Types Of Follicles Research Articles

Androgen and 19-norsteroid profiles in human preovulatory follicles from stimulated cycles: An isotope dilution-mass spectrometric study

Follicular fluid was aspirated from preovulatory follicles of women under ovarian stimulation for in vitro fertilization and analyzed by a highly specific technique based on gas chromatography-mass spectrometry associated with stable isotope dilution. 19-Nortestosterone and 19-noradrostenedione were identified and quantified for the first time in human follicular fluid. There was a strong positive correlation between 19-nortestosterone and estradiol-17β and between 19-norandrostenedione and estrone concentrations, thus indicating a common cellular origin. The accumulation of 19-norsteroids in follicular fluid confirms that they are weakly active intermediates in the multistep enzymatic conversion of androgen to estrogen. Testosterone concentrations were significantly lower than those obtained by radioimmunoassay; cross-reaction with substantially higher levels of 19-nortestosterone seems to be at the origin of this discrepancy. Androstenedione concentrations were similar to those reported in the literature and it was therefore confirmed that an estradiol/androstenedione concentration ratio above 20 is favourable for oocyte cleavage. Other and some newly estimated androgens are: testosterone sulfate, 5-androstene-3β,17β-diol 3-sulfate and disulfate, dihydrotestosterone sulfate, epitestosterone, 19-hydroxy-androstenedione, 5α-androstane-3α,17β-diol, 5α-androstane-3β,17β-diol, 5α-androstane-3,17-dione and androsterone. Dehydroepiandrosterone sulfate was by far the most abundant androgen in this type of follicles.

Effects of multiple LH injections on the secretion of estrogens from polycystic ovaries of androgen-sterilized rats.

Differences in the secretion of estrogens by follicular polycystic ovaries of androgen-sterilized rats and by normal follicular ovaries of early proestrous rats were compared. Some rats were injected i.v. with LH 30 min before bleeding. This injection of LH did not influence the secretion of estrogens by normal ovaries, but greatly increased that by polycystic ovaries, suggesting that there was abnormal steroidogenesis in cystic ovaries. In the ovaries of such androgen-sterilized rats, two types of enlarged abnormal follicles were seen. One of these was truly cystic with few or no granulosa cells (1st type). The other had a hyperplastic and infolded layer of granulosa cells with a papillary appearance (2nd type). Because it is known that the preovulatory LH surge is not found in androgen-sterilized rats, a classical approach was taken to circumvent the probable deficit in cyclic release of LH by giving an i.v. injection of LH every 4 days for 16 days, and ovarian venous blood was collected 4 days after the last injection. In consequence the 2nd type of abnormal follicle disappeared as did the abnormalities of estrogen production. These results suggest that the abnormalities of estrogen production by the polycystic ovaries of androgen-sterilized rats may be due to the 2nd type of abnormal follicle.

Changes in the kinetics of follicular growth in response to selection for large litter size in mice.

The present study examined a randomly selected control line (C) and a large litter size-selected line of mice (S1) to determine changes in the kinetics of follicular growth that have occurred in response to selection for large litter size. For each follicle type (FT), the number of healthy and atretic follicles, length of components of granulosa cell cycle, follicular growth rate, and follicular flux were determined microscopically from serially sectioned ovaries of Lines C and S1 mice. Selection for litter size significantly increased the number of small and medium, and some large follicle-size classes. While selection for litter size did not change the overall incidence of atresia at proestrus, it did decrease the incidence of atresia in the large Type 7 follicles by 19%. Selection for litter size also increased ovarian weight at proestrus. Selection for litter size increased the rate of growth through FT 3a, 5a, and 5b, and reduced the time required for follicles to grow from primordial to Graafian follicles from 39.1 days in Line C to 33.4 days in Line S1. Selection for large litter size also increased the flux of follicles through follicle Types 3a to 5b by 72%, and through follicle Types 6 and 7 by 21%. Genetic variation was found in many aspects of the kinetics of follicular growth.

A quantitative cytochemical analysis of large antral follicles in two types of rat polycystic ovaries.

Ovaries from normal mature rats, rats injected with testosterone propionate (TP), and from aged rats were removed, and large antral follicles examined by quantitative cytochemical techniques in order to analyze possible enzymatic defects that relate to follicular steroidogenesis. The ovaries from the TP-injected and the aged rats were polycystic. Lipid deposition was analyzed in frozen sections stained with Sudan black. A microdensitometer was used to measure delta 5-3 beta-hydroxysteroid dehydrogenase (3 beta OHD) activity and G-6-PD type IH generation in the theca, and in peripheral region, antral region, and corona radiata of large antral follicles. 3 beta OHD is the enzyme that converts pregnenolone to progesterone. Type IH generation is related to the conversion of androstenedione to estradiol. Lipid droplet deposition was comparable in the three types of follicles. Compared to that in normal preovulatory follicles, 3 beta OHD activity was similar in identical regions of large antral follicles in TP-injected rats, but less in the theca and peripheral region of the membrana granulosa of the aged rat. G-6-PD type IH generation was less in the peripheral region of large antral follicles of both TP-injected and aged rats than in preovulatory follicles. Type IH generation was also less in the theca of TP-injected rats than in the theca of normal rats. This study provides evidence that in spite of their normal appearance, large antral follicles in polycystic ovaries are not physiologically sound. Furthermore, the enzymatic disturbance appears to be different in different types of polycystic ovaries.

Dendritic reticulum cells in reactive lymph nodes and tonsils: an immunohistological study.

There has recently been much interest in the patterns of follicular dendritic reticulum cells (DRC) in pathological lymph nodes, particularly in relation to the phenomenon of DRC break-up (thought to be pathognomonic of AIDS-related lymphadenopathies) and to progressive transformation of germinal centres (as a possible precursor of lymphocyte predominant Hodgkin's disease). In the present study we have immunostained twenty-nine reactive lymph nodes and five tonsils with monoclonal antibody R4/23 (DAKO-DRC) in order to evaluate the frequency of such changes in lymphoid tissue unaffected by AIDS or Hodgkin's disease. Most of the specimens contained typical secondary follicles with clearly defined germinal centres and mantle zones. There were two variants in lymph nodes showing follicular hyperplasia characterized by (i) progressive transformation of germinal centres and (ii) inclusions of nests of small lymphocytes within germinal centres. In each of these types of follicles the compact evenly-distributed meshwork of DRCs, as previously described, was seen. However there were considerable variations in DRC meshwork in each category (the pattern could not be predicted from the morphology) with examples in all three of the DRC break-up previously considered specific for the AIDS related lymphadenopathy. Since none of the lymph nodes and tonsils studied had any known relationship to either Hodgkin's disease or AIDS it is argued that none of the changes in the DRC meshwork observed are specific for these conditions.

Phosvitins in Fundulus oocytes and eggs. Preliminary chromatographic and electrophoretic analyses together with biological considerations.

Vitellogenin serves as the plasma precursor for the yolk proteins, lipovitellin and phosvitin, in nonmammalian vertebrates. 32P-Vitellogenin was isolated from the plasma of the teleost, Fundulus heteroclitus, and was used both to label phosvitin in the ovary and to indicate the phosvitin region in preparative chromatographs of ovarian extracts on DEAE-cellulose. Crude [32P]phosvitin could be resolved further into two labeled components with shallow gradients on DEAE-cellulose and into eight labeled components by electrophoresis on 12% polyacrylamide gels. Only the two largest electrophoretically resolved components could be correlated with Coomassie Blue staining bands, but several of the smaller components could be indicated with the cationic carbocyanine dye, Stains-all. Stains-all-dyed components were also generally indicated as multiple bands. The ovary of a reproductively active female contains vitellogenic oocytes, postvitellogenic oocytes undergoing maturation prior to ovulation, and ovulated eggs. Examination of various types of follicles and eggs on polyacrylamide gels revealed that during maturation, the largest phosvitin components formed during vitellogenesis either disappear or diminish, while smaller phosvitin components appear. The transformation of phosvitin components can also be achieved in vitro by incubating prematurational follicles in a saline medium containing deoxycorticosterone. These preliminary results demonstrate that a complex array of phosvitin-like components are present within a single ovary of F. heteroclitus. We also postulate that one reason for the anomalous yolk proteins generally found thus far in teleost eggs is that some of the proteins derived from vitellogenin during vitellogenesis undergo further proteolysis during oocyte maturation.

Changes in aromatase activity in small ovarian follicles of the domestic fowl (Gallus domesticus) during growth and atresia.

The tritium release assay for measuring aromatase activity was adapted to measure oestrogen production in intact small ovarian follicles of the domestic fowl. The activity was measured in three types of follicle classified as normal, atretic or grossly atretic. Normal follicles were distinguished from atretic by the presence of small haemorrhages on the surface of the atretic follicles. Grossly atretic follicles were identified by their deformed shape. The aromatase activity in normal and atretic follicles was related to follicular weight, the activity in atretic follicles being less than that in normal follicles. The aromatase activity in grossly atretic follicles was independent of follicular weight. The enzyme activity in this group of follicles was significantly less than that in either normal or atretic follicles. The significance of these results is discussed in relation to the induction of atresia within the small ovarian follicles of the domestic fowl.

Effects of gonadotrophins on progesterone production by isolated granulosa cells of the adenohypophysectomized domestic fowl (Gallus domesticus).

Ovine LH and ovine FSH stimulated progesterone production in granulosa cells isolated from the F1, F2 and F3 follicles of hypophysectomized and control (sham-operation) hens when they were collected 6 h after operation, but the steroidogenic response to LH was greater for granulosa cells from hypophysectomized hens. At 15 h after operation progesterone production by granulosa cells was stimulated by LH in all 3 follicle types of control hens, but only in the F1 follicles of hypophysectomized hens. The response to FSH at 15 h was similar for control and hypophysectomized hens. The time after hypophysectomy therefore appears to affect the LH-stimulated progesterone production by granulosa cells of the F2 and F3 follicles.

Seasonal differences in ovarian activity in cows

The plasma concentrations of LH and prolactin and various parameters of ovarian function were examined in cows on known days of the oestrous cycle during May and June (autumn and winter) and during October (spring). Luteinizing hormone peak frequency and plasma prolactin concentrations were significantly higher in October than during the May-June period (LH, P less than 0.05; prolactin, P less than 0.01). The mean diameters of large healthy follicles (greater than or equal to 8 mm diameter) and the dominant oestrogen-secreting follicles were significantly larger (P less than 0.01 for both follicle types) and each follicle contained more granulosa cells (both P less than 0.01) in May-June than in October. The LH responsiveness of theca interna with respect to androstenedione production and the levels of aromatase activity in granulosa cells did not differ with time of year. The corpora lutea were heavier (P less than 0.05) and secreted more progesterone (P less than 0.01) in May-June than in October. It is concluded that seasonal differences in ovarian activity exist in cows and that these differences are probably the consequence of seasonal differences in gonadotrophin secretion.

Influence of progesterone, androstenedione and oestradiol-17 beta on the incorporation of [3H]proline in the human follicular wall.

The incorporation of proline, a precursor for collagen specific hydroxyproline, is regarded to reflect the metabolism of collagen. In vitro experiments were carried out to investigate the influence of sex steroids on the incorporation of [3H]proline into the human follicular wall. Tissue pieces from the apical wall of follicles at different stages of development were incubated in the presence of the steroids and [3H]proline, and the tissue bound radioactivity was determined. Progesterone induced a decrease of radiolabelling in both unripe and pre-ovulatory follicles. Androstenedione caused a similar effect but only in pre-ovulatory follicles. The influence of oestradiol-17 beta on the incorporation of [3H]proline was less pronounced with a tendency towards a decrease of radiolabelling in both types of follicles. It is suggested that the observed biochemical changes induced by sex steroids are of importance for connective tissue remodelling preceding human follicular rupture.

Cell proliferation in the dog (beagle) ovary during proestrus and early estrus.

Cell proliferation in the ovaries of 5 Beagle dogs was studied by autoradiography after pulse labelling with (3H)-thymidine during different periods of proestrus and early estrus. Indices of labelling and of necrosis were determined for different follicle types grouped according to their stages of atresia. Cell proliferation became most obvious during early proestrus in antral and in Graafian follicles, and during the periovulatory period in preantral follicles. By contrast, low labelling indices appeared in all follicle types during the middle of proestrus. All follicle types demonstrated a continuous decrease in labelling from early to late atresia with the exception of the granulosal layer of antral follicles during the periovulatory period. While many necrotic granulosal cells were found in antral follicles during the first 6 days of proestrus, there were few during the periovulatory period. Even Graafian follicles displayed necrotic granulosal cells now and then. It may be concluded that (1) intact ovarian follicles can show subtle changes indicating the beginning of atresia, and (2) during the proliferative phase of a long estrous cycle cell replication in the ovary is characterized by two bursts at the beginning and at the end of this period.

Histopathologic survey of ovaries of plaice, Pleuronectes platessa L., from Aber Wrac'h and Aber Benoit, Brittany, France: long‐term effects of the Amoco Cadiz crude oil spill

Abstract. Ovarian samples were collected from plaice, Pleuronectes platessa, captured in the highly oiled Aber Wrac'h and Aber Benoit estuaries at four intervals during 1979–1980, following the Amoco Cadiz crude oil spill. Reference plaice were obtained along the western and southern coasts of Brittany. Tissue samples were fixed, processed and stained by routine histologic procedures. Ovaries were examined for histopathologic changes and for four different types of follicles. The concentrations and percentages of four types of ovarian follicles (primordial, primary, growing and mature) were determined for plaice ovaries within each sampling site at each sampling interval. Reference site ovaries, which were considered representative of the ovarian cyclic events for plaice along the Brittany coast, had primordial and primary follicles in high concentration in the summer season and in low concentration in the winter season. Concentrations of mature ovarian follicles in reference site plaice ovaries were low in the summer and high in the winter seasons. Growing follicles were observed infrequently in the reference site plaice ovaries. The seasonal pattern exhibited by follicles in ovaries from Aber Benoit plaice was similar to that observed in the reference site ovaries, but both the concentrations and percentages of follicle types were lower. The seasonal pattern of follicle types in Aber Wrac'h plaice ovaries was the reverse of that seen in the reference site plaice ovaries. In the Aber Wrac'h samples, concentrations of primordial and primary follicles were high in the winter months and low in the summer season, and no mature follicles were observed at any season. Growing follicles were seen more frequently in Aber Wrac'h and Aber Benoit plaice ovaries than in reference site plaice ovaries at all sampling intervals. Other than atretic follicles with an associated leucocytosis, no histopathologic conditions were observed in any ovary.

Changes in gonadotrophin binding to rat ovaries during sexual maturation.

Changes in LH and FSH binding to rat ovaries during sexual maturation were measured by in vitro binding assay and autoradiography. Ovaries were obtained from rats at various ages ranging from 5 till 35 days of age. Animals older than 35 days of age were classified in three groups: anoestrus (uterine weight below 60 mg), early pro-oestrus (uterine weight 60-120 mg) and pro-oestrus (uterus distended with fluid). Measurements were made of specific binding of radioiodinated hCG and rFSH to ovarian homogenates (day 5-35) and to granulosa cells isolated from the largest follicles (day 35-pro-oestrus). The earliest age at which specific hCG and FSH binding could be demonstrated was day 11. Thereafter hCG binding to ovarian homogenates, expressed in CPM/microgram DNA, increased with age. Specific binding of hCG to granulosa cells increased from 1000 CPM/microgram DNA at day 35 to 4000 CPM/microgram DNA at pro-oestrus. FSH binding to ovarian homogenates increased till day 21 and remained fairly constant thereafter. Autoradiography revealed the presence of hCG binding to interstitial tissue and thecal cells. hCG binding to granulosa cells was only found in large pre-ovulatory follicles present at early pro-oestrus and pro-oestrus. FSH binding was observed to granulosa cells of all types of follicles, but not to interstitial and thecal cells. The results demonstrate that during sexual maturation FSH binding to granulosa cells remains constant, whereas LH binding to interstitium and theca and ultimately to granulosa cells increases with age suggesting an increased responsiveness of the ovaries to LH.

Follicular growth and kinetics during the estrous cycle, pregnancy and postpartum in the Indian mole rat (Bandicota bengalensis).

Follicular growth and kinetics were studied in detail in the ovaries of the Indian mole rat (Bandicota bengalensis) during various stages of the estrous cycle; days 7, 12, 15, 19, and 21 of pregnancy; and day 2 postpartum. The sizes of follicles, oocytes, nuclei, and nucleoli were measured. In all rats, regression coefficients, a, and intercepts, b, were calculated in oocyte/follicle, oocyte nucleus/follicle and oocyte nucleus/oocyte regressions. The oocyte reached its maximum size when the average follicle diameter was 117 microns in nonpregnant rats and 131 microns in pregnant rats. The oocyte nucleus attained maximum size when the follicle diameter was 110 microns during the estrous cycle and 111 microns during pregnancy and postpartum. Maximum values of the diameter of the largest antral follicle and average diameter of the four largest antral follicles were observed during proestrus (473 and 442 microns, respectively) and on day 21 of pregnancy (611 and 538 microns, respectively). Chi 2 analysis showed that distribution of various types of follicles was not independent of the stage of the estrous cycle and pregnancy. In estrus and metestrus most of the follicles were between stages I and V. However, by diestrus and proestrus, follicles of all size groups developed. The numbers of stage I and II follicles did not differ as pregnancy advanced. More stage V follicles were present on day 12 than on day 7 of pregnancy; however, their numbers decreased by day 15. Afterwards, progressive increase of stage V and (VI + VII) follicles was observed until day 21. This was accompanied by the shift of follicles from stage (III + IV) on days 19 and 21 of pregnancy and even of stage II on day 2 postpartum. Wherever possible, the results have been compared with previous observations in various rodent species.

Ovarian Follicles of New-born Merino Lambs from Genetic Lines which Differ in Fecundity

Ovaries were obtained from 78 new-born lambs (12 singletons, 25 twins, 28 triplets, 10 quadruplets and 3 quintuplets) from flocks selected for (T902, T903 and Booroola) or against (O) multiple births. Sections of the ovaries were examined with a projection microscope and the numbers of all types of follicles were estimated. There were no differences between genetic lines in the number of primordial follicles, after adjustment for litter size and sire; however, there were significantly more of these follicles in single-born lambs than in lambs born in litters of two or more within genetic lines. The number of vesicular follicles was lower in Booroola than in O lambs.

Demodex folliculorum and Demodex brevis in cutaneous biopsies

The hair follicle mites Demodex folliculorum and Demodex brevis are ubiquitous obligatory ectoparasites of man. We studied these mites in a consecutive series of skin biopsies submitted to a dermatopathology laboratory; 10% of all biopsies and 12% of all follicles contained demodectic mites. The prevalence of both species increased with age, but D. brevis had a lower prevalence. The face was most heavily infested by both species, but D. brevis had a wider distribution on the body. Males were more heavily infested than females with both species, the difference being strongest for D. brevis. Other ecologic characteristics noted include the type of follicles infested, follicular dilation, perifollicular inflammation, and the presence of a dense homogeneous eosinophilic material surrounding the mites. These data confirm ecologic and epidemiologic differences between the two species and emphasize that they should be distinguished in future studies of the roles these mites may play in a variety of disease states.

Inhibition of rat follicular androgen synthesis by dibutyryl cyclic AMP.

Abstract. LH exerts a biphasic effect on rat follicular steroidogenesis: an initial general stimulation of steroidogenesis later followed by an inhibition of androgen and oestrogen synthesis. It is known that the stimulation by LH of steroidogenesis is mediated by cAMP. The present paper examines the role of cAMP in the late inhibitory phase of steroidogenesis. Pre-ovulatory follicles were isolated from PMSG treated immature rats and incubated in modified bicarbonate buffer for different time periods. The accumulation of progesterone (P), androstenedione (A), testosterone (T) and oestradiol-17β (E2) in the medium was measured. Addition of dibutyryl cAMP (dbcAMP, 1 mm) to the medium caused a prolonged increase in P accumulation for 10 h while A and E2 accumulation was stimulated only for 4–6 h whereafter the levels were sustained. The pattern was identical to that produced by LH (10 μg/ml) in vitro. In a second type of experiments follicles were pre-incubated for 6 h with or without LH or dbcAMP and then transferred for a second 2 h incubation, in the presence or absence of unlabelled 17-hydroxyprogesterone (17-OHP) or T (1 μg/ml). Follicles pre-incubated in plain medium increased their formation of A and T in the presence of 17-OHP, while follicles exposed to LH or dbcAMP did not. Addition of T caused a similar increase in E2 formation in all groups (control, LH and dbcAMP). It is concluded that since dbcAMP mimicked the entire response of follicular steroidogenesis normally elicited by LH, cAMP seems to be the physiologic mediator of LH, both on the early stimulatory and on the late inhibitory phase of pre-ovulatory follicular steroidogenesis.

Development of preovulatory follicles and oocytes during the oestrous cycle of mature and aged rats.

Abstract. The percentage of viable granulosa cells and the viability of the oocyte of each antral follicle on each day of the oestrous cycle of mature and aged rats was determined by trypan blue exclusion and fluorescein diacetate assays, respectively. In mature rats, the number of mid-sized follicles (350–550 μm in diameter) increased from oestrus through di-oestrus. The percentage of viable granulosa cells increased in only mid-sized follicles which contained a viable oocyte. On pro-oestrus, two types of preovulatory follicles (> 550 μm in diameter) were observed in the mature pro-oestrous rat: those with a high granulosa cell viability (> 60%) and a viable oocyte and those with a lower percentage of viable granulosa cells (< 50%) and a non-viable oocyte. This suggests that in mature rats only viable oocytes would be ovulated since presumably, only those preovulatory follicles with a high percentage of viable granulosa cells are able to ovulate. In aged rats, preovulatory-sized follicles were present on each day of the cycle. The percentage of viable granulosa cells within those preovulatory-sized follicles with nonviable oocytes increased on met-oestrus, decreased on di-oestrus and increased by pro-oestrus. This pattern resulted in the development of 3 to 4 preovulatory follicles per ovary with all preovulatory-sized follicles possessing a high percentage of viable granulosa cells regardless of the functional state of the oocyte. It is proposed that in aged rats, all the preovulatory follicles would ovulate but many of the ovulated oocytes would be defective. This mechanism would account for the lower fertility of these older animals.

Steroid and adenosine 3',5'-monophosphate formation in granulosa and thecal cells from human preovulatory follicles in response to human chorionic gonadotropin.

Granulosa and thecal cells from the preovulatory follicles of 14 women were mechanically isolated and separately incubated for short term periods (0.5-4 h) in the presence and absence of hCG. After incubation, tissue cAMP levels and medium progesterone content of (P), androstenedione (A), and 17 beta-estradiol (E2) were determined. All follicles appeared healthy and mature, as judged by their number of granulosa cells, histological examination of the oocytes, athe steroid levels in antral fluid, as well as the appearance of the oocyte-cumulus complexes. Under basal conditions, both granulosa and thecal cells had the capacity to synthesize all of the steroids measured. The predominant steroid formed by the granulosa cells was P, while the thecal cells formed A as the major steroid. Both cell types produced considerable amounts of E2. The addition of hCG in various concentrations gave rise to concentration-dependent increase in cAMP formation in both cell types. Furthermore, hCG caused a statistically significant increase in the formation of P and A by the thecal cells, while steroid formation by the granulosa cells was not significantly altered. With higher concentrations of hCG, however, there was a tendency toward a stimulation of P synthesis in the granulosa cells from most of the follicles tested. It is concluded that in preovulatory follicles of human origin, follicular steroidogenesis is not rigidly compartmentalized between the two cell types. Furthermore, both granulosa and thecal cells are sensitive to stimulation with hCG in this type of follicle.

Aspects of cellular proliferation during follicular atresia in the dog ovary

Autoradiography after pulse labelling with [3H] thymidine was applied to investigate the proliferation processes in the granulosa and theca related to follicular atresia of the dog ovary during metestrus. The number of proliferating cells depends on the follicle type and its atretic stage. There is less proliferation in smaller secondary follicles than either in larger ones or tertiary follicles. While in early atresia tertiary follicles show the highest labelling indices, in advanced atresia the largest secondary follicles are those with the highest values. For each follicle type a decline in the labelling indices can be observed from early to terminal atresia. Tertiary follicles show a precipitous decrease in the labelling index between early and advanced atresia. There is a continuous gradient of proliferation from the center of the follicle over the peripheral granulosa to the theca. In tertiary follicles, an inverse correlation between labelling and necrosis of granulosa cells can be observed.