Fertilization, Implantation, and Endocrinology of Pregnancy

2011 ◽  
Author(s):  
Bruce R. Carr ◽  
Victor E. Beshay
Keyword(s):  
Fallopian Tube ◽  
Meiotic Division ◽  
Genital Tract ◽  
Polar Body ◽  
Female Genital Tract ◽  
Cumulus Cells ◽  
Hormonal Changes ◽  
Egg Maturation ◽  
Ciliary Action ◽  
First Polar Body

The complex and coordinated set of events leading to sperm and egg maturation and transport in the female genital tract that culminates in fertilization is one of the most remarkable phenomena in nature. This set of events is followed by the equally important unique processes of implantation, fetal maturation, and parturition. The hormonal changes that regulate these events are dependent on the close interaction of the fetal-placental-maternal unit. Just before ovulation, the egg, which has been arrested in the diplotene stage, completes the first meiotic division and forms the first polar body. The second meiotic division starts at the time of ovulation but ends only after fertilization by a sperm. The process of egg maturation is regulated through a closely interrelated set of hormonal events, most notably involving follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estrogen. At the time of ovulation the fimbria of the oviduct are closely applied to the surface of the ovary. The extruded oocyte and adherent granulosa cells, known as the cumulus oophorus, is collected by the ciliated fimbrial end of the fallopian tube. The transport of the egg into the end of the fallopian tube occurs within minutes and is regulated primarily by ciliary action. The cumulus cells are able to communicate with one another via a network of intercellular bridges through the zona pellucida to the perivitelline space. The cumulus cells have also been reported to play a role in nutrition and maintenance of the ovum. There are three different stages of passage of the ovum through the fallopian tube. The first stage includes the transfer of the ovum from the fimbriated end of the fallopian tube until the egg reaches and is retained at the ampullary-isthmic junction. The ampullary-isthmic junction is a functional block but is not a clearly defined anatomical structure. The ovum remains at this junction for 1–2 days, during which time fertilization occurs.

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.

The Formation of the Second Maturation Spindle in the Eggs of Limnaea, Limax, and Agriolimax

Development ◽  
1958 ◽  
Vol 6 (1) ◽  
pp. 28-51
Author(s):  
Chr. P. Raven ◽  
Frouke C. M. Escher ◽  
W. M. Herrebout ◽  
J. A. Leussink
Keyword(s):  
Polar Body ◽  
Egg Maturation ◽  
First Polar Body ◽  
Limnaea Stagnalis ◽  
Gradual Transformation ◽  
Sperm Aster

Egg maturation in Limnaea stagnalis has been described by Raven (1945, 1949). It exhibits some peculiarities, by which the Limnaea egg seemed to stand apart from all other animal eggs in which the maturation processes have been accurately studied so far. In the first place it appeared that the second maturation spindle arises by a direct and gradual transformation of the centrosphere at the inner end of the first maturation amphiaster, remaining in the egg after the extrusion of the first polar body. Secondly, the above-mentioned investigations revealed the still more astonishing fact that the inner aster of the second maturation spindle is none other than the sperm aster, which arises quite independently round about the time of the extrusion of the first polar body, grows considerably in size while the second maturation spindle is being formed, and then fuses secondarily with the latter's deep end.

98 BLASTOCYST DEVELOPMENT RATE OF CLONED CAT EMBRYOS USING SERIAL CLONING

2007 ◽  
Vol 19 (1) ◽  
pp. 166
Author(s):  
X. J. Yin ◽  
H. S. Lee ◽  
E. G. Choi ◽  
X. F. Yu ◽  
B. H. Choi ◽  
...  
Keyword(s):  
Nuclear Transfer ◽  
Second Generation ◽  
Assisted Reproductive Technologies ◽  
Polar Body ◽  
Donor Cell ◽  
Fibroblast Cell ◽  
Cumulus Cells ◽  
Reproductive Technologies ◽  
Blastocyst Development ◽  
First Polar Body

Domestic cats are a useful research model to develop assisted reproductive technologies for the conservation of endangered felids. Previously, we produced cloned offspring derived from somatic cell nuclear transfer of ear skin fibroblasts obtained from a deaf, odd-eyed, male Turkish Angora. The aim of this study was to assess the cloning efficiency of the fibroblasts derived from a cloned cat. Fibroblast cell lines were established from 6-mm skin biopsies taken from a deaf, odd-eyed, male Turkish Angora and his clone. The protocol for nuclear transfer was described previously (Yin et al. 2005 Reproduction 129, 245–249). Briefly, cumulus cells were removed from the ova by gently pipetting them into TCM-199 supplemented with 0.1% hyaluronidase. The denuded oocytes were then cultured in TCM-199 supplemented with 0.2 �g mL-1 demecolcine for 1 h and placed into TCM-199 containing 5 �g mL-1 cytochalasin B and 0.2 �g mL-1 demecolcine. The first polar body and protruded chromatin plate were removed with a beveled micropipette. Micromanipulation was used to place a single donor cell nucleus into the perivitelline space of enucleated ova. The ovum-cell couplets were fused and pulse activated. The activated couplets were cultured in 500 �L of CRI medium supplemented with 0.3% BSA for 2 days. The cleaved embryos were cultured in CRII medium supplemented with 10% FBS for 5 days. The cleavage and blastocyst development rates were 38.5% and 3.5% for second generation cloned embryos. A total of 310 second generation cloned embryos were transplanted to 9 surrogates, and 2 pregnancies at 30 days were determined by ultrasonography. One pregnancy was aborted at 40 days of gestation; the second pregnancy continued. These results indicate that the serial cloning of a cat can be generated efficiently up until pregnancy. This work was supported by KOSEF (grant #M10525010001-05N2501-00110).

74 IN VIVO SURVIVAL OF DOMESTIC CAT OOCYTES AFTER VITRIFICATION, INTRACYTOPLASMIC SPERM INJECTION, AND TRANSFER TO RECIPIENTS

2008 ◽  
Vol 20 (1) ◽  
pp. 118 ◽  
Author(s):  
M. C. Gómez ◽  
N. Kagawa ◽  
C. E. Pope ◽  
M. Kuwayama ◽  
S. P. Leibo ◽  
...  
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Oocyte Retrieval ◽  
Cumulus Cells ◽  
Domestic Cat ◽  
Minimal Amount ◽  
Vitro Fertilization ◽  
First Polar Body

The ability to cryopreserve female gametes efficiently holds immense economic and genetic implications. The purpose of the present project was to determine if domestic cat oocytes could be cryopreserved successfully by use of the Cryotop method. We evaluated (a) cleavage frequency after in vitro fertilization (IVF) v. intracytoplasmic sperm injection (ICSI) of in vivo- and in vitro-matured oocytes after vitrification, and (b) fetal development after transfer of resultant embryos into recipients. In vivo-matured cumulus–oocyte complexes (COCs) were recovered from gonadotropin-treated donors at 24 h after LH treatment, denuded of cumulus cells, and examined for the presence of the first polar body (PB). In vitro-matured COCs were obtained from ovaries donated by local clinics and placed into maturation medium for 24 h before cumulus cells were removed and PB status was determined. Oocytes were cryopreserved by the Cryotop method (Kuwayama et al. 2005 Reprod. Biomed. Online 11, 608–614) in a vitrification solution consisting of 15% DMSO, 15% ethylene glycol, and 18% sucrose. For IVF, oocytes were co-incubated with 1 � 106 motile spermatozoa mL–1 in droplets of modified Tyrode's medium in 5% CO2/air at 38�C (Pope et al. 2006 Theriogenology 66, 59–71). For ICSI, an immobilized spermatozoon was loaded into the injection pipette, which was then pushed through the zona pellucida into the ooplasm. After a minimal amount of ooplasm was aspirated into the pipette, the spermatozoon was carefully expelled, along with the aspirated ooplasm. After ICSI, or at 5 or 18 h post-insemination, in vivo- and in vitro-matured oocytes, respectively, were rinsed and placed in IVC-1 medium (Pope et al. 2006). As assessed by normal morphological appearance after liquefaction, the survival rate of both in vivo- and in vitro-matured oocytes was >90% (93–97%). For in vitro-matured oocytes, cleavage frequencies after IVF of control and vitrified oocytes were 73% (16/22) and 53% (30/57), respectively, as compared to 68% (19/28) after ICSI of vitrified oocytes (P > 0.05). For in vivo-matured oocytes, cleavage frequencies after IVF of control and vitrified oocytes were 55% (18/33) and 35% (6/17), respectively, compared to 50% (10/20) after ICSI of vitrified oocytes (P > 0.05). At 18–20 h after ICSI, 18 presumptive zygotes and four 2-cell embryos derived from vitrified in vitro-matured oocytes and 19 presumptive zygotes produced from seven in vivo-matured and 12 in vitro-matured vitrified oocytes were transferred by laparoscopy into the oviducts of two recipients at 24–26 h after oocyte retrieval. The two recipients were 9-month-old IVF/ET-derived females produced with X-sperm sorted by flow cytometry. At ultrasonography on Day 22, both recipients were pregnant, with three live fetuses observed in one recipient and one live fetus seen in the second recipient. On Day 63 and Day 66 of gestation, four live kittens were born, without assistance, to the two recipients. The one male and three female kittens weighed an average of 131 g. In summary, in vivo viability of zygotes/embryos produced by ICSI of cat oocytes vitrified by the Cryotop method was demonstrated by the birth of live kittens following transfer to recipients.

032. Cdh1: A CELL CYCLE PROTEIN INVOLVED IN FEMALE MEIOSIS AND PREVENTION OF ANEUPLOIDY

2010 ◽  
Vol 22 (9) ◽  
pp. 10
Author(s):  
K. T. Jones
Keyword(s):  
Meiotic Division ◽  
Polar Body ◽  
Single Copy ◽  
Cyclin B1 ◽  
Cell Cycle Protein ◽  
Prophase I ◽  
First Polar Body ◽  
A Cell

Mammalian oocytes are arrested at the dictyate stage of prophase I in the ovary. In growing follicles, oocytes can become responsive to Luteinising Hormone and will undergo meiotic resumption just before ovulation. During the first meiotic division, homologous chromosomes are segregated, a process that is very error prone in human oocytes. By ovulation the oocyte has extruded its first polar body and has re-arrested at metaphase of the first meiotic division. Recent work from our lab has established that the protein Cdh1 is involved uniquely in both in the process of prophase I arrest and the correct segregation of homologs in meiosis I. Thus in cultured oocytes, in vitro antisense knockdown of Cdh1 induces both meiotic resumption and high rates of aneuploidy as a result of non-disjunction during first meiosis. Cdh1 causes prophase I arrest by inducing cyclin B1 degradation and maintaining low levels of the kinase CDK1, whose activity induces meiotic resumption. Cdh1 is an activator of the Anaphase-Promoting Complex (APC), a ubiquitin ligase that earmarks proteins such as cyclin B1 for proteolysis. Cdh1 prevents aneuploidy by causing the degradation of Cdc20, a protein that is responsible for activating the APC once all homologs are correctly aligned at metaphase. Thus loss of Cdh1 seems to prematurely activate APC(Cdc20) activity. It is interesting that a single protein can affect two important meiotic transitions in oocytes. However to explore its functions more fully, and confirm that an in vitro knockdown is faithfully replicated by in vivo loss, a targeted knockout of Cdh1 is needed. Therefore we have generated an oocyte specific Cdh1 knockout by ZP3 promoter driven Cre- recombinase activity in oocytes carrying loxP insertions in the single copy Cdh1 gene. This talk will therefore focus on the effects of an in vivo Cdh1 knockout.

XXXVII.—Mendelian Action on Differentiated Sex

1909 ◽  
Vol 29 ◽  
pp. 607-618
Author(s):  
D. Berry Hart
Keyword(s):  
Fallopian Tube ◽  
Genital Tract ◽  
Female Genital Tract ◽  
Definite Form ◽  
Human Male ◽  
Prostatic Utricle ◽  
Opposite Sex ◽  
Vaginal Tract ◽  
Appendix Testis ◽  
Female Genital

(Abstract)It has long been known that the male and the female human genital tract contain not only organs characteristic of their sex proper, but also certain parts of the opposite sex in a less developed but yet perfectly definite form. Thus the female genital tract is made up of, not only its characteristic organs, the ovaries, tubes, uterus, etc., but also the epoophoron (parovarium) and its duct, the equivalent of the epididymis and ductus epididymis of the testis. In the same way, the human male has his characteristic sexual organs and also the appendix testis and prostatic utricle, the representatives of the fimbriated end of the Fallopian tube and of the lower end of the vaginal tract (hymen mainly, but varying).

Caprine blastocyst formation following intracytoplasmic sperm injection and defined culture

Zygote ◽  
1997 ◽  
Vol 5 (3) ◽  
pp. 261-265 ◽  
Author(s):  
L. Keskintepe ◽  
P.C. Morton ◽  
S.E. Smith ◽  
M.J. Tucker ◽  
A.A. Simplicio ◽  
...  
Keyword(s):  
Breeding Season ◽  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Egg Yolk ◽  
Cumulus Cells ◽  
Essential Amino Acids ◽  
Defined Medium ◽  
First Polar Body ◽  
Defined Culture ◽  
Oviduct Fluid

SummaryExperiments were undertaken to develop intracytoplasmic sperm injection (ICSI) to produce caprine embryos out of the normal breeding season. Oocytes were obtained from 2–6 mm ovarian follicles at slaughter. Selected oocytes with two to four layers of cumulus cells were incubated in 1 ml of H-TCM 199 supplemented with 10 μg each of oFSH and bLH (NHPP, NIDDK, NICHD, USDA) and 20% fetal bovine serum (FBS) in a thermos (38.5°C) for 4.5 h during transportation. Then, oocytes were transferred into 75 μl of freshly prepared maturation medium under paraffin oil and a mixture of 5% O2, 5% CO2 and 90% N2. Approximately 26 h after recovery oocytes were denuded by incubation with hyaluronidase (100 IU/ml) and pipetting and held at 38.5°C for 90 min. Spermatozoa frozen in egg yolk extender were thawed in a 37°C water bath for 15s. Motile fractions were selected by swim-up, then incubated for 90 mm in TALP with 10 μg heparin/ml. Each oocyte was positioned with its first polar body at 6 or 12 o'clock by a holding pipette. Sperm (1 μl) were added to 10 μl medium containing 10% polyvinylpyrrolidone. A sperm cell was aspirated into a pipette, and then injected head-first into the cytoplasm of an oocyte maintained in H-TCM 199 + 20% FBS at 37°C. Injected oocytes were transferred to HM and, after 90 min, cultured in 50 μl of BSA-free synthetic oviduct fluid plus polyvinyl alcohol, citrate and non-essential amino acids. Results demonstrate that caprine blastocysts can be produced outside the breeding season by the use of frozen-thawed semen and injection of sperm cells with broken tails into ova followed by culture in defined medium.

scholarly journals Immunohistochemical expression of keratan sulfate: a possible diagnostic marker for carcinomas of the female genital tract

2011 ◽  
Vol 64 (12) ◽  
pp. 1058-1063 ◽  
Author(s):  
Tsutomu Miyamoto ◽  
Keiko Ishii ◽  
Ryouichi Asaka ◽  
Akihisa Suzuki ◽  
Akiko Takatsu ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Urinary Tract ◽  
Gastrointestinal Tract ◽  
Fallopian Tube ◽  
Genital Tract ◽  
Diagnostic Marker ◽  
Female Genital Tract ◽  
Keratan Sulfate ◽  
Digestive Organs ◽  
Female Genital

AimsThe authors previously reported the expression of keratan sulfate (KS), a glycosaminoglycan, in the epithelium of normal and neoplastic endometria. The aim of this study was to evaluate its potential use as a diagnostic marker, and the expression of KS was investigated in other human epithelial tissues.MethodsExpression was examined immunohistochemically using 102 samples of normal epithelia and 110 samples of carcinomas from the female genital tract (FGT; cervix, endometrium, ovary, fallopian tube), digestive organs (gastrointestinal tract, pancreas, liver), urinary tract, lung, mammary gland, thyroid and mesothelium.ResultsIn normal tissues, KS was consistently detected in the FGT and ectopic endometrium (25/26), but was not found in the digestive organs (1/42) and urinary tract (0/6), and was only partly detected in the lung (7/10), mammary gland (3/9) and thyroid (4/4). In malignant tissues, KS was consistently observed in carcinomas of the endometrium, ovary and fallopian tube (29/32), and was partly detected in carcinomas of the lung, mammary gland, thyroid, pancreas and mesothelium, but was absent in carcinomas of the gastrointestinal tract (0/17), liver (0/5) and urinary tract (0/11). Among carcinomas of the FGT, digestive organs and urinary tract, KS positivity suggested the possibility of FGT carcinomas, with 79.5% (31/39) sensitivity and 92.9% (39/42) specificity.ConclusionsKS is a potentially useful marker for the supportive diagnosis of the primary site of metastatic carcinomas or unknown primary carcinomas, especially in the abdominal cavity.

scholarly journals Developmental competence in vivo and in vitro of in vitro-matured equine oocytes fertilized by intracytoplasmic sperm injection with fresh or frozen-thawed spermatozoa

Reproduction ◽  
2002 ◽  
pp. 455-465 ◽  
Author(s):  
YH Choi ◽  
CC Love ◽  
LB Love ◽  
DD Varner ◽  
S Brinsko ◽  
...  
Keyword(s):  
Intracytoplasmic Sperm Injection ◽  
Polar Body ◽  
Cumulus Cells ◽  
Fertilization Rate ◽  
Developmental Competence ◽  
Cleavage Rate ◽  
First Polar Body ◽  
Bovine Oocytes

This study was undertaken to evaluate the development of equine oocytes in vitro and in vivo after intracytoplasmic sperm injection (ICSI) with either fresh or frozen-thawed spermatozoa, without the use of additional activation treatments. Oocytes were collected from ovaries obtained from an abattoir and oocytes classified as having expanded cumulus cells were matured in M199 with 10% fetal bovine serum and 5 microU FSH ml(-1). After 24-26 h of in vitro maturation, oocytes with a first polar body were selected for manipulation. Fresh ejaculated stallion spermatozoa were used for the experiment after swim-up for 20 min in sperm-Tyrode's albumen lactate pyruvate. Frozen-thawed spermatozoa from the same stallion were treated in a similar way. Spermatozoa were immobilized and injected into the oocytes using a Piezo drill. Presumptive zygotes were cultured in G1.2 medium for 20 or 96 h after the injection was administered, or were transferred to the oviducts of recipient mares and recovered 96 h later. In addition, bovine oocytes with first polar bodies were injected with the two types of stallion spermatozoa and fixed 20 h after injection to examine pronuclear formation. Fertilization rate (pronucleus formation and cleavage) at 20 h after injection of spermatozoa was not significantly different between fresh and frozen-thawed sperm groups in either equine or bovine oocytes. Pronucleus formation after injection of spermatozoa into bovine oocytes was significantly higher than that for equine oocytes (P < 0.05). There were no significant differences in cleavage rate or average number of nuclei at 96 h between equine oocytes injected with fresh or frozen-thawed spermatozoa. However, embryos developed in vivo for 96 h had a significantly higher number of nuclei in both sperm treatments compared with those cultured in vitro. These results indicate that good activation rates may be obtained after injection of either fresh or frozen-thawed equine spermatozoa without additional activation treatment. Injection of frozen-thawed equine spermatozoa results in similar embryo development to that obtained with fresh equine spermatozoa. In vitro culture of equine zygotes in G1.2 medium results in a similar cleavage rate but reduced number of cells compared with in vivo culture within the oviduct. Bovine oocytes may be useful as models for assessing sperm function in horses.

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