Optimized study of an in vitro 3D culture of preantral follicles in mice
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- Journal of Veterinary Science
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- v.24 no.1
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- pp.4.1-4.16
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- 2023
Background: In vitro culture of preantral follicles is a promising technology for fertility preservation. Objectives: This study aims to investigate an optimized three-dimensional (3D) fetal bovine serum (FBS)-free preantral follicle culture system having a simple and easy operation. Methods: The isolated follicles from mouse ovaries were randomly divided in an ultra-low attachment 96-well plates supplement with FBS or bovine serum albumin (BSA) culture or encapsulated with an alginate supplement with FBS or BSA culture. Meanwhile, estradiol (E2) concentration was assessed through enzyme-linked immunosorbent assay of culture supernatants. The diameter of follicular growth was measured, and the lumen of the follicle was photographed. Spindle microtubules of oocytes were detected via immunofluorescence. The ability of oocytes to fertilize was assessed using in vitro fertilization. Results: The diameters were larger for the growing secondary follicles cultured in ultra-low attachment 96-well plates than in the alginate gel on days 6, 8, and 10 (p < 0.05). Meanwhile, the E2 concentration in the BSA-supplemented medium was significantly higher in the alginate gel than in the other three groups on days 6 and 8 (p < 0.05), and the oocytes in the FBS-free system could complete meiosis and fertilization in vitro. Conclusions: The present study furnishes insights into the mature oocytes obtained from the 3D culture of the preantral follicle by using ultra-low attachment 96-well plate with an FBS-free system in vitro and supports the clinical practices to achieve competent, mature oocytes for in vitro fertilization.
This study was conducted to examine the role of intact cumulus cells during in vitro fertilization (IVF) on sperm penetration, male pronuclear (MPN) formation and subsequent embryo development of oocytes matured and fertilized in vitro. Cumulus-oocyte complexes obtained from the slaughtered gilt ovaries were matured for 44 h in TCM199 containing 10% porcine follicular fluid, epidermal growth factor and hormones. After maturation culture, denuded oocytes or oocytes with intact cumulus cells were coincubated with frozen-thawed boar semen for 8h in a modified tris-buffered medium containing 5mM caffeine and 10mM calcium chloride. Putative zygotes were fixed and examined for sperm penetration and MPN formation (Experiments
Polycystic ovary syndrome (PCOS) is hormonal imbalance condition as the endocrine and metabolic disorder that induces the infertility and various complications in reproductive age women. Estradiol valerate (EV) is used hormone replacement therapy in menopausal women and is reported that excessive administration of EV induces the PCOS. Nerve growth factor (NGF) is the factor to regulate the survival and maturation of developing neuronal cell and is also synthesized in ovary. And NGF is overexpressed in EV-induced polycystic ovary (PCO) as previously reported. Therefore, this study examined the possibility of NGF as can be used the biological marker in diagnosis of PCOS, the hormonal imbalance condition, using PCO and CHO (chinese hamster ovarian) cell lines. The concentration of EV treatment is optimized a 1 mg as not influence on the proliferation of CHO cell but 2 mg and 3 mg of EV treatment have the inhibition effect at initial stage. The morphological change was not observed in CHO cell after dose dependent manner treatment of EV. Expression of NGF mRNA and protein is significantly increased at 30 min after EV treatment in CHO cells compared to that of control. And NGF protein expression is strongly increased in PCO tissue, which observed many follicular cysts compared to normal ovary tissue. Taken together, overexpression of NGF may be act as a molecule to induce an abnormal development of follicle, suggesting that NGF can be used as a biological marker in diagnosis of PCOS.
Seventy four patients with pituitary adenoma received radiation therapy (RT) on the pituitary area using 6 MV linear accelerator during the past 7 years at the Division of Radiation Therapy, Kangnam St. Mary's Hospital, Catholic University Medical College. Thirty nine were men and 35 were women. The age ranged from 7 to 65 years with the mean being 37 years. Sixty five (
The reproductive cycle of the small filefish, Rudarius ercodes was investigated based on the annual variations of gonadosomatic index(GSI) and hepatosomatic index(HSI) by electronic and photic microscophy. The specimens used were collected at the coastal area of Benden island, Sizuokagen, Japan, from September 1982 to August 1983. GSI began to increase from March, starting season of longer daylength and higher water temperature, and reached the maximum value between June and August. It began to decrease from September with the lowest value appearing between November and February without any evident variation. The annual variations of HSI were not distinct in male filefish and were negatively related to GSI in female : HSI decreased in the summer season when the ovary was getting mature and reached the maximum in the winter season when the ovary was getting retrogressive. The ovary consisted of a pair of saccular structure with numerous ovarian sacs branched toward the median cavity. Oogonia divided and proliferated along the germinal epithelium of the ovarian sac. Young oocytes with basophile cytoplasm showed several scattering nucleoli along the nuclear membrane. when the oocytes growing to about 300
Two-Cell mouse embryos were incubated in the anterior chamber of the rat eye, which has been known as the best place among other animals' for the mouse ovum maturation, in order to observe the capability of their early development. Within 120 hours after incubation, 71.0% of two-cell embryos have developed to the blastocysts in the male rat eye, while only 38.5% in the eye of the same mouse as donated two-cell embryos. Thus, the rat eye chamber provides more favourable environment to the embryos than the mouse itself. The results are consistent with those of the previous studies comparing the maturation of the mouse follicular oocytes in the mouse and the rat eye chamber. Although the aqueous humor which is filled in the anterior chamber of the eye is characterized by its specific properties, being of higher osmolarity, higher concentrations of ascorbic acid, pyruvate and lactate, but lower of proteins and lower temperature than those in blood or lymph serum, The embryos are able to under-take their cleavage as normal as in vivo or in vitro. Concerning with a number of studies in vitro on the development of the mouse embryos which are requiring a very limited condition, the fact that they are able to manage their further development under very different enviroment from our knowledges would provide us a moment to understand their behavior during the early development. The difference of the proportion of the developed blastocysts between in the mouse eye chamber and in the rat can possibly be resulted from the species specific difference in the physicochemical properties between their eye chambers. This assumption is based upon the findings by many investigators who chmpared the nature of the eye chamber of various animals. As a consequence, the rat eye chamber might consist of better properties for the embryonal growth than the mouse eye chamber. The mouse embryos cleaved with a delayed period. In normal development they complete almost the cleavage within 94 hours after fertilization. However, in the present studies, 81.1% of two-cell embryos developed to the blastocysts and the morula in 120 hours in the eye chamber, assumed to be about 154 hours after fertilization. Such delay in development would be caused mainly by the low temperature of the eye chamber. At present we can make two assumptions to explain the capability of the emtryonal development in the eye chambers. One is that the embryos would possess an ability to adapt themselves to the environment which provides unfavourable conditions. The other is that the embryos might remain for a certain duration in the eye chamber, which is filled with a new body fluid produced immediately after the loss of the aqueous humor and the fluid of which becomes similar to blood serum in component. The first assumption is highly reliable since the embryonal cells are mostly at the undifferentiated state and so they probably engage a simple metabolism during their early period. The second assumption is induced by the fact that the rabbit eye chamber produces a plasmoid humor which has mostly similar components to blood serum after loss of aqueous humor through cornea by puncturing. However, the plasmoid humor is substituted by the initial aqueous humor in eight hours. Even though this finding, production of the new fluid, could be applied to the rat eye, it is hardly reliabel that the plasmoid humor remains for such a long period as 120 hours. Consequently, the development of the embryos is more likely due to their adaptability to the new environment during their early developmental stages.
Objective: The aim of this study was to compare the clinical efficacy of clomiphene citrate (CC) and letrozole combined with gonadotropins for controlled ovarian stimulation (COS) in patients with CC-induced thin endometrium Methods: Fifty-one intrauterine insemination cycles performed in patients who previously had a thin endometrium (<8 mm) to ovulation induction using CC were included in this study. A CC 100 mg/day (CC+gonadotropin group, n=26) or letrozole 2.5 or 5 mg/day (letrozole+gonadotropin group, n=25) was administered on day 3~7 of the menstrual cycle, combined with gonadotropins at dose 75~150 IU every other day starting on day 5~7. We compared total dose of gonadotropin used, endometrial thickness, endometrial pattern, number of follicles
The wall shear stress in the vicinity of end-to end anastomoses under steady flow conditions was measured using a flush-mounted hot-film anemometer(FMHFA) probe. The experimental measurements were in good agreement with numerical results except in flow with low Reynolds numbers. The wall shear stress increased proximal to the anastomosis in flow from the Penrose tubing (simulating an artery) to the PTFE: graft. In flow from the PTFE graft to the Penrose tubing, low wall shear stress was observed distal to the anastomosis. Abnormal distributions of wall shear stress in the vicinity of the anastomosis, resulting from the compliance mismatch between the graft and the host artery, might be an important factor of ANFH formation and the graft failure. The present study suggests a correlation between regions of the low wall shear stress and the development of anastomotic neointimal fibrous hyperplasia(ANPH) in end-to-end anastomoses. 30523 T00401030523 ^x Air pressure decay(APD) rate and ultrafiltration rate(UFR) tests were performed on new and saline rinsed dialyzers as well as those roused in patients several times. C-DAK 4000 (Cordis Dow) and CF IS-11 (Baxter Travenol) reused dialyzers obtained from the dialysis clinic were used in the present study. The new dialyzers exhibited a relatively flat APD, whereas saline rinsed and reused dialyzers showed considerable amount of decay. C-DAH dialyzers had a larger APD(11.70
The wall shear stress in the vicinity of end-to end anastomoses under steady flow conditions was measured using a flush-mounted hot-film anemometer(FMHFA) probe. The experimental measurements were in good agreement with numerical results except in flow with low Reynolds numbers. The wall shear stress increased proximal to the anastomosis in flow from the Penrose tubing (simulating an artery) to the PTFE: graft. In flow from the PTFE graft to the Penrose tubing, low wall shear stress was observed distal to the anastomosis. Abnormal distributions of wall shear stress in the vicinity of the anastomosis, resulting from the compliance mismatch between the graft and the host artery, might be an important factor of ANFH formation and the graft failure. The present study suggests a correlation between regions of the low wall shear stress and the development of anastomotic neointimal fibrous hyperplasia(ANPH) in end-to-end anastomoses. 30523 T00401030523 ^x Air pressure decay(APD) rate and ultrafiltration rate(UFR) tests were performed on new and saline rinsed dialyzers as well as those roused in patients several times. C-DAK 4000 (Cordis Dow) and CF IS-11 (Baxter Travenol) reused dialyzers obtained from the dialysis clinic were used in the present study. The new dialyzers exhibited a relatively flat APD, whereas saline rinsed and reused dialyzers showed considerable amount of decay. C-DAH dialyzers had a larger APD(11.70