Electron Microscopic Studies of Mouse Oocytes and Two-cell Embryos exposed to Progesterone in Vitro

This experiment was undertaken in order to find out if there is any morphological change in oocytes and two-cell embryos whose development have been suppressed by progesterone for six hours in vitro. It can be observed that some part of the outer side of nuclear membrane of the suppressed oocytes was damaged. The number of nuclear pores has decreased in suppressed oocytes and this suggests that progesterone might suppress the transport of intermediary metabolites between cytoplasm and nucleus. Sometimes, closely packed aggregates of parallel or irregular endoplasmic reticula were observed in suppressed oocytes. Microvilli of suppresed oocytes showed signs of degradation and the perivitelline space became apparent. Thus it is presumed that the egg membrane has constricted during cultivation under progesterone in vitro. The other cell organelles such as mitochondria, multivesicular bodies, cortical granules and fibrillar lattices showed no difference in morphology between treated and control (intact) oocytes. In two-cell embryos, there was also no evident morphological change except for the fact that many vacuoles appeared clearly in suppressed embryonal cells. In brief, there was no fundamental morphological change in the oocytes and the embryonal cells exposed to progesterone for six hours even though it inhibits their development. The action of progesterone should be investigated thoroughly.

Urinary Estriol Determinations in Normal and Pathological Pregnancies

Estriol excreation was studied in 216 normal and 61 pathologic pregnancies. The 95% fiducial limits of the normal excretion of estriol, within which 95% out of 100 future determinations in normal pregnancies are expected to fall, were established. The estriol curve in normal pregnancy in this study agrees well in its general shape with those presented by previous investigators who used different chemical methods of determination. The estriol values in pathologic pregnancies with preeclampsia. intrauterine fetal death and antepartum hemorrage have been analyzed. The clinical significance of estriol determinations during pregnancy was discussed.

A Study of the Self Smear Method as a Screening Device for Uterine Cancer

The accuracy of Kato's self smear Kit was studied in screening a relatively unselected population group for cervical cancer. Routine cervicovaginal smears taken periodically would be the ideal method but would be unrealistic in view of the physician's time involved when considered on a large population scale. Three hundred patients were randomly selected at the Out Patient Clinic of Yonsei Medical Center and 26 cases grossly of cervical cancer were included in this group. We have here summarized the results briefly as follows. 1. Among 300 women examined in our OPD, 256 (85.3%) were classified as Pap. Class I or II, 9(0.3%) as a 'Defferred' group (Class II, Repeat), 18(6%) as Class III and 17(5.7%) as Class IV or V. 2. Fifteen of the total cases had insufficient material for evaluation (poor quality) and the major causes of the poor smears were vaginal douche within 24 hours, technical problems and cell degeneration. 3. The rate of agreement between the Self Smear Kit and cervicovaginal smear was as follows: a. The rate of complete agreement was 93.2% (280/300) b. The rate of complete or partial agreement was 96.7% (290/300). 4. The diagnostic accuracy of the Self Smear Kit was as follows: a. If we assumed the 'Deferred' group to be detection failures for uterine cancer, the diagnostic accuracy would be 82.4%. b. If we assumed the 'Deferred' group to be detection successes, the diagnostic accuracy would be 94.1%. 5. The quality of self smears was superior when taken by an aid nurse rather than by the patient herself. 6. Optimal time for good preservation with the Self Smear Kit prior to pap. stain under the following temperatures was proven experimentally to be as follows; a. At 5 degrees C: 3 days after smear b. At 15 degrees C: 3 days after smear c. At 30 degrees C: 2 days after smear

The Effect of Lecucocytes on the Maturation of Mouse Oocytes in vitro

This experiment was undertaken in order to know the effect of leucocytes on the maturation of mouse oocytes in vitro. Leucocytes obtained from heart puncture of mouse (3 X 10(4) cells/mm3) inhibited the maturation of mouse oocytes. The egg toxic activity declined with decreasing leucocyte concentration. It was found that egg toxic effect of leucocytes is not species specific. The activity of intact leucocytes or equal numbers of leucocytes that were destroyed was similar and which seems not to be influenced by the physiological stats of leucocytes.

Unruptured Cornual Pregnancy: A Case Report

Cornual pregnancy is very rare and its diagnosis is very difficult due to the distensibility of the uterus at the cornu. However, tender cystic enlargement at one horn of the uterus is suggestive of this type of ectopic gestation in this case. A case is reported of unruptured cornual pregnancy, diagnosed before operation, with a review of the literatures.

Localization of Placenta in Scanning by In113m Radiopharmaceuticals

Placenta previa is a common grave complication of late pregnancy, usually manifestated clinically by painless antenatal vaginal bleeding. Digital and rectal examinations are dangerous, due to the possibility that profuse hemorrhage from the vagina may result. Various radiological examinations have been performed in placenta previa for diagnosis and localization. However radioisotopic methods are superior due to safety, simplicity and a lower radiation dose, both fetal and maternal, compared to plain radiography. Among radiopharmaceuticals, In113m (transferrin for blood pool scan) is useful, giving more satisfactory results without any complications or untoward reactions. In our series of 88 cases from March 1971 to April 1975, In113m placental scan was performed and analysed in 62 cases which were confirmed by clinical follow up and the results are as follows: 1) Maternal age pattern. Mothers 31~35 years were 20/62 or 30.6%. 2) Maternal gravida pattern. All were multipara except 8 cases of primipara. 3) Gestational maturity on scan. 40/62 or 64% were before 36weeks or less in maturity. 4) On scan analysis Placenta previa was confirmed in 36/62 cases or 58%. 5) Fetal maturity. On delivery most were full term, 42/62 or 67.7%. 6) Vaginal delivery was done on cases where placenta was localized in the upper uterine segment, except for three who had fetal malpresentation and congenital anomaly of the maternal pelvis. All patients of placenta previa had cesarian section, except 5 cases with a minor degree of placenta previa. 7) Two cases showed false negative, which suggests 97% accuracy in the screening test of placenta previa by scan, wich is a similar result to other reports. Only 4 cases of false positive discrepancy were noted.

The in Vitro Maturation of the Mouse Oocyte

Mouse follicular oocytes, denuded and intact, were cultured in pyruvate salt sol and glutamine salt sol supplemented bovine serum albumin to compare the maturation rate. Glutamine has no effect on maturation of the denuded mouse oocyte but has an effect on maturation of the intact oocyte by increasing the maturation rate, depending on the increased concentration of glutamine (0.4 mM to 2 mM). Changes in osmolarity of the operation medium from 280 mOsm to 310 mOsm has no discernible effect on the oocyte maturation. A high frequency of abnormal 1st polar bodies was observed in pyruvate salt sol. and this may be due to the increased energy source in the cytoplasm of the 1st polar body when the po1ar body was extruded into the perivitelline space after the 1st meiosis.

Localization of Placenta in Scanning by In113m Radiopharmaceuticals

Placenta previa is a common grave complication of late pregnancy, usually manifestated clinically by painless antenatal vaginal bleeding. Digital and rectal examinations are dangerous, due to the possibility that profuse hemorrhage from the vagina may result. Various radiological examinations have been performed in placenta previa for diagnosis and localization. However radioisotopic methods are superior due to safety, simplicity and a lower radiation dose, both fetal and maternal, compared to plain radiography. Among radiopharmaceuticals, In113m (transferrin for blood pool scan) is useful, giving more satisfactory results without any complications or untoward reactions. In our series of 88 cases from March 1971 to April 1975, In113m placental scan was performed and analysed in 62 cases which were confirmed by clinical follow up and the results are as follows: 1) Maternal age pattern. Mothers 31~35 years were 20/62 or 30.6%. 2) Maternal gravida pattern. All were multipara except 8 cases of primipara. 3) Gestational maturity on scan. 40/62 or 64% were before 36weeks or less in maturity. 4) On scan analysis Placenta previa was confirmed in 36/62 cases or 58%. 5) Fetal maturity. On delivery most were full term, 42/62 or 67.7%. 6) Vaginal delivery was done on cases where placenta was localized in the upper uterine segment, except for three who had fetal malpresentation and congenital anomaly of the maternal pelvis. All patients of placenta previa had cesarian section, except 5 cases with a minor degree of placenta previa. 7) Two cases showed false negative, which suggests 97% accuracy in the screening test of placenta previa by scan, wich is a similar result to other reports. Only 4 cases of false positive discrepancy were noted.

The in Vitro Maturation of the Mouse Oocyte

Mouse follicular oocytes, denuded and intact, were cultured in pyruvate salt sol and glutamine salt sol supplemented bovine serum albumin to compare the maturation rate. Glutamine has no effect on maturation of the denuded mouse oocyte but has an effect on maturation of the intact oocyte by increasing the maturation rate, depending on the increased concentration of glutamine (0.4 mM to 2 mM). Changes in osmolarity of the operation medium from 280 mOsm to 310 mOsm has no discernible effect on the oocyte maturation. A high frequency of abnormal 1st polar bodies was observed in pyruvate salt sol. and this may be due to the increased energy source in the cytoplasm of the 1st polar body when the po1ar body was extruded into the perivitelline space after the 1st meiosis.

The Maturation in Vitro of The Rabbit Oocytes I. Amino Acids Support the Maturation In Vitro of the Rabbit Oocytes

Rabbit follicular oocytes were cultured in a medium supplemented with various elements such as bovine serum(RS), bovine serum albumin(BSA), amino acids and chorionic gonadotrophic hormone(HCG) in order to find which factors among them were most effective for oocyte maturation. The presence of BSA in the basic medium (modified Krebs-Ringer bicarbonate) did not elevate the proportion of oocyte maturation. When BS alone was added to the medium, only a few oocytes could reach to metaphase I and most of them were in degeneration. This implies that BS may act as an inhibitory or a toxic agent to the rabbit oocytes. It was found that the medium supplemented with 0.4% BSA and amino acids together raised the proportion of the oocyte maturation (54-62%). Especially the presence of proline, or of both proline and glutamine, gave a more favourable condition for the initiation of meiotic division than other amino acids. Addition of HCG to the medium did not promote the proportion of the oocyte maturation. As a consequence, it is apparent that amino acids in the medium are the most essential factors in inducing oocyte meiotic division.

Clinical Studies on Biometrics of the Placenta

With increasing concern for the study of the fetal aspects of pregnancy and child-birth the availability of normal standards becomes more and more important. It is very difficult to test the functional capacity of the placenta, and for this reason anatomic studies to determine the normal relationship between the placenta and the newborn infant have come to be of considerable importance. Three hundred seventy eight placentas randomly collected in the past three years and ranged of the twenty-fourth gestation week to term which were delivered at Severance Hospital of Yonsei Medical College. These were analysed statistically as to whether there was any correlation between the size of the placentas, weight of newborn, and age and parity of the mother and were studied to see if there was any clinical and statistical significance in the change of the placental coefficients. We have also tried to search for any relationship between the specific gravity and the size of the placenta and the newborn weight. In addition to these factors, clinical complications such as toxemia, anemia, intrauterine fetal death, giant baby and infarction of the placentas were included in this study to seek any relationship with the above mentioned factors. Another purpose was to obtain the proper values for Korean placentas. We have summarized this study briefly as follows. 1. The average weight of the 378 newborns was 3l68 gm and for the 340 normal cases it was 3270 gm. 2. The average weight of 378 placentas was 6l6 gm and for the 340 normal cases it was 621 gm. 3. The average volume of 378 placentas was 588 cc and for normal cases it was 596 cc. 4. The average placental dimensions were 17.9 X 14.2cm. 5. The average surface area of the 340 norma1 cases was 276 cm2. 6. The specific gravity of 340 normal cases was 1.0475 and there was no relationship to the gestation weeks and baby weight. 7. The average placental coefficient of 378 cases was 0.194 and for 340 normal cases it was 0.190 (varying from 0.100 to 0.333). The placental coefficients were greater in toxemia syphilis, intrauterine fetal death, prematurity twin pregnancy and immaturity. However it was lower in placental infarction and in relatively small placentas. In anemia and giant baby it was close to the normal average but in extremely large baby placenta coefficients was greater than Normal averages. (Baby wt. > 4500gm. ). 8. The weight of baby and the size of the placenta showed no demonstrable relationship to the age and parity of the mother. 9. statistically there was a high correlation between the weight of the infants and the weigh of the infants and the weight and volume of the placenta but somewhat low relationship to the surface area of the placenta.

The Effects of Follicular Fluid on in Vitro Maturation of Bovine Follicular Oocytes

The present experiments were conducted to evaluate the effects of follicular fluid on maturation of bovine follicular oocytes. TC medium 199 seemed to be in adequate for this purpose since a high proportion (ranging 84.1. to 97.0%) of the oocytes were able to resume meiotic division in both media-with or without the addition of follicular fluid. This implies a possible similarity between TC medium 199 and follicular fluid with regard to the components initiating meiosis. Actually, TC medium 199 contains amino acids, vitamins and carbohydrates most of which are also found in follicular fluid. For this reason, the effect of follicular fluid on the ovum maturation was investigated by applying Krebs-Ringer bicarbonate solution (SECM), which was main1y composed of the salts, pyruvate and lactate. When the oocytes were cultured in SECM without the addition of follicular fluid, only 7-14% of them resumed meiotic division within 30 hours. However, when follicular fluid was added, the proportion of oocytes undergoing maturation was sharply increased to about 70%. Among the groups cultured in media containing different concentrations of follicular fluid, the proportion of the oocytes in meiosis remained constant, In pure follicular fluid, 87-89% of the oocytes showed enhancement of meiotic division. The presence of the follicular fluid contributed to a decrease in the production of degenerative oocytes. As a consequence it has been noted that addition of follicular fluid to the culture medium provides a more beneficial environment for cow oocytes. Meiotic division is initiated and production of degenerative oocytes is inhibited.

Ultrastructural Studies on Mitochondria of Preimplantaion Rabbit Embryos

The ultrastructural changes of mitochondria in the ovarian oocytes from Graafian follicles, the ovulated tubal ova, and the various stages of preimplantation rabbit embryos have been observed with an electron microscope. From the ovarian oocytes to the 4-cell stage, mitochondria showed oval and round forms with a few cristae arranged concentrically and peripherally at the inner membrane. In 8-cell and 16-cell stages, mitochondria tended to change their forms to be elongated, and their sizes, and the outer membrane of the mitochondria had a tendency to become rough and irregular although there were few changes in the inner structure. In morula, some mitochondria began to show several transverse cristae proceeding into the matrix. Mitochondria rapidly increased in number at the late blastocyst stage. Matrix of mitochondria with transverse cristae found in the morula and in blastocyst stages was less dense than that of the earlier stages. The authors believe that the morphological changes of mitochondria during early embryonal development indicate the level of enzymatic activity at which this organelle is engaged in energy metabolism.

Volume Changes During the Preimplantation Stages of Mouse Egg Development

The mouse eggs in the various stages, of the development prior to implantation were collected and measurements were made on both the largest and smallest diameters of the vitellus, inner and outer surface of the zona pellucida. The various stages of development used were ovarian oocytes (germinal vesiA®e stage), ovulated but unfertilized egg, ovulated and fertilized egg, the 2-cell embryo on the second day of pregnancy, 4-8-cell embryo on the third day of pregnancy and morulablastocyst on the fourth day of pregnancy, A further comparative study on unfertilized and fertilized tubal eggs was made, The time of l2 hours after H.C.G. injection was chosen as the starting point from which to follow the collection of eggs every 3 hours for 24 hours. Since the volume gives a better comparison of size than diameter, the volume of the total eggs, intrazonal cavity, perivitelline space and the various were calculated for the various preimplantation stages of mouse egg. The volume of zona pellucida was also calsulated by subtraction of the volume of the inner zonal cavity from the volume of total egg and compared with the zona pellucida thickness. All calculations were made by computor(CEIR Time-sharing Computor). The diameter and volume of the vitellus in the ovarian oocyte is the largest one of any stage during the preimplantation stages of development, while the total volume of the entire egg as determined from the diameter of outer surface of the zona pellucida of the ovarian cocyte is the smallest one of any stage during development. The diameter and total volume of the entire egg increases from the ovarian oocytes to the first day of pregnancy and then gradually decreases until the third day of pregnancy. An increase in these parameters again takes place on the fourth day of pregnancy. The zona pellucida of the tubal ova is thicker than that of the oocyte, with the zona pellucida of the fertilized egg being definitely thinner when compared with unfertilized eggs. This phenomenon of decreased thickness in fertilized egg may be associated with zona reaction. The perivitelline space between the vitellus and zona pellucida thus formed following ovulation occupied approximately 40 percent of the total volume enclosed by the inner surface of the zona pellucida (intrazonal cavity) in the 1-cell tubal ova. Neither the cause of the rapid accumulation of fluid after ovulation which resulted in the production of the perivitelline space nor the actual time of the formation of the perivitelline space are known. Some possible reasons for the formation or origin of the perivitelline space are discussed. The size and shape of the vitellus undergo compartive reduction during preimplantation stages of development. The possible reason for the reduction of vitelline volume are discussed.

The Thickness and Volume Change of the Zona Pellucida Following Ovulation and Fertilization

Since the volume gives a better comparison of size than thickness, the volume of the zona pellucida was also calcldated by substraction of the volume of the inner zonal cavity from the volume of the total egg and compared with the zona pellucida thickness. All calculations were made by computor (CEIR timesharing computor). The zona pellucida of the tubal ova is thicker than that of the oocyte, with the zona pellucida of the fertilized egg being definite1y thinner when compared with unfertilized eggs. This phenomenon of decreased thickness in the fertilized egg may be associated with the zona reaction. The entry of the first sperm into the egg initiates a reaction in the zona pellucida the effect of which is to predude the entry of additional sperm-Braden, Austin and David (1954) defined this phenomenon as the zona reaction. The results of anatomical changes in the zona pe1lucida may also be a phenomenon such as a "zona reaction".