Priming procedure and cell isolation for study on luteal cell response to peptide hormone in the rat

1986 ◽  
Vol 111 (3) ◽  
pp. 387-393 ◽  
Author(s):  
A. Kumai ◽  
R. Asakai ◽  
S. Sakamoto ◽  
S. Sassa ◽  
R. Okamoto
Keyword(s):  
Simple Procedure ◽  
Peptide Hormone ◽  
Sucrose Density Gradient ◽  
Test Tube ◽  
Luteal Cell ◽  
Corpora Lutea ◽  
Suitable Model ◽  
Isolated Cells ◽  
Luteal Cells

Abstract. The objective of this study was to develop a method of isolating luteal cells from the ovaries of immature rats pretreated with pregnant mare serum gonadotrophin (PMSG). After the ovaries were digested by collagenase and trypsin, the corpora lutea were obtained from the tissues, gently pressed in a test tube, and then placed on a sucrose density gradient. The two bands that appeared in the tube after centrifugation were designated S1 (top band) and S2 (bottom band). Progesterone and 20α-dihydroprogesterone (20α-DHP) secreted by the isolated cells during short-term incubation were measured by radioimmunoassay (RIA). A larger amount of progesterone, i.e., 60 to 260 ng/105 cells, was secreted by S1 cells than by S2 cells during the 18-h incubation. These results suggest that this simple procedure for isolation of luteal cells may provide a suitable model for in vitro studies of the luteal function.

Inadequate function of corpora lutea following the induction of ovulation with monensin and FSH in seasonally anoestrous ewes

1988 ◽  
Vol 117 (2) ◽  
pp. 167-172 ◽  
Author(s):  
S. Atkinson
Keyword(s):  
Venous Blood ◽  
Plasma Concentrations ◽  
Ovarian Response ◽  
Treated Group ◽  
Luteal Cell ◽  
Control Group ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
The Mean

ABSTRACT Sixteen ewes in mid-seasonal anoestrus were stimulated to ovulate using sequential injections of FSH (total dose 10 mg) over a 4-day period. Half of the ewes received a dietary growth promotant (monensin) known to enhance the ovarian response to exogenous gonadotrophins. The ewes were ovariectomized on day 5 or 11 (day 0 = the initiation of FSH treatment). Serial blood samples were taken in half of the ewes to determine peripheral concentrations of LH and a single sample of ovarian venous blood was collected before ovariectomy. All luteal structures were dissected from the ovaries, counted and incubated in vitro to determine progesterone production. The luteal structures were then examined histologically for the abundance of luteal cells. The physical appearance of the ovary, along with plasma concentrations of LH and ovarian venous oestradiol indicated that the monensin-treated ewes ovulated before control ewes. The corpora lutea from control ewes produced significantly (P <0·05) more progesterone than did the corpora lutea from the monensin-treated group. Furthermore, only 7% of the remaining luteal structures in the monensin-treated group produced significant amounts of progesterone on day 11, whereas 61% of the luteal structures in the control group were actively secreting progesterone. The mean number of granulosa cells in the follicles was similar at ovulation in the two groups, but the mean numbers of large and small luteal cells were significantly (P <0·05) lower in luteal structures from the monensin-treated ewes than in those from the control ewes. It is therefore postulated that inadequate corpora lutea function following precocious ovulation is due to a lack of luteal cell development formed after premature luteinization. J. Endocr. (1988) 117, 167–172

scholarly journals Estrogen promotes luteolysis by redistributing prostaglandin F2α receptors within primate luteal cells

Reproduction ◽  
2015 ◽  
Vol 149 (5) ◽  
pp. 453-464 ◽  
Author(s):  
Soon Ok Kim ◽  
Nune Markosyan ◽  
Gerald J Pepe ◽  
Diane M Duffy
Keyword(s):  
Estrogen Receptor ◽  
Granulosa Cells ◽  
Prostaglandin F2α ◽  
Luteal Cell ◽  
Corpora Lutea ◽  
Dependent Manner ◽  
Perinuclear Region ◽  
Luteal Cells ◽  
Progesterone Production

Prostaglandin F2α (PGF2α) has been proposed as a functional luteolysin in primates. However, administration of PGF2α or prostaglandin synthesis inhibitors in vivo both initiate luteolysis. These contradictory findings may reflect changes in PGF2α receptors (PTGFRs) or responsiveness to PGF2α at a critical point during the life span of the corpus luteum. The current study addressed this question using ovarian cells and tissues from female cynomolgus monkeys and luteinizing granulosa cells from healthy women undergoing follicle aspiration. PTGFRs were present in the cytoplasm of monkey granulosa cells, while PTGFRs were localized in the perinuclear region of large, granulosa-derived monkey luteal cells by mid-late luteal phase. A PTGFR agonist decreased progesterone production in luteal cells obtained at mid-late and late luteal phases, but did not decrease progesterone production by granulosa cells or luteal cells from younger corpora lutea. These findings are consistent with a role for perinuclear PTGFRs in functional luteolysis. This concept was explored using human luteinizing granulosa cells maintained in vitro as a model for luteal cell differentiation. In these cells, PTGFRs relocated from the cytoplasm to the perinuclear area in an estrogen- and estrogen receptor-dependent manner. Similar to our findings with monkey luteal cells, human luteinizing granulosa cells with perinuclear PTGFRs responded to a PTGFR agonist with decreased progesterone production. These data support the concept that PTGFR stimulation promotes functional luteolysis only when PTGFRs are located in the perinuclear region. Estrogen receptor-mediated relocation of PTGFRs within luteal cells may be a necessary step in the initiation of luteolysis in primates.

Contrasting effects of prolactin on luteal and follicular steroidogenesis

1985 ◽  
Vol 104 (2) ◽  
pp. 241-250 ◽  
Author(s):  
B. Kalison ◽  
M. L. Warshaw ◽  
G. Gibori
Keyword(s):  
Luteal Cell ◽  
Corpora Lutea ◽  
Aromatase Activity ◽  
Luteal Cells ◽  
Progesterone Production ◽  
Progesterone Secretion ◽  
Hypophysectomized Rats ◽  
Stimulation Of

ABSTRACT To determine whether prolactin affects both luteal and follicular production of testosterone and oestradiol, pseudopregnant rats, either intact or hypophysectomized on day 8, were injected daily between days 8 and 9 with 1·5 i.u. human chorionic gonadotrophin (hCG), 250 μg prolactin or a combination of both. Control rats were given vehicle. On day 9, blood was obtained from the ovarian vein and corpora lutea and follicles were isolated and incubated in vitro for 2 h. Administration of hCG to intact rats increased ovarian secretion of testosterone and oestradiol dramatically, but did not affect progesterone secretion. Hypophysectomy on day 8 of pseudopregnancy was followed by a drop in ovarian steroid secretion. Prolactin treatment of hypophysectomized rats markedly enhanced progesterone production but had no stimulatory effect on either testosterone or oestradiol. In contrast, hCG dramatically enhanced ovarian secretion of both testosterone and oestradiol without affecting progesterone secretion. Prolactin administered together with hCG antagonized the stimulation of both testosterone and oestradiol secretion by hCG, yet increased progesterone production. When the specific effects of hCG and prolactin administration on follicles and corpora lutea were studied separately, it was found that hCG treatment in vivo greatly stimulated testosterone and oestradiol production by both tissues in vitro. Since hCG only marginally affected aromatase activity in the follicle, had no effect on aromatase activity in luteal cells and did not increase progesterone synthesis, it appears that hCG acts to increase the formation of androgen substrate for oestradiol biosynthesis. Prolactin, administered with or without hCG, inhibited both basal and hCG-stimulated testosterone and oestradiol synthesis by the follicle. In sharp contrast to its inhibitory effect on follicular production of steroids, prolactin appears to be essential for LH stimulation of testosterone and oestradiol by the corpus luteum. In the absence of prolactin, luteal cells gradually ceased to respond to LH and decreased their output of testosterone and oestradiol. Prolactin administration to hypophysectomized rats did not affect luteal cell production of either steroid. However, corpora lutea of rats treated with prolactin responded to the hCG challenge with an increase in testosterone and oestradiol synthesis. In summary, results of this investigation demonstrate that prolactin affects follicular and luteal production of testosterone and oestradiol in opposite ways. It acts on the follicle to inhibit both basal and LH-stimulated production of testosterone and oestradiol, yet it markedly enhances LH stimulation of testosterone and oestradiol synthesis by luteal cells. J. Endocr. (1985) 104, 241–250

scholarly journals HIF-1α Activation Promotes Luteolysis by Enhancing ROS Levels in the Corpus Luteum of Pseudopregnant Rats

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Zonghao Tang ◽  
Jiajie Chen ◽  
Zhenghong Zhang ◽  
Jingjing Bi ◽  
Renfeng Xu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Corpus Luteum ◽  
Cell Apoptosis ◽  
In Vitro Study ◽  
Luteal Cell ◽  
Independent Manner ◽  
Luteal Regression ◽  
Luteal Cells

The increase of oxidative stress is one of the important characteristics of mammalian luteal regression. Previous investigations have revealed the essential role of reactive oxygen species (ROS) in luteal cell death during luteolysis, while it is unknown how ROS is regulated in this process. Considering the decrease of blood flow and increase of PGF2α during luteolysis, we hypothesized that the HIF-1α pathway may be involved in the regulation of ROS in the luteal cell of the late corpus luteum (CL). Here, by using a pseudopregnant rat model, we showed that the level of both HIF-1α and its downstream BNIP3 was increased during luteal regression. Consistently, we observed the increase of autophagy level during luteolysis, which is regulated in a Beclin1-independent manner. Comparing with early (Day 7 of pseudopregnancy) and middle CL (Day 14), the level of ROS was significantly increased in late CL, indicating the contribution of oxidative stress in luteolysis. Inhibition of HIF-1α by echinomycin (Ech), a potent HIF-1α inhibitor, ameliorated the upregulation of BNIP3 and NIX, as well as the induction of autophagy and the accumulation of ROS in luteal cells on Day 21 of pseudopregnancy. Morphologically, Ech treatment delayed the atrophy of the luteal structure at the late-luteal stage. An in vitro study indicated that inhibition of HIF-1α can also attenuate PGF2α-induced ROS and luteal cell apoptosis. Furthermore, the decrease of cell apoptosis can also be observed by ROS inhibition under PGF2α treatment. Taken together, our results indicated that HIF-1α signaling is involved in the regression of CL by modulating ROS production via orchestrating autophagy. Inhibition of HIF-1α could obviously hamper the apoptosis of luteal cells and the process of luteal regression.

scholarly journals Control of Steroidogenesis in Small and Large Bovine Luteal. Cells

1987 ◽  
Vol 40 (3) ◽  
pp. 331 ◽  
Author(s):  
William Hansel ◽  
Hector W Alila ◽  
Joseph P Dowd ◽  
Xiangzhong Yang
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Corpus Luteum ◽  
Luteal Cell ◽  
Lipoxygenase Pathway ◽  
Luteal Cells ◽  
Kinase C ◽  
Bovine Corpus Luteum ◽  
Progesterone Synthesis

Evidence was cited to show that: (1) prostacyclin (PGI2) plays a luteotrophic role in the bovine corpus luteum and that products of the lipoxygenase pathway of arachidonic acid metabolism, especially 5-hydroxyeicosatetraenoic acid play luteolytic roles; (2) oxytocin of luteal cell origin plays a role in development, and possibly in regression, of the bovine corpus luteum; and (3) luteal cells arise from two sources; the characteristic small luteal cells at all stages of the o~strous cycle and pregnancy are of theca cell origin; the large cells are of granulosa cell origin early in the cycle, but a population of theca-derived large cells appears later in the cycle. Results of in vitro studies with total dispersed cells and essentially pure preparations of large and small luteal cells indicate that : (1) the recently described Ca2+ -polyphosphoinositol-protein kinase C second messenger system is involved in progesterone synthesis in the bovine corpus luteum; (2) activation of protein kinase C is stimulatory to progesterone synthesis in the small luteal cells; (3) activation of protein kinase C has no effect on progesterone synthesis in the large luteal cells; and (4) protein kinase C exerts its luteotrophic effect in total cell preparations, in part at least, by stimulating the production of prostacyclin. The protein kinase C system may cause down regulation of LH receptors in the large cells.

CHANGES IN THE DENSITY AND PROGESTERONE CONTENT OF LUTEAL TISSUE IN THE EGYPTIAN BUFFALO DURING THE OESTROUS CYCLE

1967 ◽  
Vol 39 (2) ◽  
pp. 163-171 ◽  
Author(s):  
A. S. EL-SHEIKH ◽  
FRANÇOIS B. SAKLA ◽  
SAFAA O. AMIN
Keyword(s):  
Corpus Luteum ◽  
Cell Volume ◽  
Distribution System ◽  
Oestrous Cycle ◽  
Luteal Cell ◽  
Corpora Lutea ◽  
Luteal Cells ◽  
Functional Changes ◽  
Parallel Increase ◽  
Luteal Tissue

SUMMARY The histological and functional changes of 31 corpora lutea of Egyptian buffaloes during the various phases of the oestrous cycle were studied. The volumes of the corpora lutea were calculated, the volume per cell, the cell volume and the volume of the intercellular spaces were estimated from transverse serial sections stained with haematoxylin and eosin, Mallory's triple stain or van Gieson's stain. The nuclear volumes were also determined and the cytoplasmic volume was calculated. The progesterone content was estimated using column absorption chromatography and a counter-current distribution system. It was concluded that the luteal cells increase both in volume and in number due to mitosis. The luteal cells decrease in volume after the 15th day after ovulation, the cells lose their distinct outlines in the regressive stage and disappear completely in the corpus albicans. There was a parallel increase in luteal cell volume and progesterone content until the 15th post-ovulatory day followed by a decrease in the regressive phase and disappearance of the hormone in the corpus albicans. A highly significant correlation (r = +0·875) was found between the progesterone content and the cytoplasmic volume. Progesterone concentration/g. luteal tissue increased from the corpus haemorrhagicum to the mature corpus luteum, decreased in the regressive corpus luteum and completely disappeared in the corpus albicans.

Cellular basis of luteal steroidogenesis in the human ovary

1989 ◽  
Vol 122 (1) ◽  
pp. 303-NP ◽  
Author(s):  
B. Fisch ◽  
R. A. Margara ◽  
R. M. L. Winston ◽  
S. G. Hillier
Keyword(s):  
Luteal Phase ◽  
Culture System ◽  
Direct Action ◽  
Cellular Level ◽  
Luteal Cell ◽  
Aromatase Activity ◽  
Vitro Fertilization ◽  
Luteal Cells ◽  
Expected Time

ABSTRACT A primary monolayer cell culture system was developed to investigate human corpus luteum (CL) function in vitro. Steroidogenic cells were isolated by collagenase dispersal and Percoll density-gradient fractionation from CLs enucleated at progressive stages of the luteal phase (tubal surgery patients). 'Pure' granulosa-lutein cells were aspirated from ovulatory follicles at mid-cycle (in-vitro fertilization patients). The steroidogenic capacity (progesterone/20α-dihydroprogesterone biosynthesis and aromatase activity) of isolated luteal cells was assessed in relation to CL development. Basal luteal cell steroidogenesis was maximal at around the expected time of ovulation and declined with CL age during the luteal phase. Conversely, human chorionic gonadotrophin (hCG)-responsive steroidogenesis was initially undetectable but developed as the luteal phase progressed. These results show that luteal cell steroidogenesis becomes increasingly dependent upon gonadotrophic support with CL age. This is evidence that functional luteolysis in human ovaries (1) is pre-programmed to occur at the cellular level, (2) is initiated automatically at the time of ovulation and (3) is reversed at the time of CL 'rescue' in early pregnancy by the direct action of trophoblastic hCG on steroidogenic luteal cells. The culture system described should be of value in further defining the control of human CL form and function at the cellular level. Journal of Endocrinology (1989) 122, 303–311

scholarly journals Corpora lutea induced by gonadotrophin-releasing hormone treatment of anoestrous Welsh Mountain ewes: reduced sensitivity to luteinizing hormone in vivo and to chorionic gonadotrophin in vitro

Reproduction ◽  
2005 ◽  
Vol 129 (1) ◽  
pp. 61-73 ◽  
Author(s):  
T A Bramley ◽  
D Stirling ◽  
G S Menzies ◽  
D T Baird
Keyword(s):  
Chorionic Gonadotrophin ◽  
Luteal Cell ◽  
Corpora Lutea ◽  
Lh Receptor ◽  
Group 4 ◽  
Progesterone Secretion ◽  
Group 3 ◽  
Gonadotrophin Releasing Hormone

Seasonally anoestrous Welsh Mountain ewes received 250 ng gonadotrophin-releasing hormone (GnRH) every 2 h, with (Group 1;n= 13) or without (Group 2;n= 14) progesterone priming for 48 h. Fourteen control ewes (Group 3) were studied during the luteal phase in the breeding season. Animals in Group 4 (n= 12) received progesterone priming followed by 250 ng GnRH at increasing frequency for 72 h, while ewes in Group 5 (n= 13) were given three bolus injections of 30 μg GnRH at 90-min intervals. All treatment regimens induced ovulation. However, only corpora lutea (CL) from ewes in Group 3 (breeding season) or Group 4 exhibited normal luteal function. Luteal luteinizing hormone (LH) receptor levels were significantly higher on day 12 than day 4, and CL from groups with adequate CL (3 and 4) had significantly higher125I-human chorionic gonadotrophin (hCG)-binding levels than the three groups with inadequate CL on day 12. LH-binding affinity was unchanged. Exogenous ovine LH (10 μg)in vivoon days 3 or 11 after ovulation induced a pulse of progesterone in ewes with adequate CL: however, ewes in Groups 1, 2 and 5 showed no significant response. Basal progesterone secretionin vitrowas significantly greater on day 4 than on day 12. Maximal steroidogenic responses of adequate and inadequate CL to hCG and to dibutyryl cyclic-3′,5′-AMP were similar at both stages of the luteal phase. However, the EC50for hCG on days 4 and 12 was 10-fold lower for groups with an adequate CL (0.1 IU hCG/ml) than for inadequate-CL groups (1 IU hCG/ml;P<0.05). Thus, in addition to the well-characterized premature sensitivity of GnRH-induced inadequate CL to endometrial luteolysin, we have shown (1) a marked decrease in total number of cells in the CL, a profound reduction in vascular surface area, and a decrease in mean large luteal cell volume (with no change in large luteal cell numbers), (2) decreased luteal LH receptor and progesterone content compared with adequate CL and (3) that CL that were becoming, or were destined to become, inadequate failed to respond to ovine LHin vivoand were 10-fold less sensitive to hCG in terms of luteal progesterone secretionin vitro.

Effects of prolactin on steroid production by human luteal cells in vitro

1983 ◽  
Vol 96 (3) ◽  
pp. 499-503 ◽  
Author(s):  
G. J. S. Tan ◽  
J. S. G. Biggs
Keyword(s):  
Menstrual Cycle ◽  
Ovarian Function ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Corpora Lutea ◽  
Short Term ◽  
Steroid Production ◽  
Luteal Cells ◽  
Progesterone Production

The effects of prolactin on steroidogenesis were studied in dispersed luteal cells prepared from human corpora lutea of the menstrual cycle. Prolactin, at concentrations of 0·1–1000 ng/ml, had no effect on progesterone production by luteal cells during short-term incubation (3 h). However, in two out of five corpora lutea, higher concentrations of prolactin (100 and 1000 ng/ml) significantly reduced the oestradiol-17β production induced by human chorionic gonadotrophin (hCG; 10 i.u./ml); lower doses of prolactin had little effect. In the remaining corpora lutea, prolactin failed to affect either basal or hCG-induced production of oestradiol-17β. These results are discussed in relation to the mechanism by which prolactin influences human ovarian function.

scholarly journals Cell death induced by serum deprivation in luteal cells involves the intrinsic pathway of apoptosis

Reproduction ◽  
2006 ◽  
Vol 131 (1) ◽  
pp. 103-111 ◽  
Author(s):  
Alicia A Goyeneche ◽  
Jacquelyn M Harmon ◽  
Carlos M Telleria
Keyword(s):  
Cell Death ◽  
Corpus Luteum ◽  
Simian Virus 40 ◽  
Full Length ◽  
Luteal Cell ◽  
Serum Starvation ◽  
Endocrine Gland ◽  
Luteal Cells ◽  
Defined Culture

The corpus luteum is a transient endocrine gland specializing in the production of progesterone. The regression of the corpus luteum involves an abrupt decline in its capacity for producing progesterone followed by its structural involution, which is associated with apoptosis of the luteal cells. An in vitro experimental approach is needed to study the molecular mechanisms underlying hormonal regulation of luteal cell death under defined experimental conditions. In this study, we investigated simian virus-40-transformed luteal cells to determine whether they can be driven to apoptosis and, if so, to define the intracellular pathway involved. Luteal cells were cultured in the presence or absence of fetal bovine serum for 24 or 48 h. Under serum starvation conditions, the luteal cells underwent growth arrest accompanied by cell death as evaluated by dye exclusion, and confirmed by two-color fluorescence cell viability/cytotoxicity assay. We next studied whether serum starvation-induced death of luteal cells occurred by apoptosis. Morphologic features of apoptosis were observed in cells stained with hematoxylin after being subjected to serum starvation for 48 h. The apoptotic nature was further confirmed by in situ 3′-end labeling and fragmentation of genomic DNA. Apoptosis of serum-deprived luteal cells was dependent upon caspase activation. Serum starvation induced cleavage of poly (ADP-ribose) polymerase (PARP), suggesting that caspase-3 had been activated under the stress of withdrawal of growth factors. This was confirmed by cleavage of full-length procaspase-3. Finally, the fact that serum starvation promoted the cleavage of full-length procaspase-9 and the decrease in the expression of endogenous Bid, a BH-3-only proapoptotic protein of the Bcl-2 family, indicates that the intrinsic (i.e., mitochondrial) pathway of apoptosis was activated. In summary, we have characterized an in vitro experimental model of luteal cell death that can be utilized to evaluate the role of hormones in apoptosis of luteal cells under defined culture conditions, and to study the mechanism of luteal regression.

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