SUN-244 Role of Activin, Follistatin, and Inhibin in the Regulation of KISS-1 Gene Expression in Hypothalamic Cell Models

Abstract Kisspeptin (encoded by the Kiss-1 gene) in the arcuate nucleus (ARC) of the hypothalamus governs the hypothalamic-pituitary-gonadal (HPG) axis by regulating pulsatile release of gonadotropin-releasing hormone (GnRH). Meanwhile, kisspeptin in the anteroventral periventricular nucleus (AVPV) region has been implicated in estradiol (E2)-induced GnRH surges. Kiss-1-expressing cell model mHypoA-55 exhibits characteristics of Kiss-1 neurons in the ARC region. On the other hand, Kiss-1 expressing mHypoA-50 cells originate from the AVPV region. In the mHypoA-55 ARC cells, activin significantly increased Kiss-1 gene expression. Follistatin alone reduced Kiss-1 expression within these cells. Interestingly, activin-induced Kiss-1 gene expression was completely abolished by follistatin. Inhibin A, but not inhibin B reduced Kiss-1 expression. Activin-increased Kiss-1 expression was also abolished by inhibin A. Pretreatment of the cells with follistatin or inhibin A significantly inhibited kisspeptin- or GnRH-induced Kiss-1 gene expression in mHypoA-55 cells. In contrast, in the mHypoA-50 AVPV cell model, activin, follistatin, and inhibin A did not modulate Kiss-1 gene expression. The subunits that compose activin and inhibin, as well as follistatin were expressed in both mHypoA-55 and mHypoA-50 cells. Expression of inhibin βA and βB subunits and follistatin was much higher in mHypoA-55 ARC cells. Furthermore, we found that expression of the inhibin αsubunit and follistatin genes was modulated in the presence of E2 in mHypoA-55 ARC cells. The results of this study suggest that activin, follistatin, and inhibin A within the ARC region participate in the regulation of the HPG axis under the influence of E2.

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Role of activin, follistatin, and inhibin in the regulation of Kiss-1 gene expression in hypothalamic cell models†

Biology of Reproduction ◽

10.1093/biolre/ioz094 ◽

2019 ◽

Vol 101 (2) ◽

pp. 405-415 ◽

Cited By ~ 2

Author(s):

Zolzaya Tumurgan ◽

Haruhiko Kanasaki ◽

Tuvshintugs Tumurbaatar ◽

Aki Oride ◽

Hiroe Okada ◽

...

Keyword(s):

Gene Expression ◽

Arcuate Nucleus ◽

Cell Model ◽

Gonadotropin Releasing Hormone ◽

Cell Models ◽

Inhibin A ◽

Hpg Axis ◽

Α Subunit ◽

Periventricular Nucleus

Abstract Kisspeptin (encoded by the Kiss-1 gene) in the arcuate nucleus (ARC) of the hypothalamus governs the hypothalamic-pituitary-gonadal (HPG) axis by regulating pulsatile release of gonadotropin-releasing hormone (GnRH). Meanwhile, kisspeptin in the anteroventral periventricular nucleus (AVPV) region has been implicated in estradiol (E2)-induced GnRH surges. Kiss-1–expressing cell model mHypoA-55 exhibits characteristics of Kiss-1 neurons in the ARC region. On the other hand, Kiss-1 expressing mHypoA-50 cells originate from the AVPV region. In the mHypoA-55 ARC cells, activin significantly increased Kiss-1 gene expression. Follistatin alone reduced Kiss-1 expression within these cells. Interestingly, activin-induced Kiss-1 gene expression was completely abolished by follistatin. Inhibin A, but not inhibin B reduced Kiss-1 expression. Activin-increased Kiss-1 expression was also abolished by inhibin A. Pretreatment of the cells with follistatin or inhibin A significantly inhibited kisspeptin- or GnRH-induced Kiss-1 gene expression in mHypoA-55 cells. In contrast, in the mHypoA-50 AVPV cell model, activin, follistatin, and inhibin A did not modulate Kiss-1 gene expression. The subunits that compose activin and inhibin, as well as follistatin were expressed in both mHypoA-55 and mHypoA-50 cells. Expression of inhibin βA and βB subunits and follistatin was much higher in mHypoA-55 ARC cells. Furthermore, we found that expression of the inhibin α subunit and follistatin genes was modulated in the presence of E2 in mHypoA-55 ARC cells. The results of this study suggest that activin, follistatin, and inhibin A within the ARC region participate in the regulation of the HPG axis under the influence of E2.

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Role of RFRP-3 in the Regulation of Kiss-1 Gene Expression in the AVPV Hypothalamic Cell Model mHypoA-50

Reproductive Sciences ◽

10.1177/1933719118813456 ◽

2018 ◽

Vol 26 (9) ◽

pp. 1249-1255 ◽

Cited By ~ 2

Author(s):

Haruhiko Kanasaki ◽

Tuvshintugs Tumurbaatar ◽

Aki Oride ◽

Zolzaya Tumurgan ◽

Hiroe Okada ◽

...

Keyword(s):

Gene Expression ◽

Positive Feedback ◽

Messenger Rna ◽

Cell Model ◽

Reproductive Function ◽

Feedback Regulation ◽

Gonadotropin Releasing Hormone ◽

Releasing Hormone ◽

Gonadotropin Inhibitory Hormone

Kisspeptin, encoded by the Kiss-1 gene, plays a crucial role in reproductive function by governing the hypothalamic–pituitary–gonadal axis. The recently established Kiss-1-expressing cell model mHypoA-50 displays characteristics of neuronal cells of the anteroventral periventricular (AVPV) region of the mouse hypothalamus. Because Kiss-1 gene expression in these cells is upregulated by estradiol (E2), mHypoA-50 cells are regarded as a valuable model for the study of Kiss-1-expressing neurons in the AVPV region. These cells also express RFamide-related peptide-3 (RFRP-3), a mammalian homolog of gonadotropin inhibitory hormone. The RFRP-3 expression in mHypoA-50 cells was increased by melatonin stimulation. In addition, E2 stimulation increased RFRP-3 expression in these cells. Treatment of the mHypoA-50 cells with exogenous RFRP-3 resulted in the increase of Kiss-1 messenger RNA expression within the cells; however, RFRP-3 did not modify gonadotropin-releasing hormone or kisspeptin-induced Kiss-1 gene expression in these cells. In addition, we found that RFRP-3 stimulation increased the expression of corticotropin-releasing hormone, which may be involved in E2-induced positive feedback in mHypoA-50 cells. Our observations suggest that RFRP-3 might be involved in positive feedback regulation by directly or indirectly increasing Kiss-1 gene expression.

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GABAB receptor antagonism from birth to weaning permanently modifies Kiss1 expression in hypothalamus and gonads in mice

Neuroendocrinology ◽

10.1159/000521649 ◽

2021 ◽

Author(s):

Marianne Bizzozzero Hiriart ◽

Noelia P. Di Giorgio ◽

Carlos Libertun ◽

Victoria A.R. Lux-Lantos

Keyword(s):

Arcuate Nucleus ◽

Gabab Receptor ◽

Seminiferous Tubules ◽

Testis Weight ◽

Hpg Axis ◽

Estrous Cycles ◽

Amino Butyric Acid ◽

Periventricular Nucleus ◽

Puberty Onset ◽

Ovulatory Follicles

Introduction: The kisspeptin gene Kiss1 is expressed in two hypothalamic areas: anteroventral periventricular nucleus/periventricular nucleus (AVPV/PeN) and arcuate nucleus (ARC), and also in gonads. Evidences suggest that gamma-amino butyric acid B receptors (GABAB) signaling can regulate Kiss1 expression. Here we inhibited GABAB signaling from PND2-PND21 and evaluated the hypothalamic-pituitary-gonadal (HPG) axis. Methods: BALB/c mice were treated on postnatal days 2-21 (PND2-PND21) with CGP55845 (GABAB antagonist), and evaluated in PND21 and adulthood: gene expression (qPCR) in hypothalamus and gonads, hormones by radioimmunoassay, gonad histochemistry (H&E), puberty onset, estrous cycles. Results: At PND21, CGP inhibited Kiss1 and Tac2 and increased Pdyn and Gabbr1 in the ARC of both sexes and decreased Th only in female AVPV/PeN. Serum follicle-stimulating hormone (FSH) and testis weight decreased in CGP-males and puberty onset was delayed. In adults, Kiss1, Tac2, Pdyn, Pgr, Cyp19a1, Gad1 were downregulated, while Gabbr1 was upregulated in the ARC of both sexes. In the AVPV/PeN, Kiss1, Th, Cyp19a1 and Pgr decreased while Gad1 increased in CGP-females, whereas Cyp19a1 increased in CGP-males. Serum FSH increased in CGP-males while prolactin increased in CGP-females. Testosterone and progesterone increased in ovaries from CGP-females, in which Kiss1, Cyp19a1 and Esr1 were downregulated while Hsd3b2 was upregulated, together with increased atretic and decreased ovulatory follicles. Testes from CGP-males showed decreased progesterone, increased Gabbr1, Kiss1, Kiss1r, Esr2 and decreased Cyp19a1 and clear signs of seminiferous tubules atrophy. Conclusion: These results demonstrate that appropriate GABAB signaling during this critical prepubertal period is necessary for the normal development of the HPG axis.

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Role of 1α,25-Dihydroxyvitamin D3 in Adipogenesis of SGBS Cells: New Insights into Human Preadipocyte Proliferation

Cellular Physiology and Biochemistry ◽

10.1159/000491770 ◽

2018 ◽

Vol 48 (1) ◽

pp. 397-408 ◽

Cited By ~ 8

Author(s):

Ingrid Felicidade ◽

Daniele Sartori ◽

Susan L.M. Coort ◽

Simone Cristine Semprebon ◽

Andressa Megumi Niwa ◽

...

Keyword(s):

Gene Expression ◽

Vitamin D ◽

Adipose Tissue ◽

Lipid Content ◽

Cell Model ◽

Adipogenic Differentiation ◽

Growth Arrest ◽

Proliferation And Differentiation ◽

Sgbs Cells

Background/Aims: Compared with non-obese individuals, obese individuals commonly store more vitamin D in adipose tissue. VDR expression in adipose tissue can influence adipogenesis and is therefore a target pathway deserving further study. This study aims to assess the role of 1,25(OH)2D3 in human preadipocyte proliferation and differentiation. Methods: RTCA, MTT, and trypan blue assays were used to assess the effects of 1,25(OH)2D3 on the viability, proliferation, and adipogenic differentiation of SGBS cells. Cell cycle and apoptosis analyses were performed with flow cytometry, triglycerides were quantified, and RT-qPCR was used to assess gene expression. Results: We confirmed that the SGBS cell model is suitable for studying adipogenesis and demonstrated that the differentiation protocol induces cell maturation, thereby increasing the lipid content of cells independently of treatment. 1,25(OH)2D3 treatment had different effects according to the cell stage, indicating different modes of action driving proliferation and differentiation. In preadipocytes, 1,25(OH)2D3 induced G1 growth arrest at both tested concentrations without altering CDKN1A gene expression. Treatment with 100 nM 1,25(OH)2D3 also decreased MTT absorbance and the lipid concentration. Moreover, increased normalized cell index values and decreased metabolic activity were not induced by proliferation or apoptosis. Exposure to 100 nM 1,25(OH)2D3 induced VDR, CEBPA, and CEBPB expression, even in the preadipocyte stage. During adipogenesis, 1,25(OH)2D3 had limited effects on processes such as VDR and PPARG gene expression, but it upregulated CEBPA expression. Conclusions: We demonstrated for the first time that 1,25(OH)2D3 induces changes in preadipocytes, including VDR expression and growth arrest, and increases the lipid content in adipocytes treated for 16 days. Preadipocytes are important cells in adipose tissue homeostasis, and understanding the role of 1,25(OH)2D3 in adipogenesis is a crucial step in ensuring adequate vitamin D supplementation, especially for obese individuals.

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Role of Metastasis in Hypertabastic Survival Analysis of Breast Cancer: Interaction with Clinical and Gene Expression Variables

Cancer Growth and Metastasis ◽

10.4137/cgm.s8821 ◽

2012 ◽

Vol 5 ◽

pp. CGM.S8821 ◽

Cited By ~ 3

Author(s):

Mohammad A. Tabatabai ◽

Wayne M. Eby ◽

Nadim Nimeh ◽

Karan P. Singh

Keyword(s):

Breast Cancer ◽

Gene Expression ◽

Estrogen Receptor ◽

Cancer Patients ◽

Estrogen Receptor Status ◽

The Other ◽

Primary Concern ◽

Breast Cancer Patients ◽

Receptor Status

This paper analyzes the survival of breast cancer patients, exploring the role of a metastasis variable in combination with clinical and gene expression variables. We use the hypertabastic model in a detailed analysis of 295 breast cancer patients from the Netherlands Cancer Institute given in. 1 In comparison to Cox regression the increase in accuracy is complemented by the ability to analyze the time course of the disease progression using the explicitly described hazard and survival curves. We also demonstrate the ability to compute deciles for survival and probability of survival to a given time. Our primary concern in this article is the introduction of a variable representing the existence of metastasis and the effects on the other clinical and gene expression variables. In addition to making a quantitative assessment of the impact of metastasis on the prospects for survival, we are able to look at its interactions with the other prognostic variables. The estrogen receptor status increase in importance, while the significance of the gene expression variables used in the combined model diminishes. When considering only the subgroup of patients who experienced metastasis, the covariates in the model are only the clinical variables for estrogen receptor status and tumor grade.

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Expression of the CT Antigen, SPAN-Xb, Is Regulated through Promoter Methylation.

Blood ◽

10.1182/blood.v104.11.4285.4285 ◽

2004 ◽

Vol 104 (11) ◽

pp. 4285-4285

Author(s):

Zhiqing Wang ◽

Yana Zhang ◽

Jian Zhang ◽

Seah H. Lim

Keyword(s):

Gene Expression ◽

Sequence Analysis ◽

Promoter Methylation ◽

Cpg Islands ◽

Promoter Sequence ◽

The Other ◽

Gm Csf ◽

Ct Antigen

Abstract SPAN-Xb is a spermatid protein that we have recently identified as a novel Cancer-Testis (CT) antigen in hematologic malignancies. We have also shown that SPAN-Xb expression in tumor cell lines could be upregulated by 5-azacytidine, GM-CSF and IL-7. The ability of 5-azacytidine to increase SPAN-Xb expression suggests that SPAN-Xb gene expression, like the other CT antigens, may be regulated through promoter methylation. On the other hand, the ability of GM-CSF and IL-7 to increase SPAN-Xb expression remained to be determined. In this study, we set out to determine whether or not promoter methylation regulates SPAN-Xb gene expression and the effects of GM-CSF and IL-7 on SPAN-Xb expression is due to their action on the promoter activity. We first isolated and cloned the SPAN-Xb promoter gene into the CAT (chloramphenicol acetyl transferase) reporter system, pCAT*3-Enhancer vector. In vitro methylation was achieved using SssI methylase and the recombinant vectors were transfected into the myeloma cell line, RPMI 8226 cells. CAT activity was assayed in the lysate of the transfectants 48–72 hours after gene transfer. We observed that CAT activitiy in transfectants containining demethylated recombinant pCAT*3-SPAN-Xb promoter vector. In contrast, CAT activity was abrogated once the recombinant vector was methylated in vitro, supporting the role of DNA methylation in the regulation of SPAN-Xb gene expression. CAT activity in the transfectants containing the demethylated vector could be further increased by GM-CSF and IL-7, suggesting that the increase in SPAN-Xb expression we have observed in cells treated with GM-CSF and IL-7 may be the actions of these cytokines on the SPAN-Xb promoter. These cytokines alone, however, were unable to induce CAT activity since transfectants containing the methylated promoter sequence remained negative for the CAT activity even with the addition of GM-CSF or IL-7. To further evaluate the role of DNA methylation on the expression of SPAN-Xb, we carried out the bisulfite conversion assays using genomic DNA from tumor cell lines, normal testis, blood, kidney, pancreas and spleen. Following bisulfite conversion, the modified genomic DNA was subjected to PCR amplification, cloning and sequence analysis. Five clones from each tissues were randomly picked for sequence analysis. A total of 11 CpG islands were identified within the promoter sequence. They were put together into 7 groups according to their positions in the sequence: Group I: −502; Group II: −474; Group III: −450; Group IV: −341; Group V: −311 to −300; Group VI: −226 to −222; Group VII: −184 to −181. Following sequence analysis, we observed that SPAN-Xb expressor (normal testis) was consistently demethylated within Groups V and VI CpG islands. In contrast, SPAN-Xb-negative tissues were consistently methylated at these two CpG islands, localizing the promoter activity of the sequence to these two areas of the promoter. The methylation status at the other CpG island did not predict SPAN-Xb expression. We therefore conclude that: 1. SPAN-Xb expression is regulated by promoter methylation; 2. GM-CSF and IL-7 increase SPAN-Xb expression through their action on the SPAN-Xb promoter, and; 3. The CpG islands between −311 and −300 and −226 and −222 are the regions within the SPAN-Xb promoter sequence that control gene expression.

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Ontogeny of neuropeptide Y expression in response to deprivation in lean Zucker rat pups

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1998.275.2.r466 ◽

1998 ◽

Vol 275 (2) ◽

pp. R466-R470 ◽

Cited By ~ 3

Author(s):

Timothy J. Kowalski ◽

Thomas A. Houpt ◽

Jeongwon Jahng ◽

Nori Okada ◽

Streamson C. Chua ◽

...

Keyword(s):

In Situ Hybridization ◽

Neuropeptide Y ◽

Arcuate Nucleus ◽

Maternal Deprivation ◽

The Other ◽

Zucker Rat ◽

Adult Rats ◽

Rat Pups

Hypothalamic neuropeptide Y (NPY) activity is believed to play an important role in the response to food deprivation in adult rats. Little is known, however, about the role of the hypothalamic NPY system in the control of food intake in the preweanling rat. To address this issue, we examined the effect of deprivation on arcuate nucleus preproNPY expression in lean Zucker rat pups, using in situ hybridization. PreproNPY expression within the arcuate nucleus was localized to cells in the medial portion. Twenty-four hours of food, water, and maternal deprivation significantly increased the relative abundance of preproNPY mRNA in pups on postnatal day (P) 2, P9, P12, and P15 by 14–31%. This response, however, was not observed on P5. The absence of an effect on P5 and the magnitude of the response at the other ages tested were not correlated with the amount of weight lost during deprivation.

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Specialized Subpopulations of Kisspeptin Neurons Communicate With GnRH Neurons in Female Mice

Endocrinology ◽

10.1210/en.2014-1671 ◽

2015 ◽

Vol 156 (1) ◽

pp. 32-38 ◽

Cited By ~ 22

Author(s):

Devesh Kumar ◽

Michael Candlish ◽

Vinod Periasamy ◽

Nergiz Avcu ◽

Christian Mayer ◽

...

Keyword(s):

Arcuate Nucleus ◽

Reproductive Physiology ◽

The Other ◽

Functional Specialization ◽

Neuron Population ◽

Female Mice ◽

Reproductive Maturation ◽

Neuron Populations ◽

Periventricular Nucleus ◽

Gnrh Neurons

Abstract The neuropeptide kisspeptin is a potent stimulator of GnRH neurons and has been implicated as a major regulator of the hypothalamus-pituitary-gonadal axis. There are mainly two anatomically segregated populations of neurons that express kisspeptin in the female hypothalamus: one in the anteroventral periventricular nucleus (AVPV) and the other in the arcuate nucleus (ARC). Distinct roles have been proposed for AVPV and ARC kisspeptin neurons during reproductive maturation and in mediating estrogen feedback on the hypothalamus-pituitary-gonadal axis in adults. Despite their pivotal role in the regulation of reproductive physiology, little is known about kisspeptin neuron connectivity. Although previous data suggest heterogeneity within the AVPV and ARC kisspeptin neuron populations, how many and which of these potential kisspeptin neuron subpopulations are actually communicating with GnRH neurons is not known. Here we used a combinatorial genetic transsynaptic tracing strategy to start to analyze the connectivity of individual kisspeptin neurons with the GnRH neuron population in female mice with a single-cell resolution. We find that only subsets of AVPV and ARC kisspeptin neurons are synaptically connected with GnRH neurons. We demonstrate that the majority of kisspeptin neurons within the AVPV and ARC does not communicate with GnRH neurons. Furthermore, we show that all kisspeptin neurons within the AVPV connected to GnRH neurons are estrogen sensitive and that most of these express tyrosine hydroxylase. Our data demonstrate functional specialization within the two kisspeptin neuron populations.

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Role of neuropeptide Y in the regulation of gonadotropin-releasing hormone gene expression in the rat preoptic area

Molecular Brain Research ◽

10.1016/0169-328x(94)90075-2 ◽

1994 ◽

Vol 26 (1-2) ◽

pp. 69-73 ◽

Cited By ~ 21

Author(s):

Songyun Li ◽

Min Hong ◽

Alain Fournier ◽

Serge St-Pierre ◽

Georges Pelletier

Keyword(s):

Gene Expression ◽

Neuropeptide Y ◽

Preoptic Area ◽

Gonadotropin Releasing Hormone ◽

Releasing Hormone

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Zoledronic acid promotes osteoclasts ferroptosis by inhibiting FBXO9-mediated p53 ubiquitination and degradation

PeerJ ◽

10.7717/peerj.12510 ◽

2021 ◽

Vol 9 ◽

pp. e12510

Author(s):

Xingzhou Qu ◽

Zhaoqi Sun ◽

Yang Wang ◽

Hui Shan Ong

Keyword(s):

Gene Expression ◽

Zoledronic Acid ◽

Cell Viability ◽

Cell Model ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Underlying Mechanism ◽

Cell Counting ◽

Venn Diagram

Bisphosphonates (BPs)-related osteonecrosis of jaw (BRONJ) is a severe complication of the long-term administration of BPs. The development of BRONJ is associated with the cell death of osteoclasts, but the underlying mechanism remains unclear. In the current study, the role of Zoledronic acid (ZA), a kind of bisphosphonates, in suppressing the growth of osteoclasts was investigated and its underlying mechanism was explored. The role of ZA in regulating osteoclasts function was evaluated in the RANKL-induced cell model. Cell viability was assessed by cell counting kit-8 (CCK-8) assay and fluorescein diacetate (FDA)-staining. We confirmed that ZA treatment suppressed cell viability of osteoclasts. Furthermore, ZA treatment led to osteoclasts death by facilitating osteoclasts ferroptosis, as evidenced by increased Fe2+, ROS, and malonyldialdehyde (MDA) level, and decreased glutathione peroxidase 4 (GPX4) and glutathione (GSH) level. Next, the gene expression profiles of alendronate- and risedronate-treated osteoclasts were obtained from Gene Expression Omnibus (GEO) dataset, and 18 differentially expressed genes were identified using venn diagram analysis. Among these 18 genes, the expression of F-box protein 9 (FBXO9) was inhibited by ZA treatment. Knockdown of FBXO9 resulted in osteoclasts ferroptosis. More important, FBXO9 overexpression repressed the effect of ZA on regulating osteoclasts ferroptosis. Mechanistically, FBXO9 interacted with p53 and decreased the protein stability of p53. Collectively, our study showed that ZA induced osteoclast cells ferroptosis by triggering FBXO9-mediated p53 ubiquitination and degradation.

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