IGF-I Prevents Fas Ligand-Induced Apoptosis in Granulosa Cells of Buffalo (Bubalus bubalis) Ovarian Follicles In Vitro.

2011 ◽  
Vol 85 (Suppl_1) ◽  
pp. 657-657
Author(s):  
Ravindra P. Janivara ◽  
Sunil K. Patil ◽  
Jamuna V. Kolatalu ◽  
Dhanya Joseph ◽  
Raghavendra Subbarao ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Fas Ligand ◽  
Bubalus Bubalis ◽  
Ovarian Follicles ◽  
Igf I ◽  
Induced Apoptosis

scholarly journals Serum deprivation affects secretory activity of cultured porcine ovarian follicles and granulosa cells and their response to hormones

2009 ◽  
Vol 54 (No. 10) ◽  
pp. 455-460
Author(s):  
A.V. Sirotkin
Keyword(s):  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Secretory Activity ◽  
Ovarian Follicles ◽  
Serum Deprivation ◽  
Ovarian Cells ◽  
Igf I ◽  
Granulose Cells ◽  
Vitro Model

The aim of the present study is to understand the hormonal mechanisms of the effect of malnutrition on ovarian follicle functions. For this purpose, we examined the effect of malnutrition/serum deprivation, addition of metabolic hormones and gonadotropin (IGF-I, leptin and FSH) and their combination on the release of progesterone (P<sub>4</sub>), testosterone (T), estradiol (E<sub>2</sub>) and insulin-like growth factor I (IGF-I) by cultured whole ovarian follicles and on P<sub>4</sub> and IGF-I output by cultured granulosa cells isolated from porcine ovaries. It was observed that in ovarian follicles cultured with nutrients/serum addition of IGF-I reduced release of P<sub>4</sub>, but not of T or E<sub>2</sub>. Exogenous leptin reduced output of E<sub>2</sub>, but not of P<sub>4</sub> or T, and increased IGF-I output. No significant effect of FSH on release of steroid hormones by isolated follicles was found. Serum deprivation did not affect release of P<sub>4</sub>, but reduced output of T and E<sub>2</sub>, and promoted IGF-I release by cultured ovarian follicles. Addition of hormones failed to prevent the effect of malnutrition on the secretory activity of cultured ovarian follicles. In cultured granulose cells, all the tested hormones promoted release of both P<sub>4</sub> and IGF-I. Food restriction/serum deprivation reduced both P<sub>4</sub> and IGF-I output. Additions of either IGF-I, leptin and FSH prevented the inhibitory action of malnutrition on both P<sub>4</sub> and IGF-I release. The present observations (1) confirm the involvement of the hormones IGF-I, leptin and FSH in the control of secretory activity of ovarian cells, (2) demonstrate, that both isolated ovarian granulosa cells and whole follicles cultured in the absence of serum nutrients could be an adequate in-vitro model for studying the effect of malnutrition on ovarian secretory functions, and (3) suggest, that malnutrition could affect ovarian functions through changes in the release of ovarian hormones.

Effects of recombinant bovine somatotrophin, insulin-like growth factor-I and insulin on the proliferation of bovine granulosa cells in vitro

1993 ◽  
Vol 139 (1) ◽  
pp. 67-75 ◽  
Author(s):  
J. G. Gong ◽  
D. McBride ◽  
T. A. Bramley ◽  
R. Webb
Keyword(s):  
Growth Factor ◽  
Granulosa Cells ◽  
Ovarian Follicles ◽  
Dependent Manner ◽  
Insulin Like Growth Factor ◽  
Factor I ◽  
Igf I ◽  
Dose Dependent ◽  
Bovine Somatotrophin

ABSTRACT Treatment of heifers with recombinant bovine somatotrophin (BST) significantly increases the population of small ovarian follicles and peripheral concentrations of somatotrophin, insulin-like growth factor-I (IGF-I) and insulin. To investigate the possible mechanism(s) involved in the action of BST on ovarian follicles, the effects of BST, IGF-I and insulin, given alone or in combination with either FSH or LH, on the proliferation of bovine granulosa cells in vitro were examined using a serum-free culture system. Bovine granulosa cells were obtained from antral follicles classified into three size categories according to diameter: small <5 mm; medium-sized 5–10 mm and large >10 mm. The proliferation of granulosa cells was assessed by the incorporation of [3H]thymidine into the cultured cells. Both FSH and LH (1–1000 ng/ml) inhibited the proliferation of bovine granulosa cells obtained from all three size classes of follicles in a dose-dependent manner. BST, at doses ranging from 1 to 1000 ng/ml, had no effect on the proliferation of granulosa cells from small and medium-sized follicles, but inhibited the division of granulosa cells from large follicles in a dose-dependent manner. Treatment with either IGF-I (10–3000 ng/ml) or insulin (0·5–1000 ng/ml) stimulated, in a dose-dependent manner, the proliferation of granulosa cells obtained from all three size categories of follicles. No synergistic interaction between BST (30 ng/ml) and either FSH (50 ng/ml) or LH (5 ng/ml) was observed in granulosa cells from all three size classes of follicles. In contrast, physiological concentrations of both IGF-I (100 ng/ml) and insulin (1 ng/ml) acted in synergy with both FSH (50 ng/ml) and LH (5 ng/ml) to stimulate the proliferation of granulosa cells from small follicles, whilst no such synergistic interactions were observed in granulosa cells from medium-sized and large follicles. It was concluded that the increase in the number of small ovarian follicles induced by BST treatment in heifers may be mediated by increased peripheral concentrations of IGF-I and/or insulin, possibly acting in synergy with gonadotrophins. Furthermore, insulin probably acts through its own receptor rather than acting via the type-I IGF receptor, as it can stimulate the proliferation of bovine granulosa cells at physiological concentrations. Journal of Endocrinology (1993) 139, 67–75

scholarly journals Conformation-Specific Inhibitory Anti-MMP-7 Monoclonal Antibody Sensitizes Pancreatic Ductal Adenocarcinoma Cells to Chemotherapeutic Cell Kill

Cancers ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1679
Author(s):  
Vishnu Mohan ◽  
Jean P. Gaffney ◽  
Inna Solomonov ◽  
Maxim Levin ◽  
Mordehay Klepfish ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Active Site ◽  
Drug Targets ◽  
Fas Ligand ◽  
Specific Activity ◽  
Matrix Metalloproteases ◽  
Ductal Adenocarcinoma ◽  
Enzyme Active Site ◽  
Induced Apoptosis

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity.

Encircling granulosa cells protects against di-(2-ethylhexyl)phthalate-induced apoptosis in rat oocytes cultured in vitro

Zygote ◽  
2019 ◽  
Vol 27 (4) ◽  
pp. 203-213 ◽  
Author(s):  
Anima Tripathi ◽  
Vivek Pandey ◽  
A.N. Sahu ◽  
Alok K. Singh ◽  
Pawan K. Dubey
Keyword(s):  
Oxidative Stress ◽  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Granulosa Cells ◽  
Mitochondrial Membrane ◽  
Dependent Manner ◽  
Apoptotic Markers ◽  
Induced Apoptosis ◽  
Ethylhexyl Phthalate

SummaryThe present study investigated if the presence of encircling granulosa cells protected against di(2-ethylhexyl)phthalate (DEHP)-induced oxidative stress in rat oocytes cultured in vitro. Denuded oocytes and cumulus–oocyte complexes (COCs) were treated with or without various doses of DEHP (0.0, 25.0, 50.0, 100, 200, 400 and 800 μM) in vitro. Morphological apoptotic changes, levels of oxidative stress and reactive oxygen species (ROS), mitochondrial membrane potential, and expression levels of apoptotic markers (Bcl2, Bax, cytochrome c) were analyzed. Our results showed that DEHP induced morphological apoptotic changes in a dose-dependent manner in denuded oocytes cultured in vitro. The effective dose of DEHP (400 µg) significantly (P>0.05) increased oxidative stress by elevating ROS levels and the mitochondrial membrane potential with higher mRNA expression and protein levels of apoptotic markers (Bax, cytochrome c). Encircling granulosa cells protected oocytes from DEHP-induced morphological changes, increased oxidative stress and ROS levels, as well as increased expression of apoptotic markers. Taken together our data suggested that encircling granulosa cells protected oocytes against DEHP-induced apoptosis and that the presence of granulosa cells could act positively towards the survival of oocytes under in vitro culture conditions and may be helpful during assisted reproductive technique programmes.

Presence of encircling granulosa cells protects against oxidative stress-induced apoptosis in rat eggs cultured in vitro

APOPTOSIS ◽  
2016 ◽  
Vol 22 (1) ◽  
pp. 98-107 ◽  
Author(s):  
Meenakshi Tiwari ◽  
Anima Tripathi ◽  
Shail K. Chaube
Keyword(s):  
Oxidative Stress ◽  
Granulosa Cells ◽  
Rat Eggs ◽  
Induced Apoptosis

scholarly journals MicroRNA-204-5p regulates apoptosis by targeting Bcl2 in rat ovarian granulosa cells exposed to cadmium†

2020 ◽  
Vol 103 (3) ◽  
pp. 608-619
Author(s):  
Ping Zhong ◽  
Jin Liu ◽  
Hong Li ◽  
Senbin Lin ◽  
Lingfeng Zeng ◽  
...  
Keyword(s):  
Protein Expression ◽  
Granulosa Cells ◽  
B Cell Lymphoma ◽  
Experimental Conditions ◽  
Expression Levels ◽  
Ovarian Granulosa Cells ◽  
Mrna And Protein Expression ◽  
Induced Apoptosis ◽  
Apoptosis Regulation

Abstract This study aimed to investigate whether cadmium (Cd) cytotoxicity in rat ovarian granulosa cells (OGCs) is mediated through apoptosis or autophagy and to determine the role of microRNAs (miRNAs) in Cd cytotoxicity. To test this hypothesis, rat OGCs were exposed to 0, 10, and 20 μM CdCl2 in vitro. As the Cd concentration increased, OGC apoptosis increased. In addition, Cd promoted apoptosis by decreasing the mRNA and protein expression levels of inhibition of B-cell lymphoma 2 (Bcl2). However, under our experimental conditions, no autophagic changes in rat OGCs were observed, and the mRNA and protein expression levels of the autophagic markers microtubule-associated protein 1 light chain 3 alpha (Map1lc3b) and Beclin1 (Becn1) were not changed. Microarray chip analysis, miRNA screening, and bioinformatics approaches were used to further explore the roles of apoptosis regulation-related miRNAs. In total, 19 miRNAs putatively related to Cd-induced apoptosis in rat OGCs were identified. Notably, miR-204-5p, which may target Bcl2, was identified. Then, rat OGCs were cultured in vitro and used to construct the miR-204-5p-knockdown cell line LV2-short hairpin RNA (shRNA). LV2-shRNA cells were exposed to 20 μM Cd for 12 h, and the mRNA and protein expression levels of Bcl2 were increased. Our findings suggest that Cd is cytotoxic to rat OGCs, and mitochondrial apoptosis rather than autophagy mediates Cd-induced damage to OGCs. Cd also affects apoptosis-related miRNAs, and the underlying apoptotic mechanism may involve the Bcl2 gene.

Somatomedin C augments FSH-induced differentiation of cultured rat granulosa cells

1985 ◽  
Vol 249 (1) ◽  
pp. E26-E33 ◽  
Author(s):  
J. B. Davoren ◽  
J. W. Hsueh ◽  
C. H. Li
Keyword(s):  
Granulosa Cells ◽  
Full Range ◽  
Cell Number ◽  
Cellular Protein ◽  
Aromatase Activity ◽  
Concomitant Treatment ◽  
Somatomedin C ◽  
Estrogen Production ◽  
Igf I

Growth hormone (GH) deficiency in rats is associated with decreased ovarian steroidal responsiveness to gonadotropins, possibly through a reduction in the production of the GH-dependent Somatomedin C/insulinlike growth factor I (SM C/IGF I). We have investigated the direct effects of synthetic SM C/IGF I on gonadotropin-stimulated ovarian steroidogenesis in vitro. Granulosa cells were cultured in a serum-free medium for 48 h in the presence of follicle-stimulating hormone (FSH), with or without SM C/IGF I. FSH dose-dependently increased both estrogen and progestin production. Concomitant treatment with SM C/IGF I led to a dose-dependent augmentation of progestin secretion over the full range of FSH doses tested, by a maximum of 2.3- to 2.6-fold. FSH-stimulated estrogen was enhanced by up to 2.4-fold but only at low doses of FSH. SM C/IGF I-enhanced progestin production was associated with increased pregnenolone production and 3 beta-hydroxysteroid dehydrogenase activity, whereas augmented estrogen production appeared to be due to enhanced aromatase activity. The actions of SM C/IGF I, at physiologically relevant concentrations were correlated with increased extracellular cAMP accumulation and cellular protein content but were independent of any change in cell number or viability. In contrast to SM C/IGF I, the closely related peptide multiplication-stimulating activity decreased estrogen production while increasing progestin metabolite accumulation. The present results indicate that the GH-dependent peptide SM C/IGF I may play a role in ovarian development by enhancing gonadotropin-stimulated granulosa cell steroidogenesis.

Alterations in the cell cycle of mouse cumulus granulosa cells during expansion and mucification in vivo and in vitro

1996 ◽  
Vol 8 (6) ◽  
pp. 935 ◽  
Author(s):  
AW Schuetz ◽  
DG Whittingham ◽  
R Snowden
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Granulosa Cells ◽  
Dna Content ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Ovarian Follicles ◽  
S Phase

The cell cycle characteristics of mouse cumulus granulosa cells were determined before, during and following their expansion and mucification in vivo and in vitro. Cumulus-oocyte complexes (COC) were recovered from ovarian follicles or oviducts of prepubertal mice previously injected with pregnant mare serum gonadotrophin (PMSG) or a mixture of PMSG and human chorionic gonadotrophin (PMSG+hCG) to synchronize follicle differentiation and ovulation. Cell cycle parameters were determined by monitoring DNA content of cumulus cell nuclei, collected under rigorously controlled conditions, by flow cytometry. The proportion of cumulus cells in three cell cycle-related populations (G0/G1; S; G2/M) was calculated before and after exposure to various experimental conditions in vivo or in vitro. About 30% of cumulus cells recovered from undifferentiated (compact) COC isolated 43-45 h after PMSG injections were in S phase and 63% were in G0/G1 (2C DNA content). Less than 10% of the cells were in the G2/M population. Cell cycle profiles of cumulus cells recovered from mucified COC (oviducal) after PMSG+hCG-induced ovulation varied markedly from those collected before hCG injection and were characterized by the relative absence of S-phase cells and an increased proportion of cells in G0/G1. Cell cycle profiles of cumulus cells collected from mucified COC recovered from mouse ovarian follicles before ovulation (9-10 h after hCG) were also characterized by loss of S-phase cells and an increased G0/G1 population. Results suggest that changes in cell cycle parameters in vivo are primarily mediated in response to physiological changes that occur in the intrafollicular environment initiated by the ovulatory stimulus. A similar lack of S-phase cells was observed in mucified cumulus cells collected 24 h after exposure in vitro of compact COC to dibutyryl cyclic adenosine monophosphate (DBcAMP), follicle-stimulating hormone or epidermal growth factor (EGF). Additionally, the proportion of cumulus cells in G2/M was enhanced in COC exposed to DBcAMP, suggesting that cell division was inhibited under these conditions. Thus, both the G1-->S-phase and G2-->M-phase transitions in the cell cycle appear to be amenable to physiological regulation. Time course studies revealed dose-dependent changes in morphology occurred within 6 h of exposure in vitro of COC to EGF or DBcAMP. Results suggest that the disappearance of the S-phase population is a consequence of a decline in the number of cells beginning DNA synthesis and exit of cells from the S phase following completion of DNA synthesis. Furthermore, loss of proliferative activity in cumulus cells appears to be closely associated with COC expansion and mucification, whether induced under physiological conditions in vivo or in response to a range of hormonal stimuli in vitro. The observations indicate that several signal-transducing pathways mediate changes in cell cycle parameters during cumulus cell differentiation.

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