Proliferation Of Follicular Cells Research Articles

Elevated RBP4 in Subclinical Ketosis Cows Inhibits Follicular Granulosa Cell Proliferation and Steroid Hormone Synthesis

The mechanism by which subclinical ketosis (SCK) causes postpartum reproductive disorders in dairy cows remains unclear. In this study, cows within the day 14 to 21 postpartum period were categorized into the SCK group or the control group. Subsequently, they were monitored until 45 d to 60 d postpartum and divided into the SCK anestrus group (SCK-AE, n = 12) and the control estrus group (C-E, n = 12). In comparison to the C-E group, the RBP4 and p-AKT of the SCK-AE group exhibited increased levels in serum, liver, and ovaries. In the in vitro experimental cultivation of granulosa cells (GCs), after adding RBP4, cell proliferation, steroid hormone secretion and synthesis, and GLUT4 secretion were inhibited, and cell apoptosis was exacerbated. After silencing STRA6 (RBP4 receptor), cell proliferation and steroid hormone secretion and synthesis, as well as the inhibition of GLUT4, were alleviated, and the situation of cell apoptosis also improved. The SC79 activator could promote the phosphorylation of AKT, thus alleviating the increased cell proliferation, steroid hormone secretion and synthesis, GLUT4 inhibition, and apoptosis rate in cow GCs induced by RBP4 stimulation. Our research indicates that elevated RBP4 levels in SCK cows inhibit the proliferation, apoptosis, and steroid hormone synthesis of GCs through the STRA6 receptor and the PI3K/AKT pathway.

Identification of functional circRNAs regulating ovarian follicle development in goats

BarkgroundCircular RNAs (circRNAs) play important regulatory roles in a variety of biological processes in mammals. Multiple birth-traits in goats are affected by several factors, but the expression and function of circRNAs in follicular development of goats are not clear. In this study, we aimed to investigate the possible regulatory mechanisms of circRNA and collected five groups of large follicles (Follicle diameter > 6 mm) and small follicles (1 mm < Follicle diameter < 3 mm) from Leizhou goats in estrus for RNA sequencing.ResultsRNA sequencing showed that 152 circRNAs were differentially expressed in small and large follicles. Among them, 101 circRNAs were up-regulated in large follicles and 51 circRNAs were up-regulated in small follicles. GO and KEGG enrichment analyses showed that parental genes of the differential circRNAs were significantly enriched in important pathways, such as ovarian steroidogenesis, GnRH signaling pathway, animal autophagy and oxytocin signalling pathway. BioSignal analysis revealed that 152 differentially expressed circRNAs could target 91 differential miRNAs including miR-101 family (chi-miR-101-3p, chi-miR-101-5p), miR-202 family (chi-miR-202-5p, chi-miR-202-3p),60 circRNAs with translation potential. Based on the predicted sequencing results, the ceRNA networks chicirc_008762/chi-miR-338-3p/ARHGAP18 and chicirc_040444/chi-miR-338-3p/STAR were constructed in this study. Importantly, the new gene circCFAP20DC was first discovered in goats. The EDU assay and flow cytometry results indicated that circCFAP20DC enhanced the proliferation of follicular granulosa cells(GCs). Real-time quantitative PCR and western blotting assays showed that circCFAP20DC activated the Retinoblastoma(RB) pathway and promoted the progression of granulosa cells from G1 to S phase.ConclusionDifferential circRNAs in goat size follicles may have important biological functions for follicular development. The novel gene circCFAP20DC activates the RB pathway, promoting the progression of GCs from G1 to S phase. This, in turn, enhances the proliferation of follicular GCs in goats.

Mechanisms and effects of AdipoRon, an adiponectin receptor agonist, on ovarian granulosa cells-a systematic review.

Granulosa cells, crucial components of ovarian follicles, play a fundamental role in follicle development, hormone production, and overall reproductive health. These cells are integral to steroidogenesis, including the synthesis and secretion of key hormones such as estrogen and progesterone. Dysregulation of granulosa cells can lead to reproductive disorders, including polycystic ovary syndrome and infertility. This systematic review provides a comprehensive evaluation of AdipoRon, a synthetic agonist of adiponectin receptors AdipoR1 and AdipoR2, and its effects on ovarian function, with a particular focus on granulosa cells. Due to the absence of clinical trials, the review centers on preclinical studies to explore AdipoRon's potential therapeutic benefits and to suggest future research directions. A detailed literature search across databases such as PubMed, Scopus, Web of Science, Embase, and Google Scholar was conducted using terms related to AdipoRon and ovarian function. The review encompasses four preclinical studies involving various models: primary granulosa cells from rats, laying hens' granulosa cells, human luteinized granulosa cells, and chicken ovary follicles. Findings indicate that AdipoRon enhances glucose absorption in rat granulosa cells by stimulating glucose transporter 1 expression, modulates steroid hormone secretion in laying hens' granulosa cells, and affects cell proliferation and steroidogenesis in human luteinized granulosa cells. Additionally, AdipoRon, in conjunction with recombinant chicken adiponectin, influences ovarian follicular cell proliferation and steroidogenesis in chicken ovary follicles. This review highlights the need for further investigation into AdipoRon's long-term effects and its potential applications in reproductive health and therapy.

Effects and action mechanism of gonadotropins on ovarian follicular cells: A novel role of Sphingosine-1-Phosphate (S1P). A review

Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) control antral follicular growth by regulating several processes, such as the synthesis of hormones and signaling molecules, proliferation, survival, apoptosis, luteinization, and ovulation. To exert these effects, gonadotropins bind to their respective Gs protein-coupled receptors, activating the protein kinase A (PKA) pathway or recruiting Gq proteins to activate protein kinase C (PKC) signaling. Although the action mechanism of FSH and LH is clear, recently, it has been shown that both gonadotropins promote the synthesis of sphingosine-1-phosphate (S1P) in granulosa and theca cells through the activation of sphingosine kinase 1. Moreover, the inhibition of SPHKs reduces S1P synthesis, cell viability, and the proliferation of follicular cells in response to gonadotropins, and the addition of S1P to the culture medium increases the proliferation of granulosa and theca cells without apparent effects on sexual steroid synthesis. Therefore, we consider that S1P is a crucial signaling molecule that complements the canonical gonadotropin pathway to promote the proliferation and viability of granulosa and theca cells.

Flavonoid Supplementation Is Beneficial for Polycystic Ovary Syndrome: A Systematic Review and Meta-analysis.

Polycystic ovary syndrome (PCOS) is a prevalent hormonal imbalance that predominantly affects women in their reproductive years. Previous studies have yielded conflicting conclusions. This is an updated meta-analysis aiming to explore the connection between flavonoid supplementation and PCOS. Seven databases were searched: Cochrane Library, PubMed, Web of Science, Embase, Wanfang, China Science and Technology Journal, and China National Knowledge Infrastructure, spanning from their inception to April 15, 2024. Two authors independently searched the databases using the search terms. Following strict inclusion criteria, 8 papers were ultimately included. This updated meta-analysis suggests that flavonoid supplementation could enhance follicular development, promote the proliferation and differentiation of follicular granulosa cells, elevate estradiol levels, and mitigate testosterone, C-reactive protein, and ovarian index levels. This analysis suggests that dietary flavonoids could potentially alleviate symptoms associated with PCOS. PROSPERO registration no. CRD42022382912.

Effect of progestin on thyroid function in female Wistar rats.

To characterize the influence of female-specific hormones on women's thyroid function, the study investigated the influence of extra progestin from oral contraceptives on inducing thyroid dysfunction. Sixty female Wistar rats were divided into six groups based on levonorgestrel or desogestrel administration as the main active agents: control, low (0.0039 mg*20-fold), medium (0.0039 mg*100-fold), high (0.0318 mg*100-fold) levonorgestrel (pure product); and low (0.0083 mg*20-fold) and high (0.0083 mg*100-fold) desogestrel (pure product). Progestin was administered by gavage every 4 days for 1 month. Statistical analysis was performed using one-way analysis of variance and the Kruskal-Wallis test. Following levonorgestrel gavage, serum free T4 and thyroidstimulating hormone levels were significantly lower in the experimental group than that in the control group (p=0.013 and 0.043). After desogestrel gavage, the serum free T4 and free T3 levels were lower in the experimental group than that in the control group (p=0.019 and 0.030). Thyroid hormone antibody concentrations were lower in rats administered levonorgestrel and desogestrel than that in control rats. Moreover, exposure to progestin upregulated the expression of the thyroid-stimulating hormone receptor and sodium iodide symporter in thyroid. Progestin stimulation enhanced the proliferation of follicular epithelial cells in rat thyroid tissues. Progestin exposure could cause thyroid dysfunction by upregulating the transcription of thyroid-stimulating hormone receptor and sodium iodide symporter in thyroid, thus inducing pathomorphological changes in rats' thyroid.

TAF1 is needed for the proliferation and maturation of thyroid follicle cells via Notch signaling.

Thyroid dysgenesis (TD) is the common pathogenic mechanism of congenital hypothyroidism (CH). In addition, known pathogenic genes are limited to those that are directly involved in thyroid development. To identify additional candidate pathogenetic genes, we performed forward genetic screening for TD in zebrafish, followed by positional cloning. The candidate gene was confirmed in vitro using the Nthy-ori 3.1 cell line and in vivo using a zebrafish model organism. We obtained a novel zebrafish line with thyroid dysgenesis and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1) by positional cloning. Further molecular studies revealed that taf1 was needed for the proliferation of thyroid follicular cells by binding to the NOTCH1 promoter region. Knockdown of TAF1 impaired the proliferation and maturation of thyroid cells, thereby leading to thyroid dysplasia. This study showed that TAF1 promoted Notch signaling and that this association played a pivotal role in thyroid development.NEW & NOTEWORTHY In our study, we obtained a novel zebrafish line with thyroid dysgenesis (TD) and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1). Further researches revealed that taf1 was required for thyroid follicular cells by binding to the NOTCH1 promoter region. Our findings revealed a novel role of TAF1 in thyroid morphogenesis.

Long non-coding RNA Loc105611671 promotes the proliferation of ovarian granulosa cells and steroid hormone production upregulation of CDC42.

Granulosa cells (GCs) are essential for follicular development, and long non-coding RNAs (LncRNAs) are known to support the maintenance of this process and hormone synthesis in mammals. Nevertheless, the regulatory roles of these lncRNAs within sheep follicular GCs remain largely unexplored. This study delved into the influence of a Loc105611671, on the proliferation and steroid hormone synthesis of sheep ovarian GCs and the associated target genes in vitro. Cell Counting Kit-8 (CCK-8) gain-of-function experiments indicated that overexpression of Loc105611671 significantly boosted GCs proliferation, along with estrogen (E2) and progesterone (P4) levels. Further mechanistic scrutiny revealed that Loc105611671 is primarily localized within the cytoplasm of ovarian granulosa cells and engages in molecular interplay with CDC42. This interaction results in the upregulation of CDC42 protein expression. Moreover, it was discerned that increased CDC42 levels contribute to augmented proliferation of follicular granulosa cells and the secretion of E2 and P4. Experiments involving co-transfection elucidated that the concurrent overexpression of CDC42 and Loc105611671 acted synergistically to potentiate these effects. These findings provide insights into the molecular underpinnings of fecundity in ovine species and may inform future strategies for enhancing reproductive outcomes.

MiR-128-3p Regulates Follicular Granulosa Cell Proliferation and Apoptosis by Targeting the Growth Hormone Secretagogue Receptor.

The proliferation and apoptosis of granulosa cells (GCs) affect follicle development and reproductive disorders, with microRNAs playing a crucial regulatory role. Previous studies have shown the differential expression of miR-128-3p at different stages of goat follicle development, which suggests its potential regulatory role in follicle development. In this study, through the Cell Counting Kit-8 assay, the EDU assay, flow cytometry, quantitative real-time polymerase chain reaction, Western blot, and the dual-luciferase reporter assay, we used immortal human ovarian granulosa tumor cell line (KGN) cells as materials to investigate the effects of miR-128-3p and its predicted target gene growth hormone secretagogue receptor (GHSR) on GC proliferation and apoptosis. The results show that overexpression of miR-128-3p inhibited the proliferation of KGN cells, promoted cell apoptosis, and suppressed the expression of proliferating cell nuclear antigen (PCNA) and B-cell lymphoma-2 (BCL2) while promoting that of Bcl-2 associated X protein (BAX). The dual-luciferase reporter assay revealed that miR-128-3p bound to the 3' untranslated region sequence of GHSR, which resulted in the inhibited expression of GHSR protein. Investigation of the effects of GHSR on GC proliferation and apoptosis revealed that GHSR overexpression promoted the expression of PCNA and BCL2, enhanced GC proliferation, and inhibited cell apoptosis, whereas the opposite effects were observed when GHSR expression was inhibited. In addition, miR-128-3p and GHSR can influence the expression of extracellular signal-regulated kinase 1/2 protein. In conclusion, miR-128-3p inhibits KGN cell proliferation and promotes cell apoptosis by downregulating the expression of the GHSR gene.

YTHDF2 as a Mediator in BDNF-Induced Proliferation of Porcine Follicular Granulosa Cells.

In female mammals, the proliferation and apoptosis of granulosa cells (GCs) are critical in determining the fate of follicles and are influenced by various factors, including brain-derived neurotrophic factor (BDNF). Previous research has shown that BDNF primarily regulates GC proliferation through the PI3K/AKT, NF-kB, and CREB tumour pathways; however, the role of other molecular mechanisms in mediating BDNF-induced GC proliferation remains unclear. In this study, we investigated the involvement of the m6A reader YTH domain-containing family member 2 (YTHDF2) in BDNF-stimulated GC proliferation and its underlying mechanism. GCs were cultured in DMEM medium supplemented with varying BDNF concentrations (0, 10, 30, 75, and 150 ng/mL) for 24 h. The viability, number, and cell cycle of GCs were assessed using the CCK-8 assay, cell counting, and flow cytometry, respectively. Further exploration into YTHDF2's role in BDNF-stimulated GC proliferation was conducted using RT-qPCR, Western blotting, and sequencing. Our findings indicate that YTHDF2 mediates the effect of BDNF on GC proliferation. Additionally, this study suggests for the first time that BDNF promotes YTHDF2 expression by increasing the phosphorylation level of the ERK1/2 signalling pathway. This study offers a new perspective and foundation for further elucidating the mechanism by which BDNF regulates GC proliferation.

Exploring the Metabolic and Endocrine Preconditioning Associated with Thyroid Disorders: Risk Assessment and Association with Acne Severity.

Metabolic preconditioning, characterized by conditions like obesity and insulin resistance syndrome, disrupts hormonal balance. Elevated androgen levels stimulate excessive sebum production and follicular cell proliferation, leading to acne lesions. Similarly, thyroid hormone imbalances affect sebaceous gland activity, epidermal lipid composition, and skin cell turnover, impacting acne occurrence and severity. This study aimed to assess the potential contribution of metabolic and endocrine preconditions to acne development. A total of 389 patients diagnosed with acne were included and divided into three groups: the metabolic precondition group (MPG, N = 163, 41.9%), the endocrine precondition group (EPG, N = 162, 41.65%), and the control group (CG, N = 89, 22.88%). Data related to the degree of acne severity and comorbidities of interest were collected from the patients' medical records. In the groups with concomitant diseases, moderate and severe acne were significantly more prevalent (56.44% and 41.10% in MPG, and 35.80% and 61.11% in EPG) compared to the control group (5.61% and 4.89%). The most prevalent preconditions observed were insulin resistance syndrome in MPG (63.8%) and autoimmune thyroiditis in EPG (95.06%). Significant age-related differences in acne severity were found across all study groups (p < 0.05). In MPG, the age variable was significantly higher in the presence of mild acne, while in EPG, the age variable was significantly lower for the mild acne group. A positive association was observed between the severity of acne and insulin resistance syndrome, obesity, autoimmune thyroiditis, and hypothyroidism (p < 0.05). Risk analysis indicated a significantly higher risk (RR > 1, 95% CI RR > 1, p < 0.001) of developing moderate and severe acne in the presence of these preconditions. The presence of both metabolic and endocrine preconditions significantly increased the likelihood of developing severe acne, leading to the hypothesis that both conditions may be contributing factors to the development of acne.

Withanolide derivatives: natural compounds with anticancer potential offer low toxicity to fertility and ovarian follicles in mice.

Anticancer therapy often leads to premature ovarian insufficiency (POI) and infertility due to the extreme sensitivity of the ovarian follicle reserve to the effects of chemotherapy. Withanolides are known for their cytotoxic effect on cancer cells and low cytotoxicity on non-malignant or healthy cells. Therefore, this study aimed to investigate the in vivo effects of three withanolides derivatives: 27-dehydroxy-24,25-epoxywithaferin A (WT1), 27-dehydroxywithaferin A (WT2), and withaferin A (WTA) on fertility, and the ovarian preantral follicles of young female mice. To achieve this, mice received 7 intraperitoneal doses of WT1, WT2, or WTA at a concentration of 2 mg/kg (Experiment I) and 5 or 10 mg/kg (Experiment II) over 15 alternate days. In experiment I, two days after administration of the last dose, half of the mice were mated to evaluate the effects of withanolides on fertility. The other half of the mice, as well as all mice from experiment II, were sacrificed for histological, inflammation, senescence, and immunohistochemical analyses of the follicles present in the ovary. Regardless of the administered withanolide, the concentration of 2 mg/kg did not show toxicity on the follicular morphology, ovarian function, or fertility of the mice. However, at concentrations of 5 and 10 mg/kg, the three derivatives (WT1, WT2, and WTA) increased follicular activation, cell proliferation, and ovarian senescence without affecting inflammatory cells. Furthermore, at a concentration of 10 mg/kg, the three withanolides showed intensified toxic effects, leading to DNA damage as evidenced by the labeling of γH2AX, activated Caspase 3, and TUNEL. We conclude that the cytotoxic effect of the tested withanolide derivatives (WT1, WT2, and WTA) in the concentration of 2 mg/kg did not show toxicity on the ovary. However, in higher concentrations, such as 10 mg/kg, toxic effects are potentiated, causing DNA damage.

Endocrine tumors of the female reproductive tract

Endocrine cells responsible for hormone secretion are found in virtually every organ system. The diverse neoplasms arising from endocrine cells in the female reproductive tract are not well recognized as a distinct component of endocrine oncology. Here, we integrate cellular origins with native anatomical residence to help classify neoplasms of this system. The neoplasms include steroidogenic tumors that arise usually in ovarian stroma, neuroendocrine neoplasms that can arise from normal neuroendocrine cells throughout the female reproductive tract or in ovarian germ cell tumors, and thyroid follicular cell proliferations that are exclusively a component of an ovarian teratoma and may be malignant. The neuroendocrine neoplasms run the full spectrum from indolent neuroendocrine tumors to aggressive poorly differentiated neuroendocrine carcinomas. While many of these lesions are identified as incidental findings in surgically resected tissues, others present with inappropriate hormone excess. An important consideration is the distinction of primary disease from metastatic malignancy. Genetic disorders including those caused by germline mutations of the FOXL2, GNAS, DICER1, STK11 and MEN1 genes can present with primary endocrine neoplasms of the female reproductive tract.

Progressively transformed germinal center-like follicular T-cell lymphoma:a clinicopathological analysis of 14 cases

Objective: To investigate the clinicopathologic features of progressively transformed germinal center-like follicular T-cell lymphoma (PTGC-like FTCL). Methods: The clinicopathologic data of 14 PTGC-like FTCL cases that were diagnosed at the Beijing Friendship Hospital Affiliated to the Capital Medical University from January 2017 to January 2022 were retrospectively collected. Clinicopathological features, immunophenotype, and Epstein-Barr virus (EBV) infection status were analyzed in these cases. Polymerase chain reaction (PCR) was performed to detect the clonal gene rearrangements of T cell receptor (TCR) and the immunoglobulin (Ig) in 10 and 8 cases, respectively. Results: The male to female ratio was 5∶2. The median age was 61 years (range 32-70 years). All patients had lymphadenopathy at the time of diagnosis. By using the Ann Arbor system staging, seven cases were classified as stage Ⅰ-Ⅱ, and seven cases as stage Ⅲ-Ⅳ. Seven cases had B symptoms, four cases had splenomegaly, and two cases had skin rash and pruritus. Previously, three cases were diagnosed as classic Hodgkin's lymphoma, three cases as small B-cell lymphoma, two cases as atypical lymphoid hyperplasia unable to exclude angioimmunoblastic T-cell lymphoma (AITL), one case as EBV-associated lymphoproliferative disorder, and one case as peripheral T-cell lymphoma (PTCL) associated with the proliferation of B cells. All the 14 cases showed that the large nodules were composed of mature CD20+, IgD+B lymphocytes admixed with small aggregates of neoplastic cells with pale to clear cytoplasm. Moreover, hyperplastic germinal centers (GCs) and Hodgkin/Reed-Sternberg-like (HRS-like) cells were seen within these nodules in two and five cases, respectively. The neoplastic cells expressed CD3 (14/14), CD4 (14/14), PD1 (14/14), ICOS (14/14), CD10 (9/14), bcl-6 (12/14), CXCL13 (10/14), and CD30 (10/14). The HRS-like cells in five cases expressed CD20 (2/5), PAX5 (5/5), CD30 (5/5), CD15 (2/5), LCA (0/5), OCT2 (5/5) and BOB1 (2/5). Moreover, neoplastic T cells formed rosettes around HRS-like cells. EBV-encoded RNA (EBER) in situ hybridization showed scattered, small, positive bystander B lymphocytes in 8/14 cases, including 3/5 cases containing HRS-like cells. All tested cases (including five with HRS-like cells) showed monoclonal TCR gene rearrangement and polyclonal Ig gene rearrangement. Conclusions: PTGC-like FTCL is a rare tumor originated from T-follicular helper cells. It could be distinguished from angioimmunoblastic T-cell lymphoma by the formation of follicular structure, and lack of follicular dendritic cell proliferation outside the follicles and the polymorphous inflammatory background. In addition, it should be differentiated from lymphocyte-rich classical Hodgkin's lymphoma and low-grade B cell lymphoma.

Proliferation and migration of ML1 follicular thyroid cancer cells are inhibited by IU1 targeting USP14: role of proteasome and autophagy flux.

USP14 is a deubiquitinating enzyme involved in protein degradation by interacting with the proteasome and removal of poly-ubiquitin chains on target proteins. USP14 can influence cellular processes such as cell survival, DNA repair, ER stress, endocytosis, and the inflammatory response. USP14 further plays a role in tumor growth, and the inhibition of USP14 by compounds such as IU1 may affect cancer cell migration and invasion. Here we have studied the mechanisms for the action of IU1 in ML1 follicular thyroid cancer cells, comparing them with control, primary thyroid cells. Treatment with IU1 reduced proliferation of ML1 cells in a concentration-dependent manner, and more prominently than in control cells. IU1 decreased basal migration of ML1 cells, and after stimulation of cells with the bioactive compound, sphingosine-1-phosphate. The sphingosine-1-phosphate receptor 3 was increased in ML1 cells as compared with control thyroid cells, but this was not influenced by IU1. Further studies on the mechanism, revealed that IU1 enhanced the proteasome activity as well as LC3B-dependent autophagy flux in ML1 cells with an opposite effect on control thyroid cells. This indicates that IU1 elicits a cell-type dependent autophagy response, increasing it in ML1 cancer cells. The IU1-mediated stimulation of autophagy and proteasomes can likely contribute to the reduced cell proliferation and migration observed in ML1 cells. The precise set of proteins affected by IU1 in ML1 thyroid and other cancer cells warrant further investigations.

Immunolocalization of activin and inhibin at different stages of follicular development in the lizard Sceloporus torquatus

The activins and inhibins are glycoproteins with a role in the follicular development of vertebrates, that are found in follicular fluid and somatic follicular cells, with a different pattern among taxa. The principal function of activin (Act) is to modulate the follicle-stimulating hormone (FSH) synthesis and secretion, whereas inhibin (Inh) downregulates it. Both factors are modulators of intraovarian follicular recruitment, oocyte maturation, cell proliferation, and steroidogenic activity. Our aim was to characterize the immunolocalization of Act and Inh in the ovarian follicles during the reproductive cycle of the lizard Sceloporus torquatus. Act was detected in the granulosa cells and oocyte cortex in the different stages of follicular development. On the other hand, we identified Inh in the oocyte cortex and the cytoplasm of pyriform and small cells of previtellogenic follicles. Also, we found immunoreactivity in the oocyte cortex, theca, and small cells of vitellogenic and preovulatory follicles. Our data provide evidence that Act and Inh have changes related to the stage of follicular development. This dynamic appears to be conserved among vertebrates and is fundamental to ensure an adequate follicular development in this specie.

FGF12 regulates cell cycle gene expression and promotes follicular granulosa cell proliferation through ERK phosphorylation in geese

The granulosa cells play an important role in the fate of follicular development or atresia in poultry. Fibroblast growth factor 12 (FGF12) is downregulated in atretic follicles and may be involved in regulating granulosa cell survival in previous studies, but its molecular mechanism remains unclear. In this study, FGF12 overexpression and knockdown models of goose granulosa cells were constructed to investigate its function. The downstream expression of the cell cycle pathway was analyzed by qPCR. Granulosa cell proliferative activity and apoptosis were detected by CCK8 and TUNEL. Protein phosphorylation levels of ERK and AKT were measured using Western blotting to analyze the key pathway of FGF12 regulation of granulosa cell proliferation. ERK protein phosphorylation inhibitor was added for further verification. After overexpression of FGF12, cell proliferation activity was increased, the expressions of cell cycle pathway genes CCND1, CCNA2, MAD2, and CHK1 were upregulated, the apoptosis of granulosa cell was decreased, and Caspase 3 gene and protein expression were downregulated. After the knockdown of FGF12, cell proliferation activity decreased, the expression of downstream genes in the cell cycle pathway was downregulated, the apoptosis of granulosa cells was increased, and the Bcl-2 gene and protein were downregulated. Overexpression of FGF12 promoted the synthesis of P4 and upregulates the expression of the STAR gene. Overexpression of FGF12 promoted ERK protein phosphorylation but did not affect AKT phosphorylation. The addition of ERK phosphorylation inhibitors resulted in the elimination of the increase in cell proliferative activity caused by FGF12 overexpression. In conclusion, FGF12 could promote proliferation and inhibit apoptosis of goose granulosa cells by increasing ERK phosphorylation.

P18INK4C and BRCA1 inhibit follicular cell proliferation and dedifferentiation in thyroid cancer

ABSTRACT Only 3% of thyroid cancers are medullary thyroid carcinomas (MTCs), the rest are follicular epithelial cell derived non-MTCs (NMTCs). A dysfunctional INK4-CDK4-RB pathway is detected in most of NMTCs. DNA repair defects and genome instability are associated with NMTC dedifferentiation and aggressiveness. Whether inactivation of the INK4-CDK4-RB pathway induces NMTCs and how differentiation of NMTC cells is controlled remain elusive. In this study, we generated p18Ink4c and Brca1 singly and doubly deficient mice as well as p16Ink4a and Brca1 singly and doubly deficient mice. By using these mice and human thyroid carcinoma cell lines, we discovered that loss of p18Ink4c, not p16Ink4a, in mice stimulated follicular cell proliferation and induced NMTCs. Depletion of Brca1 alone or both p16Ink4a and Brca1 did not induce thyroid tumor. Depletion of Brca1 in p18Ink4c null mice results in poorly differentiated and aggressive NMTCs with epithelial-mesenchymal transition (EMT) features and enhanced DNA damage. Knockdown of BRCA1 in thyroid carcinoma cells activated EMT and promoted tumorigenesis whereas overexpression of BRCA1 inhibited EMT. BRCA1 and EMT marker expression were inversely related in human thyroid cancers. Our finding, for the first time, demonstrates that inactivation of INK4-CDK4-RB pathway induces NMTCs and that Brca1 deficiency promotes dedifferentiation of NMTC cells. These results suggest that BRCA1 and p18INK4C collaboratively suppress thyroid tumorigenesis and progression and CDK4 inhibitors will be effective for treatment of INK4-inactivated or cyclin D-overexpressed thyroid carcinomas.

Abstract #1400018: Genetic Alterations in NRAS, TERT, PIK3CA and SPEN Genes Leading to an Aggressive Metastatic RAI Refractory Follicular Thyroid Cancer

Follicular thyroid cancer (FTC) is a well-differentiated thyroid tumor and the second most common type after papillary thyroid cancer (PTC) with higher incidence of distant metastases, more aggressive behavior, and poorer outcome than PTC Patient is 43-year-old female with 6.2 cm TR4 left thyroid nodule on Neck US. On Exam, Cricoid is deviated to the right with left thyroid firm nodule. FNA showed Follicular neoplasm. TSH level was 1.59 (0.45-4.70 mIU/L) and Free T4 level: 1.19 (0.78-2.20 ng/dl) CT scan of neck with contrast and MRI thoracic spine: Large, solid heterogenous mass in left thyroid lobe causing lateral displacement and mild narrowing of the trachea with retrosternal extension, Osteolytic lesion of T4 vertebral body and right sided pleura-based mass at the level of T7-T8 with No enlarged cervical lymph nodes. IR biopsy of vertebra: thyroid Origin Metastatic Carcinoma, Patient underwent total thyroidectomy with central neck dissection. Follicular thyroid cancer (STAGE II pT3A,pN1B,pM1) Mutational Testing: Negative BRAF and KRAS with Positive NRAS, PIK3CA, SPEN and TERT. Patient started on TSH suppression therapy. 1 month Later: Thyroglobulin: 2443.4 ng/ml (1.3-31.8), Thyroglobulin Ab< 0.9 (0.0-4.0 IU/ml), TSH 0.02 with Free T4 2.53 FDG PET: Multifocal osseous metastatic disease, predominantly in the axial skeleton Thyrogen stimulated WBS I-123 showed No evidence of residual or metastatic disease. Patient recieved 214 mCi of iodine-131. Post treatment MN scan showed accumulation of iodine in the metastatic bone lesions with small focal activity in the thyroidectomy bed. Post RAI treatment: TG trended to 1102.7 then 1307 Patient Completed 10 fractions of external beam radiation on T4 spine Ultrasound thyroid: No residual thyroid tissue or suspicious lymph nodes Oncology started Bone directed therapy with zoledronate with consideration of PIK3CA mutation targeted therapy with Everolimus or Temsirolimus FTC is more aggressive with poorer prognosis than PTC that is more likely to hematogenously disseminate, with mortality rate of 50%. The neoplastic proliferation of thyroid follicular cells develops by altering multiple molecular pathways such as the PI3K/AKT pathway involving the RAS, RAF, RET, and NTRK1 genes. RAS is a dual activator of the mitogen activated protein kinase and PI3K/AKT pathways TERT is a catalytic subunit of the telomerase complex. Reported in 11–17% of FTC. TERT promoter mutations associated with tumor aggressiveness and mortality PIK3CA encodes p110-alpha, is a catalytic subunit of phosphatidylinositol 3-kinase (PI3K) involves in thyroid cancer tumorigenesis and progression, reported in 7.7% of FTC with sensitivity to therapies targeting PI3K49-56, AKT57-58, or mTOR59-66. In our patient, pretreatment MN scan failed to show RAI avid lesions but PET showed increase FDG uptake and since receiving RAI treatment helped in lowering TG that indicate a possibility of mixture of RAI avid and non-RAI avid lesions existing at same time so RAI treatment is still recommended despite negative Diagnostic Scan for persistent elevated TG.

The effects of apelin on IGF1/FSH-induced steroidogenesis, proliferation, Bax expression, and total antioxidant capacity in granulosa cells of buffalo ovarian follicles.

Apelin (APLN) was believed to be an adipokine secreted from adipose tissue. However, studies demonstrate that it is a pleiotropic peptide and has several effects on the female reproductive system. In this study, We examined the effects of different doses of IGF1 and FSH in the presence of APLN-13 on the production of progesterone in buffalo ovary granulosa cells. Furthermore, different doses of APLN isoforms (APLN-13 and APLN-17) were tested on proliferation, Bax protein expression, and antioxidant capacity in the same cells. Granulosa cells of buffalo ovaries were cultured in the presence of different doses of IGF1 and FSH with or without APLN-13 (10-9M) to evaluate its effect on the secretion of progesterone tested by ELISA assay. The WST-1 method was used to survey the effect of APLN on granulosa cell proliferation and cytotoxicity. In addition, the antioxidant capacity of the cells in the presence of APLN was assessed using the FRAP method. mRNA and Bax protein levels were measured in granulosa cells treated with APLN using real-time PCR and western blot techniques. APLN-13 (10-9) stimulated the effect of IGF1 on the production of progesterone, and its levels were affected by APLN-13 dose-dependently. However, it did not significantly stimulate the effect of FSH on the secretion of progesterone. APLN-13 (all doses) and APLN-17 (10-8 and 10-9M) improved the proliferation of granulosa cells. Moreover, preincubation of the cells for an hour by APLN receptor antagonist (ML221, 10µM) did not significantly affect the proliferation of cells induced by APLN. Neither APLN-13 nor APLN-17 were not cytotoxic for the cells compared to the control treatment. APLN-13 at the doses of 10-6 and 10-8M substantially up and down-regulated Bax protein expression; however, such effects were not observed when the cells were preincubated with ML221. In addition, APLN-17 did not influence the expression amount of Bax. Furthermore, both APLN-13 and -17 improved the total antioxidant capacity of the ovarian granulosa cells, but such effects were not seen when the cells were preincubated with ML221. According to these results, APLN enhanced the steroidogenesis induced by IGF1 but did not affect the steroidogenesis induced by FSH. APLN also enhanced the cell proliferation and antioxidant capacity of buffalo ovaries follicular granulosa cells; however, its effect on Bax expression was different.