Ovarian Tissue Slices Research Articles

Preparation of progesterone secreting cells from rat ovaries by a density gradient

Isolation of progesterone secreting cells from the ovaries of immature rats pretreated with pregnant mare serum gonadotropin (PMS) was conducted by a simple procedure which combined the collagenase digestion with a density gradient method. After digestion of the ovarian tissue slices by the enzyme, the residue was gently pressed and placed on a sucrose density gradient. Three bands appearing in the tube after centrifugation were designated as S-1, S-2 and S-3 from the top to the bottom, respectively. The S-1 cells from the ovaries at 6 days after PMS secreted the greatest amount progesterone, i.e. approximately 430 ng per 10(5) cells during the 18th incubation. Progesterone secretion from the S-2 cells was less than 48% of that from the S-1 cells. A physiological interrelation between the S-1 and S-2 cells remains unexplained by the present experiment. The luteal cells were yellow, spheroidal and 15 to 40 mus in diameter. Many vesicle-like particles were found on the cell surfaces. Progesterone secretion from the prepared cells was stimulated significantly by hCG in vitro. This result indicates that progesterone secreting cells isolated by the collagenase-sucrose density gradient preserve their native function as luteal cells. The procedure for preparation of luteal cells in the present report may provide a suitable model for in vitro studies on the corpus luteum.

Effect of blastocysts on rat ovarian steroidogenesis in early pregnancy.

The biosynthesis of ovarian progesterone and 20alpha-hydroxypregn-4-en-3-one (20alpha-OHP) was studied in rats with pituitaries autotransplanted beneath the kidney capsule (APTr rats) 1 1/2 days after ovulation. Some animals underwent tubal ligation 1-2 weeks prior to mating. Incubated ovarian tissue slices showed an accumulation of progesterone of 64 plus or minus 11 mcg/g tissue, and a progesterone/20alpha-OHP ratio of .32 plus or minus .16. Ovarian tissue from APTr rats with viable blastocysts synthesized considerably more progesterone than tissue from tubal ligated APTr rats. The increased concentrations of progesterone were associated with decreased concentrations of 20alpha-OHP. The accumulation of progesterone in tissue from unmated APTr rats, APTr rats cervically stimulated at proestrus, and similar cervically stimulated rats with traumatized uteri was 42 plus or minus 5, 16 plus or minus 4 and 18 plus or minus 9 net mcg/g tissue, respectively. The progesterone accumulation in tissue from cervically stimulated rats was thus similar to that in tissue from tubal ligated animals. This inhibition by cervical stimulation was overcome by stimuli from the blastocysts. The results suggest that the blastocyst secretes a substance that is more active in promoting progesterone formation than pituitary prolactin.

Prostaglandins and [formula omitted] ovarian progestin biosynthesis

Prostaglandin F 2α (PGF 2α) did not alter the in vitro biosynthesis of progesterone by slices of luteinized rat ovaries when used in concentrations from 1 to 10,000 ng/ml of incubation medium; likewise, PGF 2α did not affect the incorporation of acetate-1- 14C into progestins. PGF 1α, 15-keto PGF 2α, and PGE 1 did not alter the biosynthesis of progesterone by luteinized rat ovaries; PGE 2 inhibited the production of progesterone when used at a concentration of 10 μg/ml, but not at lower doses. PGF 2α in combination with luteinizing hormone (LH) enhanced the metabolism of progesterone to 20α-hydroxypregn-4-en-3-one in luteinized rat ovaries. Interestingly, PGF 2α, at a high concentration of 10 μg/ml, did stimulate progesterone biosynthesis by slices of ovarian tissue from immature rats hormonally primed to simulate “pseudopregnancy,” suggesting a steroidogenic action of prostaglandins on the ovarian follicular or interstitial cell. PGF 2α (10 μg/ml) did not stimulate the in vitro biosynthesis of progesterone or 20α-hydroxypregn-4-en-3-one by slices of rabbit corpora lutea or rabbit ovarian interstitial tissue. It is concluded that prostaglandins do not stimulate progestin biosynthesis in rat luteal tissue.

Secretion of C19-steroids and oestrogens in the polycystic ovary syndrome. Ovarian studies in vivo and in vitro (including studies in vitro on a coincidental granulosa cell tumour).

SUMMARY Ovarian metabolism of C19-steroids and oestrogens has been assessed at ovarian wedge resection in 22 patients with polycystic ovaries. There were marked variations between different patients. High concentrations of androstenedione were found in ovarian vein plasma in some patients, and always after stimulation with human pituitary follicle-stimulating hormone (HP-FSH) in vivo. Its contribution to the daily production of testosterone has been considered. No measurable amounts of testosterone or dehydroepiandrosterone were found. Oestradiol concentration was sometimes normal or above. Large amounts of androstenedione were generally isolated from cyst fluid. Occasionally testosterone was found and also epitestosterone after FSH. That concentrations of oestrogens in cyst fluid are low was confirmed, but sometimes higher levels were found after FSH. Slices of ovarian tissue generally converted progesterone or pregnenolone to androstenedione in high yield but conversion to oestrogen appeared to be low. However, the difficulty of making quantitative comparisons by this method, in the absence of knowledge of the sizes of the pools of endogenous steroids in the tissue, has been recognized. No evidence was found in any of the 18 cases examined for a lack of 3β-hydroxysteroid dehydrogenase. In vitro synthesis of epitestosterone by both normal and polycystic ovaries has been observed. A coincidental granulosa cell tumour in one patient synthesized testosterone in high yield.

STEROID HORMONE FORMATION BY THE RAT OVARY.

ABSTRACT Ovaries were transplanted to the spleens of castrate male rats. After 120 days, slices of ovarian tissue, composed predominantly of corpora lutea, were incubated in Krebs-Ringer bicarbonate medium containing 50 μc acetate-1-14C. Radioactive steroid formation was assessed quantitatively by reverse isotope dilution. The formation of radioactive progesterone and 20α-hydroxy-pregn-4-en-3-one was established. The formation of radioactive 3β-hydroxy-pregn-5-en-20-one, androst-4-ene-3,17-dione, 17-hydroxyprogesterone, testosterone, oestrone and 17β-oestradiol could not be established. It appears that the corpus luteum of the rat, induced by endogenous gonadotrophins, forms only progestins from acetate-1-14C. Contrary to results previously obtained with ovarian tissue transplanted to female rats, radioactive steroid formation in vitro appeared to be augmented by luteinizing hormone (NIH-LH-S1) added to the incubation flasks. Administration of human chorionic gonadotrophin (200 IU/day) for 5 days prior to autopsy did not enhance acetate-1-14C incorporation in vitro.