Biosynthesis of prostaglandins by tissues of the human fetus

Human fetal tissues have been superfused and prostaglandin (PG)E2, PGF2 alpha and 6-keto-PGF1 alpha have been measured in the effluents using specific radioimmunoassays. In general, the rates of production of 6-keto-PGF by the tissues studied were greater than the rates of production of PGF2 alpha which in turn were greater than the rates of production of PGE2.     

Effect of prostaglandins on the steroidogenesis in human ovarian tissues

Although prostaglandins are luteolytic in some species, in conditions they stimulate progesterone production in the corpus luteum (1). Apart from this effect prostaglandins may also stimulate other steps in the steroidogenic sequence e.g. corticosteroidogenesis in superfused rat adrenal glands (2) and aromatization of testosterone by perfused human placenta (3). With this possibility in view and also because of paucity of data on the effect of prostaglandins on steroidogenesis in human ovarian tissues we have been studying under conditions the effect of prostaglandins on progesterone formation in human corpora lutea and on the utilization of C₂₁ steroids by the luteal and follicular compartments of the ovary. These studies are still in progress. However, the data obtained so far indicates that in addition to stimulating progesterone synthesis in the corpus luteum prostaglandins may also affect other steps in steroidogenesis in human ovarian tissues. We wish to report here in brief these preliminary results.     

Effect of prostaglandins on the in vitro steroidogenesis in human ovarian tissues

This is a brief report of preliminary findings from in vitro studies of the effect of PGs (prostaglandins) on progesterone formation in human corpora lutea and on the utilization of C21 steroids by the luteal and follicular compartments of the ovary. Ovaries were obtained from cyclic women undergoing ovariectomies for therapeutic purposes. The laboratory procedures employed in the study are explained. Results are tabulated. PGE2 stimulated progesterone biosynthesis in the corpus luteum as measured by tissue content and by de novo synthesis from acetate-1-14C. PGE2 also stimulated the biosynthesis of DPS (digitonin-precipitable sterols) from acetate. These results confirm findings of other researchers. In 1 of the experiments, PGF2alpha failed to demonstrate stimulation of progesterone biosynthesis in the human corpus luteum as measured by tissue progesterone content after incubation. Both PGF2alpha and PGE2 showed generally stimulatory effects on the utilization of exogenous labelled progesterone for the formation of androgens and estradiol by the human corpus luteum. In the follicular tissue, however, PGE2 showed an inhibitory effect on the formation of androgens and progesterone from exogenous labelled pregnenolone. No significant amounts of estrogens were biosynthesized in these experiments. These preliminary results must be confirmed by measurement of the endogenous steroidal concentrations in the tissues.     

Biosynthesis of prostaglandins in ovarian follicles and corpus luteum of sheep ovary

The biosynthetic potential of prostaglandins (PGs) was measured in ovarian follicles and corpus luteum of sheep ovary. The total prostaglandins formed under non-enzymatic conditions were much lower in comparison to that formed using native GSTs. When the GSTs of ovarian follicles were employed, the major prostaglandin formed was PGE2 (81.22%) followed by PGD2 (16.9%) and PGF2 alpha (1.87%). In case of corpus luteum, prostaglandin formed was PGF2 alpha (59.01%). Since PGF2 alpha was demonstrated to be the luteolytic factor, the present study indicates the formation of luteolytic factor in the ovarian tissue itself.     

Biosynthesis and cellular localization of embryonal prealbumin in human embryonal tissues

Immunofluorescent analysis of chorion sections and various embryonal tissues revealed localization of embryonal prealbumin (EPA) in the mesenchymal cells of the chorion, bones, umbilical cord, skin and in the cytoplasm of the epithelium distal part of the embryonal kidney canals. Synthesis of EPA slow peak in the suspension tissue culture of the chorion, umbilical cord and bones was shown. It is suggested that EPA is a mesenchyme product which is actively synthesized during the period of embryonal development. EPA is resynthesized and localized in the cells of tumors originated from the connective tissue. Cells of the tumors of non-connective origin did not contain EPA, whereas it was detected in the cells of the adjacent connective tissue. The phenomenon of embryonal reversion, probably, takes place not only in the "original" tumor cells but also in the surrounding connective tissues.     

Biosynthesis and metabolism of prostaglandins in chick epiphyseal cartilage

Microsomal fractions of cells isolated from chick epiphyseal cartilage catalyzed the synthesis of prostaglandins from radiolabeled Δ^8,11,14-eicosatrienoic and from archidonic acids. In addition, the microsomal supernatants contained both 15-hydroxyprostaglandin dehydrogenase and prostaglandin 15-keto Δ^13,14-reductase activities. Two major classes of prostaglandins (E and F) were synthesized; however, a major product which chromatographically behaves as PGA was also isolated. Synthetase activities were analyzed for pH optima and response to known stimulators and inhibitors of prostaglandin synthesis. The different activators had varying stimulatory effects on prostaglandin synthesis; the anti-inflammatory drugs were all strongl inhibitory. Synthetase activity in the growth plate was highest in the zone of hypertrophy, declining substantially in the more heavily calcified regions. Degradative enzyme activities were highest in the zone of maturation and significantly lower in the adjacent hypertrophic zone. The net effect of these opposing activities would be to elevate prostaglandin levels at the zone of hypertrophy, a finding which suggests that prostaglandins may play a role in the modulation of epiphyseal cartillage metabolism.     

Effect of prostaglandins on the in vitro steroidogenesis in human ovarian tissues

Although prostaglandins are luteolytic in some species, in $\text{in vitro}$ conditions they stimulate progesterone production in the corpus luteum (1). Apart from this effect prostaglandins may also stimulate other steps in the steroidogenic sequence e.g. corticosteroidogenesis in superfused rat adrenal glands (2) and aromatization of testosterone by perfused human placenta (3). With this possibility in view and also because of paucity of data on the effect of prostaglandins on steroidogenesis in human ovarian tissues we have been studying under $\text{in vitro}$ conditions the effect of prostaglandins on progesterone formation in human corpora lutea and on the utilization of C₂₁ steroids by the luteal and follicular compartments of the ovary. These studies are still in progress. However, the data obtained so far indicates that in addition to stimulating progesterone synthesis in the corpus luteum prostaglandins may also affect other steps in steroidogenesis in human ovarian tissues. We wish to report here in brief these preliminary results.     

Formation of prostaglandins by ovarian carcinomas

Tissue contents of prostaglandins (PG) PGE2, PGE2a and 6-keto-PGF1a (degradation product of PGI2) were determined in specimens of advanced human ovarian cancer (n = 11). The PG levels (ng/mg tissue protein) varied widley: PGE2 17-515; PGF2a 2-43 and 6-keto-PGF1a 5-105. Tumors of patients without response to chemotherapy contained more PGE2, PGF2a and 6-keto-PGF1a than did tumors responding to chemotherapy. PG production was investigated in two ovarian carcinoma-derived cell lines. The ability of these cells to synthesize PG varied depending on the cell density. An increase of cell number was associated with a decrease of PG yield. PG formation was inhibited by indomethacin in a concentration-dependent manner. The present study suggests that ovarian carcinoma cells form PG in vivo and vitro.     

Biosynthesis of prostaglandins by human spermatozoa in vitro and their role in acrosome reaction and fertilization.

Five homogenates of human sperm cells were separately incubated with [14C]arachidonic acid in the presence of reduced glutathione, L-tryptophan, and haematin as cofactors. The cyclooxygenase products of arachidonic acid metabolism were extracted, separated, and measured for their radioactivity. The rate of formation of prostaglandin (PG)D2, PGE2, PGF2 alpha, 6-keto PGF1 alpha, and thromboxane (TX)B2 were 18.0 +/- 1.11, 10.9 +/- 0.68, 5.8 +/- 0.21, 3.9 +/- 0.13 and 6.6 +/- 0.52 pmol/10(6) cells/min, respectively. These results are discussed in relation to the hypothesis that cyclooxygenase metabolites of certain polyunsaturated fatty acids play an important part in the sperm acrosome reaction and fertilization.     

Biosynthesis and metabolism of prostaglandins in chick epiphyseal cartilage.

Microsomal fractions of cells isolated from chick epiphyseal cartilage catalyzed the synthesis of prostaglandins from radiolabeled delta8,11,14-eicosatrienoic and from arachidonic acids. In addition, the microsomal supernatants contained both 15-hydroxyprostaglandin dehydrogenase and prostaglandin 15-keto delta13,14-reductase activities. Two major classes of prostaglandins (E and F) were synthesized; however, a major product which chromatographically behaves as PGA was also isolated. Synthetase activities were analyzed for pH optima and response to known stimulators and inhibitors of prostaglandin systhesis. The different activators had varying stimulatory effects on prostaglandin synthesis; the anti-inflammatory drugs were all strongly inhibitory. Synthetase activity in the growth plate was highest in the zone of hypertrophy, declining substantially in the more heavily calcified regions. Degradative enzyme activities were highest in the zone of maturation and significantly lower in the adjacent hypertrophic zone. The net effect of these opposing activities would be to elevate prostaglandin levels at the zone of hypertrophy, a finding which suggests that prostaglandins may play a role in the modulation of epiphyseal cartilage metabolism.     

Analysis of prostaglandins formed from endogenous and exogenous arachidonic acid in homogenates of human reproductive tissues

Isotope-labelled arachidonic acid has been used to study in vitro formation of prostaglandins and other products in mammalian tissue. Quantitative conclusions about cyclooxygenase activity have been drawn from such studies. However, arachidonic acid is present in all tissues, free and esterified, and therefore it can be expected that endogenous arachidonate would interfere with transformation of the radioactive exogenous substrate. (1-14C)-labelled arachidonate was, therefore, incubated with homogenates of various human tissues (amnion, chrorion, placenta and myometrium), and the two molecular forms, 12C and 14C, of arachidonic acid as well as of prostaglandin E2 and prostaglandin F2 alpha were quantitated, before and after 30 min of incubation, using gas chromatography-mass spectrometry with multiple ion detection. The results demonstrate a substantial release of arachidonic acid into the medium during incubation. There was no correlation between either the initial concentration of [12C]arachidonic acid and initial concentration of [12C]prostaglandin E2 or the percent increase of those compounds during incubation. The net formation of [12C]prostaglandin E2 and [14C]prostaglandin E2 from endogenous and exogenous precursor, respectively, were also very different. The study shows that by simply incubating (1-14C)-labelled arachidonic acid in tissue homogenates and measuring the amount of radioactivity transformed into various prostaglandins only qualitative conclusions can be drawn.     

Biosynthesis of prostaglandins from arachidonic acid by the rat ovary

Biosynthesis of prostaglandins (PGs) from ¹⁴C-arachidonic acid was studied using homogenates of the ovaries from immature rats. In ascending order of metabolizing potency were, the ovaries from untreated rats, from rats treated with pregnant mare's serum gonadotropin (PMS), and from PMS-human chorionic gonadotropin (hCG) treated rats. Among the radioactive metabolites extracted, PGE₂ and 6-keto PGF_1∝ were purified and identified by silicic acid column-, thin layer-, reversed phase partition chromatographies, and radiogaschromatography. Production of PGE₂ and 6-keto PGF_1∝ was observed in homogenates of the ovaries of intact and PMS-hCG treated rats at conversion rates of 0.72; 0.43% and 7.62; 2.31%, but not by FMS treated rat ovaries. Treatment with PMS-hCG activated metabolism of arachidonic acid into radioactive metabolites including PGE2 and 6-keto PGF_1α to a large extent. Accordingly, it is concluded that luteinizing hormone and hCG play a significant role in the biosynthesis of PGs by the rat ovarian follicle.     

Preneoplastic changes in ovarian tissues

OBJECTIVE: To test whether histologically normal epithelium within ovarian inclusion cysts and stroma exhibit changes in nuclear chromatin pattern that indicate the presence of occult ovarian lesions. STUDY DESIGN: Ovaries were collected from 10 low-risk women,from 7 high-risk women and from 3 women with ovarian cancer. Histologic sections were cut at 5 microm and hematoxylin and eosin stained. High-resolution images were recorded from the epithelium lining inclusion cysts and from the underlying stroma of ovaries from these 20 subjects. A total of 2860 epithelial nuclei and 3610 stromal nuclei were recorded. Karyometric features and nuclear abnormality were computed. Discriminant analyses and unsupervised learning algorithms defined deviations from normal that were designated "above threshold" and used to compute average nuclear abnormality of a second nuclear phenotype. RESULTS: Histologically normal epithelium from inclusion cysts of ovaries harboring a malignant lesion was shown to exhibit changes in the nuclear chromatin pattern that were statistically significant using quantitative image analysis procedures. Similar changes were seen in the inclusion cyst epithelia of high-risk ovaries. A subpopulation of cells representing a new phenotype was detected in the underlying stroma of women harboring a malignant ovarian lesion and in women at high risk of ovarian cancer. CONCLUSION: The karyometric changes observed in the epithelia lining inclusion cysts and in the underlying stroma of ovaries either with ovarian cancer or at high risk of ovarian cancer suggest the presence of preneoplastic changes in histologically normal tissue.     

Immunohistochemical demonstration of prostaglandins in various tissues of the rat

To demonstrate the tissue localization of prostaglandin (PG) E₂, PGF₂ and 6-keto-PGF₁ (a stable metabolite of PGI₂) various tissues, including decalcified periodontal tissue of 7-week-old male Wistar strain rats, were immunohistochemically examined using a streptavidin-biotin complex method. Besides tissue macrophages and endothelial cells in various tissues, hepatocytes, renal tubular cells, and parietal and chief cells in the gastric mucosa showed a positive reaction for the various PGs examined. PGs were demonstrated in the cytoplasm or in association with the cell membrane. We generally observed no difference between the localization patterns of PGE₂-, PGF₂-, and 6-keto-PGF₁-positive cells in these tissues. However, in the periodontal ligament and alveolar bone, 6-keto-PGF₁ was localized in the cytoplasm of osteocytes, osteoblasts, cementocytes, and cementoblasts, while no reaction for PGE₂ or PGF₂ was revealed in these cells. We demonstrated the immunohistochemical localization of PGs in various rat tissues including decalcified periodontal tissue and discuss the important roles of PGs in the modulation of their normal functions in these tissues.