57 RELATIONSHIP BETWEEN FETAL ABNORMALITIES AND PERIPHERAL STEROID CONCENTRATIONS DURING GESTATION IN COWS TRANSFERRED WITH EMBRYOS PRODUCED BY SOMATIC CELL NUCLEAR TRANSFER

Abstract

Cloning of mammals by nuclear transfer frequently results in gestational or neonatal failure with a variety of abnormalities that are likely caused by inappropriate epigenetic reprogramming. Early diagnosis of fetal abnormality is important for efficient production of cloned animals. Sex steroids are produced in the bovine placenta and their levels in the blood might be useful as a measure of fetal well-being, as in humans. The objective of this study was to investigate whether changes in peripheral levels of progesterone and estrogens reflect fetal abnormalities. Donor cells for nuclear transfer were obtained from subculture of cumulus cells retrieved from ovarian follicles of a Japanese Black cow. Recipient oocytes were derived from ovaries obtained at an abattoir and matured in vitro. Metaphase II oocytes were enucleated and each fused with a donor cell by DC pulses. Nuclear transferred oocytes were activated and cultured for 7 days. Embryos developed to the blastocyst stage were transferred into the uterine horn ipsilateral to the ovary bearing the CL of 32 multiparous Japanese Black and Holstein crossbred cows at 7 to 8 days after the day of standing estrus (Day 0). Blood was collected from Day 40 until parturition. Progesterone and estrogens in the blood plasma of 6 recipient cows with full-term delivery were measured by RIA. These profiles were compared with each other and with the changes in a cow made pregnant by MOET. Statistical differences were analyzed with repeated measures ANOVA. Parturition was induced on Day 290. Stillborn, dead, or euthanized calves were subjected to necropsy and histopathological analysis. Pregnant cows were 14, 13, and 9 on Days 30, 60, and 90, respectively. Thereafter, 3 aborted around Days 110, 120, and 190. Six cows delivered calves weighing 45.8 ± 2.1 kg (mean ± SEM) on Days 291 to 293. Their birth weights were greater than those of female calves (31.2 ± 0.4 kg, n = 6) produced by MOET in the same breed. Three calves grew normally until weaning. One was stillborn due to dystocia, but no abnormalities were observed except for large offspring syndrome. One was euthanized 2 days after natural delivery due to ananastasia. Thymic and thyroid hypoplasia, left ventricular dysplasia, and pulmonary fibrosis were found by necropsy. Another calf delivered by Caesarean section died of infirmity 2 days later and had thymic and thyroid hypoplasia. Changes in plasma steroid concentrations were consistent with each other and with those in a MOET cow except the last one, in which progesterone levels tended to be higher during the first trimester and estrogens were lower during the last half of gestation. Progesterone levels tended to be lower in cows bearing a healthy calf than in cows bearing a weak calf before parturition. These results imply that peripheral steroid levels may reflect fetal normality, although large offspring syndrome does not affect their concentrations.