Tuyoshi Endo

Mature adipocyte-derived dedifferentiated fat cells can transdifferentiate into skeletal myocytes in vitro

We have previously reported the establishment of preadipocyte cell lines, termed dedifferentiated fat (DFAT) cells, from mature adipocytes of various animals. DFAT cells possess long-term viability and can redifferentiate into adipocytes both in vivo and in vitro. Furthermore, DFAT cells can transdifferentiate into osteoblasts and chondrocytes under appropriate culture conditions. However, it is unclear whether DFAT cells are capable of transdifferentiating into skeletal myocytes, which is common in the mesodermal lineage. Here, we show that DFAT cells can be induced to transdifferentiate into skeletal myocytes in vitro. Myogenic induction of DFAT cells resulted in the expression of MyoD and myogenin, followed by cell fusion and formation of multinucleated cells expressing sarcomeric myosin heavy chain. These results indicate that DFAT cells derived from mature adipocytes can transdifferentiate into skeletal myocytes in vitro.

Pregnancy by Means of Tubal Insemination and Subsequent Spontaneous Pregnancy in Rabbits

The purpose of the study was to determine whether or not physical stimulation by tubal insemination had any unfavourable influences upon the tubal fimbria. Tubal insemination was carried out on 14 rabbits and subsequent pregnancy results were monitored. After a period of 69 − 123 days following tubal insemination, 10 of the rabbits were mated spontaneously and these rabbits were then monitored for pregnancy. Newborn were obtained, with normal gestation periods in six out of the 14 rabbits, following tubal insemination and all 10 of the rabbits that were mated spontaneously with males following tubal insemination subsequently delivered. It is concluded that physical stimulation by tubal insemination does not produce adhesive changes on the tubal fimbria.

Inhibitory effects of indomethacin on implantation and its related phenomena.

The dose-response relationship for the inhibitory effect of indomethacin on implantation and continuance of pregnancy was examined in four groups of rabbits administered with indomethacin (2.5, 5.0, 7.5 and 10.0 mg/kg) during the implantation period and compared with a control group. Implanted fetuses and corpora lutea were counted by laparotomy, and the number of offspring born was noted. The inhibitory effect of indomethacin on implantation was found to be dose–dependent, and the birth rate decreased in the indomethacin groups compared with the control group. As a result, even where implantation had been achieved, death of the implanted fetuses occurred at a high rate in rabbits administered with indomethacin during the implantation period.

Effects of Splenectomy on Luteal Function in Pseudopregnant Rabbits

The involvement of the immune system in changes in luteal function was evaluated in rabbits. Pseudopregnancy was induced in 10 females and the spleens (considered to be the source of macrophages) of five were excised on day 7 of pseudopregnancy, while the five controls had sham splenectomies. Subsequent changes in serum progesterone concentrations were measured as an indicator of luteal function and luteolysis. A second pseudopregnancy was induced 31 days after splenectomy. The first pseudopregnancy was prolonged and during the second pseudopregnancy the serum progesterone concentrations on days 3 and 7 were much lower in the splenectomy group than in the control group. On day 14 of the second pseudopregnancy, the serum progesterone concentration markedly decreased in the control group while it remained almost at the level of day 7 in the splenectomy group. These results suggest that splenectomy suppresses the expression of luteal function and delays luteolysis in rabbits.

Effects of anti-progesterone compound RU486 on ovulation in immature mice treated with PMSG/hCG

The effects of RU486, a competitive antagonist of progesterone, on ovulation were studied in PMSG/hCG-treated immature mice. PMSG/hCG was administered to 3-week-old female mice to induce ovulation. A dose of 0.5 mg of RU486 was administered 3 h before hCG injection, simultaneously with hCG injection, or 3, 4.5, 6, 7.5 or 9 h after hCG administration. The lowest number of ovulated eggs was observed in the group given RU486 6 h after hCG injection. Although the mean number of ovulated eggs was only 4.4 in this group, it was increased to 25.7 by supplementary administration of 2.0 mg of progesterone. It was demonstrated that RU486 had inhibitory effects on ovulation, and the inhibition could be prevented by supplementary administration of progesterone.

Cleavage of unfertilized eggs after repeated administration of human chorionic gonadotrophin to hamsters.

The proportions of unfertilized eggs in the oviducts, showing abnormal cleavage, were examined in hamsters given single or repeated doses of 30 IU human gonadotrophic hormone for the induction of ovulation. In control animals (n = 7), 1.7% of the total ovulated eggs were morphologically abnormal unfertilized eggs showing cleavage. The proportions of unfertilized eggs that were abnormal in the groups of seven hamsters treated with one, two or three doses of the gonadotrophin were 20.4%, 19.4%, and 30.4%, respectively. The proportion of unfertilized eggs showing abnormal cleavage thus appeared to increase with repeated administrations of gonadotrophin.

Reduction of Hamster Egg Abnormalities in Repeatedly Induced Ovulation by Using the Minimum Effective Dose of Human Chorionic Gonadotrophin

The minimum dose of human chorionic gonadotrophin needed to induce ovulation was determined in hamsters, in which the luteinizing hormone surge was blocked by administration of phenobarbitone. Doses of 1.0, 2.5, 5.0 and 10.0 IU human chorionic gonadotrophin were injected subcutaneously, and 5.0 IU was found to be the minimum dose needed to induce ovulation in all of a group of seven hamsters. When this minimum dose was used to induce ovulation repeatedly, one, two or three times, in groups of seven hamsters the percentages of abnormal eggs seen were 5.3%, 5.6% and 6.4%, respectively. These results indicate that the marked increases in the proportions of abnormal eggs produced when ovulation is repeatedly induced, which have been observed in previous studies, can be prevented by using the minimum effective dose of human chorionic gonadotrophin.

Phagocytotic Activity of Macrophages Separated from Corpus Lutea of Pseudopregnant Rabbits

Macrophage activity as indicated by phagocytosis in the corpus luteum at the luteolysis stage was examined in rabbits during pseudopregnancy. The day of induction of pseudopregnancy was regarded as day 0, and the phagocytotic activity of macrophages obtained from luteal tissues was examined on days 7 and 16. When sheep red blood cells (SRBCs) were used to assess phagocytotic activity there was no difference in phagocytosis between days 7 and 16 but, when sensitized SRBCs were used to assess phagocytotic activity, the activity of macrophages on day 16 was more than double that on day 7 (P < 0.05). These results indicate that immunomediated phagocytosis of luteal macrophages is enhanced at the luteolysis stage (day 16). This phenomenon may contribute to the process of luteolysis.

Relation between the number of sperms to be deposited and pregnancy rate in tubal insemination in rabbits.

SummaryPregnancy rate of tubal insemination with semen diluted to concentrations of ×100 to ×107 was studied in rabbits. The animals were laparotomized under general anesthesia, and 0.05 ml of diluted semen was injected with a glass capillary from the tubal fimbria into the oviduct. The semen was obtained from mature male rabbits by means of an artificial vagina and diluted to concentrations between ×100 and ×107. One hour after the insemination, ovulation was induced by administration of hCG. Six experimental groups were set up according to the semen concentrations. In the ×100, ×102, ×104, and ×107 groups, newborn rabbits were obtained. The best pregnancy rate was obtained with the ×102 groups, in which 105 level of sperms were deposited. The newborn rabbits were normal not only morphologically, functionally, but also in chromosomal examination. The results suggested that tubal insemination with a small number of sperms could be applied in sterility due to oligospermia in human.

Superovulation inducement by administration of LH-RH preparation in mice

Abstract We attempted to induce superovulation in mice by using luteinizing hormone-releasing hormone (LH-RH) to develop follicles. Mice were treated with 5 I.U. of hCG at 17:00 h (Day 0), and then subcutaneously injected with 0.025 μg of an LH-RH analogue one to five times from Day 0 to Day 1. After a second injection of hCG at 17:00 h on Day 2, the oviducts were recovered on Day 3 and the ampulla were examined for ovulated ova. The mean number of ova for animals in the control group ( N =17) was 13.1, while those for all the experimental groups receiving LH-RH ranged from 15.4 to 24.1. The greatest number of ova (24.1) was obtained from the group which had received LH-RH five times at 22:00 h on Day 0, and at 02:00 h, 06:00 h, 10:00 h and 14:00 h on Day 1.