RESUMO
Myofibroblast buildup and prostatic fibrosis play a crucial role in the development of benign prostatic hyperplasia (BPH). Treatments specifically targeting myofibroblasts could be a promising approach for treating BPH. Tadalafil, a phosphodiesterase type 5 (PDE5) inhibitor, holds the potential to intervene in this biological process. This study employs prostatic stromal fibroblasts to induce myofibroblast differentiation through TGFß1 stimulation. As a result, tadalafil significantly inhibited prostatic stromal fibroblast proliferation and fibrosis process, compared to the control group. Furthermore, our transcriptome sequencing results revealed that tadalafil inhibited FGF9 secretion and simultaneously improved miR-3126-3p expression via TGFß1 suppression. Overall, TGFß1 can trigger pro-fibrotic signaling through miR-3126-3p in the prostatic stroma, and the use of tadalafil can inhibit this process.
Assuntos
Fator 9 de Crescimento de Fibroblastos , Fibrose , MicroRNAs , Inibidores da Fosfodiesterase 5 , Hiperplasia Prostática , Tadalafila , Masculino , Hiperplasia Prostática/metabolismo , Hiperplasia Prostática/tratamento farmacológico , Hiperplasia Prostática/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Tadalafila/farmacologia , Inibidores da Fosfodiesterase 5/farmacologia , Humanos , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/genética , Próstata/efeitos dos fármacos , Próstata/metabolismo , Miofibroblastos/metabolismo , Miofibroblastos/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/genética , Proliferação de Células/efeitos dos fármacosRESUMO
Bone-fat balance is crucial to maintain bone homeostasis. As common progenitor cells of osteoblasts and adipocytes, bone marrow mesenchymal stem cells (BMSCs) are delicately balanced for their differentiation commitment. However, the exact mechanisms governing BMSC cell fate are unclear. In this study, we discovered that fibroblast growth factor 9 (Fgf9), a cytokine expressed in the bone marrow niche, controlled bone-fat balance by influencing the cell fate of BMSCs. Histomorphology and cytodifferentiation analysis showed that Fgf9 loss-of-function mutation (S99N) notably inhibited bone marrow adipose tissue (BMAT) formation and alleviated ovariectomy-induced bone loss and BMAT accumulation in adult mice. Furthermore, in vitro and in vivo investigations demonstrated that Fgf9 altered the differentiation potential of BMSCs, shifting from osteogenesis to adipogenesis at the early stages of cell commitment. Transcriptomic and gene expression analyses demonstrated that FGF9 upregulated the expression of adipogenic genes while downregulating osteogenic gene expression at both mRNA and protein levels. Mechanistic studies revealed that FGF9, through FGFR1, promoted adipogenic gene expression via PI3K/AKT/Hippo pathways and inhibited osteogenic gene expression via MAPK/ERK pathway. This study underscores the crucial role of Fgf9 as a cytokine regulating the bone-fat balance in adult bone, suggesting that FGF9 is a potentially therapeutic target in the treatment of osteoporosis.
Assuntos
Fator 9 de Crescimento de Fibroblastos , Células-Tronco Mesenquimais , Osteoporose , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Animais , Células-Tronco Mesenquimais/metabolismo , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/genética , Camundongos , Osteoporose/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Feminino , Diferenciação Celular , Osteogênese/genética , Sistema de Sinalização das MAP Quinases , Transdução de Sinais , Camundongos Endogâmicos C57BL , Adipogenia , Tecido Adiposo/metabolismoRESUMO
Cap-independent or eukaryotic initiation factor (eIF) 4E-independent, translation initiation in eukaryotes requires scaffolding protein eIF4G or its homolog, death-associated protein 5 (DAP5). eIF4G associates with the 40S ribosomal subunit, recruiting the ribosome to the RNA transcript. A subset of RNA transcripts, such as fibroblast growth factor 9 (FGF-9), contain 5' untranslated regions (5' UTRs) that directly bind DAP5 or eIF4GI. For viral mRNA, eIF recruitment usually utilizes RNA structure, such as a pseudoknot or stem-loops, and the RNA-helicase eIF4A is required for DAP5- or 4G-mediated translation, suggesting these 5' UTRs are structured. However, for cellular IRES-like translation, no consensus RNA structures or sequences have yet been identified for eIF binding. However, the DAP5-binding site within the FGF-9 5' UTR is unknown. Moreover, DAP5 binds to other, dissimilar 5' UTRs, some of which require an unpaired, accessible 5' end to stimulate cap-independent translation. Using SHAPE-seq, we modeled the 186 nt FGF-9 5'-UTR RNA's complex secondary structure in vitro. Further, DAP5 footprinting, toeprinting, and UV cross-linking experiments identify DAP5-RNA interactions. Modeling of FGF-9 5'-UTR tertiary structure aligns DAP5-interacting nucleotides on one face of the predicted structure. We propose that RNA structure involving tertiary folding, rather than a conserved sequence or secondary structure, acts as a DAP5-binding site. DAP5 appears to contact nucleotides near the start codon. Our findings offer a new perspective in the hunt for cap-independent translational enhancers. Structural, rather than sequence-specific, eIF-binding sites may act as attractive chemotherapeutic targets or as dosage tools for mRNA-based therapies.
Assuntos
Regiões 5' não Traduzidas , Fator de Iniciação Eucariótico 4G , Fator 9 de Crescimento de Fibroblastos , Conformação de Ácido Nucleico , Sítios de Ligação , Fator de Iniciação Eucariótico 4G/metabolismo , Fator de Iniciação Eucariótico 4G/genética , Fator de Iniciação Eucariótico 4G/química , Humanos , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/química , Biossíntese de Proteínas , Modelos Moleculares , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/química , Capuzes de RNA/metabolismo , Capuzes de RNA/genética , Capuzes de RNA/químicaRESUMO
BACKGROUND: Asthma is a respiratory disease characterized by airway remodeling. We aimed to find out the role and mechanism of lncRNA MEG3 in asthma. METHODS: We established a cellular model of asthma by inducing human airway smooth muscle cells (HASMCs) with PDGF-BB, and detected levels of lncRNA MEG3, miR-143-3p and FGF9 in HASMCs through qRT-PCR. The functions of lncRNA MEG3 or miR-143-3p on HASMCs were explored by cell transfection. The binding sites of miR-143-3p and FGF9 were subsequently analyzed with bioinformatics software, and validated with dual-luciferase reporter assay. MTT, 5-Ethynyl-2'-deoxyuridine (EdU) assay, and Transwell were used to detect the effects of lncRNA MEG3 or miR-143-3p on proliferation and migration of HASMCs. QRT-PCR and western blot assay were used to evaluate the level of proliferation-related marker PCNA in HASMCs. RESULTS: The study found that lncRNA MEG3 negatively correlated with miR-143-3p, and miR-143-3p could directly target with FGF9. Silence of lncRNA MEG3 can suppress migration and proliferation of PDGF-BB-induced HASMCs via increasing miR-143-3p. Further mechanistic studies revealed that miR-143-3p negatively regulated FGF9 expression in HASMCs. MiR-143-3p could inhibit PDGF-BB-induced HASMCs migration and proliferation through downregulating FGF9. CONCLUSION: LncRNA MEG3 silencing could inhibit the migration and proliferation of HASMCs through regulating miR-143-3p/FGF9 signaling axis. These results imply that lncRNA MEG3 plays a protective role against asthma.
Assuntos
Asma , Movimento Celular , Proliferação de Células , Fator 9 de Crescimento de Fibroblastos , MicroRNAs , Miócitos de Músculo Liso , RNA Longo não Codificante , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Proliferação de Células/genética , Asma/genética , Asma/metabolismo , Miócitos de Músculo Liso/metabolismo , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Células Cultivadas , Remodelação das Vias Aéreas/fisiologia , Remodelação das Vias Aéreas/genéticaRESUMO
Cancer-associated fibroblasts (CAFs) play a pivotal role in cancer progression. This study aimed to explore the roles of CAFs-derived Fibroblast growth factor 9 (FGF9) and Neuro-oncological ventral antigen 1 (NOVA1) in triple negative breast cancer (TNBC) progression. MDA-MB-231 and BT-549 cells were cocultured with CAF conditioned-medium (CAF-CM) or normal fibroblasts conditioned-medium (NF-CM). MTT, EdU, colony formation, wound healing, transwell migration, and invasion assays were employed to determine cell proliferation, migration and invasion, respectively. Western blot and RT-qPCR were carried out to examine the protein and mRNA expression of FGF9 and NOVA1. Xenograft tumor experiments were conducted to evaluate the effects of CAFs, FGF9, and NOVA1 on tumor growth in vivo. Our results showed that CAFs significantly promoted the proliferation, invasion, and migration of TNBC cells. FGF9 and NOVA1 were significantly upregulated in TNBC CAFs, tissues and cells. CAF-CM also could increase the expression of FGF9 and NOVA1 in TNBC cells. Knockdown of FGF9 or NOVA1 could hamper cell proliferation, invasion, migration, and EMT of TNBC cells. Moreover, CAFs with FGF9/NOVA1 knockdown also could inhibit TNBC progression. Besides, CAFs significantly accelerated tumor growth in vivo, which was blocked by FGF9/NOVA1 knockdown in nude mice. In conclusion, our results indicated the tumor-promoting role of CAFs in TNBC progression. FGF9 and NOVA1 upregulation in CAFs induced cell proliferation, migration and invasion in vitro, and facilitated tumor growth in vivo in TNBC development.
Assuntos
Fibroblastos Associados a Câncer , Movimento Celular , Proliferação de Células , Fator 9 de Crescimento de Fibroblastos , Antígeno Neuro-Oncológico Ventral , Proteínas de Ligação a RNA , Neoplasias de Mama Triplo Negativas , Animais , Feminino , Humanos , Camundongos , Fibroblastos Associados a Câncer/metabolismo , Linhagem Celular Tumoral , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Regulação Neoplásica da Expressão Gênica , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Neoplasias de Mama Triplo Negativas/metabolismo , Regulação para Cima , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Fluoride (F) and sulfur dioxide (SO2) contamination is recognized as a public health concern worldwide. Our previous research has shown that Co-exposure to F and SO2 can cause abnormal enamel mineralization. Ameloblastin (AMBN) plays a crucial role in the process of enamel mineralization. However, the process by which simultaneous exposure to F and SO2 influences enamel formation by regulating AMBN expression still needs to be understood. This study aimed to establish in vivo and in vitro models of F-SO2 Co-exposure and investigate the relationship between AMBN and abnormal enamel mineralization. By overexpressing/knocking out the Fibroblast Growth Factor 9 (FGF9) gene, we investigated the impact of FGF9-mediated Mitogen-Activated Protein Kinase (MAPK) signaling on AMBN synthesis to elucidate the mechanism underlying the induction of abnormal enamel mineralization by F-SO2 Co-exposure in rats. The results showed that F-SO2 exposure damaged the structure of rat enamel and ameloblasts. When exposed to F or SO2, gradual increases in the protein expression of FGF9 and phosphorylated p38 mitogen-activated protein kinase (p-P38) were observed. Conversely, the protein levels of AMBN, phosphorylated extracellular signal-regulated kinase (p-ERK), and phosphorylated c-Jun N-terminal kinase (p-JNK) were decreased. AMBN expression was significantly correlated with FGF9, p-ERK, and p-JNK expression in ameloblasts. Interestingly, FGF9 overexpression reduced the levels of p-ERK and p-JNK, worsening the inhibitory effect of F-SO2 on AMBN. Conversely, FGF9 knockout increased the phosphorylation of ERK and JNK, partially reversing the F-SO2-induced downregulation of AMBN. Taken together, these findings strongly demonstrate that FGF9 plays a critical role in F-SO2-induced abnormal enamel mineralization by regulating AMBN synthesis through the JNK and ERK pathways.
Assuntos
Esmalte Dentário , Fator 9 de Crescimento de Fibroblastos , Fluoretos , Sistema de Sinalização das MAP Quinases , Dióxido de Enxofre , Animais , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Ratos , Fluoretos/toxicidade , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Esmalte Dentário/efeitos dos fármacos , Dióxido de Enxofre/toxicidade , Masculino , Ratos Sprague-Dawley , Proteínas do Esmalte Dentário/genética , Proteínas do Esmalte Dentário/metabolismo , Calcificação de Dente/efeitos dos fármacos , Ameloblastos/efeitos dos fármacos , Ameloblastos/metabolismoRESUMO
Coronary atherosclerosis-induced myocardial ischemia leads to cardiomyocyte apoptosis. The regulatory mechanisms for cardiomyocyte apoptosis have not been fully understood. Circular RNAs are non-coding RNAs which play important roles in heart function maintenance and progression of heart diseases by regulating gene transcription and protein translation. Here, we reported a conserved cardiac circular RNA, which is generated from the second exon of LRP6 and named circLRP62-2 . CircLRP62-2 can protect cardiomyocyte from hypoxia-induced apoptosis. The expression of circLRP62-2 in cardiomyocytes was down-regulated under hypoxia, while forced expression of circLRP62-2 inhibited cell apoptosis. Normally, circLRP62-2 was mainly localized in the nucleus. Under hypoxia, circLRP62-2 is associated with heterogeneous nuclear ribonucleoprotein M (hnRNPM) to be translocated into the cytoplasm. It recruited hnRNPM to fibroblast growth factor 9 (FGF9) mRNA to enhance the expression of FGF9 protein, promoting hypoxia-adaption and viability of cardiomyocytes. In summary, this study uncovers a new inhibitor of apoptosis and reveals a novel anti-apoptotic pathway composed of circLRP62-2 , hnRNPM, and FGF9, which may provide therapeutic targets for coronary heart disease and ischemic myocardial injury.
Assuntos
MicroRNAs , Miócitos Cardíacos , Humanos , Miócitos Cardíacos/metabolismo , RNA Circular/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo M/metabolismo , Fator 9 de Crescimento de Fibroblastos/metabolismo , Apoptose/genética , Hipóxia/metabolismo , MicroRNAs/genéticaRESUMO
AIMS: Fibroblast growth factor (FGF) signalling is dysregulated in multiple sclerosis (MS) and other neurological and psychiatric conditions, but there is little or no consensus as to how individual FGF family members contribute to disease pathogenesis. Lesion development in MS is associated with increased expression of FGF1, FGF2 and FGF9, all of which modulate remyelination in a variety of experimental settings. However, FGF9 is also selectively upregulated in major depressive disorder (MDD), prompting us to speculate it may also have a direct effect on neuronal function and survival. METHODS: Transcriptional profiling of myelinating cultures treated with FGF1, FGF2 or FGF9 was performed, and the effects of FGF9 on cortical neurons investigated using a combination of transcriptional, electrophysiological and immunofluorescence microscopic techniques. The in vivo effects of FGF9 were explored by stereotactic injection of adeno-associated viral (AAV) vectors encoding either FGF9 or EGFP into the rat motor cortex. RESULTS: Transcriptional profiling of myelinating cultures after FGF9 treatment revealed a distinct neuronal response with a pronounced downregulation of gene networks associated with axonal transport and synaptic function. In cortical neuronal cultures, FGF9 also rapidly downregulated expression of genes associated with synaptic function. This was associated with a complete block in the development of photo-inducible spiking activity, as demonstrated using multi-electrode recordings of channel rhodopsin-transfected rat cortical neurons in vitro and, ultimately, neuronal cell death. Overexpression of FGF9 in vivo resulted in rapid loss of neurons and subsequent development of chronic grey matter lesions with neuroaxonal reduction and ensuing myelin loss. CONCLUSIONS: These observations identify overexpression of FGF9 as a mechanism by which neuroaxonal pathology could develop independently of immune-mediated demyelination in MS. We suggest targeting neuronal FGF9-dependent pathways may provide a novel strategy to slow if not halt neuroaxonal atrophy and loss in MS, MDD and potentially other neurodegenerative diseases.
Assuntos
Transtorno Depressivo Maior , Esclerose Múltipla , Animais , Ratos , Fator 1 de Crescimento de Fibroblastos , Fator 2 de Crescimento de Fibroblastos , Fator 9 de Crescimento de FibroblastosRESUMO
Granulosa cells (GC) are critical regulators of fertility. During the process of ovarian folliculogenesis, these cells undergo profound changes while producing steroid hormones that are important to control follicular growth, oocyte maturation, and ovulation. Sirtuins are enzymes that regulate several biological processes and have been associated with control of GC function. However, how sirtuins are regulated in GC during ovarian folliculogenesis remains to be unveiled. The present study was designed to investigate effects of hormones that control GC proliferation, differentiation, and steroidogenesis on expression of the seven members of the mammalian sirtuins family (SIRT1-7) and on histone deacetylase activity of nuclear sirtuins (SIRT1, 6, and 7) in GC. Bovine granulosa cells were isolated from small antral follicles (1-5 mm) and were treated with or without follicle-stimulating hormone (FSH), insulin-like growth factor 1 (IGF-1), and fibroblast growth factors 2 (FGF2) and 9 (FGF9). Following treatments, cell proliferation was determined via a cell analyzer, estradiol synthesis and histone deacetylase activity were determined via ELISA, and sirtuins mRNA expression was determined via qPCR. Treatments with FSH and IGF-1 stimulated cell proliferation while addition of FGF2 or FGF9 suppressed estradiol production stimulated by FSH plus IGF-1. In terms of treatments that regulated sirtuins expression in GC, fibroblast growth factors were the most impactful: FGF2 alone increased SIRT1 mRNA expression in comparison to several treatments and increased mRNA abundance of SIRT2 and SIRT7 when added to the combination of FSH and IGF-1; the addition of FGF9 to the combination of FSH and IGF-1 increased mRNA expression of SIRT2, SIRT3, SIRT4, SIRT6, and SIRT7 and increased mRNA expression of SIRT5 in comparison to the negative control group that received no treatment. Also, FGF2 alone increased histone deacetylase activity of sirtuins in comparison to all treatments that contained FSH and/or IGF-1. Furthermore, several correlations were observed between treatments and sirtuins expression and activity, between estradiol or GC numbers and sirtuins expression, and between expression of sirtuins. As FGF2 and FGF9 are considered anti-differentiation factors of GC that stimulate GC proliferation while suppressing estradiol production in combination with FSH and IGF-1, data of this study suggest that sirtuins are associated with control of differentiation of bovine GC.
Assuntos
Hormônio Foliculoestimulante , Fator de Crescimento Insulin-Like I , Feminino , Bovinos , Animais , Hormônio Foliculoestimulante/farmacologia , Hormônio Foliculoestimulante/metabolismo , Fator de Crescimento Insulin-Like I/farmacologia , Fator de Crescimento Insulin-Like I/metabolismo , Fator 2 de Crescimento de Fibroblastos/farmacologia , Sirtuína 1/genética , Sirtuína 1/metabolismo , Sirtuína 2/metabolismo , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/farmacologia , Progesterona/farmacologia , Células da Granulosa , Estradiol/farmacologia , Hormônio Foliculoestimulante Humano/farmacologia , RNA Mensageiro/metabolismo , Células Cultivadas , MamíferosRESUMO
Fibroblast growth factor 9 (FGF9) is crucial for the growth and development of hair follicles (HFs); however, its role in sheep wool growth is unknown. Here, we clarified the role of FGF9 in HF growth in the small-tailed Han sheep by quantifying FGF9 expression in skin tissue sections collected at different periods. Moreover, we evaluated the effects of FGF9 protein supplementation on hair shaft growth in vitro and FGF9 knockdown on cultured dermal papilla cells (DPCs). The relationship between FGF9 and the Wnt/ß-catenin signaling pathway was examined, and the underlying mechanisms of FGF9-mediated DPC proliferation were investigated. The results show that FGF9 expression varies throughout the HF cycle and participates in wool growth. The proliferation rate and cell cycle of FGF9-treated DPCs substantially increase compared to that of the control group, and the mRNA and protein expression of CTNNB1, a Wnt/ß-catenin signaling pathway marker gene, is considerably lower than that in the control group. The opposite occurs in FGF9-knockdown DPCs. Moreover, other signaling pathways are enriched in the FGF9-treated group. In conclusion, FGF9 accelerates the proliferation and cell cycle of DPCs and may regulate HF growth and development through the Wnt/ß-catenin signaling pathway.
Assuntos
Fator 9 de Crescimento de Fibroblastos , Folículo Piloso , Animais , Ovinos , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Proliferação de Células , Cabelo , Via de Sinalização WntRESUMO
The purpose of this study was to investigate whether fibroblast growth factor 4 (FGF4) and FGF9 are active in dentin differentiation. Dentin matrix protein 1 (Dmp1) -2A-Cre transgenic mice, which express the Cre-recombinase in Dmp1-expressing cells, were crossed with CAG-tdTomato mice as reporter mouse. The cell proliferation and tdTomato expressions were observed. The mesenchymal cell separated from neonatal molar tooth germ were cultured with or without FGF4, FGF9, and with or without their inhibitors ferulic acid and infigratinib (BGJ398) for 21 days. Their phenotypes were evaluated by cell count, flow cytometry, and real-time PCR. Immunohistochemistry for FGFR1, 2, and 3 expression and the expression of DMP1 were performed. FGF4 treatment of mesenchymal cells obtained promoted the expression of all odontoblast markers. FGF9 failed to enhance dentin sialophosphoprotein (Dspp) expression levels. Runt-related transcription factor 2 (Runx2) was upregulated until day 14 but was downregulated on day 21. Compared to Dmp1-negative cells, Dmp1-positive cells expressed higher levels of all odontoblast markers, except for Runx2. Simultaneous treatment with FGF4 and FGF9 had a synergistic effect on odontoblast differentiation, suggesting that they may play a role in odontoblast maturation.
Assuntos
Subunidade alfa 1 de Fator de Ligação ao Core , Fator 4 de Crescimento de Fibroblastos , Fator 9 de Crescimento de Fibroblastos , Odontoblastos , Animais , Camundongos , Diferenciação Celular , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Fator 4 de Crescimento de Fibroblastos/genética , Fator 4 de Crescimento de Fibroblastos/metabolismo , Camundongos Transgênicos , Odontoblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismoRESUMO
Background: The pathogenesis of osteoarthritis (OA) is complex and there is no specific drug for treatment. The aim of this study was to identify the molecular targets of OA therapy, focusing on the expression and biological functions of miR-182-5p and its target genes in OA. Methods: miR-182-5p and fibroblast growth factor 9 (FGF9) were overexpressed or knocked down in IL-1ß-induced chondrocytes. An OA knee model was performed by surgically destroying the medial meniscus. The gene expression of miR-182-5p and FGF9 was calculated. The protein FGF9 was tested by western blotting. Cell counting kit-8 (CCK8), plate cloning assay, and flow cytometry were conducted to evaluate cell proliferation and apoptosis. The expression of inflammatory factors, tumor necrosis factor-alpha (TNF-α), interleukin (IL)-6, and interleukin (IL)-8, was evaluated using enzyme-linked immunosorbent assay (ELISA). Dual-luciferase reporter assays validated the targeting relationship between miR-182-5p and FGF9. Hematoxylin-eosin (HE) and safranin O-fast Green (S-O) staining were utilized to access cartilage damage. Ki67 expression in cartilage was detected using immunohistochemistry (IHC). TdT-mediated dUTP nick-end labeling (TUNEL) assays were used to calculate the apoptosis rate of cartilage. Results: The expression of miR-182-5p was upregulated, and FGF9 was downregulated in the IL-1ß-induced chondrocytes. OA chondrocytes proliferation ability in the miR-182-5p mimics group was decreased, and the apoptosis rate and inflammatory factor were increased. Transfection with miR-182-5p inhibitor increased the proliferative ability and decreased the apoptosis rate in the IL-1ß-induced chondrocytes. Transfection with miR-182-5p inhibitor reversed IL-1ß-induced inflammatory factor release in chondrocytes. Targeted binding sites existed between miR-182-5p and FGF9. After overexpression of FGF9, the miR-182-5p effect on OA chondrocytes was reversed. The hyaline cartilage thickness and proteoglycan content decreased in OA rats, and this was reversed by miR-182-5p inhibitor treatment. Conclusions: miR-182-5p expression levels were increased in OA chondrocytes and regulated chondrocyte proliferation, apoptosis, and inflammation by targeting FGF9. miR-182-5p is a potential gene for OA treatment.
Assuntos
MicroRNAs , Osteoartrite , Ratos , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/farmacologia , Osteoartrite/genética , Osteoartrite/metabolismo , Osteoartrite/patologia , Inflamação/patologia , Apoptose , Interleucina-6/metabolismo , Condrócitos/metabolismo , Condrócitos/patologia , Interleucina-1beta/metabolismoRESUMO
Multiple synostoses syndrome (OMIM: #186500, #610017, #612961, #617898) is a genetically heterogeneous group of autosomal dominant diseases characterized by abnormal bone unions. The joint fusions frequently involve the hands, feet, elbows or vertebrae. Pathogenic variants in FGF9 have been associated with multiple synostoses syndrome type 3 (SYNS3). So far, only five different missense variants in FGF9 that cause SYNS3 have been reported in 18 affected individuals. Unlike other multiple synostoses syndromes, conductive hearing loss has not been reported in SYNS3. In this report, we describe the clinical and selected radiological findings in a large multigenerational family with a novel missense variant in FGF9: c.430T>C, p.(Trp144Arg). We extend the phenotypic spectrum of SYNS3 by suggesting that cleft palate and conductive hearing loss are part of the syndrome and highlight the high degree of intrafamilial phenotypic variability. These findings should be considered when counseling affected individuals.
Assuntos
Perda Auditiva Condutiva , Sinostose , Humanos , Família Estendida , Fator 9 de Crescimento de Fibroblastos , Perda Auditiva Condutiva/genética , Mutação de Sentido Incorreto , SíndromeRESUMO
The enthesis is a transitional tissue between tendon and bone that matures postnatally. The development and maturation of the enthesis involve cellular processes likened to an arrested growth plate. In this study, we explored the role of fibroblast growth factor 9 (Fgf9), a known regulator of chondrogenesis and vascularization during bone development, on the structure and function of the postnatal enthesis. First, we confirmed spatial expression of Fgf9 in the tendon and enthesis using in situ hybridization. We then used Cre-lox recombinase to conditionally knockout Fgf9 in mouse tendon and enthesis (Scx-Cre) and characterized enthesis morphology as well as mechanical properties in Fgf9ScxCre and wild-type (WT) entheses. Fgf9ScxCre mice had smaller calcaneal and humeral apophyses, thinner cortical bone at the attachment, increased cellularity, and reduced failure load in mature entheses compared to WT littermates. During postnatal development, we found reduced chondrocyte hypertrophy and disrupted type X collagen (Col X) in Fgf9ScxCre entheses. These findings support that tendon-derived Fgf9 is important for functional development of the enthesis, including its postnatal mineralization. Our findings suggest the potential role of FGF signaling during enthesis development.
Assuntos
Fator 9 de Crescimento de Fibroblastos , Tendões , Camundongos , Animais , Fator 9 de Crescimento de Fibroblastos/genética , Fator 9 de Crescimento de Fibroblastos/metabolismo , Tendões/metabolismo , Osso e Ossos , Desenvolvimento Ósseo/genética , CondrogêneseRESUMO
We developed a Biomedical Knowledge Graph model that is phenotype and biological function-aware through integrating knowledge from multiple domains in a Neo4j, graph database. All known human genes were assessed through the model to identify potential new risk genes for anterior cruciate ligament (ACL) ruptures and Achilles tendinopathy (AT). Genes were prioritised and explored in a case-control study comparing participants with ACL ruptures (ACL-R), including a sub-group with non-contact mechanism injuries (ACL-NON), to uninjured control individuals (CON). After gene filtering, 3376 genes, including 411 genes identified through previous whole exome sequencing, were found to be potentially linked to AT and ACL ruptures. Four variants were prioritised: HSPG2:rs2291826A/G, HSPG2:rs2291827G/A, ITGB2:rs2230528C/T and FGF9:rs2274296C/T. The rs2230528 CC genotype was over-represented in the CON group compared to ACL-R (p < 0.001) and ACL-NON (p < 0.001) and the TT genotype and T allele were over-represented in the ACL-R group and ACL-NON compared to CON (p < 0.001) group. Several significant differences in distributions were noted for the gene-gene interactions: (HSPG2:rs2291826, rs2291827 and ITGB2:rs2230528) and (ITGB2:rs2230528 and FGF9:rs2297429). This study substantiates the efficiency of using a prior knowledge-driven in silico approach to identify candidate genes linked to tendon and ACL injuries. Our biomedical knowledge graph identified and, with further testing, highlighted novel associations of the ITGB2 gene which has not been explored in a genetic case control association study, with ACL rupture risk. We thus recommend a multistep approach including bioinformatics in conjunction with next generation sequencing technology to improve the discovery potential of genomics technologies in musculoskeletal soft tissue injuries.HighlightsA biomedical knowledge graph was modelled for musculoskeletal soft tissue injuries to efficiently identify candidate genes for genetic susceptibility analyses.The biomedical knowledge graph and sequencing data identified potential biologically relevant variants to explore susceptibility to common tendon and ligament injuries. Specifically genetic variants within the ITGB2 and FGF9 genes were associated with ACL risk.Novel allele combinations (HSPG2-ITGB2 and ITGB2-FGF9) showcase the potential effect of ITGB2 in influencing risk of ACL rupture.
Assuntos
Tendão do Calcâneo , Lesões do Ligamento Cruzado Anterior , Tendinopatia , Humanos , Lesões do Ligamento Cruzado Anterior/genética , Ligamento Cruzado Anterior , Predisposição Genética para Doença , Estudos de Casos e Controles , Tendinopatia/genética , Loci Gênicos , Ruptura/genética , Fator 9 de Crescimento de Fibroblastos/genéticaRESUMO
Osteoarthritis (OA) is a degenerative and progressive disease that affects joints. Pathologically, it is characterized by oxidative stress-mediated excessive chondrocyte apoptosis and mitochondrial dysfunction. Fibroblast growth factor 9 (FGF9) has been shown to exert antioxidant effects and prevent degenerative diseases by activating ERK-related signaling pathways. However, the mechanism of FGF9 in the pathogenesis of OA and its relationship with anti-oxidative stress and related pathways are unclear. In this study, mice with medial meniscus instability (DMM) were used as the in vivo model whereas TBHP-induced chondrocytes served as the in vitro model to explore the mechanism underlying the effects of FGF9 in OA and its association with anti-oxidative stress. Results showed that FGF9 reduced oxidative stress, apoptosis, and mitochondrial dysfunction in TBHP-treated chondrocytes and promoted the nuclear translocation of Nrf2 to activate the Nrf2/HO1 signaling pathway. Interestingly, silencing the Nrf2 gene or blocking the ERK signaling pathway abolished the antioxidant effects of FGF9. FGF9 treatment reduced joint space narrowing, cartilage ossification, and synovial thickening in the DMM model mice. In conclusion, the present findings demonstrate that FGF9 can inhibit TBHP-induced oxidative stress in chondrocytes through the ERK and Nrf2-HO1 signaling pathways and prevent the progression of OA in vivo.
Assuntos
Antioxidantes , Osteoartrite , Animais , Camundongos , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Antioxidantes/metabolismo , Apoptose , Condrócitos , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Osteoartrite/metabolismo , Estresse Oxidativo , Transdução de Sinais , Sistema de Sinalização das MAP QuinasesRESUMO
46,XY gonadal dysgenesis (GD) is a Disorder/Difference of Sex Development (DSD) that can present with phenotypes ranging from ambiguous genitalia to complete male-to-female sex reversal. Around 50% of 46,XY DSD cases receive a molecular diagnosis. In mice, Fibroblast growth factor 9 (FGF9) is an important component of the male sex-determining pathway. Two FGF9 variants reported to date disrupt testis development in mice, but not in humans. Here, we describe a female patient with 46,XY GD harbouring the rare FGF9 variant (missense mutation), NM_002010.2:c.583G > A;p.(Asp195Asn) (D195N). By biochemical and cell-based approaches, the D195N variant disrupts FGF9 protein homodimerisation and FGF9-heparin-binding, and reduces both Sertoli cell proliferation and Wnt4 repression. XY Fgf9D195N/D195N foetal mice show a transient disruption of testicular cord development, while XY Fgf9D195N/- foetal mice show partial male-to-female gonadal sex reversal. In the general population, the D195N variant occurs at an allele frequency of 2.4 × 10-5 , suggesting an oligogenic basis for the patient's DSD. Exome analysis of the patient reveals several known and novel variants in genes expressed in human foetal Sertoli cells at the time of sex determination. Taken together, our results indicate that disruption of FGF9 homodimerization impairs testis determination in mice and, potentially, also in humans in combination with other variants.
Assuntos
Fator 9 de Crescimento de Fibroblastos , Disgenesia Gonadal 46 XY , Humanos , Masculino , Feminino , Camundongos , Animais , Dimerização , Fator 9 de Crescimento de Fibroblastos/genética , Testículo , Gônadas , Disgenesia Gonadal 46 XY/genéticaRESUMO
BACKGROUND: Fgf9 mutation was found in cleft palate patients. Our previous study indicated that Fgf9 promotes timely elevation of palate by regulating hyaluronic acid (HA) accumulation at embryonic day 13.5 (E13.5). HA is synthesized by hyaluronic acid synthases (HAS) isoforms 1, 2, or 3. However, how FGF9 regulates HA in palatogenesis is still unclear. METHODS: Using Ddx4-Cre mice, we generated the Fgf9-/- mouse model (with exon 2 deletion). Immunohistochemistry was used to detect the location and expression of HAS2 in WT and the Fgf9-/- palate at E13.5. We also predicted the association between Fgf9 and Has2 within the developing palate by performing a bioinformatics analysis. The expression of ß-catenin, HAS2, and TCF7L2 were verified by Western blotting after knockout of Fgf9. Rescue experiments were performed by ELISA in vitro. RESULTS: Fgf9-/- mice exhibited 100% penetrance of the cleft palate. A knockout of Fgf9 confirmed that HAS2 and TCF7L2 expression was positively correlated with FGF9. TCF7L2 binds to the Has2 promoter, exhibiting the high specificity predicted by JASPAR. Additionally, increased HA expression by BML-284, TCF-dependent agonist, was blocked in Fgf9-/- palate because of the significant decline in TCF7L2 expression. CONCLUSIONS: FGF9 promotes HAS2 expression via Wnt/ß-catenin/TCF7L2 pathway with TCF7L2 activating transcription of Has2 in the palate.
Assuntos
Fissura Palatina , beta Catenina , Camundongos , Animais , Fissura Palatina/genética , Ácido Hialurônico , Via de Sinalização Wnt , Fator 9 de Crescimento de Fibroblastos/genética , Proteína 2 Semelhante ao Fator 7 de Transcrição/genéticaRESUMO
Acute myocardial infarction (AMI) leads to anoxia and ischemia of cardiomyocytes, followed by apoptosis. This study investigated the protective effect of ginsenoside Rg1 (Rg1) on myocardial ischemia injury in rats with AMI. Rats were randomly divided into five groups: group A (blank control group), group B (hypoxia/reoxygenation group), group C (hypoxia/reoxygenation + 10 mg/L Rg1), group D (hypoxia/reoxygenation + 20 mg/L Rg1) and group E (hypoxia/reoxygenation + 40 mg/L Rg1). The survival rate, apoptosis rate, expression of cyclin-dependent kinase 4 (CDK4), fibroblast growth factor 9 (FGF9), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), microvessel density and myocardial infarction area of rats in each group were compared. The expressions of CDK4 and FGF9, the contents of SOD and GSH-Px in groups C, D and E injected with Rg1 were significantly promoted compared to group B without Rg1 injection (P < 0.05). The survival rate of myocardial cells was significantly increased while the apoptosis rate was significantly decreased in group C, D, E compared to group B (P < 0.05). On the 3rd, 7th and 10th day following Rg1 treatment, the infarct area of E group was significantly decreased in three groups C, D, E, and the microvessel density of infarct area was significantly increased compared with group B (P < 0.05). So, Rg1 can improve the survival rate of myocardial cells, reduce the apoptosis rate and the area of myocardial infarction, and increase the microvessel density of infarct area, thus playing a protective role in ischemic myocardial cells of AMI rats.
Assuntos
Ginsenosídeos , Infarto do Miocárdio , Animais , Ratos , Ginsenosídeos/farmacologia , Ginsenosídeos/metabolismo , Miócitos Cardíacos , Fator 9 de Crescimento de Fibroblastos/metabolismo , Fator 9 de Crescimento de Fibroblastos/farmacologia , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/farmacologia , Glutationa Peroxidase/metabolismo , Ratos Sprague-Dawley , Infarto do Miocárdio/prevenção & controle , Infarto do Miocárdio/metabolismo , Apoptose , Superóxido Dismutase/metabolismo , HipóxiaRESUMO
Neural tissue maturation is a coordinated process under tight transcriptional control. We previously analyzed the kinetics of gene expression in the medial nucleus of the trapezoid body (MNTB) in the brainstem during the critical postnatal phase of its development. While this work revealed timed execution of transcriptional programs, it was blind to the specific cells where gene expression changes occurred. Here, we utilized single-cell RNA-Seq to determine transcriptional profiles of each major MNTB cell type. We discerned directional signaling patterns between neuronal, glial, and vascular-associated cells for VEGF, TGFß, and Delta-Notch pathways during a robust period of vascular remodeling in the MNTB. Furthermore, we describe functional outcomes of the disruption of neuron-astrocyte fibroblast growth factor 9 (Fgf9) signaling. We used a conditional KO (cKO) approach to genetically delete Fgf9 from principal neurons in the MNTB, which led to an early onset of glial fibrillary acidic protein (Gfap) expression in astrocytes. In turn, Fgf9 cKO mice show increased levels of astrocyte-enriched brevican (Bcan), a component of the perineuronal net matrix that ensheaths principal neurons in the MNTB and the large calyx of Held terminal, while levels of the neuron-enriched hyaluronan and proteoglycan link protein 1 (Hapln1) were unchanged. Finally, volumetric analysis of vesicular glutamate transporters 1 and 2 (Vglut1/2), which serves as a proxy for terminal size, revealed an increase in calyx of Held volume in the Fgf9 cKO. Overall, we demonstrate a coordinated neuron-astrocyte Fgf9 signaling network that functions to regulate astrocyte maturation, perineuronal net structure, and synaptic refinement.