zebrafish embryo

Development ◽  
2002 ◽  
Vol 129 (4) ◽  
pp. 973-982 ◽  
Author(s):  
Sarah Childs ◽  
Jau-Nian Chen ◽  
Deborah M. Garrity ◽  
Mark C. Fishman
Keyword(s):  
Zebrafish Embryo ◽  
Regular Pattern ◽  
Wild Type ◽  
Cell Fates ◽  
Mutagenesis Screen ◽  
The Third ◽  
Shaped Cell ◽  
Vessel Growth ◽  
Lateral Posterior ◽  
Lineage Analysis

Little is known about how vascular patterns are generated in the embryo. The vasculature of the zebrafish trunk has an extremely regular pattern. One intersegmental vessel (ISV) sprouts from the aorta, runs between each pair of somites, and connects to the dorsal longitudinal anastomotic vessel (DLAV). We now define the cellular origins, migratory paths and cell fates that generate these metameric vessels of the trunk. Additionally, by a genetic screen we define one gene, out of bounds (obd), that constrains this angiogenic growth to a specific path. We have performed lineage analysis, using laser activation of a caged dye and mosaic construction to determine the origin of cells that constitute the ISV. Individual angioblasts destined for the ISVs arise from the lateral posterior mesoderm (LPM), and migrate to the dorsal aorta, from where they migrate between somites to their final position in the ISVs and dorsal longitudinal anastomotic vessel (DLAV). Cells of each ISV leave the aorta only between the ventral regions of two adjacent somites, and migrate dorsally to assume one of three ISV cell fates. Most dorsal is a T-shaped cell, based in the DLAV and branching ventrally; the second constitutes a connecting cell; and the third an inverted T-shaped cell, based in the aorta and branching dorsally. The ISV remains between somites during its ventral course, but changes to run mid-somite dorsally. This suggests that the pattern of ISV growth ventrally and dorsally is guided by different cues. We have also performed an ENU mutagenesis screen of 750 mutagenized genomes and identified one mutation, obd that disrupts this pattern. In obd mutant embryos, ISVs sprout precociously at abnormal sites and migrate anomalously in the vicinity of ventral somite. The dorsal extent of the ISV is less perturbed. Precocious sprouting can be inhibited in a VEGF morphant, but the anomalous site of origin of obd ISVs remains. In mosaic embryos, obd somite causes adjacent wild-type endothelial cells to assume the anomalous ISV pattern of obd embryos. Thus, the launching position of the new sprout and its initial trajectory are directed by inhibitory signals from ventral somites. Zebrafish ISVs are a tractable system for defining the origins and fates of vessels, and for dissecting elements that govern patterns of vessel growth.

Sign in / Sign up

Export Citation Format

Share Document