INHBA Research Articles

Activin levels correlate with lymphocytic infiltration in epithelial ovarian cancer.

The TGF-β superfamily member activin, a dimer of the gene products of INHBA and/or INHBB, has been implicated in immune cell maturation and recruitment, but its immune impact within epithelial ovarian cancer (EOC) is not well characterized. We sought to explore differences in activin (INHBA/ Inhibin-βA and INHBB/ Inhibin-βB) between malignant and ovarian tissues at the RNA and protein level and assess the relationship between activin and immune cells in EOC. Publicly available RNA sequencing data were accessed from GEO (#GSE143897) with normalization and quantification performed via DESeq2. Immune gene expression profile was further explored within the TCGA-OV cohort derived from The Cancer Genome Atlas (TCGA). Immunohistochemical analysis was performed to evaluate activin A and T-cell markers CD8 and FoxP3 at the protein level. ELISA to activin-A was used to assess levels in the ascites of advanced EOC patients. Kaplan-Meier curves were generated to visualize survival outcomes. Gene expression levels of components of the activin signaling pathway were elevated within EOC when compared to a benign cohort, with differences in activin type I/II receptor gene profiles identified. Additionally, INHBA gene expression was linked to lymphocytic immune markers in EOC samples. Immunohistochemistry analysis revealed a positive correlation of CD8 and FOXP3 staining with activin A at the protein level in both primary and metastatic epithelial ovarian cancer samples. Furthermore, Activin-A (inhibin-βA) is significantly elevated in EOC patient ascites. INHBA expression is elevated within EOC, correlating with worse survival, with activin protein levels correlating with specific immune infiltration. Our findings suggest that activin-A may play a role in suppressing anti-tumor immunity in EOC, highlighting its potential as a therapeutic target.

Application of RNA-Seq Technology for Screening Reproduction-Related Differentially Expressed Genes in Tibetan and Yorkshire Pig Ovarian Tissue.

The purpose of this study was to explore and verify genes that regulate the reproductive traits of Tibetan pigs at the mRNA level. The ovarian tissues of Tibetan pigs (TPs) and Yorkshire pigs (YPs) were selected as research objects, and cDNA libraries of the ovarian tissue transcripts of Tibetan pigs and Yorkshire pigs were successfully constructed by the RNA-Seq technique. A total of 651 differentially expressed genes (DEGs) were screened, including 414 up-regulated genes and 237 down-regulated genes. Through GO and KEGG enrichment analysis, it was found that these differentially expressed genes were significantly enriched in cell process, reproductive process, reproduction, cell proliferation, binding, and catalytic activity, as well as oxidative phosphorylation, endocrine resistance, thyroid hormone, Notch, and other signal transduction pathways. Genes significantly enriched in pathways closely related to reproductive regulation were analyzed and selected, and the AR, CYP11A1, CYP17A1, INHBA, ARRB2, EGFR, ETS1, HSD17B1, IGF1R, MIF, SCARB1, and SMAD4 genes were identified as important candidate genes. Twelve differentially expressed genes related to reproduction were verified by RT-qPCR. The results showed that the expression of the AR, CYP17A1, EGFR, ETS1, IGF1R, and SMAD4 genes was significantly higher in Tibetan pigs than in Yorkshire pigs, while the expression of the CYP11A1, INHBA, ARRB2, HSD17B, MIF, and SCARB1 genes in Tibetan pigs was significantly lower than in Yorkshire pigs. The purpose of this study is to provide a theoretical basis for exploring the molecular mechanism of reproductive trait effect genes and the application of molecular breeding in Tibetan pigs.

Discovery of PELATON links to the INHBA gene in the TGF-β pathway in colorectal cancer using a combination of bioinformatics and experimental investigations

Colorectal cancer (CRC) is a major worldwide health issue, with high rates of both occurrence and mortality. Dysregulation of the transforming growth factor-beta (TGF-β) signaling pathway is recognized as a pivotal factor in CRC pathogenesis. Notably, the INHBA gene and long non-coding RNAs (lncRNAs) have emerged as key contributors to CRC progression. The aim of this research is to explore the immunological roles of INHBA and PELATON in CRC through a combination of computational predictions and experimental validations, with the goal of enhancing diagnostic and therapeutic strategies. In this study, we utilized bioinformatics analyses, which involved examining differential gene expression (DEG) in the TCGA-COAD dataset and exploring the INHBA gene in relation to the TGF-β pathway. Additionally, we analyzed mutations of INHBA, evaluated the microenvironment and tumor purity, investigated the INHBA's connection to immune checkpoint inhibitors, and measured its potential as an immunotherapy target using the TIDE score. Utilizing bioinformatics analyses of the TCGA-COAD dataset beside experimental methodologies such as RT-qPCR, our investigation revealed significant upregulation of INHBA in CRC. As results, our analysis of the protein-protein interaction network associated with INHBA showed 10 interacting proteins that play a role in CRC-associated processes. We observed a notable prevalence of mutations within INHBA and explored its correlation with the response to immune checkpoint inhibitors. Our study highlights INHBA as a promising target for immunotherapy in CRC. Moreover, our study identified PELATON as a closely correlated lncRNA with INHBA, with experimental validation confirming their concurrent upregulation in CRC tissues. Thus, these findings highlight the importance of INHBA and PELATON in driving CRC progression, suggesting their potential utility as diagnostic and prognostic biomarkers. By integrating computational predictions with experimental validations, this research enhances our understanding of CRC pathogenesis and uncovers prospects for personalized therapeutic interventions.

Neutrophil-derived Activin-A moderates their pro-NETotic activity and attenuates collateral tissue damage caused by Influenza A virus infection.

Pre-neutrophils, while developing in the bone marrow, transcribe the Inhba gene and synthesize Activin-A protein, which they store and release at the earliest stage of their activation in the periphery. However, the role of neutrophil-derived Activin-A is not completely understood. To address this issue, we developed a neutrophil-specific Activin-A-deficient animal model (S100a8-Cre/Inhba fl/fl mice) and analyzed the immune response to Influenza A virus (IAV) infection. More specifically, evaluation of body weight and lung mechanics, molecular and cellular analyses of bronchoalveolar lavage fluids, flow cytometry and cell sorting of lung cells, as well as histopathological analysis of lung tissues, were performed in PBS-treated and IAV-infected transgenic animals. We found that neutrophil-specific Activin-A deficiency led to exacerbated pulmonary inflammation and widespread hemorrhagic histopathology in the lungs of IAV-infected animals that was associated with an exuberant production of neutrophil extracellular traps (NETs). Moreover, deletion of the Activin-A receptor ALK4/ACVR1B in neutrophils exacerbated IAV-induced pathology as well, suggesting that neutrophils themselves are potential targets of Activin-A-mediated signaling. The pro-NETotic tendency of Activin-A-deficient neutrophils was further verified in the context of thioglycollate-induced peritonitis, a model characterized by robust peritoneal neutrophilia. Of importance, transcriptome analysis of Activin-A-deficient neutrophils revealed alterations consistent with a predisposition for NET release. Collectively, our data demonstrate that Activin-A, secreted by neutrophils upon their activation in the periphery, acts as a feedback mechanism to moderate their pro-NETotic tendency and limit the collateral tissue damage caused by neutrophil excess activation during the inflammatory response.

Cytokine profiling and transcriptomics in mononuclear cells define immune variants in Meniere Disease.

Meniere Disease (MD) is a chronic inner ear disorder characterized by vertigo attacks, sensorineural hearing loss, tinnitus, and aural fullness. Extensive evidence supporting the inflammatory etiology of MD has been found, therefore, by using transcriptome analysis, we aim to describe the inflammatory variants of MD. We performed Bulk RNAseq on 45 patients with definite MD and 15 healthy controls. MD patients were classified according to their basal levels of IL-1β into 2 groups: high and low. Differentially expression analysis was performed using the ExpHunter Suite, and cell type proportion was evaluated using the estimation algorithms xCell, ABIS, and CIBERSORTx. MD patients showed 15 differentially expressed genes (DEG) compared to controls. The top DEGs include IGHG1 (p = 1.64 10-6) and IGLV3-21 (p = 6.28 10-3), supporting a role in the adaptative immune response. Cytokine profiling defines a subgroup of patients with high levels of IL-1β with up-regulation of IL6 (p = 7.65 10-8) and INHBA (p = 3.39 10-7) genes. Transcriptomic data from peripheral blood mononuclear cells support a proinflammatory subgroup of MD patients with high levels of IL6 and an increase in naïve B-cells, and memory CD8+ T cells.

Progesterone-independent endometrial mRNA expression in dairy cows with clinical or subclinical endometritis

Up to 50 % of dairy cows fail to resolve uterine involution and develop chronic clinical (CE) or subclinical endometritis (SE) 21 days after calving. Clinical endometritis is associated with purulent discharge, while SE is not associated with overt clinical signs. Along with numerous knowledge gaps related to its pathogenesis, SE does not allow for a straightforward and effective therapy. Therefore, it is crucial to unravel differences in the expression of genes among healthy, CE, and SE cows. This might contribute to the discovery of new drug candidates and, in consequence, a potentially effective treatment. In the present study, cows between 21 and 28 days postpartum (PP) were examined using vaginoscopy for the presence of vaginal discharge and endometrial cytology for the determination of the endometrial polymorphonuclear cell (PMN) percentage. Next, an endometrial biopsy sample was taken to investigate the expression of 13 selected candidate genes by qPCR. Uterine health status was assigned to healthy (absence of abnormal vaginal discharge and ≤5 % PMN, n = 13), SE (absence of abnormal vaginal discharge and >5 % PMN, n = 30), and CE (mucopurulent or purulent vaginal discharge and >5 % PMN, n = 9). At the same time, a blood sample was collected to assess serum progesterone concentration and to categorize cows as low (≤1 ng/mL) or high (>1 ng/mL) in progesterone. High expression of IL1B, IL6, IL17A, CXCL8, PTGES, PTGS1, PTGS2, and INHBA genes and low expression of FST was noted in the endometrium of CE compared to healthy cows. Increased endometrial INHBA expression was observed in both SE and CE compared to healthy cows. Interestingly, greater expression of PTGES and PRXL2B genes and lower expression of PTGS2 were characteristic of SE versus CE or healthy. Among cows with no overt clinical symptoms of uterine disease (healthy and SE), the endometrial expression of IL1 B, CXCL8, and PTGES was greater in cows with high versus low serum progesterone. Several genes were differentially expressed among healthy, SE, and CE cows indicating different pathways for the development of different uterine diseases. In conclusion, we found progesterone-independent SE markers, which suggests that low endometrial PTGS2 expression may be indicative of an inadequate immune response and thus contribute to the pathogenesis of SE.

THU507 Tumor Activin A Mediates Cachexia Primarily Through White Adipose Tissue Hydrolysis In Pancreatic Ductal Adenocarcinoma

Abstract Disclosure: S. Yu: None. Y. Luan: None. R. Dong: None. A. Abazarikia: None. S. Kim: None. Pancreatic ductal adenocarcinoma (PDAC) commonly harbors mutations in KRAS and TP53 genes and its overall 5-year survival rate is only 10% in the US. Among risk factors associated with the dismal PDAC prognosis, cachexia accounts for about 30% of PDAC-related death and is present in nearly 80% of PDAC patients. Significant adipose tissue loss followed by skeletal muscle loss characterizes PDAC-related cachexia in humans and mice. Accumulated evidence indicates that activin A, a homodimer of inhibin βA that is encoded by the INHBA gene, is a recognized key cachexia mediator as supported by mitigation of weight loss by activin A antagonists in PDAC-bearing mice. However, much remains unclear regarding the primary source cells of activin A and the mechanisms by which activin A mediates cachexia in PDAC. To address the unknowns, we employed CRISPR/Cas9 system for Inhba gene knock-down (KD) in mouse KPC (KrasG12D and Trp53R172H) PDAC cells and generated orthotopic mice using Inhba KD or control KPC cells. Sham group which received the same operation except for cell injection was included. Body weight was measured on a daily basis, and blood and white adipose tissues (WATs) and skeletal muscle tissues were harvested on post-surgery day 12. We found that, when compared to the Sham group, injection of control KPC cells significantly elevated serum activin A levels up to 6 ng/mL and temporally decreased the average body weight in the orthotopic mice (Ctrl group), whereas Inhba gene KD significantly reduced serum activin A levels by ∼ 2 ng/mL and prevented weight loss in the orthotopic mice (KD group). When compared to the Sham and KD groups, notable reductions in organ weights of epididymal, mesenteric or subcutaneous WATs were found in the Ctrl group, however, weights for gastrocnemius and plantaris or soleus muscle tissues were statistically similar between the Ctrl and KD groups at post-surgery day 12. Correspondingly, the average serum free fatty acids levels were higher in the Ctrl group than the KD group. Histological observations support that dramatic adipocyte size reductions in WATs were prevented by serum activin A suppression. Immunofluorescence analysis showed that WATs from the Ctrl group had considerably higher protein expression of hormone-sensitive or adipose triglyceride lipase than the KD and Sham groups. Furthermore, phosphorylation of SMAD3 and phosphorylated PKA substrate which regulate adipocyte lipolysis were intensely stained in WATs from the Ctrl group when compared to the KD and Sham groups. Collectively, our findings identify PDAC cancer cells as the major source of activin A in PDAC and suggest systemic activin A as a potential mediator of PDAC-related adipose tissue loss. The current findings warrant further investigation to address clinical relevance. Presentation: Thursday, June 15, 2023

Uncovering Tumor-Promoting Roles of Activin A in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers with high incidence rates of metastasis and cachexia. High circulating activin A, a homodimer of inhibin βA subunits that are encoded by INHBA gene, predicts poor survival among PDAC patients. However, it still raises the question of whether activin A suppression renders favorable PDAC outcomes. Here, the authors demonstrate that activin A is abundantly detected in tumor and stromal cells on PDAC tissue microarray and mouse PDAC sections. In orthotopic male mice, activin A suppression, which is acquired by tumor-targeted Inhba siRNA using cholesterol-modified polymeric nanoparticles, retards tumor growth/metastasis and cachexia and improves survival when compared to scramble siRNA-treated group. Histologically, activin A suppression coincides with decreased expression of proliferation marker Ki67 but increased accumulation of α-SMAhigh fibroblasts and cytotoxic T cells in the tumors. In vitro data demonstrate that activin A promotes KPC cell proliferation and induces the downregulation of α-SMA and upregulation of IL-6 in pancreatic stellate cells (PSC) in the SMAD3-dependent mechanism. Moreover, conditioned media from activin A-stimulated PSC promoted KPC cell growth. Collectively, our data provide a mechanistic basis for tumor-promoting roles of activin A and support therapeutic potentials of tumor activin A suppression for PDAC.

TGF-Beta Induces Activin A Production in Dermal Fibroblasts Derived from Patients with Fibrodysplasia Ossificans Progressiva.

Fibrodysplasia ossificans progressiva (FOP) is a catastrophic, ultra-rare disease of heterotopic ossification caused by genetic defects in the ACVR1 gene. The mutant ACVR1 receptor, when triggered by an inflammatory process, leads to heterotopic ossification of the muscles and ligaments. Activin A has been discovered as the main osteogenic ligand of the FOP ACVR1 receptor. However, the source of Activin A itself and the trigger of its production in FOP individuals have remained elusive. We used primary dermal fibroblasts from five FOP patients to investigate Activin A production and how this is influenced by inflammatory cytokines in FOP. FOP fibroblasts showed elevated Activin A production compared to healthy controls, both in standard culture and osteogenic transdifferentiation conditions. We discovered TGFβ1 to be an FOP-specific stimulant of Activin A, shown by the upregulation of the INHBA gene and protein expression. Activin A and TGFβ1 were both induced by BMP4 in FOP and control fibroblasts. Treatment with TNFα and IL6 produced negligible levels of Activin A and TGFβ1 in both cell groups. We present for the first time TGFβ1 as a triggering factor of Activin A production in FOP. As TGFβ1 can promote the induction of the main driver of FOP, TGFβ1 could also be considered a possible therapeutic target in FOP treatment.

Recapitulation of pro-inflammatory signature of monocytes with ACVR1A mutation using FOP patient-derived iPSCs

BackgroundFibrodysplasia ossificans progressiva (FOP) is a rare genetic disease characterized by progressive heterotopic ossification (HO) in soft tissues due to a heterozygous mutation of the ACVR1A gene (FOP-ACVR1A), which erroneously transduces the BMP signal by Activin-A. Although inflammation is known to trigger HO in FOP, the role of FOP-ACVR1A on inflammatory cells remains to be elucidated.ResultsWe generated immortalized monocytic cell lines from FOP-iPSCs (FOP-ML) and mutation rescued iPSCs (resFOP-ML). Cell morphology was evaluated during the monocyte induction and after immortalization. Fluorescence-activated cell sorting (FACS) was performed to evaluate the cell surface markers CD14 and CD16 on MLs. MLs were stimulated with lipopolysaccharide or Activin-A and the gene expression was evaluated by quantitative PCR and microarray analysis. Histological analysis was performed for HO tissue obtained from wild type mice and FOP-ACVR1A mice which conditionally express human mutant ACVR1A gene by doxycycline administration. Without any stimulation, FOP-ML showed the pro-inflammatory signature of CD16+ monocytes with an upregulation of INHBA gene, and treatment of resFOP-ML with Activin-A induced an expression profile mimicking that of FOP-ML at baseline. Treatment of FOP-ML with Activin-A further induced the inflammatory profile with an up-regulation of inflammation-associated genes, of which some, but not all, of which were suppressed by corticosteroid. Experiments using an inhibitor for TGFβ or BMP signal demonstrated that Activin-A-induced genes such as CD16 and CCL7, were regulated by both signals, indicating Activin-A transduced dual signals in FOP-ML. A comparison with resFOP-ML identified several down-regulated genes in FOP-ML including LYVE-1, which is known to suppress matrix-formation in vivo. The down-regulation of LYVE-1 in HO tissues was confirmed in FOP model mice, verifying the significance of the in vitro experiments.ConclusionThese results indicate that FOP-ML faithfully recapitulated the phenotype of primary monocytes of FOP and the combination with resFOP-ML is a useful tool to investigate molecular events at the initial inflammation stage of HO in FOP.

Regulation of DNA methylation during the testicular development of Shaziling pigs

DNA methylation is one of the key epigenetic regulatory mechanisms in development and spermatogenesis. However, the dynamic regulatory mechanisms of genome-wide DNA methylation during testicular development remain largely unknown. Herein, we generated a single-base resolution DNA methylome and transcriptome atlas of precocious porcine testicular tissues across three developmental stages (1, 75, and 150 days old). The results showed that the dynamic methylation patterns were directly related to the expression of the DNMT3A gene. Conjoint analysis revealed a negative regulatory pattern between promoter methylation and the positive regulation of 3′-untranslated region (3′UTR) methylation. Mechanistically, the decrease in promoter methylation affected the upregulation of meiosis-related genes, such as HORMAD1, SPO11, and SYCE1. Demethylation in the 3′UTR induced the downregulation of the INHBA gene and knockdown of INHBA inhibited cell proliferation by reducing the synthesis of activin A. These findings contribute to exploring the regulatory mechanisms of DNA methylation in testicular development.

Clinical features of high INHBA gene expression in colon cancer

Clinical features of high INHBA gene expression in colon cancer

RVU120 (SEL120) CDK8/19 Inhibitor - a Drug Candidate for the Treatment of MDS Can Induce Erythroid Differentiation

Background: Myelodysplastic syndromes (MDS) are a group of malignant blood disorders characterized by ineffective hematopoiesis and cytopenias and frequent evolution to acute myeloid leukemia (AML). MDS results from the expansion of genetically and epigenetically changed clones with impaired differentiation and maturation. Primary clinical goals in MDS are to achieve remissions, alleviate symptoms associated with cytopenias, and minimize the transfusion burden. While supportive red blood cell transfusions, erythropoiesis-stimulating agents and novel targeted agents may lead to clinical improvement, an allogeneic bone marrow transplant (BMT) remains the only potential curative option for patients with MDS. RVU120 (SEL120) is a specific, selective inhibitor of CDK8 and its paralog CDK19. A first-in-human Phase Ib clinical trial with RVU120 in patients with AML or high risk (HR)-MDS is currently ongoing. Preclinical studies indicated the strong antileukemic potential of RVU120 that was often associated with the multilineage commitment of CD34+ AML cells. Moreover, RVU120 could improve proliferation and induce erythroid differentiation of CD34+ cells derived from Diamond-Blackfan anemia (DBA) patients.Aims: Primary aim was to evaluate the erythroid differentiation potential of RVU120 in primary MDS and transformed cord blood CD34+ blood cells characterized with an early block in erythroid differentiation.Methods: Efficacy and mechanism of action of RVU120 and other CDK8 inhibitors were investigated in cord blood (CB) cells transduced with TLS-ERG, a fusion gene generated from t(16;21)(p11;q22). Transformed cells displayed an increased capacity for self-renewal, proliferation, and altered erythroid differentiation. Efficacy was further tested in CD34+ cells isolated from BM of MDS patients and established MDS cell lines. Cells were treated with RVU120 and global transcriptional changes and chromatin status were analyzed by RNA-seq, ATAC-seq, and ChIP-seq. Cell cycle, proliferation, and lineage-specific markers were studied by flow cytometry in liquid and semi-solid methylcellulose-based media.Results: RVU120 treatment leads to transcriptional reprogramming of transformed CD34+ CB cells. The most profound changes included decreased CDK8 occupancy followed by increased STAT5 and RNA Polymerase II loading at transcription start site and gene bodies. RVU120 treatment transcriptionally represses multiple genes associated with hematopoietic and leukemia stem cells such as CD34, FLI1, ENG and RGS18 and importantly induce the expression of genes involved in erythroid commitment, including regulators of erythroid/megakaryocytic fate, such as RGS18, KLF1, FLI1, INHBA, GATA1/2 and hemoglobin genes. Detailed analysis of transformed CB and MDS CD34+ cells by flow cytometry at early and late time points reflected sequential changes in the expression of lineage-specific surface markers, leading to erythroid differentiation.Conclusions: Presented results indicate strong erythroid differentiation potential of RVU120 in CD34+ cells that acquired genetic abnormalities leading to arrested erythroid commitment, characteristics of many MDS and AML subtypes. Observed differentiation phenotype strikingly resembles effects of RVU120 in DBA cells caused by disruption of genes encoding ribosomal proteins. Detailed transcriptomic profiling strongly links the differentiation with enrichment of genes representing regulators of erythroid commitment and hemoglobin metabolism. Further studies are warranted to investigate the efficacy of RVU120 in chronic anemias associated with bone marrow failures in AML and MDS patients. [Display omitted] DisclosuresPakulska: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Obacz: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Goller: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Combik: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Keska: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Mazan: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Juszczynski: Ryvu Therapeutics: Current equity holder in publicly-traded company. Brzozka: Ardigen: Current Employment, Membership on an entity's Board of Directors or advisory committees; Selvita SA: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees. Dziedzic: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Angelosanto: Ryvu Therapeutics: Current Employment. Shamsili: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company. Rzymski: Ryvu Therapeutics: Current Employment, Current equity holder in publicly-traded company.

P–710 The possible relation between kisspeptin and apoptosis on granulosa cells of PCOS patients

Abstract Study question Does kisspeptin administration affect the apoptotic factors of granulosa cells of Polycystic Ovary Syndrome (PCOS) patients? Summary answer Kisspeptin administration significantly increased apoptosis related factors. What is known already Kisspeptin plays a critical role in central gonadotropin secretion by acting on GnRH neurons in the hypothalamus. Kisspeptin receptors are also expressed on the ovaries. Kisspeptin–54 has recently been known as a prominent agent in oocyte maturation. Although there are many studies on the possible roles of kisspeptin administration in vivo on hypothalamic gonadotropin secretion and ovarian function and the possible roles of these effects in the pathogenesis of PCOS, there is not enough information about the effect of in vitro application of kisspeptin on ovarian function and conditions that lead to the development of PCOS. Study design, size, duration This basic research study was an in vitro experimental approach involving the use of granulosa cell from a PCOS and normal cases between July to December in 2020. 32 women were included in both control and PCOS groups. Participants/materials, setting, methods Women with (n = 16) and without (n = 16) PCOS were included in the study. Granulosa cells, follicular fluids and blood samples were collected on oocyte pick-up day. Cells were isolated from follicular fluids. Kisspeptin–54 levels were determined by ELISA in serum and follicle fluids. Also, FSH and LH levels were determined in serum. Kisspeptin–54 administrated in vitro to the half of the samples. Fluorescence spectrophotometer and laser scanning confocal microscope were used to evaluate calcium concentrations Main results and the role of chance The serum kisspeptin level was found significantly higher in PCOS patients compared to the control group (p < 0.05). Also, the level of kisspeptin in follicular fluid was statistically significantly higher in PCOS patients compared to the control group (p < 0.05). The mean serum FSH level of PCOS patients was found to be significantly lower than the control group (p < 0.05). Furthermore, the mean serum LH level of PCOS patients was significantly higher than control groups (p < 0.05). The intracellular calcium concentration in granulosa cells obtained from PCOS patients was significantly lower than the control group (p < 0.05). Limitations, reasons for caution The KISS1, KISS1R, BCL–2, CASPASE–3, FSHR, LHCGR, STAR, ESR1, ESR2, INHBA gene expression levels of apoptotic process is still under analyses. The protein expression analyses of these genes may strength the results. Wider implications of the findings: Using kisspeptin antagonists may be supposed as an additional treatment for patients with PCOS. Trial registration number OMU KAEK 2019 /724

Adrenergic-mediated increases in INHBA drive CAF phenotype and collagens.

Adrenergic-mediated increases in INHBA drive CAF phenotype and collagens.

Activin A Determines Steroid Levels and Composition in the Fetal Testis.

Activin A promotes fetal mouse testis development, including driving Sertoli cell proliferation and cord morphogenesis, but its mechanisms of action are undefined. We performed ribonucleic acid sequencing (RNA-seq) on testicular somatic cells from fetal activin A-deficient mice (Inhba KO) and wildtype littermates at embryonic day (E) E13.5 and E15.5. Analysis of whole gonads provided validation, and cultures with a pathway inhibitor discerned acute from chronic effects of altered activin A bioactivity. Activin A deficiency predominantly affects the Sertoli cell transcriptome. New candidate targets include Minar1, Sel1l3, Vnn1, Sfrp4, Masp1, Nell1, Tthy1 and Prss12. Importantly, the testosterone (T) biosynthetic enzymes present in fetal Sertoli cells, Hsd17b1 and Hsd17b3, were identified as activin-responsive. Activin-deficient testes contained elevated androstenedione (A4), displayed an Inhba gene dose-dependent A4/T ratio, and contained 11-keto androgens. The remarkable accumulation of lipid droplets in both Sertoli and germ cells at E15.5 indicated impaired lipid metabolism in the absence of activin A. This demonstrated for the first time that activin A acts on Sertoli cells to determine local steroid production during fetal testis development. These outcomes reveal how compounds that perturb fetal steroidogenesis can function through cell-specific mechanisms and can indicate how altered activin levels in utero may impact testis development.

Activin A overexpression promotes rat follicular development through SCF-kit-mediated cell signals

We explored the possible signaling pathway by which activin A induces oocyte maturation. Inhba-overexpressing lentivirus vectors were constructed and transfected into primary granulosa cells in vitro and ovary tissues in vivo in rats. The granulosa cell growth curve was drawn, and antibodies for phospho-Smad2, phospho-Erk5, phospho-Nur77, and stem cell factor (SCF) were prepared for western blot analysis. Protein expression of SCF and C-kit in the rat ovaries was detected by immunohistochemical staining. The rate of granulosa cell proliferation was higher in the Inhba gene overexpression group (INH) than in the control groups (CON group and GFP group) in vitro. Protein expression of SCF and C-kit was higher in the INH group than in the other two groups. phospho-Smad2, phospho-ERK5, P-nur77, and SCF proteins showed positive expression in rat ovarian granulosa cells in each group and were obviously increased in the INH group. Activin A overexpression may promote rat granulosa cell proliferation through Smad2/ERK5/nur77 signaling pathways, and rat granulosa cells overexpressing activin A in vitro showed increased levels of SCF and c-kit proteins.

Adrenergic-mediated increases in INHBA drive CAF phenotype and collagens.

Adrenergic-mediated increases in INHBA drive CAF phenotype and collagens.

Quantification of Inha, Inhba, and Inhbb genes expression levels in Wister rat testis during Pre- and Post-Pubertal Stages

The present study has been designed to quantitate the mRNA expression level of testicular Inha, Inhba, and Inhbb genes at pre- and post-pubertal stages of male Wister rats. Fifty male Wister rats (25 immature and 25 mature) were used in the present study. At pre-pubertal stage, 5 male rats of 25, 30, 35, 40, and 45 days old, and at post-pubertal stage, 5 male rats of 55, 60, 65, 70, and 75 days old were anasthetized and testis were obtained for molecular study to investigate the expression levels of Inha, Inhba, and Inhbb genes using qRT- PCR. The expression level of Inha gene in testis decreased as the age progress until 40 day period, and then slightly increased at 45 day period. At 55 day period, the expression significantly increased. At 60 and 65 periods, the levels recorded no increase, but 70 and 75 day periods recorded significant increase. The expression level of Inhba gene increased significantly as the age progress at the pre-pubertal stage, where the highest level was recorded at 45 day. At 55, 60, and 65 day periods, the highest expression level has been recorded, thenafter, the levels decreased at 70 and 75 day periods. The expression level of Inhbb gene increased significantly at 30, 35, 40, and 45 day periods of the pre-pubertal stage. At 55 and 60 day periods, no significant difference was recorded compared with 45 day period. At 65 day period, the highest level was recorded, thenafter, the levels decreased at 70 and 75 day periods. It can be concluded that fold changes of Inha, Inhba, and Inhbb genes in testis, correlated with the alterations of inhibins and activins concentrations at different periods of pre- and post-pubertal stages of male Wister rats life.

MP39-06 WHOLE GENOME TRANSCRIPTIONAL ANALYSIS OF CLEAR CELL RENAL CELL CARCINOMA WITH VENOUS TUMOR THROMBUS REVEALS INTRATUMORAL HETEROGENEITY AND GENES ASSOCIATED WITH POOR OUTCOME

You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology I1 Apr 2017MP39-06 WHOLE GENOME TRANSCRIPTIONAL ANALYSIS OF CLEAR CELL RENAL CELL CARCINOMA WITH VENOUS TUMOR THROMBUS REVEALS INTRATUMORAL HETEROGENEITY AND GENES ASSOCIATED WITH POOR OUTCOME Dharam Kaushik, Wasim Chowdhury, Ping Wu, Teresa Johnson-Pais, Yidong Chen, Michael A. Liss, and Ronald Rodriguez Dharam KaushikDharam Kaushik More articles by this author , Wasim ChowdhuryWasim Chowdhury More articles by this author , Ping WuPing Wu More articles by this author , Teresa Johnson-PaisTeresa Johnson-Pais More articles by this author , Yidong ChenYidong Chen More articles by this author , Michael A. LissMichael A. Liss More articles by this author , and Ronald RodriguezRonald Rodriguez More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.1180AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Overall, 4 to 10% of newly diagnosed renal cell carcinoma (RCC) patients have been found to have a venous tumor thrombus (VTT). Intratumoral heterogeneity may contribute to progression of the disease and metastases. Previous studies have focused on studying the heterogeneity of metastases, recurrence and tumor. There has been no study to date evaluating VTT and characterizing its transcriptional profile. We report results of the transcriptional analysis of primary tumor (PT), VTT and adjacent normal parenchyma (NP). METHODS We performed Whole Transcriptome Sequencing on fresh tissue specimens from 6 patients with clear cell RCC and VTT collected at the time of radical nephrectomy with tumor thrombectomy. We evaluated transcriptional alterations between PT/NP, VTT/NP and VTT/PT. We compared our data set with The Cancer Genome Atlas (TCGA) data. For multiple testing corrections, we utilized false discovery estimation and differential expression criterion. RESULTS 1. We surveyed 23,228 genes and identified differential expression in 1455, 1344 and 26 genes between tumor and normal parenchyma (PT/NP), tumor thrombus and normal parenchyma (VTT/NP) and tumor thrombus and primary tumor (VTT/PT) respectively. We identified altered key pathways (cytokine activity, regulation of apoptosis, cytoskeleton organization, immune response). 2. We compared these genes with TCGA data and identified 35 genes, which predicted poor outcome. 3. VTT demonstrated statistically significant differential expression of OSM (2.37 fold), INHBA (1.9 fold), CCL2 (2.3 fold), CCL20 (2.9 fold) and IL1B genes (4.5 fold) compared to primary tumor (TT/T), (Figure1). 4. We identified statistically significant higher expression of OSM, INHBA, CCL20, and IL1B genes in higher tumor thrombus level compared to lower tumor thrombus level. CONCLUSIONS RNA sequencing of RCC with VTT reveals significant genomic intratumoral heterogeneity. We identified key molecular pathways and differential expression of genes in VTT compared to primary tumor (VTT/PT). Furthermore, these genes were found to be upregulated in higher VTT level compared to lower VTT level. These results will require validation in a larger cohort. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e495 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Dharam Kaushik More articles by this author Wasim Chowdhury More articles by this author Ping Wu More articles by this author Teresa Johnson-Pais More articles by this author Yidong Chen More articles by this author Michael A. Liss More articles by this author Ronald Rodriguez More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...