scholarly journals Dynamic characteristics of lipid metabolism in cultured granulosa cells from geese follicles at different developmental stages

2019 ◽  
Vol 39 (12) ◽  
Author(s):  
Shanyan Gao ◽  
Xiang Gan ◽  
Hua He ◽  
Shenqiang Hu ◽  
Yan Deng ◽  
...  
Keyword(s):  
Lipid Metabolism ◽  
Granulosa Cells ◽  
Dynamic Characteristics ◽  
Developmental Stages ◽  
Lipid Synthesis ◽  
Follicular Development ◽  
Vital Role ◽  
Intracellular Lipids ◽  
Expression Of Genes

Abstract Previous studies have shown that lipid metabolism in granulosa cells (GCs) plays a vital role during mammalian ovarian follicular development. However, little research has been done on lipid metabolism in avian follicular GCs. The goal of the present study was to investigate the dynamic characteristics of lipid metabolism in GCs from geese pre-hierarchical (6–10 mm) and hierarchical (F4-F2 and F1) follicles during a 6-day period of in vitro culture. Oil red O staining showed that with the increasing incubation time, the amount of lipids accumulated in three cohorts of GCs increased gradually, reached the maxima after 96 h of culture, and then decreased. Moreover, the lipid content varied among these three cohorts, with the highest in F1 GCs. The qPCR results showed genes related to lipid synthesis and oxidation were highest expressed in pre-hierarchical GCs, while those related to lipid transport and deposition were highest expressed in hierarchical GCs. These results suggested that the amount of intracellular lipids in GCs increases with both the follicular diameter and culture time, which is accompanied by significant changes in expression of genes related to lipid metabolism. Therefore, it is postulated that the lipid accumulation capacity of geese GCs depends on the stage of follicle development and is finely regulated by the differential expression of genes related to lipid metabolism.

Functional effects of Tribbles homolog 2 in bovine ovarian granulosa cells†

2020 ◽  
Vol 102 (6) ◽  
pp. 1177-1190
Author(s):  
Aly Warma ◽  
Kalidou Ndiaye
Keyword(s):  
Western Blot ◽  
Granulosa Cells ◽  
Developmental Stages ◽  
Polyclonal Antibodies ◽  
Differentially Expressed Gene ◽  
Follicular Development ◽  
Mapk Signaling ◽  
Tissue Growth ◽  
Ovulatory Follicles

Abstract Tribbles homologs (TRIB) 1, 2, and 3 represent atypical members of the serine/threonine kinase superfamily. We previously identified TRIB2 as a differentially expressed gene in granulosa cells (GCs) of bovine preovulatory follicles. The current study aimed to further investigate TRIB2 regulation and study its function in the ovary. GCs were collected from follicles at different developmental stages: small antral follicles (SF), dominant follicles (DF) at day 5 of the estrous cycle, and hCG-induced ovulatory follicles (OFs). RT-qPCR analyses showed greater expression of TRIB2 in GC of DF as compared to OF and a significant downregulation of TRIB2 steady-state mRNA amounts by hCG/LH, starting at 6 h through 24 h post-hCG as compared to 0 h. Specific anti-TRIB2 polyclonal antibodies were generated and western blot analysis confirmed TRIB2 downregulation by hCG at the protein level. In vitro studies showed that FSH stimulates TRIB2 expression in GC. Inhibition of TRIB2 using CRISPR/Cas9 resulted in a significant increase in PCNA expression and an increase in steroidogenic enzyme CYP19A1 expression, while TRIB2 overexpression tended to decrease GC proliferation. TRIB2 inhibition also resulted in a decrease in transcription factors connective tissue growth factor (CTGF) and ankyrin repeat domain-containing protein 1 (ANKRD1) expression, while TRIB2 overexpression increased CTGF and ANKRD1. Additionally, western blot analyses showed reduction in ERK1/2 (MAPK3/1) and p38MAPK (MAPK14) phosphorylation levels following TRIB2 inhibition, while TRIB2 overexpression increased p-ERK1/2 and p-p38MAPK. These results provide evidence that TRIB2 modulates MAPK signaling in GC and that TRIB2 could act as a regulator of GC proliferation and function, which could affect steroidogenesis during follicular development.

scholarly journals Kit ligand and c-Kit are expressed during early human ovarian follicular development and their interaction is required for the survival of follicles in long-term culture

Reproduction ◽  
2006 ◽  
Vol 131 (4) ◽  
pp. 641-649 ◽  
Author(s):  
Inger B Carlsson ◽  
Mika P E Laitinen ◽  
Jennifer E Scott ◽  
Henna Louhio ◽  
Louiza Velentzis ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Developmental Stages ◽  
Culture Media ◽  
Follicular Development ◽  
Ovarian Follicles ◽  
Kit Ligand ◽  
Kit Receptor ◽  
Ovarian Follicular Development

The receptor tyrosine c-Kit and its cognate ligand, c-Kit ligand (KL, stem cell factor, SCF), are involved in ovarian follicular development in several animal species. We studied the expression of KL and c-Kit usingin situhybridization and immunohistochemistry in donated human ovarian cortical tissue. The KL transcripts were expressed in granulosa cells of primary follicles, whereas the expression of c-Kit was confined to the oocyte and granulosa cells in primary and secondary follicles. We employed an ovarian organ culture using firstly serum-containing and then serum-free medium to study the effects of KL and an anti-c-Kit antibody, ACK2, on the development and survival of ovarian folliclesin vitro. Culture of ovarian cortical slices for 7 days resulted in a 37% increase in the number of primary follicles and a 6% increase in secondary follicles. The proportion of viable follicles decreased in all cultures. The addition of KL (1, 10 and 100 ng/ml) into the culture media did not affect the developmental stages of the follicles or the proportion of atretic follicles. Inclusion of ACK2 (800 ng/ml) in the culture medium significantly increased the proportion of atretic follicles on days 7 (49 vs 28% in control cultures) and 14 (62 vs 38%) of culture. In conclusion, c-Kit and KL are expressed in human ovaries during follicular development. Blocking the c-Kit receptor induces follicular atresia. The KL/c-Kit signaling system is likely to control the survival of human ovarian follicles during early follicular development.

scholarly journals The Differential Metabolomes in Cumulus and Mural Granulosa Cells from Human Preovulatory Follicles

2021 ◽  
Author(s):  
Er-Meng Gao ◽  
Bongkoch Turathum ◽  
Ling Wang ◽  
Di Zhang ◽  
Yu-Bing Liu ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Cumulus Cells ◽  
Cholesterol Transport ◽  
Vitro Fertilization ◽  
Expression Of Genes ◽  
Preovulatory Follicles ◽  
Morphological Differences

AbstractThis study evaluated the differences in metabolites between cumulus cells (CCs) and mural granulosa cells (MGCs) from human preovulatory follicles to understand the mechanism of oocyte maturation involving CCs and MGCs. CCs and MGCs were collected from women who were undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. The differences in morphology were determined by immunofluorescence. The metabolomics of CCs and MGCs was measured by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) followed by quantitative polymerase chain reaction (qPCR) and western blot analysis to further confirm the genes and proteins involved in oocyte maturation. CCs and MGCs were cultured for 48 h in vitro, and the medium was collected for detection of hormone levels. There were minor morphological differences between CCs and MGCs. LC-MS/MS analysis showed that there were differences in 101 metabolites between CCs and MGCs: 7 metabolites were upregulated in CCs, and 94 metabolites were upregulated in MGCs. The metabolites related to cholesterol transport and estradiol production were enriched in CCs, while metabolites related to antiapoptosis were enriched in MGCs. The expression of genes and proteins involved in cholesterol transport (ABCA1, LDLR, and SCARB1) and estradiol production (SULT2B1 and CYP19A1) was significantly higher in CCs, and the expression of genes and proteins involved in antiapoptosis (CRLS1, LPCAT3, and PLA2G4A) was significantly higher in MGCs. The level of estrogen in CCs was significantly higher than that in MGCs, while the progesterone level showed no significant differences. There are differences between the metabolomes of CCs and MGCs. These differences may be involved in the regulation of oocyte maturation.

Characterization of isolated bovine preantral follicles based on morphology, diameter and cell number

Zygote ◽  
2020 ◽  
Vol 28 (2) ◽  
pp. 154-159
Author(s):  
Juliana I. Candelaria ◽  
Anna C. Denicol
Keyword(s):  
Granulosa Cells ◽  
Developmental Stages ◽  
Assisted Reproductive Technologies ◽  
Reproductive Technologies ◽  
Cell Number ◽  
Culture Conditions ◽  
Preantral Follicles ◽  
Secondary Stage

SummaryPreantral follicles are a potential reservoir of oocytes to be used in assisted reproductive technologies. With the increasing interest in developing techniques to grow preantral follicles in vitro, and as the bovine emerges as an appropriate model species to understand human folliculogenesis, the establishment of an accurate classification of developmental stages is needed. Classification of bovine preantral follicles has been mostly based on histological analysis and estimation models, which may not translate well to correctly characterize preantral follicles isolated from the ovary. In this study, we classified bovine preantral follicles by morphology upon isolation, determined diameter and number of granulosa cells by direct counting, and compared our results with previous studies reporting bovine preantral follicle classification. Follicles were isolated via homogenization of ovary tissue and classified into primary, early secondary and secondary stage based on morphology and number of layers of granulosa cells. Diameter was individually measured and Hoechst 33342 was used as a nuclear stain to count granulosa cells. We found that follicles classified by morphology into primary, early secondary, and secondary had different mean diameter and cell number (P < 0.01); cell number and diameter were positively correlated, as were cell density and cell number in each developmental stage (P < 0.01). Results obtained here were mostly in agreement with previous classifications based on histological sections and on isolated follicles, with some discrepancies. The present data add accuracy to classification of bovine preantral follicles that is critical to optimize culture conditions to produce developmentally competent oocytes.

scholarly journals AKT is involved in granulosa cell autophagy regulation via mTOR signaling during rat follicular development and atresia

Reproduction ◽  
2014 ◽  
Vol 147 (1) ◽  
pp. 73-80 ◽  
Author(s):  
JongYeob Choi ◽  
MinWha Jo ◽  
EunYoung Lee ◽  
DooSeok Choi
Keyword(s):  
Cell Death ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Apoptotic Cell ◽  
Follicular Development ◽  
Negative Regulator ◽  
Metabolic Clearance ◽  
Akt Inhibitor ◽  
Western Blots

In this study, we examined whether granulosa cell autophagy during follicular development and atresia was regulated by the class I phosphoinositide-3 kinase/protein kinase B (AKT) pathway, which is known to control the activity of mammalian target of rapamycin (mTOR), a major negative regulator of autophagy. Ovaries and granulosa cells were obtained using an established gonadotropin-primed immature rat model that induces follicular development and atresia. Autophagy was evaluated by measuring the expression level of microtubule-associated protein light chain 3-II (LC3-II) using western blots and immunohistochemistry. The activity of AKT and mTOR was also examined by observing the phosphorylation of AKT and ribosomal protein S6 kinase (S6K) respectively. After gonadotropin injection, LC3-II expression was suppressed and phosphorylation of AKT and S6K increased in rat granulosa cells. By contrast, gonadotropin withdrawal by metabolic clearance promoted LC3-II expression and decreased phosphorylation of AKT and S6K. In addition,in-vitroFSH treatment of rat granulosa cells also indicated inhibition of LC3-II expression accompanied by a marked increase in phosphorylation of AKT and S6K. Inhibition of AKT phosphorylation using AKT inhibitor VIII suppressed FSH-mediated phosphorylation of S6K, followed by an increase in LC3-II expression. Furthermore, co-treatment with FSH and AKT inhibitor increased the levels of apoptosis and cell death of granulosa cells compared with the single treatment with FSH. Taken together, our findings indicated that AKT-mediated activation of mTOR suppresses granulosa cell autophagy during follicular development and is involved in the regulation of apoptotic cell death.

Identification of Gαs messenger ribonucleic acid splice variants in human granulosa cells

1997 ◽  
Vol 18 (1) ◽  
pp. 27-35 ◽  
Author(s):  
G N Europe-Finner ◽  
E Cartwright ◽  
J Bellinger ◽  
H J Mardon ◽  
D H Barlow ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Ovarian Function ◽  
Splice Variants ◽  
Consensus Sequence ◽  
Phosphorylation Site ◽  
Follicular Development ◽  
Protein Isoforms ◽  
Mrna Isoforms ◽  
Messenger Ribonucleic Acid

ABSTRACT Granulosa cells are essential for follicular development and corpus luteum formation and their functions are regulated by gonadotrophins through G protein-coupled receptors. The dominant second messenger pathway involves the stimulation of cyclic AMP formation by Gαs-linked receptors. In this paper we have investigated the expression of Gαs mRNA splice variants in relation to expression of Gαs protein isoforms in granulosa cells obtained from patients undergoing in vitro fertilization. We have carried out ribonuclease protection assays using cRNA riboprobes which are capable of detecting all Gαs mRNA isoforms as well as quantifying total amounts of Gαs mRNA. Granulosa cells express the message for Gαs-Large and Gαs-Small and the presence of two distinct protein products was confirmed by immunoblotting using the antibody RM/1. Moreover, the data show that a significant fraction of Gαs-Large and Gαs-Small mRNAs contain an extra CAG codon. This should generate proteins with an extra serine residue, resulting in Gαs variants with the consensus sequence of a protein kinase C phosphorylation site. These results highlight the possible interaction between different signalling pathways in the control of cAMP production and the need to investigate the relationship between Gαs variants and different adenylyl cyclase isozymes in patients with normal and abnormal ovarian function.

scholarly journals INTRODUCING IN VITRO EXPERIMENTS OF OXYGEN BUBBLES SHOCKWAVES TRIGGERING INTRACELLULAR LIPIDS LUMINESCENCE: IMPLICATIONS IN CANCER ETIOLOGY

2019 ◽  
Vol 7 (4) ◽  
pp. 355-364
Author(s):  
Abraham A.
Keyword(s):  
Lipid Metabolism ◽  
Lipid Droplets ◽  
Cancer Metabolism ◽  
Light Emission ◽  
Direct Consequence ◽  
Static Electricity ◽  
Cancer Etiology ◽  
H2o2 Decomposition ◽  
Intracellular Lipids

Background The main purpose of this manuscript is to introduce a mechanism supporting a previously hypothesized factor in cancer origin, where endogenous energy emission during cell respiration was identified as additional factor in cancer origin. Recent published reports identify the pressure profile of shockwaves as causing lipid droplets membrane deformation. Lipid metabolism has been highlighted to have a key role in cancer metabolism, and metastasis; for example, several publications have suggested targeting lipid metabolism of cancer cells as a strategy to control metastasis. New studies have revealed that lipid layers are responsible for the storage and discharge of static electricity. This manuscript introduces shockwaves from oxygen bubbles bursts as a mechanism causing intracellular lipids discharge or static electricity. The effect causes shape changes of lipid droplets up to a light emission stage. Materials and Methods Cheek cells intracellular material, including DNA strands and lipid droplets were precipitated in a test tube by following written instructions on DNA precipitation published online by The University of Michigan. The DNA precipitate was transferred onto a clean glass slide and covered by a similar one and dubbed a sandwich (SDW).  A slide assembly was developed where the effect of oxygen bubbles cavitation-induced shockwaves on the trapped DNA precipitate and lipid droplets were recorded. Microphotographs and video recordings were stored in a computer via a video-microscope. Results Lipid droplets exposed to prolonged shockwaves energy were documented to undergo recurrent expanding architectural deformation up to a final contracting phase where light was emitted.  Conclusions Intracellular lipid droplets are ubiquitously present in cells; and recent research has shown their expanded roles in cellular signaling in both mitotic and non-mitotic cells. In cancer, one highlighted key role is the potential of lipid metabolism in metastatic colonization. Data introduced in this manuscript demonstrates a direct consequence of ROS (H2O2) decomposition (via oxygen bubbles bursts) as a trigger for lipid intracellular droplets emission of light radiation, thus supporting a previously proposed biophysical mechanism in cancer origin.

GPR120 agonist ameliorated insulin resistance and improved ovarian function

Zygote ◽  
2021 ◽  
pp. 1-6
Author(s):  
Yang Liu ◽  
Jiayi Ding ◽  
Xiaofang Tan ◽  
Ya Shen ◽  
Li Xu ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Lipid Metabolism ◽  
Rat Model ◽  
Lipid Accumulation ◽  
Ovarian Function ◽  
Polycystic Ovary ◽  
Vital Role ◽  
Expression Levels ◽  
Glucose And Lipid Metabolism

Summary GPR120 is implicated in the regulation of glucose and lipid metabolism, and insulin resistance. In the current study, we aimed to investigate the role of GPR120 in polycystic ovary syndrome (PCOS). With the adoption of dehydroepiandrosterone, a rat model was established to simulate PCOS in vitro. mRNA and protein expression levels of GPR120 were measured using RT-qPCR and western blot, respectively. In addition, expression levels of testosterone, estradiol, luteinizing hormone and follicle-stimulating hormone, serum total cholesterol and triglyceride were assessed using the corresponding kits. Moreover, haematoxylin and eosin staining was used to detect pathological changes in ovary or liver and oil red staining was utilized to evaluate lipid accumulation. In the present study, GPR120 was downregulated in plasma, liver and ovary in the PCOS rat model. In addition, the GPR120 agonist regulated lipid metabolism in the liver and weight in the PCOS rat model. Furthermore, the GPR120 agonist decreased insulin resistance in the PCOS rat model but improved the ovarian function. It is suggested that GPR120 plays a vital role in suppressing insulin resistance, regulating ovary function and decreasing lipid accumulation in the liver, demonstrating that targeting GPR120 could be an effective method for the improvement of PCOS.

scholarly journals SCAP/SREBPs are Central Players in Lipid Metabolism and Novel Metabolic Targets in Cancer Therapy

2018 ◽  
Vol 18 (6) ◽  
pp. 484-493 ◽  
Author(s):  
Xiang Cheng ◽  
Jianying Li ◽  
Deliang Guo
Keyword(s):  
Lipid Metabolism ◽  
Cancer Therapy ◽  
Transcriptional Activation ◽  
Molecular Mechanisms ◽  
Regulatory Element ◽  
Lipid Synthesis ◽  
Feedback Regulation ◽  
Negative Feedback Regulation ◽  
Expression Of Genes ◽  
New Findings

Lipid metabolism reprogramming emerges as a new hallmark of malignancies. Sterol regulatory element-binding proteins (SREBPs), which are central players in lipid metabolism, are endoplasmic reticulum (ER)-bound transcription factors that control the expression of genes important for lipid synthesis and uptake. Their transcriptional activation requires binding to SREBP cleavageactivating protein (SCAP) to translocate their inactive precursors from the ER to the Golgi to undergo cleavage and subsequent nucleus translocation of their NH2-terminal forms. Recent studies have revealed that SREBPs are markedly upregulated in human cancers, providing the mechanistic link between lipid metabolism alterations and malignancies. Pharmacological or genetic inhibition of SCAP or SREBPs significantly suppresses tumor growth in various cancer models, demonstrating that SCAP/SREBPs could serve as promising metabolic targets for cancer therapy. In this review, we will summarize recent progress in our understanding of the underlying molecular mechanisms regulating SCAP/SREBPs and lipid metabolism in malignancies, discuss new findings about SREBP trafficking, which requires SCAP N-glycosylation, and introduce a newly identified microRNA-29-mediated negative feedback regulation of the SCAP/SREBP pathway. Moreover, we will review recently developed inhibitors targeting the SCAP/SREBP pathway for cancer treatment.

scholarly journals Pentachloronitrobenzene alters progesterone production and primordial follicle recruitment in cultured granulosa cells and rat ovary†

2019 ◽  
Vol 102 (2) ◽  
pp. 511-520
Author(s):  
Yanrong Kuai ◽  
Xiaobo Gao ◽  
Huixia Yang ◽  
Haiyan Luo ◽  
Yang Xu ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Granulosa Cells ◽  
Crop Production ◽  
Follicular Development ◽  
Mitogen Activated Protein Kinase ◽  
Dependent Manner ◽  
Serum Progesterone ◽  
Primordial Follicles ◽  
Progesterone Production

Abstract Pentachloronitrobenzene (PCNB) is an organochlorine fungicide widely used for crop production and has become an environmental concern. Little is known about the effect of PCNB on ovarian steroidogenesis and follicular development. We found that PCNB stimulated Star expression and progesterone production in cultured rat granulosa cells in a dose-dependent manner. PCNB activated mitogen-activated protein kinase (MAPK3/1) extracellulat regulated kinase (ERK1/2), thus inhibition of either protein kinase A (PKA) or MAPK3/1 signaling pathway significantly attenuated progesterone biosynthesis caused by PCNB, suggesting that PCNB induced progesterone production by activating the cyclic adenosine monophosphate (cAMP/PKA) and MAPK3/1 signaling pathways. Further investigation demonstrated that PCNB induced Star expression and altered MAPK3/1 signaling in ovary tissues of immature SD rats treated with PCNB at the dose of 100, 200, or 300 mg/kg by daily gavage for 7 days, while serum progesterone level was dose-dependently decreased. We demonstrated that PCNB exposure accelerated the recruitment of primordial follicles into the growing follicle pool in ovary tissues, accompanied by increased levels of anti-Mullerian hormone (AMH) in both ovary tissues and serum. Taken together, our data demonstrate for the first time that PCNB stimulated Star expression, altered MAPK3/1 signaling and progesterone production in vivo and in vitro, and accelerated follicular development with a concomitant increase in AMH in ovary tissues and serum. Our findings provide novel insight into the toxicity of PCNB to animal ovary function.

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