Morphological studies of polycystic mouse ovaries induced by dehydroepiandrosterone

Summary Morphological alterations induced by dehydroepiandrosterone (DHA) were studied in polycystic mouse ovaries (PCO). Treated mice showed ovulatory failure and cystic changes; cysts and follicles in various stages of growth and atresia were present although corpora lutea were absent. The levels of testosterone, dihydrotestosterone, 3- and 3-androstanediol, estrone and androstenedione increased, whereas estradiol was not detectable.The ultrastructure of granulosa cells in healthy and atretic follicles was similar to that of control animals, although the membrana granulosa in cysts was reduced to a monolayer of flattened cells. The theca interna of healthy and atretic follicles and ovarian cysts showed ultrastructural signs of abnormal steroidogenic stimulation.No significant differences (0.7<P<0.8) were found between the extensive surface area of gap junctions of healthy follicles of control and DHA-treated animals. On the P-face of granulosa cells of large healthy follicles, meandering strands of tight junctional particles were observed; their average length was significantly longer than those in healthy follicles of control animals (P<0.001). This increase was probably related to the large amounts of androgens present in the treated animals.Theca interna cells possessed small gap junctions; no significant differences (P>0.9) in gap-junction surface area were observed between DHA-treated and control animals. These results suggest that the size of gap junctions is probably unrelated to the steroidogenic activities of theca cells.The following trivial names have been used: Dihydrotestosterone: 5-androstan 17 ol-13 one; 3-androstanediol: 5-androstan 3,17 diol; 3-androstanediol: 5-androstan 3,17 diol     

Secretion of 17 alpha-hydroxyprogesterone, androstenedione, and estrogens by porcine granulosa and theca interna cells in culture

The steroid secreting activities of dispersed granulosa and theca interna cells from preovulatory follicles of prepubertal gilts 72 h after pregnant mare's serum gonadotropin treatment (750 IU) were compared. The cells were cultured for 24 h with or without steroid substrate (10(-8) to 10(-5) M progesterone, 17 alpha-hydroxyprogesterone, or androstenedione), FSH (100 ng/mL), LH (100 ng/mL), and cyanoketone (0.25 microM, an inhibitor of 3 beta-hydroxysteroid dehydrogenase). Granulosa cells cultured alone secreted mainly progesterone. Theca interna cells secreted mainly 17 alpha-hydroxyprogesterone and androstenedione, with secretion being markedly enhanced by LH. In the presence of cyanoketone, which inhibited endogenous progesterone production, theca interna but not granulosa cells were able to convert exogenous progesterone to 17 alpha-hydroxyprogesterone and androstenedione, and exogenous 17 alpha-hydroxyprogesterone to androstenedione and estradiol-17 beta in high yield. The secretion of the latter steroids from exogenous substrates was unaffected by LH. Theca interna cells secreted more estradiol-17 beta than did granulosa cells in the absence of aromatizable substrate, but estradiol-17 beta secretion by the latter was markedly increased after the addition of androstenedione. These apparent differences in steroid secreting activity between the cell types suggest that the enzymes responsible for conversion of C21 to C19 steroids, i.e., 17 alpha-hydroxylase and C17,20-lyase, reside principally in the theca interna cells. However, aromatase activity appears to be much higher in granulosa cells.     

Das histochemische Verhalten der Azolesterasen sowie weiterer Oxydoreduktasen bei der Follikelatresie im Ovarium einiger Nager

Zusammenfassung Histochemische Untersuchungen am Ratten-, Mäuse- und Meerschweinchenovarium ergaben ein unterschiedliches Verhalten der Azolesterasen (= Cholinesterase und Acetylcholinesterase) in Granulosa, Theca und Thecaorgan. Die Differenzierung der genannten Fermente wurde mit Hilfe von DFP, iso-OMPA und 284 C 51 durchgeführt. Die -Hydroxybuttersäuredehydrogenase gleicht in der Verteilung weitgehend der NADH-Cytochrom-c-Reduktase, zeigt aber quantitative Speziesunterschiede. Bei der -Ketoglutaratoxydase ist im Ratten- und Mäuseovarium erst beim atresierenden Follikel eine deutliche Aktivität der Granulosa und eine Zunahme in der Theca externa feststellbar, während im Meerschweinchenovarium die Theca interna immer, die Granulosa aber nur beim Abbau reagiert.

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The histochemical behaviour of the azolesterases and other oxydoreductases during atresia of follicles in the ovaries of some rodents

Summary Histochemical investigations on the ovaries of rats, mice and guinea pigs revealed a variable behaviour of the azolesterases (= cholinesterase and acetylcholinesterase) in granulosa, theca and theca organ. The differentiation of these enzymes was carried out with DFP, iso-OMPA and 284 C 51. The distribution of -hydroxybutyrate dehydrogenase closely parallels that of NADH-cytochrome-c-reductase but shows quantitative variations in the different species. In the ovaries of rats and mice -ketoglutarate oxidase shows a marked activity only in atretic follicles and an increased activity in the theca externa. In guinea pig ovaries, however, the theca interna always reacts, but the granulosa only does so in process of decomposing.

     

Localization of type 5 17β-hydroxysteroid dehydrogenase mRNA in mouse tissues as studied by in situ hybridization

The mouse enzyme type 5 17-hydroxysteroid dehydrogenase (17-HSD) catalyzes the conversion of androstenedione to testosterone and, to a lesser degree, the conversion of estrone to estradiol. In order to determine the exact sites of action of type 5 17-HSD, we studied the cellular localization of the mRNA of the enzyme in mouse tissues by using in situ hybridization. Specific hybridization signal was found in the liver, ovary, adrenal cortex, and kidney. In the liver of mice of both sexes, a strong signal was observed in all hepatocytes. In the ovary, specific labeling was detected in the granulosa and theca interna cells in growing follicles and in luteal cells. In the female adrenal cortex, intense labeling was restricted to the zona reticularis, whereas no type 5 17-HSD mRNA expression could be found in the male adrenal cortex. In the kidney of mice of both sexes, type 5 17-HSD mRNA was expressed in epithelial cells in both the proximal and distal convoluted tubules. The data indicate that androgens and estrogens are formed via the action of type 5 17-HSD in specific cell types in the liver, ovary, adrenal cortex, and kidney.This work was supported by Genome Canada and Genome Québec.     

Steroid synthesizing cellular sites in the ovary of the domestic pigeon Columba livia (Gmelin): a histochemical study.

Synopsis The ovary of the domestic pigeon,Columba livia, has been assayed histochemically for the localization of ⁵-3-hydroxysteroid dehydrogenase (⁵-3-HSDH), 17-hydroxysteroid dehydrogenase (17-HSDH), 11-hydroxysteroid dehydrogenase (11-HSDH), glucose-6-phosphate dehydrogenase (G6P-DH) and NADH-diaphorase activities during different periods of the reproductive cycle. ⁵-3-HSDH, 17-HSDH, 11-HSDH, G6P-DH and NADH-diaphorase activity was found in the theca interna of growing, atretic and postovulatory follicles, the granulosa of ovulatory, atretic and postovulatory follicles, and interstitial gland cells during the pre-incubation and the laying periods. During the incubation and squab feeding periods only ⁵-3-HSDH, G6P-DH and NADH-diaphorase activities were observed in the above mentioned cells. The steroidogenic potential of atretic follicles depends upon the type of atresia a follicle undergoes.     

Histochemical demonstration of Δ5-3 ß-OHD activity in the granulosa cells of ovarian follicles of immature and mature mice correlated with some ultrastructural observations

Summary A systematic study of ⁵-3-hydroxysteroid dehydrogenase in the granulosa cells of immature and mature mice was made. The histochemical results were compared with the ultrastructural findings on the same cells in an attempt to determine whether the granulosa cells are capable of a steroidogenic role.In newborn and immature mice the granulosa cells of a great amount of follicles demonstrated a moderate or strong histochemical activity. In mature mice the granulosa cells demonstrated a weak or moderate activity normally only in preovulatory follicles and in some other atretic follicles. The granulosa cells of the normal developing follicles did not show such activity. In addition the histological control of numerous parallel sections demonstrated particularly in immature ovaries the presence of a great amount of atretic follicles.In the cytoplasm of the granulosa cells of the follicles in immature ovaries only clusters of lipid droplets with ribosomes were noted; while in the preovulatory follicles of mature animals there started to appear mitochondria with tubular cristae, smooth membranes of the endoplasmic reticulum and irregular lipid droplets. In the obviously atretic follicles several granulosa cells as well as theca interna cells showed numerous lipid droplets and ribosomes together with different degenerating organelles. The granulosa cells of the normal developing follicles showed a well developed Golgi complex, granular endoplasmic reticulum and free ribosomes.The histochemical and ultrastructural findings suggest a steroidogenic role of the granulosa cells only in the larger preovulatory follicles (probably related to an early luteinization of this layer) but this role was not demonstrated in the same cells in normal developing follicles.In addition, since an histochemical positivity was demonstrated also in the granulosa cells of some obviously atretic follicles, it is possible that many of the follicles having granulosa cells filled with lipid droplets and attached ribosomes and histochemically positive might be, in the immature ovaries, in a very precocious stage of atresia.It is to precise for these cells whether a cytoplasm with these two strictly correlated components (lipid droplets and attached ribosomes) and showing an histochemical positivity could carry-on all the biochemical steps involved in steroid biosynthesis or only has only a temporary capability to produce some precursors of steroids.The present results were partially presented to the 56^ème Congrès de l'Association des Anatomistes (Nantes, 4–8/4/1971) and to the 66° Verhandlungen der Anatomischen Gesellschaft (Zagreb, 2/4/1971).     

Follicle growth in the ovary of the rabbit after ovulation-inducing application of human chorionic gonadotropin

Summary The growth of tertiary follicles, i.e., the proliferation of cells in the stratum granulosum and in the capillary network of the theca interna, after injection of ovulation-inducing human chorionic gonadotropin (HCG), was investigated in the rabbit by means of autoradiographic and morphometric methods.Based on the frequency distribution of follicles with different sizes and on the labeling index (LI) of granulosa cells as a function of follicle size and of time prior to and after HCG stimulation, two groups of tertiary tollicles can be distinguished: growing (250–900 m in diameter) and mature (>900 m in diameter) elements. The growth of both groups is influenced by the release of gonadotropins.After HCG stimulation, follicles belonging to the first group grow rapidly. During, and a short time after ovulation, almost all non-ruptured follicles larger than 600 m in diameter become atretic. Within 35–50 h the ruptured and atretic mature follicles (>900 m in diameter) are replaced by follicles out of the group of growing follicles.From these results the following concept for regulation of follicle growth is derived: In principle, all growing follicles possess the potential to develop into mature follicles. When a sufficient number of mature follicles is generated, these mature follicles determine the number of succeeding growing follicles. Follicles that are not required for providing mature follicles become atretic as soon as they reach a diameter of 700 m. When the majority of mature follicles is lost during ovulation (by rupture or atresia), this inhibition regulated by mature follicles is abolished, and all of the growing follicles again are capable to develop into mature follicles.The relative amount of capillaries in the theca interna of growing and mature follicles remains constant with increasing follicle size. This means that the capillary network grows parallel to the increasing size of follicles. No differences are found between intact and atretic follicles; advanced atretic follicles were excluded from this study.The labeling index (LI) of granulosa cells in the stratum granulosum and of endothelial cells in the theca interna, as a function of follicle size and of time after HCG stimulation, are closely correlated. A change in the LI of granulosa cells is usually followed with a certain delay by a similar alteration of the LI of endothelial cells in the theca interna. This suggests that granulosa cells have a certain regulatory function on capillary growth.     

Deleterious effects of androstenedione on growth and cell morphology of Schizosaccharomyces pombe

Androgens (androstenedione and testosterone) belong to the most important compounds in human steroidogenesis. The 17-hydroxysteroid dehydrogenase responsible for interconversion of the oxygenic group on C-17 of androgens ring is involved in steroid hormone synthesis. The fission yeast Schizosaccharomyces pombe 972 h^- was found to contain constitutive 17-hydroxysteroid dehydrogenase that was able to reduce androstenedione to testosterone and oxidize testosterone to androstenedione. The reductive pathway was found to be predominant while the oxidative one was carried out with much lower activity. Exogenous androstenedione, contrary to testosterone, inhibited S. pombe growth and stimulated the formation of aberrant swollen cells with slighter cell wall sensitivity to the action of the lytic enzyme Novozym. It is postulated that the 17hydroxysteroid dehydrogenase prevents the deleterious effects of androstenedione on the morphology and growth of the yeast's cells by androstedione reduction to testosterone.     

Role of mitogen-activated protein kinase pathway in reactive oxygen species-mediated endothelin-1-induced β-myosin heavy chain gene expression and cardiomyocyte hypertrophy

Endothelin-1 (ET-1) has been found to increase cardiac -myosin heavy chain (-MyHC) gene expression and induce hypertrophy in cardiomyocytes. ET-1 has been demonstrated to increase intracellular reactive oxygen species (ROS) in cardiomyocytes. The exact molecular mechanism by which ROS regulate ET-1-induced -MyHC gene expression and hypertrophy in cardiomyocytes, however, has not yet been fully described. We aim to elucidate the molecular regulatory mechanism of ROS on ET-1-induced -MyHC gene expression and hypertrophic signaling in neonatal rat cardiomyocytes. Following stimulation with ET-1, cultured neonatal rat cardiomyocytes were examined for ³H-leucine incorporation and -MyHC promoter activities. The effects of antioxidant pretreatment on ET-1-induced cardiac hypertrophy and mitogen-activated protein kinase (MAPKs) phosphorylation were studied to elucidate the redox-sensitive pathway in cardiomyocyte hypertrophy and -MyHC gene expression. ET-1 increased ³H-leucine incorporation and -MyHC promoter activities, which were blocked by the specific ET_A receptor antagonist BQ-485. Antioxidants significantly reduced ET-1-induced ³H-leucine incorporation, -MyHC gene promoter activities and MAPK (extracellular signal-regulated kinase, p38, and c-Jun NH₂ -terminal kinase) phosphorylation. Both PD98059 and SB203580 inhibited ET-1-increased ³H-leucine incorporation and -MyHC promoter activities. Co-transfection of the dominant negative mutant of Ras, Raf, and MEK1 decreased the ET-1-induced -MyHC promoter activities, suggesting that the Ras-Raf-MAPK pathway is required for ET-1 action. Truncation analysis of the -MyHC gene promoter showed that the activator protein-2 (AP-2)/specificity protein-1 (SP-1) binding site(s) were(was) important cis-element(s) in ET-1-induced -MyHC gene expression. Moreover, ET-1-induced AP-2 and SP-1 binding activities were also inhibited by antioxidant. These data demonstrate the involvement of ROS in ET-1-induced hypertrophic responses and -MyHC expression. ROS mediate ET-1-induced activation of MAPK pathways, which culminates in hypertrophic responses and -MyHC expression. Tzu-Hurng Cheng, Neng-Lang Shih: These authors have equally contributed to this work     

β-Adrenoceptors in the Tree Shrew Brain. I. Distribution and Characterization of [125I]Iodocyanopindolol Binding Sites

1. The number and distribution pattern of -adrenergic receptors in the brain have been reported to be species specific. The aim of the present study was to describe binding of the -adrenoceptor ligand [¹²⁵I]iodocyanopindolol in the brain of the tree shrew (Tupaia belangeri), a species which provides an appropriate model for studies of psychosocial stress and its consequences on central nervous processes.2. ¹²⁵I-Iodocyanopindolol (¹²⁵ICYP) labeling revealed a high degree of nonspecific binding, which was due mainly to interactions of this ligand with serotonin binding sites. For a quantitative evaluation of ₁- and ₂-adrenoceptors, serotonin binding sites had to be blocked by 100 M 5HT.3. Binding of the radioligand to ₁- and ₂-adrenoceptors was characterized using the ₁-specific antagonist CGP20712A and the ₂-specific antagonist ICI118.551. ₁-adrenoceptor binding is present in the whole brain, revealing low receptor numbers in most brain regions (up to 1.5 to 2.7 fmol/mg). A slight enrichment was observed in cortical areas (lateral orbital cortex: 4.0±0.7 fmol/mg) and in the cerebellar molecular layer (8.7±1.0 fmol/mg).4. Competition experiments demonstrated high- and low-affinity binding sites with considerable variations in K _i values for CGP20712A, showing that various affinity states of ₁-adrenoceptors are present in the brain (K _i: 0.61 nM to 67.1 M). In the hippocampus, only low-affinity ₁-adrenoceptors were detected (K _i: 1.3±0.2 M). Since it is known that ¹²⁵ICYP labels not only membrane bound but also internalized -adrenoceptors, it can be assumed that the large population of the low-affinity sites represents internalized receptors which may be abundant due to a high sequestration rate.5. High numbers of ₂-adrenoceptors are present in only a few brain structures of tree shrews (external layer of the olfactory bulb, 15.8±2.0 fmol/mg; claustrum, 19.3±1.5 fmol/mg; anteroventral thalamic nucleus, 19.4±1.5 fmol/mg; cerebellar molecular layer, 55.0±4.3 fmol/mg). Also for this class of -adrenoceptors, high- and low-affinity binding sites for the ₂-selective antagonist ICI118.551 were observed, indicating that ¹²⁵ICYP labels membrane bound and internalized ₂-adrenoceptors. Only in the cerebellar molecular layer was a high percentage of high-affinity ₂-adrenoceptors detected (K _i for ICI118.551 was 1.8±0.3 nM for 90% of the receptors).6. In conclusion, ₁- and ₂-adrenoceptor binding can be localized and quantified by in vitro receptor autoradiography in the brains of tree shrews when serotonergic binding sites are blocked. Modulatory effects of long-term psychosocial conflict on the central nervous -adrenoceptor system in male tree shrews are described in the following paper.     

Theca interna: the other side of bovine follicular atresia

Currently, histological classifications of ovarian follicular atresia are almost exclusively based on the morphology of the membrana granulosa without reference to the theca interna. Atresia in the bovine small antral ovarian follicle has been redefined into antral or basal atresia where cell death commences initially within antral or basal regions of the membrana granulosa, respectively. To examine cell death in the theca interna in the two types of atretic follicles, bovine ovaries were collected and processed for immunohistochemistry and light microscopy. Follicles were classified as healthy, antral atretic, or basal atretic. Follicle diameter was recorded and sections stained with lectin from Bandeiraea simplicifolia to identify endothelial cells or with an antibody to cytochrome P450 cholesterol side-chain cleavage to identify steroidogenic cells and combined with TUNEL labeling to identify dead cells. The numerical density of steroidogenic cells within the theca interna was significantly reduced (P < 0.001) in basal atretic follicles in comparison with other follicles. Cell death was greater in both endothelial cells (P < 0.05) and steroidogenic cells (P < 0.01) of the theca interna of basal atretic follicles compared with healthy and antral atretic follicles. Thus, we conclude that the theca interna is susceptible to cell death early in atresia, particularly in basal atretic follicles.     

The in vivo effects of tetrahydrocannabinol on the activity of lactate and succinate dehydrogenases in rat preovulatory follicles

Summary The effects of ⁹-tetrahydrocannabinol on the activities of lactate and succinate dehydrogenases in the theca interna and membrana granulosa of rat preovulatory follicles have been analysed microdensitometrically using the same injection regime employed in a previous study on steroidogenic enzymes. A small but statistically significant (18%) decrease in succinate dehydrogenase activity was observed in the theca interna, but none in any region of the membrana granulosa. Lactate dehydrogenase activity was unaffected by THC administration. Thus, a dosage and regimen sufficient to cause significant decreases in the activities of steroidogenic enzymes had little effect on succinate and lactate dehydrogenases in rat preovulatory follicles.     

3α-Hydroxysteroid dehydrogenase and 3β-hydroxysteroid dehydrogenase in the ovary of young Mongolian gerbils

Summary The ovaries of sexually mature, pregnant mare serum gonadotropin (PMSG) stimulated, 12 week old Mongolian gerbils were investigated morphologically and enzyme histochemically for the appearance of the 3-hydroxysteroid and the 3-hydroxysteroid dehydrogenase during the estrous cycle. Up to ovulation, on day 3 of the estrous cycle, the number of vesicular follicles increases continuously. Primarily atretic follicles can be seen on day 4. On day 5 corpora lutea appear, but they degenerate already by day 6.During the entire estrous cycle, 3-hydroxysteroid dehydrogenase and 3-hydroxysteroid dehydrogenase activity can be found in the theca of tertiary follicles and in the interstitial cells, whereas the theca of secondary follicles and the granulosa of healthy follicles do not exhibit any enzyme activity. The activity decreases from day 1 till day 6. The granulosa of atretic follicles and the cells of corpora lutea show only weak activity. It may be significant that the intensity of enzyme activity in the ovary and the estrogen level in the plasma are differently correlated to the estrous cycle.This investigation was supported by the Deutsche Forschungsgemeinschaft     

Bacillus subtilis bacteriophages SP beta c1 is a deletion mutant of SP beta

Summary The restriction fragment patterns of two mutant forms of the temperate Bacillus subtilis bacteriophage SP have been examined. The DNA of a heat-inducible mutant, SPc2, which has a molecular size of 128 kilobases (kb), yields the same restriction pattern as the wild type SPc⁺ DNA. The DNA of a clear-plaque mutant, SPc1, has a molecular size of 117 kb, and is deleted for an 11 kb region of phage DNA. Neither SPc1 nor SPc2 DNA is cleaved by the endonuclease HaeIII.     

The role of angiotensin II,endothelin-1 and transforming growth factor-β as autocrine/paracrine mediators of stretch-induced cardiomyocyte hypertrophy

Cardiac hypertrophy is a compensatory response of myocardial tissue upon increased mechanical load. Of the mechanical factors, stretch is rapidly followed by hypertrophic responses. We tried to elucidate the role of angiotensin II (AII), endothelin-1 (ET-1) and transforming growth factor- (TGF-) as autocrine/paracrine mediators of stretch-induced cardiomyocyte hypertrophy. We collected conditioned medium (CM) from stretched cardiomyocytes and from other stretched cardiac cells, such as cardiac fibroblasts, endothelial cells and vascular smooth muscle cells (VSMCs). These CMs were administered to stationary cardiomyocytes with or without an AII type 1 (AT₁) receptor antagonist (losartan), an ET-1 type A (ET_A) receptor antagonist (BQ610), or anti-TGF- antibodies. By measuring the mRNA levels of the proto-oncogene c-fos and the hypertrophy marker gene atrial natriuretic peptide (ANP), the molecular phenotype of the CM-treated stationary cardiomyocytes was characterized.Our results showed that c-fos and ANP expression in stationary cardiomyocytes was increased by AII release from cardiomyocytes that had been stretched for 60 min. Stretched cardiomyocytes, cardiac fibroblasts and endothelial cells released ET-1 which led to increased c-fos and ANP expression in stationary cardiomyocytes. ET-1 released by stretched VSMCs, and TGF- released by stretched cardiac fibroblasts and endothelial cells, appeared to be paracrine mediators of ANP expression in stationary cardiomyocytes.These results indicate that AII, ET-1 and TGF- (released by cardiac and vascular cell types) act as autocrine/paracrine mediators of stretch-induced cardiomyocyte hypertrophy. Therefore, it is likely that in stretched myocardium the cardiomyocytes, cardiac fibroblasts, endothelial cells and VSMCs take part in intercellular interactions contributing to cardiomyocyte hypertrophy.     

Correlation of plasma estradiol-17β and progesterone levels with ultrastructure and histochemistry of ovarian follicles in the white-spotted char,Salvelinus leucomaenis

Summary Plasma estradiol-17 and progesterone profiles were correlated with morphological changes in ovarian follicles during the preovulatory and postovulatory periods in the white-spotted char, Salvelinus leucomaenis. Plasma estradiol levels were highest in September, and were followed by a sharp drop in October; they remained very low throughout the postovulatory period. There was a good correlation between plasma estradiol levels and the gonadosomatic index, thus suggesting that estradiol is responsible for the synthesis of vitellogenic proteins. Plasma progesterone levels were very low in August, began to rise in September and reached a peak soon after ovulation; progesterone remained high for several days after ovulation. A preovulatory rise in plasma progesterone levels was recorded, and this is discussed in relation to the induction of oocyte maturation.In the preovulatory follicles, neither granulosa cells nor special thecal cells (ST cells) showed ⁵, 3-hydroxysteroid dehydrogenase (3-HSD) activity. In the young postovulatory follicles, in contrast, the ST cells showed intense 3-HSD activity with extensive agranular endoplasmic reticulum and numerous large mitochondria, while granulosa cells did not show 3-HSD activity. These results strongly suggest that the ST cells are the major sites of progesterone synthesis during the postovulatory period.Nanae Fish Culture Experimental Station Contribution No 14     

Electron microscopy of cytodifferentiation and its subcellular steroidogenic sites in the theca cell of the human ovary

Summary The cytodifferentiation and subcellular steroidogenic sites in the theca cell of the human ovary during the follicular phase were investigated using the electron microscopic cytochemistry. Only fibroblast-like cells were seen around or near the primordial follicle. In the theca interna of the secondary and Graafian follicle however there were three different cell types: fibroblast-like cells, theca gland cells (steroid-secreting cells) and transitional cells (partially or incompletely differentiated theca cells). On the other hand the theca externa of these follicles consisted mainly of fibroblast-like cells. The hallmarks of the cytodifferentiation of the theca cells were: 1) the appearance of lipid droplets, 2) a structural change of the mitochondrial cristae from lamellar to tubular form and 3) the appearance and development of smooth endoplasmic reticulum. Reaction products of 3-hydroxysteroid ferricyanide reductase, indicating 3-hydroxysteroid dehydrogenase activity, were localized on tubular or lamellar cristae and inner membrane of the mitochondria, and on the membranes of smooth endoplasmic reticulum in the transitional cell as well as in the theca gland cell of the secondary and Graafian follicle. From these data, it is suggested that the transitional cell has a steroid-secreting activity and also plays an important role in follicular development in human reproduction.Supported by a grant from the Japanese Educational Ministry     

Effects of tetrahydrocannabinol on rat preovulatory follicles: a quantitative cytochemical analysis

Summary Using a microdensitometric histochemical assay, ⁵-3-hydroxysteroid dehydrogenase activity and glucose-6-phosphate dehydrogenase Types I and II hydrogen generation were measured in preovulatory follicles from normal rats, and in follicles from rats given tetrahydrocannabinol for three days prior to sacrifice. Hydroxysteroid dehydrogenase and Type I hydrogen generation are involved in steroidogenesis, whereas Type II hydrogen generation is involved with general cellular metabolism. All ovaries were removed on pro-oestrus, frozen, sectioned and the sections reacted with the appropriate media. Enzyme activity was measured in the theca and in three regions of the membrana granulosa: peripheral antral and corona radiata. Compared to control animals, the hydroxysteroid dehydrogenase activity was significantly reduced in all follicular regions in rats exposed to tetrahydrocannabinol. Type I hydrogen generation was significantly less in the theca and peripheral region of preovulatory follicles from rats given tetrahydrocannabinol, but the same in the antral region and corona radiata. In all follicular regions examined, Type II hydrogen generation was unchanged following tetrahydrocannabinol administration. Thus, only the enzymes specifically associated with follicular steroidogenesis were affected by administration of the drug.     

Enzymatic synthesis of lactic acid derivatives with emulsifying properties

Novozym 435 (50 g l^–1) catalyzed the synthesis of n-octyl--d-glucopyranoside lactate by transesterification between n-octyl--d-glucopyranoside (30 g l^–1) and ethyl lactate (100 g l^–1) in acetone. In 12 h, a molar yield of 87% n-octyl -d-glucopiranoside lactate was obtained at a overall conversion of 90%.