Effects of vitrification procedures on subsequent development and ultrastructure of in vitro-matured swamp buffalo (Bubalus bubalis) oocytes

2007 ◽  
Vol 19 (2) ◽  
pp. 383 ◽  
Author(s):  
Duangjai Boonkusol ◽  
Tassanee Faisaikarm ◽  
Andras Dinnyes ◽  
Yindee Kitiyanant
Keyword(s):  
Electron Microscopy ◽  
Mitochondrial Membrane ◽  
Subsequent Development ◽  
Bubalus Bubalis ◽  
Ultrastructural Changes ◽  
Cortical Granules ◽  
Transmission Electron ◽  
Developmental Capacity ◽  
Swamp Buffalo

The purpose of the present study was to investigate the effects of two vitrification procedures on developmental capacity and ultrastructural changes of matured swamp buffalo oocytes. In vitro-matured oocytes were vitrified by using 35 and 40% ethylene glycol as vitrification solution for solid surface vitrification (SSV) and in-straw vitrification (ISV), respectively. Survival rate of vitrified–warmed oocytes, evaluated on the basis of ooplasm homogeneity, oolemma integrity and zona pellucida intactness, as well as parthenogenetic blastocyst rates of vitrified–warmed oocytes were significantly higher with SSV (89.3 and 13.6%, respectively) than ISV (81.8 and 5.5%, respectively). However, they were still significantly lower than that of control oocytes (100 and 34.2%, respectively). For examining the ultrastructural changes, fresh, VS-exposed (ISV and SSV), and vitrified–warmed (ISV and SSV) oocytes were processed for transmission electron microscopy. In VS-exposed oocytes, reduction of microvilli abundance and damage of mitochondrial membrane were found only in the ISV group. In vitrified–warmed oocytes, however, it was clear that both methods of vitrification induced profound ultrastructural modifications to microvilli, mitochondria, oolemma and cortical granules as well as to the size and position of vesicles. Damaged mitochondria were, however, more abundant in ISV vitrified oocytes than in SSV vitrified oocytes, which correlated with the developmental data, showing the superiority of the SSV method. The present study demonstrated the feasibility of vitrification of in vitro-matured swamp buffalo oocytes.

109 EFFECTS OF VITRIFICATION PROCEDURES ON SUBSEQUENT DEVELOPMENT AND ULTRASTRUCTURE OF IN VITRO-MATURED SWAMP BUFFALO (BUBALUS BUBALIS) OOCYTES

2007 ◽  
Vol 19 (1) ◽  
pp. 172
Author(s):  
D. Boonkusol ◽  
T. Faisaikarm ◽  
A. Dinnyes ◽  
Y. Kitiyanant
Keyword(s):  
Membrane Integrity ◽  
Subsequent Development ◽  
Bubalus Bubalis ◽  
Ultrastructural Changes ◽  
Cortical Granules ◽  
Chi Square ◽  
Developmental Capacity ◽  
Swamp Buffalo ◽  
Chi Square Analysis

The purpose of this study was to investigate the effects of 2 vitrification procedures on the developmental capacity and ultrastructural changes of matured swamp buffalo (Bubalus bubalis) oocytes. In vitro-matured (IVM) oocytes were vitrified by using 35% and 40% ethylene glycol (EG) as vitrification solution (VS) for solid surface vitrification (SSV) and in-straw vitrification (ISV), respectively. Survival rate of vitrified–warmed oocytes was evaluated on the basis of homogeneous cytoplasm, membrane integrity, and complete zona pellucida. All developmental data were analyzed using chi-square analysis. P < 0.05 was considered significant. The blastocyst rates of parthenogenetic vitrified–warmed oocytes were significantly higher with SSV (89.3% and 13.6%, respectively) than with ISV (81.8% and 5.5%, respectively). However, they were still significantly lower than those of control (100% and 34.2%, respectively). For examining the ultrastructural changes, fresh VS-exposed (ISV and SSV), and vitrified–warmed oocytes were processed for transmission electron microscopy. In VS-exposed oocytes, reduction of microvilli abundance and damage of mitochondrial membrane were found only in the ISV group. In vitrified–warmed oocytes, however, it was clear that both methods of vitrification induced profound ultrastructural modifications to microvilli, mitochondria, oolemma, and cortical granules as well as to the size and position of vesicles. Damaged mitochondria were, however, more abundant in ISV vitrified oocytes than in SSV vitrified oocytes, which correlated with the developmental data, showing the superiority of the SSV method. This study demonstrated for the first time the feasibility of vitrification of IVM swamp buffalo oocytes.

scholarly journals Ultrastructure of in vitro oocyte maturation in buffalo (Bubalus bubalis)

Zygote ◽  
2010 ◽  
Vol 18 (4) ◽  
pp. 309-314 ◽  
Author(s):  
Rafael Gianella Mondadori ◽  
Tiago Rollemberg Santin ◽  
Andrei Antonioni Guedes Fidelis ◽  
Khesller Patrícia Olázia Name ◽  
Juliana Souza da Silva ◽  
...  
Keyword(s):  
Polar Body ◽  
Bubalus Bubalis ◽  
Ultrastructural Changes ◽  
Oocyte Nucleus ◽  
Cortical Granules ◽  
Immature Oocytes ◽  
Transmission Electron ◽  
High Lipid ◽  
First Polar Body

SummaryThe objective of the present study was to describe ultrastructural changes in the nucleus and cytoplasmic organelles during in vitro maturation (IVM) of buffalo cumulus–oocyte complexes (COCs). The structures were collected by ovum pick-up (OPU). Some COCs, removed from maturation medium at 0, 6, 12, 18 and 24 h, were processed for transmission electron microscopy. The average number of COCs collected by OPU/animal/session was 6.4, and 44% of them were viable. Immature oocytes had a peripherally located nucleus, Golgi complex and mitochondrial clusters, as well as a large number of coalescent lipid vacuoles. After 6 h of IVM, the oocyte nucleus morphology changed from round to a flatter shape, and the granulosa cells (GC) lost most of their contact with zona pellucida (ZP). At 12 h the first polar body was extruded and the aspect of lipid droplet changed to dark, probably denoting lipid oxidation. Cortical granules were clearly visible at 18 h of maturation, always located along the oocyte periphery. At 24 h of IVM the number of cortical granules increased. Ultrastructure studies revealed that: (1) immature oocytes have a high lipid content; (2) the perivitelline space (PS) increases during IVM; (3) Golgi complexes and mitochondrial clusters migrate to oocyte periphery during IVM; (4) 6 h of IVM are enough to lose contact between GC and ZP; (5) the oocyte lipid droplets’ appearance changes between 6 and 12 h of IVM.

scholarly journals The ultrastructure changes of Haemonchus contortus exposed to bamboo leaves (Gigantochloa apus) aqueous extract under in vitro condition

2020 ◽  
Vol 22 (1) ◽  
Author(s):  
Budi Purwo Widiarso ◽  
WISNU NURCAHYO ◽  
KURNIASIH KURNIASIH ◽  
JOKO PRASTOWO
Keyword(s):  
Electron Microscopy ◽  
Aqueous Extract ◽  
Haemonchus Contortus ◽  
Leaf Extract ◽  
Structural Changes ◽  
Ultrastructural Changes ◽  
Bamboo Leaves ◽  
Transmission Electron ◽  
External Part

Abstract. Widiarso, Nurcahyo W, Kurniasih, Prastowo J. 2021. The ultrastructure changes of Haemonchus contortus exposed to bamboo leaves (Gigantochloa apus) aqueous extract under in vitro condition. Biodiversitas 22: 1-5. The ultrastructural changes induced in adult Haemonchus contortus in vitro using the aqueous extract of bamboo leaves were assessed using scanning electron microscopy (SEM). The H. contortus adult females were obtained from three groups and treatment was repeated thrice. The first group (T0) was not treated with bamboo leaves; 100% of the worms lived. The second group (T1), treated with 0.1% bamboo leaf-extract, had 50% mortality 4 h after examination. The third group (T2), treated with 1% bamboo leaf-extract, had 100% mortality 4 h after examination. Five worms used per treatment were submerged in ethanol and incubated for 24 hours. The ultrastructural changes observed by SEM revealed structural alteration of the worm surface after in vitro contact with the bamboo leaf aqueous extract and compared to the control worms. The main changes concerned the anterior end or cephalic region, cuticle surface, and vulva flap area. The structural modification of the external part of the female reproductive system was found only in vitro. The structural changes found in the worms exposed to the bamboo leaves might affect their motility and nutrition with possible consequences on their reproduction. Transmission electron microscopy may help to understand the external changes observed in H. contortus.

scholarly journals Diabetes aggravates renal ischemia-reperfusion injury by repressing mitochondrial function and PINK1/Parkin-mediated mitophagy

2019 ◽  
Vol 317 (4) ◽  
pp. F852-F864 ◽  
Author(s):  
Yuan-Yuan Yang ◽  
Dao-Jing Gong ◽  
Jian-Jian Zhang ◽  
Xiu-Heng Liu ◽  
Lei Wang
Keyword(s):  
Electron Microscopy ◽  
Mitochondrial Function ◽  
High Glucose ◽  
Mitochondrial Membrane ◽  
Ischemia Reperfusion ◽  
Oxygen Species ◽  
Transmission Electron ◽  
Normal Glucose

Diabetes could aggravate ischemia-reperfusion (I/R) injury, but the underlying mechanism is unclear. In the present study, we aimed to investigate whether diabetes exacerbates renal I/R injury and its possible mechanism. In vitro, HK-2 cells under normal or high glucose conditions were subjected to hypoxia (12 h) followed by reoxygenation (3 h) (H/R). Cell viability, intracellular ATP content, mitochondrial membrane potential, reactive oxygen species production, and apoptosis were measured. In vivo, streptozotocin-induced diabetic and nondiabetic rats were subjected to I/R. Renal pathology, function, and apoptosis were evaluated by hematoxylin and eosin staining, transmission electron microscopy, and Western blot analysis. Compared with the normal glucose + H/R group, mitochondrial function (ATP, mitochondrial membrane potential, and reactive oxygen species) and mitophagy were reduced in the high glucose + H/R group, as was expression of phosphatase and tensin homolog-induced putative kinase 1 (PINK1) and Parkin. Also, cells in the high glucose + H/R group exhibited more apoptosis compared with the normal glucose + H/R group, as assessed by flow cytometry, TUNEL staining, and Western blot analysis. Compared with normal rats that underwent I/R, diabetic rats that underwent I/R exhibited more severe tubular damage and renal dysfunction as well as expression of the apoptotic protein caspase-3. Meanwhile, diabetes alleviated mitophagy-associated protein expression in rats subjected to I/R, including expression of PINK1 and Parkin. Transmission electron microscopy indicated that the mitophagosome could be hardly observed and that mitochondrial morphology and structure were obviously damaged in the diabetes + I/R group. In conclusion, our results, for the first time, indicate that diabetes could aggravate I/R injury by repressing mitochondrial function and PINK1/Parkin-mediated mitophagy in vivo and in vitro.

Biological approaches to characterize the mode of action of two 5-nitroindazolinone prototypes on Trypanosoma cruzi bloodstream trypomastigotes

Parasitology ◽  
2016 ◽  
Vol 143 (11) ◽  
pp. 1469-1478 ◽  
Author(s):  
CRISTINA FONSECA-BERZAL ◽  
CRISTIANE FRANÇA DA SILVA ◽  
RUBEM F. S. MENNA-BARRETO ◽  
MARCOS MEUSER BATISTA ◽  
JOSÉ A. ESCARIO ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Trypanosoma Cruzi ◽  
Mode Of Action ◽  
Mitochondrial Membrane ◽  
Drug Resistant ◽  
Cellular Mechanisms ◽  
Transmission Electron ◽  
Intracellular Vesicles

SUMMARYThe phenotypic activity of two 5-nitroindazolinones, i.e. 2-benzyl-1-propyl (22) and 2-benzyl-1-butyl (24) derivatives, previously proposed as anti-Trypanosoma cruzi prototypes, was presently assayed on bloodstream trypomastigotes (BT) of the moderately drug-resistant Y strain. Further exploration of putative targets and cellular mechanisms involved in their activity was also carried out. Therefore, transmission electron microscopy, high-resolution respirometry and flow cytometry procedures were performed on BT treated for up to 24 h with the respective EC50 value of each derivative. Results demonstrated that although 22 and 24 were not as active as benznidazole in this in vitro assay on BT, both compounds triggered important damages in T. cruzi that lead to the parasite death. Ultrastructural alterations included shedding events, detachment of plasma membrane and nuclear envelope, loss of mitochondrial integrity, besides the occurrence of a large number of intracellular vesicles and profiles of endoplasmic reticulum surrounding cytoplasmic organelles such as mitochondrion. Moreover, both derivatives affected mitochondrion leading to this organelle dysfunction, as reflected by the inhibition in oxygen consumption and the loss of mitochondrial membrane potential. Altogether, the findings exposed in the present study propose autophagic processes and mitochondrial machinery as part of the mode of action of both 5-nitroindazolinones 22 and 24 on T. cruzi trypomastigotes.

scholarly journals Cardiomyocyte mitochondrial oxidative stress and cytoskeletal breakdown in the heart with a primary volume overload

2015 ◽  
Vol 308 (6) ◽  
pp. H651-H663 ◽  
Author(s):  
Danielle M. Yancey ◽  
Jason L. Guichard ◽  
Mustafa I. Ahmed ◽  
Lufang Zhou ◽  
Michael P. Murphy ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Mitochondrial Membrane ◽  
Volume Overload ◽  
Ros Production ◽  
Isolated Cardiomyocytes ◽  
Transmission Electron ◽  
Cyclical Stretch

Left ventricular (LV) volume overload (VO) results in cardiomyocyte oxidative stress and mitochondrial dysfunction. Because mitochondria are both a source and target of ROS, we hypothesized that the mitochondrially targeted antioxidant mitoubiquinone (MitoQ) will improve cardiomyocyte damage and LV dysfunction in VO. Isolated cardiomyocytes from Sprague-Dawley rats were exposed to stretch in vitro and VO of aortocaval fistula (ACF) in vivo. ACF rats were treated with and without MitoQ. Isolated cardiomyocytes were analyzed after 3 h of cyclical stretch or 8 wk of ACF with MitoSox red or 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate to measure ROS and with tetramethylrhodamine to measure mitochondrial membrane potential. Transmission electron microscopy and immunohistochemistry were used for cardiomyocyte structural assessment. In vitro cyclical stretch and 8-wk ACF resulted in increased cardiomyocyte mitochondrial ROS production and decreased mitochondrial membrane potential, which were significantly improved by MitoQ. ACF had extensive loss of desmin and β2-tubulin that was paralleled by mitochondrial disorganization, loss of cristae, swelling, and clustering identified by mitochondria complex IV staining and transmission electron microscopy. MitoQ improved mitochondrial structural damage and attenuated desmin loss/degradation evidenced by immunohistochemistry and protein expression. However, LV dilatation and fractional shortening were unaffected by MitoQ treatment in 8-wk ACF. In conclusion, although MitoQ did not affect LV dilatation or function in ACF, these experiments suggest a connection of cardiomyocyte mitochondria-derived ROS production with cytoskeletal disruption and mitochondrial damage in the VO of ACF.

Arterial Wall Changes in Cerebral Vasospasm

Neurosurgery ◽  
1989 ◽  
Vol 25 (5) ◽  
pp. 736-746 ◽  
Author(s):  
J. M. Findlay ◽  
B. K. A. Weir ◽  
K. Kanamaru ◽  
F. Espinosa
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Vessel Wall ◽  
Cerebral Arteries ◽  
Fold Increase ◽  
Ultrastructural Changes ◽  
Wall Thickening ◽  
Vessel Wall Thickness ◽  
Transmission Electron

Abstract A right-sided subarachnoid hemorrhage (SAH) was created in 12 monkeys. Only the right (clot-side) cerebral arteries developed angiographic vasospasm (VSP), which was maximal 7 days after SAH. Eight animals were killed at this time and the remainder at 14 days. At the time of killing the middle cerebral arteries (MCAs) were harvested, and four normal, left (non-clot-side) MCAs were vasoconstricted in vitro with prostaglandin F2… All MCAs were studied with scanning and transmission electron microscopy. Right MCAs in maximal VSP 7 days from SAH were undistinguishable on scanning electron microscopy from normal arteries vasoconstricted in vitro: both groups demonstrated a mean 57% reduction in vessel caliber and a 5-fold increase in vessel wall thickness compared to normal, nonvasoconstricted left MCAs. On transmission electron microscopy, however, arteries in SAH-induced VSP showed degenerative changes in the tunica intima and media. These changes were still evident at 14 days. despite considerable resolution of VSP. These findings, as well as those from other pathological studies of animal and human cerebral arteries in VSP, suggest that the arterial narrowing and vessel wall thickening seen within several weeks of SAH is due primarily to medial contraction, but unlike simple vasoconstruction, is associated with degenerative ultrastructural changes in the endothelium and vascular smooth muscle cells which may denote a temporarily irreversible state.

Maytansine-Induced Mitotic Arrest in Human Lymphoblasts

1977 ◽  
Vol 35 ◽  
pp. 460-461
Author(s):  
Tai-Te Chao ◽  
John Sullivan ◽  
Awtar Krishan
Keyword(s):  
Electron Microscopy ◽  
Cell Cycle ◽  
Transmission Electron Microscopy ◽  
Tubulin Polymerization ◽  
Vinca Alkaloids ◽  
Antimitotic Activity ◽  
Transmission Electron ◽  
Flow Microfluorometry ◽  
Brain Tubulin

Maytansine, a novel ansa macrolide (1), has potent anti-tumor and antimitotic activity (2, 3). It blocks cell cycle traverse in mitosis with resultant accumulation of metaphase cells (4). Inhibition of brain tubulin polymerization in vitro by maytansine has also been reported (3). The C-mitotic effect of this drug is similar to that of the well known Vinca- alkaloids, vinblastine and vincristine. This study was carried out to examine the effects of maytansine on the cell cycle traverse and the fine struc- I ture of human lymphoblasts.Log-phase cultures of CCRF-CEM human lymphoblasts were exposed to maytansine concentrations from 10-6 M to 10-10 M for 18 hrs. Aliquots of cells were removed for cell cycle analysis by flow microfluorometry (FMF) (5) and also processed for transmission electron microscopy (TEM). FMF analysis of cells treated with 10-8 M maytansine showed a reduction in the number of G1 cells and a corresponding build-up of cells with G2/M DNA content.

Correlation Between Cellular Functions and Morphological Changes of Human Trophoblast in Vitro

1975 ◽  
Vol 33 ◽  
pp. 600-601
Author(s):  
John C. Garancis ◽  
Robert O. Hussa ◽  
Michael T. Story ◽  
Donald Yorde ◽  
Roland A. Pattillo
Keyword(s):  
Electron Microscopy ◽  
Cellular Proliferation ◽  
Morphological Changes ◽  
Culture Fluid ◽  
Cellular Functions ◽  
Human Trophoblast ◽  
Transmission Electron ◽  
Marked Activity ◽  
Malignant Trophoblast

Human malignant trophoblast cells in continuous culture were incubated for 3 days in medium containing 1 mM N6-O2'-dibutyryl cyclic adenosine 3':5'-monophosphate (dibutyryl cyclic AMP) and 1 mM theophylline. The culture fluid was replenished daily. Stimulated cultures secreted many times more chorionic gonadotropin and estrogens than did control cultures in the absence of increased cellular proliferation. Scanning electron microscopy revealed remarkable surface changes of stimulated cells. Control cells (not stimulated) were smooth or provided with varying numbers of microvilli (Fig. 1). The latter, usually, were short and thin. The surface features of stimulated cells were considerably different. There was marked increase of microvilli which appeared elongated and thick. Many cells were covered with confluent polypoid projections (Fig. 2). Transmission electron microscopy demonstrated marked activity of cytoplasmic organelles. Mitochondria were increased in number and size; some giant forms with numerous cristae were observed.

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