Inhibin Subunits Research Articles

The role of steroidogenic factor 1 (SF-1) in steroidogenic cell function of the testes and ovaries of mature mice.

The nuclear receptor steroidogenic factor 1 (SF-1) is essential for mature mouse gonad steroidogenic gene expression, for Leydig and Sertoli cell function, and depletion of SF-1 in steroidogenic cells of the testis compromises steroidogenesis, spermatogenesis and male fertility. Steroidogenic factor 1 (SF-1 or NR5A1) plays an essential role in the development of fetal gonads and regulates genes involved in steroid biosynthesis. Since SF-1 is expressed in multiple cell types in mouse gonads, we developed three novel conditional knockout (cKO) mouse models employing Cre-recombinase and floxed alleles of SF-1 (Nr5a1f/f) to identify its role in testes and ovaries of mature mice: Cytochrome P450 17α-hydroxylase (Cyp17Cre/+;Nr5a1f/f, Leydig and theca cell-specific), aromatase (Cyp19Cre/+;Nr5a1f/f, Sertoli and granulosa cell-specific), as well as a combination of both (Cyp17+Cyp19-Cre;Nr5a1f/f). Compared to control animals, Cyp19-Cre;Nr5a1f/f cKO males showed normal fertility and testicular function. The Cyp17Cre/+;Nr5a1f/f cKO males had smaller testis, with drastically reduced Leydig cell volumes and impaired steroidogenesis, though their reproductive performance remained comparable to controls. Some 50% of Cyp17Cre/++Cyp19Cre/+;Nr5a1f/f double-cKO (dKO) males were infertile, while the remaining 50% showed significantly reduced fertility. These dKO males also had smaller testis with degenerative seminiferous tubules, abnormal Leydig cell morphology and lower levels of intra-testicular testosterone. Abnormal Sertoli cell localization was noted in dKO testes, with increased Sox9, p27 and inhibin subunit ßb and decreased androgen receptor expression. Female mice from all genotypes showed normal reproductive capacity, though steroidogenic gene expression levels were significantly decreased in both Cyp17Cre/+;Nr5a1f/f cKO and dKO females. These results show the essential role of SF-1 in mature mouse gonad steroidogenic gene expression, for Leydig and Sertoli cell function, and that depletion SF-1 in all steroidogenic cells of the testis compromises steroidogenesis, spermatogenesis and male fertility.

Exploration the role of INHBA in Hu sheep granulosa cells using RNA-Seq

Activin/inhibin is an important factor for the fecundity of Hu sheep, and it is involved in follicular development in ovaries. Inhibin subunit beta A (INHBA) participates in the synthesis of activin A and inhibin A. In this study, we also noted a positive correlation between INHBA level and the secretion of both activin A and inhibin A in culture medium. Nevertheless, both knockdown and overexpression of INHBA downregulated the expression of Inhibin Subunit Alpha (INHA). Based on RNA-Sequencing, we further examined the effect and molecular mechanism of INHBA knockdown in GCs on mRNA expression. A total of 1,687 differentially expressed genes (DEGs) were identified (Fold change ≥ 2; False-discovory-rates (FDR) ≤ 0.01), of which 602 genes were upregulated and 1,087 genes were downregulated in the INHBA interference group compared with the control groups. Gene Ontology (GO) enrichment indicated that these DEGs were mainly involved in the regulation of cell cycle, protein serine/threonine kinase activity, and actin cytoskeleton reorganization. Moreover, DEGs were significantly enriched in 40 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, including P53, progesterone-mediated oocyte maturation, and PI3K-AKT signaling pathways. We also noted a positive correlation between INHBA level and many PI3K/Akt/mTOR pathway-related genes at the gene or/and protein expression. Overall, this study may contribute to a better understanding of the roles of INHBA on GCs of prolific sheep, as well as the molecular effect of low INHBA expression on GCs, clarifying some reproductive failures.

RF34 | PMON19 Biallelic Variants in the INHA Gene Are the Cause of Gonadal Dysgenesis, Primary Amenorrhea and Absence of Puberty in the Setting of Hypergonadotropic Hypogonadism

Abstract Background Hypergonadotropic hypogonadism can be caused by congenital or acquired causes. These can range from chromosomal abnormalities, enzymatic defects or mutations that cause gonadotropin resistance, trauma, chemotherapy, infections etc. It is uncommon in females other than in Turner syndrome and not all the genetic etiologies are known. Clinical Case A 15 10/12 year old female with primary amenorrhea and absent pubertal development presented with FSH and LH levels of 35.24 mIU/mL and 14.38 mIU/mL, respectively and non-pubertal estradiol levels of 4 pg/mL consistent with hypergonadotropic hypogonadism. Her evaluation showed a normal 46,XX female karyotype and no evidence of hyperandrogenism or thyroid dysfunction. Ultrasound showed no evidence of ovaries and a rudimentary uterus measuring 0.9 cm×1.4 cm×1.2 cm with a delayed bone age of 12 years. Family history was positive for mother having delayed puberty with menarche at 16.5 years of age. Patient was started on hormone replacement therapy with transdermal estrogen. Four months after treatment she developed breast buds and vaginal discharge. After 1.5 years, the uterus measurements were 5.7 cm x3.7 cm x2.2 cm although no ovarian tissue or follicles were identified on MRI. Her biochemical profile also showed decreases in FSH (3.48 mIU/mL) and LH (0.99 mIU/mL) and an increase in estradiol (38pg/mL). Exome sequencing was performed and revealed biallelic missense variants c.766C>T, p.(Pro256Ser) and c.1036G>C p.(Gly346Arg) in exon 2 of the INHA (NM_002191) gene. Both variants are rare in the human population, seen in a heterozygous state with a global allele frequency (gnomADv2.1) of 0 and 0.00002790 (rs780833794), respectively, and predicted to be damaging by bioinformatic tools. INHA, a member of the transforming growth factor-β (TGF-β) superfamily, encodes the alpha subunit of glycoprotein hormones inhibin A and B. Studies in animal models indicate that inhibin A negatively regulates follicle stimulating hormone (FSH) secretion by suppressing FSHR expression and plays a crucial role in ovarian steroid hormone production, follicular development, proliferation and apoptosis of granulosa cells, and oocyte maturation. Heterozygous pathogenic variants in the INHA gene have been reported in patients with primary ovarian insufficiency (POI) and secondary amenorrhea1, while our patient with biallelic variants in INHA is affected by gonadal dysgenesis, primary amenorrhea, absence of puberty, a more severe POI phenotype. Conclusion To our knowledge, this is the first case of biallelic mutations in INHA gene leading to a severe phenotype (gonadal dysgenesis, primary amenorrhea and absence of puberty) compared to heterozygous mutations that lead to POI with secondary amenorrhea. 1

Identification of a Biomarker Combination for Survival Stratification in pStage II/III Gastric Cancer after Curative Resection.

Simple SummaryGastric cancer (GC) is the fifth most common cancer worldwide and the fourth most common cause of cancer-related deaths, with a high frequency of recurrence and metastasis, and a poor prognosis. This study presents a novel combination of four proteins (PDGFRB, INHBA, MMP11, and galectin-10) in GC tissues that have been identified as useful survival stratification markers in patients with pStage II/III GC after curative resection by quantitative polymerase chain reaction (qPCR), proteomic analysis, and immunohistochemistry (IHC).Background: We sought to identify an optimal combination of survival risk stratification markers in patients with pathological (p) stage II/III gastric cancer (GC) after curative resection. Methods: We measured the expression levels of 127 genes in pStage II/III GC tissues of two patient cohorts by quantitative polymerase chain reaction (qPCR) and the expression of 1756 proteins between two prognosis (good and poor) groups by proteomic analysis to identify candidate survival stratification markers. Further, immunohistochemistry (IHC) using tumor microarrays (TMAs) in another cohort of patients was performed to identify an optimal biomarker combination for survival stratification in GC patients. Results: secreted protein acidic and rich in cysteine (SPARC), erb-b2 receptor tyrosine kinase 2 (ERBB2), inhibin subunit beta A (INHBA), matrix metallopeptidase-11 (MMP11), tumor protein p53 (TP53), and platelet-derived growth factor receptor-beta (PDGFRB) were identified as candidate biomarkers from qPCR analysis, and SPARC and galectin-10 were obtained from the proteomic analysis. The combination of PDGFRB, INHBA, MMP11, and galectin-10 was identified as the optimal combination of survival risk stratification markers. Conclusions: A combination of four proteins in GC tissues may serve as useful survival risk stratification markers in patients with pStage II/III GC following curative resection. Our results may facilitate future multicenter prospective clinical trials.

Prognostic impact of activin subunit inhibin beta A in gastric and esophageal adenocarcinomas

PurposeAdenocarcinomas of the esophagus (AEG) and stomach (AS) are among the most common cancers worldwide. Novel markers for risk stratification and guiding treatment are strongly needed. Activin is a multi-functional cytokine with context specific pro- and anti-tumorigenic effects. We aimed to investigate the prognostic role of activin tumor protein expression in AEG/ASs.MethodsTissue from a retrospective cohort of 277 patients with AEG/AS treated primarily by surgery at the Charité - Universitätsmedizin Berlin was collected and analyzed by immunohistochemistry using a specific antibody to the activin homodimer inhibin beta A. Additionally, we evaluated T-cell infiltration and PD1 expression as well as expression of PD-L1 by immunohistochemistry as possible confounding factors. Clinico-pathologic data were collected and correlated with activin protein expression.ResultsOut of 277 tumor samples, 72 (26.0%) exhibited high activin subunit inhibin beta A protein expression. Higher expression was correlated with lower Union for International Cancer Control (UICC) stage and longer overall survival. Interestingly, activin subunit expression correlated with CD4+ T-cell infiltration, and the correlation with higher overall survival was exclusively seen in tumors with high CD4+ T-cell infiltration, pointing towards a role of activin in the tumor immune response in AEG/ASs.ConclusionIn our cohort of AEG/AS, higher activin subunit levels were correlated with longer overall survival, an effect exclusively seen in tumors with high CD4+ cell infiltration. Further mechanistic research is warranted discerning the exact effect of this context specific cytokine.

INHBB is a novel prognostic biomarker and correlated with immune infiltrates in gastric cancer.

Inhibin subunit beta B (INHBB) is a potential prognostic biomarker for a variety of cancers. However, its role in gastric cancer (GC) remains elusive. The differential expression data of INHBB in tumor and normal tissues were extracted from several databases and genetic alterations of INHBB were assessed by cBioPortal. Kaplan-Meier analysis was used to evaluate the survival rate of patients with GC with INHBB and association with clinical features in GC. Cox regression analysis was used to explore the prognostic value of clinical indicators and INHBB in GC, and a nomogram prognostic model was established. In addition, the predictive validity of the nomogram model was assessed by time-depended receiver operating characteristic (ROC) and calibration curves. Functional enrichment analyses were conducted to functionally annotate INHBB. Notably, we found that the quantitative assessment of immune cell subpopulation infiltration correlated with INHBB expression. INHBB expression is upregulated in GC and is correlated with several clinical features including prognostic indicators and a histological type. Genetic alterations were observed in INHBB, its DNA methylation level was negatively correlated with INHBB expression. High INHBB expression is associated with a poor prognosis and is an independent risk factor for prognosis in GC, along with age and residual tumor. The nomogram model showed a good prediction ability and was validated by time-depended ROC and calibration curves. Functional enrichment analysis indicated that INHBB-associated genes were enriched in tumor microenvironment Gene Ontology (GO) terms and were correlated with tumor-associated pathways. INHBB has a regulatory function in immune cell infiltration, especially macrophage infiltration in GC. Specifically, patients with GC with high INHBB expression and high macrophage infiltration have a worse prognosis. INHBB expression was negatively correlated with the expression of chemokines/chemokine receptors and plays a regulatory role in immunoinhibitor/immunostimulator-involved pathways. INHBB is a potential prognostic biomarker for GC and may drive the abnormal activity of critical cancer-associated pathways, potentially contributing to immune cell infiltration to promote GC development.

Inflammation in Preeclampsia: Genetic Biomarkers, Mechanisms, and Therapeutic Strategies.

ObjectivePreeclampsia is a common and serious complication of pregnancy, posing a threat to maternal and fetal safety due to the lack of effective biomarkers and treatment strategies. This study aimed to identify potential biomarkers that can be used to predict preeclampsia and identify the molecular mechanisms of preeclampsia pathogenesis and drug prediction at the transcriptome level.MethodsWe analyzed differential expression genes (DEGs) in preeclampsia and non-preeclampsia groups in the GSE75010 dataset, cross-linking with extracted inflammatory response-related genes to obtain differentially expressed inflammation-related genes (DINRGs). Enrichment analysis and protein-protein interaction (PPI) networks were constructed to understand the functions and enrichment pathways. Machine learning models were used to identify key genes associated with preeclampsia and build a nomogram in the training set, which was validated in the validation set. The R package RcisTarget was used to predict transcription factors, and Cytoscape was used to construct miRNA-mRNA pathways, which could identify the molecular mechanisms. Then, we conducted molecular docking of the obtained key genes INHBA (inhibin subunit beta A), OPRK1 (opioid receptor kappa 1), and TPBG (trophoblast glycoprotein), as well as predicted transcription factors with drug molecules. Additionally, the CIBERSORT method explored the differences in immune cell infiltration between preeclampsia and non-preeclampsia samples based on the GSE75010 dataset.ResultsA total of 69 DINRGs associated with preeclampsia patients were screened. INHBA, OPRK1, and TPBG were the key genes based on machine learning models. A nomogram for prediction was further constructed, and the receiver operating curves (ROCs) showed good performance. Based on the transcriptome level of key genes, we proposed that RELA-miR-548K/miR-1206-TPBG may be a potential RNA regulatory pathway regulating the progression of early preeclampsia. Molecular docking suggested the effectiveness of curcumin in the treatment of preeclampsia. Additionally, regulatory T cells (Tregs) and resting mast cells were significantly different between the two groups.ConclusionIn summary, we identified three key inflammation-associated genes, namely INHBA, OPRK1, and TPBG, which can be used as potential genetic biomarkers for preeclampsia prediction and treatment, and established a nomogram as a predictive model. Additionally, we provided insights into the mechanisms of preeclampsia development at the transcriptome level and performed corresponding drug predictions.

Comprehensive analysis of INHBA: A biomarker for anti-TGFβ treatment in head and neck cancer.

Inhibin subunit βA (INHBA) is a protein-coding gene belonging to the transforming growth factor β (TGFβ) superfamily, which is associated with the development of a variety of cancers. However, the role of INHBA in head and neck squamous cell carcinoma (HNSC) remains unclear. The expression profile and prognostic significance of INHBA in HNSC were assessed using a variety of informatics methods. The level of INHBA expression was significantly higher in patients with HNSC, and it was correlated with sex, tumor-node-metastasis (TNM) stage, histological grade, and human papillomavirus (HPV) status. Kaplan-Meier (K-M) analysis indicated that poor overall survival (OS) and disease-free survival (DFS) were significantly associated with INHBA upregulation in HNSC. INHBA overexpression was validated as an independent poor prognostic factor by multivariate Cox regression, and including INHBA expression level in the prognostic model could increase prediction accuracy. In addition, copy number alterations (CNAs) of INHBA and miR-217-5p downregulation are potential mechanisms for elevated INHBA expression in HNSC. In conclusion, INHBA may represent a promising predictive biomarker and candidate target for anti-TGFβ therapy in HNSC.

Correlation Study between Immunohistochemical Expression of Testicular Inhibin and Seminal Fluid Analysis

The inhibin family of dimeric peptide hormones are produced in the testis and are postulated to have paracrine and autocrine roles in the regulation of steroidogenesis, Testis is a reproductive organ composed of the seminiferous tubules which is the site of spermatogenesis, while the Sertoli and Leydig cells are responsible for secreting hormone; (inhibin), to maintain spermatogenesis. Dissimilarity of activin expression and plasma level it can cause variation to the parameters (sperm count, motility, germinal layer width). Aim of study was evaluation of the inhibin immunohistochemical activity in testicular tissue and the association between the inhibin expression and spermatogenesis. This study was performed in Al-Nahrain University / College of Medicine from November 2020 to June 2021. A total of 20 adult male rabbit. All animals were examined and monitored in order to exclude unhealthy animal. The animals were euthanized by inhalation of chloroform. The animals’ behavior observed during the experiment, in addition, we measured the weights of animals before and after the experiment. Dissection of the Rabbit was done to obtain the testis. We collected seminal fluid by milking the epididymis and then seminal fluid analysis was performed by professional lab technician. The volume of testis was measured. The specimens are kept in 10% formalin for (24-48) hours for histological preparation. In comparison between the two groups of the study (A, B) showed a statistically significant differences in means of sperm count, motility of sperm, germinal layer width, Leydig cell number, and Sertoli cell number between them. Regarding the means of inhibin expression and plasma level in the parameters (sperm count, motility of sperm, germinal layer width) were suggestively advanced in group B compared to that in group A. In conclusion, the expression and level of inhibin subunits, is correlated with parameters (sperm count, motility, germinal layer width), Inhibin peptide hormone acts as inhibitor for FSH-pituitary secretion that in turn promote spermatogenesis. The study shown a highly significant differences of inhibin manufacture and action through spermatogenesis is occupied in the fine regulator of the seminiferous epithelium of the cycle. Also, this study showed a negative correlation between the parameters with inhibin.

Homozygosity for a novel INHA mutation in two male siblings with hypospadias, primary hypogonadism, and high-normal testicular volume.

BackgroundThe human INHA gene encodes the inhibin subunit alpha protein, which is common to both inhibin A and B. The functional importance of inhibins in male sex development, sexual function, and reproduction remain largely unknown.ObjectiveWe report for the first time two male siblings with homozygous INHAmutations.MethodsThe medical files were examined for clinical, biochemical, and imaging data. Genetic analysis was performed using next-generation and Sanger sequencing methods.ResultsTwo brothers complained of gynecomastia, testicular pain, and had a history of hypospadias. Biochemistry revealed low serum testosterone, high gonadotropin and anti-Mullerian hormone, and very low/undetectable inhibin concentrations, where available. Both patients had azoospermia in the spermiogram. We have identified a homozygous 2 bp deletion (c.208_209delAG, R70Gfs*3) variant, which leads to a truncated INHA protein in both patients, and confirmed heterozygosity in the parents. The external genital development, pubertal onset and progression, reproductive functions, serum gonadotropins, and sex hormones of mother and father, who were heterozygous carriers of the identified mutation, were normal.ConclusionHomozygosity for INHA mutations causes decreased prenatal and postnatal testosterone production and infertility in males, while the heterozygous female and male carriers of INHA mutations do not have any abnormality in sex development and reproduction.

CircTHBS1 drives gastric cancer progression by increasing INHBA mRNA expression and stability in a ceRNA- and RBP-dependent manner

Circular RNAs (circRNAs) play vital regulatory roles in the progression of multiple cancers. In our study, transcriptome analysis and self-organizing maps (SOM) were applied to screen backbone circRNAs in gastric cancer (GC). Upon validation of the expression patterns of screened circRNAs, gain- and loss-of-function assays were performed in vitro and in vivo. Underlying mechanisms were investigated using RNA pull-down, luciferase reporter assay and RNA immunoprecipitation. The expression of circTHBS1 was significantly increased in GC and associated with poor prognosis. CircTHBS1 facilitated the malignant behavior and epithelial-to-mesenchymal transition of GC cells. Mechanistically, circTHBS1 sponged miR-204-5p to promote the expression of Inhibin Subunit Beta A (INHBA). Moreover, circTHBS1 could enhance the HuR-mediated mRNA stability of INHBA, which subsequently activated the TGF-β pathway. Our research identified circTHBS1 as an oncogenic circRNA that enhances GC malignancy by elevating INHBA expression, providing new insight and a feasible target for the diagnosis and treatment of GC.

COUP-TFI deletion affects angiogenesis and apoptosis related gene expression in mouse placenta: results of an explorative study

Background: Chicken Ovalbumin Upstream Promoter-Transcription Factor I (COUP-TFI) is a member of the steroid/thyroid nuclear receptor superfamily. The aim of this study was to investigate whether absence of this gene affects placental development and fetal growth in a COUP-TFI knockout mouse model. Methods: Placentas of COUP-TFI-knockout (COUP-TFI KO) and wild-type (WT) were collected at 18.5 days post-coitum. The expression level of the following genes known to be involved in different key molecular pathways was evaluated: BCL2 Associated X (Bax) and B-cell lymphoma 2 (Bcl-2) (apoptosis), p21, p53 and α subunit of inhibin (INHA) (proliferation and apoptosis), vascular endothelial growth factor A (VEGF-A), placental growth factor (PlGF), hypoxia-inducible factor 1-alpha (HIF1α), Fms related receptor tyrosine kinase 1 (Flt-1), and endoglin (ENG) (angiogenesis). Mouse litter weight at birth was also assessed. Results: RT-qPCR analysis showed increased mRNA expression of VEGF-A and Bax in placental tissue of COUP-TFI KO mice compared to WT mice. We also found a loss in the positive correlation between Bcl-2 and INHA, p21 and ENG, as well as HIF1α and Flt-1 mRNA expression in COUP-TFI mutants. Finally, KO mice were lighter than WT littermates (respectively, the mean weight of COUP-TFI KO mice was 1.3 grams, ± 0.13, compared to 1.6 g, ± 0.14 of WT mice, p < 0.05). Conclusions: Our results show that COUP-TFI deletion is associated with a lower birth weight in mice and increased placental transcript expression of pro-apoptotic Bax and pro-angiogenetic VEGF-A genes.

Inhibin subunit beta A promotes cell proliferation and metastasis of breast cancer through Wnt/β-catenin signaling pathway

ABSTRACT Emerging evidence has demonstrated that inhibin subunit beta A (INHBA) is dysregulated and plays a critical role in various cancers. With the development of sequencing technology, studies have discovered that INHBA is overexpressed in breast cancer tissues. However, the biological roles of INHBA in breast cancer are still far to clear. In the present study, we analyzed the INHBA expression in the Cancer Genome Atlas (TCGA) database. Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to assess the expression of INHBA in breast cancer cell lines. Cell proliferation, invasion and epithelial–mesenchymal transition (EMT) were determined by using CCK-8, EdU, Transwell and western blot assays. The result showed that INHBA was highly expressed in breast cancer cell lines. Functional analysis revealed that silence or elevation of INHBA inhibited or promoted the proliferation, migration, invasion and EMT and Wnt/β-catenin signaling pathway-related markers of MCF-7 cells. Mechanically, blocking of Wnt/β-catenin pathway by XAV939 reversed the promotion effect of INHBA overexpression on breast cancer cells’ proliferation, migration and invasion. Our findings emphasized that INHBA may act as an oncogene via activating the Wnt/β-catenin pathway, which may provide a potential therapeutic target for the treatment of breast cancer.

Tubule-derived INHBB promotes interstitial fibroblast activation and renal fibrosis.

Upstream stimuli for myofibroblast activation are of considerable interest for understanding the mechanisms underlying renal fibrosis. Activin B, a member of the TGF-β family, exists as a homodimer of inhibin subunit beta B (INHBB), but its role in renal fibrosis remains unknown. We found that INHBB expression was significantly increased in various renal fibrosis models and human chronic kidney disease specimens with renal fibrosis. Notably, the increase of INHBB occurred mainly in the tubular epithelial cells (TECs). In vivo, inhibiting INHBB blocked the activation of interstitial fibroblasts and ameliorated the renal fibrosis induced by unilateral ureteral obstruction or ischemia-reperfusion injury, while ectopic expression of INHBB in the TECs was able to activate interstitial fibroblasts and initiate interstitial fibrosis. In vitro, overexpression of INHBB in TECs led to the secretion of activin B, thereby promoting the proliferation and activation of interstitial fibroblasts through activin B/Smad signaling. Furthermore, inhibition of activin B/Smad signaling attenuated the fibrotic response caused by tubular INHBB. Mechanistically, the upregulation of INHBB depended on the transcription factor Sox9 in the injured TECs. Clinical analyses also identified a positive correlation between Sox9 and INHBB expression in human specimens, suggesting the Sox9/INHBB axis as a positive regulator of renal fibrosis. In conclusion, tubule-derived INHBB is implicated in the pathogenesis of renal fibrosis by activating the surrounding fibroblasts in a paracrine manner, thereby exhibiting as a potential therapeutic target. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Exosomal lncRNA SNHG10 derived from colorectal cancer cells suppresses natural killer cell cytotoxicity by upregulating INHBC

BackgroundExosome-mediated crosstalk between cancer cells and immune cells contributes to tumor growth. In this study, we investigated the mechanism underlying the exosome-mediated immune escape of colorectal cancer (CRC) cells from natural killer (NK) cells via the transfer of long noncoding RNAs (lncRNAs).MethodsAn epithelial–mesenchymal transition (EMT) model of SW480 cells was established by transforming growth factor beta (TGF-β), followed by the assessment of the effect of EMT-derived exosomes (EMT-exo) on the functions of NK cells. RNA sequencing was performed to identify exosomal lncRNAs and target genes. The function of exosomal lncRNAs in tumor growth was further verified in vivo.ResultsEMT-exo suppressed the proliferation, cytotoxicity, IFN-γ production, and perforin-1 and granzyme B secretion of NK cells. RNA sequencing revealed that SNHG10 expression was upregulated in EMT-exo compared with that in non-EMT-exo. Moreover, SNHG10 expression was upregulated in tumor tissues in CRC, which was associated with poor prognosis. Overexpression of SNHG10 in exosomes (oe-lnc-SNHG10 exo) significantly suppressed the viability and cytotoxicity of NK cells. Transcriptome sequencing of NK cells revealed that the expression levels of 114 genes were upregulated in the oe-lnc-SNHG10 exo group, including inhibin subunit beta C (INHBC), which was involved in the TGF-β signaling pathway. Si-INHBC treatment abrogated the effect of oe-lnc-SNHG10 exo on NK cells. oe-lnc-SNHG10 exo induced tumor growth and upregulated INHBC expression in mice and downregulated the expression of perforin, granzyme B, and NK1.1 in tumor tissues.ConclusionsThe CRC cell-derived exosomal lncRNA SNHG10 suppresses the function of NK cells by upregulating INHBC expression. This study provides evidence that exosomal lncRNAs contribute to immune escape by inducing NK cell inhibition and proposes a potential treatment strategy for CRC.

INHBA transfection regulates proliferation, apoptosis and hormone synthesis in sheep granulosa cells

Inhibin subunit beta A (INHBA) participates in the synthesis of inhibin A, activin A and activin AB. Here we investigated the effect and molecular mechanism of INHBA on proliferation, apoptosis and hormone synthesis in sheep granulosa cells (GCs) using in vitro transfection. We first noticed that INHBA expression increased with follicle diameter and was widely distributed in ovarian tissue. The proliferation rate of GCs was significantly increased and decreased with overexpression and silence of INHBA, respectively, compared with the negative controls. INHBA transfection affected GC proliferation and apoptosis, regulating the expression of many cell cycle-related and apoptosis-related genes. INHBA overexpression significantly decreased activin and estradiol secretion while increasing inhibin and progesterone secretion. The expression of follicle-stimulating hormone beta subunit was significantly decreased and increased with INHBA overexpression and knockdown, respectively. Notably, silence of INHBA inhibited the expression of many transforming growth factor beta-related genes. Overall, the functional molecule of INHBA gene may be associated with follicular development via regulating proliferation, apoptosis and folliculogenesis-related hormone secretion of sheep GCs. In addition, our findings may contribute to a better understanding of the law of follicular development and thus improve the reproductive performance of female animals.

CircRNA-Mediated Inhibin-Activin Balance Regulation in Ovarian Granulosa Cell Apoptosis and Follicular Atresia.

Ovarian granulosa cells (GC) play an essential role in the development and atresia of follicles. Emerging studies suggest that non-coding RNAs are involved in the regulation of GC apoptosis. Here, we aimed to analyze the function of ssc-circINHA-001, coded by the first exon of the inhibin subunit α gene (INHA), in resisting GC apoptosis and follicular atresia by enhancing the expression of the inhibin subunit β A (INHBA) through a cluster of miRNAs. A higher expression of ssc-circINHA-001 in healthy follicles compared to early atretic follicles was detected by qRT-PCR. Its circular structure was confirmed by RNase R treatment and reversed PCR. The function of ssc-circINHA-001 in GC resistance to apoptosis was detected by in vitro transfection of its si-RNA. Furthermore, the dual-luciferase reporter assay suggested that ssc-circINHA-001 adsorbed three miRNAs, termed miR-214-5p, miR-7144-3p, and miR-9830-5p, which share the common target INHBA. A low expression of ssc-circINHA-001 increased the levels of the free miRNAs, inhibited INHBA expression, and thus raised GCs apoptosis through a shift from the secretion of activin to that of inhibin. Our study demonstrated the existence of a circRNA–microRNAs–INHBA regulatory axis in follicular GC apoptosis and provides insight into the relationship between circRNA function and its coding gene in inhibin/activin balance and ovarian physiological functions.

Second trimester maternal serum biomarkers and the risk of cerebral palsy.

To investigate whether second trimester maternal serum screening (2TMSS) biomarkers are associated with cerebral palsy (CP) and identify CP characteristics associated with abnormal biomarker levels. In this retrospective case-control data linkage study, we linked mothers of 129 singleton CP cases from a population register to their 2TMSS records and selected 10 singleton pregnancy controls per case (n=1290). We compared mean and abnormal levels of alpha-fetoprotein (AFP), beta subunit of human chorionic gonadotrophin (β-hCG), unconjugated estriol (uE3), and inhibin between cases and controls and within CP subgroups. Compared to control pregnancies, CP pregnancies had higher mean levels of AFP (1.10 vs. 1.01 multiple of the population median [MoM], p=0.01) and inhibin (1.10 vs. 0.98 MoM, p≤0.01). CP pregnancies were 2.5 times more likely to be associated with high levels of AFP (OR 2.52 [95% confidence interval [CI] 1.30, 4.65]; p<0.01) and 2.6 times for inhibin (OR 2.63 [95% CI 1.37, 4.77]; p<0.01), and 6.8 times when AFP and inhibin were both elevated (OR 6.75 [95% CI 2.41, 18.94]; p<0.01). In CP cases, high AFP and high inhibin levels were associated with preterm birth and low birthweight. Abnormal second-trimester biomarker levels suggest abnormal placentation plays a role in the causal pathway of some CP cases.

INHBA promotes the proliferation, migration and invasion of colon cancer cells through the upregulation of VCAN.

ObjectiveColon cancer has high morbidity and mortality rates, and proliferation, invasion and migration play an important role in colon cancer progression. Here, the effects of inhibin subunit beta A (INHBA) on cell proliferation, invasion and migration were investigated.MethodsThe UALCAN database was used to assess INHBA expression in colon cancer tissues and predict the survival of patients with high and low INHBA expression. The relevant proteins were detected by RT-qPCR and western blot. Cell transfection was performed to overexpress or inhibit INHBA and versican (VCAN). The high correlation between INHBA and VCAN found through LinkedOmics and StarBase databases was verified by immunoprecipitation assays. Cell proliferation was detected by cell counting kit-8 and colony formation assays. Wound healing and Transwell assays were used to assess migration and invasion.ResultsINHBA expression was upregulated in colon cancer tissues and cells. INHBA inhibition impaired the proliferation, migration and invasion of these cells. In addition, we confirmed the correlation between INHBA and VCAN in colon cancer cells. Finally, we found that INHBA interference inhibited the aggressive behavior of colon cancer cells by downregulating VCAN.ConclusionINHBA promotes the proliferation, migration and invasion of colon cancer cells through the upregulation of VCAN.

Effect of NAD+ boosting on kidney ischemia-reperfusion injury.

Acute kidney injury (AKI) is associated with a very high mortality and an increased risk for progression to chronic kidney disease (CKD). Ischemia-reperfusion injury (IRI) is a model for AKI, which results in tubular damage, dysfunction of the mitochondria and autophagy, and in decreased cellular nicotinamide adenine dinucleotide (NAD+) with progressing fibrosis resulting in CKD. NAD+ is a co-enzyme for several proteins, including the NAD+ dependent sirtuins. NAD+ augmentation, e.g. by use of its precursor nicotinamide riboside (NR), improves mitochondrial homeostasis and organismal metabolism in many species. In the present investigation the effects of prophylactic administration of NR on IRI-induced AKI were studied in the rat. Bilateral IRI reduced kidney tissue NAD+, caused tubular damage, reduced α-Klotho (klotho), and altered autophagy flux. AKI initiated progression to CKD, as shown by induced profibrotic Periostin (postn) and Inhibin subunit beta-A, (activin A / Inhba), both 24 hours and 14 days after surgery. NR restored tissue NAD+ to that of the sham group, increased autophagy (reduced p62) and sirtuin1 (Sirt1) but did not ameliorate renal tubular damage and profibrotic genes in the 24 hours and 14 days IRI models. AKI induced NAD+ depletion and impaired autophagy, while augmentation of NAD+ by NR restored tissue NAD+ and increased autophagy, possibly serving as a protective response. However, prophylactic administration of NR did not ameliorate tubular damage of the IRI rats nor rescued the initiation of fibrosis in the long-term AKI to CKD model, which is a pivotal event in CKD pathogenesis.